WorldWideScience

Sample records for therapeutic gene delivery

  1. Gene therapy prospects--intranasal delivery of therapeutic genes.

    Science.gov (United States)

    Podolska, Karolina; Stachurska, Anna; Hajdukiewicz, Karolina; Małecki, Maciej

    2012-01-01

    Gene therapy is recognized to be a novel method for the treatment of various disorders. Gene therapy strategies involve gene manipulation on broad biological processes responsible for the spreading of diseases. Cancer, monogenic diseases, vascular and infectious diseases are the main targets of gene therapy. In order to obtain valuable experimental and clinical results, sufficient gene transfer methods are required. Therapeutic genes can be administered into target tissues via gene carriers commonly defined as vectors. The retroviral, adenoviral and adeno-associated virus based vectors are most frequently used in the clinic. So far, gene preparations may be administered directly into target organs or by intravenous, intramuscular, intratumor or intranasal injections. It is common knowledge that the number of gene therapy clinical trials has rapidly increased. However, some limitations such as transfection efficiency and stable and long-term gene expression are still not resolved. Consequently, great effort is focused on the evaluation of new strategies of gene delivery. There are many expectations associated with intranasal delivery of gene preparations for the treatment of diseases. Intranasal delivery of therapeutic genes is regarded as one of the most promising forms of pulmonary gene therapy research. Gene therapy based on inhalation of gene preparations offers an alternative way for the treatment of patients suffering from such lung diseases as cystic fibrosis, alpha-1-antitrypsin defect, or cancer. Experimental and first clinical trials based on plasmid vectors or recombinant viruses have revealed that gene preparations can effectively deliver therapeutic or marker genes to the cells of the respiratory tract. The noninvasive intranasal delivery of gene preparations or conventional drugs seems to be very encouraging, although basic scientific research still has to continue.

  2. Gene delivery to skeletal muscle results in sustained expression and systemic delivery of a therapeutic protein.

    Science.gov (United States)

    Kessler, P D; Podsakoff, G M; Chen, X; McQuiston, S A; Colosi, P C; Matelis, L A; Kurtzman, G J; Byrne, B J

    1996-11-26

    Somatic gene therapy has been proposed as a means to achieve systemic delivery of therapeutic proteins. However, there is limited evidence that current methods of gene delivery can practically achieve this goal. In this study, we demonstrate that, following a single intramuscular administration of a recombinant adeno-associated virus (rAAV) vector containing the beta-galactosidase (AAV-lacZ) gene into adult BALB/c mice, protein expression was detected in myofibers for at least 32 weeks. A single intramuscular administration of an AAV vector containing a gene for human erythropoietin (AAV-Epo) into mice resulted in dose-dependent secretion of erythropoietin and corresponding increases in red blood cell production that persisted for up to 40 weeks. Primary human myotubes transduced in vitro with the AAV-Epo vector also showed dose-dependent production of Epo. These results demonstrate that rAAV vectors are able to transduce skeletal muscle and are capable of achieving sustained expression and systemic delivery of a therapeutic protein following a single intramuscular administration. Gene therapy using AAV vectors may provide a practical strategy for the treatment of inherited and acquired protein deficiencies.

  3. Intracellular delivery of potential therapeutic genes: prospects in cancer gene therapy.

    Science.gov (United States)

    Bakhtiar, Athirah; Sayyad, Mustak; Rosli, Rozita; Maruyama, Atsushi; Chowdhury, Ezharul H

    2014-01-01

    Conventional therapies for malignant cancer such as chemotherapy and radiotherapy are associated with poor survival rates owing to the development of cellular resistance to cancer drugs and the lack of targetability, resulting in unwanted adverse effects on healthy cells and necessitating the lowering of therapeutic dose with consequential lower efficacy of the treatment. Gene therapy employing different types of viral and non-viral carriers to transport gene(s) of interest and facilitating production of the desirable therapeutic protein(s) has tremendous prospects in cancer treatments due to the high-level of specificity in therapeutic action of the expressed protein(s) with diminished off-target effects, although cancer cell-specific delivery of transgene(s) still poses some challenges to be addressed. Depending on the potential therapeutic target genes, cancer gene therapy could be categorized into tumor suppressor gene replacement therapy, immune gene therapy and enzyme- or prodrug-based therapy. This review would shed light on the current progress of delivery of potentially therapeutic genes into various cancer cells in vitro and animal models utilizing a variety of viral and non-viral vectors.

  4. Irradiation promotes Akt-targeting therapeutic gene delivery to the tumor vasculature

    International Nuclear Information System (INIS)

    Sonveaux, Pierre; Frerart, Francoise; Bouzin, Caroline; Brouet, Agnes; Wever, Julie de; Jordan, Benedicte F.; Gallez, Bernard; Feron, Olivier

    2007-01-01

    Purpose: To determine whether radiation-induced increases in nitric oxide (NO) production can influence tumor blood flow and improve delivery of Akt-targeting therapeutic DNA lipocomplexes to the tumor. Methods and Materials: The contribution of NO to the endothelial response to radiation was identified using NO synthase (NOS) inhibitors and endothelial NOS (eNOS)-deficient mice. Reporter-encoding plasmids complexed with cationic lipids were used to document the tumor vascular specificity and the efficacy of in vivo lipofection after irradiation. A dominant-negative Akt gene construct was used to evaluate the facilitating effects of radiotherapy on the therapeutic transgene delivery. Results: The abundance of eNOS protein was increased in both irradiated tumor microvessels and endothelial cells, leading to a stimulation of NO release and an associated increase in tumor blood flow. Transgene expression was subsequently improved in the irradiated vs. nonirradiated tumor vasculature. This effect was not apparent in eNOS-deficient mice and could not be reproduced in irradiated cultured endothelial cells. Finally, we combined low-dose radiotherapy with a dominant-negative Akt gene construct and documented synergistic antitumor effects. Conclusions: This study offers a new rationale to combine radiotherapy with gene therapy, by directly exploiting the stimulatory effects of radiation on NO production by tumor endothelial cells. The preferential expression of the transgene in the tumor microvasculature underscores the potential of such an adjuvant strategy to limit the angiogenic response of irradiated tumors

  5. Catalytic immunoglobulin gene delivery in a mouse model of Alzheimer's disease: prophylactic and therapeutic applications.

    Science.gov (United States)

    Kou, Jinghong; Yang, Junling; Lim, Jeong-Eun; Pattanayak, Abhinandan; Song, Min; Planque, Stephanie; Paul, Sudhir; Fukuchi, Ken-Ichiro

    2015-02-01

    Accumulation of amyloid beta-peptide (Aβ) in the brain is hypothesized to be a causal event leading to dementia in Alzheimer's disease (AD). Aβ vaccination removes Aβ deposits from the brain. Aβ immunotherapy, however, may cause T cell- and/or Fc-receptor-mediated brain inflammation and relocate parenchymal Aβ deposits to blood vessels leading to cerebral hemorrhages. Because catalytic antibodies do not form stable immune complexes and Aβ fragments produced by catalytic antibodies are less likely to form aggregates, Aβ-specific catalytic antibodies may have safer therapeutic profiles than reversibly-binding anti-Aβ antibodies. Additionally, catalytic antibodies may remove Aβ more efficiently than binding antibodies because a single catalytic antibody can hydrolyze thousands of Aβ molecules. We previously isolated Aβ-specific catalytic antibody, IgVL5D3, with strong Aβ-hydrolyzing activity. Here, we evaluated the prophylactic and therapeutic efficacy of brain-targeted IgVL5D3 gene delivery via recombinant adeno-associated virus serotype 9 (rAAV9) in an AD mouse model. One single injection of rAAV9-IgVL5D3 into the right ventricle of AD model mice yielded widespread, high expression of IgVL5D3 in the unilateral hemisphere. IgVL5D3 expression was readily detectable in the contralateral hemisphere but to a much lesser extent. IgVL5D3 expression was also confirmed in the cerebrospinal fluid. Prophylactic and therapeutic injection of rAAV9-IgVL5D3 reduced Aβ load in the ipsilateral hippocampus of AD model mice. No evidence of hemorrhages, increased vascular amyloid deposits, increased proinflammatory cytokines, or infiltrating T-cells in the brains was found in the experimental animals. AAV9-mediated anti-Aβ catalytic antibody brain delivery can be prophylactic and therapeutic options for AD.

  6. Delivery strategies of the CRISPR-Cas9 gene-editing system for therapeutic applications.

    Science.gov (United States)

    Liu, Chang; Zhang, Li; Liu, Hao; Cheng, Kun

    2017-11-28

    The CRISPR-Cas9 genome-editing system is a part of the adaptive immune system in archaea and bacteria to defend against invasive nucleic acids from phages and plasmids. The single guide RNA (sgRNA) of the system recognizes its target sequence in the genome, and the Cas9 nuclease of the system acts as a pair of scissors to cleave the double strands of DNA. Since its discovery, CRISPR-Cas9 has become the most robust platform for genome engineering in eukaryotic cells. Recently, the CRISPR-Cas9 system has triggered enormous interest in therapeutic applications. CRISPR-Cas9 can be applied to correct disease-causing gene mutations or engineer T cells for cancer immunotherapy. The first clinical trial using the CRISPR-Cas9 technology was conducted in 2016. Despite the great promise of the CRISPR-Cas9 technology, several challenges remain to be tackled before its successful applications for human patients. The greatest challenge is the safe and efficient delivery of the CRISPR-Cas9 genome-editing system to target cells in human body. In this review, we will introduce the molecular mechanism and different strategies to edit genes using the CRISPR-Cas9 system. We will then highlight the current systems that have been developed to deliver CRISPR-Cas9 in vitro and in vivo for various therapeutic purposes. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Magnetic nanoparticles for targeted therapeutic gene delivery and magnetic-inducing heating on hepatoma

    International Nuclear Information System (INIS)

    Yuan, Chenyan; Zhang, Jia; Li, Hongbo; Zhang, Hao; Wang, Ling; Zhang, Dongsheng; An, Yanli

    2014-01-01

    Gene therapy holds great promise for treating cancers, but their clinical applications are being hampered due to uncontrolled gene delivery and expression. To develop a targeted, safe and efficient tumor therapy system, we constructed a tissue-specific suicide gene delivery system by using magnetic nanoparticles (MNPs) as carriers for the combination of gene therapy and hyperthermia on hepatoma. The suicide gene was hepatoma-targeted and hypoxia-enhanced, and the MNPs possessed the ability to elevate temperature to the effective range for tumor hyperthermia as imposed on an alternating magnetic field (AMF). The tumoricidal effects of targeted gene therapy associated with hyperthermia were evaluated in vitro and in vivo. The experiment demonstrated that hyperthermia combined with a targeted gene therapy system proffer an effective tool for tumor therapy with high selectivity and the synergistic effect of hepatoma suppression. (paper)

  8. Ultrasound and microbubble-targeted delivery of therapeutic compounds : ICIN Report Project 49: Drug and gene delivery through ultrasound and microbubbles

    NARCIS (Netherlands)

    Juffermans, L J M; Meijering, D B M; van Wamel, A; Henning, R H; Kooiman, K; Emmer, M; de Jong, N; van Gilst, W H; Musters, R; Paulus, W J; van Rossum, A C; Deelman, L E; Kamp, O

    The molecular understanding of diseases has been accelerated in recent years, producing many new potential therapeutic targets. A noninvasive delivery system that can target specific anatomical sites would be a great boost for many therapies, particularly those based on manipulation of gene

  9. Lipopolyplex for therapeutic gene delivery and its application for the treatment of Parkinson’s disease

    Directory of Open Access Journals (Sweden)

    Wei eChen

    2016-04-01

    Full Text Available Abstract: Lipopolyplex is a core-shell structure composed of nucleic acid, polycation and lipid. As a non-viral gene delivery vector, lipopolyplex combining the advantages of polyplex and lipoplex has shown superior colloidal stability, reduced cytotoxicity, extremely high gene transfection efficiency. Following intravenous administration, there are many strategies based on lipopolyplex to overcome the complex biological barriers in systemic gene delivery including condensation of nucleic acids into nanoparticles, long circulation, cell targeting, endosomal escape, release to cytoplasm and entry into cell nucleus. Parkinson’s disease is the second most common neurodegenerative disorder and severely influences the patients’ life quality. Current gene therapy clinical trials for Parkinson’s disease employing viral vectors didn’t achieve satisfactory efficacy. However, lipopolyplex may become a promising alternative approach owing to its stability in blood, ability to cross the blood-brain barrier and specific targeting to diseased brain cells.

  10. Self-focusing therapeutic gene delivery with intelligent gene vector swarms: intra-swarm signalling through receptor transgene expression in targeted cells.

    Science.gov (United States)

    Tolmachov, Oleg E

    2015-01-01

    Gene delivery in vivo that is tightly focused on the intended target cells is essential to maximize the benefits of gene therapy and to reduce unwanted side-effects. Cell surface markers are immediately available for probing by therapeutic gene vectors and are often used to direct gene transfer with these vectors to specific target cell populations. However, it is not unusual for the choice of available extra-cellular markers to be too scarce to provide a reliable definition of the desired therapeutically relevant set of target cells. Therefore, interrogation of intra-cellular determinants of cell-specificity, such as tissue-specific transcription factors, can be vital in order to provide detailed cell-guiding information to gene vector particles. An important improvement in cell-specific gene delivery can be achieved through auto-buildup in vector homing efficiency using intelligent 'self-focusing' of swarms of vector particles on target cells. Vector self-focusing was previously suggested to rely on the release of diffusible chemo-attractants after a successful target-specific hit by 'scout' vector particles. I hypothesize that intelligent self-focusing behaviour of swarms of cell-targeted therapeutic gene vectors can be accomplished without the employment of difficult-to-use diffusible chemo-attractants, instead relying on the intra-swarm signalling through cells expressing a non-diffusible extra-cellular receptor for the gene vectors. In the proposed model, cell-guiding information is gathered by the 'scout' gene vector particles, which: (1) attach to a variety of cells via a weakly binding (low affinity) receptor; (2) successfully facilitate gene transfer into these cells; (3) query intra-cellular determinants of cell-specificity with their transgene expression control elements and (4) direct the cell-specific biosynthesis of a vector-encoded strongly binding (high affinity) cell-surface receptor. Free members of the vector swarm loaded with therapeutic cargo

  11. RNAi-based therapeutic nanostrategy: IL-8 gene silencing in pancreatic cancer cells using gold nanorods delivery vehicles

    International Nuclear Information System (INIS)

    Panwar, Nishtha; Yang, Chengbin; Yin, Feng; Chuan, Tjin Swee; Yong, Ken-Tye; Yoon, Ho Sup

    2015-01-01

    RNA interference (RNAi)-based gene silencing possesses great ability for therapeutic intervention in pancreatic cancer. Among various oncogene mutations, Interleukin-8 (IL-8) gene mutations are found to be overexpressed in many pancreatic cell lines. In this work, we demonstrate IL-8 gene silencing by employing an RNAi-based gene therapy approach and this is achieved by using gold nanorods (AuNRs) for efficient delivery of IL-8 small interfering RNA (siRNA) to the pancreatic cell lines of MiaPaCa-2 and Panc-1. Upon comparing to Panc-1 cells, we found that the dominant expression of the IL-8 gene in MiaPaCa-2 cells resulted in an aggressive behavior towards the processes of cell invasion and metastasis. We have hence investigated the suitability of using AuNRs as novel non-viral nanocarriers for the efficient uptake and delivery of IL-8 siRNA in realizing gene knockdown of both MiaPaCa-2 and Panc-1 cells. Flow cytometry and fluorescence imaging techniques have been applied to confirm transfection and release of IL-8 siRNA. The ratio of AuNRs and siRNA has been optimized and transfection efficiencies as high as 88.40 ± 2.14% have been achieved. Upon successful delivery of IL-8 siRNA into cancer cells, the effects of IL-8 gene knockdown are quantified in terms of gene expression, cell invasion, cell migration and cell apoptosis assays. Statistical comparative studies for both MiaPaCa-2 and Panc-1 cells are presented in this work. IL-8 gene silencing has been demonstrated with knockdown efficiencies of 81.02 ± 10.14% and 75.73 ± 6.41% in MiaPaCa-2 and Panc-1 cells, respectively. Our results are then compared with a commercial transfection reagent, Oligofectamine, serving as positive control. The gene knockdown results illustrate the potential role of AuNRs as non-viral gene delivery vehicles for RNAi-based targeted cancer therapy applications. (paper)

  12. Gene therapy-mediated delivery of targeted cytotoxins for glioma therapeutics.

    Science.gov (United States)

    Candolfi, Marianela; Xiong, Weidong; Yagiz, Kader; Liu, Chunyan; Muhammad, A K M G; Puntel, Mariana; Foulad, David; Zadmehr, Ali; Ahlzadeh, Gabrielle E; Kroeger, Kurt M; Tesarfreund, Matthew; Lee, Sharon; Debinski, Waldemar; Sareen, Dhruv; Svendsen, Clive N; Rodriguez, Ron; Lowenstein, Pedro R; Castro, Maria G

    2010-11-16

    Restricting the cytotoxicity of anticancer agents by targeting receptors exclusively expressed on tumor cells is critical when treating infiltrative brain tumors such as glioblastoma multiforme (GBM). GBMs express an IL-13 receptor (IL13Rα2) that differs from the physiological IL4R/IL13R receptor. We developed a regulatable adenoviral vector (Ad.mhIL-4.TRE.mhIL-13-PE) encoding a mutated human IL-13 fused to Pseudomonas exotoxin (mhIL-13-PE) that specifically binds to IL13Rα2 to provide sustained expression, effective anti-GBM cytotoxicity, and minimal neurotoxicity. The therapeutic Ad also encodes mutated human IL-4 that binds to the physiological IL4R/IL13R without interacting with IL13Rα2, thus inhibiting potential binding of mhIL-13-PE to normal brain cells. Using intracranial GBM xenografts and syngeneic mouse models, we tested the Ad.mhIL-4.TRE.mhIL-13-PE and two protein formulations, hIL-13-PE used in clinical trials (Cintredekin Besudotox) and a second-generation mhIL-13-PE. Cintredekin Besudotox doubled median survival without eliciting long-term survival and caused severe neurotoxicity; mhIL-13-PE led to ∼40% long-term survival, eliciting severe neurological toxicity at the high dose tested. In contrast, Ad-mediated delivery of mhIL-13-PE led to tumor regression and long-term survival in over 70% of the animals, without causing apparent neurotoxicity. Although Cintredekin Besudotox was originally developed to target GBM, when tested in a phase III trial it failed to achieve clinical endpoints and revealed neurotoxicity. Limitations of Cintredekin Besudotox include its short half-life, which demanded frequent or continued administration, and binding to IL4R/IL13R, present in normal brain cells. These shortcomings were overcome by our therapeutic Ad, thus representing a significant advance in the development of targeted therapeutics for GBM.

  13. Synthetic sustained gene delivery systems.

    Science.gov (United States)

    Agarwal, Ankit; Mallapragada, Surya K

    2008-01-01

    Gene therapy today is hampered by the need of a safe and efficient gene delivery system that can provide a sustained therapeutic effect without cytotoxicity or unwanted immune responses. Bolus gene delivery in solution results in the loss of delivered factors via lymphatic system and may cause undesired effects by the escape of bioactive molecules to distant sites. Controlled gene delivery systems, acting as localized depot of genes, provide an extended sustained release of genes, giving prolonged maintenance of the therapeutic level of encoded proteins. They also limit the DNA degradation in the nuclease rich extra-cellular environment. While attempts have been made to adapt existing controlled drug delivery technologies, more novel approaches are being investigated for controlled gene delivery. DNA encapsulated in nano/micro spheres of polymers have been administered systemically/orally to be taken up by the targeted tissues and provide sustained release once internalized. Alternatively, DNA entrapped in hydrogels or scaffolds have been injected/implanted in tissues/cavities as platforms for gene delivery. The present review examines these different modalities for sustained delivery of viral and non-viral gene-delivery vectors. Design parameters and release mechanisms of different systems made with synthetic or natural polymers are presented along with their prospective applications and opportunities for continuous development.

  14. Therapeutic Efficacy of Vectored PGT121 Gene Delivery in HIV-1-Infected Humanized Mice.

    Science.gov (United States)

    Badamchi-Zadeh, Alexander; Tartaglia, Lawrence J; Abbink, Peter; Bricault, Christine A; Liu, Po-Ting; Boyd, Michael; Kirilova, Marinela; Mercado, Noe B; Nanayakkara, Ovini S; Vrbanac, Vladimir D; Tager, Andrew M; Larocca, Rafael A; Seaman, Michael S; Barouch, Dan H

    2018-04-01

    Broadly neutralizing antibodies (bNAbs) are being explored for HIV-1 prevention and cure strategies. However, administration of purified bNAbs poses challenges in resource-poor settings, where the HIV-1 disease burden is greatest. In vivo vector-based production of bNAbs represents an alternative strategy. We investigated adenovirus serotype 5 (Ad5) and adeno-associated virus serotype 1 (AAV1) vectors to deliver the HIV-1-specific bNAb PGT121 in wild-type and immunocompromised C57BL/6 mice as well as in HIV-1-infected bone marrow-liver-thymus (BLT) humanized mice. Ad5.PGT121 and AAV1.PGT121 produced functional antibody in vivo Ad5.PGT121 produced PGT121 rapidly within 6 h, whereas AAV1.PGT121 produced detectable PGT121 in serum by 72 h. Serum PGT121 levels were rapidly reduced by the generation of anti-PGT121 antibodies in immunocompetent mice but were durably maintained in immunocompromised mice. In HIV-1-infected BLT humanized mice, Ad5.PGT121 resulted in a greater reduction of viral loads than did AAV1.PGT121. Ad5.PGT121 also led to more-sustained virologic control than purified PGT121 IgG. Ad5.PGT121 afforded more rapid, robust, and durable antiviral efficacy than AAV1.PGT121 and purified PGT121 IgG in HIV-1-infected humanized mice. Further evaluation of vector delivery of HIV-1 bNAbs is warranted, although approaches to prevent the generation of antiantibody responses may also be required. IMPORTANCE Broadly neutralizing antibodies (bNAbs) are being explored for HIV-1 prevention and cure strategies, but delivery of purified antibodies may prove challenging. We investigated adenovirus serotype 5 (Ad5) and adeno-associated virus serotype 1 (AAV1) vectors to deliver the HIV-1-specific bNAb PGT121. Ad5.PGT121 afforded more rapid, robust, and durable antiviral efficacy than AAV1.PGT121 and purified PGT121 IgG in HIV-1-infected humanized mice. Copyright © 2018 Badamchi-Zadeh et al.

  15. Gene delivery of therapeutic polypeptides to brain capillary endothelial cells for protein secretion

    DEFF Research Database (Denmark)

    Larsen, Annette Burkhart; Thomsen, Louiza Bohn; Moos, Torben

    . Results: mRNA expression of proteins with neuroprotective potential in RBEC were enabled. Their expression patters were compared with those of RBE4 and HeLa cells using RT-qPCR analyzes. The evidence for protein synthesis and secretion was obtained by detection of FLAG-tagged to the C-terminal of any......Background: The potential for treatment of chronic disorders affecting the CNS is complicated by the inability of several drugs to cross the blood-brain barrier (BBB). None-viral gene therapy applied to brain capillary endothelial cells (BCECs) denotes a novel approach to overcome the restraints...... in this passage, as turning BCECs into recombinant protein factories by transfection could result in protein secretion into the brain. Aim: The aim of the present study was to investigate the possibility of transfection to primary rat brain capillary endothelial cells (RBEC) for recombinant protein synthesis...

  16. Protein nanoparticles for therapeutic protein delivery.

    Science.gov (United States)

    Herrera Estrada, L P; Champion, J A

    2015-06-01

    Therapeutic proteins can face substantial challenges to their activity, requiring protein modification or use of a delivery vehicle. Nanoparticles can significantly enhance delivery of encapsulated cargo, but traditional small molecule carriers have some limitations in their use for protein delivery. Nanoparticles made from protein have been proposed as alternative carriers and have benefits specific to therapeutic protein delivery. This review describes protein nanoparticles made by self-assembly, including protein cages, protein polymers, and charged or amphipathic peptides, and by desolvation. It presents particle fabrication and delivery characterization for a variety of therapeutic and model proteins, as well as comparison of the features of different protein nanoparticles.

  17. Protein based therapeutic delivery agents: Contemporary developments and challenges.

    Science.gov (United States)

    Yin, Liming; Yuvienco, Carlo; Montclare, Jin Kim

    2017-07-01

    As unique biopolymers, proteins can be employed for therapeutic delivery. They bear important features such as bioavailability, biocompatibility, and biodegradability with low toxicity serving as a platform for delivery of various small molecule therapeutics, gene therapies, protein biologics and cells. Depending on size and characteristic of the therapeutic, a variety of natural and engineered proteins or peptides have been developed. This, coupled to recent advances in synthetic and chemical biology, has led to the creation of tailor-made protein materials for delivery. This review highlights strategies employing proteins to facilitate the delivery of therapeutic matter, addressing the challenges for small molecule, gene, protein and cell transport. Copyright © 2017 Elsevier Ltd. All rights reserved.

  18. Cationic Bolaamphiphiles for Gene Delivery

    Science.gov (United States)

    Tan, Amelia Li Min; Lim, Alisa Xue Ling; Zhu, Yiting; Yang, Yi Yan; Khan, Majad

    2014-05-01

    Advances in medical research have shed light on the genetic cause of many human diseases. Gene therapy is a promising approach which can be used to deliver therapeutic genes to treat genetic diseases at its most fundamental level. In general, nonviral vectors are preferred due to reduced risk of immune response, but they are also commonly associated with low transfection efficiency and high cytotoxicity. In contrast to viral vectors, nonviral vectors do not have a natural mechanism to overcome extra- and intracellular barriers when delivering the therapeutic gene into cell. Hence, its design has been increasingly complex to meet challenges faced in targeting of, penetration of and expression in a specific host cell in achieving more satisfactory transfection efficiency. Flexibility in design of the vector is desirable, to enable a careful and controlled manipulation of its properties and functions. This can be met by the use of bolaamphiphile, a special class of lipid. Unlike conventional lipids, bolaamphiphiles can form asymmetric complexes with the therapeutic gene. The advantage of having an asymmetric complex lies in the different purposes served by the interior and exterior of the complex. More effective gene encapsulation within the interior of the complex can be achieved without triggering greater aggregation of serum proteins with the exterior, potentially overcoming one of the great hurdles faced by conventional single-head cationic lipids. In this review, we will look into the physiochemical considerations as well as the biological aspects of a bolaamphiphile-based gene delivery system.

  19. Gene therapy of cancer and development of therapeutic target gene

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Chang Min; Kwon, Hee Chung

    1998-04-01

    We applied HSV-tk/GCV strategy to orthotopic rat hepatoma model and showed anticancer effects of hepatoma. The increased expression of Lac Z gene after adenovirus-mediated gene delivery throughout hepatic artery was thought that is increased the possibility of gene therapy for curing hepatoma. With the construction of kGLP-laboratory, it is possible to produce a good quantity and quality of adenovirus in lage-scale production and purification of adenovirus vector. Also, the analysis of hepatoma related genes by PCR-LOH could be used for the diagnosis of patients and the development of therapeutic gene.

  20. Gene therapy of cancer and development of therapeutic target gene

    International Nuclear Information System (INIS)

    Kim, Chang Min; Kwon, Hee Chung

    1998-04-01

    We applied HSV-tk/GCV strategy to orthotopic rat hepatoma model and showed anticancer effects of hepatoma. The increased expression of Lac Z gene after adenovirus-mediated gene delivery throughout hepatic artery was thought that is increased the possibility of gene therapy for curing hepatoma. With the construction of kGLP-laboratory, it is possible to produce a good quantity and quality of adenovirus in lage-scale production and purification of adenovirus vector. Also, the analysis of hepatoma related genes by PCR-LOH could be used for the diagnosis of patients and the development of therapeutic gene

  1. Silk constructs for delivery of muskuloskeletal therapeutics

    Science.gov (United States)

    Meinel, Lorenz; Kaplan, David L.

    2012-01-01

    Silk fibroin (SF) is a biopolymer with distinguishing features from many other bio- as well as synthetic polymers. From a biomechanical and drug delivery perspective, SF combines remarkable versatility for scaffolding (solid implants, hydrogels, threads, solutions), with advanced mechanical properties and good stabilization and controlled delivery of entrapped protein and small molecule drugs, respectively. It is this combination of mechanical and pharmaceutical features which render SF so exciting for biomedical applications. his pattern along with the versatility of this biopolymer have been translated into progress for musculoskeletal applications. We review the use and potential of silk fibroin for systemic and localized delivery of therapeutics in diseases affecting the musculoskeletal system. We also present future directions for this biopolymer as well as the necessary research and development steps for their achievement. PMID:22522139

  2. Long-term delivery of protein therapeutics.

    Science.gov (United States)

    Vaishya, Ravi; Khurana, Varun; Patel, Sulabh; Mitra, Ashim K

    2015-03-01

    Proteins are effective biotherapeutics with applications in diverse ailments. Despite being specific and potent, their full clinical potential has not yet been realized. This can be attributed to short half-lives, complex structures, poor in vivo stability, low permeability, frequent parenteral administrations and poor adherence to treatment in chronic diseases. A sustained release system, providing controlled release of proteins, may overcome many of these limitations. This review focuses on recent development in approaches, especially polymer-based formulations, which can provide therapeutic levels of proteins over extended periods. Advances in particulate, gel-based formulations and novel approaches for extended protein delivery are discussed. Emphasis is placed on dosage form, method of preparation, mechanism of release and stability of biotherapeutics. Substantial advancements have been made in the field of extended protein delivery via various polymer-based formulations over last decade despite the unique delivery-related challenges posed by protein biologics. A number of injectable sustained-release formulations have reached market. However, therapeutic application of proteins is still hampered by delivery-related issues. A large number of protein molecules are under clinical trials, and hence, there is an urgent need to develop new methods to deliver these highly potent biologics.

  3. Structuring polymers for delivery of DNA-based therapeutics: updated insights.

    Science.gov (United States)

    Gupta, Madhu; Tiwari, Shailja; Vyas, Suresh

    2012-01-01

    Gene therapy offers greater opportunities for treating numerous incurable diseases from genetic disorders, infections, and cancer. However, development of appropriate delivery systems could be one of the most important factors to overcome numerous biological barriers for delivery of various therapeutic molecules. A number of nonviral polymer-mediated vectors have been developed for DNA delivery and offer the potential to surmount the associated problems of their viral counterpart. To address the concerns associated with safety issues, a wide range of polymeric vectors are available and have been utilized successfully to deliver their therapeutics in vivo. Today's research is mainly focused on the various natural or synthetic polymer-based delivery carriers that protect the DNA molecule from degradation, which offer specific targeting to the desired cells after systemic administration, have transfection efficiencies equivalent to virus-mediated gene delivery, and have long-term gene expression through sustained-release mechanisms. This review explores an updated overview of different nonviral polymeric delivery system for delivery of DNA-based therapeutics. These polymeric carriers have been evaluated in vitro and in vivo and are being utilized in various stages of clinical evaluation. Continued research and understanding of the principles of polymer-based gene delivery systems will enable us to develop new and efficient delivery systems for the delivery of DNA-based therapeutics to achieve the goal of efficacious and specific gene therapy for a vast array of clinical disorders as the therapeutic solutions of tomorrow.

  4. Real time sensor for therapeutic radiation delivery

    International Nuclear Information System (INIS)

    Bliss, M.; Craig, R.A.; Reeder, P.L.

    1998-01-01

    The invention is a real time sensor for therapeutic radiation. A probe is placed in or near the patient that senses in real time the dose at the location of the probe. The strength of the dose is determined by either an insertion or an exit probe. The location is determined by a series of vertical and horizontal sensing elements that gives the operator a real time read out dose location relative to placement of the patient. The increased accuracy prevents serious tissue damage to the patient by preventing overdose or delivery of a dose to a wrong location within the body. 14 figs

  5. Nonviral Delivery Systems For Cancer Gene Therapy: Strategies And Challenges.

    Science.gov (United States)

    Shim, Gayong; Kim, Dongyoon; Le, Quoc-Viet; Park, Gyu Thae; Kwon, Taekhyun; Oh, Yu-Kyoung

    2018-01-19

    Gene therapy has been receiving widespread attention due to its unique advantage in regulating the expression of specific target genes. In the field of cancer gene therapy, modulation of gene expression has been shown to decrease oncogenic factors in cancer cells or increase immune responses against cancer. Due to the macromolecular size and highly negative physicochemical features of plasmid DNA, efficient delivery systems are an essential ingredient for successful gene therapy. To date, a variety of nanostructures and materials have been studied as nonviral gene delivery systems. In this review, we will cover nonviral delivery strategies for cancer gene therapy, with a focus on target cancer genes and delivery materials. Moreover, we will address current challenges and perspectives for nonviral delivery-based cancer gene therapeutics. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  6. Liver cell-targeted delivery of therapeutic molecules.

    Science.gov (United States)

    Kang, Jeong-Hun; Toita, Riki; Murata, Masaharu

    2016-01-01

    The liver is the largest internal organ in mammals and is involved in metabolism, detoxification, synthesis of proteins and lipids, secretion of cytokines and growth factors and immune/inflammatory responses. Hepatitis, alcoholic or non-alcoholic liver disease, hepatocellular carcinoma, hepatic veno-occlusive disease, and liver fibrosis and cirrhosis are the most common liver diseases. Safe and efficient delivery of therapeutic molecules (drugs, genes or proteins) into the liver is very important to increase the clinical efficacy of these molecules and to reduce their side effects in other organs. Several liver cell-targeted delivery systems have been developed and tested in vivo or ex vivo/in vitro. In this review, we discuss the literature concerning liver cell-targeted delivery systems, with a particular emphasis on the results of in vivo studies.

  7. An industry update: the latest developments in Therapeutic delivery.

    Science.gov (United States)

    Steinbach, Oliver C

    2018-05-01

    The present industry update covers the period of 1 January-31 January 2018, with information sourced from company press releases, regulatory and patent agencies as well as scientific literature. Several public offerings (Gecko, Insmed), licensing (Foresee) and commercialization agreements (Alnylam, Collegium Pharmaceutical) as well as patent filings (Elute) continue to prove the sustained investments in the drug delivery market. In increasing numbers, more effective ways to deliver the active ingredient to the right location and the right dose through devices (Boehringer Ingelheim's Respimat, Medtronics' SynchroMedII) or improved compound properties through formulation (Aquestive Therapeutics' PharmFilm, Noven Pharmaceuticals' transdermal patch) are reaching the market. Furthering biologics and gene delivery (Avacta, Bracco) proves that novel drug delivery technologies are successfully addressing more challenging drug formats.

  8. Selenium nanoparticles: potential in cancer gene and drug delivery.

    Science.gov (United States)

    Maiyo, Fiona; Singh, Moganavelli

    2017-05-01

    In recent decades, colloidal selenium nanoparticles have emerged as exceptional selenium species with reported chemopreventative and therapeutic properties. This has sparked widespread interest in their use as a carrier of therapeutic agents with results displaying synergistic effects of selenium with its therapeutic cargo and improved anticancer activity. Functionalization remains a critical step in selenium nanoparticles' development for application in gene or drug delivery. In this review, we highlight recent developments in the synthesis and functionalization strategies of selenium nanoparticles used in cancer drug and gene delivery systems. We also provide an update of recent preclinical studies utilizing selenium nanoparticles in cancer therapeutics.

  9. Current issues of RNAi therapeutics delivery and development.

    Science.gov (United States)

    Haussecker, D

    2014-12-10

    12 years following the discovery of the RNAi mechanism in Man, a number of RNAi therapeutics development candidates have emerged with profiles suggesting that they could become drugs of significant medical importance for diseases like TTR amyloidosis, HBV, solid cancers, and hemophilia. Despite this robust progress, the perception of RNAi therapeutics has been on a roller-coaster ride driven not only by science, but also regulatory trends, the stock markets, and Big Pharma business development decisions [1]. This presentation provides an update on the current state of RNAi therapeutics development with a particular focus on what RNAi delivery can achieve today and key challenges to be overcome to expand therapeutic opportunities. The delivery of RNAi triggers to disease-relevant cell types clearly represents the rate-limiting factor in broadly expanding the applicability of RNAi therapeutics. Today, with at least 3 delivery options (lipid nanoparticles/LNPs, GalNAc-siRNA conjugates, Dynamic PolyConjugates/DPCs) for which profound gene knockdowns have been demonstrated in non-human primates and in the clinic, RNAi therapeutics should in principle be able to address most diseases related to gene expression in the liver. Given the central importance of the liver in systemic physiology, this already represents a significant therapeutic and commercial opportunity rivaling that of e.g. monoclonal antibodies. Beyond the liver, there is a reason to believe that current RNAi therapeutics technologies can address a number of solid tumors (e.g. LNPs), diseases of the eye (e.g. self-delivering RNAi triggers) as well as diseases involving the respiratory epithelium (e.g. aerosolized LNPs), certain phagocytic cells (LNPs), hematopoietic stem cells and their progeny (lentiviral DNA-directed RNAi), vascular endothelial cells (cationic lipoplexes), and certain cell types in the kidney (self-delivering RNAi triggers, DPCs; Table 1). Despite this success, there has been a sense that

  10. Gene doping: gene delivery for olympic victory.

    Science.gov (United States)

    Gould, David

    2013-08-01

    With one recently recommended gene therapy in Europe and a number of other gene therapy treatments now proving effective in clinical trials it is feasible that the same technologies will soon be adopted in the world of sport by unscrupulous athletes and their trainers in so called 'gene doping'. In this article an overview of the successful gene therapy clinical trials is provided and the potential targets for gene doping are highlighted. Depending on whether a doping gene product is secreted from the engineered cells or is retained locally to, or inside engineered cells will, to some extent, determine the likelihood of detection. It is clear that effective gene delivery technologies now exist and it is important that detection and prevention plans are in place. © 2012 The Author. British Journal of Clinical Pharmacology © 2012 The British Pharmacological Society.

  11. Targeted Delivery of siRNA Therapeutics to Malignant Tumors

    Directory of Open Access Journals (Sweden)

    Qixin Leng

    2017-01-01

    Full Text Available Over the past 20 years, a diverse group of ligands targeting surface biomarkers or receptors has been identified with several investigated to target siRNA to tumors. Many approaches to developing tumor-homing peptides, RNA and DNA aptamers, and single-chain variable fragment antibodies by using phage display, in vitro evolution, and recombinant antibody methods could not have been imagined by researchers in the 1980s. Despite these many scientific advances, there is no reason to expect that the ligand field will not continue to evolve. From development of ligands based on novel or existing biomarkers to linking ligands to drugs and gene and antisense delivery systems, several fields have coalesced to facilitate ligand-directed siRNA therapeutics. In this review, we discuss the major categories of ligand-targeted siRNA therapeutics for tumors, as well as the different strategies to identify new ligands.

  12. Complexity in the therapeutic delivery of RNAi medicines

    DEFF Research Database (Denmark)

    Colombo, Stefano; Zeng, Xianghui; Ragelle, Héloïse

    2014-01-01

    of this review is to reflect on the complexity in the therapeutic delivery of RNA interference-based drugs emerging from the recent clinical experiences and report the actual technological and analytical advances introduced to solve it. EXPERT OPINION: The complexity in the therapeutic delivery of nucleic acids...

  13. Liposomes for Use in Gene Delivery

    Directory of Open Access Journals (Sweden)

    Daniel A. Balazs

    2011-01-01

    Full Text Available Liposomes have a wide array of uses that have been continuously expanded and improved upon since first being observed to self-assemble into vesicular structures. These arrangements can be found in many shapes and sizes depending on lipid composition. Liposomes are often used to deliver a molecular cargo such as DNA for therapeutic benefit. The lipids used to form such lipoplexes can be cationic, anionic, neutral, or a mixture thereof. Herein physical packing parameters and specific lipids used for gene delivery will be discussed, with lipids classified according to overall charge.

  14. An intestinal Trojan horse for gene delivery.

    Science.gov (United States)

    Peng, Haisheng; Wang, Chao; Xu, Xiaoyang; Yu, Chenxu; Wang, Qun

    2015-03-14

    The intestinal epithelium forms an essential element of the mucosal barrier and plays a critical role in the pathophysiological response to different enteric disorders and diseases. As a major enteric dysfunction of the intestinal tract, inflammatory bowel disease is a genetic disease which results from the inappropriate and exaggerated mucosal immune response to the normal constituents in the mucosal microbiota environment. An intestine targeted drug delivery system has unique advantages in the treatment of inflammatory bowel disease. As a new concept in drug delivery, the Trojan horse system with the synergy of nanotechnology and host cells can achieve better therapeutic efficacy in specific diseases. Here, we demonstrated the feasibility of encapsulating DNA-functionalized gold nanoparticles into primary isolated intestinal stem cells to form an intestinal Trojan horse for gene regulation therapy of inflammatory bowel disease. This proof-of-concept intestinal Trojan horse will have a wide variety of applications in the diagnosis and therapy of enteric disorders and diseases.

  15. Advances in RNAi therapeutic delivery to leukocytes using lipid nanoparticles.

    Science.gov (United States)

    Ramishetti, Srinivas; Landesman-Milo, Dalit; Peer, Dan

    2016-11-01

    Small interfering RNAs (siRNAs) therapeutics has advanced into clinical trials for liver diseases and solid tumors, but remain a challenge for manipulating leukocytes fate due to lack of specificity and safety issues. Leukocytes ingest pathogens and defend the body through a complex network. They are also involved in the pathogeneses of inflammation, viral infection, autoimmunity and cancers. Modulating gene expression in leukocytes using siRNAs holds great promise to treat leukocyte-mediated diseases. Leukocytes are notoriously hard to transduce with siRNAs and are spread throughout the body often located deep in tissues, therefore developing an efficient systemic delivery strategy is still a challenge. Here, we discuss recent advances in siRNA delivery to leukocyte subsets such as macrophages, monocytes, dendritic cells and lymphocytes. We focus mainly on lipid-based nanoparticles (LNPs) comprised of new generation of ionizable lipids and their ability to deliver siRNA to primary or malignant leukocytes in a targeted manner. Special emphasis is made on LNPs targeted to subsets of leukocytes and we detail a novel microfluidic mixing technology that could aid in changing the landscape of process development of LNPs from a lab tool to a potential novel therapeutic modality.

  16. How controlled release technology can aid gene delivery.

    Science.gov (United States)

    Jo, Jun-Ichiro; Tabata, Yasuhiko

    2015-01-01

    Many types of gene delivery systems have been developed to enhance the level of gene expression. Controlled release technology is a feasible gene delivery system which enables genes to extend the expression duration by maintaining and releasing them at the injection site in a controlled manner. This technology can reduce the adverse effects by the bolus dose administration and avoid the repeated administration. Biodegradable biomaterials are useful as materials for the controlled release-based gene delivery technology and various biodegradable biomaterials have been developed. Controlled release-based gene delivery plays a critical role in a conventional gene therapy and genetic engineering. In the gene therapy, the therapeutic gene is released from biodegradable biomaterial matrices around the tissue to be treated. On the other hand, the intracellular controlled release of gene from the sub-micro-sized matrices is required for genetic engineering. Genetic engineering is feasible for cell transplantation as well as research of stem cells biology and medicine. DNA hydrogel containing a sequence of therapeutic gene and the exosome including the individual specific nucleic acids may become candidates for controlled release carriers. Technologies to deliver genes to cell aggregates will play an important role in the promotion of regenerative research and therapy.

  17. A new electrospray method for targeted gene delivery.

    Science.gov (United States)

    Boehringer, Stephan; Ruzgys, Paulius; Tamò, Luca; Šatkauskas, Saulius; Geiser, Thomas; Gazdhar, Amiq; Hradetzky, David

    2018-03-05

    A challenge for gene therapy is absence of safe and efficient local delivery of therapeutic genetic material. An efficient and reproducible physical method of electrospray for localized and targeted gene delivery is presented. Electrospray works on the principle of coulombs repulsion, under influence of electric field the liquid carrying genetic material is dispersed into micro droplets and is accelerated towards the targeted tissue, acting as a counter electrode. The accelerated droplets penetrate the targeted cells thus facilitating the transfer of genetic material into the cell. The work described here presents the principle of electrospray for gene delivery, the basic instrument design, and the various optimized parameters to enhance gene transfer in vitro. We estimate a transfection efficiency of up to 60% was achieved. We describe an efficient gene transfer method and a potential electrospray-mediated gene transfer mechanism.

  18. Dendrimer Advances for the Central Nervous System Delivery of Therapeutics

    Science.gov (United States)

    2013-01-01

    The effectiveness of noninvasive treatment for central nervous system (CNS) diseases is generally limited by the poor access of therapeutic agents into the CNS. Most CNS drugs cannot permeate into the brain parenchyma because of the blood-brain barrier (BBB), and overcoming this has become one of the most significant challenges in the development of CNS therapeutics. Rapid advances in nanotechnology have provided promising solutions to this challenge. This review discusses the latest applications of dendrimers in the treatment of CNS diseases with an emphasis on brain tumors. Dendrimer-mediated drug delivery, imaging, and diagnosis are also reviewed. The toxicity, biodistribution, and transport mechanisms in dendrimer-mediated delivery of CNS therapeutic agents bypassing or crossing the BBB are also discussed. Future directions and major challenges of dendrimer-mediated delivery of CNS therapeutic agents are included. PMID:24274162

  19. Dendrimer advances for the central nervous system delivery of therapeutics.

    Science.gov (United States)

    Xu, Leyuan; Zhang, Hao; Wu, Yue

    2014-01-15

    The effectiveness of noninvasive treatment for central nervous system (CNS) diseases is generally limited by the poor access of therapeutic agents into the CNS. Most CNS drugs cannot permeate into the brain parenchyma because of the blood-brain barrier (BBB), and overcoming this has become one of the most significant challenges in the development of CNS therapeutics. Rapid advances in nanotechnology have provided promising solutions to this challenge. This review discusses the latest applications of dendrimers in the treatment of CNS diseases with an emphasis on brain tumors. Dendrimer-mediated drug delivery, imaging, and diagnosis are also reviewed. The toxicity, biodistribution, and transport mechanisms in dendrimer-mediated delivery of CNS therapeutic agents bypassing or crossing the BBB are also discussed. Future directions and major challenges of dendrimer-mediated delivery of CNS therapeutic agents are included.

  20. Lipoprotein Nanoplatform for Targeted Delivery of Diagnostic and Therapeutic Agents

    Directory of Open Access Journals (Sweden)

    Jerry D. Glickson

    2008-03-01

    Full Text Available Low-density lipoprotein (LDL provides a highly versatile natural nanoplatform for delivery of visible or near-infrared fluorescent optical and magnetic resonance imaging (MRI contrast agents and photodynamic therapy and chemotherapeutic agents to normal and neoplastic cells that overexpress low-density lipoprotein receptors (LDLRs. Extension to other lipoproteins ranging in diameter from about 10 nm (high-density lipoprotein [HDL] to over a micron (chylomicrons is feasible. Loading of contrast or therapeutic agents onto or into these particles has been achieved by protein loading (covalent attachment to protein side chains, surface loading (intercalation into the phospholipid monolayer, and core loading (extraction and reconstitution of the triglyceride/cholesterol ester core. Core and surface loading of LDL have been used for delivery of optical imaging agents to tumor cells in vivo and in culture. Surface loading was used for delivery of gadolinium-bis-stearylamide contrast agents for in vivo MRI detection in tumor-bearing mice. Chlorin and phthalocyanine near-infrared photodynamic therapy agents (≤ 400/LDL have been attached by core loading. Protein loading was used to reroute the LDL from its natural receptor (LDLR to folate receptors and could be used to target other receptors. A semisynthetic nanoparticle has been constructed by coating magnetite iron oxide nanoparticles with carboxylated cholesterol and overlaying a monolayer of phospholipid to which apolipoprotein A1 or E was adsorbed for targeting HDL or adsorbing synthetic amphipathic helical peptides ltargeting LDL or folate receptors. These particles can be used for in situ loading of magnetite into cells for MRI-monitored cell tracking or gene expression.

  1. Stimuli-responsive nanomaterials for therapeutic protein delivery.

    Science.gov (United States)

    Lu, Yue; Sun, Wujin; Gu, Zhen

    2014-11-28

    Protein therapeutics have emerged as a significant role in treatment of a broad spectrum of diseases, including cancer, metabolic disorders and autoimmune diseases. The efficacy of protein therapeutics, however, is limited by their instability, immunogenicity and short half-life. In order to overcome these barriers, tremendous efforts have recently been made in developing controlled protein delivery systems. Stimuli-triggered release is an appealing and promising approach for protein delivery and has made protein delivery with both spatiotemporal- and dosage-controlled manners possible. This review surveys recent advances in controlled protein delivery of proteins or peptides using stimuli-responsive nanomaterials. Strategies utilizing both physiological and external stimuli are introduced and discussed. Copyright © 2014 Elsevier B.V. All rights reserved.

  2. Au nanoinjectors for electrotriggered gene delivery into the cell nucleus.

    Science.gov (United States)

    Kang, Mijeong; Kim, Bongsoo

    2015-01-01

    Intracellular delivery of exogenous materials is an essential technique required for many fundamental biological researches and medical treatments. As our understanding of cell structure and function has been improved and diverse therapeutic agents with a subcellular site of action have been continuously developed, there is a demand to enhance the performance of delivering devices. Ideal intracellular delivery devices should convey various kinds of exogenous materials without deteriorating cell viability regardless of cell type and, furthermore, precisely control the location and the timing of delivery as well as the amount of delivered materials for advanced researches.In this chapter the development of a new intracellular delivery device, a nanoinjector made of a Au (gold) nanowire (a Au nanoinjector) is described in which delivery is triggered by external application of an electric pulse. As a model study, a gene was delivered directly into the nucleus of a neuroblastoma cell, and successful delivery without cell damage was confirmed by the expression of the delivered gene. The insertion of a Au nanoinjector directly into a cell can be generally applied to any kind of cell, and a high degree of surface modification of Au allows attachment of diverse materials such as proteins, small molecules, or nanoparticles as well as genes on Au nanoinjectors. This expands their applicability, and it is expected that they will provide important information on the effects of delivered exogenous materials and consequently contribute to the development of related therapeutic or clinical technologies.

  3. Star-Shaped Polypeptides: Synthesis and Opportunities for Delivery of Therapeutics.

    Science.gov (United States)

    Byrne, Mark; Murphy, Robert; Kapetanakis, Antonios; Ramsey, Joanne; Cryan, Sally-Ann; Heise, Andreas

    2015-09-17

    Significant advances in the synthesis of polypeptides by N-carboxyanhydride (NCA) polymerisation over the last decade have enabled the design of advanced polypeptide architectures such as star-shaped polypeptides. These materials combine the functionality offered by amino acids with the flexibility of creating stable nanoparticles with adjustable cargo space for therapeutic delivery. This review highlights recent advances in the synthesis of star polypeptides by NCA polymerisation followed by a critical review of the applications of this class of polymer in the delivery of therapeutic agents. This includes examples of traditional small-molecule drugs as well as the emerging class of biologics such as genetic therapeutics (gene delivery). © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Gene doping: gene delivery for olympic victory

    OpenAIRE

    Gould, David

    2012-01-01

    With one recently recommended gene therapy in Europe and a number of other gene therapy treatments now proving effective in clinical trials it is feasible that the same technologies will soon be adopted in the world of sport by unscrupulous athletes and their trainers in so called ‘gene doping’. In this article an overview of the successful gene therapy clinical trials is provided and the potential targets for gene doping are highlighted. Depending on whether a doping gene product is secreted...

  5. Noninvasive ocular drug delivery: potential transcorneal and other alternative delivery routes for therapeutic molecules in glaucoma.

    Science.gov (United States)

    Foldvari, Marianna

    2014-01-01

    Drug delivery to the eye is made difficult by multiple barriers (such as the tear film, cornea, and vitreous) between the surface of the eye and the treatment site. These barriers are difficult to surmount for the purposes of drug delivery without causing toxicity. Using nanotechnology tools to control, manipulate, and study delivery systems, new approaches to delivering drugs, genes, and antigens that are effective and safe can be developed. Topical administration to the ocular surface would be the safest method for delivery, as it is noninvasive and painless compared with other delivery methods. However, there is only limited success using topical delivery methods, especially for gene therapy. Current thinking on treatments of the future enabled by nanodelivery systems and the identification of target specificity parameters that require deeper understanding to develop successful topical delivery systems for glaucoma is highlighted.

  6. Chitosan for gene delivery and orthopedic tissue engineering applications.

    Science.gov (United States)

    Raftery, Rosanne; O'Brien, Fergal J; Cryan, Sally-Ann

    2013-05-15

    Gene therapy involves the introduction of foreign genetic material into cells in order exert a therapeutic effect. The application of gene therapy to the field of orthopaedic tissue engineering is extremely promising as the controlled release of therapeutic proteins such as bone morphogenetic proteins have been shown to stimulate bone repair. However, there are a number of drawbacks associated with viral and synthetic non-viral gene delivery approaches. One natural polymer which has generated interest as a gene delivery vector is chitosan. Chitosan is biodegradable, biocompatible and non-toxic. Much of the appeal of chitosan is due to the presence of primary amine groups in its repeating units which become protonated in acidic conditions. This property makes it a promising candidate for non-viral gene delivery. Chitosan-based vectors have been shown to transfect a number of cell types including human embryonic kidney cells (HEK293) and human cervical cancer cells (HeLa). Aside from its use in gene delivery, chitosan possesses a range of properties that show promise in tissue engineering applications; it is biodegradable, biocompatible, has anti-bacterial activity, and, its cationic nature allows for electrostatic interaction with glycosaminoglycans and other proteoglycans. It can be used to make nano- and microparticles, sponges, gels, membranes and porous scaffolds. Chitosan has also been shown to enhance mineral deposition during osteogenic differentiation of MSCs in vitro. The purpose of this review is to critically discuss the use of chitosan as a gene delivery vector with emphasis on its application in orthopedic tissue engineering.

  7. Advances in the delivery of RNA therapeutics: from concept to clinical reality.

    Science.gov (United States)

    Kaczmarek, James C; Kowalski, Piotr S; Anderson, Daniel G

    2017-06-27

    The rapid expansion of the available genomic data continues to greatly impact biomedical science and medicine. Fulfilling the clinical potential of genetic discoveries requires the development of therapeutics that can specifically modulate the expression of disease-relevant genes. RNA-based drugs, including short interfering RNAs and antisense oligonucleotides, are particularly promising examples of this newer class of biologics. For over two decades, researchers have been trying to overcome major challenges for utilizing such RNAs in a therapeutic context, including intracellular delivery, stability, and immune response activation. This research is finally beginning to bear fruit as the first RNA drugs gain FDA approval and more advance to the final phases of clinical trials. Furthermore, the recent advent of CRISPR, an RNA-guided gene-editing technology, as well as new strides in the delivery of messenger RNA transcribed in vitro, have triggered a major expansion of the RNA-therapeutics field. In this review, we discuss the challenges for clinical translation of RNA-based therapeutics, with an emphasis on recent advances in delivery technologies, and present an overview of the applications of RNA-based drugs for modulation of gene/protein expression and genome editing that are currently being investigated both in the laboratory as well as in the clinic.

  8. Enabling Anticancer Therapeutics by Nanoparticle Carriers: The Delivery of Paclitaxel

    Directory of Open Access Journals (Sweden)

    Bing Yan

    2011-07-01

    Full Text Available Anticancer drugs, such as paclitaxel (PTX, are indispensable for the treatment of a variety of malignancies. However, the application of most drugs is greatly limited by the low water solubility, poor permeability, or high efflux from cells. Nanoparticles have been widely investigated to enable drug delivery due to their low toxicity, sustained drug release, molecular targeting, and additional therapeutic and imaging functions. This review takes paclitaxel as an example and compares different nanoparticle-based delivery systems for their effectiveness in cancer chemotherapy.

  9. Engineered modular biomaterial logic gates for environmentally triggered therapeutic delivery

    Science.gov (United States)

    Badeau, Barry A.; Comerford, Michael P.; Arakawa, Christopher K.; Shadish, Jared A.; Deforest, Cole A.

    2018-03-01

    The successful transport of drug- and cell-based therapeutics to diseased sites represents a major barrier in the development of clinical therapies. Targeted delivery can be mediated through degradable biomaterial vehicles that utilize disease biomarkers to trigger payload release. Here, we report a modular chemical framework for imparting hydrogels with precise degradative responsiveness by using multiple environmental cues to trigger reactions that operate user-programmable Boolean logic. By specifying the molecular architecture and connectivity of orthogonal stimuli-labile moieties within material cross-linkers, we show selective control over gel dissolution and therapeutic delivery. To illustrate the versatility of this methodology, we synthesized 17 distinct stimuli-responsive materials that collectively yielded all possible YES/OR/AND logic outputs from input combinations involving enzyme, reductant and light. Using these hydrogels we demonstrate the first sequential and environmentally stimulated release of multiple cell lines in well-defined combinations from a material. We expect these platforms will find utility in several diverse fields including drug delivery, diagnostics and regenerative medicine.

  10. Biominetic High Density Lipoproteins for the Delivery of Therapeutic Oligonucleotides

    Science.gov (United States)

    Tripathy, Sushant

    Advances in nanotechnology have brought about novel inorganic and hybrid nanoparticles with unique physico-chemical properties that make them suitable for a broad range of applications---from nano-circuitry to drug delivery. A significant part of those advancements have led to ground-breaking discoveries that have changed the approaches to formulation of therapeutics against diseases, such as cancer. Now-a-days the focus does not lie solely on finding a candidate small-molecule therapeutic with minimal adverse effects, but researchers are looking up to nanoparticles to improve biodistribution and biocompatibility profile of clinically proven therapeutics. The plethora of conjugation chemistries offered by currently extant inorganic nanoparticles have, in recent years, led to great leaps in the field of biomimicry---a modality that promises high biocompatibility. Further, in the pursuit of highly specific therapeutic molecules, researchers have turned to silencing oligonucleotides and some have already brought together the strengths of nanoparticles and silencing oligonucleotides in search of an efficacious therapy for cancer with minimal adverse effects. This dissertation work focuses on such a biomimetic platform---a gold nanoparticle based high density lipoprotein biomimetic (HDL NP), for the delivery of therapeutic oligonucleotides. The first chapter of this body of work introduces the molecular target of the silencing oligonucleotides---VEGFR2, and its role in the progression of solid tumor cancers. The background information also covers important aspects of natural high density lipoproteins (HDL), especially their innate capacity to bind and deliver exogenous and endogenous silencing oligonucleotides to tissues that express their high affinity receptor SRB1. We subsequently describe the synthesis of the biomimetic HDL NP and its oligonucleotide conjugates, and establish their biocompatibility. Further on, experimental data demonstrate the efficacy of silencing

  11. Ultrasound-Mediated Local Drug and Gene Delivery Using Nanocarriers

    Science.gov (United States)

    Zhou, Qiu-Lan; Chen, Zhi-Yi; Yang, Feng

    2014-01-01

    With the development of nanotechnology, nanocarriers have been increasingly used for curative drug/gene delivery. Various nanocarriers are being introduced and assessed, such as polymer nanoparticles, liposomes, and micelles. As a novel theranostic system, nanocarriers hold great promise for ultrasound molecular imaging, targeted drug/gene delivery, and therapy. Nanocarriers, with the properties of smaller particle size, and long circulation time, would be advantageous in diagnostic and therapeutic applications. Nanocarriers can pass through blood capillary walls and cell membrane walls to deliver drugs. The mechanisms of interaction between ultrasound and nanocarriers are not clearly understood, which may be related to cavitation, mechanical effects, thermal effects, and so forth. These effects may induce transient membrane permeabilization (sonoporation) on a single cell level, cell death, and disruption of tissue structure, ensuring noninvasive, targeted, and efficient drug/gene delivery and therapy. The system has been used in various tissues and organs (in vitro or in vivo), including tumor tissues, kidney, cardiac, skeletal muscle, and vascular smooth muscle. In this review, we explore the research progress and application of ultrasound-mediated local drug/gene delivery with nanocarriers. PMID:25202710

  12. Ultrasound-Mediated Local Drug and Gene Delivery Using Nanocarriers

    Directory of Open Access Journals (Sweden)

    Qiu-Lan Zhou

    2014-01-01

    Full Text Available With the development of nanotechnology, nanocarriers have been increasingly used for curative drug/gene delivery. Various nanocarriers are being introduced and assessed, such as polymer nanoparticles, liposomes, and micelles. As a novel theranostic system, nanocarriers hold great promise for ultrasound molecular imaging, targeted drug/gene delivery, and therapy. Nanocarriers, with the properties of smaller particle size, and long circulation time, would be advantageous in diagnostic and therapeutic applications. Nanocarriers can pass through blood capillary walls and cell membrane walls to deliver drugs. The mechanisms of interaction between ultrasound and nanocarriers are not clearly understood, which may be related to cavitation, mechanical effects, thermal effects, and so forth. These effects may induce transient membrane permeabilization (sonoporation on a single cell level, cell death, and disruption of tissue structure, ensuring noninvasive, targeted, and efficient drug/gene delivery and therapy. The system has been used in various tissues and organs (in vitro or in vivo, including tumor tissues, kidney, cardiac, skeletal muscle, and vascular smooth muscle. In this review, we explore the research progress and application of ultrasound-mediated local drug/gene delivery with nanocarriers.

  13. Recent Trends of Polymer Mediated Liposomal Gene Delivery System

    Directory of Open Access Journals (Sweden)

    Shyamal Kumar Kundu

    2014-01-01

    Full Text Available Advancement in the gene delivery system have resulted in clinical successes in gene therapy for patients with several genetic diseases, such as immunodeficiency diseases, X-linked adrenoleukodystrophy (X-ALD blindness, thalassemia, and many more. Among various delivery systems, liposomal mediated gene delivery route is offering great promises for gene therapy. This review is an attempt to depict a portrait about the polymer based liposomal gene delivery systems and their future applications. Herein, we have discussed in detail the characteristics of liposome, importance of polymer for liposome formulation, gene delivery, and future direction of liposome based gene delivery as a whole.

  14. An industry update: the latest news in therapeutic delivery.

    Science.gov (United States)

    Harris, Elaine

    2017-11-01

    The present industry update covers the period 1-31 July 2017. Information was sourced primarily from company press releases, regulatory and patent agencies, scientific literature and various news websites. There was positive approval news this month for GlaxoSmithKline for its new self-injecting treatment for systemic lupus erythematosus but less positive news for Ocular Therapeutix, a new drug application for its treatment for postoperative ocular pain, DEXTENZA™ was rejected for a second time. Endo Pharmaceuticals agreed to withdraw its opioid formulation Opana ® Er due to abuse concerns. Collaborations for novel therapeutic delivery research were announced this month by Takeda and BioSurfaces, Catelent and Rutgers University, Lilly and Purdue University and Titan Pharmaceuticals with Walter Reed and the Southwest Research Institute. A number of companies announced significant financing deals to allow for the clinical development of products with enhanced delivery options including Sebacia and Diasome.

  15. Design and Fabrication of N-Alkyl-Polyethylenimine-Stabilized Iron Oxide Nanoclusters for Gene Delivery

    OpenAIRE

    Liu, Gang; Wang, Zhiyong; Lee, Seulki; Ai, Hua; Chen, Xiaoyuan

    2012-01-01

    With the rapid development of nanotechnology, inorganic magnetic nanoparticles, especially iron oxide nanoparticles (IOs), have emerged as great vehicles for biomedical diagnostic and therapeutic applications. In order to rationally design IO-based gene delivery nanovectors, surface modification is essential and determines the loading and release of the gene of interest. Here we highlight the basic concepts and applications of nonviral gene delivery vehicles based on low molecular weight N-al...

  16. Modified montmorillonite as vector for gene delivery.

    Science.gov (United States)

    Lin, Feng-Huei; Chen, Chia-Hao; Cheng, Winston T K; Kuo, Tzang-Fu

    2006-06-01

    Currently, gene delivery systems can be divided into two parts: viral or non-viral vectors. In general, viral vectors have a higher efficiency on gene delivery. However, they may sometimes provoke mutagenesis and carcinogenesis once re-activating in human body. Lots of non-viral vectors have been developed that tried to solve the problems happened on viral vectors. Unfortunately, most of non-viral vectors showed relatively lower transfection rate. The aim of this study is to develop a non-viral vector for gene delivery system. Montmorillonite (MMT) is one of clay minerals that consist of hydrated aluminum with Si-O tetrahedrons on the bottom of the layer and Al-O(OH)2 octahedrons on the top. The inter-layer space is about 12 A. The room is not enough to accommodate DNA for gene delivery. In the study, the cationic hexadecyltrimethylammonium (HDTMA) will be intercalated into the interlayer of MMT as a layer expander to expand the layer space for DNA accommodation. The optimal condition for the preparation of DNA-HDTMA-MMT is as follows: 1 mg of 1.5CEC HDTMA-MMT was prepared under pH value of 10.7 and with soaking time for 2 h. The DNA molecules can be protected from nuclease degradation, which can be proven by the electrophoresis analysis. DNA was successfully transfected into the nucleus of human dermal fibroblast and expressed enhanced green fluorescent protein (EGFP) gene with green fluorescence emission. The HDTMA-MMT has a great potential as a vector for gene delivery in the future.

  17. Carbon Nanotubes in Drug and Gene Delivery

    Science.gov (United States)

    Karimi, Mahdi; Ghasemi, Amir; Mirkiani, Soroush; Moosavi Basri, Seyed Masoud; Hamblin, Michael R.

    2017-10-01

    Recent important discoveries and developments in nanotechnology have had a remarkable and ever-increasing impact on many industries, especially materials science, pharmaceuticals, and biotechnology. Within this book, the authors describe different features of carbon nanotubes, survey the properties of both the multi-walled and single-walled varieties, and cover their applications in drug and gene delivery.

  18. Therapeutic genes for anti-HIV/AIDS gene therapy.

    Science.gov (United States)

    Bovolenta, Chiara; Porcellini, Simona; Alberici, Luca

    2013-01-01

    The multiple therapeutic approaches developed so far to cope HIV-1 infection, such as anti-retroviral drugs, germicides and several attempts of therapeutic vaccination have provided significant amelioration in terms of life-quality and survival rate of AIDS patients. Nevertheless, no approach has demonstrated efficacy in eradicating this lethal, if untreated, infection. The curative power of gene therapy has been proven for the treatment of monogenic immunodeficiensies, where permanent gene modification of host cells is sufficient to correct the defect for life-time. No doubt, a similar concept is not applicable for gene therapy of infectious immunodeficiensies as AIDS, where there is not a single gene to be corrected; rather engineered cells must gain immunotherapeutic or antiviral features to grant either short- or long-term efficacy mostly by acquisition of antiviral genes or payloads. Anti-HIV/AIDS gene therapy is one of the most promising strategy, although challenging, to eradicate HIV-1 infection. In fact, genetic modification of hematopoietic stem cells with one or multiple therapeutic genes is expected to originate blood cell progenies resistant to viral infection and thereby able to prevail on infected unprotected cells. Ultimately, protected cells will re-establish a functional immune system able to control HIV-1 replication. More than hundred gene therapy clinical trials against AIDS employing different viral vectors and transgenes have been approved or are currently ongoing worldwide. This review will overview anti-HIV-1 infection gene therapy field evaluating strength and weakness of the transgenes and payloads used in the past and of those potentially exploitable in the future.

  19. The evolution of heart gene delivery vectors

    Science.gov (United States)

    Wasala, Nalinda B.; Shin, Jin-Hong; Duan, Dongsheng

    2012-01-01

    Gene therapy holds promise for treating numerous heart diseases. A key premise for the success of cardiac gene therapy is the development of powerful gene transfer vehicles that can achieve highly efficient and persistent gene transfer specifically in the heart. Other features of an ideal vector include negligible toxicity, minimal immunogenicity and easy manufacturing. Rapid progress in the fields of molecular biology and virology has offered great opportunities to engineer various genetic materials for heart gene delivery. Several nonviral vectors (e.g. naked plasmids, plasmid lipid/polymer complexes and oligonucleotides) have been tested. Commonly used viral vectors include lentivirus, adenovirus and adeno-associated virus. Among these, adeno-associated virus has shown many attractive features for pre-clinical experimentation in animal models of heart diseases. We review the history and evolution of these vectors for heart gene transfer. PMID:21837689

  20. Receptor-Mediated Endocytosis and Brain Delivery of Therapeutic Biologics

    Directory of Open Access Journals (Sweden)

    Guangqing Xiao

    2013-01-01

    Full Text Available Transport of macromolecules across the blood-brain-barrier (BBB requires both specific and nonspecific interactions between macromolecules and proteins/receptors expressed on the luminal and/or the abluminal surfaces of the brain capillary endothelial cells. Endocytosis and transcytosis play important roles in the distribution of macromolecules. Due to the tight junction of BBB, brain delivery of traditional therapeutic proteins with large molecular weight is generally not possible. There are multiple pathways through which macromolecules can be taken up into cells through both specific and nonspecific interactions with proteins/receptors on the cell surface. This review is focused on the current knowledge of receptor-mediated endocytosis/transcytosis and brain delivery using the Angiopep-2-conjugated system and the molecular Trojan horses. In addition, the role of neonatal Fc receptor (FcRn in regulating the efflux of Immunoglobulin G (IgG from brain to blood, and approaches to improve the pharmacokinetics of therapeutic biologics by generating Fc fusion proteins, and increasing the pH dependent binding affinity between Fc and FcRn, are discussed.

  1. Polymeric Gene Delivery for Diabetic Treatment

    Directory of Open Access Journals (Sweden)

    Sung Wan Kim

    2011-08-01

    Full Text Available Several polymers were used to delivery genes to diabetic animals. Polyaminobutyl glycolic acid was utilized to deliver IL-10 plasmid DNA to prevent autoimmune insulitis of non-obese diabetic (NOD mouse. Polyethylene glycol grafted polylysine was combined with antisense glutamic acid decarboxylase (GAD MRNA to represent GAD autoantigene expression. GLP1 and TSTA (SP-EX4 were delivered by bioreducible polymer to stop diabetic progression. Fas siRNA delivery was carried out to treat diabetic NOD mice animal.

  2. Enhancement of therapeutic drug and DNA delivery into cells by electroporation

    Energy Technology Data Exchange (ETDEWEB)

    Rabussay, Dietmar [Genetronics, Inc., Department of Research and Development, 11199 Sorrento Valley Road, San Diego, CA (United States); Dev, Nagendu B [Genetronics, Inc., Department of Research and Development, 11199 Sorrento Valley Road, San Diego, CA (United States); Fewell, Jason [Valentis, Inc., 8301 New Trails Drive, The Woodlands, TX (United States); Smith, Louis C [Valentis, Inc., 8301 New Trails Drive, The Woodlands, TX (United States); Widera, Georg [Genetronics, Inc., Department of Research and Development, 11199 Sorrento Valley Road, San Diego, CA (United States); Zhang Lei [Genetronics, Inc., Department of Research and Development, 11199 Sorrento Valley Road, San Diego, CA (United States)

    2003-02-21

    The effectiveness of potentially powerful therapeutics, including DNA, is often limited by their inability to permeate the cell membrane efficiently. Electroporation (EP) also referred to as 'electropermeabilization' of the outer cell membrane renders this barrier temporarily permeable by inducing 'pores' across the lipid bilayer. For in vivo EP, the drug or DNA is delivered into the interstitial space of the target tissue by conventional means, followed by local EP. EP pulses of micro- to millisecond duration and field strengths of 100-1500 V cm{sup -1} generally enhance the delivery of certain chemotherapeutic drugs by three to four orders of magnitude and intracellular delivery of DNA several hundred-fold. We have used EP in clinical studies for human cancer therapy and in animals for gene therapy and DNA vaccination. Late stage squamous cell carcinomas of the head and neck were treated with intratumoural injection of bleomycin and subsequent EP. Of the 69 tumours treated, 25% disappeared completely and another 32% were reduced in volume by more than half. Residence time of bleomycin in electroporated tumours was significantly greater than in non-electroporated lesions. Histological findings and gene expression patterns after bleomycin-EP treatment indicated rapid apoptosis of the majority of tumour cells. In animals, we demonstrated the usefulness of EP for enhanced DNA delivery by achieving normalization of blood clotting times in haemophilic dogs, and by substantially increasing transgene expression in smooth muscle cells of arterial walls using a novel porous balloon EP catheter. Finally, we have found in animal experiments that the immune response to DNA vaccines can be dramatically enhanced and accelerated by EP and co-injection of micron-sized particles. We conclude that EP represents an effective, economical and safe approach to enhance the intracellular delivery, and thus potency, of important drugs and genes for therapeutic purposes

  3. Enhancement of therapeutic drug and DNA delivery into cells by electroporation

    International Nuclear Information System (INIS)

    Rabussay, Dietmar; Dev, Nagendu B; Fewell, Jason; Smith, Louis C; Widera, Georg; Zhang Lei

    2003-01-01

    The effectiveness of potentially powerful therapeutics, including DNA, is often limited by their inability to permeate the cell membrane efficiently. Electroporation (EP) also referred to as 'electropermeabilization' of the outer cell membrane renders this barrier temporarily permeable by inducing 'pores' across the lipid bilayer. For in vivo EP, the drug or DNA is delivered into the interstitial space of the target tissue by conventional means, followed by local EP. EP pulses of micro- to millisecond duration and field strengths of 100-1500 V cm -1 generally enhance the delivery of certain chemotherapeutic drugs by three to four orders of magnitude and intracellular delivery of DNA several hundred-fold. We have used EP in clinical studies for human cancer therapy and in animals for gene therapy and DNA vaccination. Late stage squamous cell carcinomas of the head and neck were treated with intratumoural injection of bleomycin and subsequent EP. Of the 69 tumours treated, 25% disappeared completely and another 32% were reduced in volume by more than half. Residence time of bleomycin in electroporated tumours was significantly greater than in non-electroporated lesions. Histological findings and gene expression patterns after bleomycin-EP treatment indicated rapid apoptosis of the majority of tumour cells. In animals, we demonstrated the usefulness of EP for enhanced DNA delivery by achieving normalization of blood clotting times in haemophilic dogs, and by substantially increasing transgene expression in smooth muscle cells of arterial walls using a novel porous balloon EP catheter. Finally, we have found in animal experiments that the immune response to DNA vaccines can be dramatically enhanced and accelerated by EP and co-injection of micron-sized particles. We conclude that EP represents an effective, economical and safe approach to enhance the intracellular delivery, and thus potency, of important drugs and genes for therapeutic purposes. The safety and pharmaco

  4. Nanoparticles as conjugated delivery agents for therapeutic applications

    Science.gov (United States)

    Muroski, Megan Elizabeth

    This dissertation explores the use of nanoparticles as conjugated delivery agents. Chapter 1 is a general introduction. Chapter 2 discusses the delivery by a nanoparticle platform provides a method to manipulate gene activation, by taking advantage of the high surface area of a nanoparticle and the ability to selectively couple a desired biological moiety to the NP surface. The nanoparticle based transfection approach functions by controlled release of gene regulatory elements from a 6 nm AuNP (gold nanoparticle) surface. The endosomal release of the regulatory elements from the nanoparticle surface results in endogenous protein knockdown simultaneously with exogenous protein expression for the first 48 h. The use of fluorescent proteins as the endogenous and exogenous signals for protein expression enables the efficiency of co-delivery of siRNA (small interfering RNA) for GFP (green fluorescent protein) knockdown and a dsRed-express linearized plasmid for induction to be optically analyzed in CRL-2794, a human kidney cell line expressing an unstable green fluorescent protein. Delivery of the bimodal nanoparticle in cationic liposomes results in 20% GFP knockdown within 24 h of delivery and continues exhibiting knockdown for up to 48 h for the bimodal agent. Simultaneous dsRed expression is observed to initiate within the same time frame with expression levels reaching 34% after 25 days although cells have divided approximately 20 times, implying daughter cell transfection has occurred. Fluorescence cell sorting results in a stable colony, as demonstrated by Western blot analysis. The simultaneous delivery of siRNA and linearized plasmid DNA on the surface of a single nanocrystal provides a unique method for definitive genetic control within a single cell and leads to a very efficient cell transfection protocol. In Chapter 3, we wanted to understand the NP complex within the cell, and to look at the dynamics of release utilizing nanometal surface energy transfer as

  5. A sight on the current nanoparticle-based gene delivery vectors

    Science.gov (United States)

    Dizaj, Solmaz Maleki; Jafari, Samira; Khosroushahi, Ahmad Yari

    2014-05-01

    Nowadays, gene delivery for therapeutic objects is considered one of the most promising strategies to cure both the genetic and acquired diseases of human. The design of efficient gene delivery vectors possessing the high transfection efficiencies and low cytotoxicity is considered the major challenge for delivering a target gene to specific tissues or cells. On this base, the investigations on non-viral gene vectors with the ability to overcome physiological barriers are increasing. Among the non-viral vectors, nanoparticles showed remarkable properties regarding gene delivery such as the ability to target the specific tissue or cells, protect target gene against nuclease degradation, improve DNA stability, and increase the transformation efficiency or safety. This review attempts to represent a current nanoparticle based on its lipid, polymer, hybrid, and inorganic properties. Among them, hybrids, as efficient vectors, are utilized in gene delivery in terms of materials (synthetic or natural), design, and in vitro/ in vivo transformation efficiency.

  6. Recent Advancements in Targeted Delivery of Therapeutic Molecules in Neurodegenerative Disease - Spinocerebellar Ataxia - Opportunities and Challenges

    Directory of Open Access Journals (Sweden)

    Satya Prakash

    2008-01-01

    Full Text Available Drug discovery and its methodologies have been very effective in terms of treating cancers and immunological disorders but have not been able to stop genetic diseases as most of the drugs target at the protein level. They merely mitigate the symptoms of the disease. Spinocerebellar ataxia is a neurological genetic disorder that is caused by the formation of an abnormal protein. There have been several reports on ataxic drug development but actual clinical treatment is yet to be achieved. Oligonucleotide therapy called sequence specific siRNA mediated gene silencing has evolved with promising results. This approach emphasizes on suppressing the expression of the diseased gene at mRNA level. However, there is a limitation in delivery of siRNA to the target site. Several methods have been developed over the last decade to enhance the target specific delivery of DNA, siRNA, protein and small drug molecules for therapeutic purpose with less or no side effects. This review discusses the latest upcoming technologies in the field that focus on a number of nonviral nanocarriers for targeted delivery. In this review, we explore the promise and potential of novel therapeutics with interest on ataxia therapy.

  7. Layered double hydroxide nanoparticles in gene and drug delivery.

    Science.gov (United States)

    Ladewig, Katharina; Xu, Zhi Ping; Lu, Gao Qing Max

    2009-09-01

    Layered double hydroxides (LDHs) have been known for many decades as catalyst and ceramic precursors, traps for anionic pollutants, catalysts and additives for polymers, but their successful synthesis on the nanometer scale a few years ago opened up a whole new field for their application in nanomedicine. The delivery of drugs and other therapeutic/bioactive molecules (e.g., peptides, proteins, nucleic acids) to mammalian cells is an area of research that is of tremendous importance to medicine and provides manifold applications for any new developments in the area of nanotechnology. Among the many different nanoparticles that have been shown to facilitate gene and/or drug delivery, LDH nanoparticles have attracted particular attention owing to their many desirable properties. This review aims to report recent progress in gene and drug delivery using LDH nanoparticles. It summarizes the advantages and disadvantages of using LDH nanoparticles as carriers for nucleic acids and drugs against the general background of bottlenecks that are encountered by cellular delivery systems. It describes further the models that have been proposed for the internalization of LDH nanoparticles into cells so far and discusses the intracellular fate of the particles and their cargo. The authors offer some remarks on how this field of research will progress in the near future and which challenges need to be overcome before LDH nanoparticles can be used in a clinical setting.

  8. Nanotechnological strategies for nerve growth factor delivery: Therapeutic implications in Alzheimer's disease.

    Science.gov (United States)

    Faustino, Célia; Rijo, Patrícia; Reis, Catarina Pinto

    2017-06-01

    Alzheimer's disease (AD) is a progressive neurodegenerative disorder associated with amyloid-β peptide misfolding and aggregation. Neurotrophic factors, such as nerve growth factor (NGF), can prevent neuronal damage and rescue the cholinergic neurons that undergo cell death in AD, reverse deposition of extracellular amyloid plaques and improve cognitive deficits. However, NGF administration is hampered by the poor pharmacokinetic profile of the therapeutic protein and its inability to cross the blood-brain barrier, which requires specialised drug delivery systems (DDS) for efficient NGF delivery to the brain. This review covers the main therapeutic approaches that have been developed for NGF delivery targeting the brain, from polymeric implants to gene and cell-based therapies, focusing on the role of nanoparticulate systems for the sustained release of NGF in the brain as a neuroprotective and disease-modifying approach toward AD. Lipid- and polymer-based delivery systems, magnetic nanoparticles and quantum dots are specifically addressed as promising nanotechnological strategies to overcome the current limitations of NGF-based therapies. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Hybrid Nanomaterial Complexes for Advanced Phage-guided Gene Delivery

    Directory of Open Access Journals (Sweden)

    Teerapong Yata

    2014-01-01

    Full Text Available Developing nanomaterials that are effective, safe, and selective for gene transfer applications is challenging. Bacteriophages (phage, viruses that infect bacteria only, have shown promise for targeted gene transfer applications. Unfortunately, limited progress has been achieved in improving their potential to overcome mammalian cellular barriers. We hypothesized that chemical modification of the bacteriophage capsid could be applied to improve targeted gene delivery by phage vectors into mammalian cells. Here, we introduce a novel hybrid system consisting of two classes of nanomaterial systems, cationic polymers and M13 bacteriophage virus particles genetically engineered to display a tumor-targeting ligand and carry a transgene cassette. We demonstrate that the phage complex with cationic polymers generates positively charged phage and large aggregates that show enhanced cell surface attachment, buffering capacity, and improved transgene expression while retaining cell type specificity. Moreover, phage/polymer complexes carrying a therapeutic gene achieve greater cancer cell killing than phage alone. This new class of hybrid nanomaterial platform can advance targeted gene delivery applications by bacteriophage.

  10. Biodegradable polymer nanocarriers for therapeutic antisense microRNA delivery in living animals

    Science.gov (United States)

    Paulmurugan, Ramasamy; Sekar, Narayana M.; Sekar, Thillai V.

    2012-03-01

    MicroRNAs are endogenous regulators of gene expression, deregulated in several cellular diseases including cancer. Altering the cellular microenvironment by modulating the microRNAs functions can regulate different genes involved in major cellular processes, and this approach is now being investigated as a promising new generation of molecularly targeted anti-cancer therapies. AntagomiRs (Antisense-miRNAs) are a novel class of chemically modified stable oligonucleotides used for blocking the functions of endogenous microRNAs, which are overexpressed. A key challenge in achieving effective microRNAbased therapeutics lies in the development of an efficient delivery system capable of specifically delivering antisense oligonucleotides and target cancer cells in living animals. We are now developing an effective delivery system designed to selectively deliver antagomiR- 21 and antagomiR-10b to triple negative breast cancer cells, and to revert tumor cell metastasis and invasiveness. The FDA-approved biodegradable PLGA-nanoparticles were selected as a carrier for antagomiRs delivery. Chemically modified antagomiRs (antagomiR-21 and antagomiR-10b) were co-encapsulated in PEGylated-PLGA-nanoparticles by using the double-emulsification (W/O/W) solvent evaporation method, and the resulting average particle size of 150-200nm was used for different in vitro and in vivo experiments. The antagomiR encapsulated PLGA-nanoparticles were evaluated for their in vitro antagomiRs delivery, intracellular release profile, and antagomiRs functional effects, by measuring the endogenous cellular targets, and the cell growth and metastasis. The xenografts of tumor cells in living mice were used for evaluating the anti-metastatic and anti-invasive properties of cells. The results showed that the use of PLGA for antagomiR delivery is not only efficient in crossing cell membrane, but can also maintain functional intracellular antagomiRs level for a extended period of time and achieve

  11. Application of Mesenchymal Stem Cells for Therapeutic Agent Delivery in Anti-tumor Treatment

    Directory of Open Access Journals (Sweden)

    Daria S. Chulpanova

    2018-03-01

    Full Text Available Mesenchymal stem cells (MSCs are non-hematopoietic progenitor cells, which can be isolated from different types of tissues including bone marrow, adipose tissue, tooth pulp, and placenta/umbilical cord blood. There isolation from adult tissues circumvents the ethical concerns of working with embryonic or fetal stem cells, whilst still providing cells capable of differentiating into various cell lineages, such as adipocytes, osteocytes and chondrocytes. An important feature of MSCs is the low immunogenicity due to the lack of co-stimulatory molecules expression, meaning there is no need for immunosuppression during allogenic transplantation. The tropism of MSCs to damaged tissues and tumor sites makes them a promising vector for therapeutic agent delivery to tumors and metastatic niches. MSCs can be genetically modified by virus vectors to encode tumor suppressor genes, immunomodulating cytokines and their combinations, other therapeutic approaches include MSCs priming/loading with chemotherapeutic drugs or nanoparticles. MSCs derived membrane microvesicles (MVs, which play an important role in intercellular communication, are also considered as a new therapeutic agent and drug delivery vector. Recruited by the tumor, MSCs can exhibit both pro- and anti-oncogenic properties. In this regard, for the development of new methods for cancer therapy using MSCs, a deeper understanding of the molecular and cellular interactions between MSCs and the tumor microenvironment is necessary. In this review, we discuss MSC and tumor interaction mechanisms and review the new therapeutic strategies using MSCs and MSCs derived MVs for cancer treatment.

  12. Junk DNA enhances pEI-based non-viral gene delivery

    NARCIS (Netherlands)

    Gaal, E.V.B. van; Oosting, R.S.; Hennink, W.E.; Crommelin, D.J.A.; Mastrobattista, E.

    Gene therapy aims at delivering exogenous DNA into the nuclei of target cells to establish expression of a therapeutic protein. Non-viral gene delivery is examined as a safer alternative to viral approaches, but is presently characterized by a low efficiency. In the past years several non-viral

  13. Solidified self-nanoemulsifying formulation for oral delivery of combinatorial therapeutic regimen

    DEFF Research Database (Denmark)

    Jain, Amit K; Thanki, Kaushik; Jain, Sanyog

    2014-01-01

    PURPOSE: The present work reports rationalized development and characterization of solidified self-nanoemulsifying drug delivery system for oral delivery of combinatorial (tamoxifen and quercetin) therapeutic regimen. METHODS: Suitable oil for the preparation of liquid SNEDDS was selected based...

  14. Barriers to Liposomal Gene Delivery: from Application Site to the Target.

    Science.gov (United States)

    Saffari, Mostafa; Moghimi, Hamid Reza; Dass, Crispin R

    2016-01-01

    Gene therapy is a therapeutic approach to deliver genetic material into cells to alter their function in entire organism. One promising form of gene delivery system (DDS) is liposomes. The success of liposome-mediated gene delivery is a multifactorial issue and well-designed liposomal systems might lead to optimized gene transfection particularly in vivo. Liposomal gene delivery systems face different barriers from their site of application to their target, which is inside the cells. These barriers include presystemic obstacles (epithelial barriers), systemic barriers in blood circulation and cellular barriers. Epithelial barriers differ depending on the route of administration. Systemic barriers include enzymatic degradation, binding and opsonisation. Both of these barriers can act as limiting hurdles that genetic material and their vector should overcome before reaching the cells. Finally liposomes should overcome cellular barriers that include cell entrance, endosomal escape and nuclear uptake. These barriers and their impact on liposomal gene delivery will be discussed in this review.

  15. Field distribution and DNA transport in solid tumors during electric field-mediated gene delivery.

    Science.gov (United States)

    Henshaw, Joshua W; Yuan, Fan

    2008-02-01

    Gene therapy has a great potential in cancer treatment. However, the efficacy of cancer gene therapy is currently limited by the lack of a safe and efficient means to deliver therapeutic genes into the nucleus of tumor cells. One method under investigation for improving local gene delivery is based on the use of pulsed electric field. Despite repeated demonstration of its effectiveness in vivo, the underlying mechanisms behind electric field-mediated gene delivery remain largely unknown. Without a thorough understanding of these mechanisms, it will be difficult to further advance the gene delivery. In this review, the electric field-mediated gene delivery in solid tumors will be examined by following individual transport processes that must occur in vivo for a successful gene transfer. The topics of examination include: (i) major barriers for gene delivery in the body, (ii) distribution of electric fields at both cell and tissue levels during the application of external fields, and (iii) electric field-induced transport of genes across each of the barriers. Through this approach, the review summarizes what is known about the mechanisms behind electric field-mediated gene delivery and what require further investigations in future studies.

  16. Design of multimodal degradable hydrogels for controlled therapeutic delivery

    Science.gov (United States)

    Kharkar, Prathamesh Madhav

    Hydrogels are of growing interest for the delivery of therapeutics to specific sites in the body. For localized drug delivery, hydrophilic polymeric precursors often are laden with bioactive moieties and then directly injected to the site of interest for in situ gel formation. The release of physically entrapped cargo is dictated by Fickian diffusion, degradation of the drug carrier, or a combination of both. The goal of this work was to design and characterize degradable hydrogel formulations that are responsive to multiple biologically relevant stimuli for degradation-mediated delivery of cargo molecules such as therapeutic proteins, growth factors, and immunomodulatory agents. We began by demonstrating the use of cleavable click linkages formed by Michael-type addition reactions in conjunction with hydrolytically cleavable functionalities for the degradation of injectable hydrogels by endogenous stimuli for controlled protein release. Specifically, the reaction between maleimides and thiols was utilized for hydrogel formation, where thiol selection dictates the degradability of the resulting linkage under thiol-rich reducing conditions. Relevant microenvironments where degradation would occur in vivo include those rich in glutathione (GSH), a tripeptide that is found at elevated concentrations in carcinoma tissues. Degradation of the hydrogels was monitored with rheometry and volumetric swelling measurements. Arylthiol-based thioether succinimide linkages underwent degradation via click cleavage and thiol exchange reaction in the presence of GSH and via ester hydrolysis, whereas alkylthiol-based thioether succinimide linkages only undergo degradation by only ester hydrolysis. The resulting control over the degradation rate within a reducing microenvironment resulted in 2.5 fold differences in the release profile of the model protein, a fluorescently-labeled bovine serum albumin, from dually degradable hydrogels compared to non-degradable hydrogels, where the

  17. Biosensor-controlled gene therapy/drug delivery with nanoparticles for nanomedicine

    Science.gov (United States)

    Prow, Tarl W.; Rose, William A.; Wang, Nan; Reece, Lisa M.; Lvov, Yuri; Leary, James F.

    2005-04-01

    Nanomedicine involves cell-by-cell regenerative medicine, either repairing cells one at a time or triggering apoptotic pathways in cells that are not repairable. Multilayered nanoparticle systems are being constructed for the targeted delivery of gene therapy to single cells. Cleavable shells containing targeting, biosensing, and gene therapeutic molecules are being constructed to direct nanoparticles to desired intracellular targets. Therapeutic gene sequences are controlled by biosensor-activated control switches to provide the proper amount of gene therapy on a single cell basis. The central idea is to set up gene therapy "nanofactories" inside single living cells. Molecular biosensors linked to these genes control their expression. Gene delivery is started in response to a biosensor detected problem; gene delivery is halted when the cell response indicates that more gene therapy is not needed. Cell targeting of nanoparticles, both nanocrystals and nanocapsules, has been tested by a combination of fluorescent tracking dyes, fluorescence microscopy and flow cytometry. Intracellular targeting has been tested by confocal microscopy. Successful gene delivery has been visualized by use of GFP reporter sequences. DNA tethering techniques were used to increase the level of expression of these genes. Integrated nanomedical systems are being designed, constructed, and tested in-vitro, ex-vivo, and in small animals. While still in its infancy, nanomedicine represents a paradigm shift in thinking-from destruction of injured cells by surgery, radiation, chemotherapy to cell-by-cell repair within an organ and destruction of non-repairable cells by natural apoptosis.

  18. Biodegradable Polyphosphazene Biomaterials for Tissue Engineering and Delivery of Therapeutics

    Directory of Open Access Journals (Sweden)

    Amanda L. Baillargeon

    2014-01-01

    Full Text Available Degradable biomaterials continue to play a major role in tissue engineering and regenerative medicine as well as for delivering therapeutic agents. Although the chemistry of polyphosphazenes has been studied extensively, a systematic review of their applications for a wide range of biomedical applications is lacking. Polyphosphazenes are synthesized through a relatively well-known two-step reaction scheme which involves the substitution of the initial linear precursor with a wide range of nucleophiles. The ease of substitution has led to the development of a broad class of materials that have been studied for numerous biomedical applications including as scaffold materials for tissue engineering and regenerative medicine. The objective of this review is to discuss the suitability of poly(amino acid esterphosphazene biomaterials in regard to their unique stimuli responsive properties, tunable degradation rates and mechanical properties, as well as in vitro and in vivo biocompatibility. The application of these materials in areas such as tissue engineering and drug delivery is discussed systematically. Lastly, the utility of polyphosphazenes is further extended as they are being employed in blend materials for new applications and as another method of tailoring material properties.

  19. Membrane-Mimic Nanoparticles for Drug and Gene Delivery

    KAUST Repository

    Alamoudi, Kholod

    2017-12-01

    Nanoscale organic particles have gained a prominent role in drug and gene delivery field. As the nature of the nanoparticle’s (NPs) surface plays a major role in their targeting efficiency, bioavailability, and cytotoxicity, membrane-mimic nanoparticles are considered highly attractive materials for in vivo and in vitro applications. Synthetic membrane vesicles (liposomes) and nanoconstructs built with native cancer cellular membrane are excellent scaffolds to improve cellular delivery. Liposomes have been extensively used due to their high loading capacity, biocompatibility and biodegradability. However, modifications with stimuli responsive materials are highly needed to improve their stability and turn them active participants in controlled delivery. Towards a nature inspired approach, reconstructed bilayers from cell membrane are a good candidate to enhance NP’s targeting ability and biocompatibility. The primary focus of this research is to develop smart responsive (lipid) membrane coated NPs with surface modifications for controlled and targeted drug and/or gene delivery for application in cancer therapy. Three approaches have been developed, namely i) liposomes as thermoresponsive nanocarriers for the delivery of genetic material; ii) magnetically photosensitive liposome hybrids and iii) biomimetic periodic mesoporous organo silica engineered for better a biocompatibility and targeting capabilities. In the first project synthetic liposomes were loaded with ammonium bicarbonate salt (ABC) and siRNA. The combination of lipids chosen and the relative ratios allowed the rapid release of the genetic material to the multi drug resistant cancer cells studied, upon external heat trigger. This design has improved the gene silencing efficiency via successful endosomal escape. In the second project, SPIO@Au nanoparticles were imbedded in the lipid bilayer to produce a photo/thermal responsive carrier that could be also used in cell imaging besides gene transfection

  20. Matricellular proteins in drug delivery: Therapeutic targets, active agents, and therapeutic localization.

    Science.gov (United States)

    Sawyer, Andrew J; Kyriakides, Themis R

    2016-02-01

    Extracellular matrix is composed of a complex array of molecules that together provide structural and functional support to cells. These properties are mainly mediated by the activity of collagenous and elastic fibers, proteoglycans, and proteins such as fibronectin and laminin. ECM composition is tissue-specific and could include matricellular proteins whose primary role is to modulate cell-matrix interactions. In adults, matricellular proteins are primarily expressed during injury, inflammation and disease. Particularly, they are closely associated with the progression and prognosis of cardiovascular and fibrotic diseases, and cancer. This review aims to provide an overview of the potential use of matricellular proteins in drug delivery including the generation of therapeutic agents based on the properties and structures of these proteins as well as their utility as biomarkers for specific diseases. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. Dual delivery systems based on polyamine analog BENSpm as prodrug and gene delivery vectors

    Science.gov (United States)

    Zhu, Yu

    Combination drug and gene therapy shows promise in cancer treatment. However, the success of such strategy requires careful selection of the therapeutic agents, as well as development of efficient delivery vectors. BENSpm (N 1, N11-bisethylnorspermine), a polyamine analogue targeting the intracellular polyamine pathway, draws our special attention because of the following reasons: (1) polyamine pathway is frequently dysregulated in cancer; (2) BENSpm exhibits multiple functions to interfere with the polyamine pathway, such as to up-regulate polyamine metabolism enzymes and down-regulate polyamine biosynthesis enzymes. Therefore BENSpm depletes all natural polyamines and leads to apoptosis and cell growth inhibition in a wide range of cancers; (3) preclinical studies proved that BENSpm can act synergistically with various chemotherapy agents, making it a promising candidate in combination therapy; (4) multiple positive charges in BENSpm enable it as a suitable building block for cationic polymers, which can be further applied to gene delivery. In this dissertation, our goal was to design dual-function delivery vector based on BENSpm that can function as a gene delivery vector and, after intracellular degradation, as an active anticancer agent targeting dysregulated polyamine metabolism. We first demonstrated strong synergism between BENSpm and a potential therapeutic gene product TRAIL. Strong synergism was obtained in both estrogen-dependent MCF-7 breast cancer cells and triple-negative MDA-MB-231 breast cancer cells. Significant dose reduction of TRAIL in combination with BENSpm in MDA-MB-231 cells, together with the fact that BENSpm rendered MCF-7 cells more sensitive to TRAIL treatment verified our rationale of designing BENSpm-based delivery platform. This was expected to be beneficial for overcoming drug resistance in chemotherapy, as well as boosting the therapeutic effect of therapeutic genes. We first designed a lipid-based BENSpm dual vector (Lipo

  2. Cell mediated therapeutics for cancer treatment: Tumor homing cells as therapeutic delivery vehicles

    Science.gov (United States)

    Balivada, Sivasai

    Many cell types were known to have migratory properties towards tumors and different research groups have shown reliable results regarding cells as delivery vehicles of therapeutics for targeted cancer treatment. Present report discusses proof of concept for 1. Cell mediated delivery of Magnetic nanoparticles (MNPs) and targeted Magnetic hyperthermia (MHT) as a cancer treatment by using in vivo mouse cancer models, 2. Cells surface engineering with chimeric proteins for targeted cancer treatment by using in vitro models. 1. Tumor homing cells can carry MNPs specifically to the tumor site and tumor burden will decrease after alternating magnetic field (AMF) exposure. To test this hypothesis, first we loaded Fe/Fe3O4 bi-magnetic NPs into neural progenitor cells (NPCs), which were previously shown to migrate towards melanoma tumors. We observed that NPCs loaded with MNPs travel to subcutaneous melanoma tumors. After alternating magnetic field (AMF) exposure, the targeted delivery of MNPs by the NPCs resulted in a mild decrease in tumor size (Chapter-2). Monocytes/macrophages (Mo/Ma) are known to infiltrate tumor sites, and also have phagocytic activity which can increase their uptake of MNPs. To test Mo/Ma-mediated MHT we transplanted Mo/Ma loaded with MNPs into a mouse model of pancreatic peritoneal carcinomatosis. We observed that MNP-loaded Mo/Ma infiltrated pancreatic tumors and, after AMF treatment, significantly prolonged the lives of mice bearing disseminated intraperitoneal pancreatic tumors (Chapter-3). 2. Targeted cancer treatment could be achieved by engineering tumor homing cell surfaces with tumor proteases cleavable, cancer cell specific recombinant therapeutic proteins. To test this, Urokinase and Calpain (tumor specific proteases) cleavable; prostate cancer cell (CaP) specific (CaP1 targeting peptide); apoptosis inducible (Caspase3 V266ED3)- rCasp3V266ED3 chimeric protein was designed in silico. Hypothesized membrane anchored chimeric protein (rCasp3V

  3. Innovations in gene and growth factor delivery systems for diabetic wound healing

    Science.gov (United States)

    Laiva, Ashang Luwang; O'Brien, Fergal J.

    2017-01-01

    Abstract The rise in lower extremity amputations due to nonhealing of foot ulcers in diabetic patients calls for rapid improvement in effective treatment regimens. Administration of growth factors (GFs) are thought to offer an off‐the‐shelf treatment; however, the dose‐ and time‐dependent efficacy of the GFs together with the hostile environment of diabetic wound beds impose a major hindrance in the selection of an ideal route for GF delivery. As an alternative, the delivery of therapeutic genes using viral and nonviral vectors, capable of transiently expressing the genes until the recovery of the wounded tissue offers promise. The development of implantable biomaterial dressings capable of modulating the release of either single or combinatorial GFs/genes may offer solutions to this overgrowing problem. This article reviews the state of the art on gene and protein delivery and the strategic optimization of clinically adopted delivery strategies for the healing of diabetic wounds. PMID:28482114

  4. Gene delivery to skeletal muscle results in sustained expression and systemic delivery of a therapeutic protein

    Science.gov (United States)

    Kessler, Paul D.; Podsakoff, Gregory M.; Chen, Xiaojuan; McQuiston, Susan A.; Colosi, Peter C.; Matelis, Laura A.; Kurtzman, Gary J.; Byrne, Barry J.

    1996-01-01

    Somatic gene therapy has been proposed as a means to achieve systemic delivery of therapeutic proteins. However, there is limited evidence that current methods of gene delivery can practically achieve this goal. In this study, we demonstrate that, following a single intramuscular administration of a recombinant adeno-associated virus (rAAV) vector containing the β-galactosidase (AAV-lacZ) gene into adult BALB/c mice, protein expression was detected in myofibers for at least 32 weeks. A single intramuscular administration of an AAV vector containing a gene for human erythropoietin (AAV-Epo) into mice resulted in dose-dependent secretion of erythropoietin and corresponding increases in red blood cell production that persisted for up to 40 weeks. Primary human myotubes transduced in vitro with the AAV-Epo vector also showed dose-dependent production of Epo. These results demonstrate that rAAV vectors are able to transduce skeletal muscle and are capable of achieving sustained expression and systemic delivery of a therapeutic protein following a single intramuscular administration. Gene therapy using AAV vectors may provide a practical strategy for the treatment of inherited and acquired protein deficiencies. PMID:8943064

  5. Chitosan-Based Nanoparticles for Mucosal Delivery of RNAi Therapeutics

    DEFF Research Database (Denmark)

    Martirosyan, Alina; Olesen, Morten Jarlstad; Howard, Kenneth A.

    2014-01-01

    of the polysaccharide chitosan have been used to facilitate delivery of siRNA across mucosal surfaces following local administration. This chapter describes the mucosal barriers that need to be addressed in order to design an effective mucosal delivery strategy and the utilization of the mucoadhesive properties...... of chitosan. Focus is given to preparation methods and the preclinical application of chitosan nanoparticles for respiratory and oral delivery of siRNA....

  6. Cell based-gene delivery approaches for the treatment of spinal cord injury and neurodegenerative disorders.

    Science.gov (United States)

    Taha, Masoumeh Fakhr

    2010-03-01

    Cell based-gene delivery has provided an important therapeutic strategy for different disorders in the recent years. This strategy is based on the transplantation of genetically modified cells to express specific genes and to target the delivery of therapeutic factors, especially for the treatment of cancers and neurological, immunological, cardiovascular and heamatopoietic disorders. Although, preliminary reports are encouraging, and experimental studies indicate functionally and structurally improvements in the animal models of different disorders, universal application of this strategy for human diseases requires more evidence. There are a number of parameters that need to be evaluated, including the optimal cell source, the most effective gene/genes to be delivered, the optimal vector and method of gene delivery into the cells and the most efficient route for the delivery of genetically modified cells into the patient. Also, some obstacles have to be overcome, including the safety and usefulness of the approaches and the stability of the improvements. Here, recent studies concerning with the cell-based gene delivery for spinal cord injury and some neurodegenerative disorders such as amyotrophic lateral sclerosis, Parkinson's disease and Alzheimer's disease are briefly reviewed, and their exciting consequences are discussed.

  7. Coating nanocarriers with hyaluronic acid facilitates intravitreal drug delivery for retinal gene therapy

    NARCIS (Netherlands)

    Martens, Thomas F.; Remaut, Katrien; Deschout, Hendrik; Engbersen, Johan F J; Hennink, Wim E.; Van Steenbergen, Mies J.; Demeester, Jo; De Smedt, Stefaan C.; Braeckmans, Kevin

    2015-01-01

    Retinal gene therapy could potentially affect the lives of millions of people suffering from blinding disorders. Yet, one of the major hurdles remains the delivery of therapeutic nucleic acids to the retinal target cells. Due to the different barriers that need to be overcome in case of topical or

  8. Targeted drug delivery to magnetic implants for therapeutic applications

    International Nuclear Information System (INIS)

    Yellen, Benjamin B.; Forbes, Zachary G.; Halverson, Derek S.; Fridman, Gregory; Barbee, Kenneth A.; Chorny, Michael; Levy, Robert; Friedman, Gary

    2005-01-01

    A new method for locally targeted drug delivery is proposed that employs magnetic implants placed directly in the cardiovascular system to attract injected magnetic carriers. Theoretical simulations and experimental results support the assumption that using magnetic implants in combination with externally applied magnetic field will optimize the delivery of magnetic drug to selected sites within a subject

  9. Optimization of Intracellular Transportation of Gene Therapeutic DNA in Small Cell Lung Cancer (Ph.d.)

    DEFF Research Database (Denmark)

    Cramer, Frederik

    2013-01-01

    Small cell lung cancer (SCLC) is a highly malignant disease characterized as being very aggressive and metastasizing at a rapid pace. The malevolent pace of SCLC cell migration results in almost three out of four SCLC patients having disseminated SCLC at the time of diagnosis. Unfortunately...... has to be able to repeated systemic delivery of gene therapy to cancer cells in a both safe and efficient way. Non-viral delivery vectors fulfill many of these requirements except the latter. It is currently very difficult to systemically transport sufficient amounts of therapeutic DNA, by a non......-viral delivery system, to the nuclei of the SCLC cells. As a result, the gene therapy expression obtained is too low to have any clinical relevance. We have at the Department of Radiation Biology developed a transcriptionally targeting suicide gene therapy system which is built on a double stranded DNA plasmid...

  10. Preclinical evaluation of gene delivery methods for the treatment of loco-regional disease in breast cancer.

    LENUS (Irish Health Repository)

    Rajendran, Simon

    2011-04-01

    Preclinical results with various gene therapy strategies indicate significant potential for new cancer treatments. However, many therapeutics fail at clinical trial, often due to differences in tissue physiology between animal models and humans, and tumor phenotype variation. Clinical data relevant to treatment strategies may be generated prior to clinical trial through experimentation using intact patient tissue ex vivo. We developed a novel tumor slice model culture system that is universally applicable to gene delivery methods, using a realtime luminescence detection method to assess gene delivery. Methods investigated include viruses (adenovirus [Ad] and adeno-associated virus), lipofection, ultrasound (US), electroporation and naked DNA. Viability and tumor populations within the slices were well maintained for seven days, and gene delivery was qualitatively and quantitatively examinable for all vectors. Ad was the most efficient gene delivery vector with transduction efficiency >50%. US proved the optimal non-viral gene delivery method in human tumor slices. The nature of the ex vivo culture system permitted examination of specific elements. Parameters shown to diminish Ad gene delivery included blood, regions of low viability and secondary disease. US gene delivery was significantly reduced by blood and skin, while tissue hyperthermia improved gene delivery. US achieved improved efficacy for secondary disease. The ex vivo model was also suitable for examination of tissue-specific effects on vector expression, with Ad expression mediated by the CXCR4 promoter shown to provide a tumor selective advantage over the ubiquitously active cytomegalovirus promoter. In conclusion, this is the first study incorporating patient tissue models in comparing gene delivery from various vectors, providing knowledge on cell-type specificity and examining the crucial biological factors determining successful gene delivery. The results highlight the importance of in

  11. Preclinical evaluation of gene delivery methods for the treatment of loco-regional disease in breast cancer.

    LENUS (Irish Health Repository)

    Rajendran, Simon

    2012-01-31

    Preclinical results with various gene therapy strategies indicate significant potential for new cancer treatments. However, many therapeutics fail at clinical trial, often due to differences in tissue physiology between animal models and humans, and tumor phenotype variation. Clinical data relevant to treatment strategies may be generated prior to clinical trial through experimentation using intact patient tissue ex vivo. We developed a novel tumor slice model culture system that is universally applicable to gene delivery methods, using a realtime luminescence detection method to assess gene delivery. Methods investigated include viruses (adenovirus [Ad] and adeno-associated virus), lipofection, ultrasound (US), electroporation and naked DNA. Viability and tumor populations within the slices were well maintained for seven days, and gene delivery was qualitatively and quantitatively examinable for all vectors. Ad was the most efficient gene delivery vector with transduction efficiency >50%. US proved the optimal non-viral gene delivery method in human tumor slices. The nature of the ex vivo culture system permitted examination of specific elements. Parameters shown to diminish Ad gene delivery included blood, regions of low viability and secondary disease. US gene delivery was significantly reduced by blood and skin, while tissue hyperthermia improved gene delivery. US achieved improved efficacy for secondary disease. The ex vivo model was also suitable for examination of tissue-specific effects on vector expression, with Ad expression mediated by the CXCR4 promoter shown to provide a tumor selective advantage over the ubiquitously active cytomegalovirus promoter. In conclusion, this is the first study incorporating patient tissue models in comparing gene delivery from various vectors, providing knowledge on cell-type specificity and examining the crucial biological factors determining successful gene delivery. The results highlight the importance of in

  12. Dendrimers as Carriers for siRNA Delivery and Gene Silencing: A Review

    Directory of Open Access Journals (Sweden)

    Jiangyu Wu

    2013-01-01

    Full Text Available RNA interference (RNAi was first literaturally reported in 1998 and has become rapidly a promising tool for therapeutic applications in gene therapy. In a typical RNAi process, small interfering RNAs (siRNA are used to specifically downregulate the expression of the targeted gene, known as the term “gene silencing.” One key point for successful gene silencing is to employ a safe and efficient siRNA delivery system. In this context, dendrimers are emerging as potential nonviral vectors to deliver siRNA for RNAi purpose. Dendrimers have attracted intense interest since their emanating research in the 1980s and are extensively studied as efficient DNA delivery vectors in gene transfer applications, due to their unique features based on the well-defined and multivalent structures. Knowing that DNA and RNA possess a similar structure in terms of nucleic acid framework and the electronegative nature, one can also use the excellent DNA delivery properties of dendrimers to develop effective siRNA delivery systems. In this review, the development of dendrimer-based siRNA delivery vectors is summarized, focusing on the vector features (siRNA delivery efficiency, cytotoxicity, etc. of different types of dendrimers and the related investigations on structure-activity relationship to promote safe and efficient siRNA delivery system.

  13. Dendrimers as Carriers for siRNA Delivery and Gene Silencing: A Review

    Science.gov (United States)

    Huang, Weizhe; He, Ziying

    2013-01-01

    RNA interference (RNAi) was first literaturally reported in 1998 and has become rapidly a promising tool for therapeutic applications in gene therapy. In a typical RNAi process, small interfering RNAs (siRNA) are used to specifically downregulate the expression of the targeted gene, known as the term “gene silencing.” One key point for successful gene silencing is to employ a safe and efficient siRNA delivery system. In this context, dendrimers are emerging as potential nonviral vectors to deliver siRNA for RNAi purpose. Dendrimers have attracted intense interest since their emanating research in the 1980s and are extensively studied as efficient DNA delivery vectors in gene transfer applications, due to their unique features based on the well-defined and multivalent structures. Knowing that DNA and RNA possess a similar structure in terms of nucleic acid framework and the electronegative nature, one can also use the excellent DNA delivery properties of dendrimers to develop effective siRNA delivery systems. In this review, the development of dendrimer-based siRNA delivery vectors is summarized, focusing on the vector features (siRNA delivery efficiency, cytotoxicity, etc.) of different types of dendrimers and the related investigations on structure-activity relationship to promote safe and efficient siRNA delivery system. PMID:24288498

  14. Nanoparticle-mediated delivery of suicide genes in cancer therapy.

    Science.gov (United States)

    Vago, Riccardo; Collico, Veronica; Zuppone, Stefania; Prosperi, Davide; Colombo, Miriam

    2016-09-01

    Conventional chemotherapeutics have been employed in cancer treatment for decades due to their efficacy in killing the malignant cells, but the other side of the coin showed off-target effects, onset of drug resistance and recurrences. To overcome these limitations, different approaches have been investigated and suicide gene therapy has emerged as a promising alternative. This approach consists in the introduction of genetic materials into cancerous cells or the surrounding tissue to cause cell death or retard the growth of the tumor mass. Despite promising results obtained both in vitro and in vivo, this innovative approach has been limited, for long time, to the treatment of localized tumors, due to the suboptimal efficiency in introducing suicide genes into cancer cells. Nanoparticles represent a valuable non-viral delivery system to protect drugs in the bloodstream, to improve biodistribution, and to limit side effects by achieving target selectivity through surface ligands. In this scenario, the real potential of suicide genes can be translated into clinically viable treatments for patients. In the present review, we summarize the recent advances of inorganic nanoparticles as non-viral vectors in terms of therapeutic efficacy, targeting capacity and safety issues. We describe the main suicide genes currently used in therapy, with particular emphasis on toxin-encoding genes of bacterial and plant origin. In addition, we discuss the relevance of molecular targeting and tumor-restricted expression to improve treatment specificity to cancer tissue. Finally, we analyze the main clinical applications, limitations and future perspectives of suicide gene therapy. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. Adenoviral Gene Delivery to Primary Human Cutaneous Cells and Burn Wounds

    OpenAIRE

    Hirsch, Tobias; von Peter, Sebastian; Dubin, Grzegorz; Mittler, Dominik; Jacobsen, Frank; Lehnhardt, Markus; Eriksson, Elof; Steinau, Hans-Ulrich; Steinstraesser, Lars

    2006-01-01

    The adenoviral transfer of therapeutic genes into epidermal and dermal cells is an interesting approach to treat skin diseases and to promote wound healing. The aim of this study was to assess the in vitro and in vivo transfection efficacy in skin and burn wounds after adenoviral gene delivery. Primary keratinocytes (HKC), fibroblasts (HFB), and HaCaT cells were transfected using different concentrations of an adenoviral construct (eGFP). Transfection efficiency and cytotoxicity was determine...

  16. Chitosan-based delivery systems for protein therapeutics and antigens

    NARCIS (Netherlands)

    Amidi, M.; Mastrobattista, E.; Jiskoot, W.; Hennink, W.E.

    Therapeutic peptides/proteins and protein-based antigens are chemically and structurally labile compounds, which are almost exclusively administered by parenteral injections. Recently, non-invasive mucosal routes have attracted interest for administration of these biotherapeutics. Chitosan-based

  17. Investigation of a thiolated polymer in gene delivery

    Science.gov (United States)

    Bacalocostantis, Irene

    Thiol-containing bioreducible polymers show significant potential as delivery vectors in gene therapy, a rapidly growing field which seeks to treat genetic-based disorders by delivering functional synthetic genes to diseased cells. Studies have shown that thiolated polymers exhibit improved biodegradability and prolonged in vivo circulation times over non-thiolated polymers. However, the extent to which thiol concentrations impact the carrier's delivery potential has not been well explored. The aim of this dissertation is to investigate how relative concentrations of free thiols and disulfide crosslinks impact a polymeric carriers delivery performance with respect to DNA packaging, complex stability, cargo protection, gene release, internalization efficiency and cytotoxicity. To accomplish this goal, several fluorescent polymers containing varying concentrations of thiol groups were synthesized by conjugating thiol-pendant chains onto the primary amines of cationic poly(allylamine). In vitro delivery assays and characterization techniques were employed to assess the effect of thiols in gene delivery.

  18. Nanostructured Mineral Coatings Stabilize Proteins for Therapeutic Delivery.

    Science.gov (United States)

    Yu, Xiaohua; Biedrzycki, Adam H; Khalil, Andrew S; Hess, Dalton; Umhoefer, Jennifer M; Markel, Mark D; Murphy, William L

    2017-09-01

    Proteins tend to lose their biological activity due to their fragile structural conformation during formulation, storage, and delivery. Thus, the inability to stabilize proteins in controlled-release systems represents a major obstacle in drug delivery. Here, a bone mineral inspired protein stabilization strategy is presented, which uses nanostructured mineral coatings on medical devices. Proteins bound within the nanostructured coatings demonstrate enhanced stability against extreme external stressors, including organic solvents, proteases, and ethylene oxide gas sterilization. The protein stabilization effect is attributed to the maintenance of protein conformational structure, which is closely related to the nanoscale feature sizes of the mineral coatings. Basic fibroblast growth factor (bFGF) released from a nanostructured mineral coating maintains its biological activity for weeks during release, while it maintains activity for less than 7 d during release from commonly used polymeric microspheres. Delivery of the growth factors bFGF and vascular endothelial growth factor using a mineral coated surgical suture significantly improves functional Achilles tendon healing in a rabbit model, resulting in increased vascularization, more mature collagen fiber organization, and a two fold improvement in mechanical properties. The findings of this study demonstrate that biomimetic interactions between proteins and nanostructured minerals provide a new, broadly applicable mechanism to stabilize proteins in the context of drug delivery and regenerative medicine. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Industry Update: the latest developments in therapeutic delivery.

    Science.gov (United States)

    Steinbach, Oliver C

    2013-05-01

    The present industry update covers the period 2-28 February 2013, with information sourced from company press releases, regulatory and patent agencies, as well as from scientific literature. With the US presidential inauguration in January, the inevitable 'fiscal cliff' and with the healthcare reform taking shape, there are many challenges ahead for the world's largest healthcare market. Healthcare providers are preparing for the 'accountable care organizations' paradigm shift and it has been reported that the pharmaceutical industry is low in public perception; only 'for-profit' health insurers rank lower according to a survey recently published. Although in 2012 one could still observe a decline in the overall number of drug delivery partnering deals announced, there has been a slight recovery, thus, meaning a repeat of the lows of 2011 have not been witnessed. The experts expect a reversed trend in 2013 as larger companies seek to refill pipelines with new candidates and drug-delivery solutions. There is a lot of emphasis on altered oral delivery profiles to provide patient compliance. This is indicated by the recent licensing deals, such as that between Radius Health Inc. and 3M, TissueGen and Biomedical Structures, and new financing rounds of companies, such as SKL, Catalent and BioActiva. Injectable technology, including needle-free delivery, continues to be prevalent in deals, such as those between Clearside and Kala until compelling alternatives to the delivery of biologicals are available. Alternative routes of administration, such as nasal, buccal (Kala), sublingual (Stallergenes) see growing interest; however, bioavailability and reproducible dosing will continue to be a challenge in the foreseeable future.

  20. Targeting Potassium Channels for Increasing Delivery of Imaging Agents and Therapeutics to Brain Tumors

    Directory of Open Access Journals (Sweden)

    Nagendra Sanyasihally Ningaraj

    2013-05-01

    Full Text Available Every year in the US, 20,000 new primary and nearly 200,000 metastatic brain tumor cases are reported. The cerebral microvessels/ capillaries that form the blood–brain barrier (BBB not only protect the brain from toxic agents in the blood but also pose a significant hindrance to the delivery of small and large therapeutic molecules. Different strategies have been employed to circumvent the physiological barrier posed by blood-brain tumor barrier (BTB. Studies in our laboratory have identified significant differences in the expression levels of certain genes and proteins between normal and brain tumor capillary endothelial cells. In this study, we validated the non-invasive and clinically relevant Dynamic Contrast Enhancing-Magnetic Resonance Imaging (DCE-MRI method with invasive, clinically irrelevant but highly accurate Quantitative Autoradiography (QAR method using rat glioma model. We also showed that DCE-MRI metric of tissue vessel perfusion-permeability is sensitive to changes in blood vessel permeability following administration of calcium-activated potassium (BKCa channel activator NS-1619. Our results show that human gliomas and brain tumor endothelial cells that overexpress BKCa channels can be targeted for increased BTB permeability for MRI enhancing agents to brain tumors. We conclude that monitoring the outcome of increased MRI enhancing agents’ delivery to microsatellites and leading tumor edges in glioma patients would lead to beneficial clinical outcome.

  1. Applications of lipid based formulation technologies in the delivery of biotechnology-based therapeutics.

    Science.gov (United States)

    du Plessis, Lissinda H; Marais, Etienne B; Mohammed, Faruq; Kotzé, Awie F

    2014-01-01

    In the last decades several new biotechnologically-based therapeutics have been developed due to progress in genetic engineering. A growing challenge facing pharmaceutical scientists is formulating these compounds into oral dosage forms with adequate bioavailability. An increasingly popular approach to formulate biotechnology-based therapeutics is the use of lipid based formulation technologies. This review highlights the importance of lipid based drug delivery systems in the formulation of oral biotechnology based therapeutics including peptides, proteins, DNA, siRNA and vaccines. The different production procedures used to achieve high encapsulation efficiencies of the bioactives are discussed, as well as the factors influencing the choice of excipient. Lipid based colloidal drug delivery systems including liposomes and solid lipid nanoparticles are reviewed with a focus on recent advances and updates. We further describe microemulsions and self-emulsifying drug delivery systems and recent findings on bioactive delivery. We conclude the review with a few examples on novel lipid based formulation technologies.

  2. Intra Articular Therapeutic Delivery for Post Traumatic Osteoarthritis

    Science.gov (United States)

    2016-10-01

    size distribution therapeutic timepoints EPIC-µCT Articular cartilage Subchondral bone Osteophytes Proteoglycans 3. OVERALL PROJECT SUMMARY: In...joint degeneration induced by MMT. Previously documented in Year 1 annual report: Changes in articular cartilage and subchondral bone morphology...and resulted in increased cartilage thickness at 3 weeks. The majority of alterations to subchondral bone (density, thickness) were detected at 3

  3. Optimising oral systems for the delivery of therapeutic proteins and ...

    African Journals Online (AJOL)

    Therapeutic proteins/peptides are mostly administered as parenteral (injectable) preparations as a result of their poor oral bioavailability which is due to degradation by proteolytic enzymes, poor membrane permeability and large molecular size. However, the oral route would be preferred to the parenteral administration ...

  4. Recent Advances in Skin Penetration Enhancers for Transdermal Gene and Drug Delivery.

    Science.gov (United States)

    Amjadi, Morteza; Mostaghaci, Babak; Sitti, Metin

    2017-01-01

    There is a growing interest in transdermal delivery systems because of their noninvasive, targeted, and on-demand delivery of gene and drugs. However, efficient penetration of therapeutic compounds into the skin is still challenging largely due to the impermeability of the outermost layer of the skin, known as stratum corneum. Recently, there have been major research activities to enhance the skin penetration depth of pharmacological agents. This article reviews recent advances in the development of various strategies for skin penetration enhancement. We show that approaches such as ultrasound waves, laser, and microneedle patches have successfully been employed to physically disrupt the stratum corneum structure for enhanced transdermal delivery. Rather than physical approaches, several non-physical route have also been utilized for efficient transdermal delivery across the skin barrier. Finally, we discuss some clinical applications of transdermal delivery systems for gene and drug delivery. This paper shows that transdermal delivery devices can potentially function for diverse healthcare and medical applications while further investigations are still necessary for more efficient skin penetration of gene and drugs. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  5. P-glycoprotein trafficking as a therapeutic target to optimize CNS drug delivery.

    Science.gov (United States)

    Davis, Thomas P; Sanchez-Covarubias, Lucy; Tome, Margaret E

    2014-01-01

    The primary function of the blood-brain barrier (BBB)/neurovascular unit is to protect the central nervous system (CNS) from potentially harmful xenobiotic substances and maintain CNS homeostasis. Restricted access to the CNS is maintained via a combination of tight junction proteins as well as a variety of efflux and influx transporters that limits the transcellular and paracellular movement of solutes. Of the transporters identified at the BBB, P-glycoprotein (P-gp) has emerged as the transporter that is the greatest obstacle to effective CNS drug delivery. In this chapter, we provide data to support intracellular protein trafficking of P-gp within cerebral capillary microvessels as a potential target for improved drug delivery. We show that pain-induced changes in P-gp trafficking are associated with changes in P-gp's association with caveolin-1, a key scaffolding/trafficking protein that colocalizes with P-gp at the luminal membrane of brain microvessels. Changes in colocalization with the phosphorylated and nonphosphorylated forms of caveolin-1, by pain, are accompanied by dynamic changes in the distribution, relocalization, and activation of P-gp "pools" between microvascular endothelial cell subcellular compartments. Since redox-sensitive processes may be involved in signaling disassembly of higher-order structures of P-gp, we feel that manipulating redox signaling, via specific protein targeting at the BBB, may protect disulfide bond integrity of P-gp reservoirs and control trafficking to the membrane surface, providing improved CNS drug delivery. The advantage of therapeutic drug "relocalization" of a protein is that the physiological impact can be modified, temporarily or long term, despite pathology-induced changes in gene transcription. © 2014 Elsevier Inc. All rights reserved.

  6. [Smart therapeutics based on synthetic gene circuits].

    Science.gov (United States)

    Peng, Shuguang; Xie, Zhen

    2017-03-25

    Synthetic biology has an important impact on biology research since its birth. Applying the thought and methods that reference from electrical engineering, synthetic biology uncovers many regulatory mechanisms of life systems, transforms and expands a series of biological components. Therefore, it brings a wide range of biomedical applications, including providing new ideas for disease diagnosis and treatment. This review describes the latest advances in the field of disease diagnosis and therapy based on mammalian cell or bacterial synthetic gene circuits, and provides new ideas for future smart therapy design.

  7. Polymersome-based drug-delivery strategies for cancer therapeutics.

    Science.gov (United States)

    Anajafi, Tayebeh; Mallik, Sanku

    2015-01-01

    Polymersomes are stable vesicles prepared from amphiphilic polymers and are more stable compared with liposomes. Although these nanovesicles have many attractive properties for in vitro/in vivo applications, liposome-based drug delivery systems are still prevalent in the market. In order to expedite the translational potential and to provide medically valuable formulations, the polymersomes need to be biocompatible and biodegradable. In this review, recent developments for biocompatible and biodegradable polymersomes, including the design of intelligent, targeted, and stimuli-responsive vesicles are summarized.

  8. Malaria Prevention by New Technology: Vectored Delivery of Antibody Genes

    Science.gov (United States)

    2017-10-01

    AWARD NUMBER: W81XWH-15-1-0401 TITLE: Malaria Prevention by New Technology : Vectored Delivery of Antibody Genes PRINCIPAL INVESTIGATOR: Gary...CONTRACT NUMBER Malaria Prevention by New Technology : Vectored Delivery of Antibody Genes 5b. GRANT NUMBER W81XWH-15-1-0401 5c. PROGRAM ELEMENT...whole animals. Using a specific technology originally applied to expression of HIV antibodies, we demonstrated that mice can be protected from

  9. Novel electric power-driven hydrodynamic injection system for gene delivery: safety and efficacy of human factor IX delivery in rats.

    Science.gov (United States)

    Yokoo, T; Kamimura, K; Suda, T; Kanefuji, T; Oda, M; Zhang, G; Liu, D; Aoyagi, Y

    2013-08-01

    The development of a safe and reproducible gene delivery system is an essential step toward the clinical application of the hydrodynamic gene delivery (HGD) method. For this purpose, we have developed a novel electric power-driven injection system called the HydroJector-EM, which can replicate various time-pressure curves preloaded into the computer program before injection. The assessment of the reproducibility and safety of gene delivery system in vitro and in vivo demonstrated the precise replication of intravascular time-pressure curves and the reproducibility of gene delivery efficiency. The highest level of luciferase expression (272 pg luciferase per mg of proteins) was achieved safely using the time-pressure curve, which reaches 30 mm Hg in 10 s among various curves tested. Using this curve, the sustained expression of a therapeutic level of human factor IX protein (>500 ng ml(-1)) was maintained for 2 months after the HGD of the pBS-HCRHP-FIXIA plasmid. Other than a transient increase in liver enzymes that recovered in a few days, no adverse events were seen in rats. These results confirm the effectiveness of the HydroJector-EM for reproducible gene delivery and demonstrate that long-term therapeutic gene expression can be achieved by automatic computer-controlled hydrodynamic injection that can be performed by anyone.

  10. Rapid endosomal escape of prickly nanodiamonds: implications for gene delivery

    Science.gov (United States)

    Chu, Zhiqin; Miu, Kaikei; Lung, Pingsai; Zhang, Silu; Zhao, Saisai; Chang, Huan-Cheng; Lin, Ge; Li, Quan

    2015-06-01

    The prickly nanodiamonds easily entered cells via endocytosis followed by unique intracellular translocation characteristics—quick endosomal escape followed by stable residence in cytoplasm. Endosomal membrane rupturing is identified as the major route of nanodiamonds’ escaping the vesicle confinement and to the cytoplasm. Little cytotoxicity is observed to associate with the nanodiamonds’ cytosolic release. Such features enable its application for gene delivery, which requires both effective cellular uptake and cytosolic release of the gene. Taking green fluorescent protein gene as an example, we demonstrate the successful cytosolic delivery and expression of such a gene using the prickly nanodiamonds as carrier.

  11. Rapid endosomal escape of prickly nanodiamonds: implications for gene delivery

    Science.gov (United States)

    Chu, Zhiqin; Miu, Kaikei; Lung, Pingsai; Zhang, Silu; Zhao, Saisai; Chang, Huan-Cheng; Lin, Ge; Li, Quan

    2015-01-01

    The prickly nanodiamonds easily entered cells via endocytosis followed by unique intracellular translocation characteristics—quick endosomal escape followed by stable residence in cytoplasm. Endosomal membrane rupturing is identified as the major route of nanodiamonds’ escaping the vesicle confinement and to the cytoplasm. Little cytotoxicity is observed to associate with the nanodiamonds’ cytosolic release. Such features enable its application for gene delivery, which requires both effective cellular uptake and cytosolic release of the gene. Taking green fluorescent protein gene as an example, we demonstrate the successful cytosolic delivery and expression of such a gene using the prickly nanodiamonds as carrier. PMID:26123532

  12. Rapid endosomal escape of prickly nanodiamonds: implications for gene delivery.

    Science.gov (United States)

    Chu, Zhiqin; Miu, Kaikei; Lung, Pingsai; Zhang, Silu; Zhao, Saisai; Chang, Huan-Cheng; Lin, Ge; Li, Quan

    2015-06-30

    The prickly nanodiamonds easily entered cells via endocytosis followed by unique intracellular translocation characteristics—quick endosomal escape followed by stable residence in cytoplasm. Endosomal membrane rupturing is identified as the major route of nanodiamonds' escaping the vesicle confinement and to the cytoplasm. Little cytotoxicity is observed to associate with the nanodiamonds' cytosolic release. Such features enable its application for gene delivery, which requires both effective cellular uptake and cytosolic release of the gene. Taking green fluorescent protein gene as an example, we demonstrate the successful cytosolic delivery and expression of such a gene using the prickly nanodiamonds as carrier.

  13. An industry update: the latest developments in therapeutic delivery.

    Science.gov (United States)

    Steinbach, Oliver C

    2017-10-01

    The present industry update covers the period 1-30 June 2017, with information sourced from company press releases, regulatory and patent agencies as well as scientific literature. The combination of drug and devices such as improved, safer injectables (see various market reports, companies Adamis and Baxter), patches (Microdermis) and (nano)carriers are moving increasingly from the R&D stage into clinical trials and toward the market. This addresses increased safety and effectiveness requirements, limiting physico-chemical properties of active ingredients, cost-effectiveness and patient comfort through ease of use. Further attention in the market is on local delivery methods (such as intraocular by Icon Bioscience, Glaukos) and the sheer infinite possibilities of nanotechnology such as LDL nanocarriers, microneedles and hydrogel cubes. Another 21st century key technology area is mobile applications (Vital Art and Science) and connected devices (SmartPill, Pop Test Devices) which are increasingly finding their way into the drug delivery field to enable, for example, closed loop monitoring of drug dosing in trials and of patients with their care providers. Not surprisingly companies are increasingly utilizing convergence to combine their diverse capabilities (Vetter Pharma/Microdermis, TXCell/Lentigen Technology).

  14. Gastroretentive drug delivery systems for therapeutic management of peptic ulcer.

    Science.gov (United States)

    Garg, Tarun; Kumar, Animesh; Rath, Goutam; Goyal, Amit K

    2014-01-01

    A peptic ulcer, stomach ulcer, or gastric ulcer, also known as peptic ulcer disease (PUD), is a very common chronic disorder of the stomach which is mainly caused by damage or impairment of the stomach lining. Various factors such as pepsin, gastric acid, H. pylori, NSAIDs, prostaglandins, mucus, bicarbonate, and blood flow to mucosa play an important role in causing peptic ulcers. In this review article, our main focus is on some important gastroretentive drug delivery systems (GRDDS) (floating, bioadhesive, high density, swellable, raft forming, superporous hydrogel, and magnetic systems) which will be helpful in gastroretention of different dosage forms for treatment of peptic ulcer. GRDDS provides a mean for controlled release of compounds that are absorbed by active transport in the upper intestine. It also enables controlled delivery for paracellularly absorbed drugs without a decrease in bioavailability. The above approaches are specific for targeting and leading to a marked improvement in the quality of life for a large number of patients. In the future, it is expected that they will become of growing significance, finally leading to improved efficiencies of various types of pharmacotherapies.

  15. An industry update: the latest developments in therapeutic delivery.

    Science.gov (United States)

    Simpson, Iain

    2018-01-01

    This industry update covers the period from 1 September through 30 September 2017, and is based on information sourced from company press releases, scientific literature, patents and various news websites. The month saw the US FDA approve three new molecular entities, Aliqopa (copanlisib dihydrochloride) (Bayer Healthcare); Solosec (secnidazole) (Symbiomix Therapeutics) and Verzenio (abemaciclib) (Eli Lilly and Co). Intarcia Therapeutics Inc. has its application for approval of a novel drug device combination of exenatide for the treatment of diabetes rejected by FDA but said that it will work to address the concerns and refile the application. The impact of biosimilars in the market is steadily increasing with seven biosimilars approved in the USA and Sandoz hoping to add to this with its announcement that FDA has accepted its Biologics License Application for a biosimilar version of Roche's Rituxan. Circassia announced positive top line results of a respiratory drug, Duaklir (for the treatment of chronic obstructive pulmonary disease) and Sarepta (for its new treatment for Duchenne muscular dystrophy). Axovant Sciences Ltd announced the failure if its drug Intepirdine in the treatment of Alzheimer's, adding to a growing list of drug failures in this area. There were a number of developments in the area of oncology with Bristol-Myers Squibb and Infinity Pharmaceuticals announcing an expansion of their collaboration looking at combination treatments, as well as Eli Lilly and Co's approval for Verzenio. Rani Therapeutics and Intra-Cellular Therapies announced successful funding rounds to support their drug programs. Allergan announced a novel licensing deal for its dry eye drug, Restasis, which it hopes would allow it to stave off patent challenges from several companies looking to develop generic versions of the drug. New research suggests that loss of sense of smell can be linked to an increased risk of developing Parkinson's disease.

  16. Mesenchymal stem cells as therapeutic delivery vehicles targeting tumor stroma

    DEFF Research Database (Denmark)

    Serakinci, Nedime; Christensen, Rikke; Sørensen, Flemming Brandt

    2011-01-01

    The field of stem cell biology continues to evolve by characterization of further types of stem cells and by exploring their therapeutic potential for experimental and clinical applications. Human mesenchymal stem cells (hMSCs) are one of the most promising candidates simply because...... better understanding and in vivo supporting data. The homing ability of hMSCs was investigated by creating a human xenograft model by transplanting an ovarian cancer cell line into immunocompromised mice. Then, genetically engineered hMSC-telo1 cells were injected through the tail vein...

  17. Recent Advancements in Targeted Delivery of Therapeutic Molecules in Neurodegenerative Disease–-Spinocerebellar Ataxia–-Opportunities and Challenges

    Directory of Open Access Journals (Sweden)

    Satya Prakash

    2008-01-01

    Full Text Available Drug discovery and its methodologies have been very effective in terms of treating cancers and immunological disorders but have not been able to stop genetic diseases as most of the drugs target at the protein level. They merely mitigate the symptoms of the disease. Spinocerebellar ataxia is a neurological genetic disorder that is caused by the formation of an abnormal protein. There have been several reports on ataxic drug development but actual clinical treatment is yet to be achieved. Oligonucleotide therapy called sequence specific siRNA mediated gene silencing has evolved with promising results. This approach emphasizes on suppressing the expression of the diseased gene at mRNA level. However, there is a limitation in delivery of siRNA to the target site. Several methods have been developed over the last decade to enhance the target specific delivery of DNA, siRNA, protein and small drug molecules for therapeutic purpose with less or no side effects. This review discusses the latest upcoming technologies in the field that focus on a number of nonviral nanocarriers for targeted delivery. In this review, we explore the promise and potential of novel therapeutics with interest on ataxia therapy.

  18. Therapeutic Ultrasound Enhancement of Drug Delivery to Soft Tissues

    Science.gov (United States)

    Lewis, George; Wang, Peng; Lewis, George; Olbricht, William

    2009-04-01

    Effects of exposure to 1.58 MHz focused ultrasound on transport of Evans Blue Dye (EBD) in soft tissues are investigated when an external pressure gradient is applied to induce convective flow through the tissue. The magnitude of the external pressure gradient is chosen to simulate conditions in brain parenchyma during convection-enhanced drug delivery (CED) to the brain. EBD uptake and transport are measured in equine brain, avian muscle and agarose brain-mimicking phantoms. Results show that ultrasound enhances EBD uptake and transport, and the greatest enhancement occurs when the external pressure gradient is applied. The results suggest that exposure of the brain parenchyma to ultrasound could enhance penetration of material infused into the brain during CED therapy.

  19. In vitro and in vivo delivery of therapeutic proteins using cell penetrating peptides.

    Science.gov (United States)

    Bolhassani, Azam; Jafarzade, Behnaz Sadat; Mardani, Golnaz

    2017-01-01

    The failure of proteins to penetrate mammalian cells or target tumor cells restricts their value as therapeutic tools in a variety of diseases such as cancers. Recently, protein transduction domains (PTDs) or cell penetrating peptides (CPPs) have been shown to promote the delivery of therapeutic proteins or peptides into live cells. The successful delivery of proteins mainly depends on their physicochemical properties. Although, linear cell penetrating peptides are one of the most effective delivery vehicles; but currently, cyclic CPPs has been developed to potently transport bioactive full-length proteins into cells. Up to now, several small protein transduction domains from viral proteins including Tat or VP22 could be fused to other peptides or proteins to entry them in various cell types at a dose-dependent approach. A major disadvantage of PTD-fusion proteins is primary uptake into endosomal vesicles leading to inefficient release of the fusion proteins into the cytosol. Recently, non-covalent complex formation (Chariot) between proteins and CPPs has attracted a special interest to overcome some delivery limitations (e.g., toxicity). Many preclinical and clinical trials of CPP-based delivery are currently under evaluation. Generally, development of more efficient protein transduction domains would significantly increase the potency of protein therapeutics. Moreover, the synergistic or combined effects of CPPs with other delivery systems for protein/peptide drug delivery would promote their therapeutic effects in cancer and other diseases. In this review, we will describe the functions and implications of CPPs for delivering the therapeutic proteins or peptides in preclinical and clinical studies. Copyright © 2016 Elsevier Inc. All rights reserved.

  20. Current and future technological advances in transdermal gene delivery.

    Science.gov (United States)

    Chen, Xianfeng

    2017-12-19

    Transdermal gene delivery holds significant advantages as it is able to minimize the problems of systemic administration such as enzymatic degradation, systemic toxicity, and poor delivery to target tissues. This technology has the potential to transform the treatment and prevention of a range of diseases. However, the skin poses a great barrier for gene delivery because of the "bricks-and-mortar" structure of the stratum corneum and the tight junctions between keratinocytes in the epidermis. This review systematically summarizes the typical physical and chemical approaches to overcome these barriers and facilitate gene delivery via skin for applications in vaccination, wound healing, skin cancers and skin diseases. Next, the advantages and disadvantages of different approaches are discussed and the insights for future development are provided. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Functionalized nanoparticles for AMF-induced gene and drug delivery

    Science.gov (United States)

    Biswas, Souvik

    The properties and broad applications of nano-magnetic colloids have generated much interest in recent years. Specially, Fe3O4 nanoparticles have attracted a great deal of attention since their magnetic properties can be used for hyperthermia treatment or drug targeting. For example, enhanced levels of intracellular gene delivery can be achieved using Fe3O4 nano-vectors in the presence of an external magnetic field, a process known as 'magnetofection'. The low cytotoxicity, tunable particle size, ease of surface functionalization, and ability to generate thermal energy using an external alternating magnetic field (AMF) are properties have propelled Fe3O4 research to the forefront of nanoparticle research. The strategy of nanoparticle-mediated, AMF-induced heat generation has been used to effect intracellular hyperthermia. One application of this 'magnetic hyperthermia' is heat activated local delivery of a therapeutic effector (e.g.; drug or polynucleotide). This thesis describes the development of a magnetic nano-vector for AMF-induced, heat-activated pDNA and small molecule delivery. The use of heat-inducible vectors, such as heat shock protein ( hsp) genes, is a promising mode of gene therapy that would restrict gene expression to a local region by focusing a heat stimulus only at a target region. We thus aimed to design an Fe3O4 nanoparticle-mediated gene transfer vehicle for AMF-induced localized gene expression. We opted to use 'click' oximation techniques to assemble the magnetic gene transfer vector. Chapter 2 describes the synthesis, characterization, and transfection studies of the oxime ether lipid-based nano-magnetic vectors MLP and dMLP. The synthesis and characterization of a novel series of quaternary ammonium aminooxy reagents (2.1--2.4) is described. These cationic aminooxy compounds were loaded onto nanoparticles for ligation with carbonyl groups and also to impart a net positive charge on the nanoparticle surface. Our studies indicated that the

  2. Multirate delivery of multiple therapeutic agents from metal-organic frameworks

    Directory of Open Access Journals (Sweden)

    Alistair C. McKinlay

    2014-12-01

    Full Text Available The highly porous nature of metal-organic frameworks (MOFs offers great potential for the delivery of therapeutic agents. Here, we show that highly porous metal-organic frameworks can be used to deliver multiple therapeutic agents—a biologically active gas, an antibiotic drug molecule, and an active metal ion—simultaneously but at different rates. The possibilities offered by delivery of multiple agents with different mechanisms of action and, in particular, variable timescales may allow new therapy approaches. Here, we show that the loaded MOFs are highly active against various strains of bacteria.

  3. Recent Progress in Functional Micellar Carriers with Intrinsic Therapeutic Activities for Anticancer Drug Delivery.

    Science.gov (United States)

    Qu, Ying; Chu, BingYang; Shi, Kun; Peng, JinRong; Qian, ZhiYong

    2017-12-01

    Polymeric micelles have presented superior delivery properties for poorly water-soluble chemotherapeutic agents. However, it remains discouraging that there may be some additional short or long-term toxicities caused by the metabolites of high quantities of carriers. If carriers had simultaneous therapeutic effects with the drug, these issues would not be a concern. For this, carriers not only simply act as drug carriers, but also exert an intrinsic therapeutic effect as a therapeutic agent. The functional micellar carriers would be beneficial to maximize the anticancer effect, overcome the drug resistance and reduce the systemic toxicity. In this review, we aim to summarize the recent progress on the development of functional micellar carriers with intrinsic anticancer activities for the delivery of anticancer drugs. This review focuses on the design strategies, properties of carriers and the drug loading behavior. In addition, the combinational therapeutic effects between carriers and chemotherapeutic agents are also discussed.

  4. Gold nanoparticle trapping and delivery for therapeutic applications

    Directory of Open Access Journals (Sweden)

    Aziz MS

    2011-12-01

    Full Text Available MS Aziz1, Nathaporn Suwanpayak3,4, Muhammad Arif Jalil2, R Jomtarak4, T Saktioto2, Jalil Ali1, PP Yupapin41Institute of Advanced Photonics Science, 2Ibnu Sina Institute of Fundamental Science Studies, Nanotechnology Research Alliance, Universiti Teknologi Malaysia, Johor Bahru, Malaysia; 3King Mongkut's Institute of Technology Ladkrabang, Chump on Campus, Chumphon, 4Nanoscale Science and Engineering Research Alliance (N'SERA, Faculty of Science, King Mongkut's Institute of Technology Ladkrabang, Bangkok, ThailandAbstract: A new optical trapping design to transport gold nanoparticles using a PANDA ring resonator system is proposed. Intense optical fields in the form of dark solitons controlled by Gaussian pulses are used to trap and transport nanoscopic volumes of matter to the desired destination via an optical waveguide. Theoretically, the gradient and scattering forces are responsible for this trapping phenomenon, where in practice such systems can be fabricated and a thin-film device formed on the specific artificial medical materials, for instance, an artificial bone. The dynamic behavior of the tweezers can be tuned by controlling the optical pulse input power and parameters of the ring resonator system. Different trap sizes can be generated to trap different gold nanoparticles sizes, which is useful for gold nanoparticle therapy. In this paper, we have shown the utility of gold nanoparticle trapping and delivery for therapy, which may be useful for cosmetic therapy and related applications.Keywords: gold nanoparticle trapping, particle trapping, therapy, transport

  5. Recent advances in dendrimer-based nanovectors for tumor-targeted drug and gene delivery

    Science.gov (United States)

    Kesharwani, Prashant; Iyer, Arun K.

    2015-01-01

    Advances in the application of nanotechnology in medicine have given rise to multifunctional smart nanocarriers that can be engineered with tunable physicochemical characteristics to deliver one or more therapeutic agent(s) safely and selectively to cancer cells, including intracellular organelle-specific targeting. Dendrimers having properties resembling biomolecules, with well-defined 3D nanopolymeric architectures, are emerging as a highly attractive class of drug and gene delivery vector. The presence of numerous peripheral functional groups on hyperbranched dendrimers affords efficient conjugation of targeting ligands and biomarkers that can recognize and bind to receptors overexpressed on cancer cells for tumor-cell-specific delivery. The present review compiles the recent advances in dendrimer-mediated drug and gene delivery to tumors by passive and active targeting principles with illustrative examples. PMID:25555748

  6. Systemic gene delivery to the central nervous system using Adeno-associated virus

    Directory of Open Access Journals (Sweden)

    Mathieu eBOURDENX

    2014-06-01

    Full Text Available Adeno-associated virus (AAV-mediated gene delivery has emerged as an effective and safe tool for both preclinical and clinical studies of neurological disorders. The recent discovery that several serotypes are able to cross the blood-brain-barrier when administered systemically has been a real breakthrough in the field of neurodegenerative diseases. Widespread transgene expression after systemic injection could spark interest as a therapeutic approach. Such strategy will avoid invasive brain surgery and allow non-focal gene therapy promising for CNS diseases affecting large portion of the brain. Here, we will review the recent results achieved through different systemic routes of injection generated in the last decade using systemic AAV-mediated delivery and propose a brief assessment of their values. In particular, we emphasize how the methods used for virus engineering could improve brain transduction after peripheral delivery.

  7. Biopolymer-Based Nanoparticles for Drug/Gene Delivery and Tissue Engineering

    Science.gov (United States)

    Nitta, Sachiko Kaihara; Numata, Keiji

    2013-01-01

    There has been a great interest in application of nanoparticles as biomaterials for delivery of therapeutic molecules such as drugs and genes, and for tissue engineering. In particular, biopolymers are suitable materials as nanoparticles for clinical application due to their versatile traits, including biocompatibility, biodegradability and low immunogenicity. Biopolymers are polymers that are produced from living organisms, which are classified in three groups: polysaccharides, proteins and nucleic acids. It is important to control particle size, charge, morphology of surface and release rate of loaded molecules to use biopolymer-based nanoparticles as drug/gene delivery carriers. To obtain a nano-carrier for therapeutic purposes, a variety of materials and preparation process has been attempted. This review focuses on fabrication of biocompatible nanoparticles consisting of biopolymers such as protein (silk, collagen, gelatin, β-casein, zein and albumin), protein-mimicked polypeptides and polysaccharides (chitosan, alginate, pullulan, starch and heparin). The effects of the nature of the materials and the fabrication process on the characteristics of the nanoparticles are described. In addition, their application as delivery carriers of therapeutic drugs and genes and biomaterials for tissue engineering are also reviewed. PMID:23344060

  8. Biopolymer-Based Nanoparticles for Drug/Gene Delivery and Tissue Engineering

    Directory of Open Access Journals (Sweden)

    Keiji Numata

    2013-01-01

    Full Text Available There has been a great interest in application of nanoparticles as biomaterials for delivery of therapeutic molecules such as drugs and genes, and for tissue engineering. In particular, biopolymers are suitable materials as nanoparticles for clinical application due to their versatile traits, including biocompatibility, biodegradability and low immunogenicity. Biopolymers are polymers that are produced from living organisms, which are classified in three groups: polysaccharides, proteins and nucleic acids. It is important to control particle size, charge, morphology of surface and release rate of loaded molecules to use biopolymer-based nanoparticles as drug/gene delivery carriers. To obtain a nano-carrier for therapeutic purposes, a variety of materials and preparation process has been attempted. This review focuses on fabrication of biocompatible nanoparticles consisting of biopolymers such as protein (silk, collagen, gelatin, β-casein, zein and albumin, protein-mimicked polypeptides and polysaccharides (chitosan, alginate, pullulan, starch and heparin. The effects of the nature of the materials and the fabrication process on the characteristics of the nanoparticles are described. In addition, their application as delivery carriers of therapeutic drugs and genes and biomaterials for tissue engineering are also reviewed.

  9. Toward a new generation of therapeutics: artificial cell targeted delivery of live cells for therapy.

    Science.gov (United States)

    Prakash, Satya; Martoni, Christopher

    2006-01-01

    Scientific evidence in the prevention and treatment of various disorders is accumulating regarding probiotics. The health benefits supported by adequate clinical data include increased resistance to infectious disease, decreased duration of diarrhea, management of inflammatory bowel disease, reduction of serum cholesterol, prevention of allergy, modulation of cytokine gene expression, and suppression of carcinogen production. Recent ventures in metabolic engineering and heterologous protein expression have enhanced the enzymatic and immunomodulatory effects of probiotics and, with time, may allow more active intervention among critical care patients. In addition, a number of approaches are currently being explored, including the physical and chemical protection of cells, to increase probiotic viability and its health benefits. Traditional immobilization of probiotics in gel matrices, most notably calcium alginate and kappa-carrageenan, has frequently been employed, with noted improvements in viability during freezing and storage. Conflicting reports exist, however, on the protection offered by immobilization from harsh physiologic environments. An alternative approach, microencapsulation in "artificial cells," builds on immobilization technologies by combining enhanced mechanical stability of the capsule membrane with improved mass transport, increased cell loading, and greater control of parameters. This review summarizes the current clinical status of probiotics, examines the promises and challenges of current immobilization technologies, and presents the concept of artificial cells for effective delivery of therapeutic bacterial cells.

  10. Gene Delivery for Metastatic Prostate Cancer Cells

    National Research Council Canada - National Science Library

    Pang, Shen

    2001-01-01

    .... Enhanced by the bystander effect, the specific expression of the DTA gene causes significant cell death in prostate cancer cell cultures, with very low background cell eradication in control cell lines...

  11. PLGA-Chitosan nanoparticle-mediated gene delivery for oral cancer treatment: A brief review

    Science.gov (United States)

    Bakar, L. M.; Abdullah, M. Z.; Doolaanea, A. A.; Ichwan, S. J. A.

    2017-08-01

    Cancer becomes a serious issue on society with increasing of their growth and proliferation, either in well economic developed countries or not. Recent years, oral cancer is one of the most threatening diseases impairing the quality of life of the patient. Scientists have emphasised on application of gene therapy for oral cancer by using nanoparticle as transportation vectors as a new alternative platform in order to overcome the limitations of conventional approaches. In modern medicine, nanotechnologies’ application, such as nanoparticles-mediated gene delivery, is one of promising tool for therapeutic devices. The objective of this article is to present a brief review summarizes on the current progress of nanotechnology-based gene delivery treatment system targeted for oral cancer.

  12. Engineered nonviral nanocarriers for intracellular gene delivery applications

    International Nuclear Information System (INIS)

    Ojea-Jiménez, Isaac; Puntes, Victor F; Tort, Olivia; Lorenzo, Julia

    2012-01-01

    The efficient delivery of nucleic acids into mammalian cells is a central aspect of cell biology and of medical applications, including cancer therapy and tissue engineering. Non-viral chemical methods have been received with great interest for transfecting cells. However, further development of nanocarriers that are biocompatible, efficient and suitable for clinical applications is still required. In this paper, the different material platforms for gene delivery are comparatively addressed, and the mechanisms of interaction with biological systems are discussed carefully. (paper)

  13. Electrospun Composites of Polycaprolactone and Porous Silicon Nanoparticles for the Tunable Delivery of Small Therapeutic Molecules

    Directory of Open Access Journals (Sweden)

    Steven J. P. McInnes

    2018-03-01

    Full Text Available This report describes the use of an electrospun composite of poly(ε-caprolactone (PCL fibers and porous silicon (pSi nanoparticles (NPs as an effective system for the tunable delivery of camptothecin (CPT, a small therapeutic molecule. Both materials are biodegradable, abundant, low-cost, and most importantly, have no known cytotoxic effects. The composites were treated with and without sodium hydroxide (NaOH to investigate the wettability of the porous network for drug release and cell viability measurements. CPT release and subsequent cell viability was also investigated. We observed that the cell death rate was not only affected by the addition of our CPT carrier, pSi, but also by increasing the rate of dissolution via treatment with NaOH. This is the first example of loading pSi NPs as a therapeutics nanocarrier into electronspun PCL fibers and this system opens up new possibilities for the delivery of molecular therapeutics.

  14. Advances in sarcoma gene mutations and therapeutic targets.

    Science.gov (United States)

    Gao, Peng; Seebacher, Nicole A; Hornicek, Francis; Guo, Zheng; Duan, Zhenfeng

    2018-01-01

    Sarcomas are rare and complex malignancies that have been associated with a poor prognostic outcome. Over the last few decades, traditional treatment with surgery and/or chemotherapy has not significantly improved outcomes for most types of sarcomas. In recent years, there have been significant advances in the understanding of specific gene mutations that are important in driving the pathogenesis and progression of sarcomas. Identification of these new gene mutations, using next-generation sequencing and advanced molecular techniques, has revealed a range of potential therapeutic targets. This, in turn, may lead to the development of novel agents targeted to different sarcoma subtypes. In this review, we highlight the advances made in identifying sarcoma gene mutations, including those of p53, RB, PI3K and IDH genes, as well as novel therapeutic strategies aimed at utilizing these mutant genes. In addition, we discuss a number of preclinical studies and ongoing early clinical trials in sarcoma targeting therapies, as well as gene editing technology, which may provide a better choice for sarcoma patient management. Published by Elsevier Ltd.

  15. Effect of adrenomedullin gene delivery on insulin resistance in type 2 diabetic rats

    Directory of Open Access Journals (Sweden)

    Hoda Y. Henein

    2011-01-01

    Full Text Available Type 2 diabetes mellitus is one of the common metabolic disorders that ultimately afflicts large number of individuals. Adrenomedullin (AM is a potent vasodilator peptide; previous studies reported development of insulin resistance in aged AM deficient mice. In this study, we employed a gene delivery approach to explore its potential role in insulin resistance. Four groups were included: control, diabetic, non-diabetic injected with the AM gene and diabetic injected with the AM gene. One week following gene delivery, serum glucose, insulin, triglycerides, leptin, adiponectin and corticosterone were measured as well as the insulin resistance index (HOMA-IR. Soleus muscle glucose uptake and RT-PCR of both AM and glucose transporter-4 (GLUT 4 gene expressions were assessed. A single tail vein injection of adrenomedullin gene in type 2 diabetic rats improved skeletal muscle insulin responsiveness with significant improvement of soleus muscle glucose uptake, HOMA-IR, serum glucose, insulin and triglycerides and significant increase in muscle GLUT 4 gene expression (P < 0.05 compared with the non-injected diabetic rats. The beneficial effects of AM gene delivery were accompanied by a significant increase in the serum level of adiponectin (2.95 ± 0.09 versus 2.33 ± 0.17 μg/ml in the non-injected diabetic group as well as a significant decrease in leptin and corticosterone levels (7.51 ± 0.51 and 262.88 ± 10.34 versus 10.63 ± 1.4 and 275.86 ± 11.19 ng/ml respectively in the non-injected diabetic group. The conclusion of the study is that AM gene delivery can improve insulin resistance and may have significant therapeutic applications in type 2 diabetes mellitus.

  16. Recent trends in the transdermal delivery of therapeutic agents used for the management of neurodegenerative diseases.

    Science.gov (United States)

    Ita, Kevin

    2017-06-01

    With the increasing proportion of the global geriatric population, it becomes obvious that neurodegenerative diseases will become more widespread. From an epidemiological standpoint, it is necessary to develop new therapeutic agents for the management of Alzheimer's disease, Parkinson's disease, multiple sclerosis and other neurodegenerative disorders. An important approach in this regard involves the use of the transdermal route. With transdermal drug delivery systems (TDDS), it is possible to modulate the pharmacokinetic profiles of these medications and improve patient compliance. Transdermal drug delivery has also been shown to be useful for drugs with short half-life and low or unpredictable bioavailability. In this review, several transdermal drug delivery enhancement technologies are being discussed in relation to the delivery of medications used for the management of neurodegenerative disorders.

  17. Targeted delivery of genes to endothelial cells and cell- and gene-based therapy in pulmonary vascular diseases.

    Science.gov (United States)

    Suen, Colin M; Mei, Shirley H J; Kugathasan, Lakshmi; Stewart, Duncan J

    2013-10-01

    Pulmonary arterial hypertension (PAH) is a devastating disease that, despite significant advances in medical therapies over the last several decades, continues to have an extremely poor prognosis. Gene therapy is a method to deliver therapeutic genes to replace defective or mutant genes or supplement existing cellular processes to modify disease. Over the last few decades, several viral and nonviral methods of gene therapy have been developed for preclinical PAH studies with varying degrees of efficacy. However, these gene delivery methods face challenges of immunogenicity, low transduction rates, and nonspecific targeting which have limited their translation to clinical studies. More recently, the emergence of regenerative approaches using stem and progenitor cells such as endothelial progenitor cells (EPCs) and mesenchymal stem cells (MSCs) have offered a new approach to gene therapy. Cell-based gene therapy is an approach that augments the therapeutic potential of EPCs and MSCs and may deliver on the promise of reversal of established PAH. These new regenerative approaches have shown tremendous potential in preclinical studies; however, large, rigorously designed clinical studies will be necessary to evaluate clinical efficacy and safety. © 2013 American Physiological Society. Compr Physiol 3:1749-1779, 2013.

  18. Delivery systems and local administration routes for therapeutic siRNA.

    Science.gov (United States)

    Vicentini, Fabiana Testa Moura de Carvalho; Borgheti-Cardoso, Lívia Neves; Depieri, Lívia Vieira; de Macedo Mano, Danielle; Abelha, Thais Fedatto; Petrilli, Raquel; Bentley, Maria Vitória Lopes Badra

    2013-04-01

    With the increasing number of studies proposing new and optimal delivery strategies for the efficacious silencing of gene-related diseases by the local administration of siRNAs, the present review aims to provide a broad overview of the most important and latest developments of non-viral siRNA delivery systems for local administration. Moreover, the main disease targets for the local delivery of siRNA to specific tissues or organs, including the skin, the lung, the eye, the nervous system, the digestive system and the vagina, were explored.

  19. Ex vivo culture of patient tissue & examination of gene delivery.

    LENUS (Irish Health Repository)

    Rajendran, Simon

    2012-01-31

    This video describes the use of patient tissue as an ex vivo model for the study of gene delivery. Fresh patient tissue obtained at the time of surgery is sliced and maintained in culture. The ex vivo model system allows for the physical delivery of genes into intact patient tissue and gene expression is analysed by bioluminescence imaging using the IVIS detection system. The bioluminescent detection system demonstrates rapid and accurate quantification of gene expression within individual slices without the need for tissue sacrifice. This slice tissue culture system may be used in a variety of tissue types including normal and malignant tissue and allows us to study the effects of the heterogeneous nature of intact tissue and the high degree of variability between individual patients. This model system could be used in certain situations as an alternative to animal models and as a complementary preclinical mode prior to entering clinical trial.

  20. Pancreatic Cancer Gene Therapy: From Molecular Targets to Delivery Systems

    Energy Technology Data Exchange (ETDEWEB)

    Fillat, Cristina, E-mail: cristina.fillat@crg.es; Jose, Anabel; Ros, Xavier Bofill-De; Mato-Berciano, Ana; Maliandi, Maria Victoria; Sobrevals, Luciano [Programa Gens i Malaltia, Centre de Regulació Genòmica-CRG, UPF, Parc de Recerca Biomedica de Barcelona-PRBB and Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER), Barcelona (Spain)

    2011-01-18

    The continuous identification of molecular changes deregulating critical pathways in pancreatic tumor cells provides us with a large number of novel candidates to engineer gene-targeted approaches for pancreatic cancer treatment. Targets—both protein coding and non-coding—are being exploited in gene therapy to influence the deregulated pathways to facilitate cytotoxicity, enhance the immune response or sensitize to current treatments. Delivery vehicles based on viral or non-viral systems as well as cellular vectors with tumor homing characteristics are a critical part of the design of gene therapy strategies. The different behavior of tumoral versus non-tumoral cells inspires vector engineering with the generation of tumor selective products that can prevent potential toxic-associated effects. In the current review, a detailed analysis of the different targets, the delivery vectors, the preclinical approaches and a descriptive update on the conducted clinical trials are presented. Moreover, future possibilities in pancreatic cancer treatment by gene therapy strategies are discussed.

  1. Intrathecal delivery of protein therapeutics to the brain: a critical reassessment.

    Science.gov (United States)

    Calias, Pericles; Banks, William A; Begley, David; Scarpa, Maurizio; Dickson, Patricia

    2014-11-01

    Disorders of the central nervous system (CNS), including stroke, neurodegenerative diseases, and brain tumors, are the world's leading causes of disability. Delivery of drugs to the CNS is complicated by the blood-brain barriers that protect the brain from the unregulated leakage and entry of substances, including proteins, from the blood. Yet proteins represent one of the most promising classes of therapeutics for the treatment of CNS diseases. Many strategies for overcoming these obstacles are in development, but the relatively straightforward approach of bypassing these barriers through direct intrathecal administration has been largely overlooked. Originally discounted because of its lack of usefulness for delivering small, lipid-soluble drugs to the brain, the intrathecal route has emerged as a useful, in some cases perhaps the ideal, route of administration for certain therapeutic protein and targeted disease combinations. Here, we review blood-brain barrier functions and cerebrospinal fluid dynamics and their relevance to drug delivery via the intrathecal route, discuss animal and human studies that have investigated intrathecal delivery of protein therapeutics, and outline several characteristics of protein therapeutics that can allow them to be successfully delivered intrathecally. Copyright © 2014 Elsevier Inc. All rights reserved.

  2. Physical non-viral gene delivery methods for tissue engineering

    Science.gov (United States)

    Mellott, Adam J.; Forrest, M. Laird; Detamore, Michael S.

    2016-01-01

    The integration of gene therapy into tissue engineering to control differentiation and direct tissue formation is not a new concept; however, successful delivery of nucleic acids into primary cells, progenitor cells, and stem cells has proven exceptionally challenging. Viral vectors are generally highly effective at delivering nucleic acids to a variety of cell populations, both dividing and non-dividing, yet these viral vectors are marred by significant safety concerns. Non-viral vectors are preferred for gene therapy, despite lower transfection efficiencies, and possess many customizable attributes that are desirable for tissue engineering applications. However, there is no single non-viral gene delivery strategy that “fits-all” cell types and tissues. Thus, there is a compelling opportunity to examine different non-viral vectors, especially physical vectors, and compare their relative degrees of success. This review examines the advantages and disadvantages of physical non-viral methods (i.e., microinjection, ballistic gene delivery, electroporation, sonoporation, laser irradiation, magnetofection, and electric field-induced molecular vibration), with particular attention given to electroporation because of its versatility, with further special emphasis on Nucleofection™. In addition, attributes of cellular character that can be used to improve differentiation strategies are examined for tissue engineering applications. Ultimately, electroporation exhibits a high transfection efficiency in many cell types, which is highly desirable for tissue engineering applications, but electroporation and other physical non-viral gene delivery methods are still limited by poor cell viability. Overcoming the challenge of poor cell viability in highly efficient physical non-viral techniques is the key to using gene delivery to enhance tissue engineering applications. PMID:23099792

  3. Physical non-viral gene delivery methods for tissue engineering.

    Science.gov (United States)

    Mellott, Adam J; Forrest, M Laird; Detamore, Michael S

    2013-03-01

    The integration of gene therapy into tissue engineering to control differentiation and direct tissue formation is not a new concept; however, successful delivery of nucleic acids into primary cells, progenitor cells, and stem cells has proven exceptionally challenging. Viral vectors are generally highly effective at delivering nucleic acids to a variety of cell populations, both dividing and non-dividing, yet these viral vectors are marred by significant safety concerns. Non-viral vectors are preferred for gene therapy, despite lower transfection efficiencies, and possess many customizable attributes that are desirable for tissue engineering applications. However, there is no single non-viral gene delivery strategy that "fits-all" cell types and tissues. Thus, there is a compelling opportunity to examine different non-viral vectors, especially physical vectors, and compare their relative degrees of success. This review examines the advantages and disadvantages of physical non-viral methods (i.e., microinjection, ballistic gene delivery, electroporation, sonoporation, laser irradiation, magnetofection, and electric field-induced molecular vibration), with particular attention given to electroporation because of its versatility, with further special emphasis on Nucleofection™. In addition, attributes of cellular character that can be used to improve differentiation strategies are examined for tissue engineering applications. Ultimately, electroporation exhibits a high transfection efficiency in many cell types, which is highly desirable for tissue engineering applications, but electroporation and other physical non-viral gene delivery methods are still limited by poor cell viability. Overcoming the challenge of poor cell viability in highly efficient physical non-viral techniques is the key to using gene delivery to enhance tissue engineering applications.

  4. Self-Assembly of pH-Responsive Microspheres for Intestinal Delivery of Diverse Lipophilic Therapeutics.

    Science.gov (United States)

    Zhou, Xing; Zhao, Yang; Chen, Siyu; Han, Songling; Xu, Xiaoqiu; Guo, Jiawei; Liu, Mengyu; Che, Ling; Li, Xiaohui; Zhang, Jianxiang

    2016-08-08

    Targeted delivery of therapeutics to the intestine is preferred for the management of many diseases due to its diverse advantages. Currently, there are still challenges in creating cost-effective and translational pH-responsive microspheres for intestinal delivery of various hydrophobic drugs. Herein we report a multiple noncovalent interactions-mediated assembly strategy in which carboxyl-bearing compounds (CBCs) are guest molecules, while poly(N-isopropylacrylamide) (PNIPAm) serves as a host polymer. Formation of microparticles and therapeutic packaging can be achieved simultaneously by this assembly approach, leading to well-shaped microspheres with extremely higher drug loading capacity as compared to microspheres based on two FDA-approved materials of poly(d,l-lactide-co-glycolide) (PLGA) and an enteric coating polymer EudragitS 100 (S100). Also, carboxyl-deficient hydrophobic drugs can be effectively entrapped. These assembled microspheres, with excellent reconstitution capability as well as desirable scalability, could selectively release drug molecules under intestinal conditions. By significantly enhancing drug dissolution/release in the intestine, these pH-responsive assemblies may notably improve the oral bioavailability of loaded therapeutics. Moreover, the assembled microspheres possessed superior therapeutic performance in rodent models of inflammation and tumor over the control microspheres derived from PLGA and S100. Therapy with newly developed microspheres did not cause undesirable side effects. Furthermore, in vivo evaluation in mice revealed the carrier material PNIPAm was safe for oral delivery at doses as high as 10 g/kg. Collectively, our findings demonstrated that this type of pH-responsive microsphere may function as superior and translational intestine-directed delivery systems for a diverse array of therapeutics.

  5. Neuropathic Pain and Lung Delivery of Nanoparticulate Drugs: An Emerging Novel Therapeutic Strategy.

    Science.gov (United States)

    Islam, Nazrul; Abbas, Muzaffar; Rahman, Shafiqur

    2017-01-01

    Neuropathic pain is a chronic neurological disorder affecting millions of people around the world. The currently available pharmacologic agents for the treatment of neuropathic pain have limited efficacy and are associated with dose related unwanted adverse effects. Due to the limited access of drug molecules across blood-brain barrier, a small percentage of drug that is administered systematically, reaches the central nervous system in active form. These therapeutic agents also require daily treatment regimen that is inconvenient and potentially impact patient compliance. Application of nanoparticulate drugs for enhanced delivery system has been explored extensively in the last decades. Pulmonary delivery of nanomedicines for the management of various diseases has become an emerging treatment strategy that ensures the targeted delivery of drugs both for systemic and local effects with low dose and limited adverse effects. To the best of our knowledge, there are no inhaled drug products available on market for the treatment of neuropathic pain. The advantages of delivering therapeutics into deep lungs include non-invasive drug delivery, higher bioavailability with low dose, lower systemic toxicity, and potentially greater blood-brain barrier penetration. This review discusses and highlights the important issues on the application of emerging nanoparticulate lung delivery of drugs for the effective treatment of neuropathic pain. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  6. Enhanced Delivery of Gold Nanoparticles with Therapeutic Potential for Targeting Human Brain Tumors

    Science.gov (United States)

    Etame, Arnold B.

    The blood brain barrier (BBB) remains a major challenge to the advancement and application of systemic anti-cancer therapeutics into the central nervous system. The structural and physiological delivery constraints of the BBB significantly limit the effectiveness of conventional chemotherapy, thereby making systemic administration a non-viable option for the vast majority of chemotherapy agents. Furthermore, the lack of specificity of conventional systemic chemotherapy when applied towards malignant brain tumors remains a major shortcoming. Hence novel therapeutic strategies that focus both on targeted and enhanced delivery across the BBB are warranted. In recent years nanoparticles (NPs) have emerged as attractive vehicles for efficient delivery of targeted anti-cancer therapeutics. In particular, gold nanoparticles (AuNPs) have gained prominence in several targeting applications involving systemic cancers. Their enhanced permeation and retention within permissive tumor microvasculature provide a selective advantage for targeting. Malignant brain tumors also exhibit transport-permissive microvasculature secondary to blood brain barrier disruption. Hence AuNPs may have potential relevance for brain tumor targeting. However, the permeation of AuNPs across the BBB has not been well characterized, and hence is a potential limitation for successful application of AuNP-based therapeutics within the central nervous system (CNS). In this dissertation, we designed and characterized AuNPs and assessed the role of polyethylene glycol (PEG) on the physical and biological properties of AuNPs. We established a size-dependent permeation profile with respect to core size as well as PEG length when AuNPs were assessed through a transport-permissive in-vitro BBB. This study was the first of its kind to systematically examine the influence of design on permeation of AuNPs through transport-permissive BBB. Given the significant delivery limitations through the non

  7. Non-viral gene delivery strategies for cancer therapy, tissue engineering and regenerative medicine

    Science.gov (United States)

    Bhise, Nupura S.

    Gene therapy involves the delivery of deoxyribonucleic acid (DNA) into cells to override or replace a malfunctioning gene for treating debilitating genetic diseases, including cancer and neurodegenerative diseases. In addition to its use as a therapeutic, it can also serve as a technology to enable regenerative medicine strategies. The central challenge of the gene therapy research arena is developing a safe and effective delivery agent. Since viral vectors have critical immunogenic and tumorogenic safety issues that limit their clinical use, recent efforts have focused on developing non-viral biomaterial based delivery vectors. Cationic polymers are an attractive class of gene delivery vectors due to their structural versatility, ease of synthesis, biodegradability, ability to self-complex into nanoparticles with negatively charged DNA, capacity to carry large cargo, cellular uptake and endosomal escape capacity. In this thesis, we hypothesized that developing a biomaterial library of poly(betaamino esters) (PBAE), a newer class of cationic polymers consisting of biodegradable ester groups, would allow investigating vector design parameters and formulating effective non-viral gene delivery strategies for cancer drug delivery, tissue engineering and stem cell engineering. Consequently, a high-throughput transfection assay was developed to screen the PBAE-based nanoparticles in hard to transfect fibroblast cell lines. To gain mechanistic insights into the nanoparticle formulation process, biophysical properties of the vectors were characterized in terms of molecular weight (MW), nanoparticle size, zeta potential and plasmid per particle count. We report a novel assay developed for quantifying the plasmid per nanoparticle count and studying its implications for co-delivery of multiple genes. The MW of the polymers ranged from 10 kDa to 100 kDa, nanoparticle size was about 150 run, zeta potential was about 30 mV in sodium acetate buffer (25 mM, pH 5) and 30 to 100

  8. AAV vectors as gene delivery vehicles in the central nervous system

    NARCIS (Netherlands)

    Broekman, M.L.D.

    2006-01-01

    Recombinant gene delivery vehicles based on the replication-defective AAV have gained a preeminent position in the field of gene delivery to the brain. Efficient global gene delivery to the CNS is beneficial for the study of gene products is the entire CNS as well as for introducing and expressing

  9. Protein kinase A inhibition modulates the intracellular routing of gene delivery vehicles in HeLa cells, leading to productive transfection

    NARCIS (Netherlands)

    Rehman, Zia Ur; Hoekstra, Dick; Zuhorn, Inge S.

    2011-01-01

    Cellular entry of nanoparticles for drug- and gene delivery relies on various endocytic pathways, including clathrin-and caveolae-mediated endocytosis. To improve delivery, i.e., the therapeutic and/or cell biological impact, current efforts are aimed at avoiding processing of the carriers along the

  10. Conceptual and technical aspects of transfection and gene delivery.

    Science.gov (United States)

    Kaestner, Lars; Scholz, Anke; Lipp, Peter

    2015-03-15

    Genetically modified animals are state of the art in biomedical research as gene therapy is a promising perspective in the attempt to cure hereditary diseases. Both approaches have in common that modified or corrected genetic information must be transferred into cells in general or into particular cell types of an organism. Here we give an overview of established and emerging methods of transfection and gene delivery and provide conceptual and technical advantages and drawbacks of their particular use. Additionally, based on a flow chart, we compiled a rough guideline to choose a gene transfer method for a particular field of application. Copyright © 2015 Elsevier Ltd. All rights reserved.

  11. Application of Ferriferous Oxide Modified by Chitosan in Gene Delivery

    Directory of Open Access Journals (Sweden)

    Yu Kuang

    2012-01-01

    Full Text Available New approaches to improve the traditional gene carriers are still required. Here we explore Fe3O4 modified with degradable polymers that enhances gene delivery and target delivery using permanent magnetic field. Two magnetic Fe3O4 nanoparticles coated with chitosan (CTS and polyethylene glycol (PEG were synthesized by means of controlled chemical coprecipitation. Plasmid pEGFP was encapsulated as a reported gene. The ferriferous oxide complexes were approximately spherical; surface charge of CTS-Fe3O4 and PEG-Fe3O4 was about 20 mv and 0 mv, respectively. The controlled release of DNA from the CTS-Fe3O4 nanoparticles was observed. Concurrently, a desired Fe3O4 concentration of less than 2 mM was verified as safe by means of a cytotoxicity test in vitro. Presence of the permanent magnetic field significantly increased the transfection efficiency. Furthermore, the passive target property and safety of magnetic nanoparticles were also demonstrated in an in vivo test. The novel gene delivery system was proved to be an effective tool required for future target expression and gene therapy in vivo.

  12. Recent Trends in Nanotechnology-Based Drugs and Formulations for Targeted Therapeutic Delivery.

    Science.gov (United States)

    Iqbal, Hafiz M N; Rodriguez, Angel M V; Khandia, Rekha; Munjal, Ashok; Dhama, Kuldeep

    2017-01-01

    In the recent past, a wider spectrum of nanotechnologybased drugs or drug-loaded devices and systems has been engineered and investigated with high interests. The key objective is to help for an enhanced/better quality of patient life in a secure way by avoiding/limiting drug abuse, or severe adverse effects of some in practice traditional therapies. Various methodological approaches including in vitro, in vivo, and ex vivo techniques have been exploited, so far. Among them, nanoparticles-based therapeutic agents are of supreme interests for an enhanced and efficient delivery in the current biomedical sector of the modern world. The development of new types of novel, effective and highly reliable therapeutic drug delivery system (DDS) for multipurpose applications is essential and a core demand to tackle many human health related diseases. In this context, nanotechnology-based several advanced DDS have been engineered with novel characteristics for biomedical, pharmaceutical and cosmeceutical applications that include but not limited to the enhanced/improved bioactivity, bioavailability, drug efficacy, targeted delivery, and therapeutically safer with an extra advantage of overcoming demerits of traditional drug formulations/designs. This review work is focused on recent trends/advances in nanotechnology-based drugs and formulations designed for targeted therapeutic delivery. Moreover, information is also reviewed and given from recent patents and summarized or illustrated diagrammatically to depict a better understanding. Recent patents covering various nanotechnology-based approaches for several applications have also been reviewed. The drug-loaded nanoparticles are among versatile candidates with multifunctional characteristics for potential applications in biomedical, and tissue engineering sector. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  13. Targeting Potassium Channels for Increasing Delivery of Imaging Agents and Therapeutics to Brain Tumors

    OpenAIRE

    Nagendra Sanyasihally Ningaraj; Divya eKhaitan

    2013-01-01

    Every year in the US, 20,000 new primary and nearly 200,000 metastatic brain tumor cases are reported. The cerebral microvessels/ capillaries that form the blood–brain barrier (BBB) not only protect the brain from toxic agents in the blood but also pose a significant hindrance to the delivery of small and large therapeutic molecules. Different strategies have been employed to circumvent the physiological barrier posed by blood-brain tumor barrier (BTB). Studies in our laboratory have identifi...

  14. Multidisciplinary perspectives for Alzheimer's and Parkinson's diseases: hydrogels for protein delivery and cell-based drug delivery as therapeutic strategies.

    Science.gov (United States)

    Giordano, Carmen; Albani, Diego; Gloria, Antonio; Tunesi, Marta; Batelli, Sara; Russo, Teresa; Forloni, Gianluigi; Ambrosio, Luigi; Cigada, Alberto

    2009-12-01

    This review presents two intriguing multidisciplinary strategies that might make the difference in the treatment of neurodegenerative disorders such as Alzheimer's and Parkinson's diseases. The first proposed strategy is based on the controlled delivery of recombinant proteins known to play a key role in these neurodegenerative disorders that are released in situ by optimized polymer-based systems. The second strategy is the use of engineered cells, encapsulated and delivered in situ by suitable polymer-based systems, that act as drug reservoirs and allow the delivery of selected molecules to be used in the treatment of Alzheimer's and Parkinson's diseases. In both these scenarios, the design and development of optimized polymer-based drug delivery and cell housing systems for central nervous system applications represent a key requirement. Materials science provides suitable hydrogel-based tools to be optimized together with suitably designed recombinant proteins or drug delivering-cells that, once in situ, can provide an effective treatment for these neurodegenerative disorders. In this scenario, only interdisciplinary research that fully integrates biology, biochemistry, medicine and materials science can provide a springboard for the development of suitable therapeutic tools, not only for the treatment of Alzheimer's and Parkinson's diseases but also, prospectively, for a wide range of severe neurodegenerative disorders.

  15. BDNF gene delivery mediated by neuron-targeted nanoparticles is neuroprotective in peripheral nerve injury.

    Science.gov (United States)

    Lopes, Cátia D F; Gonçalves, Nádia P; Gomes, Carla P; Saraiva, Maria J; Pêgo, Ana P

    2017-03-01

    Neuron-targeted gene delivery is a promising strategy to treat peripheral neuropathies. Here we propose the use of polymeric nanoparticles based on thiolated trimethyl chitosan (TMCSH) to mediate targeted gene delivery to peripheral neurons upon a peripheral and minimally invasive intramuscular administration. Nanoparticles were grafted with the non-toxic carboxylic fragment of the tetanus neurotoxin (HC) to allow neuron targeting and were explored to deliver a plasmid DNA encoding for the brain-derived neurotrophic factor (BDNF) in a peripheral nerve injury model. The TMCSH-HC/BDNF nanoparticle treatment promoted the release and significant expression of BDNF in neural tissues, which resulted in an enhanced functional recovery after injury as compared to control treatments (vehicle and non-targeted nanoparticles), associated with an improvement in key pro-regenerative events, namely, the increased expression of neurofilament and growth-associated protein GAP-43 in the injured nerves. Moreover, the targeted nanoparticle treatment was correlated with a significantly higher density of myelinated axons in the distal stump of injured nerves, as well as with preservation of unmyelinated axon density as compared with controls and a protective role in injury-denervated muscles, preventing them from denervation. These results highlight the potential of TMCSH-HC nanoparticles as non-viral gene carriers to deliver therapeutic genes into the peripheral neurons and thus, pave the way for their use as an effective therapeutic intervention for peripheral neuropathies. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Designing the nanoparticle-biomolecule interface for "targeting and therapeutic delivery".

    Science.gov (United States)

    Mahon, Eugene; Salvati, Anna; Baldelli Bombelli, Francesca; Lynch, Iseult; Dawson, Kenneth A

    2012-07-20

    The endogenous transport mechanisms which occur in living organisms have evolved to allow selective transport and processing operate on a scale of tens of nanometers. This presents the possibility of unprecedented access for engineered nanoscale materials to organs and sub-cellular locations, materials which may in principle be targeted to precise locations for diagnostic or therapeutic gain. For this reason, nano-architectures could represent a truly radical departure as delivery agents for drugs, genes and therapies to treat a host of diseases. Thus, for active targeting, unlike the case of small molecular drugs where molecular structure has evolved to promote higher physiochemical affinity to specific sites, one aims to exploit these energy dependant endogenous processes. Many active targeting strategies have been developed, but despite this truly remarkable potential, in applications they have met with mixed success to date. This situation may have more to do with our current understanding and integration of knowledge across disciplines, than any intrinsic limitation on the vision itself. In this review article we suggest that much more fundamental and detailed control of the nanoparticle-biomolecule interface is required for sustained and general success in this field. In the simplest manifestation, pristine nanoparticles in biological fluids act as a scaffold for biomolecules, which adsorb rapidly to the nanoparticles' surface, conferring a new biological identity to the nanoparticles. It is this nanoparticle-biomolecule interface that is 'read' and acted upon by the cellular machinery. Moreover, where targeting moieties are grafted onto nanoparticles, they may not retain their function as a result of poor orientation, and structural or conformational disruption. Further surface adsorption of biomolecules from the surrounding environment i.e. the formation of a biomolecule corona may also obscure specific surface recognition. To transfer the remarkable

  17. Delivery of therapeutics for deep-seated ocular conditions - status quo.

    Science.gov (United States)

    Nguyen, Hubert; Eng, Shawn; Ngo, Thanh; Dass, Crispin R

    2018-04-19

    There is a need for research into designing effective pharmaceutical systems for delivering therapeutic drugs to the posterior of the eye for glaucoma-related pathology, macular degeneration, diabetic retinopathy, macular oedema, retinitis and choroiditis. Conventionally, eye drops have been extensively utilised for topical drug delivery to the anterior segment of the eye, but are less effective for delivery of therapeutics to the back of the eye due to significant barriers hampering drug penetration into the target intraocular tissue. This review explores some of the current and novel delivery systems employed to deliver therapeutics to the back of the eye such as those using liposomes, ocular implants, in situ gels, and nanoparticles, and how they can overcome some of these limitations. Issues such as blinking, precorneal fluid drainage, tear dilution and turnover, conjunctiva and nasal drug absorption, the corneal epithelium, vitreous drug clearance, and the blood-ocular barriers are reviewed and discussed. Further studies are needed to address their shortcomings such as drug compatibility and stability, economic viability and patient compliance. © 2018 Royal Pharmaceutical Society.

  18. Advanced Therapeutic Strategies for Chronic Lung Disease Using Nanoparticle-Based Drug Delivery

    Directory of Open Access Journals (Sweden)

    Ji Young Yhee

    2016-09-01

    Full Text Available Chronic lung diseases include a variety of obstinate and fatal diseases, including asthma, chronic obstructive pulmonary disease (COPD, cystic fibrosis (CF, idiopathic pulmonary fibrosis (IPF, and lung cancers. Pharmacotherapy is important for the treatment of chronic lung diseases, and current progress in nanoparticles offers great potential as an advanced strategy for drug delivery. Based on their biophysical properties, nanoparticles have shown improved pharmacokinetics of therapeutics and controlled drug delivery, gaining great attention. Herein, we will review the nanoparticle-based drug delivery system for the treatment of chronic lung diseases. Various types of nanoparticles will be introduced, and recent innovative efforts to utilize the nanoparticles as novel drug carriers for the effective treatment of chronic lung diseases will also be discussed.

  19. Adenoviral gene delivery to primary human cutaneous cells and burn wounds.

    Science.gov (United States)

    Hirsch, Tobias; von Peter, Sebastian; Dubin, Grzegorz; Mittler, Dominik; Jacobsen, Frank; Lehnhardt, Markus; Eriksson, Elof; Steinau, Hans-Ulrich; Steinstraesser, Lars

    2006-01-01

    The adenoviral transfer of therapeutic genes into epidermal and dermal cells is an interesting approach to treat skin diseases and to promote wound healing. The aim of this study was to assess the in vitro and in vivo transfection efficacy in skin and burn wounds after adenoviral gene delivery. Primary keratinocytes (HKC), fibroblasts (HFB), and HaCaT cells were transfected using different concentrations of an adenoviral construct (eGFP). Transfection efficiency and cytotoxicity was determined up to 30 days. Expression was quantified by FACS analysis and fluorimeter. Cytotoxicity was measured using the trypan blue exclusion method. 45 male Sprague Dawley rats received 2x10(8) pfu of Ad5-CMV-LacZ or carrier control intradermally into either superficial partial thickness scald burn or unburned skin. Animals were euthanized after 48 h, 7 or 14 days posttreatment. Transgene expression was assessed using immunohistochemistry and bioluminescent assays. The highest transfection rate was observed 48 h posttransfection: 79% for HKC, 70% for HFB, and 48% for HaCaT. The eGFP expression was detectable in all groups over 30 days (P>0.05). Cytotoxic effects of the adenoviral vector were observed for HFB after 10 days and HaCaT after 30 days. Reporter gene expression in vivo was significantly higher in burned skin compared with unburned skin (P=0,004). Gene expression decreases from 2 to 7 days with no significant expression after 14 days. This study demonstrates that effective adenoviral-mediated gene transfer of epidermal primary cells and cell-lines is feasible. Ex vivo gene transfer in epithelial cells might have promise for the use in severely burned patients who receive autologous keratinocyte sheets. Transient cutaneous gene delivery in burn wounds using adenoviral vectors causes significant concentrations in the wound tissue for at least 1 week. Based on these findings, we hypothesize that transient cutaneous adenoviral gene delivery of wound healing promoting factors has

  20. Noninvasive gene delivery to foveal cones for vision restoration

    Science.gov (United States)

    Khabou, Hanen; Garita-Hernandez, Marcela; Jaillard, Céline; Brazhnikova, Elena; Bertin, Stéphane; Forster, Valérie; Desrosiers, Mélissa; Winckler, Céline; Goureau, Olivier; Duebel, Jens; Sahel, José-Alain

    2018-01-01

    Intraocular injection of adeno-associated viral (AAV) vectors has been an evident route for delivering gene drugs into the retina. However, gaps in our understanding of AAV transduction patterns within the anatomically unique environments of the subretinal and intravitreal space of the primate eye impeded the establishment of noninvasive and efficient gene delivery to foveal cones in the clinic. Here, we establish new vector-promoter combinations to overcome the limitations associated with AAV-mediated cone transduction in the fovea with supporting studies in mouse models, human induced pluripotent stem cell–derived organoids, postmortem human retinal explants, and living macaques. We show that an AAV9 variant provides efficient foveal cone transduction when injected into the subretinal space several millimeters away from the fovea, without detaching this delicate region. An engineered AAV2 variant provides gene delivery to foveal cones with a well-tolerated dose administered intravitreally. Both delivery modalities rely on a cone-specific promoter and result in high-level transgene expression compatible with optogenetic vision restoration. The model systems described here provide insight into the behavior of AAV vectors across species to obtain safety and efficacy needed for gene therapy in neurodegenerative disorders. PMID:29367457

  1. Lipid nanoparticles as drug/gene delivery systems to the retina.

    Science.gov (United States)

    del Pozo-Rodríguez, Ana; Delgado, Diego; Gascón, Alicia R; Solinís, Maria Ángeles

    2013-03-01

    This review highlights the application of lipid nanoparticles (Solid Lipid Nanoparticles, Nanostructured Lipid Carriers, or Lipid Drug Conjugates) as effective drug/gene delivery systems for retinal diseases. Most drug products for ocular disease treatment are marketed as eye drop formulations but, due to ocular barriers, the drug concentration in the retina hardly ever turns out to be effective. Up to this date, several delivery systems have been designed to deliver drugs to the retina. Drug delivery strategies may be classified into 3 groups: noninvasive techniques, implants, and colloidal carriers. The best known systems for drug delivery to the posterior eye are intravitreal implants; in fact, some of them are being clinically used. However, their long-term accumulation might impact the patient's vision. On the contrary, colloidal drug delivery systems (microparticles, liposomes, or nanoparticles) can be easily administered in a liquid form. Nanoparticular systems diffuse rapidly and are better internalized in ocular tissues than microparticles. In comparison with liposomes, nanoparticles have a higher loading capacity and are more stable in biological fluids and during storage. In addition, their capacity to adhere to the ocular surface and interact with the endothelium makes these drug delivery systems interesting as new therapeutic tools in ophthalmology. Within the group of nanoparticles, those composed of lipids (Solid Lipid Nanoparticles, Nanostructred Lipid Carriers, and Lipid Drug Conjugates) are more biocompatible, easy to produce at large scale, and they may be autoclaved or sterilized. The present review summarizes scientific results that evidence the potential application of lipid nanoparticles as drug delivery systems for the retina and also as nonviral vectors in gene therapy of retina disorders, although much more effort is still needed before these lipidic systems could be available in the market.

  2. Theory and in vivo application of electroporative gene delivery.

    Science.gov (United States)

    Somiari, S; Glasspool-Malone, J; Drabick, J J; Gilbert, R A; Heller, R; Jaroszeski, M J; Malone, R W

    2000-09-01

    Efficient and safe methods for delivering exogenous genetic material into tissues must be developed before the clinical potential of gene therapy will be realized. Recently, in vivo electroporation has emerged as a leading technology for developing nonviral gene therapies and nucleic acid vaccines (NAV). Electroporation (EP) involves the application of pulsed electric fields to cells to enhance cell permeability, resulting in exogenous polynucleotide transit across the cytoplasmic membrane. Similar pulsed electrical field treatments are employed in a wide range of biotechnological processes including in vitro EP, hybridoma production, development of transgenic animals, and clinical electrochemotherapy. Electroporative gene delivery studies benefit from well-developed literature that may be used to guide experimental design and interpretation. Both theory and experimental analysis predict that the critical parameters governing EP efficacy include cell size and field strength, duration, frequency, and total number of applied pulses. These parameters must be optimized for each tissue in order to maximize gene delivery while minimizing irreversible cell damage. By providing an overview of the theory and practice of electroporative gene transfer, this review intends to aid researchers that wish to employ the method for preclinical and translational gene therapy, NAV, and functional genomic research.

  3. Non-viral gene delivery strategies for gene therapy: a “ménage à trois” among nucleic acids, materials, and the biological environment

    International Nuclear Information System (INIS)

    Pezzoli, Daniele; Candiani, Gabriele

    2013-01-01

    Gene delivery is the science of transferring genetic material into cells by means of a vector to alter cellular function or structure at a molecular level. In this context, a number of nucleic acid-based drugs have been proposed and experimented so far and, as they act on distinct steps along the gene transcription–translation pathway, specific delivery strategies are required to elicit the desired outcome. Cationic lipids and polymers, collectively known as non-viral delivery systems, have thus made their breakthrough in basic and medical research. Albeit they are promising alternatives to viral vectors, their therapeutic application is still rather limited as high transfection efficiencies are normally associated to adverse cytotoxic side effects. In this scenario, drawing inspiration from processes naturally occurring in vivo, major strides forward have been made in the development of more effective materials for gene delivery applications. Specifically, smart vectors sensitive to a variety of physiological stimuli such as cell enzymes, redox status, and pH are substantially changing the landscape of gene delivery by helping to overcome some of the systemic and intracellular barriers that viral vectors naturally evade. Herein, after summarizing the state-of-the-art information regarding the use of nucleic acids as drugs, we review the main bottlenecks still limiting the overall effectiveness of non-viral gene delivery systems. Finally, we provide a critical outline of emerging stimuli-responsive strategies and discuss challenges still existing on the road toward conceiving more efficient and safer multifunctional vectors.

  4. Recent Advances in Non-viral Vectors for Gene Delivery

    Science.gov (United States)

    Guo, Xia; Huang, Leaf

    2011-01-01

    CONSPECTUS Non-viral vectors, typically based on cationic lipids or polymers, are preferred due to safety concerns with viral vectors. So far, non-viral vectors can proficiently transfect cells in culture, but obtaining efficient nanomedicines is far from evident. To overcome the hurdles associated with non-viral vectors is significant for improving delivery efficiency and therapeutic effect of nucleic acid. The drawbacks include the strong interaction of cationic delivery vehicles with blood components, uptake by the reticuloendothelial system (RES), toxicity, targeting ability of the carriers to the cells of interest, and so on. PEGylation is the predominant method used to reduce the binding of plasma proteins with non-viral vectors and minimize the clearance by RES after intravenous administration. The nanoparticles that are not rapidly cleared from the circulation accumulate in the tumors due to the enhanced permeability and retention effect, and the targeting ligands attached to the distal end of the PEGylated components allow binding to the receptors on the target cell surface. Neutral or anionic liposomes have been also developed for systemic delivery of nucleic acids in experimental animal model. Designing and synthesizing novel cationic lipids and polymers, and binding nucleic acid with peptides, targeting ligands, polymers, or environmentally sensitive moieties also attract many attentions for resolving the problems encountered by non-viral vectors. The application of inorganic nanoparticles in nucleic acid delivery is an emerging field, too. Recently, different classes of non-viral vectors appear to be converging and the features of different classes of non-viral vectors could be combined in one strategy. More hurdles associated with efficient nucleic acid delivery therefore might be expected to be overcome. In this account, we will focus on these novel non-viral vectors, which are classified into multifunctional hybrid nucleic acid vectors, novel

  5. Tailoring the dendrimer core for efficient gene delivery.

    Science.gov (United States)

    Hu, Jingjing; Hu, Ke; Cheng, Yiyun

    2016-04-15

    Dendrimers have been widely used as non-viral gene vectors due to well-defined chemical structures, high density of cationic charges and ease of surface modification. Although a large number of studies have reported the important roles of dendrimer architecture, component, generation and surface functionality in gene delivery, the effect of dendrimer core on this issue still remains unclear. Recent literatures suggest that a slight alternation in dendrimer core has a profound effect in the transfection efficacy and biocompatibility. In this review, we will discuss the transfection mechanism of dendrimers with different types of cores in respect of flexibility, hydrophobicity and functionality. We hope to open a possibility of designing efficient dendrimers for gene delivery by choosing a proper dendrimer core. As a branch of researches on dendrimers and dendritic polymers, the design of biocompatible and high efficient polymeric gene carriers has attracted increasing attentions during these years. Although the effect of dendrimer generation, species, architecture and surface functionality on gene delivery have been widely reported, the effect of dendrimer core on this issue still remains unclear. Recent literatures suggest that a minor variation on the dendrimer core has a profound effect in the transfection efficacy and biocompatibility. This critical review summarized the dendrimers with different types of cores and discussed the transfection mechanism with particular focus on the flexibility, hydrophobicity, and functionality. It is hoped to provide a new insight to design efficient and safe dendrimer-based gene vectors by choosing a proper core. To the best of our knowledge, this is the first review on the effect of dendrimer core on gene delivery. The findings obtained in this filed are of central importance in the design of efficient polymeric gene vectors. This article will appeal a wide readership such as physical chemist, dendrimer chemist, biological

  6. A Potential Therapeutic Strategy for Malignant Mesothelioma with Gene Medicine

    Directory of Open Access Journals (Sweden)

    Yuji Tada

    2013-01-01

    Full Text Available Malignant mesothelioma, closely linked with occupational asbestos exposure, is relatively rare in the frequency, but the patient numbers are going to increase in the next few decades all over the world. The current treatment modalities are not effective in terms of the overall survival and the quality of life. Mesothelioma mainly develops in the thoracic cavity and infrequently metastasizes to extrapleural organs. A local treatment can thereby be beneficial to the patients, and gene therapy with an intrapleural administration of vectors is one of the potential therapeutics. Preclinical studies demonstrated the efficacy of gene medicine for mesothelioma, and clinical trials with adenovirus vectors showed the safety of an intrapleural injection and a possible involvement of antitumor immune responses. Nevertheless, low transduction efficiency remains the main hurdle that hinders further clinical applications. Moreover, rapid generation of antivector antibody also inhibits transgene expressions. In this paper, we review the current status of preclinical and clinical gene therapy for malignant mesothelioma and discuss potential clinical directions of gene medicine in terms of a combinatory use with anticancer agents and with immunotherapy.

  7. Polyethyleneimine grafted short halloysite nanotubes for gene delivery.

    Science.gov (United States)

    Long, Zheru; Zhang, Jun; Shen, Yan; Zhou, Changren; Liu, Mingxian

    2017-12-01

    Inorganic nanoparticles have attracted much attentions in gene delivery because of their desirable characteristics including low toxicity, well-controlled characteristics, high gene delivery efficiency, and multi-functionalities. Here, natural occurred halloysite nanotubes (HNTs) were developed as a novel non-viral gene vector. To increase the efficiency of endocytosis, HNTs were firstly shortened into an appropriate size (~200nm). Then polyethyleneimine (PEI) was grafted onto HNTs to bind green fluorescence protein (GFP) labeled pDNA. The structure and physical-chemical properties of PEI grafted HNTs (PEI-g-HNTs) were characterized by various methods. PEI-g-HNTs show lower cytotoxicity than PEI. PEI-g-HNTs are positively charged and can bind DNA tightly at designed N/P ratio from 5:1 to 40:1. PEI-g-HNTs/pDNA complexes show much higher transfection efficiency towards both 293T and HeLa cells compared with PEI/pDNA complexes at the equivalent N/P ratio. The transfection efficiencies of PEI-g-HNTs/pDNA complex towards HeLa cell can reach to 44.4% at N/P ratio of 20. PEI-g-HNTs/pDNA complexes possess a higher GFP protein expression than PEI/pDNA from simple western immunoblots. So, PEI-g-HNTs are potential gene vectors with good biocompatibility and high transfection efficiency, which have promising applications in cancer gene therapy. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. Gene therapy for human glioblastoma using neurotropic JC virus-like particles as a gene delivery vector.

    Science.gov (United States)

    Chao, Chun-Nun; Yang, Yu-Hsuan; Wu, Mu-Sheng; Chou, Ming-Chieh; Fang, Chiung-Yao; Lin, Mien-Chun; Tai, Chien-Kuo; Shen, Cheng-Huang; Chen, Pei-Lain; Chang, Deching; Wang, Meilin

    2018-02-02

    Glioblastoma multiforme (GBM), the most common malignant brain tumor, has a short period of survival even with recent multimodality treatment. The neurotropic JC polyomavirus (JCPyV) infects glial cells and oligodendrocytes and causes fatal progressive multifocal leukoencephalopathy in patients with AIDS. In this study, a possible gene therapy strategy for GBM using JCPyV virus-like particles (VLPs) as a gene delivery vector was investigated. We found that JCPyV VLPs were able to deliver the GFP reporter gene into tumor cells (U87-MG) for expression. In an orthotopic xenograft model, nude mice implanted with U87 cells expressing the near-infrared fluorescent protein and then treated by intratumoral injection of JCPyV VLPs carrying the thymidine kinase suicide gene, combined with ganciclovir administration, exhibited significantly prolonged survival and less tumor fluorescence during the experiment compared with controls. Furthermore, JCPyV VLPs were able to protect and deliver a suicide gene to distal subcutaneously implanted U87 cells in nude mice via blood circulation and inhibit tumor growth. These findings show that metastatic brain tumors can be targeted by JCPyV VLPs carrying a therapeutic gene, thus demonstrating the potential of JCPyV VLPs to serve as a gene therapy vector for the far highly treatment-refractory GBM.

  9. Nanoparticle-mediated delivery of pioglitazone enhances therapeutic neovascularization in a murine model of hindlimb ischemia.

    Science.gov (United States)

    Nagahama, Ryoji; Matoba, Tetsuya; Nakano, Kaku; Kim-Mitsuyama, Shokei; Sunagawa, Kenji; Egashira, Kensuke

    2012-10-01

    Critical limb ischemia is a severe form of peripheral artery disease (PAD) for which neither surgical revascularization nor endovascular therapy nor current medicinal therapy has sufficient therapeutic effects. Peroxisome proliferator activated receptor-γ agonists present angiogenic activity in vitro; however, systemic administration of peroxisome proliferator-activated receptor-γ agonists is hampered by its side effects, including heart failure. Here, we demonstrate that the nanoparticle (NP)-mediated delivery of the peroxisome proliferator activated receptor-γ agonist pioglitazone enhances its therapeutic efficacy on ischemia-induced neovascularization in a murine model. In a nondiabetic murine model of hindlimb ischemia, a single intramuscular injection of pioglitazone-incorporated NP (1 µg/kg) into ischemic muscles significantly improved the blood flow recovery in the ischemic limbs, significantly increasing the number of CD31-positive capillaries and α-smooth muscle actin-positive arterioles. The therapeutic effects of pioglitazone-incorporated NP were diminished by the peroxisome proliferator activated receptor-γ antagonist GW9662 and were not observed in endothelial NO synthase-deficient mice. Pioglitazone-incorporated NP induced endothelial NO synthase phosphorylation, as demonstrated by Western blot analysis, as well as expression of multiple angiogenic growth factors in vivo, including vascular endothelial growth factor-A, vascular endothelial growth factor-B, and fibroblast growth factor-1, as demonstrated by real-time polymerase chain reaction. Intramuscular injection of pioglitazone (1 µg/kg) was ineffective, and oral administration necessitated a >500 μg/kg per day dose to produce therapeutic effects equivalent to those of pioglitazone-incorporated NP. NP-mediated drug delivery is a novel modality that may enhance the effectiveness of therapeutic neovascularization, surpassing the effectiveness of current treatments for peripheral artery

  10. Retroviral packaging cells encapsulated in TheraCyte immunoisolation devices enable long-term in vivo gene delivery.

    Science.gov (United States)

    Krupetsky, Anna; Parveen, Zahida; Marusich, Elena; Goodrich, Adrienne; Dornburg, Ralph

    2003-05-01

    The method of delivering a therapeutic gene into a patient is still one of the major obstacles towards successful human gene therapy. Here we describe a novel gene delivery approach using TheraCyte immunoisolation devices. Retroviral vector producing cells, derived from the avian retrovirus spleen necrosis virus, SNV, were encapsulated in TheraCyte devices and tested for the release of retroviral vectors. In vitro experiments show that such devices release infectious retroviral vectors into the tissue culture medium for up to 4 months. When such devices were implanted subcutaneously in SCID mice, infectious virus was released into the blood stream. There, the vectors were transported to and infected tumors, which had been induced by subcutaneous injection of tissue culture cells. Thus, this novel concept of a continuous, long-term gene delivery may constitute an attractive approach for future in vivo human gene therapy.

  11. Fiberoptic microneedles: novel optical diffusers for interstitial delivery of therapeutic light.

    Science.gov (United States)

    Kosoglu, Mehmet A; Hood, Robert L; Rossmeisl, John H; Grant, David C; Xu, Yong; Robertson, John L; Rylander, Marissa Nichole; Rylander, Christopher G

    2011-11-01

    Photothermal therapies have limited efficacy and application due to the poor penetration depth of light inside tissue. In earlier work, we described the development of novel fiberoptic microneedles to provide a means to mechanically penetrate dermal tissue and deliver light directly into a localized target area.This paper presents an alternate fiberoptic microneedle design with the capability of delivering more diffuse, but therapeutically useful photothermal energy. Laser lipolysis is envisioned as a future clinical application for this design. A novel fiberoptic microneedle was developed using hydrofluoric acid etching of optical fiber to permit diffuse optical delivery. Microneedles etched for 10, 30, and 50 minutes, and an optical fiber control were compared with three techniques. First, red light delivery from the microneedles was evaluated by imaging the reflectance of the light from a white paper.Second, spatial temperature distribution of the paper in response to near-IR light (1,064 nm, 1 W CW) was recorded using infrared thermography. Third, ex vivo adipose tissue response during 1,064 nm, (5 W CW)irradiation was recorded with bright field microscopy. Acid etching exposed a 3 mm length of the fiber core, allowing circumferential delivery of light along this length. Increasing etching time decreased microneedle diameter, resulting in increased uniformity of red and 1,064 nm light delivery along the microneedle axis. For equivalent total energy delivery, thinner microneedles reduced carbonization in the adipose tissue experiments. We developed novel microscale optical diffusers that provided a more homogeneous light distribution from their surfaces, and compared performance to a flat-cleaved fiber, a device currently utilized in clinical practice. These fiberoptic microneedles can potentially enhance clinical laser procedures by providing direct delivery of diffuse light to target chromophores, while minimizing undesirable photothermal damage in adjacent

  12. Sendai viroplexes for epidermal growth factor receptor-directed delivery of interleukin-12 and salmosin genes to cancer cells.

    Science.gov (United States)

    Kim, Jung Seok; Kim, Min Woo; Jeong, Hwa Yeon; Kang, Seong Jae; Park, Sang Il; Lee, Yeon Kyung; Kim, Hong Sung; Kim, Keun Sik; Park, Yong Serk

    2016-07-01

    The effective delivery of therapeutic genes to target cells has been a fundamental goal in cancer gene therapy because of its advantages with respect to both safety and transfection efficiency. In the present, study we describe a tumor-directed gene delivery system that demonstrates remarkable efficacy in gene delivery and minimizes the off-target effects of gene transfection. The system consists of a well-verified cationic O,O'-dimyristyl-N-lysyl glutamate (DMKE), Sendai virus fusion (F) protein and hemagglutinin-neuraminidase (HN) protein, referred to as cationic Sendai F/HN virosomes. To achieve tumor-specific recognition, anti-epidermal growth factor (EGF) receptor antibody was coupled to the surface of the virosomes containing interleukin-12 (IL-12) and/or salmosin genes that have potent anti-angiogenetic functions. Among the virosomal formulations, the anti-EGF receptor (EGFR) viroplexes, prepared via complexation of plasmid DNA (pDNA) with cationic DMKE lipid, exhibited more efficient gene transfection to tumor cells over-expressing EGF receptors compared to the neutrally-charged anti-EGFR virosomes encapsulating pDNA. In addition, the anti-EGFR viroplexes with IL-12 and salmosin genes exhibited the most effective therapeutic efficacy in a mouse tumor model. Especially when combined with doxorubicin, transfection of the two genes via the anti-EGFR viroplexes exhibited an enhanced inhibitory effect on tumor growth and metastasis in lungs. The results of the present study suggest that anti-EGFR viroplexes can be utilized as an effective strategy for tumor-directed gene delivery. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  13. Newly engineered magnetic erythrocytes for sustained and targeted delivery of anti-cancer therapeutic compounds.

    Directory of Open Access Journals (Sweden)

    Caterina Cinti

    Full Text Available Cytotoxic chemotherapy of cancer is limited by serious, sometimes life-threatening, side effects that arise from toxicities to sensitive normal cells because the therapies are not selective for malignant cells. So how can they be selectively improved? Alternative pharmaceutical formulations of anti-cancer agents have been investigated in order to improve conventional chemotherapy treatment. These formulations are associated with problems like severe toxic side effects on healthy organs, drug resistance and limited access of the drug to the tumor sites suggested the need to focus on site-specific controlled drug delivery systems. In response to these concerns, we have developed a new drug delivery system based on magnetic erythrocytes engineered with a viral spike fusion protein. This new erythrocyte-based drug delivery system has the potential for magnetic-controlled site-specific localization and highly efficient fusion capability with the targeted cells. Here we show that the erythro-magneto-HA virosomes drug delivery system is able to attach and fuse with the target cells and to efficiently release therapeutic compounds inside the cells. The efficacy of the anti-cancer drug employed is increased and the dose required is 10 time less than that needed with conventional therapy.

  14. Newly Engineered Magnetic Erythrocytes for Sustained and Targeted Delivery of Anti-Cancer Therapeutic Compounds

    Science.gov (United States)

    Taranta, Monia; Naldi, Ilaria

    2011-01-01

    Cytotoxic chemotherapy of cancer is limited by serious, sometimes life-threatening, side effects that arise from toxicities to sensitive normal cells because the therapies are not selective for malignant cells. So how can they be selectively improved? Alternative pharmaceutical formulations of anti-cancer agents have been investigated in order to improve conventional chemotherapy treatment. These formulations are associated with problems like severe toxic side effects on healthy organs, drug resistance and limited access of the drug to the tumor sites suggested the need to focus on site-specific controlled drug delivery systems. In response to these concerns, we have developed a new drug delivery system based on magnetic erythrocytes engineered with a viral spike fusion protein. This new erythrocyte-based drug delivery system has the potential for magnetic-controlled site-specific localization and highly efficient fusion capability with the targeted cells. Here we show that the erythro-magneto-HA virosomes drug delivery system is able to attach and fuse with the target cells and to efficiently release therapeutic compounds inside the cells. The efficacy of the anti-cancer drug employed is increased and the dose required is 10 time less than that needed with conventional therapy. PMID:21373641

  15. Therapeutic Role and Drug Delivery Potential of Neuroinflammation as a Target in Neurodegenerative Disorders.

    Science.gov (United States)

    Singh, Abhijeet; Chokriwal, Ankit; Sharma, Madan Mohan; Jain, Devendra; Saxena, Juhi; Stephen, Bjorn John

    2017-08-16

    Neuroinflammation, the condition associated with the hyperactivity of immune cells within the CNS (central nervous system), has recently been linked to a host range of neurodegenerative disorders. Targeting neuroinflammation could be of prime importance as recent research highlights the beneficial aspects associated with modulating the inflammatory mediators associated with the CNS. One of the main obstructions in neuroinflammatory treatments is the hindrance posed by the blood-brain barrier for the delivery of drugs. Hence, research has focused on novel modes of transport for drugs to cross the barrier through drug delivery and nanotechnology approaches. In this Review, we highlight the therapeutic advancement made in the field of neurodegenerative disorders by focusing on the effect neuroinflammation treatment has on these conditions.

  16. Macrophages offer a paradigm switch for CNS delivery of therapeutic proteins.

    Science.gov (United States)

    Klyachko, Natalia L; Haney, Matthew J; Zhao, Yuling; Manickam, Devika S; Mahajan, Vivek; Suresh, Poornima; Hingtgen, Shawn D; Mosley, R Lee; Gendelman, Howard E; Kabanov, Alexander V; Batrakova, Elena V

    2014-07-01

    Active targeted transport of the nanoformulated redox enzyme, catalase, in macrophages attenuates oxidative stress and as such increases survival of dopaminergic neurons in animal models of Parkinson's disease. Optimization of the drug formulation is crucial for the successful delivery in living cells. We demonstrated earlier that packaging of catalase into a polyion complex micelle ('nanozyme') with a synthetic polyelectrolyte block copolymer protected the enzyme against degradation in macrophages and improved therapeutic outcomes. We now report the manufacture of nanozymes with superior structure and therapeutic indices. Synthesis, characterization and therapeutic efficacy of optimal cell-based nanoformulations are evaluated. A formulation design for drug carriers typically works to avoid entrapment in monocytes and macrophages focusing on small-sized nanoparticles with a polyethylene glycol corona (to provide a stealth effect). By contrast, the best nanozymes for delivery in macrophages reported in this study have a relatively large size (≈ 200 nm), which resulted in improved loading capacity and release from macrophages. Furthermore, the cross-linking of nanozymes with the excess of a nonbiodegradable linker ensured their low cytotoxicity, and efficient catalase protection in cell carriers. Finally, the 'alternatively activated' macrophage phenotype (M2) utilized in these studies did not promote further inflammation in the brain, resulting in a subtle but statistically significant effect on neuronal regeneration and repair in vivo. The optimized cross-linked nanozyme loaded into macrophages reduced neuroinflammatory responses and increased neuronal survival in mice. Importantly, the approach for nanoformulation design for cell-mediated delivery is different from the common requirements for injectable formulations.

  17. Systems-level thinking for nanoparticle-mediated therapeutic delivery to neurological diseases.

    Science.gov (United States)

    Curtis, Chad; Zhang, Mengying; Liao, Rick; Wood, Thomas; Nance, Elizabeth

    2017-03-01

    Neurological diseases account for 13% of the global burden of disease. As a result, treating these diseases costs $750 billion a year. Nanotechnology, which consists of small (~1-100 nm) but highly tailorable platforms, can provide significant opportunities for improving therapeutic delivery to the brain. Nanoparticles can increase drug solubility, overcome the blood-brain and brain penetration barriers, and provide timed release of a drug at a site of interest. Many researchers have successfully used nanotechnology to overcome individual barriers to therapeutic delivery to the brain, yet no platform has translated into a standard of care for any neurological disease. The challenge in translating nanotechnology platforms into clinical use for patients with neurological disease necessitates a new approach to: (1) collect information from the fields associated with understanding and treating brain diseases and (2) apply that information using scalable technologies in a clinically-relevant way. This approach requires systems-level thinking to integrate an understanding of biological barriers to therapeutic intervention in the brain with the engineering of nanoparticle material properties to overcome those barriers. To demonstrate how a systems perspective can tackle the challenge of treating neurological diseases using nanotechnology, this review will first present physiological barriers to drug delivery in the brain and common neurological disease hallmarks that influence these barriers. We will then analyze the design of nanotechnology platforms in preclinical in vivo efficacy studies for treatment of neurological disease, and map concepts for the interaction of nanoparticle physicochemical properties and pathophysiological hallmarks in the brain. WIREs Nanomed Nanobiotechnol 2017, 9:e1422. doi: 10.1002/wnan.1422 For further resources related to this article, please visit the WIREs website. © 2016 Wiley Periodicals, Inc.

  18. Macrophages offer a paradigm switch for CNS delivery of therapeutic proteins

    Science.gov (United States)

    Klyachko, Natalia L; Haney, Matthew J; Zhao, Yuling; Manickam, Devika S; Mahajan, Vivek; Suresh, Poornima; Hingtgen, Shawn D; Mosley, R Lee; Gendelman, Howard E; Kabanov, Alexander V; Batrakova, Elena V

    2013-01-01

    Aims Active targeted transport of the nanoformulated redox enzyme, catalase, in macrophages attenuates oxidative stress and as such increases survival of dopaminergic neurons in animal models of Parkinson’s disease. Optimization of the drug formulation is crucial for the successful delivery in living cells. We demonstrated earlier that packaging of catalase into a polyion complex micelle (‘nanozyme’) with a synthetic polyelectrolyte block copolymer protected the enzyme against degradation in macrophages and improved therapeutic outcomes. We now report the manufacture of nanozymes with superior structure and therapeutic indices. Methods Synthesis, characterization and therapeutic efficacy of optimal cell-based nanoformulations are evaluated. Results A formulation design for drug carriers typically works to avoid entrapment in monocytes and macrophages focusing on small-sized nanoparticles with a polyethylene glycol corona (to provide a stealth effect). By contrast, the best nanozymes for delivery in macrophages reported in this study have a relatively large size (~200 nm), which resulted in improved loading capacity and release from macrophages. Furthermore, the cross-linking of nanozymes with the excess of a nonbiodegradable linker ensured their low cytotoxicity, and efficient catalase protection in cell carriers. Finally, the ‘alternatively activated’ macrophage phenotype (M2) utilized in these studies did not promote further inflammation in the brain, resulting in a subtle but statistically significant effect on neuronal regeneration and repair in vivo. Conclusion The optimized cross-linked nanozyme loaded into macrophages reduced neuroinflammatory responses and increased neuronal survival in mice. Importantly, the approach for nanoformulation design for cell-mediated delivery is different from the common requirements for injectable formulations. PMID:24237263

  19. Water insoluble and soluble lipids for gene delivery.

    Science.gov (United States)

    Mahato, Ram I

    2005-04-05

    Among various synthetic gene carriers currently in use, liposomes composed of cationic lipids and co-lipids remain the most efficient transfection reagents. Physicochemical properties of lipid/plasmid complexes, such as cationic lipid structure, cationic lipid to co-lipid ratio, charge ratio, particle size and zeta potential have significant influence on gene expression and biodistribution. However, most cationic lipids are toxic and cationic liposomes/plasmid complexes do not disperse well inside the target tissues because of their large particle size. To overcome the problems associated with cationic lipids, we designed water soluble lipopolymers for gene delivery to various cells and tissues. This review provides a critical discussion on how the components of water insoluble and soluble lipids affect their transfection efficiency and biodistribution of lipid/plasmid complexes.

  20. Mannan-Modified PLGA Nanoparticles for Targeted Gene Delivery

    Directory of Open Access Journals (Sweden)

    Fansheng Kong

    2012-01-01

    Full Text Available The studies of targeted gene delivery nanocarriers have gained increasing attention during the past decades. In this study, mannan modified DNA loaded bioadhesive PLGA nanoparticles (MAN-DNA-NPs were investigated for targeted gene delivery to the Kupffer cells (KCs. Bioadhesive PLGA nanoparticles were prepared and subsequently bound with pEGFP. Following the coupling of the mannan-based PE-grafted ligands (MAN-PE with the DNA-NPs, the MAN-DNA-NPs were delivered intravenously to rats. The transfection efficiency was determined from the isolated KCs and flow cytometry was applied for the quantitation of gene expression after 48 h post transfection. The size of the MAN-DNA-NPs was found to be around 190 nm and the Zeta potential was determined to be −15.46mV. The pEGFP binding capacity of MAN-DNA-NPs was (88.9±5.8% and the in vitro release profiles of the MAN-DNA-NPs follow the Higuchi model. When compared with non-modified DNA-NPs and Lipofectamine 2000-DNA, MAN-DNA-NPs produced the highest gene expressions, especially in vivo. The in vivo data from flow cytometry analysis showed that MAN-DNA-NPs displayed a remarkably higher transfection efficiency (39% than non-modified DNA-NPs (25% and Lipofectamine 2000-DNA (23% in KCs. The results illustrate that MAN-DNA-NPs have the ability to target liver KCs and could function as promising active targeting drug delivery vectors.

  1. Influence of FCGRT gene polymorphisms on pharmacokinetics of therapeutic antibodies.

    Science.gov (United States)

    Passot, Christophe; Azzopardi, Nicolas; Renault, Sylvaine; Baroukh, Nadine; Arnoult, Christophe; Ohresser, Marc; Boisdron-Celle, Michèle; Gamelin, Erick; Watier, Hervé; Paintaud, Gilles; Gouilleux-Gruart, Valérie

    2013-01-01

    The neonatal Fc receptor (FcRn) encoded by FCGRT is known to be involved in the pharmacokinetics (PK) of therapeutic monoclonal antibodies (mAbs). Variability in the expression of FCGRT gene and consequently in the FcRn protein level could explain differences in PK observed between patients treated with mAbs. We studied whether the previously described variable number tandem repeat (VNTR) or copy number variation (CNV) of FCGRT are associated with individual variations of PK parameters of cetuximab. VNTR and CNV were assessed on genomic DNA of 198 healthy individuals and of 94 patients treated with the therapeutic mAb. VNTR and CNV were analyzed by allele-specific PCR and duplex real-time PCR with Taqman (®) technology, respectively. The relationship between FCGRT polymorphisms (VNTR and CNV) and PK parameters of patients treated with cetuximab was studied. VNTR3 homozygote patients had a lower cetuximab distribution clearance than VNTR2/VNTR3 and VNTR3/VNTR4 patients (p = 0.021). We observed no affects of VNTR genotype on elimination clearance. One healthy person (0.5%) and 1 patient (1.1%) had 3 copies of FCGRT. The PK parameters of this patient did not differ from those of patients with 2 copies. The FCGRT promoter VNTR may influence mAbs' distribution in the body. CNV of FCGRT cannot be used as a relevant pharmacogenetic marker because of its low frequency.

  2. Long circulating polymeric nanoparticles for gene/drug delivery.

    Science.gov (United States)

    Hu, Jiaming; Sheng, Yan; Shi, Junfeng; Yu, Bohao; Yu, Zhiqiang; Liao, Guochao

    2017-12-07

    The major limitation in the improving polymeric nanoparticles into an efficient gene/drug delivery carrier is the rapid opsonization, phagocytic uptake by mononuclear phagocyte system and subsequent clearance from the bloodstream. The prolonged circulation time of nanoparticles in the blood is a prerequisite to realizing a controlled and targeted (passive or active targeting) release of the encapsulated gene/drug at the desired site of action. In this review, the factors such as biological barriers and physical barriers including particle size, shape, zeta potential, and hydrophilicity will be discussed, which can cause effects on blood clearance and organ accumulation. Some natural and synthetic polymers utilized in long-circulating nanoparticles will also be discussed. The most popular method to mask or camouflage nanoparticles is the adsorbed, grafted or conjugated of poly (ethylene glycol) (PEG) or other hydrophilic polymers (e.g. polysaccharides) to the particle surface. Surface modification of nanoparticles with these polymers results in an increased blood circulation time by several orders of magnitude in comparison to the bare nanoparticles. However, the circulation half-life of nanoparticles still cannot satisfy the need for clinical use. At present, identification of novel potential coating materials is an emerging field of interest in the design of long-circulating polymer-based nanoparticulate gene/drug delivery. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  3. Selective Inhibition of Histone Deacetylation in Melanoma Increases Targeted Gene Delivery by a Bacteriophage Viral Vector

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    Samuel Campbell

    2018-04-01

    Full Text Available The previously developed adeno-associated virus/phage (AAVP vector, a hybrid between M13 bacteriophage (phage viruses that infect bacteria only and human Adeno-Associated Virus (AAV, is a promising tool in targeted gene therapy against cancer. AAVP can be administered systemically and made tissue specific through the use of ligand-directed targeting. Cancer cells and tumor-associated blood vessels overexpress the αν integrin receptors, which are involved in tumor angiogenesis and tumor invasion. AAVP is targeted to these integrins via a double cyclic RGD4C ligand displayed on the phage capsid. Nevertheless, there remain significant host-defense hurdles to the use of AAVP in targeted gene delivery and subsequently in gene therapy. We previously reported that histone deacetylation in cancer constitutes a barrier to AAVP. Herein, to improve AAVP-mediated gene delivery to cancer cells, we combined the vector with selective adjuvant chemicals that inhibit specific histone deacetylases (HDAC. We examined the effects of the HDAC inhibitor C1A that mainly targets HDAC6 and compared this to sodium butyrate, a pan-HDAC inhibitor with broad spectrum HDAC inhibition. We tested the effects on melanoma, known for HDAC6 up-regulation, and compared this side by side with a normal human kidney HEK293 cell line. Varying concentrations were tested to determine cytotoxic levels as well as effects on AAVP gene delivery. We report that the HDAC inhibitor C1A increased AAVP-mediated transgene expression by up to ~9-fold. These findings indicate that selective HDAC inhibition is a promising adjuvant treatment for increasing the therapeutic value of AAVP.

  4. Transdermal therapeutic systems for memantine delivery. Comparison of passive and iontophoretic transport.

    Science.gov (United States)

    Del Río-Sancho, S; Serna-Jiménez, C E; Sebastián-Morelló, M; Calatayud-Pascual, M A; Balaguer-Fernández, C; Femenía-Font, A; Kalia, Y N; Merino, V; López-Castellano, A

    2017-01-30

    Memantine is a non-competitive N-methyl-d-aspartate (NMDA) receptor antagonist used in the treatment of moderate to severe dementia including the symptoms of Alzheimer's disease (AD). It is administered orally but compliance, swallowing problems and the routine use of multiple medications in elderly AD patients means that an alternative route of administration would be of interest. The aim of the present study was to develop memantine hydrochloride occlusive transdermal therapeutic systems (TTS) for passive and iontophoretic delivery across the skin. Polyvinyl pyrrolidone (PVP) and a mixture with polyvinyl alcohol (PVA) were employed as polymeric matrices. The study involved the TTS characterization in addition to quantification of the memantine transport across porcine skin in vitro. The evaluation of the TTS physical properties suggested that systems were made more mechanically resistant by including PVA (6%) or high concentrations of PVP (24%). Moreover, a linear correlation was observed between the concentration of PVP and the bioadhesion of the systems. Drug delivery experiments showed that the highest transdermal flux provided by a passive TTS (PVP 24% w/w limonene) was 8.89±0.81μgcm -2 h -1 whereas the highest iontophoretic transport was 46.4±3.6μgcm -2 h -1 . These innovative TTS would enable two dosage regimens that could lead to therapeutic plasma concentrations. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. Nanoparticles for the delivery of therapeutic antibodies: Dogma or promising strategy?

    Science.gov (United States)

    Sousa, Flávia; Castro, Pedro; Fonte, Pedro; Kennedy, Patrick J; Neves-Petersen, Maria Teresa; Sarmento, Bruno

    2017-10-01

    Over the past two decades, therapeutic antibodies have demonstrated promising results in the treatment of a wide array of diseases. However, the application of antibody-based therapy implies multiple administrations and a high cost of antibody production, resulting in costly therapy. Another disadvantage inherent to antibody-based therapy is the limited stability of antibodies and the low level of tissue penetration. The use of nanoparticles as delivery systems for antibodies allows for a reduction in antibody dosing and may represent a suitable alternative to increase antibody stability Areas covered: We discuss different nanocarriers intended for the delivery of antibodies as well as the corresponding encapsulation methods. Recent developments in antibody nanoencapsulation, particularly the possible toxicity issues that may arise from entrapment of antibodies into nanocarriers, are also assessed. In addition, this review will discuss the alterations in antibody structure and bioactivity that occur with nanoencapsulation. Expert opinion: Nanocarriers can protect antibodies from degradation, ensuring superior bioavailability. Encapsulation of therapeutic antibodies may offer some advantages, including potential targeting, reduced immunogenicity and controlled release. Furthermore, antibody nanoencapsulation may aid in the incorporation of the antibodies into the cells, if intracellular components (e.g. intracellular enzymes, oncogenic proteins, transcription factors) are to be targeted.

  6. Therapeutic misconception in early phase gene transfer trials.

    Science.gov (United States)

    Henderson, Gail E; Easter, Michele M; Zimmer, Catherine; King, Nancy M P; Davis, Arlene M; Rothschild, Barbra Bluestone; Churchill, Larry R; Wilfond, Benjamin S; Nelson, Daniel K

    2006-01-01

    Many subjects in early phase clinical trials expect to benefit in some way from the research intervention. It is understandable that people hope for improvement in their condition, no matter what the evidence. Yet unreasonable expectation of medical benefit may reflect problems with informed consent: Investigators may not disclose clearly that direct medical benefit from an early phase experimental intervention is unlikely or impossible, or subjects may not appreciate the differences between treatment and research. This paper presents findings from recent interviews with researchers and subjects and analysis of consent forms in early phase gene transfer research, a cutting-edge technology often called 'gene therapy'. We use three variables to construct a composite measure of therapeutic misconception TM, tapping misconceptions about the purposes of early phase research and the potential for direct medical benefit in these trials. Our multivariate model demonstrates the importance of both subject- and study-level factors as predictors of this TM index: education, disease type, and communication by study personnel about the likelihood of benefit. We hope that this work will deepen the discussion of how to define and measure TM, and refine the specification of factors that are related to subjects' TM.

  7. Robust Lentiviral Gene Delivery But Limited Transduction Capacity of Commonly Used Adeno-Associated Viral Serotypes in Xenotransplanted Human Skin.

    Science.gov (United States)

    Jakobsen, Maria; Askou, Anne Louise; Stenderup, Karin; Rosada, Cecilia; Dagnæs-Hansen, Frederik; Jensen, Thomas G; Corydon, Thomas J; Mikkelsen, Jacob Giehm; Aagaard, Lars

    2015-08-01

    Skin is an easily accessible organ, and therapeutic gene transfer to skin remains an attractive alternative for the treatment of skin diseases. Although we have previously documented potent lentiviral gene delivery to human skin, vectors based on adeno-associated virus (AAV) rank among the most promising gene delivery tools for in vivo purposes. Thus, we compared the potential usefulness of various serotypes of recombinant AAV vectors and lentiviral vectors for gene transfer to human skin in a xenotransplanted mouse model. Vector constructs encoding firefly luciferase were packaged in AAV capsids of serotype 1, 2, 5, 6, 8, and 9 and separately administered by intradermal injection in human skin transplants. For all serotypes, live bioimaging demonstrated low levels of transgene expression in the human skin graft, and firefly luciferase expression was observed primarily in neighboring tissue outside of the graft. In contrast, gene delivery by intradermally injected lentiviral vectors was efficient and led to extensive and persistent firefly luciferase expression within the human skin graft only. The study demonstrates the limited capacity of single-stranded AAV vectors of six commonly used serotypes for gene delivery to human skin in vivo.

  8. A unique carrier for delivery of therapeutic compounds beyond the blood-brain barrier.

    Directory of Open Access Journals (Sweden)

    Delara Karkan

    Full Text Available BACKGROUND: Therapeutic intervention in many neurological diseases is thwarted by the physical obstacle formed by the blood-brain barrier (BBB that excludes most drugs from entering the brain from the blood. Thus, identifying efficacious modes of drug delivery to the brain remains a "holy grail" in molecular medicine and nanobiotechnology. Brain capillaries, that comprise the BBB, possess an endogenous receptor that ferries an iron-transport protein, termed p97 (melanotransferrin, across the BBB. Here, we explored the hypothesis that therapeutic drugs "piggybacked" as conjugates of p97 can be shuttled across the BBB for treatment of otherwise inoperable brain tumors. APPROACH: Human p97 was covalently linked with the chemotherapeutic agents paclitaxel (PTAX or adriamycin (ADR and following intravenous injection, measured their penetration into brain tissue and other organs using radiolabeled and fluorescent derivatives of the drugs. In order to establish efficacy of the conjugates, we used nude mouse models to assess p97-drug conjugate activity towards glioma and mammary tumors growing subcutaneously compared to those growing intracranially. PRINCIPAL FINDINGS: Bolus-injected p97-drug conjugates and unconjugated p97 traversed brain capillary endothelium within a few minutes and accumulated to 1-2% of the injected by 24 hours. Brain delivery with p97-drug conjugates was quantitatively 10 fold higher than with free drug controls. Furthermore, both free-ADR and p97-ADR conjugates equally inhibited the subcutaneous growth of gliomas growing outside the brain. Evocatively, only p97-ADR conjugates significantly prolonged the survival of animals bearing intracranial gliomas or mammary tumors when compared to similar cumulated doses of free-ADR. SIGNIFICANCE: This study provides the initial proof of concept for p97 as a carrier capable of shuttling therapeutic levels of drugs from the blood to the brain for the treatment of neurological disorders

  9. Human gene therapy: novel approaches to improve the current gene delivery systems.

    Science.gov (United States)

    Cucchiarini, Magali

    2016-06-01

    Even though gene therapy made its way through the clinics to treat a number of human pathologies since the early years of experimental research and despite the recent approval of the first gene-based product (Glybera) in Europe, the safe and effective use of gene transfer vectors remains a challenge in human gene therapy due to the existence of barriers in the host organism. While work is under active investigation to improve the gene transfer systems themselves, the use of controlled release approaches may offer alternative, convenient tools of vector delivery to achieve a performant gene transfer in vivo while overcoming the various physiological barriers that preclude its wide use in patients. This article provides an overview of the most significant contributions showing how the principles of controlled release strategies may be adapted for human gene therapy.

  10. Efficient gene delivery using chitosan-polyethylenimine hybrid systems

    Energy Technology Data Exchange (ETDEWEB)

    Jiang, Hu-Lin; Kim, Tae-Hee; Kim, You-Kyoung; Park, In-Young; Cho, Chong-Su [Department of Agricultural Bioechnology, Seoul National University, Seoul 151-921 (Korea, Republic of); Cho, Myung-Haing [Laboratory of Toxicology, College of Veterinary Medicine, Seoul National University, Seoul 151-742 (Korea, Republic of)], E-mail: chocs@plaza.snu.ac.kr

    2008-06-01

    Chitosan and chitosan derivatives have been investigated as non-viral vectors because they have several advantages, such as biocompatibility, biodegradability, low cytotoxicity and low immunogenicity. However, low transfection efficiency and low cell specificity must be solved for their use in clinical trials. In this paper, chitosan-polyethylenimine (PEI) hybrid systems such as chitosan/PEI blend and chitosan-graft-PEI are described for efficient gene delivery because the PEI has high transfection efficiency owing to a proton sponge effect and chitosan has biocompatibility. Also, hepatocyte specificity of the galactosylated chitosan is explained after combination with PEI.

  11. Efficient gene delivery using chitosan-polyethylenimine hybrid systems

    International Nuclear Information System (INIS)

    Jiang, Hu-Lin; Kim, Tae-Hee; Kim, You-Kyoung; Park, In-Young; Cho, Chong-Su; Cho, Myung-Haing

    2008-01-01

    Chitosan and chitosan derivatives have been investigated as non-viral vectors because they have several advantages, such as biocompatibility, biodegradability, low cytotoxicity and low immunogenicity. However, low transfection efficiency and low cell specificity must be solved for their use in clinical trials. In this paper, chitosan-polyethylenimine (PEI) hybrid systems such as chitosan/PEI blend and chitosan-graft-PEI are described for efficient gene delivery because the PEI has high transfection efficiency owing to a proton sponge effect and chitosan has biocompatibility. Also, hepatocyte specificity of the galactosylated chitosan is explained after combination with PEI

  12. Therapeutic Efficacy of an ω-3-Fatty Acid-Containing 17-β Estradiol Nano-Delivery System against Experimental Atherosclerosis.

    Directory of Open Access Journals (Sweden)

    Dipti Deshpande

    Full Text Available Atherosclerosis and its consequences remain prevalent clinical challenges throughout the world. Initiation and progression of atherosclerosis involves a complex, dynamic interplay among inflammation, hyperlipidemia, and endothelial dysfunction. A multicomponent treatment approach targeted for delivery within diseased vessels could prove beneficial in treating atherosclerosis. This study was undertaken to evaluate the multimodal effects of a novel ω-3-fatty acid-rich, 17-β-estradiol (17-βE-loaded, CREKA-peptide-modified nanoemulsion system on experimental atherosclerosis. In vitro treatment of cultured human aortic endothelial cells (ECs with the 17-βE-loaded, CREKA-peptide-modified nanoemulsion system increased cellular nitrate/nitrite, indicating improved nitric oxide formation. In vivo, systemic administration of this nanoemulsion system to apolipoprotein-E knock out (ApoE-/- mice fed a high-fat diet significantly improved multiple parameters related to the etiology and development of occlusive atherosclerotic vasculopathy: lesion area, circulating plasma lipid levels, and expression of aortic-wall inflammatory markers. These salutary effects were attributed selectively to the 17-βE and/or ω-3 polyunsaturated fatty acid components of the nano-delivery system. At therapeutic doses, the 17-βE-loaded, CREKA-peptide modified nanoemulsion system appeared to be biocompatible in that it elicited no apparent adverse/toxic effects, as indexed by body weight, plasma alanine aminotransferase/aspartate aminotransferase levels, and liver and kidney histopathology. The study demonstrates the therapeutic potential of a novel, 17-βE-loaded, CREKA-peptide-modified nanoemulsion system against atherosclerosis in a multimodal fashion by reducing lesion size, lowering the levels of circulating plasma lipids and decreasing the gene expression of inflammatory markers associated with the disease.

  13. Ultrasound enhanced delivery of molecular imaging and therapeutic agents in Alzheimer's disease mouse models.

    Directory of Open Access Journals (Sweden)

    Scott B Raymond

    Full Text Available Alzheimer's disease is a neurodegenerative disorder typified by the accumulation of a small protein, beta-amyloid, which aggregates and is the primary component of amyloid plaques. Many new therapeutic and diagnostic agents for reducing amyloid plaques have limited efficacy in vivo because of poor transport across the blood-brain barrier. Here we demonstrate that low-intensity focused ultrasound with a microbubble contrast agent may be used to transiently disrupt the blood-brain barrier, allowing non-invasive, localized delivery of imaging fluorophores and immunotherapeutics directly to amyloid plaques. We administered intravenous Trypan blue, an amyloid staining red fluorophore, and anti-amyloid antibodies, concurrently with focused ultrasound therapy in plaque-bearing, transgenic mouse models of Alzheimer's disease with amyloid pathology. MRI guidance permitted selective treatment and monitoring of plaque-heavy anatomical regions, such as the hippocampus. Treated brain regions exhibited 16.5+/-5.4-fold increase in Trypan blue fluorescence and 2.7+/-1.2-fold increase in anti-amyloid antibodies that localized to amyloid plaques. Ultrasound-enhanced delivery was consistently reproduced in two different transgenic strains (APPswe:PSEN1dE9, PDAPP, across a large age range (9-26 months, with and without MR guidance, and with little or no tissue damage. Ultrasound-mediated, transient blood-brain barrier disruption allows the delivery of both therapeutic and molecular imaging agents in Alzheimer's mouse models, which should aid pre-clinical drug screening and imaging probe development. Furthermore, this technique may be used to deliver a wide variety of small and large molecules to the brain for imaging and therapy in other neurodegenerative diseases.

  14. Amphiphilic block co-polymers: preparation and application in nanodrug and gene delivery.

    Science.gov (United States)

    Xiong, Xiao-Bing; Binkhathlan, Ziyad; Molavi, Ommoleila; Lavasanifar, Afsaneh

    2012-07-01

    Self-assembly of amphiphilic block co-polymers composed of poly(ethylene oxide) (PEO) as the hydrophilic block and poly(ether)s, poly(amino acid)s, poly(ester)s and polypropyleneoxide (PPO) as the hydrophobic block can lead to the formation of nanoscopic structures of different morphologies. These structures have been the subject of extensive research in the past decade as artificial mimics of lipoproteins and viral vectors for drug and gene delivery. The aim of this review is to provide an overview of the synthesis of commonly used amphiphilic block co-polymers. It will also briefly go over some pharmaceutical applications of amphiphilic block co-polymers as "nanodelivery systems" for small molecules and gene therapeutics. Copyright © 2012 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  15. Gene Delivery into Plant Cells for Recombinant Protein Production

    Directory of Open Access Journals (Sweden)

    Qiang Chen

    2015-01-01

    Full Text Available Recombinant proteins are primarily produced from cultures of mammalian, insect, and bacteria cells. In recent years, the development of deconstructed virus-based vectors has allowed plants to become a viable platform for recombinant protein production, with advantages in versatility, speed, cost, scalability, and safety over the current production paradigms. In this paper, we review the recent progress in the methodology of agroinfiltration, a solution to overcome the challenge of transgene delivery into plant cells for large-scale manufacturing of recombinant proteins. General gene delivery methodologies in plants are first summarized, followed by extensive discussion on the application and scalability of each agroinfiltration method. New development of a spray-based agroinfiltration and its application on field-grown plants is highlighted. The discussion of agroinfiltration vectors focuses on their applications for producing complex and heteromultimeric proteins and is updated with the development of bridge vectors. Progress on agroinfiltration in Nicotiana and non-Nicotiana plant hosts is subsequently showcased in context of their applications for producing high-value human biologics and low-cost and high-volume industrial enzymes. These new advancements in agroinfiltration greatly enhance the robustness and scalability of transgene delivery in plants, facilitating the adoption of plant transient expression systems for manufacturing recombinant proteins with a broad range of applications.

  16. Self-Assembling Multifunctional Peptide Dimers for Gene Delivery Systems

    Directory of Open Access Journals (Sweden)

    Kitae Ryu

    2015-01-01

    Full Text Available Self-assembling multifunctional peptide was designed for gene delivery systems. The multifunctional peptide (MP consists of cellular penetrating peptide moiety (R8, matrix metalloproteinase-2 (MMP-2 specific sequence (GPLGV, pH-responsive moiety (H5, and hydrophobic moiety (palmitic acid (CR8GPLGVH5-Pal. MP was oxidized to form multifunctional peptide dimer (MPD by DMSO oxidation of thiols in terminal cysteine residues. MPD could condense pDNA successfully at a weight ratio of 5. MPD itself could self-assemble into submicron micelle particles via hydrophobic interaction, of which critical micelle concentration is about 0.01 mM. MPD showed concentration-dependent but low cytotoxicity in comparison with PEI25k. MPD polyplexes showed low transfection efficiency in HEK293 cells expressing low level of MMP-2 but high transfection efficiency in A549 and C2C12 cells expressing high level of MMP-2, meaning the enhanced transfection efficiency probably due to MMP-induced structural change of polyplexes. Bafilomycin A1-treated transfection results suggest that the transfection of MPD is mediated via endosomal escape by endosome buffering ability. These results show the potential of MPD for MMP-2 targeted gene delivery systems due to its multifunctionality.

  17. [Application of ultrasound-enhanced gene and drug delivery to the ocular tissue].

    Science.gov (United States)

    Sonoda, Shozo; Yamashita, Toshifumi; Suzuki, Ryo; Maruyama, Kazuo; Sakamoto, Taiji

    2013-01-01

    Visual images provide an immensely rich source of information about the external world. Eye has characteristic structure sensory cells are arranged along the eye wall, and is filled inside with vitreous body. In recent years, intravitreal injection of anti-vascular endothelial growth factor (VEGF) agent had widely spread, and numerous number of patients who suffered ocular angiogenic disease such as diabetic retinopathy, age-related macular degeneration and retinal vascular occlusion for the disease, were treated and spared the blindness. Vitreous cavity was regarded as reservoir of drug, intravitreal injection is thought a sort of drug delivery. However, with regard to the administration of a selective drug deliver, it has not yet been solved. Our aim is to establish a new method of gene transfer, drug delivery using low-energy ultrasound to the eye, to date, we confirmed drug and gene deliver to the ocular tissue such as cornea, conjunctiva and retina with high efficiency. In addition, tissue damage was minimal. We have also shown that ultrasound irradiation with combination of a microbubbles or bubble liposome could be introduced drug and gene more effectively. Based on these knowledge, we will focus on development of a new device for intraocular ultrasound exposure and potential for therapeutic application of ultrasound to humans retinal disease such as retinal artery obstruction.

  18. Gene delivery to the lungs: pulmonary gene therapy for cystic fibrosis.

    Science.gov (United States)

    Villate-Beitia, Ilia; Zarate, Jon; Puras, Gustavo; Pedraz, José Luis

    2017-07-01

    Cystic fibrosis (CF) is a monogenic autosomal recessive disorder where the defective gene, the cystic fibrosis transmembrane conductance regulator (CFTR), is well identified. Moreover, the respiratory tract can be targeted through noninvasive aerosolized formulations for inhalation. Therefore, gene therapy is considered a plausible strategy to address this disease. Conventional gene therapy strategies rely on the addition of a correct copy of the CFTR gene into affected cells in order to restore the channel activity. In recent years, genome correction strategies have emerged, such as zinc-finger nucleases, transcription activator-like effector nucleases and clustered regularly interspaced short palindromic repeats associated to Cas9 nucleases. These gene editing tools aim to repair the mutated gene at its original genomic locus with high specificity. Besides, the success of gene therapy critically depends on the nucleic acids carriers. To date, several clinical studies have been carried out to add corrected copies of the CFTR gene into target cells using viral and non-viral vectors, some of them with encouraging results. Regarding genome editing systems, preliminary in vitro studies have been performed in order to repair the CFTR gene. In this review, after briefly introducing the basis of CF, we discuss the up-to-date gene therapy strategies to address the disease. The review focuses on the main factors to take into consideration when developing gene delivery strategies, such as the design of vectors and plasmid DNA, in vitro/in vivo tests, translation to human use, administration methods, manufacturing conditions and regulatory issues.

  19. Tumor Microenvironment Gene Signature as a Prognostic Classifier and Therapeutic Target

    Science.gov (United States)

    2016-06-01

    AWARD NUMBER: W81XWH-14-1-0107 TITLE: Tumor Microenvironment Gene Signature as a Prognostic Classifier and Therapeutic Target PRINCIPAL...AND SUBTITLE Tumor Microenvironment Gene Signature as a 5a. CONTRACT NUMBER W81XWH-14-1-0107 Prognostic Classifier and Therapeutic Target 5b...gene signature that correlates with poor survival in ovarian cancer patients. We are refining this gene signature to develop biomarkers for the

  20. Intracellular Delivery of Proteins with Cell-Penetrating Peptides for Therapeutic Uses in Human Disease.

    Science.gov (United States)

    Dinca, Ana; Chien, Wei-Ming; Chin, Michael T

    2016-02-22

    Protein therapy exhibits several advantages over small molecule drugs and is increasingly being developed for the treatment of disorders ranging from single enzyme deficiencies to cancer. Cell-penetrating peptides (CPPs), a group of small peptides capable of promoting transport of molecular cargo across the plasma membrane, have become important tools in promoting the cellular uptake of exogenously delivered proteins. Although the molecular mechanisms of uptake are not firmly established, CPPs have been empirically shown to promote uptake of various molecules, including large proteins over 100 kiloDaltons (kDa). Recombinant proteins that include a CPP tag to promote intracellular delivery show promise as therapeutic agents with encouraging success rates in both animal and human trials. This review highlights recent advances in protein-CPP therapy and discusses optimization strategies and potential detrimental effects.

  1. Preparation and characterization of magnetic gene vectors for targeting gene delivery

    Energy Technology Data Exchange (ETDEWEB)

    Zheng, S.W.; Liu, G. [College of Chemistry, Chemical Engineering and Materials Science and Key Laboratory of Organic Synthesis of Jiangsu Province, Soochow University, SIP, Suzhou 215123 (China); Hong, R.Y., E-mail: rhong@suda.edu.cn [College of Chemistry, Chemical Engineering and Materials Science and Key Laboratory of Organic Synthesis of Jiangsu Province, Soochow University, SIP, Suzhou 215123 (China); State Key Laboratory of Multi-phase Complex Systems, Institute of Process Engineering, Chinese Academy of Sciences, Beijing 100080 (China); Li, H.Z. [State Key Laboratory of Multi-phase Complex Systems, Institute of Process Engineering, Chinese Academy of Sciences, Beijing 100080 (China); Li, Y.G., E-mail: ilguoliang@sohu.com [Department of radiology, the First Affiliated Hospital of Soochow University, Suzhou 215007 (China); Wei, D.G., E-mail: dougwei@deas.harvard.edu [Center for Nanoscale Systems, School of Engineering and Applied Science, Harvard University, 11 Oxford Street, Cambridge, MA 02139 (United States)

    2012-10-15

    Highlights: Black-Right-Pointing-Pointer PEI is ideal candidate polymer for the design of gene delivery systems. Black-Right-Pointing-Pointer PEI-CMD-MNPs exhibited a typical superparamagnetic behavior. Black-Right-Pointing-Pointer PEI-CMD-MNPs were well stable over the entire range of pH and NaCl concentration. Black-Right-Pointing-Pointer DNA-PEI-CMD-MNPs transfected cells by a magnet have higher transfection efficiency and gene expression efficiency. - Abstract: The PEI-CMD-MNPs were successfully prepared by the surface modification of magnetic Fe{sub 3}O{sub 4} nanoparticles with carboxymethyl dextran (CMD) and polyethyleneimine (PEI). The PEI-CMD-MNPs polyplexes exhibited a typical superparamagnetic behavior and were well stable over the entire range of pH and NaCl concentration. These PEI-CMD-MNPs were used as magnetic gene vectors for targeting gene delivery. The prepared MNPs at different surface modification stages were characterized using Fourier transform infrared (FT-IR), thermogravimetric analysis (TGA), field emissions canning electron microscopy (FE-SEM), powder X-ray diffraction (XRD) and dynamic laser light scattering (DLS) analysis. The magnetic properties were studied by vibrating sample magnetometer (VSM). To evaluate the performance of the magnetic nanoparticles as gene transfer vector, the PEI-CMD-MNPs were used to delivery green fluorescent protein (GFP) gene into BHK21 cells. The expression of GFP gene was detected by fluorescence microscope. DNA-PEI-CMD-MNPs polyplexes absorbed by the cells were also monitored by Magnetic resonance imaging (MRI). The transfection efficiency and gene expression efficiency of that transfected with a magnet were much higher than that of standard transfection.

  2. Colon-targeted delivery of piceatannol enhances anti-colitic effects of the natural product: potential molecular mechanisms for therapeutic enhancement

    Directory of Open Access Journals (Sweden)

    Yum S

    2015-08-01

    Full Text Available Soohwan Yum, Seongkeun Jeong, Sunyoung Lee, Joon Nam, Wooseong Kim, Jin-Wook Yoo, Min-Soo Kim, Bok Luel Lee, Yunjin Jung College of Pharmacy, Pusan National University, Busan, Republic of Korea Abstract: Piceatannol (PCT, an anti-colitic natural product, undergoes extensive Phase II hepatic metabolism, resulting in very low bioavailability. We investigated whether colon-targeted delivery of PCT could enhance anti-colitic effects and how therapeutic enhancement occurred at the molecular level. Molecular effects of PCT were examined in human colon carcinoma cells and inflamed colons. The anti-colitic effects of PCT in a colon-targeted capsule (colon-targeted PCT were compared with PCT in a gelatin capsule (conventional PCT in a trinitrobenzene sulfonic acid-induced rat colitis model. Colon-targeted PCT elicited greatly enhanced recovery of the colonic inflammation. In HCT116 cells, PCT inhibited nuclear factor kappaB while activating anti-colitic transcription factors, nuclear factor-erythroid 2 (NF-E2 p45-related factor 2, and hypoxia-inducible factor-1. Colon-targeted PCT, but not conventional PCT, modulated production of the target gene products of the transcription factors in the inflamed colonic tissues. Rectal administration of PCT, which simulates the therapeutic action of colon-targeted PCT, also ameliorated rat colitis and reproduced the molecular effects in the inflamed colonic tissues. Colon-targeted delivery increased therapeutic efficacy of PCT against colitis, likely resulting from multitargeted effects exerted by colon-targeted PCT. The drug delivery technique may be useful for therapeutic optimization of anti-colitic lead compounds including natural products. Keywords: piceatannol, colitis, colon-targeted delivery, multitarget, polypharmacology

  3. Lipid Phases Eye View to Lipofection. Cationic Phosphatidylcholine Derivatives as Efficient DNA Carriers for Gene Delivery

    OpenAIRE

    Rumiana Koynova

    2008-01-01

    Efficient delivery of genetic material to cells is needed for tasks of utmost importance in laboratory and clinic, such as gene transfection and gene silencing. Synthetic cationic lipids can be used as delivery vehicles for nucleic acids and are now considered the most promising non-viral gene carriers. They form complexes (lipoplexes) with the polyanionic nucleic acids. A critical obstacle for clinical application of the lipid-mediated DNA delivery (lipofection) is its unsatisfactory efficie...

  4. Design of dendrimer-based drug delivery nanodevices with enhanced therapeutic efficacies

    Science.gov (United States)

    Kannan, Rangaramanujam

    2007-03-01

    Dendrimers and hyperbranched polymers possess highly branched architectures, with a large number of controllable, tailorable, `peripheral' functionalities. Since the surface chemistry of these materials can be modified with relative ease, these materials have tremendous potential in targeted drug delivery. They have significant potential compared to liposomes and nanoparticles, because of the reduced macrophage update, increased cellular transport, and the ability to modulate the local environment through functional groups. We are developing nanodevices based on dendritic systems for drug delivery, that contain a high drug payload, ligands, and imaging agents, resulting in `smart' drug delivery devices that can target, deliver, and signal. In collaboration with the Children's Hospital of Michigan, Karmanos Cancer Institute, and College of Pharmacy, we are testing the in vitro and in vivo response of these nanodevices, by adapting the chemistry for specific clinical applications such as asthma and cancer. These materials are characterized by UV/Vis spectroscopy, flow cytometry, fluorescence/confocal microscopy, and appropriate animal models. Our results suggest that: (1) We can prepare drug-dendrimer conjugates with drug payloads of greater than 50%, for a variety of drugs; (2) The dendritic polymers are capable of transporting and delivering drugs into cells faster than free drugs, with superior therapeutic efficiency. This can be modulated by the surface functionality of the dendrimer; (3) For chemotherapy drugs, the conjugates are a factor of 6-20 times more effective even in drug-resistant cell lines; (4) For corticosteroidal drugs, the dendritic polymers provide higher drug residence times in the lung, allowing for passive targeting. The ability of the drug-dendrimer-ligand conjugates to target specific asthma and cancer cells is currently being explored using in vitro and in vivo animal models.

  5. Perinatal systemic gene delivery using adeno-associated viral vectors

    Directory of Open Access Journals (Sweden)

    Rajvinder eKarda

    2014-11-01

    Full Text Available Neurodegenerative monogenic diseases can also affect a broad range of tissues and organs throughout the body. An effective treatment would require a systemic approach. The intravenous administration of novel therapies is ideal but is hampered by the inability of such drugs to cross the blood-brain barrier and precludes efficacy in the central nervous system. A number of these early lethal intractable diseases also present devastating irreversible pathology at birth or soon after. Therefore, any therapy would ideally be administered during the perinatal period to prevent, stop or ameliorate disease progression. The concept of perinatal gene therapy has moved a step further towards being a feasible approach to treating such disorders. This has primarily been driven by the recent discoveries that particular serotypes of adeno-associated virus (AAV gene delivery vectors have the ability to cross the blood-brain barrier following intravenous administration. Furthermore, this has been safely demonstrated in perinatal mice and non-human primates. This review focuses on the progress made in using AAV to achieve systemic transduction and what this means for developing perinatal gene therapy for early lethal neurodegenerative diseases.

  6. Nebulisation of receptor-targeted nanocomplexes for gene delivery to the airway epithelium.

    Science.gov (United States)

    Manunta, Maria D I; McAnulty, Robin J; Tagalakis, Aristides D; Bottoms, Stephen E; Campbell, Frederick; Hailes, Helen C; Tabor, Alethea B; Laurent, Geoffrey J; O'Callaghan, Christopher; Hart, Stephen L

    2011-01-01

    Gene therapy mediated by synthetic vectors may provide opportunities for new treatments for cystic fibrosis (CF) via aerosolisation. Vectors for CF must transfect the airway epithelium efficiently and not cause inflammation so they are suitable for repeated dosing. The inhaled aerosol should be deposited in the airways since the cystic fibrosis transmembrane conductance regulator gene (CFTR) is expressed predominantly in the epithelium of the submucosal glands and in the surface airway epithelium. The aim of this project was to develop an optimised aerosol delivery approach applicable to treatment of CF lung disease by gene therapy. The vector suspension investigated in this study comprises receptor-targeting peptides, cationic liposomes and plasmid DNA that self-assemble by electrostatic interactions to form a receptor-targeted nanocomplex (RTN) of approximately 150 nm with a cationic surface charge of +50 mV. The aerodynamic properties of aerosolised nanocomplexes produced with three different nebulisers were compared by determining aerosol deposition in the different stages of a Next Generation Pharmaceutical Impactor (NGI). We also investigated the yield of intact plasmid DNA by agarose gel electrophoresis and densitometry, and transfection efficacies in vitro and in vivo. RTNs nebulised with the AeroEclipse II BAN were the most effective, compared to other nebulisers tested, for gene delivery both in vitro and in vivo. The biophysical properties of the nanocomplexes were unchanged after nebulisation while the deposition of RTNs suggested a range of aerosol aerodynamic sizes between 5.5 µm-1.4 µm cut off (NGI stages 3-6) compatible with deposition in the central and lower airways. RTNs showed their ability at delivering genes via nebulisation, thus suggesting their potential applications for therapeutic interventions of cystic fibrosis and other respiratory disorders.

  7. Nebulisation of receptor-targeted nanocomplexes for gene delivery to the airway epithelium.

    Directory of Open Access Journals (Sweden)

    Maria D I Manunta

    Full Text Available Gene therapy mediated by synthetic vectors may provide opportunities for new treatments for cystic fibrosis (CF via aerosolisation. Vectors for CF must transfect the airway epithelium efficiently and not cause inflammation so they are suitable for repeated dosing. The inhaled aerosol should be deposited in the airways since the cystic fibrosis transmembrane conductance regulator gene (CFTR is expressed predominantly in the epithelium of the submucosal glands and in the surface airway epithelium. The aim of this project was to develop an optimised aerosol delivery approach applicable to treatment of CF lung disease by gene therapy.The vector suspension investigated in this study comprises receptor-targeting peptides, cationic liposomes and plasmid DNA that self-assemble by electrostatic interactions to form a receptor-targeted nanocomplex (RTN of approximately 150 nm with a cationic surface charge of +50 mV. The aerodynamic properties of aerosolised nanocomplexes produced with three different nebulisers were compared by determining aerosol deposition in the different stages of a Next Generation Pharmaceutical Impactor (NGI. We also investigated the yield of intact plasmid DNA by agarose gel electrophoresis and densitometry, and transfection efficacies in vitro and in vivo.RTNs nebulised with the AeroEclipse II BAN were the most effective, compared to other nebulisers tested, for gene delivery both in vitro and in vivo. The biophysical properties of the nanocomplexes were unchanged after nebulisation while the deposition of RTNs suggested a range of aerosol aerodynamic sizes between 5.5 µm-1.4 µm cut off (NGI stages 3-6 compatible with deposition in the central and lower airways.RTNs showed their ability at delivering genes via nebulisation, thus suggesting their potential applications for therapeutic interventions of cystic fibrosis and other respiratory disorders.

  8. Ionically crosslinked alginate–carboxymethyl cellulose beads for the delivery of protein therapeutics

    International Nuclear Information System (INIS)

    Kim, Min Sup; Park, Sang Jun; Gu, Bon Kang; Kim, Chun-Ho

    2012-01-01

    Highlights: ► We prepared Fe 3+ crosslinked alginate–carboxymethyl cellulose (AC) beads. ► Different surface and inner morphology of AC beads were observed on volume of CMC. ► AC beads showed minimum swelling degree in acidic condition. ► Protein release from AC beads was to control in gastrointestinal condition. - Abstract: We developed Fe 3+ -crosslinked alginate–carboxymethyl cellulose (AC) beads in various volume ratios by dropping an AC solution into a ferric chloride solution to form protein therapeutic carrier beads. Scanning electron microscopy revealed that the roughness and pore size of the crosslinked beads increased with the volume ratio of the carboxymethyl cellulose. Fourier transform-infrared analysis revealed the formation of a three-dimensional bonding structure between the anionic polymeric chains of AC and the Fe 3+ ions. The degree of swelling and the release profile of albumin from the beads were investigated under simulated gastrointestinal conditions (pH 1.2, 4.5, and 7.4). The Fe 3+ -crosslinked AC beads displayed different degrees of swelling and albumin release for the various AC volume ratios and under various pH conditions. An in vitro release test was used to monitor the controlled release of albumin from the AC beads under simulated gastrointestinal conditions over 24 h. The Fe 3+ -crosslinked AC beads protected and controlled the release of protein, demonstrating that such beads present a promising protein therapeutic carrier for the oral delivery.

  9. "Clicking" Gene Therapeutics: A Successful Union of Chemistry and Biomedicine for New Solutions

    DEFF Research Database (Denmark)

    Astakhova, Kira; Ray, Roslyn; Taskova, Maria

    2018-01-01

    The use of nucleic acid, DNA and RNA, based strategies to disrupt gene expression as a therapeutic is quickly emerging. Indeed, synthetic oligonucleotides represent a major component of modern gene therapeutics. However, the efficiency and specificity of intracellular uptake for nonmodified oligo...

  10. Delivery of Therapeutic Proteins via Extracellular Vesicles: Review and Potential Treatments for Parkinson's Disease, Glioma, and Schwannoma.

    Science.gov (United States)

    Hall, Justin; Prabhakar, Shilpa; Balaj, Leonora; Lai, Charles P; Cerione, Richard A; Breakefield, Xandra O

    2016-04-01

    Extracellular vesicles present an attractive delivery vehicle for therapeutic proteins. They intrinsically contain many proteins which can provide information to other cells. Advantages include reduced immune reactivity, especially if derived from the same host, stability in biologic fluids, and ability to target uptake. Those from mesenchymal stem cells appear to be intrinsically therapeutic, while those from cancer cells promote tumor progression. Therapeutic proteins can be loaded into vesicles by overexpression in the donor cell, with oligomerization and membrane sequences increasing their loading. Examples of protein delivery for therapeutic benefit in pre-clinical models include delivery of: catalase for Parkinson's disease to reduce oxidative stress and thus help neurons to survive; prodrug activating enzymes which can convert a prodrug which crosses the blood-brain barrier into a toxic chemotherapeutic drug for schwannomas and gliomas; and the apoptosis-inducing enzyme, caspase-1 under a Schwann cell specific promoter for schwannoma. This therapeutic delivery strategy is novel and being explored for a number of diseases.

  11. A novel therapeutic strategy for cartilage diseases based on lipid nanoparticle-RNAi delivery system

    Science.gov (United States)

    Wang, Shaowei; Wei, Xiaochun; Sun, Xiaojuan; Chen, Chongwei; Zhou, Jingming; Zhang, Ge; Wu, Heng; Guo, Baosheng

    2018-01-01

    Background Cartilage degeneration affects millions of people but preventing its degeneration is a big challenge. Although RNA interference (RNAi) has been used in human trials via silencing specific genes, the cartilage RNAi has not been possible to date because the cartilage is an avascular and very dense tissue with very low permeability. Purpose The objective of this study was to develop and validate a novel lipid nanoparticle (LNP)-siRNA delivery system that can prevent cartilage degeneration by knocking down specific genes. Methods LNP transfection efficiency was evaluated in vitro and ex vivo. Indian Hedgehog (Ihh) has been correlated with cartilage degeneration. The in vivo effects of LNP-Ihh siRNA complexes on cartilage degeneration were evaluated in a rat model of surgery-induced osteoarthritis (OA). Results In vitro, 100% of chondrocytes were transfected with siRNA in the LNP-siRNA group. In accordance with the cell culture results, red positive signals could be detected even in the deep layer of cartilage tissue cultures treated by LNP-beacon. In vivo data showed that LNP is specific for cartilage, since positive signals were detected by fluorescence molecular tomography and confocal microscopy in joint cartilage injected with LNP-beacon, but not on the surface of the synovium. In the rat model of OA, intraarticular injection of LNP-Ihh siRNA attenuated OA progression, and PCR results showed LNP-Ihh siRNA exerted a positive impact on anabolic metabolism and negative impact on catabolic metabolism. Conclusion This study demonstrates that our LNP-RNAi delivery system has a significantly chondroprotective effect that attenuates cartilage degeneration and holds great promise as a powerful tool for treatment of cartilage diseases by knocking down specific genes. PMID:29440889

  12. A novel therapeutic strategy for cartilage diseases based on lipid nanoparticle-RNAi delivery system.

    Science.gov (United States)

    Wang, Shaowei; Wei, Xiaochun; Sun, Xiaojuan; Chen, Chongwei; Zhou, Jingming; Zhang, Ge; Wu, Heng; Guo, Baosheng; Wei, Lei

    2018-01-01

    Cartilage degeneration affects millions of people but preventing its degeneration is a big challenge. Although RNA interference (RNAi) has been used in human trials via silencing specific genes, the cartilage RNAi has not been possible to date because the cartilage is an avascular and very dense tissue with very low permeability. The objective of this study was to develop and validate a novel lipid nanoparticle (LNP)-siRNA delivery system that can prevent cartilage degeneration by knocking down specific genes. LNP transfection efficiency was evaluated in vitro and ex vivo. Indian Hedgehog ( Ihh ) has been correlated with cartilage degeneration. The in vivo effects of LNP-Ihh siRNA complexes on cartilage degeneration were evaluated in a rat model of surgery-induced osteoarthritis (OA). In vitro, 100% of chondrocytes were transfected with siRNA in the LNP-siRNA group. In accordance with the cell culture results, red positive signals could be detected even in the deep layer of cartilage tissue cultures treated by LNP-beacon. In vivo data showed that LNP is specific for cartilage, since positive signals were detected by fluorescence molecular tomography and confocal microscopy in joint cartilage injected with LNP-beacon, but not on the surface of the synovium. In the rat model of OA, intraarticular injection of LNP-Ihh siRNA attenuated OA progression, and PCR results showed LNP-Ihh siRNA exerted a positive impact on anabolic metabolism and negative impact on catabolic metabolism. This study demonstrates that our LNP-RNAi delivery system has a significantly chondroprotective effect that attenuates cartilage degeneration and holds great promise as a powerful tool for treatment of cartilage diseases by knocking down specific genes.

  13. Neutrophil-Mediated Delivery of Therapeutic Nanoparticles across Blood Vessel Barrier for Treatment of Inflammation and Infection

    OpenAIRE

    Chu, Dafeng; Gao, Jin; Wang, Zhenjia

    2015-01-01

    Endothelial cells form a monolayer in lumen of blood vessels presenting a great barrier for delivery of therapeutic nanoparticles (NPs) into extravascular tissues where most diseases occur, such as inflammation disorders and infection. Here, we report a strategy for delivering therapeutic NPs across this blood vessel barrier by nanoparticle in situ hitchhiking activated neutrophils. Using intravital microscopy of TNF-α-induced inflammation of mouse cremaster venules and a mouse model of acute...

  14. Optimization of conditions for gene delivery system based on PEI

    Directory of Open Access Journals (Sweden)

    Roya Cheraghi

    2017-01-01

    Full Text Available Objective(s: PEI based nanoparticle (NP due to dual capabilities of proton sponge and DNA binding is known as powerful tool for nucleic acid delivery to cells. However, serious cytotoxicity and complicated conditions, which govern NPs properties and its interactions with cells practically, hindered achievement to high transfection efficiency. Here, we have tried to optimize the properties of PEI/ firefly luciferase plasmid complexes and cellular condition to improve transfection efficiency. Materials and Methods: For this purpose, firefly luciferase, as a robust gene reporter, was complexed with PEI to prepare NPs with different size and charge. The physicochemical properties of nanoparticles were evaluated using agarose gel retardation and dynamic light scattering.  MCF7 and BT474 cells at different confluency were also transfected with prepared nanoparticles at various concentrations for short and long times. Results: The branched PEI can instantaneously bind to DNA and form cationic NPs. The results demonstrated the production of nanoparticles with size about 100-500 nm dependent on N/P ratio. Moreover, increase of nanoparticles concentration on the cell surface drastically improved the transfection rate, so at a concentration of 30 ng/ìl, the highest transfection efficiency was achieved. On the other side, at confluency between 40-60%, the maximum efficiency was obtained. The result demonstrated that N/P ratio of 12 could establish an optimized ratio between transfection efficiency and cytotoxicity of PEI/plasmid nanoparticles. The increase of NPs N/P ratio led to significant cytotoxicity. Conclusion: Obtained results verified the optimum conditions for PEI based gene delivery in different cell lines.

  15. A novel therapeutic strategy for cartilage diseases based on lipid nanoparticle-RNAi delivery system

    Directory of Open Access Journals (Sweden)

    Wang S

    2018-01-01

    Full Text Available Shaowei Wang,1 Xiaochun Wei,1 Xiaojuan Sun,1 Chongwei Chen,1 Jingming Zhou,2 Ge Zhang,3 Heng Wu,3 Baosheng Guo,3 Lei Wei1,2 1Department of Orthopaedics, The 2nd Hospital of Shanxi Medical University, Taiyuan, Shanxi, China; 2Department of Orthopaedics, Rhode Island Hospital, The Warren Alpert Medical School of Brown University, Providence, RI, USA; 3Integrated Traditional Chinese and Western Medicine, School of Chinese Medicine, Hong Kong Baptist University, Hong Kong Background: Cartilage degeneration affects millions of people but preventing its degeneration is a big challenge. Although RNA interference (RNAi has been used in human trials via silencing specific genes, the cartilage RNAi has not been possible to date because the cartilage is an avascular and very dense tissue with very low permeability. Purpose: The objective of this study was to develop and validate a novel lipid nanoparticle (LNP-siRNA delivery system that can prevent cartilage degeneration by knocking down specific genes. Methods: LNP transfection efficiency was evaluated in vitro and ex vivo. Indian Hedgehog (Ihh has been correlated with cartilage degeneration. The in vivo effects of LNP-Ihh siRNA complexes on cartilage degeneration were evaluated in a rat model of surgery-induced osteoarthritis (OA. Results: In vitro, 100% of chondrocytes were transfected with siRNA in the LNP-siRNA group. In accordance with the cell culture results, red positive signals could be detected even in the deep layer of cartilage tissue cultures treated by LNP-beacon. In vivo data showed that LNP is specific for cartilage, since positive signals were detected by fluorescence molecular tomography and confocal microscopy in joint cartilage injected with LNP-beacon, but not on the surface of the synovium. In the rat model of OA, intraarticular injection of LNP-Ihh siRNA attenuated OA progression, and PCR results showed LNP-Ihh siRNA exerted a positive impact on anabolic metabolism and negative

  16. A Review of Gene Delivery and Stem Cell Based Therapies for Regenerating Inner Ear Hair Cells

    Directory of Open Access Journals (Sweden)

    Michael S. Detamore

    2011-09-01

    Full Text Available Sensory neural hearing loss and vestibular dysfunction have become the most common forms of sensory defects, affecting millions of people worldwide. Developing effective therapies to restore hearing loss is challenging, owing to the limited regenerative capacity of the inner ear hair cells. With recent advances in understanding the developmental biology of mammalian and non-mammalian hair cells a variety of strategies have emerged to restore lost hair cells are being developed. Two predominant strategies have developed to restore hair cells: transfer of genes responsible for hair cell genesis and replacement of missing cells via transfer of stem cells. In this review article, we evaluate the use of several genes involved in hair cell regeneration, the advantages and disadvantages of the different viral vectors employed in inner ear gene delivery and the insights gained from the use of embryonic, adult and induced pluripotent stem cells in generating inner ear hair cells. Understanding the role of genes, vectors and stem cells in therapeutic strategies led us to explore potential solutions to overcome the limitations associated with their use in hair cell regeneration.

  17. A review of gene delivery and stem cell based therapies for regenerating inner ear hair cells.

    Science.gov (United States)

    Devarajan, Keerthana; Staecker, Hinrich; Detamore, Michael S

    2011-09-13

    Sensory neural hearing loss and vestibular dysfunction have become the most common forms of sensory defects, affecting millions of people worldwide. Developing effective therapies to restore hearing loss is challenging, owing to the limited regenerative capacity of the inner ear hair cells. With recent advances in understanding the developmental biology of mammalian and non-mammalian hair cells a variety of strategies have emerged to restore lost hair cells are being developed. Two predominant strategies have developed to restore hair cells: transfer of genes responsible for hair cell genesis and replacement of missing cells via transfer of stem cells. In this review article, we evaluate the use of several genes involved in hair cell regeneration, the advantages and disadvantages of the different viral vectors employed in inner ear gene delivery and the insights gained from the use of embryonic, adult and induced pluripotent stem cells in generating inner ear hair cells. Understanding the role of genes, vectors and stem cells in therapeutic strategies led us to explore potential solutions to overcome the limitations associated with their use in hair cell regeneration.

  18. Gene Editing and CRISPR Therapeutics: Strategies Taught by Cell and Gene Therapy.

    Science.gov (United States)

    Ramirez, Juan C

    2017-01-01

    A few years ago, we assisted in the demonstration for the first time of the revolutionary idea of a type of adaptive-immune system in the bacteria kingdom. This system, named CRISPR, and variants engineered in the lab, have been demonstrated as functional with extremely high frequency and fidelity in almost all eukaryotic cells studied to date. The capabilities of this RNA-guided nuclease have added to the interest that was announced with the advent of previous technologies for genome editing tools, such as ZFN and TALEN. The capabilities exhibited by these gene editors, opens up a novel scenario that indicates the promise of a next-generation medicine based on precision and personalized objectives, mostly due to the change in the paradigm regarding gene-surgery. This has certainly attracted, like never before, the attention of the biotech business and investor community. This chapter offers a brief overview of some of the factors that have contributed to a rapid entry into the biotech and pharmaceutical company's pipeline, focusing on how cell and gene therapies (CGT), collectively known as advanced therapies, have become the driving forces toward the therapeutic uses of gene editing technology. The sum of all those efforts for more than 30years has contributed to the new paradigm of considering genes as medicines. Copyright © 2017. Published by Elsevier Inc.

  19. Therapeutics: Gene Therapy for Alpha-1 Antitrypsin Deficiency.

    Science.gov (United States)

    Gruntman, Alisha M; Flotte, Terence R

    2017-01-01

    This review seeks to give an overview of alpha-1 antitrypsin deficiency, including the different disease phenotypes that it encompasses. We then describe the different therapeutic endeavors that have been undertaken to address these different phenotypes. Lastly we discuss future potential therapeutics, such as genome editing, and how they may play a role in treating alpha-1 antitrypsin deficiency.

  20. Ultrasound-Stimulated Drug Delivery Using Therapeutic Reconstituted High-Density Lipoprotein Nanoparticles.

    Science.gov (United States)

    Xiong, Fangyuan; Nirupama, Sabnis; Sirsi, Shashank R; Lacko, Andras; Hoyt, Kenneth

    2017-01-01

    method for monitoring the effects of US-stimulated drug delivery of IR-780 dye-loaded rHDL NPs in living animals. No change in optical imaging data was found in the control animals. However, there was considerable dye accumulation (surrogate drug) within 48 h in the low (5 µg), moderate (10 µg), and high (50 µg) rHDL NP concentration-dosed group animals ( p 0.69, p < 0.003). IR-780 dye extraction from the tumor tissue samples confirmed the in vivo and ex vivo US therapy findings. Overall, the addition of US therapy considerably improved local rHDL NP accumulation in tumor tissue. This study concludes that US-mediated drug delivery can facilitate tumor uptake of rHDL NPs and more research is warranted to optimize the drug dosing schedule and the respective therapeutic protocols.

  1. Cholesterol tethered bioresponsive polycation as a candidate for gene delivery

    Energy Technology Data Exchange (ETDEWEB)

    Zhu Ying [Second Affiliated Hospital, Medical College, Zhejiang University, Hangzhou 310009 (China); Wang Youxiang, E-mail: yx_wang@zju.edu.cn [Department of Polymer Science and Engineering, Zhejiang University, Hangzhou 310027 (China); Key Laboratory of Macromolecular Synthesis and Functionalization, Ministry of Education, Zhejiang University, Hangzhou 310027 (China); Hu Qiaoling; Shen Jiacong [Department of Polymer Science and Engineering, Zhejiang University, Hangzhou 310027 (China); Key Laboratory of Macromolecular Synthesis and Functionalization, Ministry of Education, Zhejiang University, Hangzhou 310027 (China)

    2009-04-30

    The efficient unpacking of viral protein shell gave the inspiration for the synthesized vectors. In this research, novel cholesterol tethered bioresponsive polyethylenimine (PEI) was specially designed via disulfide-containing cross-linker. The cholesterol lipid had proved to increase the permeability of gene vector through cell membrane. The acid-base titration indicated that the synthesized polycation possessed efficient proton sponge effect, which was suggested to increase endosomal release of pDNA complexes into the cytoplasm. The cholesterol tethered polycation could effectively induce DNA condensation and form spherical particles with diameter about 200 nm at N/P ratio of 10. At glutathione concentration of 3 mM, the polyplexes were unpacked due to the bioresponsive cleavage of the disulfide bonds. The in-vitro experiment indicated that the polyplexes showed efficient transfection efficiency to HEK293T cells. All the results indicated that the bioresponsive polycation could be served as an effective trigger to control the release of DNA at the intracellular environment. The novel bioresponsive polycation might have great potential in non-viral gene delivery research and application.

  2. Progranulin gene delivery reduces plaque burden and synaptic atrophy in a mouse model of Alzheimer's disease.

    Directory of Open Access Journals (Sweden)

    Jackalina M Van Kampen

    Full Text Available Progranulin (PGRN is a multifunctional protein that is widely expressed throughout the brain, where it has been shown to act as a critical regulator of CNS inflammation and also functions as an autocrine neuronal growth factor, important for long-term neuronal survival. PGRN has been shown to activate cell signaling pathways regulating excitoxicity, oxidative stress, and synaptogenesis, as well as amyloidogenesis. Together, these critical roles in the CNS suggest that PGRN has the potential to be an important therapeutic target for the treatment of various neurodegenerative disorders, particularly Alzheimer's disease (AD. AD is the leading cause of dementia and is marked by the appearance of extracellular plaques consisting of aggregates of amyloid-β (Aβ, as well as neuroinflammation, oxidative stress, neuronal loss and synaptic atrophy. The ability of PGRN to target multiple key features of AD pathophysiology suggests that enhancing its expression may benefit this disease. Here, we describe the application of PGRN gene transfer using in vivo delivery of lentiviral expression vectors in a transgenic mouse model of AD. Viral vector delivery of the PGRN gene effectively enhanced PGRN expression in the hippocampus of Tg2576 mice. This elevated PGRN expression significantly reduced amyloid plaque burden in these mice, accompanied by reductions in markers of inflammation and synaptic atrophy. The overexpression of PGRN was also found to increase activity of neprilysin, a key amyloid beta degrading enzyme. PGRN regulation of neprilysin activity could play a major role in the observed alterations in plaque burden. Thus, PGRN may be an effective therapeutic target for the treatment of AD.

  3. Poly(NIPAAm-co-AAm)-gold nanoshell composites for optically-triggered cancer therapeutic delivery

    Science.gov (United States)

    Strong, Laura Elizabeth

    Cancer is currently the second leading cause of death in the United States. Although many treatment options exist, some of the most common, including radiotherapy and chemotherapy, are restricted by dose-limiting toxicities. For example, for 2-6% of patients receiving a regimen of the chemotherapeutic doxorubicin, irreversible cardiotoxicity will occur1, causing these patients to immediately cease treatment. In addition, the largest hurdle for translating novel biological therapies such as siRNA into the clinic is lack of an efficient delivery mechanism to get the therapeutic into malignant cells2. Both of these situations would benefit from a minimally invasive controlled release system that only delivers a therapeutic to the site of malignant tissue. This thesis presents work towards the creation of such a delivery platform using two novel material components: a thermally responsive poly[N-isopropylacrylamide-co-acrylamide] (NIPAAm-co-AAm) hydrogel and gold-silica nanoshells. Thermally responsive hydrogels go through a physical property transition at their lower critical solution temperature (LCST). When transitioning from below to above the LCST, the hydrogel material expels large amounts of water and absorbed molecules. This phase change can be optically triggered by embedded gold-silica nanoshells, which rapidly transfer near-infrared (NIR) light energy into heat energy due to the surface plasmon resonance phenomena. When this material is loaded with absorbed drug molecules, drug release can be externally triggered by exposure to an NIR laser. Initial characterization of this material was done using bulk hydrogel-nanoshell composites. Poly(NIPAAm-co-AAm)-nanoshell composites were synthesized via free radical polymerization. The LCST of the poly(NIPAAm-co-AAm) hydrogels was determined to be from 39-45 °C, or slightly above physiologic temperature. The material was swollen in a drug solution of either doxorubicin (a common chemotherapeutic) or a 21bp ds

  4. Tuberculosis therapeutics: Engineering of nanomedicinal systems for local delivery of targeted drug cocktails

    Science.gov (United States)

    D'Addio, Suzanne M.

    In this thesis, a multifunctional nanocarrier drug delivery system was investigated and optimized to improve tuberculosis therapy by promoting the intracellular delivery of high payloads of antibiotics. To meet the needs of a patient population which continues to grow by close to 10 million people a year, innovative therapeutics must be formulated by robust and scalable processes. We use Flash NanoPrecipitation for the continuous precipitation of nanocarriers by block copolymer directed assembly, which enables the development of nanocarriers with tunable properties. Stable nanocarriers of Rifampicin and a hydrophobic Rifampicin prodrug have efficacy against tuberculosis in vitro that is equivalent to the soluble Rifampicin. To overcome poor in vivo efficacy of the recently discovered antitubercular drug SQ641, we co-encapsulate SQ641 and Cyclosporine A in a stable aqueous nanocarrier suspension, which enables drug administration and also enhances intracellular accumulation and antitubercular efficacy relative to SQ641 in solution. Since the mannose receptor is involved in the phagocytosis of tuberculosis bacilli, we modify the surface of nanocarriers with mannoside residues to target specific intracellular accumulation in macrophages. The surface density of mannoside terminated polyethylene glycol chains was controlled between 0 and 75% and in vitro cellular association reveals a 9% surface density is optimal for internalization mediated by the mannose receptor. We explore the preparation of large, porous aerosol carrier particles of with tunable deposition characteristics by spray freeze drying with ultrasonic atomization for direct dosing to the lungs. Nanocarriers are loaded at 3 - 50 wt% in mannitol particles with constant size, limited nanocarrier aggregation, and 63% dose delivered to the lungs, as determined by in vitro cascade impaction. There has been a lag in the development of new technologies to facilitate development and commercialization of

  5. Zein-alginate based oral drug delivery systems: Protection and release of therapeutic proteins.

    Science.gov (United States)

    Lee, Sungmun; Kim, Yeu-Chun; Park, Ji-Ho

    2016-12-30

    Reactive oxygen species (ROS) play an important role in the development of inflammatory bowel diseases. Superoxide dismutase (SOD) has a great therapeutic potential by scavenging superoxide that is one of ROS; however, in vivo application is limited especially when it is orally administered. SOD is easily degraded in vivo by the harsh conditions of gastrointestinal tract. Here, we design a zein-alginate based oral drug delivery system that protects SOD from the harsh conditions of gastrointestinal tract and releases it in the environment of the small intestine. SOD is encapsulated in zein-alginate nanoparticles (ZAN) via a phase separation method. We demonstrate that ZAN protect SOD from the harsh conditions of the stomach or small intestine condition. ZAN (200:40) at the weight ratio of 200mg zein to 40mg of alginate releases SOD in a pH dependent manner, and it releases 90.8±1.2% of encapsulated SOD at pH 7.4 in 2h, while only 11.4±0.4% of SOD was released at pH 1.3. The encapsulation efficiency of SOD in ZAN (200:40) was 62.1±2.0%. SOD in ZAN (200:40) reduced the intracellular ROS level and it saved 88.9±7.5% of Caco-2 cells from the toxic superoxide in 4 hours. Based on the results, zein-alginate based oral drug delivery systems will have numerous applications to drugs that are easily degradable in the harsh conditions of gastrointestinal tract. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. Solid Lipid Nanoparticles as Efficient Drug and Gene Delivery Systems: Recent Breakthroughs

    Directory of Open Access Journals (Sweden)

    Jafar Ezzati Nazhad Dolatabadi

    2015-06-01

    Full Text Available In recent years, nanomaterials have been widely applied as advanced drug and gene delivery nanosystems. Among them, solid lipid nanoparticles (SLNs have attracted great attention as colloidal drug delivery systems for incorporating hydrophilic or lipophilic drugs and various macromolecules as well as proteins and nucleic acids. Therefore, SLNs offer great promise for controlled and site specific drug and gene delivery. This article includes general information about SLN structures and properties, production procedures, characterization. In addition, recent progress on development of drug and gene delivery systems using SLNs was reviewed.

  7. Self-Assembling Peptide Amphiphiles for Therapeutic Delivery of Proteins, Drugs, and Stem Cells

    Science.gov (United States)

    Lee, Sungsoo Seth

    Biomaterials are used to help regenerate or replace the structure and function of damaged tissues. In order to elicit desired therapeutic responses in vivo, biomaterials are often functionalized with bioactive agents, such as growth factors, small molecule drugs, or even stem cells. Therefore, the strategies used to incorporate these bioactive agents in the microstructures and nanostructures of biomaterials can strongly influence the their therapeutic efficacy. Using self-assembling peptide amphiphiles (PAs), this work has investigated supramolecular nanostructures with improved interaction with three types of therapeutic agents: bone morphogenetic protein 2 (BMP-2) which promotes osteogenic differentiation and bone growth, anti-inflammatory drug naproxen which is used to treat osteo- and rheumatoid arthritis, and neural stem cells that could differentiate into neurons to treat neurodegenerative diseases. For BMP-2 delivery, two specific systems were investigated with affinity for BMP-2: 1) heparin-binding nanofibers that display the natural ligand of the osteogenic protein, and 2) nanofibers that display a synthetic peptide ligand discovered in our laboratory through phage display to directly bind BMP-2. Both systems promoted enhanced osteoblast differentiation of pluripotent C2C12 cells and augmented bone regeneration in two in vivo models, a rat critical-size femur defect model and spinal arthrodesis model. The thesis also describes the use of PA nanofibers to improve the delivery of the anti-inflammatory drug naproxen. To promote a controlled release, naproxen was chemically conjugated to the nanofiber surface via an ester bond that would only be cleaved by esterases, which are enzymes found naturally in the body. In the absence of esterases, the naproxen remained conjugated to the nanofibers and was non-bioactive. On the other hand, in the presence of esterases, naproxen was slowly released and inhibited cyclooxygenase-2 (COX-2) activity, an enzyme responsible

  8. Design of PEI-conjugated bio-reducible polymer for efficient gene delivery.

    Science.gov (United States)

    Nam, Joung-Pyo; Kim, Soyoung; Kim, Sung Wan

    2018-07-10

    The poly(cystaminebis(acrylamide)-diaminohexane) (poly(CBA-DAH)) was designed previously as a bio-reducible efficient gene delivery carrier. However, the high weight ratio required to form the polyplexes between poly(CBA-DAH) with pDNA is still a problem that needs to be addressed. To solve this problem and increase the transfection efficiency, poly(ethylenimine) (PEI, 1.8 kDa) was conjugated to poly(CBA-DAH) via disulfide bond. The PEI conjugated poly(CBA-DAH) (PCDP) can bind with pDNA at a very low weight ratio of 0.5 and above, like PEI 25 kDa, and form the polyplexes with nano-size (102-128 nm) and positive surface charge (27-34 mV). PCDP and PCDP polyplexes had negligible cytotoxicity and indicated similar or better cellular uptake than the comparison groups such as PEI 25 kDa and Lipofectamine® polyplexes. To confirm the transfection efficiency, the plasmid DNA (pDNA) encoded with the luciferase reporter gene (gWiz-Luc) and green fluorescent protein reporter gene (GFP) were used and treated with PCDP into the A549, Huh-7, and Mia PaCa-2 cells. PCDP/pDNA polyplexes showed highest transfection efficiency in all tested cell lines. In the luciferase assay, PCDP polyplexes showed 10.2 times higher gene transfection efficiency than Lipofectamine® polyplexes in mimic in vivo conditions (30% FBS, A549 cells). The VEGF siRNA expressing plasmid (pshVEGF), which is constructed as a therapeutic gene by our previous work, was delivered by PCDP into the cancer cells. The VEGF gene expression of PCDP/pshVEGF polyplexes was dramatically lower than control and the VEGF gene silencing efficiencies of PCDP/pshVEGF (w/w; 10/1) polyplexes were 54% (A549 cells), 77% (Huh-7 cells), and 66% (Mia PaCa-2 cells). In addition, PCDP/pshVEGF had reduced cell viability rates of about 31% (A549 cells), 39% (Huh-7 cells), and 42% (Mia PaCa-2 cells) and showed better results than all comparison groups. In the transfection efficiency and VEGF silencing assay, PCDP polyplexes showed

  9. Bioreducible poly(amido amine)s for non-viral gene delivery

    NARCIS (Netherlands)

    Lin, C.

    2008-01-01

    This thesis describes the design and development of bioreducible poly(amido amine)s as non-viral vectors for gene delivery in vitro and in vivo. The structural influences of these polymers on their physico-chemical properties and gene delivery properties, transfection capability and cytotoxicity in

  10. Assessing delivery and quantifying efficacy of small interfering ribonucleic acid therapeutics in the skin using a dual-axis confocal microscope

    Science.gov (United States)

    Ra, Hyejun; Gonzalez-Gonzalez, Emilio; Smith, Bryan R.; Gambhir, Sanjiv S.; Kino, Gordon S.; Solgaard, Olav; Kaspar, Roger L.; Contag, Christopher H.

    2010-05-01

    Transgenic reporter mice and advances in imaging instrumentation are enabling real-time visualization of cellular mechanisms in living subjects and accelerating the development of novel therapies. Innovative confocal microscope designs are improving their utility for microscopic imaging of fluorescent reporters in living animals. We develop dual-axis confocal (DAC) microscopes for such in vivo studies and create mouse models where fluorescent proteins are expressed in the skin for the purpose of advancing skin therapeutics and transdermal delivery tools. Three-dimensional image volumes, through the different skin compartments of the epidermis and dermis, can be acquired in several seconds with the DAC microscope in living mice, and are comparable to histologic analyses of reporter protein expression patterns in skin sections. Intravital imaging with the DAC microscope further enables visualization of green fluorescent protein (GFP) reporter gene expression in the skin over time, and quantification of transdermal delivery of small interfering RNA (siRNA) and therapeutic efficacy. Visualization of transdermal delivery of nucleic acids will play an important role in the development of innovative strategies for treating skin pathologies.

  11. Immuno-Oncology-The Translational Runway for Gene Therapy: Gene Therapeutics to Address Multiple Immune Targets.

    Science.gov (United States)

    Weß, Ludger; Schnieders, Frank

    2017-12-01

    Cancer therapy is once again experiencing a paradigm shift. This shift is based on extensive clinical experience demonstrating that cancer cannot be successfully fought by addressing only single targets or pathways. Even the combination of several neo-antigens in cancer vaccines is not sufficient for successful, lasting tumor eradication. The focus has therefore shifted to the immune system's role in cancer and the striking abilities of cancer cells to manipulate and/or deactivate the immune system. Researchers and pharma companies have started to target the processes and cells known to support immune surveillance and the elimination of tumor cells. Immune processes, however, require novel concepts beyond the traditional "single-target-single drug" paradigm and need parallel targeting of diverse cells and mechanisms. This review gives a perspective on the role of gene therapy technologies in the evolving immuno-oncology space and identifies gene therapy as a major driver in the development and regulation of effective cancer immunotherapy. Present challenges and breakthroughs ranging from chimeric antigen receptor T-cell therapy, gene-modified oncolytic viruses, combination cancer vaccines, to RNA therapeutics are spotlighted. Gene therapy is recognized as the most prominent technology enabling effective immuno-oncology strategies.

  12. Gene delivery systems by the combination of lipid bubbles and ultrasound.

    Science.gov (United States)

    Negishi, Yoichi; Endo-Takahashi, Yoko; Maruyama, Kazuo

    2016-11-28

    Gene therapy is promising for the treatment of many diseases including cancers and genetic diseases. From the viewpoint of safety, ultrasound (US)-mediated gene delivery with nano/ microbubbles was recently developed as a novel non-viral vector system. US-mediated gene delivery using nano/microbubbles are able to produce transient changes in the permeability of the cell membrane after US-induced cavitation while reducing cellular damage and enables the tissue-specific or the site-specific intracellular delivery of gene both in vitro and in vivo. We have recently developed novel lipid nanobubbles (Lipid Bubbles). These nanobubbles can also be used to enhance the efficacy of the US-mediated genes (plasmid DNA, siRNA, and miRNA etc.) delivery. In this review, we describe US-mediated delivery systems combined with nano/microbubbles and discuss their feasibility as non-viral vector systems.

  13. Three-layered polyplex as a microRNA targeted delivery system for breast cancer gene therapy

    Science.gov (United States)

    Li, Yan; Dai, Yu; Zhang, Xiaojin; Chen, Jihua

    2017-07-01

    MicroRNAs (miRNAs), small non-coding RNAs, play an important role in modulating cell proliferation, migration, and differentiation. Since miRNAs can regulate multiple cancer-related genes simultaneously, regulating miRNAs could target a set of related oncogenic genes or pathways. Owing to their reduced immune response and low toxicity, miRNAs with small size and low molecular weight have become increasingly promising therapeutic drugs in cancer therapy. However, one of the major challenges of miRNAs-based cancer therapy is to achieve specific, effective, and safe delivery of therapeutic miRNAs into cancer cells. Here we provide a strategy using three-layered polyplex with folic acid as a targeting group to systemically deliver miR-210 into breast cancer cells, which results in breast cancer growth being inhibited.

  14. PLGA Nanoparticles for Ultrasound-Mediated Gene Delivery to Solid Tumors

    Directory of Open Access Journals (Sweden)

    Marxa Figueiredo

    2012-01-01

    Full Text Available This paper focuses on novel approaches in the field of nanotechnology-based carriers utilizing ultrasound stimuli as a means to spatially target gene delivery in vivo, using nanoparticles made with either poly(lactic-co-glycolic acid (PLGA or other polymers. We specifically discuss the potential for gene delivery by particles that are echogenic (amenable to destruction by ultrasound composed either of polymers (PLGA, polystyrene or other contrast agent materials (Optison, SonoVue microbubbles. The use of ultrasound is an efficient tool to further enhance gene delivery by PLGA or other echogenic particles in vivo. Echogenic PLGA nanoparticles are an attractive strategy for ultrasound-mediated gene delivery since this polymer is currently approved by the US Food and Drug Administration for drug delivery and diagnostics in cancer, cardiovascular disease, and also other applications such as vaccines and tissue engineering. This paper will review recent successes and the potential of applying PLGA nanoparticles for gene delivery, which include (a echogenic PLGA used with ultrasound to enhance local gene delivery in tumors or muscle and (b PLGA nanoparticles currently under development, which could benefit in the future from ultrasound-enhanced tumor targeted gene delivery.

  15. Dual-therapeutic reporter genes fusion for enhanced cancer gene therapy and imaging.

    Science.gov (United States)

    Sekar, T V; Foygel, K; Willmann, J K; Paulmurugan, R

    2013-05-01

    Two of the successful gene-directed enzyme prodrug therapies include herpes simplex virus-thymidine kinase (HSV1-TK) enzyme-ganciclovir prodrug and the Escherichia coli nitroreductase (NTR) enzyme-CB1954 prodrug strategies; these enzyme-prodrug combinations produce activated cytotoxic metabolites of the prodrugs capable of tumor cell death by inhibiting DNA synthesis and killing quiescent cells, respectively. Both these strategies also affect significant bystander cell killing of neighboring tumor cells that do not express these enzymes. We have developed a dual-combination gene strategy, where we identified HSV1-TK and NTR fused in a particular orientation can effectively kill tumor cells when the tumor cells are treated with a fusion HSV1-TK-NTR gene- along with a prodrug combination of GCV and CB1954. In order to determine whether the dual-system demonstrate superior therapeutic efficacy than either HSV1-TK or NTR systems alone, we conducted both in vitro and in vivo tumor xenograft studies using triple negative SUM159 breast cancer cells, by evaluating the efficacy of cell death by apoptosis and necrosis upon treatment with the dual HSV1-TK genes-GCV-CB1954 prodrugs system, and compared the efficiency to HSV1-TK-GCV and NTR-CB1954. Our cell-based studies, tumor regression studies in xenograft mice, histological analyses of treated tumors and bystander studies indicate that the dual HSV1-TK-NTR-prodrug system is two times more efficient even with half the doses of both prodrugs than the respective single gene-prodrug system, as evidenced by enhanced apoptosis and necrosis of tumor cells in vitro in culture and xenograft of tumor tissues in animals.

  16. Polyamidoamine-Decorated Nanodiamonds as a Hybrid Gene Delivery Vector and siRNA Structural Characterization at the Charged Interfaces.

    Science.gov (United States)

    Lim, Dae Gon; Rajasekaran, Nirmal; Lee, Dukhee; Kim, Nam Ah; Jung, Hun Soon; Hong, Sungyoul; Shin, Young Kee; Kang, Eunah; Jeong, Seong Hoon

    2017-09-20

    Nanodiamonds have been discovered as a new exogenous material source in biomedical applications. As a new potent form of nanodiamond (ND), polyamidoamine-decorated nanodiamonds (PAMAM-NDs) were prepared for E7 or E6 oncoprotein-suppressing siRNA gene delivery for high risk human papillomavirus-induced cervical cancer, such as types 16 and 18. It is critical to understand the physicochemical properties of siRNA complexes immobilized on cationic solid ND surfaces in the aspect of biomolecular structural and conformational changes, as the new inert carbon material can be extended into the application of a gene delivery vector. A spectral study of siRNA/PAMAM-ND complexes using differential scanning calorimetry and circular dichroism spectroscopy proved that the hydrogen bonding and electrostatic interactions between siRNA and PAMAM-NDs decreased endothermic heat capacity. Moreover, siRNA/PAMAM-ND complexes showed low cell cytotoxicity and significant suppressing effects for forward target E6 and E7 oncogenic genes, proving functional and therapeutic efficacy. The cellular uptake of siRNA/PAMAM-ND complexes at 8 h was visualized by macropinocytes and direct endosomal escape of the siRNA/PAMAM-ND complexes. It is presumed that PAMAM-NDs provided a buffering cushion to adjust the pH and hard mechanical stress to escape endosomes. siRNA/PAMAM-ND complexes provide a potential organic/inorganic hybrid material source for gene delivery carriers.

  17. Application of pulsed-magnetic field enhances non-viral gene delivery in primary cells from different origins

    Energy Technology Data Exchange (ETDEWEB)

    Kamau Chapman, Sarah W. [Institute of Veterinary Biochemistry and Molecular Biology, University of Zurich, Winterthurerstr. 190, 8057 Zurich (Switzerland); Hassa, Paul O. [Institute of Veterinary Biochemistry and Molecular Biology, University of Zurich, Winterthurerstr. 190, 8057 Zurich (Switzerland); European Molecular Biology Laboratory (EMBL) Heidelberg, Meyerhofstrasse 1, 69117 Heidelberg (Germany); Koch-Schneidemann, Sabine; Rechenberg, Brigitte von [Musculoskeletal Research Unit, Equine Hospital, Vetsuisse Faculty Zurich, University of Zurich, Winterthurerstr. 260, 8057 Zurich (Switzerland); Hofmann-Amtenbrink, Margarethe [MatSearch, Chemin Jean Pavillard 14, 1009 Pully (Switzerland); Steitz, Benedikt; Petri-Fink, Alke; Hofmann, Heinrich [Laboratory of Powder Technology, Ecole Polytechnique Federale de Lausanne (EPFL), Lausanne (Switzerland); Hottiger, Michael O. [Institute of Veterinary Biochemistry and Molecular Biology, University of Zurich, Winterthurerstr. 190, 8057 Zurich (Switzerland)], E-mail: hottiger@vetbio.uzh.ch

    2008-04-15

    Primary cell lines are more difficult to transfect when compared to immortalized/transformed cell lines, and hence new techniques are required to enhance the transfection efficiency in these cells. We isolated and established primary cultures of synoviocytes, chondrocytes, osteoblasts, melanocytes, macrophages, lung fibroblasts, and embryonic fibroblasts. These cells differed in several properties, and hence were a good representative sample of cells that would be targeted for expression and delivery of therapeutic genes in vivo. The efficiency of gene delivery in all these cells was enhanced using polyethylenimine-coated polyMAG magnetic nanoparticles, and the rates (17-84.2%) surpassed those previously achieved using other methods, especially in cells that are difficult to transfect. The application of permanent and pulsating magnetic fields significantly enhanced the transfection efficiencies in synoviocytes, chondrocytes, osteoblasts, melanocytes and lung fibroblasts, within 5 min of exposure to these magnetic fields. This is an added advantage for future in vivo applications, where rapid gene delivery is required before systemic clearance or filtration of the gene vectors occurs.

  18. The sigma-2 receptor as a therapeutic target for drug delivery in triple negative breast cancer

    International Nuclear Information System (INIS)

    Makvandi, Mehran; Tilahun, Estifanos D.; Lieberman, Brian P.; Anderson, Redmond-Craig; Zeng, Chenbo; Xu, Kuiying; Hou, Catherine; McDonald, Elizabeth S.; Pryma, Daniel A.; Mach, Robert H.

    2015-01-01

    Background: Triple-negative breast cancer (TNBC) is associated with high relapse rates and increased mortality when compared with other breast cancer subtypes. In contrast to receptor positive breast cancers, there are no approved targeted therapies for TNBC. Identifying biomarkers for TNBC is of high importance for the advancement of patient care. The sigma-2 receptor has been shown to be overexpressed in triple negative breast cancer in vivo and has been characterized as a marker of proliferation. The aim of the present study was to define the sigma-2 receptor as a target for therapeutic drug delivery and biomarker in TNBC. Methods: Three TNBC cell lines were evaluated: MDA-MB-231, HCC1937 and HCC1806. Sigma-2 compounds were tested for pharmacological properties specific to the sigma-2 receptor through competitive inhibition assays. Sigma-2 receptor expression was measured through radioligand receptor saturation studies. Drug sensitivity for taxol was compared to a sigma-2 targeting compound conjugated to a cytotoxic payload, SW IV-134. Cell viability was assessed after treatments for 2 or 48 h. Sigma-2 blockade was assessed to define sigma-2 mediated cytotoxicity of SW IV-134. Caspase 3/7 activation induced by SW IV-134 was measured at corresponding treatment time points. Results: SW IV-134 was the most potent compound tested in two of the three cell lines and was similarly effective in all three. MDA-MB-231 displayed a statistically significant higher sigma-2 receptor expression and also was the most sensitive cell line evaluated to SW IV-134. Conclusion: Targeting the sigma-2 receptor with a cytotoxic payload was effective in all the three cell lines evaluated and provides the proof of concept for future development of a therapeutic platform for the treatment of TNBC. - Highlights: • TNBC cells are sensitive to sigma-2 receptor targeted drug conjugate SW IV-134. • MDA-MB-231 displayed the highest amount of sigma-2 receptors and corresponded well with

  19. The sigma-2 receptor as a therapeutic target for drug delivery in triple negative breast cancer

    Energy Technology Data Exchange (ETDEWEB)

    Makvandi, Mehran; Tilahun, Estifanos D.; Lieberman, Brian P.; Anderson, Redmond-Craig; Zeng, Chenbo; Xu, Kuiying; Hou, Catherine; McDonald, Elizabeth S.; Pryma, Daniel A.; Mach, Robert H., E-mail: rmach@mail.med.upenn.edu

    2015-11-27

    Background: Triple-negative breast cancer (TNBC) is associated with high relapse rates and increased mortality when compared with other breast cancer subtypes. In contrast to receptor positive breast cancers, there are no approved targeted therapies for TNBC. Identifying biomarkers for TNBC is of high importance for the advancement of patient care. The sigma-2 receptor has been shown to be overexpressed in triple negative breast cancer in vivo and has been characterized as a marker of proliferation. The aim of the present study was to define the sigma-2 receptor as a target for therapeutic drug delivery and biomarker in TNBC. Methods: Three TNBC cell lines were evaluated: MDA-MB-231, HCC1937 and HCC1806. Sigma-2 compounds were tested for pharmacological properties specific to the sigma-2 receptor through competitive inhibition assays. Sigma-2 receptor expression was measured through radioligand receptor saturation studies. Drug sensitivity for taxol was compared to a sigma-2 targeting compound conjugated to a cytotoxic payload, SW IV-134. Cell viability was assessed after treatments for 2 or 48 h. Sigma-2 blockade was assessed to define sigma-2 mediated cytotoxicity of SW IV-134. Caspase 3/7 activation induced by SW IV-134 was measured at corresponding treatment time points. Results: SW IV-134 was the most potent compound tested in two of the three cell lines and was similarly effective in all three. MDA-MB-231 displayed a statistically significant higher sigma-2 receptor expression and also was the most sensitive cell line evaluated to SW IV-134. Conclusion: Targeting the sigma-2 receptor with a cytotoxic payload was effective in all the three cell lines evaluated and provides the proof of concept for future development of a therapeutic platform for the treatment of TNBC. - Highlights: • TNBC cells are sensitive to sigma-2 receptor targeted drug conjugate SW IV-134. • MDA-MB-231 displayed the highest amount of sigma-2 receptors and corresponded well with

  20. Human Growth Hormone Delivery with a Microneedle Transdermal System: Preclinical Formulation, Stability, Delivery and PK of Therapeutically Relevant Doses

    Directory of Open Access Journals (Sweden)

    Mahmoud Ameri

    2014-05-01

    Full Text Available This study evaluated the feasibility of coating formulated recombinant human growth hormone (rhGH on a titanium microneedle transdermal delivery system, Zosano Pharma (ZP-hGH, and assessed preclinical patch delivery performance. Formulation rheology and surface activity were assessed by viscometry and contact angle measurement. rhGH liquid formulation was coated onto titanium microneedles by dip-coating and drying. The stability of coated rhGH was determined by size exclusion chromatography-high performance liquid chromatography (SEC-HPLC. Preclinical delivery and pharmacokinetic studies were conducted in female hairless guinea pigs (HGP using rhGH coated microneedle patches at 0.5 and 1 mg doses and compared to Norditropin® a commercially approved rhGH subcutaneous injection. Studies demonstrated successful rhGH formulation development and coating on microneedle arrays. The ZP-hGH patches remained stable at 40 °C for six months with no significant change in % aggregates. Pharmacokinetic studies showed that the rhGH-coated microneedle patches, delivered with high efficiency and the doses delivered indicated linearity with average Tmax of 30 min. The absolute bioavailability of the microneedle rhGH patches was similar to subcutaneous Norditropin® injections. These results suggest that ZP-transdermal microneedle patch delivery of rhGH is feasible and may offer an effective and patient-friendly alternative to currently marketed rhGH injectables.

  1. Human Growth Hormone Delivery with a Microneedle Transdermal System: Preclinical Formulation, Stability, Delivery and PK of Therapeutically Relevant Doses.

    Science.gov (United States)

    Ameri, Mahmoud; Kadkhodayan, Miryam; Nguyen, Joe; Bravo, Joseph A; Su, Rebeca; Chan, Kenneth; Samiee, Ahmad; Daddona, Peter E

    2014-05-15

    This study evaluated the feasibility of coating formulated recombinant human growth hormone (rhGH) on a titanium microneedle transdermal delivery system, Zosano Pharma (ZP)-hGH, and assessed preclinical patch delivery performance. Formulation rheology and surface activity were assessed by viscometry and contact angle measurement. rhGH liquid formulation was coated onto titanium microneedles by dip-coating and drying. The stability of coated rhGH was determined by size exclusion chromatography-high performance liquid chromatography (SEC-HPLC). Preclinical delivery and pharmacokinetic studies were conducted in female hairless guinea pigs (HGP) using rhGH coated microneedle patches at 0.5 and 1 mg doses and compared to Norditropin® a commercially approved rhGH subcutaneous injection. Studies demonstrated successful rhGH formulation development and coating on microneedle arrays. The ZP-hGH patches remained stable at 40 °C for six months with no significant change in % aggregates. Pharmacokinetic studies showed that the rhGH-coated microneedle patches, delivered with high efficiency and the doses delivered indicated linearity with average Tmax of 30 min. The absolute bioavailability of the microneedle rhGH patches was similar to subcutaneous Norditropin® injections. These results suggest that ZP-transdermal microneedle patch delivery of rhGH is feasible and may offer an effective and patient-friendly alternative to currently marketed rhGH injectables.

  2. Production of bioinspired and rationally designed polymer hydrogels for controlled delivery of therapeutic proteins

    Science.gov (United States)

    Kim, Sung Hye

    Hydrogel systems for controlled delivery therapeutic growth factors have been developed in a wide spectrum of strategies: these systems aim for the release of growth factors via a passive diffusion, electrostatic interaction, degradation of hydrogels, and responsiveness to external stimuli. Heparin, a highly sulfated glycosaminoglycan (GAG), was employed for a targeted delivery system of vascular endothelial growth factor (VEGF) to endothelial cells overexpressing a relevant receptor VEGFR-2. Addition of dimeric VEGF to 4-arm star-shaped poly(ethylene glycol) (PEG) immobilized with low-molecular weight heparin (LMWH) afforded a non-covalently assembled hydrogel via interaction between heparin and VEGF, with storage modulus 10 Pa. The release of VEGF and hydrogel erosion reached maximum 100 % at day 4 in the presence of VEGFR-2 overexpressing pocine aortic endothelial cell (PAE/KDR), while those of 80% were achieved via passive release at day 5 in the presence of PAE cell lacking VEGFR-2 or in the absence of cell, indicating that the release of VEGF was in targeted manner toward cell receptor. The proliferation of PAE/KDR in the presence of [PEG-LMWH/VEGF] hydrogel was greater by ca. 30% at day 4 compared to that of PAE, confirming that the release of VEGF was in response to the cellular demand. The phosphorylation fraction of VEGFR-2 on PAE/KDR was greater in the presence of [PEG-LMWH/VEGF] hydrogel, increasing from 0.568 at day 1 to 0.790 at day 4, whereas it was maintained at 0.230 at day 4 in the presence of [PEG-LMWH] hydrogel. This study has proven that this hydrogel, assembled via bio-inspired non-covalent interaction, liberating VEGFon celluar demand to target cell, eroding upon VEGF release, and triggering endothelial cell proliferation, could be used in multiple applications including targeted delivery and angiogenesis. Heparin has been widely exploited in growth factor delivery systems owing to its ability to bind many growth factors through the flexible

  3. Discovery of Cationic Polymers for Non-viral Gene Delivery using Combinatorial Approaches

    Science.gov (United States)

    Barua, Sutapa; Ramos, James; Potta, Thrimoorthy; Taylor, David; Huang, Huang-Chiao; Montanez, Gabriela; Rege, Kaushal

    2015-01-01

    Gene therapy is an attractive treatment option for diseases of genetic origin, including several cancers and cardiovascular diseases. While viruses are effective vectors for delivering exogenous genes to cells, concerns related to insertional mutagenesis, immunogenicity, lack of tropism, decay and high production costs necessitate the discovery of non-viral methods. Significant efforts have been focused on cationic polymers as non-viral alternatives for gene delivery. Recent studies have employed combinatorial syntheses and parallel screening methods for enhancing the efficacy of gene delivery, biocompatibility of the delivery vehicle, and overcoming cellular level barriers as they relate to polymer-mediated transgene uptake, transport, transcription, and expression. This review summarizes and discusses recent advances in combinatorial syntheses and parallel screening of cationic polymer libraries for the discovery of efficient and safe gene delivery systems. PMID:21843141

  4. Effect of a Novel Nonviral Gene Delivery of BMP-2 on Bone Healing

    Directory of Open Access Journals (Sweden)

    P. Schwabe

    2012-01-01

    Full Text Available Background. Gene therapeutic drug delivery approaches have been introduced to improve the efficiency of growth factors at the site of interest. This study investigated the efficacy and safety of a new nonviral copolymer-protected gene vector (COPROG for the stimulation of bone healing. Methods. In vitro, rat osteoblasts were transfected with COPROG + luciferase plasmid or COPROG + hBMP-2 plasmid. In vivo, rat tibial fractures were intramedullary stabilized with uncoated versus COPROG+hBMP-2-plasmid-coated titanium K-wires. The tibiae were prepared for biomechanical and histological analyses at days 28 and 42 and for transfection/safety study at days 2, 4, 7, 28, and 42. Results. In vitro results showed luciferase expression until day 21, and hBMP-2-protein was measured from day 2 – day 10. In vivo, the local application of hBMP-2-plasmid showed a significantly higher maximum load after 42 days compared to that in the control. The histomorphometric analysis revealed a significantly less mineralized periosteal callus area in the BMP-2 group compared to the control at day 28. The rt-PCR showed no systemic biodistribution of luciferase RNA. Conclusion. A positive effect on fracture healing by nonviral BMP-2 plasmid application from COPROG-coated implants could be shown in this study; however, the effect of the vector may be improved with higher plasmid concentrations. Transfection showed no biodistribution to distant organs and was considered to be safe.

  5. Direct visualization of electroporation-assisted in vivo gene delivery to tumors using intravital microscopy – spatial and time dependent distribution

    Directory of Open Access Journals (Sweden)

    Dachs Gabi U

    2004-11-01

    Full Text Available Abstract Background Electroporation is currently receiving much attention as a way to increase drug and DNA delivery. Recent studies demonstrated the feasibility of electrogene therapy using a range of therapeutic genes for the treatment of experimental tumors. However, the transfection efficiency of electroporation-assisted DNA delivery is still low compared to viral methods and there is a clear need to optimize this approach. In order to optimize treatment, knowledge about spatial and time dependency of gene expression following delivery is of utmost importance in order to improve gene delivery. Intravital microscopy of tumors growing in dorsal skin fold window chambers is a useful method for monitoring gene transfection, since it allows non-invasive dynamic monitoring of gene expression in tumors in a live animal. Methods Intravital microscopy was used to monitor real time spatial distribution of the green fluorescent protein (GFP and time dependence of transfection efficiency in syngeneic P22 rat tumor model. DNA alone, liposome-DNA complexes and electroporation-assisted DNA delivery using two different sets of electric pulse parameters were compared. Results Electroporation-assisted DNA delivery using 8 pulses, 600 V/cm, 5 ms, 1 Hz was superior to other methods and resulted in 22% increase in fluorescence intensity in the tumors up to 6 days post-transfection, compared to the non-transfected area in granulation tissue. Functional GFP was detected within 5 h after transfection. Cells expressing GFP were detected throughout the tumor, but not in the surrounding tissue that was not exposed to electric pulses. Conclusions Intravital microscopy was demonstrated to be a suitable method for monitoring time and spatial distribution of gene expression in experimental tumors and provided evidence that electroporation-assisted gene delivery using 8 pulses, 600 V/cm, 5 ms, 1 Hz is an effective method, resulting in early onset and homogenous

  6. Direct visualization of electroporation-assisted in vivo gene delivery to tumors using intravital microscopy – spatial and time dependent distribution

    International Nuclear Information System (INIS)

    Cemazar, Maja; Wilson, Ian; Dachs, Gabi U; Tozer, Gillian M; Sersa, Gregor

    2004-01-01

    Electroporation is currently receiving much attention as a way to increase drug and DNA delivery. Recent studies demonstrated the feasibility of electrogene therapy using a range of therapeutic genes for the treatment of experimental tumors. However, the transfection efficiency of electroporation-assisted DNA delivery is still low compared to viral methods and there is a clear need to optimize this approach. In order to optimize treatment, knowledge about spatial and time dependency of gene expression following delivery is of utmost importance in order to improve gene delivery. Intravital microscopy of tumors growing in dorsal skin fold window chambers is a useful method for monitoring gene transfection, since it allows non-invasive dynamic monitoring of gene expression in tumors in a live animal. Intravital microscopy was used to monitor real time spatial distribution of the green fluorescent protein (GFP) and time dependence of transfection efficiency in syngeneic P22 rat tumor model. DNA alone, liposome-DNA complexes and electroporation-assisted DNA delivery using two different sets of electric pulse parameters were compared. Electroporation-assisted DNA delivery using 8 pulses, 600 V/cm, 5 ms, 1 Hz was superior to other methods and resulted in 22% increase in fluorescence intensity in the tumors up to 6 days post-transfection, compared to the non-transfected area in granulation tissue. Functional GFP was detected within 5 h after transfection. Cells expressing GFP were detected throughout the tumor, but not in the surrounding tissue that was not exposed to electric pulses. Intravital microscopy was demonstrated to be a suitable method for monitoring time and spatial distribution of gene expression in experimental tumors and provided evidence that electroporation-assisted gene delivery using 8 pulses, 600 V/cm, 5 ms, 1 Hz is an effective method, resulting in early onset and homogenous distribution of gene expression in the syngeneic P22 rat tumor model

  7. Amino acid-substituted gemini surfactant-based nanoparticles as safe and versatile gene delivery agents.

    Science.gov (United States)

    Singh, Jagbir; Yang, Peng; Michel, Deborah; Verrall, Ronald E; Foldvari, Marianna; Badea, Ildiko

    2011-05-01

    Gene based therapy represents an important advance in the treatment of diseases that heretofore have had either no treatment or cure. To capitalize on the true potential of gene therapy, there is a need to develop better delivery systems that can protect these therapeutic biomolecules and deliver them safely to the target sites. Recently, we have designed and developed a series of novel amino acid-substituted gemini surfactants with the general chemical formula C(12)H(25) (CH(3))(2)N(+)-(CH(2))(3)-N(AA)-(CH(2))(3)-N(+) (CH(3))(2)-C(12)H(25) (AA= glycine, lysine, glycyl-lysine and, lysyl-lysine). These compounds were synthesized and tested in rabbit epithelial cells using a model plasmid and a helper lipid. Plasmid/gemini/lipid (P/G/L) nanoparticles formulated using these novel compounds achieved higher gene expression than the nanoparticles containing the parent unsubstituted compound. In this study, we evaluated the cytotoxicity of P/G/L nanoparticles and explored the relationship between transfection efficiency/toxicity and their physicochemical characteristics (such as size, binding properties, etc.). An overall low toxicity is observed for all complexes with no significant difference among substituted and unsubstituted compounds. An interesting result revealed by the dye exclusion assay suggests a more balanced protection of the DNA by the glycine and glycyl-lysine substituted compounds. Thus, the higher transfection efficiency is attributed to the greater biocompatibility and flexibility of the amino acid/peptide-substituted gemini surfactants and demonstrates the feasibility of using amino acid-substituted gemini surfactants as gene carriers for the treatment of diseases affecting epithelial tissue.

  8. Transdermal and intradermal delivery of therapeutic agents: application of physical technologies

    National Research Council Canada - National Science Library

    Banga, Ajay K

    2011-01-01

    .... Commercialization of transdermal drug delivery requires technology from many disciplines beyond pharmaceutical sciences, such as polymer chemistry, adhesion sciences, mass transport, web film coating...

  9. Advanced drug and gene delivery systems based on functional biodegradable polycarbonates and copolymers

    NARCIS (Netherlands)

    Chen, Wei; Meng, Fenghua; Cheng, R.; Deng, C.; Feijen, Jan; Zhong, Zhiyuan

    2014-01-01

    Biodegradable polymeric nanocarriers are one of the most promising systems for targeted and controlled drug and gene delivery. They have shown several unique advantages such as excellent biocompatibility, prolonged circulation time, passive tumor targeting via the enhanced permeability and retention

  10. Non-electrostatic complexes with DNA: towards novel synthetic gene delivery systems.

    Science.gov (United States)

    Soto, J; Bessodes, M; Pitard, B; Mailhe, P; Scherman, D; Byk, G

    2000-05-01

    We have developed new DNA complexing amphiphile based on Hoechst 33258 interaction with DNA grooves. The synthesis and physicochemical characterisation of lipid/DNA complexes, as compared to that of classical lipopolyamine for gene delivery, are described and discussed.

  11. Reducible poly(amido ethylenimine)s designed for triggered intracellular gene delivery

    NARCIS (Netherlands)

    Christensen, Lane V.; Christensen, L.; Chang, Chien-Wen; Kim, Won Jong; Kim, Sung Wan; Zhong, Zhiyuan; Lin, C.; Engbersen, Johannes F.J.; Feijen, Jan

    2006-01-01

    Poly(amido ethylenimine) polymers, a new type of peptidomimetic polymer, containing multiple disulfide bonds (SS-PAEIs) designed to degrade after delivery of plasmid DNA (pDNA) into the cell were synthesized and investigated as new carriers for triggered intracellular gene delivery. More

  12. Combining bio-electrospraying with gene therapy: a novel biotechnique for the delivery of genetic material via living cells.

    Science.gov (United States)

    Ward, Eliot; Chan, Emma; Gustafsson, Kenth; Jayasinghe, Suwan N

    2010-05-01

    The investigations reported in this article demonstrate the ability of bio-electrosprays and cell electrospinning to deliver a genetic construct in association with living cells. Previous studies on both bio-electrosprays and cell electrospinning demonstrated great promise for tissue engineering and regenerative biology/medicine. The investigations described herein widen the applicability of these biotechniques by combining gene therapy protocols, resulting in a novel drug delivery methodology previously unexplored. In these studies a human cell line was transduced with recombinant self-inactivating lentiviral particles. These particles incorporated a green fluorescent protein fused to an endosomal targeting construct. This construct encodes a peptide, which can subsequently be detected on the surface of cells by specific T-cells. The transduced cell line was subsequently manipulated in association with either bio-electrospraying or cell electrospinning. Hence this demonstrates (i) the ability to safely handle genetically modified living cells and (ii) the ability to directly form pre-determined architectures bearing living therapeutic cells. This merged technology demonstrates a unique approach for directly forming living therapeutic architectures for controlled and targeted release of experimental cells/genes, as well as medical cell/gene therapeutics for a plethora of biological and medical applications. Hence, such developments could be applied to personalised medicine.

  13. Layered nanoemulsions as mucoadhesive buccal systems for controlled delivery of oral cancer therapeutics

    Directory of Open Access Journals (Sweden)

    Gavin A

    2015-02-01

    Full Text Available Amy Gavin,1 Jimmy TH Pham,2 Dawei Wang,2 Bill Brownlow,3 Tamer A Elbayoumi3 1College of Dental Medicine, 2Arizona College of Osteopathic Medicine, 3Department of Pharmaceutical Sciences, College of Pharmacy-Glendale, Midwestern University, Glendale, AZ, USA Abstract: Oral cavity and oropharyngeal cancers are considered the eighth most common cancer worldwide, with relatively poor prognosis (62% of patients surviving 5 years, after diagnosis. The aim of this study was to develop a proof-of-concept mucoadhesive lozenge/buccal tablet, as a potential platform for direct sustained delivery of therapeutic antimitotic nanomedicines. Our system would serve as an adjuvant therapy for oral cancer patients undergoing full-scale diagnostic and operative treatment plans. We utilized lipid-based nanocarriers, namely nanoemulsions (NEs, containing mixed-polyethoxylated emulsifiers and a tocopheryl moiety–enriched oil phase. Prototype NEs, loaded with the proapoptotic lipophilic drug genistein (Gen, were further processed into buccal tablet formulations. The chitosan polyelectrolyte solution overcoat rendered NE droplets cationic, by acting as a mucoadhesive interfacial NE layer. With approximate size of 110 nm, the positively charged chitosan-layered NE (+25 mV vs negatively charged chitosan-free/primary aqueous NE (-28 mV exhibited a controlled-release profile and effective mucoadhesion for liquid oral spray prototypes. When punch-pressed, porous NE-based buccal tablets were physically evaluated for hardness, friability, and swelling in addition to ex vivo tissue mucoadhesion force and retention time measurements. Chitosan-containing NE tablets were found equivalent to primary NE and placebo tablets in compression tests, yet significantly superior in all ex vivo adhesion and in vitro release assays (P≤0.05. Following biocompatibility screening of prototype chitosan-layered NEs, substantial anticancer activity of selected cationic Gen-loaded NE

  14. BDNF gene delivery mediated by neuron-targeted nanoparticles is neuroprotective in peripheral nerve injury

    OpenAIRE

    Lopes, CDF; Gonçalves, NP; Gomes, CP; Saraiva, MJ; Pêgo, AP

    2017-01-01

    Neuron-targeted gene delivery is a promising strategy to treat peripheral neuropathies. Here we propose the use of polymeric nanoparticles based on thiolated trimethyl chitosan (TMCSH) to mediate targeted gene delivery to peripheral neurons upon a peripheral and minimally invasive intramuscular administration. Nanoparticles were grafted with the non-toxic carboxylic fragment of the tetanus neurotoxin (HC) to allow neuron targeting and were explored to deliver a plasmid DNA encoding for the br...

  15. Membrane-Mimic Nanoparticles for Drug and Gene Delivery

    KAUST Repository

    Alamoudi, Kholod

    2017-01-01

    -mimic nanoparticles are considered highly attractive materials for in vivo and in vitro applications. Synthetic membrane vesicles (liposomes) and nanoconstructs built with native cancer cellular membrane are excellent scaffolds to improve cellular delivery. Liposomes

  16. Gene therapy strategy for long-term myocardial protection using adeno-associated virus-mediated delivery of heme oxygenase gene.

    Science.gov (United States)

    Melo, Luis G; Agrawal, Reitu; Zhang, Lunan; Rezvani, Mojgan; Mangi, Abeel A; Ehsan, Afshin; Griese, Daniel P; Dell'Acqua, Giorgio; Mann, Michael J; Oyama, Junichi; Yet, Shaw-Fang; Layne, Matthew D; Perrella, Mark A; Dzau, Victor J

    2002-02-05

    Ischemia and oxidative stress are the leading mechanisms for tissue injury. An ideal strategy for preventive/protective therapy would be to develop an approach that could confer long-term transgene expression and, consequently, tissue protection from repeated ischemia/reperfusion injury with a single administration of a therapeutic gene. In the present study, we used recombinant adeno-associated virus (rAAV) as a vector for direct delivery of the cytoprotective gene heme oxygenase-1 (HO-1) into the rat myocardium, with the purpose of evaluating this strategy as a therapeutic approach for long-term protection from ischemia-induced myocardial injury. Human HO-1 gene (hHO-1) was delivered to normal rat hearts by intramyocardial injection. AAV-mediated transfer of the hHO-1 gene 8 weeks before acute coronary artery ligation and release led to a dramatic reduction (>75%) in left ventricular myocardial infarction. The reduction in infarct size was accompanied by decreases in myocardial lipid peroxidation and in proapoptotic Bax and proinflammatory interleukin-1beta protein abundance, concomitant with an increase in antiapoptotic Bcl-2 protein level. This suggested that the transgene exerts its cardioprotective effects in part by reducing oxidative stress and associated inflammation and apoptotic cell death. This study documents the beneficial therapeutic effect of rAAV-mediated transfer, before myocardial injury, of a cytoprotective gene that confers long-term myocardial protection from ischemia/reperfusion injury. Our data suggest that this novel "pre-event" gene transfer approach may provide sustained tissue protection from future repeated episodes of injury and may be beneficial as preventive therapy for patients with or at risk of developing coronary ischemic events.

  17. A Novel Nonviral Gene Delivery System: Multifunctional Envelope-Type Nano Device

    Science.gov (United States)

    Hatakeyama, Hiroto; Akita, Hidetaka; Kogure, Kentaro; Harashima, Hideyoshi

    In this review we introduce a new concept for developing a nonviral gene delivery system which we call "Programmed Packaging." Based on this concept, we succeeded in developing a multifunctional envelope-type nano device (MEND), which exerts high transfection activities equivalent to those of an adenovirus in a dividing cell. The use of MEND has been extended to in vivo applications. PEG/peptide/DOPE ternary conjugate (PPD)-MEND, a new in vivo gene delivery system for the targeting of tumor cells that dissociates surface-modified PEG in tumor tissue by matrix metalloproteinase (MMP) and exerts significant transfection activities, was developed. In parallel with the development of MEND, a quantitative gene delivery system, Confocal Image-assisted 3-dimensionally integrated quantification (CIDIQ), also was developed. This method identified the rate-limiting step of the nonviral gene delivery system by comparing it with adenoviral-mediated gene delivery. The results of this analysis provide a new direction for the development of rational nonviral gene delivery systems.

  18. Nano-scale gene delivery systems; current technology, obstacles, and future directions.

    Science.gov (United States)

    Garcia-Guerra, Antonio; Dunwell, Thomas L; Trigueros, Sonia

    2018-01-07

    Within the different applications of nanomedicine currently being developed, nano-gene delivery is appearing as an exciting new technique with the possibility to overcome recognised hurdles and fulfill several biological and medical needs. The central component of all delivery systems is the requirement for the delivery of genetic material into cells, and for them to eventually reside in the nucleus where their desired function will be exposed. However, genetic material does not passively enter cells; thus, a delivery system is necessary. The emerging field of nano-gene delivery exploits the use of new materials and the properties that arise at the nanometre-scale to produce delivery vectors that can effectively deliver genetic material into a variety of different types of cells. The novel physicochemical properties of the new delivery vectors can be used to address the current challenges existing in nucleic acid delivery in vitro and in vivo. While there is a growing interest in nanostructure-based gene delivery, the field is still in its infancy, and there is yet much to discover about nanostructures and their physicochemical properties in a biological context. We carry out an organized and focused search of bibliographic databases. Our results suggest that despite new breakthroughs in nanostructure synthesis and advanced characterization techniques, we still face many barriers in producing highly efficient and non-toxic delivery systems. In this review, we overview the types of systems currently used for clinical and biomedical research applications along with their advantages and disadvantages, as well as discussing barriers that arise from nano-scale interactions with biological material. In conclusion, we hope that by bringing the far reaching multidisciplinary nature of nano-gene delivery to light, new targeted nanotechnology-bases strategies are developed to overcome the major challenges covered in this review. Copyright© Bentham Science Publishers; For

  19. Exploring the role of peptides in polymer-based gene delivery.

    Science.gov (United States)

    Sun, Yanping; Yang, Zhen; Wang, Chunxi; Yang, Tianzhi; Cai, Cuifang; Zhao, Xiaoyun; Yang, Li; Ding, Pingtian

    2017-09-15

    Polymers are widely studied as non-viral gene vectors because of their strong DNA binding ability, capacity to carry large payload, flexibility of chemical modifications, low immunogenicity, and facile processes for manufacturing. However, high cytotoxicity and low transfection efficiency substantially restrict their application in clinical trials. Incorporating functional peptides is a promising approach to address these issues. Peptides demonstrate various functions in polymer-based gene delivery systems, such as targeting to specific cells, breaching membrane barriers, facilitating DNA condensation and release, and lowering cytotoxicity. In this review, we systematically summarize the role of peptides in polymer-based gene delivery, and elaborate how to rationally design polymer-peptide based gene delivery vectors. Polymers are widely studied as non-viral gene vectors, but suffer from high cytotoxicity and low transfection efficiency. Incorporating short, bioactive peptides into polymer-based gene delivery systems can address this issue. Peptides demonstrate various functions in polymer-based gene delivery systems, such as targeting to specific cells, breaching membrane barriers, facilitating DNA condensation and release, and lowering cytotoxicity. In this review, we highlight the peptides' roles in polymer-based gene delivery, and elaborate how to utilize various functional peptides to enhance the transfection efficiency of polymers. The optimized peptide-polymer vectors should be able to alter their structures and functions according to biological microenvironments and utilize inherent intracellular pathways of cells, and consequently overcome the barriers during gene delivery to enhance transfection efficiency. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  20. Mass transfer of therapeutics through natural human plaque biofilms: a model for therapeutic delivery to pathological bacterial biofilms.

    Science.gov (United States)

    Robinson, Colin

    2011-09-01

    Bacterial biofilms in the mouth are prime mediators of the destruction of the dental and oral tissues. This brief review summarises recent work using a device for generating intact plaque in the mouth on natural enamel surfaces such that quantitative studies of mass transfer through natural plaque biofilms could be carried out in relation to plaque architecture. This data is discussed against the background of existing information. The device revealed complex plaque architecture with high a surface area to mass ratio decreasing from the exterior of the biofilm towards the tissue surface. Fluoride, a potent inhibitor of caries was concentrated in the outer regions of the biofilm. This implies some restriction of diffusion and possibly binding to the high surface area of the outer biofilm. Whilst all components examined conformed to this distribution pattern, some relatively uncharged materials penetrated the bacterial biomass whilst other, more highly charged materials tended to be restricted to the channels or biomass surface. Plaque architecture was robust but could be altered using detergent indicating that biomass architecture and chemistry could be manipulated as a possible means of facilitating mass transport of therapeutics. Copyright © 2011 Elsevier Ltd. All rights reserved.

  1. Targeted Delivery of CRISPR/Cas9-Mediated Cancer Gene Therapy via Liposome-Templated Hydrogel Nanoparticles.

    Science.gov (United States)

    Chen, Zeming; Liu, Fuyao; Chen, Yanke; Liu, Jun; Wang, Xiaoying; Chen, Ann T; Deng, Gang; Zhang, Hongyi; Liu, Jie; Hong, Zhangyong; Zhou, Jiangbing

    2017-12-08

    Due to its simplicity, versatility, and high efficiency, the clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 technology has emerged as one of the most promising approaches for treatment of a variety of genetic diseases, including human cancers. However, further translation of CRISPR/Cas9 for cancer gene therapy requires development of safe approaches for efficient, highly specific delivery of both Cas9 and single guide RNA to tumors. Here, novel core-shell nanostructure, liposome-templated hydrogel nanoparticles (LHNPs) that are optimized for efficient codelivery of Cas9 protein and nucleic acids is reported. It is demonstrated that, when coupled with the minicircle DNA technology, LHNPs deliver CRISPR/Cas9 with efficiency greater than commercial agent Lipofectamine 2000 in cell culture and can be engineered for targeted inhibition of genes in tumors, including tumors the brain. When CRISPR/Cas9 targeting a model therapeutic gene, polo-like kinase 1 (PLK1), is delivered, LHNPs effectively inhibit tumor growth and improve tumor-bearing mouse survival. The results suggest LHNPs as versatile CRISPR/Cas9-delivery tool that can be adapted for experimentally studying the biology of cancer as well as for clinically translating cancer gene therapy.

  2. Development of Liposomal Bubbles with Perfluoropropane Gas as Gene Delivery Carriers

    Science.gov (United States)

    Maruyama, Kazuo; Suzuki, Ryo; Sawamura, Kaori; Takizawa, Tomoko; Utoguchi, Naoki; Negishi, Yoichi

    2007-05-01

    Liposomes have some advantages as drug, antigen and gene delivery carriers. Their size can be easily controlled and they can be modified to add a targeting function. Based on liposome technology, we developed novel liposomal bubbles (Bubble liposomes) containing the ultrasound imaging gas, perfluoropropane. We assessed the feasibility of Bubble liposomes as carriers for gene delivery after cavitation induced by ultrasound. At first, we investigated their ability to deliver genes with Bubble liposomes and ultrasound to various types of cells such as mouse sarcoma cells, mouse melanoma cells, human T cell line and human umbilical vein endothelial cells. The results showed that the Bubble liposomes could deliver plasmid DNA to many cell types without cytotoxicity. In addition, we found that Bubble liposomes could effectively deliver plasmid DNA into mouse femoral artery in vivo. The gene transduction with Bubble liposomes was more effectively than conventional lipofection. We conclude that Bubble liposomes are unique and efficient gene delivery carriers in vitro and in vivo.

  3. ScreenFect A: an efficient and low toxic liposome for gene delivery to mesenchymal stem cells.

    Science.gov (United States)

    Li, Li-Ming; Ruan, Gui-Xin; HuangFu, Ming-Yi; Chen, Zhi-Lan; Liu, Hui-Na; Li, Lin-Xian; Hu, Yu-Lan; Han, Min; Davidson, Gary; Levkin, Pavel A; Gao, Jian-Qing

    2015-07-05

    Mesenchymal stem cells (MSCs) hold great promise in variety of therapeutic applications including tissue engineering and cancer therapy. Genetic modification of MSCs can be used to enhance the therapeutic effect of MSCs by facilitating a specific function or by transforming MSCs into more effective gene therapy tools. However, the successful generation of genetically modified MSCs is often limited by the poor transfection efficiency or high toxicity of available transfection reagents. In our previous study, we used thiol-yne click chemistry to develop new liposomal vectors, including ScreenFect(®) A (SF) (Li et al., 2012). In this study, we investigated the transfection performance of SF on MSCs. A comparative evaluation of transfection efficiency, cell viability and cellular DNA uptake was performed using the Lipofectamine™ 2000 (L2K) as a control, and the results show that SF is superior to L2K for MSC transfection. The presence of serum did not significantly influence the transfection efficiency of either SF or L2K but greatly reduced the viability of MSC transfected by L2K. The higher efficiency of SF-mediated transfection compared to L2K was also correlated with better proliferation of cells. These results were supported by monitoring the intracellular fate of DNA, which confirmed stable transportation of DNA from lysosomes and efficient nuclear localization. TGF-β1 gene delivery by SF promoted MSC osteogenic differentiation in an osteogenic induction condition. As the first study of SF lipofection on stem cells, this study highlights a promising role of SF in gene delivery to MSCs as well as other stem cells to facilitate tissue engineering and other therapeutic effects based on genetically modified stem cells. Copyright © 2015 Elsevier B.V. All rights reserved.

  4. Three-dimensional tumor spheroids for in vitro analysis of bacteria as gene delivery vectors in tumor therapy.

    Science.gov (United States)

    Osswald, Annika; Sun, Zhongke; Grimm, Verena; Ampem, Grace; Riegel, Karin; Westendorf, Astrid M; Sommergruber, Wolfgang; Otte, Kerstin; Dürre, Peter; Riedel, Christian U

    2015-12-12

    Several studies in animal models demonstrated that obligate and facultative anaerobic bacteria of the genera Bifidobacterium, Salmonella, or Clostridium specifically colonize solid tumors. Consequently, these and other bacteria are discussed as live vectors to deliver therapeutic genes to inhibit tumor growth. Therapeutic approaches for cancer treatment using anaerobic bacteria have been investigated in different mouse models. In the present study, solid three-dimensional (3D) multicellular tumor spheroids (MCTS) of the colorectal adenocarcinoma cell line HT-29 were generated and tested for their potential to study prodrug-converting enzyme therapies using bacterial vectors in vitro. HT-29 MCTS resembled solid tumors displaying all relevant features with an outer zone of proliferating cells and hypoxic and apoptotic regions in the core. Upon incubation with HT-29 MCTS, Bifidobacterium bifidum S17 and Salmonella typhimurium YB1 selectively localized, survived and replicated in hypoxic areas inside MCTS. Furthermore, spores of the obligate anaerobe Clostridium sporogenes germinated in these hypoxic areas. To further evaluate the potential of MCTS to investigate therapeutic approaches using bacteria as gene delivery vectors, recombinant bifidobacteria expressing prodrug-converting enzymes were used. Expression of a secreted cytosine deaminase in combination with 5-fluorocytosine had no effect on growth of MCTS due to an intrinsic resistance of HT-29 cells to 5-fluorouracil, i.e. the converted drug. However, a combination of the prodrug CB1954 and a strain expressing a secreted chromate reductase effectively inhibited MCTS growth. Collectively, the presented results indicate that MCTS are a suitable and reliable model to investigate live bacteria as gene delivery vectors for cancer therapy in vitro.

  5. Pediatric Multiple Sclerosis: Genes, Environment, and a Comprehensive Therapeutic Approach.

    Science.gov (United States)

    Cappa, Ryan; Theroux, Liana; Brenton, J Nicholas

    2017-10-01

    Pediatric multiple sclerosis is an increasingly recognized and studied disorder that accounts for 3% to 10% of all patients with multiple sclerosis. The risk for pediatric multiple sclerosis is thought to reflect a complex interplay between environmental and genetic risk factors. Environmental exposures, including sunlight (ultraviolet radiation, vitamin D levels), infections (Epstein-Barr virus), passive smoking, and obesity, have been identified as potential risk factors in youth. Genetic predisposition contributes to the risk of multiple sclerosis, and the major histocompatibility complex on chromosome 6 makes the single largest contribution to susceptibility to multiple sclerosis. With the use of large-scale genome-wide association studies, other non-major histocompatibility complex alleles have been identified as independent risk factors for the disease. The bridge between environment and genes likely lies in the study of epigenetic processes, which are environmentally-influenced mechanisms through which gene expression may be modified. This article will review these topics to provide a framework for discussion of a comprehensive approach to counseling and ultimately treating the pediatric patient with multiple sclerosis. Copyright © 2017 Elsevier Inc. All rights reserved.

  6. Gene electrotransfer in clinical trials

    DEFF Research Database (Denmark)

    Gehl, Julie

    2014-01-01

    Electroporation is increasingly being used for delivery of chemotherapy to tumors. Likewise, gene delivery by electroporation is rapidly gaining momentum for both vaccination purposes and for delivery of genes coding for other therapeutic molecules, such as chronic diseases or cancer. This chapter...... describes how gene therapy may be performed using electric pulses to enhance uptake and expression....

  7. Synergistic effect of amino acids modified on dendrimer surface in gene delivery.

    Science.gov (United States)

    Wang, Fei; Wang, Yitong; Wang, Hui; Shao, Naimin; Chen, Yuanyuan; Cheng, Yiyun

    2014-11-01

    Design of an efficient gene vector based on dendrimer remains a great challenge due to the presence of multiple barriers in gene delivery. Single-functionalization on dendrimer cannot overcome all the barriers. In this study, we synthesized a list of single-, dual- and triple-functionalized dendrimers with arginine, phenylalanine and histidine for gene delivery using a one-pot approach. The three amino acids play different roles in gene delivery: arginine is essential in formation of stable complexes, phenylalanine improves cellular uptake efficacy, and histidine increases pH-buffering capacity and minimizes cytotoxicity of the cationic dendrimer. A combination of these amino acids on dendrimer generates a synergistic effect in gene delivery. The dual- and triple-functionalized dendrimers show minimal cytotoxicity on the transfected NIH 3T3 cells. Using this combination strategy, we can obtain triple-functionalized dendrimers with comparable transfection efficacy to several commercial transfection reagents. Such a combination strategy should be applicable to the design of efficient and biocompatible gene vectors for gene delivery. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. GFP expression by intracellular gene delivery of GFP-coding fragments using nanocrystal quantum dots

    International Nuclear Information System (INIS)

    Hoshino, Akiyoshi; Manabe, Noriyoshi; Fujioka, Kouki; Hanada, Sanshiro; Yamamoto, Kenji; Yasuhara, Masato; Kondo, Akihiko

    2008-01-01

    Gene therapy is an attractive approach to supplement a deficient gene function. Although there has been some success with specific gene delivery using various methods including viral vectors and liposomes, most of these methods have a limited efficiency or also carry a risk for oncogenesis. We herein report that quantum dots (QDs) conjugated with nuclear localizing signal peptides (NLSP) successfully introduced gene-fragments with promoter elements, which promoted the expression of the enhanced green fluorescent protein (eGFP) gene in mammalian cells. The expression of eGFP protein was observed when the QD/gene-construct was added to the culture media. The gene-expression efficiency varied depending on multiple factors around QDs, such as (1) the reading direction of the gene-fragments, (2) the quantity of gene-fragments attached on the surface of the QD-constructs, (3) the surface electronic charges varied according to the structure of the QD/gene-constructs, and (4) the particle size of QD/gene complex varied according to the structure and amounts of gene-fragments. Using this QD/gene-construct system, eGFP protein could be detected 28 days after the gene-introduction whereas the fluorescence of QDs had disappeared. This system therefore provides another method for the intracellular delivery of gene-fragments without using either viral vectors or specific liposomes.

  9. Transdermal and intradermal delivery of therapeutic agents: application of physical technologies

    National Research Council Canada - National Science Library

    Banga, Ajay K

    2011-01-01

    .... Advancements in science combined with the need for diverse drug delivery modalities have introduced a variety of transdermal and intradermal products for existing drugs at a fraction of the cost of new drug development...

  10. Nanovectors for Targeting and Delivery of Therapeutics to HER-2 NEU Positive Breast Cancer Cells

    National Research Council Canada - National Science Library

    Serda, Rita E

    2008-01-01

    ...s to reach the tumor site. To overcome these biological barriers, we have developed a multistage delivery system comprised of biocompatible porous silicon particles that encapsulate iron oxide nanoparticles for protection...

  11. Neutrophil-Mediated Delivery of Therapeutic Nanoparticles across Blood Vessel Barrier for Treatment of Inflammation and Infection.

    Science.gov (United States)

    Chu, Dafeng; Gao, Jin; Wang, Zhenjia

    2015-12-22

    Endothelial cells form a monolayer in lumen of blood vessels presenting a great barrier for delivery of therapeutic nanoparticles (NPs) into extravascular tissues where most diseases occur, such as inflammation disorders and infection. Here, we report a strategy for delivering therapeutic NPs across this blood vessel barrier by nanoparticle in situ hitchhiking activated neutrophils. Using intravital microscopy of TNF-α-induced inflammation of mouse cremaster venules and a mouse model of acute lung inflammation, we demonstrated that intravenously (iv) infused NPs made from denatured bovine serum albumin (BSA) were specifically internalized by activated neutrophils, and subsequently, the neutrophils containing NPs migrated across blood vessels into inflammatory tissues. When neutrophils were depleted using anti-Gr-1 in a mouse, the transport of albumin NPs across blood vessel walls was robustly abolished. Furthermore, it was found that albumin nanoparticle internalization did not affect neutrophil mobility and functions. Administration of drug-loaded albumin NPs markedly mitigated the lung inflammation induced by LPS (lipopolysaccharide) or infection by Pseudomonas aeruginosa. These results demonstrate the use of an albumin nanoparticle platform for in situ targeting of activated neutrophils for delivery of therapeutics across the blood vessel barriers into diseased sites. This study demonstrates our ability to hijack neutrophils to deliver nanoparticles to targeted diseased sites.

  12. rAAV Vectors as Safe and Efficient Tools for the Stable Delivery of Genes to Primary Human Chondrosarcoma Cells In Vitro and In Situ

    Directory of Open Access Journals (Sweden)

    Henning Madry

    2012-01-01

    Full Text Available Treatment of chondrosarcoma remains a major challenge in orthopaedic oncology. Gene transfer strategies based on recombinant adenoassociated viral (rAAV vectors may provide powerful tools to develop new, efficient therapeutic options against these tumors. In the present study, we tested the hypothesis that rAAV is adapted for a stable and safe delivery of foreign sequences in human chondrosarcoma tissue by transducing primary human chondrosarcoma cells in vitro and in situ with different reporter genes (E. coli lacZ, firefly luc, Discosoma sp. RFP. The effects of rAAV administration upon cell survival and metabolic activities were also evaluated to monitor possibly detrimental effects of the gene transfer method. Remarkably, we provide evidence that efficient and prolonged expression of transgene sequences via rAAV can be safely achieved in all the systems investigated, demonstrating the potential of the approach of direct application of therapeutic gene vectors as a means to treat chondrosarcoma.

  13. Synthetic polyspermine imidazole-4, 5-amide as an efficient and cytotoxicity-free gene delivery system

    Directory of Open Access Journals (Sweden)

    Duan S

    2012-07-01

    Full Text Available Shi-Yue Duan, Xue-Mei Ge, Nan Lu, Fei Wu, Weien Yuan, Tuo JinSchool of Pharmacy, Shanghai Jiao Tong University, Shanghai, 200240, People's Republic of ChinaAbstract: A chemically dynamic spermine-based polymer: polyspermine imidazole-4, 5-amide (PSIA, Mw > 7 kDa was designed, synthesized, and evaluated in terms of its ability to deliver nucleic acids. This polymer was made from an endogenous monomer professionally condensing genes in sperms, spermine, and a known safety drug metabolite, imidazole-4, 5-dicarboxylic acid, through a bis-amide bond conjugated with the imidazole ring. This polymer can condense pDNA at a W/W ratio above 10 to form polyplexes (100–200 nm in diameter, which is consistent with the observation by transmission electron microscopy (TEM, and the zeta potential was in the range of 10–20 mV. The pDNA packaged polymer was stable in phosphate buffer solution (PBS at pH 7.4 (simulated body fluid while the polyplexes were releasing pDNA into the solution at pH 5.8 (simulated endo-lysosomes due to the degradation of the bis-amide linkages in response to changes in pH values. PSIA-polyplexes were able to achieve efficient cellular uptake and luciferase gene silencing by co-transfection of pDNA and siRNA in COS-7 cells and HepG2 cells with negligible cytotoxicity. Biodistribution of Rhodamine B-labeled PSIA-polyplexes after being systemically injected in BALB/c nude-mice showed that the polyplexes circulated throughout the body, accumulated mainly in the kidney at 4 hours of sample administration, and moved to the liver and spleen after 24 hours. All the results suggested that PSIA offered a promising example to balance the transfection efficiency and toxicity of a synthetic carrier system for the delivery of therapeutic nucleic acids.Keywords: gene delivery, polyspermine, cytotoxicity, transfection efficiency, biodistribution

  14. Polylysine-modified polyethylenimine (PEI-PLL) mediated VEGF gene delivery protects dopaminergic neurons in cell culture and in rat models of Parkinson's Disease (PD).

    Science.gov (United States)

    Sheikh, Muhammad Abid; Malik, Yousra Saeed; Xing, Zhenkai; Guo, Zhaopei; Tian, Huayu; Zhu, Xiaojuan; Chen, Xuesi

    2017-05-01

    Parkinson's Disease (PD) is a chronic neurodegenerative disorder characterized by motor deficits which result from the progressive loss of dopaminergic neurons. Gene therapy using growth factors such as VEGF seems to be a viable approach for potential therapeutic treatment of PD. In this study, we utilized a novel non-viral gene carrier designated as PEI-PLL synthesized by our laboratory to deliver VEGF gene to study its effect by using both cell culture as well as animal models of PD. For cell culture experiments, we utilized 6-hydroxydopamine (6-OHDA) mediated cell death model of MN9D cells following transfection with either a control plasmid or VEGF expressing plasmid. As compared to control transfected cells, PEI-PLL mediated VEGF gene delivery to MN9D cells resulted in increased cell viability, increase in the number of Tyrosine hydroxylase (TH) positive cells and decreased apoptosis following 6-OHDA insult. Next, we studied the therapeutic potential of PEI-PLL mediated VEGF gene delivery in SNPc by using unilateral 6-OHDA Medial forebrain bundle (MFB) lesion model of PD in rats. VEGF administration prevented the loss of motor functions induced by 6-OHDA as determined by behavior analysis. Similarly, VEGF inhibited the 6-OHDA mediated loss of DA neurons in Substantia Nigra Pars Compacta (SNPc) as well as DA nerve fibers in striatum as determined by TH immunostaining. In addition, PEI-PLL mediated VEGF gene delivery also prevented apoptosis and microglial activation in PD rat models. Together, these results clearly demonstrated the beneficial effects of PEI-PLL mediated VEGF gene delivery on dopaminergic system in both cell culture and animal models of PD. In this report, we exploited the potential of PEI-PLL to deliver VEGF gene for the potential therapeutic treatment of PD by using both cell culture and animal models of PD. To the best of our knowledge, this is the first report describing the use of novel polymeric gene carriers for the delivery of VEGF gene

  15. Effective Nanoparticle-based Gene Delivery by a Protease Triggered Charge Switch

    DEFF Research Database (Denmark)

    Gjetting, Torben; Jølck, Rasmus Irming; Andresen, Thomas Lars

    2014-01-01

    Gene carriers made from synthetic materials are of interest in relation to gene therapy but suffer from lack of transfection efficiency upon systemic delivery. To address this problem, a novel lipo-peptide-PEG conjugate constituted by a lipid-anchor, a peptide sensitive to proteases and a poly (e...

  16. Quaternized Polyamidoamine Dendrimers as Novel Gene Delivery System: Relationship between Degree of Quaternization and Their Influences

    International Nuclear Information System (INIS)

    Lee, Jung Hoon; Lim, Yong beom; Choi, Joon Sig; Choi, Myung Un; Yang, Chul Hak; Park, Jong Sang

    2003-01-01

    Quaternary ammonium groups were introduced to Starburst polyamidoamine (PAMAM) dendrimers for a gene carrier. These quaternary dendritic carriers exhibited reduced cytotoxicity on 293T cells compared to parent dendrimers examined and their transfection efficiency were similar with parent dendrimers. Quaternization could be a promising tool to improve properties of dendrimers as a gene delivery carrier

  17. Horizontal gene transfer from macrophages to ischemic muscles upon delivery of naked DNA with Pluronic block copolymers.

    Science.gov (United States)

    Mahajan, Vivek; Gaymalov, Zagit; Alakhova, Daria; Gupta, Richa; Zucker, Irving H; Kabanov, Alexander V

    2016-01-01

    Intramuscular administration of plasmid DNA (pDNA) with non-ionic Pluronic block copolymers increases gene expression in injected muscles and lymphoid organs. We studied the role of immune cells in muscle transfection upon inflammation. Local inflammation in murine hind limb ischemia model (MHLIM) drastically increased DNA, RNA and expressed protein levels in ischemic muscles injected with pDNA/Pluronic. The systemic inflammation (MHLIM or peritonitis) also increased expression of pDNA/Pluronic in the muscles. When pDNA/Pluronic was injected in ischemic muscles the reporter gene, Green Fluorescent Protein (GFP) co-localized with desmin(+) muscle fibers and CD11b(+) macrophages (MØs), suggesting transfection of MØs along with the muscle cells. P85 enhanced (∼ 4 orders) transfection of MØs with pDNA in vitro. Moreover, adoptively transferred MØs were shown to pass the transgene to inflamed muscle cells in MHLIM. Using a co-culture of myotubes (MTs) and transfected MØs expressing a reporter gene under constitutive (cmv-luciferase) or muscle specific (desmin-luciferase) promoter we demonstrated that P85 enhances horizontal gene transfer from MØ to MTs. Therefore, MØs can play an important role in muscle transfection with pDNA/Pluronic during inflammation, with both inflammation and Pluronic contributing to the increased gene expression. pDNA/Pluronic has potential for therapeutic gene delivery in muscle pathologies that involve inflammation. Copyright © 2015 Elsevier Ltd. All rights reserved.

  18. Efficient intranuclear gene delivery by CdSe aqueous quantum dots electrostatically-coated with polyethyleneimine

    International Nuclear Information System (INIS)

    Au, Giang H T; Shih, Wan Y; Shih, Wei-Heng

    2015-01-01

    Quantum dots (QDs) are semiconducting nanoparticles with photoluminescence properties that do not photobleach. Due to these advantages, using QDs for non-viral gene delivery has the additional benefit of being able to track the delivery of the genes in real time as it happens. We investigate the efficacy of mercaptopropionic acid (MPA)-capped CdSe aqueous quantum dots (AQDs) electrostatically complexed with branched polyethyleneimine (PEI) both as a non-viral gene delivery vector and as a fluorescent probe for tracking the delivery of genes into nuclei. The MPA-capped CdSe AQDs that were completely synthesized in water were the model AQDs. A nominal MPA:Cd:Se = 4:3:1 was chosen for optimal photoluminescence and zeta potential. The gene delivery study was carried out in vitro using a human colon cancer cell line, HT29 (ATCC). The model gene was a plasmid DNA (pDNA) that can express red fluorescent protein (RFP). Positively charged branched PEI was employed to provide a proton buffer to the AQDs to allow for endosomal escape. It is shown that by using a PEI-AQD complex with a PEI/AQD molar ratio of 300 and a nominal pDNA/PEI-AQD ratio of 6, we can achieve 75 ± 2.6% RFP expression efficiency with cell vitality remaining at 78 ± 4% of the control. (paper)

  19. Delivery of Therapeutic Proteins Using Electrospun Fibers-Recent Developments and Current Challenges.

    Science.gov (United States)

    Seif, Salem; Planz, Viktoria; Windbergs, Maike

    2017-10-01

    Proteins play a vital role within the human body by regulating various functions and even serving as structural constituent of many body parts. In this context, protein-based therapeutics have attracted a lot of attention in the last few decades as potential treatment of different diseases. Due to the steadily increasing interest in protein-based therapeutics, different dosage forms were investigated for delivering such complex macromolecules to the human body. Here, electrospun fibers hold a great potential for embedding proteins without structural damage and for controlled release of the protein for therapeutic applications. This review provides a comprehensive overview of the current state of protein-based carrier systems using electrospun fibers, with special emphasis on discussing their potential and key challenges in developing such therapeutic strategies, along with a prospective view of anticipated future directions. © 2017 Deutsche Pharmazeutische Gesellschaft.

  20. Bifidobacterium breve as a delivery vector of IL-24 gene therapy for head and neck squamous cell carcinoma in vivo.

    Science.gov (United States)

    Wang, L; Vuletic, I; Deng, D; Crielaard, W; Xie, Z; Zhou, K; Zhang, J; Sun, H; Ren, Q; Guo, C

    2017-11-01

    Beneficial bacteria are becoming ever more popular gene delivery method for hypoxia-tumor targeting in vivo. In this study we investigated the therapeutic effect of new recombinant Bifidobacterium breve strain expressing interleukin (IL)-24 gene (B. breve-IL24) on head and neck tumor xenograft in mice. Briefly, B. breve transformants were obtained through electro-transformation. Bacteria-tumor-targeting ability were analyzed in vivo over different time points (1, 3 and 7 days post-bacteria injection). Furthermore, the therapeutic effect of bacteria on tumor cells in vivo were analyzed as follows: 30 Balb/c nude mice bearing subcutaneous tumor were randomly divided in three groups (Drug group, green fluorescent protein (GFP) group and Saline group). The therapy lasted for 2 weeks and included B. breve-IL24 administration via tail vein for Drug group, B. breve-GFP for GFP group and phosphate buffered saline for Saline group. The tumor growth was monitored using standard caliper technique, while the apoptosis induction in vivo was analyzed by Real-time Positron Emission Tomography/Computed Tomography (PET/CT) imaging ([18F]-ML-10 tracer). At the end of the experiment, tumor tissues were collected and analyzed by western blotting. Briefly, our results suggested that our new recombinant bacterium has the capability of targeting tumor tissue in vivo. As for the therapeutic effect, our new strain has revealed to be a promising therapeutic approach against tumor growth in vivo. Briefly, higher tumor growth inhibition and higher tumor cell apoptosis induction were observed in Drug group compared with the GFP and Saline groups. To conclude, a new recombinant strain B. breve-IL24 offers a novel, safe and clinically acceptable therapeutic approach for tumor therapy in vivo.

  1. Ivacaftor: A Novel Gene-Based Therapeutic Approach for Cystic Fibrosis

    OpenAIRE

    Condren, Michelle E.; Bradshaw, Marquita D.

    2013-01-01

    Ivacaftor is a new therapeutic agent that acts at the cystic fibrosis transmembrane conductance regulator (CFTR) channel to alter activity. It is approved for use in patients 6 years and older with cystic fibrosis who have at least 1 G551D mutation in the CFTR gene. It is unlike any other current pharmacologic agent for cystic fibrosis in that it specifically targets the gene defect associated with cystic fibrosis as opposed to treating resulting symptomology. Mucoactive agents, antibiotics, ...

  2. Gene Therapy for Advanced Melanoma: Selective Targeting and Therapeutic Nucleic Acids

    Directory of Open Access Journals (Sweden)

    Joana R. Viola

    2013-01-01

    Full Text Available Despite recent advances, the treatment of malignant melanoma still results in the relapse of the disease, and second line treatment mostly fails due to the occurrence of resistance. A wide range of mutations are known to prevent effective treatment with chemotherapeutic drugs. Hence, approaches with biopharmaceuticals including proteins, like antibodies or cytokines, are applied. As an alternative, regimens with therapeutically active nucleic acids offer the possibility for highly selective cancer treatment whilst avoiding unwanted and toxic side effects. This paper gives a brief introduction into the mechanism of this devastating disease, discusses the shortcoming of current therapy approaches, and pinpoints anchor points which could be harnessed for therapeutic intervention with nucleic acids. We bring the delivery of nucleic acid nanopharmaceutics into perspective as a novel antimelanoma therapeutic approach and discuss the possibilities for melanoma specific targeting. The latest reports on preclinical and already clinical application of nucleic acids in melanoma are discussed.

  3. Self-assembled pentablock copolymers for selective and sustained gene delivery

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Bingqi [Iowa State Univ., Ames, IA (United States)

    2011-05-15

    The poly(diethylaminoethyl methacrylate) (PDEAEM) - Pluronic F127 - PDEAEM pentablock copolymer (PB) gene delivery vector system has been found to possess an inherent selectivity in transfecting cancer cells over non-cancer cells in vitro, without attaching any targeting ligands. In order to understand the mechanism of this selective transfection, three possible intracellular barriers to transfection were investigated in both cancer and non-cancer cells. We concluded that escape from the endocytic pathway served as the primary intracellular barrier for PB-mediated transfection. Most likely, PB vectors were entrapped and rendered non-functional in acidic lysosomes of non-cancer cells, but survived in less acidic lysosomes of cancer cells. The work highlights the importance of identifying intracellular barriers for different gene delivery systems and provides a new paradigm for designing targeting vectors based on intracellular differences between cell types, rather than through the use of targeting ligands. The PB vector was further developed to simultaneously deliver anticancer drugs and genes, which showed a synergistic effect demonstrated by significantly enhanced gene expression in vitro. Due to the thermosensitive gelation behavior, the PB vector packaging both drug and gene was also investigated for its in vitro sustained release properties by using polyethylene glycol diacrylate as a barrier gel to mimic the tumor matrix in vivo. Overall, this work resulted in the development of a gene delivery vector for sustained and selective gene delivery to tumor cells for cancer therapy.

  4. The emerging pathogenic and therapeutic importance of the anaplastic lymphoma kinase gene.

    LENUS (Irish Health Repository)

    Kelleher, Fergal C

    2012-02-01

    The anaplastic lymphoma kinase gene (ALK) is a gene on chromosome 2p23 that has expression restricted to the brain, testis and small intestine but is not expressed in normal lymphoid tissue. It has similarity to the insulin receptor subfamily of kinases and is emerging as having increased pathologic and potential therapeutic importance in malignant disease. This gene was originally established as being implicated in the pathogenesis of rare diseases including inflammatory myofibroblastic tumour (IMT) and ALK-positive anaplastic large cell lymphoma, which is a subtype of non-Hodgkin\\'s lymphoma. Recently the number of diseases in which ALK is implicated in their pathogenesis has increased. In 2007, an inversion of chromosome 2 involving ALK and a fusion partner gene in a subset of non-small cell lung cancer was discovered. In 2008, publications emerged implicating ALK in familial and sporadic cases of neuroblastoma, a childhood cancer of the sympatho-adrenal system. Chromosomal abnormalities involving ALK are translocations, amplifications or mutations. Chromosomal translocations are the longest recognised ALK genetic abnormality. When translocations occur a fusion gene is created between ALK and a gene partner. This has been described in ALK-positive anaplastic large cell lymphoma in which ALK is fused to NPM (nucleolar protein gene) and in non-small cell lung cancer where ALK is fused to EML4 (Echinoderm microtubule-associated protein 4). The most frequently described partner genes in inflammatory myofibroblastic tumour are tropomyosin 3\\/4 (TMP3\\/4), however in IMTs a diversity of ALK fusion partners have been found, with the ability to homodimerise a common characteristic. Point mutations and amplification of the ALK gene occur in the childhood cancer neuroblastoma. Therapeutic targeting of ALK fusion genes using tyrosine kinase inhibition, vaccination using an ALK specific antigen and treatment using viral vectors for RNAi are emerging potential therapeutic

  5. Tropism-Modification Strategies for Targeted Gene Delivery Using Adenoviral Vectors

    Directory of Open Access Journals (Sweden)

    Andrew H. Baker

    2010-10-01

    Full Text Available Achieving high efficiency, targeted gene delivery with adenoviral vectors is a long-standing goal in the field of clinical gene therapy. To achieve this, platform vectors must combine efficient retargeting strategies with detargeting modifications to ablate native receptor binding (i.e. CAR/integrins/heparan sulfate proteoglycans and “bridging” interactions. “Bridging” interactions refer to coagulation factor binding, namely coagulation factor X (FX, which bridges hepatocyte transduction in vivo through engagement with surface expressed heparan sulfate proteoglycans (HSPGs. These interactions can contribute to the off-target sequestration of Ad5 in the liver and its characteristic dose-limiting hepatotoxicity, thereby significantly limiting the in vivo targeting efficiency and clinical potential of Ad5-based therapeutics. To date, various approaches to retargeting adenoviruses (Ad have been described. These include genetic modification strategies to incorporate peptide ligands (within fiber knob domain, fiber shaft, penton base, pIX or hexon, pseudotyping of capsid proteins to include whole fiber substitutions or fiber knob chimeras, pseudotyping with non-human Ad species or with capsid proteins derived from other viral families, hexon hypervariable region (HVR substitutions and adapter-based conjugation/crosslinking of scFv, growth factors or monoclonal antibodies directed against surface-expressed target antigens. In order to maximize retargeting, strategies which permit detargeting from undesirable interactions between the Ad capsid and components of the circulatory system (e.g. coagulation factors, erythrocytes, pre-existing neutralizing antibodies, can be employed simultaneously. Detargeting can be achieved by genetic ablation of native receptor-binding determinants, ablation of “bridging interactions” such as those which occur between the hexon of Ad5 and coagulation factor X (FX, or alternatively, through the use of polymer

  6. In Vitro and In Vivo Effective Gene Delivery with Novel Liposomal Bubbles

    Science.gov (United States)

    Nishiie, Norihito; Suzuki, Ryo; Oda, Yusuke; Hirata, Keiichi; Taira, Yuichiro; Utoguchi, Naoki; Negishi, Yoichi; Maruyama, Kazuo

    2010-03-01

    Microbubbles, which were ultrasound contrast agents, could improve the transfection efficiency by cavitation with ultrasound exposure. However, conventional microbubbles had some problems regarding size and targeting ability. To solve these problems, we paid attention to liposomes that had many advantages as drug, antigen and gene delivery carriers. Because they can easily be controlled their size and added a targeting function. And we succeeded to prepare novel liposomal bubbles (Bubble liposomes) entrapping perfluoropropane which was utilized for contrast enhancement in ultrasonography. In this study, we assessed the feasibility of Bubble liposomes as gene delivery tools utilized cavitation by ultrasound exposure. In vitro gene delivery, Bubble liposomes could deliver plasmid DNA to many cell types such as tumor cells, T cell line and endothelial cells without cytotoxicity. In vivo gene delivery, Bubble liposomes could effectively deliver plasmid DNA into mouse femoral artery. This method was more effectively than conventional lipofection. We conclude that Bubble liposomes are unique and efficient gene delivery tools in vitro and in vivo.

  7. Extracellular vesicles as a platform for membrane-associated therapeutic protein delivery.

    Science.gov (United States)

    Yang, Yoosoo; Hong, Yeonsun; Cho, Eunji; Kim, Gi Beom; Kim, In-San

    2018-01-01

    Membrane proteins are of great research interest, particularly because they are rich in targets for therapeutic application. The suitability of various membrane proteins as targets for therapeutic formulations, such as drugs or antibodies, has been studied in preclinical and clinical studies. For therapeutic application, however, a protein must be expressed and purified in as close to its native conformation as possible. This has proven difficult for membrane proteins, as their native conformation requires the association with an appropriate cellular membrane. One solution to this problem is to use extracellular vesicles as a display platform. Exosomes and microvesicles are membranous extracellular vesicles that are released from most cells. Their membranes may provide a favourable microenvironment for membrane proteins to take on their proper conformation, activity, and membrane distribution; moreover, membrane proteins can cluster into microdomains on the surface of extracellular vesicles following their biogenesis. In this review, we survey the state-of-the-art of extracellular vesicle (exosome and small-sized microvesicle)-based therapeutics, evaluate the current biological understanding of these formulations, and forecast the technical advances that will be needed to continue driving the development of membrane protein therapeutics.

  8. Method for Targeted Therapeutic Delivery of Proteins into Cells | NCI Technology Transfer Center | TTC

    Science.gov (United States)

    The Protein Expression Laboratory at the National Cancer Institute in Frederick, MD is seeking statements of capability or interest from parties interested in collaborative research to further develop a platform technology for the targeted intra-cellular delivery of proteins using virus-like particles (VLPs).

  9. Lipid-polymer hybrid nanoparticles as a new generation therapeutic delivery platform: a review.

    Science.gov (United States)

    Hadinoto, Kunn; Sundaresan, Ajitha; Cheow, Wean Sin

    2013-11-01

    Lipid-polymer hybrid nanoparticles (LPNs) are core-shell nanoparticle structures comprising polymer cores and lipid/lipid-PEG shells, which exhibit complementary characteristics of both polymeric nanoparticles and liposomes, particularly in terms of their physical stability and biocompatibility. Significantly, the LPNs have recently been demonstrated to exhibit superior in vivo cellular delivery efficacy compared to that obtained from polymeric nanoparticles and liposomes. Since their inception, the LPNs have advanced significantly in terms of their preparation strategy and scope of applications. Their preparation strategy has undergone a shift from the conceptually simple two-step method, involving preformed polymeric nanoparticles and lipid vesicles, to the more principally complex, yet easier to perform, one-step method, relying on simultaneous self-assembly of the lipid and polymer, which has resulted in better products and higher production throughput. The scope of LPNs' applications has also been extended beyond single drug delivery for anticancer therapy, to include combinatorial and active targeted drug deliveries, and deliveries of genetic materials, vaccines, and diagnostic imaging agents. This review details the current state of development for the LPNs preparation and applications from which we identify future research works needed to bring the LPNs closer to its clinical realization. Copyright © 2013 Elsevier B.V. All rights reserved.

  10. Cerebellomedullary Cistern Delivery for AAV-Based Gene Therapy: A Technical Note for Nonhuman Primates

    OpenAIRE

    Samaranch, Lluis; Bringas, John; Pivirotto, Philip; Sebastian, Waldy San; Forsayeth, John; Bankiewicz, Krystof

    2015-01-01

    Accessing cerebrospinal fluid (CSF) from the craniocervical junction through the posterior atlanto-occipital membrane via cerebellomedullary injection (also known as cisternal puncture or cisterna magna injection) has become a standard procedure in preclinical studies. Such delivery provides broader coverage to the central and peripheral nervous system unlike local parenchymal delivery alone. As a clinical application, this approach offers a more reliable method for neurological gene replacem...

  11. Safety and Efficacy of AAV Retrograde Pancreatic Ductal Gene Delivery in Normal and Pancreatic Cancer Mice.

    Science.gov (United States)

    Quirin, Kayla A; Kwon, Jason J; Alioufi, Arafat; Factora, Tricia; Temm, Constance J; Jacobsen, Max; Sandusky, George E; Shontz, Kim; Chicoine, Louis G; Clark, K Reed; Mendell, Joshua T; Korc, Murray; Kota, Janaiah

    2018-03-16

    Recombinant adeno-associated virus (rAAV)-mediated gene delivery shows promise to transduce the pancreas, but safety/efficacy in a neoplastic context is not well established. To identify an ideal AAV serotype, route, and vector dose and assess safety, we have investigated the use of three AAV serotypes (6, 8, and 9) expressing GFP in a self-complementary (sc) AAV vector under an EF1α promoter (scAAV.GFP) following systemic or retrograde pancreatic intraductal delivery. Systemic delivery of scAAV9.GFP transduced the pancreas with high efficiency, but gene expression did not exceed >45% with the highest dose, 5 × 10 12 viral genomes (vg). Intraductal delivery of 1 × 10 11 vg scAAV6.GFP transduced acini, ductal cells, and islet cells with >50%, ∼48%, and >80% efficiency, respectively, and >80% pancreatic transduction was achieved with 5 × 10 11 vg. In a Kras G12D -driven pancreatic cancer mouse model, intraductal delivery of scAAV6.GFP targeted acini, epithelial, and stromal cells and exhibited persistent gene expression 5 months post-delivery. In normal mice, intraductal delivery induced a transient increase in serum amylase/lipase that resolved within a day of infusion with no sustained pancreatic inflammation or fibrosis. Similarly, in PDAC mice, intraductal delivery did not increase pancreatic intraepithelial neoplasia progression/fibrosis. Our study demonstrates that scAAV6 targets the pancreas/neoplasm efficiently and safely via retrograde pancreatic intraductal delivery.

  12. Safety and Efficacy of AAV Retrograde Pancreatic Ductal Gene Delivery in Normal and Pancreatic Cancer Mice

    Directory of Open Access Journals (Sweden)

    Kayla A. Quirin

    2018-03-01

    Full Text Available Recombinant adeno-associated virus (rAAV-mediated gene delivery shows promise to transduce the pancreas, but safety/efficacy in a neoplastic context is not well established. To identify an ideal AAV serotype, route, and vector dose and assess safety, we have investigated the use of three AAV serotypes (6, 8, and 9 expressing GFP in a self-complementary (sc AAV vector under an EF1α promoter (scAAV.GFP following systemic or retrograde pancreatic intraductal delivery. Systemic delivery of scAAV9.GFP transduced the pancreas with high efficiency, but gene expression did not exceed >45% with the highest dose, 5 × 1012 viral genomes (vg. Intraductal delivery of 1 × 1011 vg scAAV6.GFP transduced acini, ductal cells, and islet cells with >50%, ∼48%, and >80% efficiency, respectively, and >80% pancreatic transduction was achieved with 5 × 1011 vg. In a KrasG12D-driven pancreatic cancer mouse model, intraductal delivery of scAAV6.GFP targeted acini, epithelial, and stromal cells and exhibited persistent gene expression 5 months post-delivery. In normal mice, intraductal delivery induced a transient increase in serum amylase/lipase that resolved within a day of infusion with no sustained pancreatic inflammation or fibrosis. Similarly, in PDAC mice, intraductal delivery did not increase pancreatic intraepithelial neoplasia progression/fibrosis. Our study demonstrates that scAAV6 targets the pancreas/neoplasm efficiently and safely via retrograde pancreatic intraductal delivery.

  13. DNA Nanotechnology for Precise Control over Drug Delivery and Gene Therapy.

    Science.gov (United States)

    Angell, Chava; Xie, Sibai; Zhang, Liangfang; Chen, Yi

    2016-03-02

    Nanomedicine has been growing exponentially due to its enhanced drug targeting and reduced drug toxicity. It uses the interactions where nanotechnological components and biological systems communicate with each other to facilitate the delivery performance. At this scale, the physiochemical properties of delivery systems strongly affect their capacities. Among current delivery systems, DNA nanotechnology shows many advantages because of its unprecedented engineering abilities. Through molecular recognition, DNA nanotechnology can be used to construct a variety of nanostructures with precisely controllable size, shape, and surface chemistry, which can be appreciated in the delivery process. In this review, different approaches that are currently used for the construction of DNA nanostructures are reported. Further, the utilization of these DNA nanostructures with the well-defined parameters for the precise control in drug delivery and gene therapy is discussed. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Ligand-Modified Human Serum Albumin Nanoparticles for Enhanced Gene Delivery.

    Science.gov (United States)

    Look, Jennifer; Wilhelm, Nadine; von Briesen, Hagen; Noske, Nadja; Günther, Christine; Langer, Klaus; Gorjup, Erwin

    2015-09-08

    The development of nonviral gene delivery systems is a great challenge to enable safe gene therapy. In this study, ligand-modified nanoparticles based on human serum albumin (HSA) were developed and optimized for an efficient gene therapy. Different glutaraldehyde cross-linking degrees were investigated to optimize the HSA nanoparticles for gene delivery. The peptide sequence arginine-glycine-aspartate (RGD) and the HIV-1 transactivator of transduction sequence (Tat) are well-known as promising targeting ligands. Plasmid DNA loaded HSA nanoparticles were covalently modified on their surface with these different ligands. The transfection potential of the obtained plasmid DNA loaded RGD- and Tat-modified nanoparticles was investigated in vitro, and optimal incubation conditions for these preparations were studied. It turned out that Tat-modified HSA nanoparticles with the lowest cross-linking degree of 20% showed the highest transfection potential. Taken together, ligand-functionalized HSA nanoparticles represent promising tools for efficient and safe gene therapy.

  15. Solidified self-nanoemulsifying formulation for oral delivery of combinatorial therapeutic regimen

    DEFF Research Database (Denmark)

    Jain, Amit K; Thanki, Kaushik; Jain, Sanyog

    2014-01-01

    formulation in contrast to that of clinically available tamoxifen citrate when measured as function of hepatotoxicity markers and histopathological changes. CONCLUSIONS: In nutshell, co-encapsulation of quercetin with tamoxifen in solid SNEDDS poses great potential in improving the therapeutic efficacy...

  16. 78 FR 77471 - Prospective Grant of Exclusive License for: Convection Enhanced Delivery of a Therapeutic Agent...

    Science.gov (United States)

    2013-12-23

    ...; nationalized), U.S. Patent Application 7,371,225, European Patent Application 03756863.1, Australian Patent 2003299140, to Medicenna Therapeutics, Inc. having a principle place of business in 1075 West Georgia St... date of this published notice, NIH receives written evidence and argument that establishes that the...

  17. RGD-based strategies for selective delivery of therapeutics and imaging agents to the tumour vasculature

    NARCIS (Netherlands)

    Temming, K; Molema, G; Kok, RJ

    2005-01-01

    During the past decade, RGD-peptides have become a popular tool for the targeting of drugs and imaging agents to a(v)beta(3)-integrin expressing tumour vasculature. RGD-peptides have been introduced by recombinant means into therapeutic proteins and viruses. Chemical means have been applied to

  18. A sight on protein-based nanoparticles as drug/gene delivery systems.

    Science.gov (United States)

    Salatin, Sara; Jelvehgari, Mitra; Maleki-Dizaj, Solmaz; Adibkia, Khosro

    2015-01-01

    Polymeric nanomaterials have extensively been applied for the preparation of targeted and controlled release drug/gene delivery systems. However, problems involved in the formulation of synthetic polymers such as using of the toxic solvents and surfactants have limited their desirable applications. In this regard, natural biomolecules including proteins and polysaccharide are suitable alternatives due to their safety. According to literature, protein-based nanoparticles possess many advantages for drug and gene delivery such as biocompatibility, biodegradability and ability to functionalize with targeting ligands. This review provides a general sight on the application of biodegradable protein-based nanoparticles in drug/gene delivery based on their origins. Their unique physicochemical properties that help them to be formulated as pharmaceutical carriers are also discussed.

  19. Therapeutic gene editing in CD34+ hematopoietic progenitors from Fanconi anemia patients.

    Science.gov (United States)

    Diez, Begoña; Genovese, Pietro; Roman-Rodriguez, Francisco J; Alvarez, Lara; Schiroli, Giulia; Ugalde, Laura; Rodriguez-Perales, Sandra; Sevilla, Julian; Diaz de Heredia, Cristina; Holmes, Michael C; Lombardo, Angelo; Naldini, Luigi; Bueren, Juan Antonio; Rio, Paula

    2017-11-01

    Gene targeting constitutes a new step in the development of gene therapy for inherited diseases. Although previous studies have shown the feasibility of editing fibroblasts from Fanconi anemia (FA) patients, here we aimed at conducting therapeutic gene editing in clinically relevant cells, such as hematopoietic stem cells (HSCs). In our first experiments, we showed that zinc finger nuclease (ZFN)-mediated insertion of a non-therapeutic EGFP-reporter donor in the AAVS1 "safe harbor" locus of FA-A lymphoblastic cell lines (LCLs), indicating that FANCA is not essential for the editing of human cells. When the same approach was conducted with therapeutic FANCA donors, an efficient phenotypic correction of FA-A LCLs was obtained. Using primary cord blood CD34 + cells from healthy donors, gene targeting was confirmed not only in in vitro cultured cells, but also in hematopoietic precursors responsible for the repopulation of primary and secondary immunodeficient mice. Moreover, when similar experiments were conducted with mobilized peripheral blood CD34 + cells from FA-A patients, we could demonstrate for the first time that gene targeting in primary hematopoietic precursors from FA patients is feasible and compatible with the phenotypic correction of these clinically relevant cells. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

  20. Potent spinal parenchymal AAV9-mediated gene delivery by subpial injection in adult rats and pigs

    Directory of Open Access Journals (Sweden)

    Atsushi Miyanohara

    2016-01-01

    Full Text Available Effective in vivo use of adeno-associated virus (AAV-based vectors to achieve gene-specific silencing or upregulation in the central nervous system has been limited by the inability to provide more than limited deep parenchymal expression in adult animals using delivery routes with the most clinical relevance (intravenous or intrathecal. Here, we demonstrate that the spinal pia membrane represents the primary barrier limiting effective AAV9 penetration into the spinal parenchyma after intrathecal AAV9 delivery. We develop a novel subpial AAV9 delivery technique and AAV9-dextran formulation. We use these in adult rats and pigs to show (i potent spinal parenchymal transgene expression in white and gray matter including neurons, glial and endothelial cells after single bolus subpial AAV9 delivery; (ii delivery to almost all apparent descending motor axons throughout the length of the spinal cord after cervical or thoracic subpial AAV9 injection; (iii potent retrograde transgene expression in brain motor centers (motor cortex and brain stem; and (iv the relative safety of this approach by defining normal neurological function for up to 6 months after AAV9 delivery. Thus, subpial delivery of AAV9 enables gene-based therapies with a wide range of potential experimental and clinical utilizations in adult animals and human patients.

  1. Coordination conjugates of therapeutic proteins with drug carriers: A new approach for versatile advanced drug delivery

    Czech Academy of Sciences Publication Activity Database

    Kejík, Z.; Bříza, T.; Králová, Jarmila; Poučková, P.; Kral, A.; Martásek, P.; Král, V.

    2011-01-01

    Roč. 21, č. 18 (2011), s. 5514-5520 ISSN 0960-894X R&D Projects: GA ČR GA203/09/1311 Grant - others:MPO(CZ) FR-TI3/521; AV ČR(CZ) KAN200100801 Program:FR; KA Institutional research plan: CEZ:AV0Z50520514 Keywords : combined cancer therapy * photodynamic therapy * targeted drug delivery Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.554, year: 2011

  2. 3D Printed Microtransporters: Compound Micromachines for Spatiotemporally Controlled Delivery of Therapeutic Agents.

    Science.gov (United States)

    Huang, Tian-Yun; Sakar, Mahmut Selman; Mao, Angelo; Petruska, Andrew J; Qiu, Famin; Chen, Xue-Bo; Kennedy, Stephen; Mooney, David; Nelson, Bradley J

    2015-11-01

    Functional compound micromachines are fabricated by a design methodology using 3D direct laser writing and selective physical vapor deposition of magnetic materials. Microtransporters with a wirelessly controlled Archimedes screw pumping mechanism are engineered. Spatiotemporally controlled collection, transport, and delivery of micro particles, as well as magnetic nanohelices inside microfluidic channels are demonstrated. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Convection-enhanced drug delivery to the brain: therapeutic potential and neuropathological considerations.

    Science.gov (United States)

    Barua, Neil U; Gill, Steven S; Love, Seth

    2014-03-01

    Convection-enhanced delivery (CED) describes a direct method of drug delivery to the brain through intraparenchymal microcatheters. By establishing a pressure gradient at the tip of the infusion catheter in order to exploit bulk flow through the interstitial spaces of the brain, CED offers a number of advantages over conventional drug delivery methods-bypass of the blood-brain barrier, targeted distribution through large brain volumes and minimization of systemic side effects. Despite showing early promise, CED is yet to fulfill its potential as a mainstream strategy for the treatment of neurological disease. Substantial research effort has been dedicated to optimize the technology for CED and identify the parameters, which govern successful drug distribution. It seems likely that successful clinical translation of CED will depend on suitable catheter technology being used in combination with drugs with optimal physicochemical characteristics, and on neuropathological analysis in appropriate preclinical models. In this review, we consider the factors most likely to influence the success or failure of CED, and review its application to the treatment of high-grade glioma, Parkinson's disease (PD) and Alzheimer's disease (AD). © 2013 International Society of Neuropathology.

  4. Minimally invasive and targeted therapeutic cell delivery to the skin using microneedle devices.

    Science.gov (United States)

    Gualeni, B; Coulman, S A; Shah, D; Eng, P F; Ashraf, H; Vescovo, P; Blayney, G J; Piveteau, L-D; Guy, O J; Birchall, J C

    2018-03-01

    Translation of cell therapies to the clinic is accompanied by numerous challenges, including controlled and targeted delivery of the cells to their site of action, without compromising cell viability and functionality. To explore the use of hollow microneedle devices (to date only used for the delivery of drugs and vaccines into the skin and for the extraction of biological fluids) to deliver cells into skin in a minimally invasive, user-friendly and targeted fashion. Melanocyte, keratinocyte and mixed epidermal cell suspensions were passed through various types of microneedles and subsequently delivered into the skin. Cell viability and functionality are maintained after injection through hollow microneedles with a bore size ≥ 75 μm. Healthy cells are delivered into the skin at clinically relevant depths. Hollow microneedles provide an innovative and minimally invasive method for delivering functional cells into the skin. Microneedle cell delivery represents a potential new treatment option for cell therapy approaches including skin repigmentation, wound repair, scar and burn remodelling, immune therapies and cancer vaccines. © 2017 British Association of Dermatologists.

  5. Lumbar spine intervertebral disc gene delivery: a pilot study in lewis rats.

    Science.gov (United States)

    Damle, Sheela R; Rawlins, Bernard A; Boachie-Adjei, Oheneba; Crystal, Ronald G; Hidaka, Chisa; Cunningham, Matthew E

    2013-02-01

    Basic research toward understanding and treating disc pathology in the spine has utilized numerous animal models, with delivery of small molecules, purified factors, and genes of interest. To date, gene delivery to the rat lumbar spine has only been described utilizing genetically programmed cells in a matrix which has required partial disc excision, and expected limitation of treatment diffusion into the disc. This study was designed to develop and describe a surgical technique for lumbar spine exposure and disc space preparation, and use of a matrix-free method for gene delivery. Naïve or genetically programmed isogeneic bone marrow stromal cells were surgically delivered to adolescent male Lewis rat lumbar discs, and utilizing quantitative biochemical and qualitative immunohistological assessments, the implanted cells were detected 3 days post-procedure. Statistically significant differences were noted for recovery of the β-galactosidase marker gene comparing delivery of naïve or labeled cells (10(5) cells per disc) from the site of implantation, and between delivery of 10(5) or 10(6) labeled cells per disc at the site of implantation and the adjacent vertebral body. Immunohistology confirmed that the β-galactosidase marker was detected in the adjacent vertebra bone in the zone of surgical implantation. The model requires further testing in larger cohorts and with biologically active genes of interest, but the observations from the pilot experiments are very encouraging that this will be a useful comparative model for basic spine research involving gene or cell delivery, or other locally delivered therapies to the intervertebral disc or adjacent vertebral bodies in rats.

  6. Artificial Virus as Trump-card to Resolve Exigencies in Targeted Gene Delivery.

    Science.gov (United States)

    Ajithkumar, K C; Pramod, Kannissery

    2018-01-01

    Viruses are potent pathogens that can effectively deliver the genetic material to susceptible host cells. This capability is beneficially utilized to successfully deliver the genetic material. However, the use of virus mediated gene delivery is considered divisive, because the potentially replicable genomes recombine or integrate with the cell DNA resulting in immunogenicity, ranging from inflammation to death. Thus, the need for potentially effective non-viral gene delivery vehicles arises. Non-viral vectors, protein only particles and virus like particles (VLP) can be constructed which contain all the necessary functional moieties. These resemble viruses and are called artificial or synthetic virus. The artificial virus eliminates the disadvantages of viral vectors but retain the beneficial effects of the viruses. Need for further functionalization can be avoided by this approach because incorporation of requisite agents such as cell ligands, membrane active peptides, etc. into proteins is possible. The protein- DNA complexes resemble bacterial inclusion bodies. Nucleic acids influence conformation of protein units which subsequently result in cell uptake and finally to the cell nucleus. Such tunable systems mimic the activities of infected viruses and are used for the safe and effective delivery of drugs and genetic material in gene therapy. The versatility, stability and biocompatible nature of artificial virus along with high transfection efficacy have made it favorite for gene delivery purposes, in addition to being useful for various biomedical and drug delivery applications. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  7. Gene therapy for carcinoma of the breast: Therapeutic genetic correction strategies

    International Nuclear Information System (INIS)

    Obermiller, Patrice S; Tait, David L; Holt, Jeffrey T

    2000-01-01

    Gene therapy is a therapeutic approach that is designed to correct specific molecular defects that contribute to the cause or progression of cancer. Genes that are mutated or deleted in cancers include the cancer susceptibility genes p53 and BRCA1. Because mutational inactivation of gene function is specific to tumor cells in these settings, cancer gene correction strategies may provide an opportunity for selective targeting without significant toxicity for normal nontumor cells. Both p53 and BRCA1 appear to inhibit cancer cells that lack mutations in these genes, suggesting that the so-called gene correction strategies may have broader potential than initially believed. Increasing knowledge of cancer genetics has identified these and other genes as potential targets for gene replacement therapy. Initial patient trials of p53 and BRCA1 gene therapy have provided some indications of potential efficacy, but have also identified areas of basic and clinical research that are needed before these approaches may be widely used in patient care

  8. DNA-transporting nanoparticles : design and in vitro evaluation of DNA and formulation for non-viral gene delivery

    NARCIS (Netherlands)

    van Gaal, E.V.B.

    2010-01-01

    The aim of gene therapy is to treat, cure or prevent a disease by replacing defective genes, introducing new genes or changing the expression of a person’s genes. Success of gene therapy is dependent on successful delivery of DNA from the site of administration into cell nuclei. Naturally occurring

  9. Development of Gold Nanoparticle towards Radioenhancement Therapy, Renal Clearance, siRNA Delivery and Light-Controlled Gene Silencing

    Science.gov (United States)

    Wang, Jianxin

    Gold nanoparticles (GNPs) have been widely studied and used in research for diagnostic, prophylactic or therapeutic purposes. However, they still face many technical challenges before they can be used to effectively address unmet biomedical needs. The theme of this dissertation is focused on addressing challenges of GNPs in clinical translation, and to improve their potential for application in radioenhancement therapy and siRNA delivery. We demonstrate the facile self-assembly of micellar gold nanocapsules using zwitterionic surfactants, with hydrodynamic diameters below 10 nm, which holds promise for good renal clearance to promote the excretion of GNPs in human body. We also prepared PEI- and PEG-coated GNPs and demonstrated their uptake into HeLa cells with exposure to soft X-rays (120 kVp), based on the consideration that the proximity of GNPs to nuclear DNA may be beneficial for enhancing low-energy ionizing radiotherapy. GNP-mediated siRNA delivery may be challenged by nonspecific siRNA desorption during circulation, which can cause off-target effects and immunogenicity. The use of gold nanorods (GNRs) for siRNA delivery also faces challenges like reduced dispersion stability during siRNA functionalization. We developed an effective way to load siRNA onto GNRs at high density, using oleylsulfobetaine (OSB) as an intermediate surfactant and dithiocarbamates (DTCs) as desorption-resistant anchors for siRNA. The GNR?siRNA complexes provided excellent control for laser-triggered gene silencing.

  10. Electric pulse-mediated gene delivery to various animal tissues

    DEFF Research Database (Denmark)

    Mir, Lluis M; Moller, Pernille H; André, Franck

    2005-01-01

    therapy, termed electrogenetherapy (EGT as well). By transfecting cells with a long lifetime, such as muscle fibers, a very long-term expression of genes can be obtained. A great variety of tissues have been transfected successfully, from muscle as the most extensively used, to both soft (e.g., spleen...

  11. Magnetotactic Bacterial Cages as Safe and Smart Gene Delivery Vehicles

    KAUST Repository

    Alsaiari, Shahad K.

    2016-07-27

    In spite of the huge advances in the area of synthetic carriers, their efficiency still poorly compares to natural vectors. Herein, we report the use of unmodified magnetotactic bacteria as a guidable delivery vehicle for DNA functionalized gold nanoparticles (AuNPs). High cargo loading is established under anaerobic conditions (bacteria is alive) through endocytosis where AuNPs are employed as transmembrane proteins mimics (facilitate endocytosis) as well as imaging agents to verify and quantify loading and release. The naturally bio-mineralized magnetosomes, within the bacteria, induce heat generation inside bacteria through magnetic hyperthermia. Most importantly after exposing the system to air (bacteria is dead) the cell wall stays intact providing an efficient bacterial vessel. Upon incubation with THP-1 cells, the magnetotactic bacterial cages (MBCs) adhere to the cell wall and are directly engulfed through the phagocytic activity of these cells. Applying magnetic hyperthermia leads to the dissociation of the bacterial microcarrier and eventual release of cargo.

  12. Magnetotactic Bacterial Cages as Safe and Smart Gene Delivery Vehicles

    KAUST Repository

    Alsaiari, Shahad K.; Ezzedine, Alaa H.; Abdallah, Abdallah; Sougrat, Rachid; Khashab, Niveen M.

    2016-01-01

    In spite of the huge advances in the area of synthetic carriers, their efficiency still poorly compares to natural vectors. Herein, we report the use of unmodified magnetotactic bacteria as a guidable delivery vehicle for DNA functionalized gold nanoparticles (AuNPs). High cargo loading is established under anaerobic conditions (bacteria is alive) through endocytosis where AuNPs are employed as transmembrane proteins mimics (facilitate endocytosis) as well as imaging agents to verify and quantify loading and release. The naturally bio-mineralized magnetosomes, within the bacteria, induce heat generation inside bacteria through magnetic hyperthermia. Most importantly after exposing the system to air (bacteria is dead) the cell wall stays intact providing an efficient bacterial vessel. Upon incubation with THP-1 cells, the magnetotactic bacterial cages (MBCs) adhere to the cell wall and are directly engulfed through the phagocytic activity of these cells. Applying magnetic hyperthermia leads to the dissociation of the bacterial microcarrier and eventual release of cargo.

  13. Lignin nanotubes as vehicles for gene delivery into human cells.

    Science.gov (United States)

    Ten, Elena; Ling, Chen; Wang, Yuan; Srivastava, Arun; Dempere, Luisa Amelia; Vermerris, Wilfred

    2014-01-13

    Lignin nanotubes (LNTs) synthesized from the aromatic plant cell wall polymer lignin in a sacrificial alumina membrane template have as useful features their flexibility, ease of functionalization due to the availability of many functional groups, label-free detection by autofluorescence, and customizable optical properties. In this report we show that the physicochemical properties of LNTs can be varied over a wide range to match requirements for specific applications by using lignin with different subunit composition, a function of plant species and genotype, and by choosing the lignin isolation method (thioglycolic acid, phosphoric acid, sulfuric acid (Klason), sodium hydroxide lignin), which influences the size and reactivity of the lignin fragments. Cytotoxicity studies with human HeLa cells showed that concentrations of up to 90 mg/mL are tolerated, which is a 10-fold higher concentration than observed for single- or multiwalled carbon nanotubes (CNTs). Confocal microscopy imaging revealed that all LNT formulations enter HeLa cells without auxiliary agents and that LNTs made from NaOH-lignin penetrate the cell nucleus. We further show that DNA can adsorb to LNTs. Consequently, exposure of HeLa cells to LNTs coated with DNA encoding the green fluorescent protein (GFP) leads to transfection and expression of GFP. The highest transfection efficiency was obtained with LNTs made from NaOH-lignin due to a combination of high DNA binding capacity and DNA delivery directly into the nucleus. These combined features of LNTs make LNTs attractive as smart delivery vehicles of DNA without the cytotoxicity associated with CNTs or the immunogenicity of viral vectors.

  14. Murine bone marrow-derived mesenchymal stem cells as vehicles for interleukin-12 gene delivery into Ewing sarcoma tumors.

    Science.gov (United States)

    Duan, Xiaoping; Guan, Hui; Cao, Ying; Kleinerman, Eugenie S

    2009-01-01

    This study evaluated the therapeutic efficacy of interleukin 12 (IL-12) gene therapy in Ewing sarcoma and whether murine mesenchymal stem cells (MSCs) could serve as vehicles for IL-12 gene delivery. MSCs were isolated from murine bone marrow cells. Cells were phenotyped using flow cytometry. Cultured MSCs differentiated into osteocytes and adipocytes using the appropriate media. Freshly isolated MSCs were transfected with adenoviral vectors containing either the beta-galactosidase (Ad:beta-gal) or the IL-12 (Ad:IL-12) gene. Expression of IL-12 was confirmed using reverse transcription polymerase chain reaction. Mice with TC71 Ewing sarcoma tumors were then treated intravenously with MSCs transfected with Ad:beta-gal or Ad:IL-12. Tumors were measured and analyzed by immunohistochemical analysis for expression of IL-12 protein. Expression of both p35 and p40 IL-12 subunits was demonstrated in MSCs transfected in vitro with Ad:IL-12. IL-12 expression was seen in tumors from mice treated with MSCs transfected with Ad:IL-12. Tumor growth was also significantly inhibited compared with that in mice treated with MSCs transfected with Ad:beta-gal. MSCs can be transfected with the IL-12 gene. These transfected cells localize to tumors after intravenous injection and induce local IL-12 protein production and the regression of established tumors. Copyright (c) 2008 American Cancer Society.

  15. A Promising Combo Gene Delivery System Developed from (3-Aminopropyl)triethoxysilane-Modified Iron Oxide Nanoparticles and Cationic Polymers

    Science.gov (United States)

    Zhang, Zubin; Song, Lina; Dong, Jinlai; Guo, Dawei; Du, Xiaolin; Cao, Biyin; Zhang, Yu; Gu, Ning; Mao, Xinliang

    2013-05-01

    (3-Aminopropyl)triethoxysilane-modified iron oxide nanoparticles (APTES-IONPs) have been evaluated for various biomedical applications, including medical imaging and drug delivery. Cationic polymers (CPs) such as Lipofectamine and TurboFect are widely used for research in gene delivery, but their toxicity and low in vivo efficiency limited their further application. In the present study, we synthesized water-soluble APTES-IONPs and developed a combo gene delivery system based on APTES-IONPs and CPs. This system significantly increased gene-binding capacity, protected genes from degradation, and improved gene transfection efficiency for DNA and siRNA in both adherent and suspension cells. Because of its great biocompatibility, high gene-carrying ability, and very low cytotoxicity, this combo gene delivery system will be expected for a wide application, and it might provide a new method for gene therapy.

  16. A Promising Combo Gene Delivery System Developed from (3-Aminopropyl)triethoxysilane-Modified Iron Oxide Nanoparticles and Cationic Polymers

    International Nuclear Information System (INIS)

    Zhang Zubin; Song Lina; Dong Jinlai; Guo Dawei; Du Xiaolin; Cao Biyin; Zhang Yu; Gu Ning; Mao Xinliang

    2013-01-01

    (3-Aminopropyl)triethoxysilane-modified iron oxide nanoparticles (APTES-IONPs) have been evaluated for various biomedical applications, including medical imaging and drug delivery. Cationic polymers (CPs) such as Lipofectamine and TurboFect are widely used for research in gene delivery, but their toxicity and low in vivo efficiency limited their further application. In the present study, we synthesized water-soluble APTES-IONPs and developed a combo gene delivery system based on APTES-IONPs and CPs. This system significantly increased gene-binding capacity, protected genes from degradation, and improved gene transfection efficiency for DNA and siRNA in both adherent and suspension cells. Because of its great biocompatibility, high gene-carrying ability, and very low cytotoxicity, this combo gene delivery system will be expected for a wide application, and it might provide a new method for gene therapy.

  17. Myocardial gene delivery using molecular cardiac surgery with recombinant adeno-associated virus vectors in vivo

    Science.gov (United States)

    White, JD; Thesier, DM; Swain, JBD; Katz, MG; Tomasulo, C; Henderson, A; Wang, L; Yarnall, C; Fargnoli, A; Sumaroka, M; Isidro, A; Petrov, M; Holt, D; Nolen-Walston, R; Koch, WJ; Stedman, HH; Rabinowitz, J; Bridges, CR

    2013-01-01

    We use a novel technique that allows for closed recirculation of vector genomes in the cardiac circulation using cardiopulmonary bypass, referred to here as molecular cardiac surgery with recirculating delivery (MCARD). We demonstrate that this platform technology is highly efficient in isolating the heart from the systemic circulation in vivo. Using MCARD, we compare the relative efficacy of single-stranded (ss) adeno-associated virus (AAV)6, ssAAV9 and self-complimentary (sc)AAV6-encoding enhanced green fluorescent protein, driven by the constitutive cytomegalovirus promoter to transduce the ovine myocardium in situ. MCARD allows for the unprecedented delivery of up to 48 green fluorescent protein genome copies per cell globally in the sheep left ventricular (LV) myocardium. We demonstrate that scAAV6-mediated MCARD delivery results in global, cardiac-specific LV gene expression in the ovine heart and provides for considerably more robust and cardiac-specific gene delivery than other available delivery techniques such as intramuscular injection or intracoronary injection; thus, representing a potential, clinically translatable platform for heart failure gene therapy. PMID:21228882

  18. Chitosan nanoparticles as non-viral gene delivery systems: determination of loading efficiency.

    Science.gov (United States)

    Carrillo, Carolina; Suñé, Josep Maria; Pérez-Lozano, Pilar; García-Montoya, Encarna; Sarrate, Rocío; Fàbregas, Anna; Miñarro, Montserrat; Ticó, Josep Ramon

    2014-07-01

    Chitosan has been studied for use in particle delivery systems for therapeutic purposes, since one of its most important applications is as a non-viral vector in gene therapy. Due to its positive charge, it is capable of forming DNA complexes (polyplexes) obtained through several methods and with the property of protecting nucleic acids. Two methods for obtaining the nanoparticles of chitosan-nucleic acids are reported in this study: simple complexation (of depolymerized chitosan or of different chitosan salts with plasmid) and ionic gelation (by adsorption of plasmid in the nanoparticles or by encapsulation of plasmid into nanoparticles). The determination of the loading efficiency of chitosan nanoparticles with the plasmid is carried out by electrophoretic mobility of the samples on agarose gel. Furthermore, the nanoparticles have been characterized according to their morphology, size and surface charge using AFM, TEM, laser diffraction and dynamic light scattering techniques. The polyplexes obtained have been found to be spherical and nanometric in size (between 100-230nm) with a zeta potential between 37 and 48mV. Positive results have been obtained by agarose gel electrophoresis for all studied cases: a concentration of between 20 and 30μg/mL of chitosan salts is required while for the remaining chitosan samples studied, 100% loading efficiency does not occur until a concentration equal to 100μg/mL (regardless of previous depolymerisation and the method performed). Chitosan-plasmid nanocapsules have been obtained at the polymer concentrations worked with (between 0.025 and 0.2%). Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  19. Synthesis and evaluation of cationic nanomicelles for in vitro and in vivo gene delivery

    Science.gov (United States)

    Mandke, Rhishikesh Subhash

    The goal of proposed study was to contribute towards the development of a nano size, high efficiency and low toxicity non-viral polymeric vector for gene delivery in vitro and in vivo. A series of fatty acid grafted low-molecular-weight chitosan (N-acyl LMWCs) were synthesized, purified and characterized for their physicochemical properties using various analytical techniques such as infrared spectroscopy, elemental analysis and dynamic light scattering. The formulation parameters including pH, sonication duration, and filtration altered the physicochemical characteristics of N-acyl LMWC nanomicelles. The acyl chain length and degree of unsaturation in fatty acids also had an impact on the physicochemical properties and the transfection efficiency of nanomicelles. N-acyl LMWC nanomicelles showed efficient in vitro transfection as visualized and quantified using a reporter plasmid (encoding green fluorescent protein), and therapeutic plasmids (encoding for interleukin-4 and interleukin-10), respectively. The in vitro transfection efficiencies of N-acyl LMWCs with 18:1 and 18:2 grafts (oleic and linoleic acids) were comparable with FuGENERTM HD (marketed non-viral vector) but were ˜8-fold and 35-fold higher as compared to LMWC and naked DNA, respectively. The in vivo transfection efficiency of N-acyl LMWC to deliver plasmids individually encoding IL-4 and IL-10 as well as a bicistronic plasmid encoding both IL-4 and IL-10 was studied in a multiple, low-dose streptozotocin induced diabetic mouse model. The transfection efficiency of pDNA/N-acyl LMWC polyplexes injected via intramuscular route showed significant improvement (ptumor necrosis factor-alpha (TNF-alpha) levels. The pancreas of pDNA/N-acyl LMWC polyplex treated animals exhibited protection from streptozotocin-induced insulitis and the delivery systems were biocompatible. Histological studies revealed that there were no signs of chronic inflammation at the injection site. The bicistronic plasmid exhibited

  20. Probing suitable therapeutic nanoparticles for controlled drug delivery and diagnostic reproductive health biomarker development

    Energy Technology Data Exchange (ETDEWEB)

    Jha, Rakhi [School of Medical Science and Technology, Indian Institute of Technology Kharagpur, Kharagpur 721 302 (India); National Institute of Animal Welfare, Ministry of Environment, Forest and Climate Change, Faridabad, Haryana 121 004 (India); Jha, Pradeep K., E-mail: jha.rk.pk@gmail.com [School of Medical Science and Technology, Indian Institute of Technology Kharagpur, Kharagpur 721 302 (India); Gupta, Santosh; Bhuvaneshwaran, S.P. [School of Medical Science and Technology, Indian Institute of Technology Kharagpur, Kharagpur 721 302 (India); Hossain, Maidul [Department of Chemistry & Chemical Technology, Vidyasagar University, Midnapore 721102 (India); Guha, Sujoy K. [School of Medical Science and Technology, Indian Institute of Technology Kharagpur, Kharagpur 721 302 (India)

    2016-04-01

    Nanomaterial mediated drug delivery represents a highly promising technique while its selectivity for reproductive healthcare application still remains a challenge. Since the delicate structure and functional role of reproductive tissue and gametes require the use of biocompatible nanomedicine/devices that do not affect fertility or the development of resulting offspring, this paper reports an intercomparative study of human spermatozoa interaction with three different nanoparticles (NPs) namely; iron oxide (Fe{sub 3}O{sub 4)}, multiwalled carbon nanotubes (MWCNT) and graphene platelet nanopowder (GPN) to probe their suitability for drug delivery carrier and biomarker development purposes. ATR–FTIR results revealed that the sperm cell interaction with GPN had maximum amide I absorption for cell proteins and C=O stretching of the peptide backbone at the band around 1657 cm{sup −1} followed by iron oxide NPs whereas MWCNT had no absorption. These results showed that GPN followed by iron oxide NPs got maximally entrapped by cell membrane protein with maximum disruption but MWCNT exhibited less entrapment but significantly higher internalization which was further validated by morphological analysis of these cell NP interaction by SEM, HRTEM and fluorescence microscopy. The uptake kinetics and penetration mechanism of NPs were examined with isothermal titration calorimetry (ITC). Interestingly, ITC results confirmed ATR–FTIR and morphological observations that the binding of GPN and Fe{sub 3}O{sub 4} NPs with cell was exothermic and their bindings were favored by both negative enthalpy and positive entropy whereas in the case of MWCNT it was endothermic supported by unfavorable positive enthalpy and a favorable entropy change. Hence, it was evident that MWCNT had better internalization efficiency without disrupting the sperm lipid membrane compared to Fe{sub 3}O{sub 4} and GPN NPs. Therefore, this work proposes CNT as promising means. - Highlights: • Biophysical

  1. Monocyte Trafficking, Engraftment, and Delivery of Nanoparticles and an Exogenous Gene into the Acutely Inflamed Brain Tissue - Evaluations on Monocyte-Based Delivery System for the Central Nervous System.

    Directory of Open Access Journals (Sweden)

    Hsin-I Tong

    Full Text Available The ability of monocytes and monocyte-derived macrophages (MDM to travel towards chemotactic gradient, traverse tissue barriers, and accumulate precisely at diseased sites makes them attractive candidates as drug carriers and therapeutic gene delivery vehicles targeting the brain, where treatments are often hampered by the blockade of the blood brain barrier (BBB. This study was designed to fully establish an optimized cell-based delivery system using monocytes and MDM, by evaluating their homing efficiency, engraftment potential, as well as carriage and delivery ability to transport nano-scaled particles and exogenous genes into the brain, following the non-invasive intravenous (IV cell adoptive transfer in an acute neuroinflammation mouse model induced by intracranial injection of Escherichia coli lipopolysaccharides. We demonstrated that freshly isolated monocytes had superior inflamed-brain homing ability over MDM cultured in the presence of macrophage colony stimulating factor. In addition, brain trafficking of IV infused monocytes was positively correlated with the number of adoptive transferred cells, and could be further enhanced by transient disruption of the BBB with IV administration of Mannitol, Bradykinin or Serotonin right before cell infusion. A small portion of transmigrated cells was detected to differentiate into IBA-1 positive cells with microglia morphology in the brain. Finally, with the use of superparamagnetic iron oxide nanoparticles SHP30, the ability of nanoscale agent-carriage monocytes to enter the inflamed brain region was validated. In addition, lentiviral vector DHIV-101 was used to introduce green fluorescent protein (GFP gene into monocytes, and the exogenous GFP gene was detected in the brain at 48 hours following IV infusion of the transduced monocytes. All together, our study has set up the optimized conditions for the more-in-depth tests and development of monocyte-mediated delivery, and our data supported

  2. Ultrasound-Mediated Drug/Gene Delivery in Solid Tumor Treatment

    Directory of Open Access Journals (Sweden)

    Yufeng Zhou

    2013-01-01

    Full Text Available Ultrasound is an emerging modality for drug delivery in chemotherapy. This paper reviews this novel technology by first introducing the designs and characteristics of three classes of drug/gene vehicles, microbubble (including nanoemulsion, liposomes, and micelles. In comparison to conventional free drug, the targeted drug-release and delivery through vessel wall and interstitial space to cancerous cells can be activated and enhanced under certain sonication conditions. In the acoustic field, there are several reactions of these drug vehicles, including hyperthermia, bubble cavitation, sonoporation, and sonodynamics, whose physical properties are illustrated for better understanding of this approach. In vitro and in vivo results are summarized, and future directions are discussed. Altogether, ultrasound-mediated drug/gene delivery under imaging guidance provides a promising option in cancer treatment with enhanced agent release and site specificity and reduced toxicity.

  3. Efficient Gene Delivery to Pig Airway Epithelia and Submucosal Glands Using Helper-Dependent Adenoviral Vectors

    Directory of Open Access Journals (Sweden)

    Huibi Cao

    2013-01-01

    Full Text Available Airway gene delivery is a promising strategy to treat patients with life-threatening lung diseases such as cystic fibrosis (CF. However, this strategy has to be evaluated in large animal preclinical studies in order to translate it to human applications. Because of anatomic and physiological similarities between the human and pig lungs, we utilized pig as a large animal model to examine the safety and efficiency of airway gene delivery with helper-dependent adenoviral vectors. Helper-dependent vectors carrying human CFTR or reporter gene LacZ were aerosolized intratracheally into pigs under bronchoscopic guidance. We found that the LacZ reporter and hCFTR transgene products were efficiently expressed in lung airway epithelial cells. The transgene vectors with this delivery can also reach to submucosal glands. Moreover, the hCFTR transgene protein localized to the apical membrane of both ciliated and nonciliated epithelial cells, mirroring the location of wild-type CF transmembrane conductance regulator (CFTR. Aerosol delivery procedure was well tolerated by pigs without showing systemic toxicity based on the limited number of pigs tested. These results provide important insights into developing clinical strategies for human CF lung gene therapy.

  4. A fusion of minicircle DNA and nanoparticle delivery technologies facilitates therapeutic genetic engineering of autologous canine olfactory mucosal cells.

    Science.gov (United States)

    Delaney, Alexander M; Adams, Christopher F; Fernandes, Alinda R; Al-Shakli, Arwa F; Sen, Jon; Carwardine, Darren R; Granger, Nicolas; Chari, Divya M

    2017-06-29

    Olfactory ensheathing cells (OECs) promote axonal regeneration and improve locomotor function when transplanted into the injured spinal cord. A recent clinical trial demonstrated improved motor function in domestic dogs with spinal injury following autologous OEC transplantation. Their utility in canines offers promise for human translation, as dogs are comparable to humans in terms of clinical management and genetic/environmental variation. Moreover, the autologous, minimally invasive derivation of OECs makes them viable for human spinal injury investigation. Genetic engineering of transplant populations may augment their therapeutic potential, but relies heavily on viral methods which have several drawbacks for clinical translation. We present here the first proof that magnetic particles deployed with applied magnetic fields and advanced DNA minicircle vectors can safely bioengineer OECs to secrete a key neurotrophic factor, with an efficiency approaching that of viral vectors. We suggest that our alternative approach offers high translational potential for the delivery of augmented clinical cell therapies.

  5. Status of therapeutic gene transfer to treat canine dilated cardiomyopathy in dogs.

    Science.gov (United States)

    Sleeper, Meg M; Bish, Lawrence T; Sweeney, H Lee

    2010-07-01

    Therapeutic gene transfer holds promise as a way to treat dilated cardiomyopathy from any underlying cause because the approach attempts to address metabolic disturbances that occur at the molecular level of the failing heart. Calcium-handling abnormalities and increased rates of apoptosis are abnormalities that occur in many types of heart disease, and gene therapies that target these metabolic defects have proven to be beneficial in numerous rodent models of heart disease. The authors are currently evaluating this approach to treat canine idiopathic dilated cardiomyopathy.

  6. Gelatin nanoparticles enhance delivery of hepatitis C virus recombinant NS2 gene.

    Science.gov (United States)

    Sabet, Salwa; George, Marina A; El-Shorbagy, Haidan M; Bassiony, Heba; Farroh, Khaled Y; Youssef, Tareq; Salaheldin, Taher A

    2017-01-01

    Development of an effective non-viral vaccine against hepatitis C virus infection is of a great importance. Gelatin nanoparticles (Gel.NPs) have an attention and promising approach as a viable carrier for delivery of vaccine, gene, drug and other biomolecules in the body. The present study aimed to develop stable Gel.NPs conjugated with nonstructural protein 2 (NS2) gene of Hepatitis C Virus genotype 4a (HCV4a) as a safe and an efficient vaccine delivery system. Gel.NPs were synthesized and characterized (size: 150±2 nm and zeta potential +17.6 mv). NS2 gene was successfully cloned and expressed into E. coli M15 using pQE-30 vector. Antigenicity of the recombinant NS2 protein was confirmed by Western blotting to verify the efficiency of NS2 as a possible vaccine. Then NS2 gene was conjugated to gelatin nanoparticles and a successful conjugation was confirmed by labeling and imaging using Confocal Laser Scanning Microscope (CLSM). Interestingly, the transformation of the conjugated NS2/Gel.NPs complex into E. coli DH5-α was 50% more efficient than transformation with the gene alone. In addition, conjugated NS2/Gel.NPs with ratio 1:100 (w/w) showed higher transformation efficiency into E. coli DH5-α than the other ratios (1:50 and 2:50). Gel.NPs effectively enhanced the gene delivery in bacterial cells without affecting the structure of NS2 gene and could be used as a safe, easy, rapid, cost-effective and non-viral vaccine delivery system for HCV.

  7. Optimization of Acetalated Dextran-Based Nanocomposite Microparticles for Deep Lung Delivery of Therapeutics via Spray-Drying.

    Science.gov (United States)

    Wang, Zimeng; Meenach, Samantha A

    2017-12-01

    Nanocomposite microparticle (nCmP) systems exhibit promising potential in the application of therapeutics for pulmonary drug delivery. This work aimed at identifying the optimal spray-drying condition(s) to prepare nCmP with specific drug delivery properties including small aerodynamic diameter, effective nanoparticle (NP) redispersion upon nCmP exposure to an aqueous solution, high drug loading, and low water content. Acetalated dextran (Ac-Dex) was used to form NPs, curcumin was used as a model drug, and mannitol was the excipient in the nCmP formulation. Box-Behnken design was applied using Design-Expert software for nCmP parameter optimization. NP ratio (NP%) and feed concentration (Fc) are significant parameters that affect the aerodynamic diameters of nCmP systems. NP% is also a significant parameter that affects the drug loading. Fc is the only parameter that influenced the water content of the particles significantly. All nCmP systems could be completely redispersed into the parent NPs, indicating that none of the factors have an influence on this property within the design range. The optimal spray-drying condition to prepare nCmP with a small aerodynamic diameter, redispersion of the NPs, low water content, and high drug loading is 80% NP%, 0.5% Fc, and an inlet temperature lower than 130°C. Copyright © 2017 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

  8. Development and Evaluation of Naproxen Sodium Gel Using Piper cubeba for Enhanced Transdermal Drug Delivery and Therapeutic Facilitation.

    Science.gov (United States)

    Patwardhan, Sunetra; Patil, Manohar; Sockalingam, Anbazhagan

    2017-01-01

    The absorption of drug through skin avoids many side effects of oral route like gastric irritation, nausea, systemic toxicity etc and thus improves patient compliance. Naproxen sodium (NPRS) is one of the potent NSAID agents. The present study was aimed to develop and evaluate the gel formulation containing NPRS for transdermal drug delivery reducing the side effects and improving patient compliance. The patents on topical delivery of NSAIDS (US 9012402 B1, US 9072659 B2, US 20150258196 A1) and patents indicating use of herbal penetration enhancers (US 20100273746A1, WO 2005009510 A2, US 6004969 A) helped in selecting the drug, excipients. Current protocol employs various extracts of Piper cubeba fruit to evaluate its role in absorption of NPRS. Various batches containing 1% NPRS and varying concentrations of synthetic permeation enhancers or the extracts were formulated in carbopol gel. Gel was evaluated for parameters like organoleptic parameters, pH, viscosity and spreadability. An ex-vivo percutaneous absorption of NPRS from gel was investigated and compared with best performing synthetic enhancer, transcutol P (TP). The batch containing 2% n-hexane extract (NHE) of Piper cubeba showed higher permeation than TP and Chloroform (CE), Methanolic (ME) and aqueous (AE) extracts as well. It showed improved % cumulative release (85.09%) and flux (278.61μg/cm2.h), as compared to TP and other extracts. Histopathology indicated the formulation safer as compared to that with synthetic enhancer. It suggests P. cubeba as effective and safer tool for transdermal delivery and acts as therapeutic facilitator for naproxen. GC-MS analysis indicates lignans & terpenes in NHE to which this permeation enhancement activity may be attributed. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  9. Vascular nanomedicine: Site specific delivery of elastin stabilizing therapeutics to damaged arteries

    Science.gov (United States)

    Sinha, Aditi

    Elastin, a structural protein in the extra-cellular matrix, plays a critical role in the normal functioning of blood vessels. Apart from performing its primary function of providing resilience to arteries, it also plays major role in regulating cell-cell and cell-matrix interactions, response to injury, and morphogenesis. Medial arterial calcification (MAC) and abdominal aortic aneurysm (AAA) are two diseases where the structural and functional integrity of elastin is severely compromised. Although the clinical presentation of MAC and AAA differ, they have one common underlying causative mechanism---pathological degradation of elastin. Hence prevention of elastin degradation in the early stages of MAC and AAA can mitigate, partially if not wholly, the fatal consequences of both the diseases. The work presented here is motivated by the overwhelming statistics of people afflicted by elastin associated cardiovascular diseases and the unavailability of cure for the same. Overall goal of our research is to understand role of elastin degradation in cardiovascular diseases and to develop a targeted vascular drug delivery system that is minimally invasive, biodegradable, and non-toxic, that prevents elastin from degradation. Our hope is that such treatment will also help regenerate elastin, thereby providing a multi-fold treatment option for elasto-degenerative vascular diseases. For this purpose, we have first confirmed the combined role of degraded elastin and hyperglycemia in the pathogenesis of MAC. We have shown that in the absence of degraded elastin and TGF-beta1 (abundantly present in diabetic arteries) vascular smooth muscle cells maintain their homeostatic state, regardless of environmental glucose concentrations. However simultaneous exposure to glucose, elastin peptides and TGF-beta1 causes the pathological transgenesis of vascular cells to osteoblast-like cells. We show that plant derived polyphenols bind to vascular elastin with great affinity resulting in

  10. Formulation, functional evaluation and ex vivo performance of thermoresponsive soluble gels - A platform for therapeutic delivery to mucosal sinus tissue.

    Science.gov (United States)

    Pandey, Preeti; Cabot, Peter J; Wallwork, Benjamin; Panizza, Benedict J; Parekh, Harendra S

    2017-01-01

    Mucoadhesive in situ gelling systems (soluble gels) have received considerable attention recently as effective stimuli-transforming vectors for a range of drug delivery applications. Considering this fact, the present work involves systematic formulation development, optimization, functional evaluation and ex vivo performance of thermosensitive soluble gels containing dexamethasone 21-phosphate disodium salt (DXN) as the model therapeutic. A series of in situ gel-forming systems comprising the thermoreversible polymer poloxamer-407 (P407), along with hydroxypropyl methyl cellulose (HPMC) and chitosan were first formulated. The optimized soluble gels were evaluated for their potential to promote greater retention at the mucosal surface, for improved therapeutic efficacy, compared to existing solution/suspension-based steroid formulations used clinically. Optimized soluble gels demonstrated a desirable gelation temperature with Newtonian fluid behaviour observed under storage conditions (4-8°C), and pseudoplastic fluid behaviour recorded at nasal cavity/sinus temperature (≈34°C). The in vitro characterization of formulations including rheological evaluation, textural analysis and mucoadhesion studies of the gel form were investigated. Considerable improvement in mechanical properties and mucoadhesion was observed with incorporation of HPMC and chitosan into the gelling systems. The lead poloxamer-based soluble gels, PGHC4 and PGHC7, which were carried through to ex vivo permeation studies displayed extended drug release profiles in conditions mimicking the human nasal cavity, which indicates their suitability for treating a range of conditions affecting the nasal cavity/sinuses. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Hydrogel-Assisted Antisense LNA Gapmer Delivery for In Situ Gene Silencing in Spinal Cord Injury

    DEFF Research Database (Denmark)

    Moreno, Pedro M.D.; Ferreira, Ana R.; Salvador, Daniela

    2018-01-01

    )-modified AON gapmers in combination with a fibrin hydrogel bridging material to induce gene silencing in situ at a SCI lesion site. LNA gapmers were effectively developed against two promising gene targets aiming at enhancing axonal regeneration—RhoA and GSK3β. The fibrin-matrix-assisted AON delivery system......After spinal cord injury (SCI), nerve regeneration is severely hampered due to the establishment of a highly inhibitory microenvironment at the injury site, through the contribution of multiple factors. The potential of antisense oligonucleotides (AONs) to modify gene expression at different levels...

  12. Temperature-Responsive Smart Nanocarriers for Delivery Of Therapeutic Agents: Applications and Recent Advances.

    Science.gov (United States)

    Karimi, Mahdi; Sahandi Zangabad, Parham; Ghasemi, Alireza; Amiri, Mohammad; Bahrami, Mohsen; Malekzad, Hedieh; Ghahramanzadeh Asl, Hadi; Mahdieh, Zahra; Bozorgomid, Mahnaz; Ghasemi, Amir; Rahmani Taji Boyuk, Mohammad Reza; Hamblin, Michael R

    2016-08-24

    Smart drug delivery systems (DDSs) have attracted the attention of many scientists, as carriers that can be stimulated by changes in environmental parameters such as temperature, pH, light, electromagnetic fields, mechanical forces, etc. These smart nanocarriers can release their cargo on demand when their target is reached and the stimulus is applied. Using the techniques of nanotechnology, these nanocarriers can be tailored to be target-specific, and exhibit delayed or controlled release of drugs. Temperature-responsive nanocarriers are one of most important groups of smart nanoparticles (NPs) that have been investigated during the past decades. Temperature can either act as an external stimulus when heat is applied from the outside, or can be internal when pathological lesions have a naturally elevated termperature. A low critical solution temperature (LCST) is a special feature of some polymeric materials, and most of the temperature-responsive nanocarriers have been designed based on this feature. In this review, we attempt to summarize recent efforts to prepare innovative temperature-responsive nanocarriers and discuss their novel applications.

  13. Dual AAV/IL-10 Plus STAT3 Anti-Inflammatory Gene Delivery Lowers Atherosclerosis in LDLR KO Mice, but without Increased Benefit

    Directory of Open Access Journals (Sweden)

    Maohua Cao

    2012-01-01

    Full Text Available Both IL-10 and STAT3 are in the same signal transduction pathway, with IL-10-bound IL10 receptor (R acting through STAT3 for anti-inflammatory effect. To investigate possible therapeutic synergism, we delivered both full-length wild-type human (h STAT3 and hIL-10 genes by separate adenoassociated virus type 8 (AAV8 tail vein injection into LDLR KO on HCD. Compared to control Neo gene-treated animals, individual hSTAT3 and hIL-10 delivery resulted in significant reduction in atherogenesis, as determined by larger aortic lumen size, thinner aortic wall thickness, and lower blood velocity (all statistically significant. However, dual hSTAT3/hIL-10 delivery offered no improvement in therapeutic effect. Plasma cholesterol levels in dual hSTAT3/hIL-10-treated animals were statistically higher compared to hIL-10 alone. While no advantage was seen in this case, we consider that the dual gene approach has intrinsic merit, but properly chosen partnered genes must be used.

  14. Local Gene Delivery System by Bubble Liposomes and Ultrasound Exposure into Joint Synovium

    Directory of Open Access Journals (Sweden)

    Yoichi Negishi

    2011-01-01

    Full Text Available Recently, we have developed novel polyethylene glycol modified liposomes (bubble liposomes; BL entrapping an ultrasound (US imaging gas, which can work as a gene delivery tool with US exposure. In this study, we investigated the usefulness of US-mediated gene transfer systems with BL into synoviocytes in vitro and joint synovium in vivo. Highly efficient gene transfer could be achieved in the cultured primary synoviocytes transfected with the combination of BL and US exposure, compared to treatment with plasmid DNA (pDNA alone, pDNA plus BL, or pDNA plus US. When BL was injected into the knee joints of mice, and US exposure was applied transcutaneously to the injection site, highly efficient gene expression could be observed in the knee joint transfected with the combination of BL and US exposure, compared to treatment with pDNA alone, pDNA plus BL, or pDNA plus US. The localized and prolonged gene expression was also shown by an in vivo luciferase imaging system. Thus, this local gene delivery system into joint synovium using the combination of BL and US exposure may be an effective means for gene therapy in joint disorders.

  15. An industry update: what is the latest news in therapeutic delivery?

    Science.gov (United States)

    Simpson, Iain

    2017-07-01

    This Industry Update covers the period from 1 through 30 April 2017, and is based on information sourced from company press releases, scientific literature, patents and various news websites. Biogen expanded its portfolio of developmental drugs targeting neurological diseases by licensing a drug from Bristol-Myers Squibb which is currently under investigation for the treatment of progressive supranuclear palsy. Takeda announced a deal with NuBiyota focused on developing treatments for gastrointestinal disorders, based on the manipulation of microbes living in the gut. Novartis announced an expansion of its neuroscience collaboration with Amgen as well as licensing of a developmental compound to treat dry eye. EmulateBio announced a collaboration with the US FDA to explore and evaluate its organs-on-chip technology in toxicological testing of food products, but which has wider opportunities in drug development. In the treatment of pulmonary hypertension, Vectura announced the commercial launch of its Fox nebulizer technology to deliver a Bayer drug and Respira announced a deal in which it exclusively licensed an inhaled drug-device product to United Therapeutics. Published research suggested that the benefit of pumps over multiple injections in the treatment of diabetes maybe more due to training than technology, and a group from the University of Pennsylvania showed that electrical stimulation of the brain can improve memory function but only when applied at the right time. Alzheon published work that explains the mechanism behind its developmental Alzheimer's drug.

  16. Current state of a dual behaviour of antimicrobial peptides-Therapeutic agents and promising delivery vectors.

    Science.gov (United States)

    Piotrowska, Urszula; Sobczak, Marcin; Oledzka, Ewa

    2017-12-01

    Micro-organism resistance is an important challenge in modern medicine due to the global uncontrolled use of antibiotics. Natural and synthetic antimicrobial peptides (AMPs) symbolize a new family of antibiotics, which have stimulated research and clinical interest as new therapeutic options for infections. They represent one of the most promising antimicrobial substances, due to their broad spectrum of biological activity, against bacteria, fungi, protozoa, viruses, yeast and even tumour cells. Besides, being antimicrobial, AMPs have been shown to bind and neutralize bacterial endotoxins, as well as possess immunomodulatory, anti-inflammatory, wound-healing, angiogenic and antitumour properties. In contrast to conventional antibiotics, which have very defined and specific molecular targets, host cationic peptides show varying, complex and very rapid mechanisms of actions that make it difficult to form an effective antimicrobial defence. Importantly, AMPs display their antimicrobial activity at micromolar concentrations or less. To do this, many peptide-based drugs are commercially available for the treatment of numerous diseases, such as hepatitis C, myeloma, skin infections and diabetes. Herein, we present an overview of the general mechanism of AMPs action, along with recent developments regarding carriers of AMPs and their potential applications in medical fields. © 2017 John Wiley & Sons A/S.

  17. An efficient parallel stochastic simulation method for analysis of nonviral gene delivery systems

    KAUST Repository

    Kuwahara, Hiroyuki

    2011-01-01

    Gene therapy has a great potential to become an effective treatment for a wide variety of diseases. One of the main challenges to make gene therapy practical in clinical settings is the development of efficient and safe mechanisms to deliver foreign DNA molecules into the nucleus of target cells. Several computational and experimental studies have shown that the design process of synthetic gene transfer vectors can be greatly enhanced by computational modeling and simulation. This paper proposes a novel, effective parallelization of the stochastic simulation algorithm (SSA) for pharmacokinetic models that characterize the rate-limiting, multi-step processes of intracellular gene delivery. While efficient parallelizations of the SSA are still an open problem in a general setting, the proposed parallel simulation method is able to substantially accelerate the next reaction selection scheme and the reaction update scheme in the SSA by exploiting and decomposing the structures of stochastic gene delivery models. This, thus, makes computationally intensive analysis such as parameter optimizations and gene dosage control for specific cell types, gene vectors, and transgene expression stability substantially more practical than that could otherwise be with the standard SSA. Here, we translated the nonviral gene delivery model based on mass-action kinetics by Varga et al. [Molecular Therapy, 4(5), 2001] into a more realistic model that captures intracellular fluctuations based on stochastic chemical kinetics, and as a case study we applied our parallel simulation to this stochastic model. Our results show that our simulation method is able to increase the efficiency of statistical analysis by at least 50% in various settings. © 2011 ACM.

  18. Preparation and Characterization of Gelatin-Based Mucoadhesive Nanocomposites as Intravesical Gene Delivery Scaffolds

    Directory of Open Access Journals (Sweden)

    Ching-Wen Liu

    2014-01-01

    Full Text Available This study aimed to develop optimal gelatin-based mucoadhesive nanocomposites as scaffolds for intravesical gene delivery to the urothelium. Hydrogels were prepared by chemically crosslinking gelatin A or B with glutaraldehyde. Physicochemical and delivery properties including hydration ratio, viscosity, size, yield, thermosensitivity, and enzymatic degradation were studied, and scanning electron microscopy (SEM was carried out. The optimal hydrogels (H, composed of 15% gelatin A175, displayed an 81.5% yield rate, 87.1% hydration ratio, 42.9 Pa·s viscosity, and 125.8 nm particle size. The crosslinking density of the hydrogels was determined by performing pronase degradation and ninhydrin assays. In vitro lentivirus (LV release studies involving p24 capsid protein analysis in 293T cells revealed that hydrogels containing lentivirus (H-LV had a higher cumulative release than that observed for LV alone (3.7-, 2.3-, and 2.3-fold at days 1, 3, and 5, resp.. Lentivirus from lentivector constructed green fluorescent protein (GFP was then entrapped in hydrogels (H-LV-GFP. H-LV-GFP showed enhanced gene delivery in AY-27 cells in vitro and to rat urothelium by intravesical instillation in vivo. Cystometrogram showed mucoadhesive H-LV reduced peak micturition and threshold pressure and increased bladder compliance. In this study, we successfully developed first optimal gelatin-based mucoadhesive nanocomposites as intravesical gene delivery scaffolds.

  19. In silico prediction of novel therapeutic targets using gene-disease association data.

    Science.gov (United States)

    Ferrero, Enrico; Dunham, Ian; Sanseau, Philippe

    2017-08-29

    Target identification and validation is a pressing challenge in the pharmaceutical industry, with many of the programmes that fail for efficacy reasons showing poor association between the drug target and the disease. Computational prediction of successful targets could have a considerable impact on attrition rates in the drug discovery pipeline by significantly reducing the initial search space. Here, we explore whether gene-disease association data from the Open Targets platform is sufficient to predict therapeutic targets that are actively being pursued by pharmaceutical companies or are already on the market. To test our hypothesis, we train four different classifiers (a random forest, a support vector machine, a neural network and a gradient boosting machine) on partially labelled data and evaluate their performance using nested cross-validation and testing on an independent set. We then select the best performing model and use it to make predictions on more than 15,000 genes. Finally, we validate our predictions by mining the scientific literature for proposed therapeutic targets. We observe that the data types with the best predictive power are animal models showing a disease-relevant phenotype, differential expression in diseased tissue and genetic association with the disease under investigation. On a test set, the neural network classifier achieves over 71% accuracy with an AUC of 0.76 when predicting therapeutic targets in a semi-supervised learning setting. We use this model to gain insights into current and failed programmes and to predict 1431 novel targets, of which a highly significant proportion has been independently proposed in the literature. Our in silico approach shows that data linking genes and diseases is sufficient to predict novel therapeutic targets effectively and confirms that this type of evidence is essential for formulating or strengthening hypotheses in the target discovery process. Ultimately, more rapid and automated target

  20. Reduction-sensitive lipopolyamines as a novel nonviral gene delivery system for modulated release of DNA with improved transgene expression.

    Science.gov (United States)

    Byk, G; Wetzer, B; Frederic, M; Dubertret, C; Pitard, B; Jaslin, G; Scherman, D

    2000-11-16

    We have designed and synthesized original cationic lipids for modulated release of DNA from cationic lipid/DNA complexes. Our rationale was that modulated degradation of the lipids during or after penetration into the cell could improve the trafficking of DNA to the nucleus resulting in increased transgene expression. The new reduction-sensitive lipopolyamines (RSL) harbor a disulfide bridge within different positions in the backbone of the lipids as biosensitive function. A useful synthetic method was developed to obtain, with very good yields and reproducibility, unsymmetrical disulfide-bridged molecules, starting from symmetrical disulfides and thiols. The new lipopolyamines are good candidates as carriers of therapeutic genes for in vivo gene delivery. To optimize the transfection efficiency in these novel series, we have carried out structure-activity relationship studies by placing the disulfide bridge at different positions in the backbone of the cationic lipid and by systematic variation of lipid chain length. Results indicate that the transfection level can be modulated as a function of the location of the disulfide bridge in the molecule. We suggest that an early release of DNA during or after penetration into the cell, probably promoted by reduction of a disulfide bridge placed between the polyamine and the lipid, implies a total loss of transfection efficiency. On the other hand, proper modulation of DNA release by inserting the disulfide bridge between one lipid chain and the rest of the molecule brings about increased transfection efficiency as compared to previously described nondegradable lipopolyamine analogues. Finally, preliminary physicochemical characterization of the complexes demonstrates that DNA release from complexes can be modulated as a function of the surrounding reducing conditions of the complexes and of the localization of the disulfide bridge within the lipopolyamine. Our results suggest that RSL is a promising new approach for gene

  1. The next step in gene delivery: molecular engineering of adeno-associated virus serotypes.

    Science.gov (United States)

    Wang, Jinhui; Faust, Susan M; Rabinowitz, Joseph E

    2011-05-01

    Delivery is at the heart of gene therapy. Viral DNA delivery systems are asked to avoid the immune system, transduce specific target cell types while avoiding other cell types, infect dividing and non-dividing cells, insert their cargo within the host genome without mutagenesis or to remain episomal, and efficiently express transgenes for a substantial portion of a lifespan. These sought-after features cannot be associated with a single delivery system, or can they? The Adeno-associated virus family of gene delivery vehicles has proven to be highly malleable. Pseudotyping, using AAV serotype 2 terminal repeats to generate designer shells capable of transducing selected cell types, enables the packaging of common genomes into multiple serotypes virions to directly compare gene expression and tropism. In this review the ability to manipulate this virus will be examined from the inside out. The influence of host cell factors and organism biology including the immune response on the molecular fate of the viral genome will be discussed as well as differences in cellular trafficking patterns and uncoating properties that influence serotype transduction. Re-engineering the prototype vector AAV2 using epitope insertion, chemical modification, and molecular evolution not only demonstrated the flexibility of the best-studied serotype, but now also expanded the tool kit for molecular modification of all AAV serotypes. Current AAV research has changed its focus from examination of wild-type AAV biology to the feedback of host cell/organism on the design and development of a new generation of recombinant AAV delivery vehicles. This article is part of a Special Section entitled "Special Section: Cardiovascular Gene Therapy". Copyright © 2010 Elsevier Ltd. All rights reserved.

  2. Part II: Functional delivery of a neurotherapeutic gene to neural stem cells using minicircle DNA and nanoparticles: Translational advantages for regenerative neurology.

    Science.gov (United States)

    Fernandes, Alinda R; Chari, Divya M

    2016-09-28

    Both neurotrophin-based therapy and neural stem cell (NSC)-based strategies have progressed to clinical trials for treatment of neurological diseases and injuries. Brain-derived neurotrophic factor (BDNF) in particular can confer neuroprotective and neuro-regenerative effects in preclinical studies, complementing the cell replacement benefits of NSCs. Therefore, combining both approaches by genetically-engineering NSCs to express BDNF is an attractive approach to achieve combinatorial therapy for complex neural injuries. Current genetic engineering approaches almost exclusively employ viral vectors for gene delivery to NSCs though safety and scalability pose major concerns for clinical translation and applicability. Magnetofection, a non-viral gene transfer approach deploying magnetic nanoparticles and DNA with magnetic fields offers a safe alternative but significant improvements are required to enhance its clinical application for delivery of large sized therapeutic plasmids. Here, we demonstrate for the first time the feasibility of using minicircles with magnetofection technology to safely engineer NSCs to overexpress BDNF. Primary mouse NSCs overexpressing BDNF generated increased daughter neuronal cell numbers post-differentiation, with accelerated maturation over a four-week period. Based on our findings we highlight the clinical potential of minicircle/magnetofection technology for therapeutic delivery of key neurotrophic agents. Copyright © 2016 Elsevier B.V. All rights reserved.

  3. Administration of HPV DNA vaccine via electroporation elicits the strongest CD8+ T cell immune responses compared to intramuscular injection and intradermal gene gun delivery

    Science.gov (United States)

    Best, Simon R.; Peng, Shiwen; Juang, Chi-Mou; Hung, Chien-Fu; Hannaman, Drew; Saunders, John R.; Wu, T.-C.; Pai, Sara I.

    2009-01-01

    DNA vaccines are an attractive approach to eliciting antigen-specific immunity. Intracellular targeting of tumor antigens through its linkage to immunostimulatory molecules such as calreticulin (CRT) can improve antigen processing and presentation through the MHC Class I pathway and increase cytotoxic CD8+ T cell production. However, even with these enhancements, the efficacy of such immunotherapeutic strategies is dependent on the identification of an effective route and method of DNA administration. Electroporation and gene gun-mediated particle delivery are leading methods of DNA vaccine delivery that can generate protective and therapeutic levels of immune responses in experimental models. In this study, we perform a head-to-head comparison of three methods of vaccination – conventional intramuscular injection, electroporation mediated intramuscular delivery, and epidermal gene gun-mediated particle delivery - in the ability to generate antigen specific cytotoxic CD8+ T cell responses as well as anti-tumor immune responses against an HPV-16 E7 expressing tumor cell line using the pNGVL4a-CRT/E7(detox) DNA vaccine. Vaccination via electroporation generated the highest number of E7-specific cytotoxic CD8+ T cells, which correlated to improved outcomes in the treatment of growing tumors. In addition, we demonstrate that electroporation results in significantly higher levels of circulating protein compared to gene gun or intramuscular vaccination, which likely enhances calreticulin’s role as a local tumor anti-angiogenesis agent. We conclude that electroporation is a promising method for delivery of HPV DNA vaccines and should be considered for DNA vaccine delivery in human clinical trials. PMID:19622402

  4. Hydrogel Design for Supporting Neurite Outgrowth and Promoting Gene Delivery to Maximize Neurite Extension

    Science.gov (United States)

    Shepard, Jaclyn A.; Stevans, Alyson C.; Holland, Samantha; Wang, Christine E.; Shikanov, Ariella; Shea, Lonnie D.

    2012-01-01

    Hydrogels capable of gene delivery provide a combinatorial approach for nerve regeneration, with the hydrogel supporting neurite outgrowth and gene delivery inducing the expression of inductive factors. This report investigates the design of hydrogels that balance the requirements for supporting neurite growth with those requirements for promoting gene delivery. Enzymatically-degradable PEG hydrogels encapsulating dorsal root ganglia explants, fibroblasts, and lipoplexes encoding nerve growth factor were gelled within channels that can physically guide neurite outgrowth. Transfection of fibroblasts increased with increasing concentration of Arg-Gly-Asp (RGD) cell adhesion sites and decreasing PEG content. The neurite length increased with increasing RGD concentration within 10% PEG hydrogels, yet was maximal within 7.5% PEG hydrogels at intermediate RGD levels. Delivering lipoplexes within the gel produced longer neurites than culture in NGF-supplemented media or co-culture with cells exposed to DNA prior to encapsulation. Hydrogels designed to support neurite outgrowth and deliver gene therapy vectors locally may ultimately be employed to address multiple barriers that limit regeneration. PMID:22038654

  5. Non-Viral Transfection Methods Optimized for Gene Delivery to a Lung Cancer Cell Line

    Science.gov (United States)

    Salimzadeh, Loghman; Jaberipour, Mansooreh; Hosseini, Ahmad; Ghaderi, Abbas

    2013-01-01

    Background Mehr-80 is a newly established adherent human large cell lung cancer cell line that has not been transfected until now. This study aims to define the optimal transfection conditions and effects of some critical elements for enhancing gene delivery to this cell line by utilizing different non-viral transfection Procedures. Methods In the current study, calcium phosphate (CaP), DEAE-dextran, superfect, electroporation and lipofection transfection methods were used to optimize delivery of a plasmid construct that expressed Green Fluorescent Protein (GFP). Transgene expression was detected by fluorescent microscopy and flowcytometry. Toxicities of the methods were estimated by trypan blue staining. In order to evaluate the density of the transfected gene, we used a plasmid construct that expressed the Stromal cell-Derived Factor-1 (SDF-1) gene and measured its expression by real-time PCR. Results Mean levels of GFP-expressing cells 48 hr after transfection were 8.4% (CaP), 8.2% (DEAE-dextran), 4.9% (superfect), 34.1% (electroporation), and 40.1% (lipofection). Lipofection had the highest intense SDF-1 expression of the analyzed methods. Conclusion This study has shown that the lipofection and electroporation methods were more efficient at gene delivery to Mehr-80 cells. The quantity of DNA per transfection, reagent concentration, and incubation time were identified as essential factors for successful transfection in all of the studied methods. PMID:23799175

  6. Non-viral transfection methods optimized for gene delivery to a lung cancer cell line.

    Science.gov (United States)

    Salimzadeh, Loghman; Jaberipour, Mansooreh; Hosseini, Ahmad; Ghaderi, Abbas

    2013-04-01

    Mehr-80 is a newly established adherent human large cell lung cancer cell line that has not been transfected until now. This study aims to define the optimal transfection conditions and effects of some critical elements for enhancing gene delivery to this cell line by utilizing different non-viral transfection Procedures. In the current study, calcium phosphate (CaP), DEAE-dextran, superfect, electroporation and lipofection transfection methods were used to optimize delivery of a plasmid construct that expressed Green Fluorescent Protein (GFP). Transgene expression was detected by fluorescent microscopy and flowcytometry. Toxicities of the methods were estimated by trypan blue staining. In order to evaluate the density of the transfected gene, we used a plasmid construct that expressed the Stromal cell-Derived Factor-1 (SDF-1) gene and measured its expression by real-time PCR. Mean levels of GFP-expressing cells 48 hr after transfection were 8.4% (CaP), 8.2% (DEAE-dextran), 4.9% (superfect), 34.1% (electroporation), and 40.1% (lipofection). Lipofection had the highest intense SDF-1 expression of the analyzed methods. This study has shown that the lipofection and electroporation methods were more efficient at gene delivery to Mehr-80 cells. The quantity of DNA per transfection, reagent concentration, and incubation time were identified as essential factors for successful transfection in all of the studied methods.

  7. Technical Improvement and Application of Hydrodynamic Gene Delivery in Study of Liver Diseases

    Directory of Open Access Journals (Sweden)

    Mei Huang

    2017-08-01

    Full Text Available Development of an safe and efficient in vivo gene delivery method is indispensable for molecular biology research and the progress in the following gene therapy. Over the past few years, hydrodynamic gene delivery (HGD with naked DNA has drawn increasing interest in both research and potential clinic applications due to its high efficiency and low risk in triggering immune responses and carcinogenesis in comparison to viral vectors. This method, involving intravenous injection (i.v. of massive DNA in a short duration, gives a transient but high in vivo gene expression especially in the liver of small animals. In addition to DNA, it has also been shown to deliver other substance such as RNA, proteins, synthetic small compounds and even viruses in vivo. Given its ability to robustly mimic in vivo hepatitis B virus (HBV production in liver, HGD has become a fundamental and important technology on HBV studies in our group and many other groups. Recently, there have been interesting reports about the applications and further improvement of this technology in other liver research. Here, we review the principle, safety, current application and development of hydrodynamic delivery in liver disease studies, and discuss its future prospects, clinical potential and challenges.

  8. A novel Listeria monocytogenes-based DNA delivery system for cancer gene therapy.

    LENUS (Irish Health Repository)

    van Pijkeren, Jan Peter

    2012-01-31

    Bacteria-mediated transfer of plasmid DNA to mammalian cells (bactofection) has been shown to have significant potential as an approach to express heterologous proteins in various cell types. This is achieved through entry of the entire bacterium into cells, followed by release of plasmid DNA. In a murine model, we show that Listeria monocytogenes can invade and spread in tumors, and establish the use of Listeria to deliver genes to tumors in vivo. A novel approach to vector lysis and release of plasmid DNA through antibiotic administration was developed. Ampicillin administration facilitated both plasmid transfer and safety control of vector. To further improve on the gene delivery system, we selected a Listeria monocytogenes derivative that is more sensitive to ampicillin, and less pathogenic than the wild-type strain. Incorporation of a eukaryotic-transcribed lysin cassette in the plasmid further increased bacterial lysis. Successful gene delivery of firefly luciferase to growing tumors in murine models and to patient breast tumor samples ex vivo was achieved. The model described encompasses a three-phase treatment regimen, involving (1) intratumoral administration of vector followed by a period of vector spread, (2) systemic ampicillin administration to induce vector lysis and plasmid transfer, and (3) systemic administration of combined moxifloxacin and ampicillin to eliminate systemic vector. For the first time, our results reveal the potential of Listeria monocytogenes for in vivo gene delivery.

  9. Ultrasound-guided drug delivery in cancer

    Energy Technology Data Exchange (ETDEWEB)

    Chowdhury, Sayan Mullick; Lee, Tae Hwa; Willmann, Jugen K. [Dept. of Radiology, Stanford University School of Medicine, Stanford (United States)

    2017-07-15

    Recent advancements in ultrasound and microbubble (USMB) mediated drug delivery technology has shown that this approach can improve spatially confined delivery of drugs and genes to target tissues while reducing systemic dose and toxicity. The mechanism behind enhanced delivery of therapeutics is sonoporation, the formation of openings in the vasculature, induced by ultrasound-triggered oscillations and destruction of microbubbles. In this review, progress and challenges of USMB mediated drug delivery are summarized, with special focus on cancer therapy.

  10. Ultrasound-guided drug delivery in cancer

    Directory of Open Access Journals (Sweden)

    Sayan Mullick Chowdhury

    2017-07-01

    Full Text Available Recent advancements in ultrasound and microbubble (USMB mediated drug delivery technology has shown that this approach can improve spatially confined delivery of drugs and genes to target tissues while reducing systemic dose and toxicity. The mechanism behind enhanced delivery of therapeutics is sonoporation, the formation of openings in the vasculature, induced by ultrasound-triggered oscillations and destruction of microbubbles. In this review, progress and challenges of USMB mediated drug delivery are summarized, with special focus on cancer therapy.

  11. Hypoxia-regulated therapeutic gene as a preemptive treatment strategy against ischemia/reperfusion tissue injury.

    Science.gov (United States)

    Pachori, Alok S; Melo, Luis G; Hart, Melanie L; Noiseux, Nicholas; Zhang, Lunan; Morello, Fulvio; Solomon, Scott D; Stahl, Gregory L; Pratt, Richard E; Dzau, Victor J

    2004-08-17

    Ischemia and reperfusion represent major mechanisms of tissue injury and organ failure. The timing of administration and the duration of action limit current treatment approaches using pharmacological agents. In this study, we have successfully developed a preemptive strategy for tissue protection using an adenoassociated vector system containing erythropoietin hypoxia response elements for ischemia-regulated expression of the therapeutic gene human heme-oxygenase-1 (hHO-1). We demonstrate that a single administration of this vector several weeks in advance of ischemia/reperfusion injury to multiple tissues such as heart, liver, and skeletal muscle yields rapid and timely induction of hHO-1 during ischemia that resulted in dramatic reduction in tissue damage. In addition, overexpression of therapeutic transgene prevented long-term pathological tissue remodeling and normalized tissue function. Application of this regulatable system using an endogenous physiological stimulus for expression of a therapeutic gene may be a feasible strategy for protecting tissues at risk of ischemia/reperfusion injury.

  12. Hypoxia-regulated therapeutic gene as a preemptive treatment strategy against ischemia/reperfusion tissue injury

    Science.gov (United States)

    Pachori, Alok S.; Melo, Luis G.; Hart, Melanie L.; Noiseux, Nicholas; Zhang, Lunan; Morello, Fulvio; Solomon, Scott D.; Stahl, Gregory L.; Pratt, Richard E.; Dzau, Victor J.

    2004-08-01

    Ischemia and reperfusion represent major mechanisms of tissue injury and organ failure. The timing of administration and the duration of action limit current treatment approaches using pharmacological agents. In this study, we have successfully developed a preemptive strategy for tissue protection using an adenoassociated vector system containing erythropoietin hypoxia response elements for ischemia-regulated expression of the therapeutic gene human heme-oxygenase-1 (hHO-1). We demonstrate that a single administration of this vector several weeks in advance of ischemia/reperfusion injury to multiple tissues such as heart, liver, and skeletal muscle yields rapid and timely induction of hHO-1 during ischemia that resulted in dramatic reduction in tissue damage. In addition, overexpression of therapeutic transgene prevented long-term pathological tissue remodeling and normalized tissue function. Application of this regulatable system using an endogenous physiological stimulus for expression of a therapeutic gene may be a feasible strategy for protecting tissues at risk of ischemia/reperfusion injury.

  13. Early pregnancy peripheral blood gene expression and risk of preterm delivery: a nested case control study

    Directory of Open Access Journals (Sweden)

    Muhie Seid Y

    2009-12-01

    Full Text Available Abstract Background Preterm delivery (PTD is a significant public health problem associated with greater risk of mortality and morbidity in infants and mothers. Pathophysiologic processes that may lead to PTD start early in pregnancy. We investigated early pregnancy peripheral blood global gene expression and PTD risk. Methods As part of a prospective study, ribonucleic acid was extracted from blood samples (collected at 16 weeks gestational age from 14 women who had PTD (cases and 16 women who delivered at term (controls. Gene expressions were measured using the GeneChip® Human Genome U133 Plus 2.0 Array. Student's T-test and fold change analysis were used to identify differentially expressed genes. We used hierarchical clustering and principle components analysis to characterize signature gene expression patterns among cases and controls. Pathway and promoter sequence analyses were used to investigate functions and functional relationships as well as regulatory regions of differentially expressed genes. Results A total of 209 genes, including potential candidate genes (e.g. PTGDS, prostaglandin D2 synthase 21 kDa, were differentially expressed. A set of these genes achieved accurate pre-diagnostic separation of cases and controls. These genes participate in functions related to immune system and inflammation, organ development, metabolism (lipid, carbohydrate and amino acid and cell signaling. Binding sites of putative transcription factors such as EGR1 (early growth response 1, TFAP2A (transcription factor AP2A, Sp1 (specificity protein 1 and Sp3 (specificity protein 3 were over represented in promoter regions of differentially expressed genes. Real-time PCR confirmed microarray expression measurements of selected genes. Conclusions PTD is associated with maternal early pregnancy peripheral blood gene expression changes. Maternal early pregnancy peripheral blood gene expression patterns may be useful for better understanding of PTD

  14. Targeted nanoparticle delivery of therapeutic antisense microRNAs presensitizes glioblastoma cells to lower effective doses of temozolomide in vitro and in a mouse model.

    Science.gov (United States)

    Malhotra, Meenakshi; Sekar, Thillai Veerapazham; Ananta, Jeyarama S; Devulapally, Rammohan; Afjei, Rayhaneh; Babikir, Husam A; Paulmurugan, Ramasamy; Massoud, Tarik F

    2018-04-20

    Temozolomide (TMZ) chemotherapy for glioblastoma (GBM) is generally well tolerated at standard doses but it can cause side effects. GBMs overexpress microRNA-21 and microRNA-10b, two known oncomiRs that promote cancer development, progression and resistance to drug treatment. We hypothesized that systemic injection of antisense microRNAs (antagomiR-21 and antagomiR-10b) encapsulated in cRGD-tagged PEG-PLGA nanoparticles would result in high cellular delivery of intact functional antagomiRs, with consequent efficient therapeutic response and increased sensitivity of GBM cells to lower doses of TMZ. We synthesized both targeted and non-targeted nanoparticles, and characterized them for size, surface charge and encapsulation efficiency of antagomiRs. When using targeted nanoparticles in U87MG and Ln229 GBM cells, we showed higher uptake-associated improvement in sensitivity of these cells to lower concentrations of TMZ in medium. Co-inhibition of microRNA-21 and microRNA-10b reduced the number of viable cells and increased cell cycle arrest at G2/M phase upon TMZ treatment. We found a significant increase in expression of key target genes for microRNA-21 and microRNA-10b upon using targeted versus non-targeted nanoparticles. There was also significant reduction in tumor volume when using TMZ after pre-treatment with loaded nanoparticles in human GBM cell xenografts in mice. In vivo targeted nanoparticles plus different doses of TMZ showed a significant therapeutic response even at the lowest dose of TMZ, indicating that preloading cells with antagomiR-21 and antagomiR-10b increases cellular chemosensitivity towards lower TMZ doses. Future clinical applications of this combination therapy may result in improved GBM response by using lower doses of TMZ and reducing nonspecific treatment side effects.

  15. Live-Attenuated Bacterial Vectors: Tools for Vaccine and Therapeutic Agent Delivery

    Directory of Open Access Journals (Sweden)

    Ivan Y. C. Lin

    2015-11-01

    Full Text Available Genetically attenuated microorganisms, including pathogenic and commensal bacteria, can be engineered to carry and deliver heterologous antigens to elicit host immunity against both the vector as well as the pathogen from which the donor gene is derived. These live attenuated bacterial vectors have been given much attention due to their capacity to induce a broad range of immune responses including localized mucosal, as well as systemic humoral and/or cell-mediated immunity. In addition, the unique tumor-homing characteristics of these bacterial vectors has also been exploited for alternative anti-tumor vaccines and therapies. In such approach, tumor-associated antigen, immunostimulatory molecules, anti-tumor drugs, or nucleotides (DNA or RNA are delivered. Different potential vectors are appropriate for specific applications, depending on their pathogenic routes. In this review, we survey and summarize the main features of the different types of live bacterial vectors and discussed the clinical applications in the field of vaccinology. In addition, different approaches for using live attenuated bacterial vectors for anti-cancer therapy is discussed, and some promising pre-clinical and clinical studies in this field are outlined.

  16. Towards prostate cancer gene therapy: Development of a chlorotoxin-targeted nanovector for toxic (melittin) gene delivery.

    Science.gov (United States)

    Tarokh, Zahra; Naderi-Manesh, Hossein; Nazari, Mahboobeh

    2017-03-01

    Prostate cancer is the second leading cause of death due to cancer in men. Owing to shortcomings in the current treatments, other therapies are being considered. Toxic gene delivery is one of the most effective methods for cancer therapy. Cationic polymers are able to form stable nanoparticles via interaction with nucleic acids electrostatically. Branched polyethylenimine that contains amine groups has notable buffering capacity and the ability to escape from endosome through the proton sponge effect. However, the cytotoxicity of this polymer is high, and modification is one of the applicable strategies to overcome this problem. In this study, PEI was targeted with chlorotoxin (CTX) via N-succinimidyl 3-(2-pyridyldithio) propionate (SPDP) cross-linker. CTX can bind specifically to matrix metalloproteinase-2 that is overexpressed in certain cancers. Melittin as the major component of bee venom has been reported to have anti-cancer activity. This was thus selected to deliver to PC3 cell line. Flow cytometry analysis revealed that transfection efficiency of targeted nanoparticles is significantly higher compared to non-targeted nanoparticles. Targeted nanoparticles carrying the melittin gene also decreased cell viability of PC3 cells significantly while no toxic effects were observed on NIH3T3 cell line. Therefore, CTX-targeted nanoparticles carrying the melittin gene could serve as an appropriate gene delivery system for prostate and other MMP-2 positive cancer cells. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. Towards boron neutron capture therapy: the formulation and preliminary in vitro evaluation of liposomal vehicles for the therapeutic delivery of the dequalinium salt of bis-nido-carborane.

    Science.gov (United States)

    Theodoropoulos, Dimitrios; Rova, Aikaterini; Smith, James R; Barbu, Eugen; Calabrese, Gianpiero; Vizirianakis, Ioannis S; Tsibouklis, John; Fatouros, Dimitrios G

    2013-11-15

    Liposomes of phosphatidylcholine or of dimyristoylphosphatidylcholine that incorporate bis-nido-carborane dequalinium salt are stable in physiologically relevant media and have in vitro toxicity profiles that appear to be compatible with potential therapeutic applications. These features render the structures suitable candidate boron-delivery vehicles for evaluation in the boron neutron capture therapy of cancer. Copyright © 2013 Elsevier Ltd. All rights reserved.

  18. Illuminating the gateway of gene silencing: perspective of RNA interference technology in clinical therapeutics.

    Science.gov (United States)

    Sindhu, Annu; Arora, Pooja; Chaudhury, Ashok

    2012-07-01

    A novel laboratory revolution for disease therapy, the RNA interference (RNAi) technology, has adopted a new era of molecular research as the next generation "Gene-targeted prophylaxis." In this review, we have focused on the chief technological challenges associated with the efforts to develop RNAi-based therapeutics that may guide the biomedical researchers. Many non-curable maladies, like neurodegenerative diseases and cancers have effectively been cured using this technology. Rapid advances are still in progress for the development of RNAi-based technologies that will be having a major impact on medical research. We have highlighted the recent discoveries associated with the phenomenon of RNAi, expression of silencing molecules in mammals along with the vector systems used for disease therapeutics.

  19. Muscle myeloid type I interferon gene expression may predict therapeutic responses to rituximab in myositis patients.

    Science.gov (United States)

    Nagaraju, Kanneboyina; Ghimbovschi, Svetlana; Rayavarapu, Sree; Phadke, Aditi; Rider, Lisa G; Hoffman, Eric P; Miller, Frederick W

    2016-09-01

    To identify muscle gene expression patterns that predict rituximab responses and assess the effects of rituximab on muscle gene expression in PM and DM. In an attempt to understand the molecular mechanism of response and non-response to rituximab therapy, we performed Affymetrix gene expression array analyses on muscle biopsy specimens taken before and after rituximab therapy from eight PM and two DM patients in the Rituximab in Myositis study. We also analysed selected muscle-infiltrating cell phenotypes in these biopsies by immunohistochemical staining. Partek and Ingenuity pathway analyses assessed the gene pathways and networks. Myeloid type I IFN signature genes were expressed at higher levels at baseline in the skeletal muscle of rituximab responders than in non-responders, whereas classic non-myeloid IFN signature genes were expressed at higher levels in non-responders at baseline. Also, rituximab responders have a greater reduction of the myeloid and non-myeloid type I IFN signatures than non-responders. The decrease in the type I IFN signature following administration of rituximab may be associated with the decreases in muscle-infiltrating CD19(+) B cells and CD68(+) macrophages in responders. Our findings suggest that high levels of myeloid type I IFN gene expression in skeletal muscle predict responses to rituximab in PM/DM and that rituximab responders also have a greater decrease in the expression of these genes. These data add further evidence to recent studies defining the type I IFN signature as both a predictor of therapeutic responses and a biomarker of myositis disease activity. Published by Oxford University Press on behalf British Society for Rheumatology 2016. This work is written by US Government employees and is in the public domain in the US.

  20. De-repressing LncRNA-Targeted Genes to Upregulate Gene Expression: Focus on Small Molecule Therapeutics

    Directory of Open Access Journals (Sweden)

    Roya Pedram Fatemi

    2014-01-01

    Full Text Available Non-protein coding RNAs (ncRNAs make up the overwhelming majority of transcripts in the genome and have recently gained attention for their complex regulatory role in cells, including the regulation of protein-coding genes. Furthermore, ncRNAs play an important role in normal development and their expression levels are dysregulated in several diseases. Recently, several long noncoding RNAs (lncRNAs have been shown to alter the epigenetic status of genomic loci and suppress the expression of target genes. This review will present examples of such a mechanism and focus on the potential to target lncRNAs for achieving therapeutic gene upregulation by de-repressing genes that are epigenetically silenced in various diseases. Finally, the potential to target lncRNAs, through their interactions with epigenetic enzymes, using various tools, such as small molecules, viral vectors and antisense oligonucleotides, will be discussed. We suggest that small molecule modulators of a novel class of drug targets, lncRNA-protein interactions, have great potential to treat some cancers, cardiovascular disease, and neurological disorders.

  1. Low cytotoxicity fluorescent PAMAM dendrimer as gene carriers for monitoring the delivery of siRNA

    Energy Technology Data Exchange (ETDEWEB)

    Guan, Lingmei [Sichuan University, State Key Laboratory of Bio-resources and Eco-environment, The Ministry of Education, College of Life Sciences (China); Huang, Saipeng [Chinese Academy of Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, Center for Molecular Sciences, Institute of Chemistry (China); Chen, Zhao [Xi’an Jiaotong University, School of Science (China); Li, Yanchao [Chinese Academy of Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, Center for Molecular Sciences, Institute of Chemistry (China); Liu, Ke [Sichuan University, State Key Laboratory of Bio-resources and Eco-environment, The Ministry of Education, College of Life Sciences (China); Liu, Yang, E-mail: yliu@iccas.ac.cn; Du, Libo, E-mail: dulibo@iccas.ac.cn [Chinese Academy of Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, Center for Molecular Sciences, Institute of Chemistry (China)

    2015-09-15

    Visual detection of gene vectors has attracted a great deal of attention due to the application of these vectors in monitoring and evaluating the effect of gene carriers in living cells. A non-viral vector, the fluorescent PAMAM dendrimer (F-PAMAM), was synthesized through conjugation of PAMAM dendrimers and fluorescein. In vitro and ex vivo experiments show that F-PAMAM exhibits superphotostability, low cytotoxicity and facilitates endocytosis by A549 cells. The vector has a high siRNA binding affinity and it increases the efficiency of cy5-siRNA delivery in A549 cells, in comparison with a cy5-siRNA monomer. Our results provide a new method for simultaneously monitoring the delivery of siRNA and its non-viral carriers in living cells.

  2. Hydrodynamic gene delivery in human skin using a hollow microneedle device.

    Science.gov (United States)

    Dul, M; Stefanidou, M; Porta, P; Serve, J; O'Mahony, C; Malissen, B; Henri, S; Levin, Y; Kochba, E; Wong, F S; Dayan, C; Coulman, S A; Birchall, J C

    2017-11-10

    Microneedle devices have been proposed as a minimally invasive delivery system for the intradermal administration of nucleic acids, both plasmid DNA (pDNA) and siRNA, to treat localised disease or provide vaccination. Different microneedle types and application methods have been investigated in the laboratory, but limited and irreproducible levels of gene expression have proven to be significant challenges to pre-clinical to clinical progression. This study is the first to explore the potential of a hollow microneedle device for the delivery and subsequent expression of pDNA in human skin. The regulatory approved MicronJet600® (MicronJet hereafter) device was used to deliver reporter plasmids (pCMVβ and pEGFP-N1) into viable excised human skin. Exogenous gene expression was subsequently detected at multiple locations that were distant from the injection site but within the confines of the bleb created by the intradermal bolus. The observed levels of gene expression in the tissue are at least comparable to that achieved by the most invasive microneedle application methods e.g. lateral application of a microneedle. Gene expression was predominantly located in the epidermis, although also evident in the papillary dermis. Optical coherence tomography permitted real time visualisation of the sub-surface skin architecture and, unlike a conventional intradermal injection, MicronJet administration of a 50μL bolus appears to create multiple superficial microdisruptions in the papillary dermis and epidermis. These were co-localised with expression of the pCMVβ reporter plasmid. We have therefore shown, for the first time, that a hollow microneedle device can facilitate efficient and reproducible gene expression of exogenous naked pDNA in human skin using volumes that are considered to be standard for intradermal administration, and postulate a hydrodynamic effect as the mechanism of gene delivery. Copyright © 2017 Elsevier B.V. All rights reserved.

  3. Non-Viral Transfection Methods Optimized for Gene Delivery to a Lung Cancer Cell Line

    OpenAIRE

    Salimzadeh, Loghman; Jaberipour, Mansooreh; Hosseini, Ahmad; Ghaderi, Abbas

    2013-01-01

    Background Mehr-80 is a newly established adherent human large cell lung cancer cell line that has not been transfected until now. This study aims to define the optimal transfection conditions and effects of some critical elements for enhancing gene delivery to this cell line by utilizing different non-viral transfection Procedures. Methods In the current study, calcium phosphate (CaP), DEAE-dextran, superfect, electroporation and lipofection transfection methods were used to optimize deliver...

  4. Translational Advancement of Somatostatin Gene Delivery for Disease Modification and Cognitive Sparing in Intractable Epilepsy

    Science.gov (United States)

    2015-09-01

    seizures. A thoroughly characterized rodent epilepsy model will be used as a platform to test the hypotheses. In this model temporal lobe electrical...expression in the hippocampus resected from a young temporal lobe epilepsy patient. Post-baccalaureate student Andrew Moss has since expanded this project...somatostatin gene delivery persistently reduces seizure severity in a rat model of temporal lobe epilepsy , Gowri Natarajan, Jessica Anne McElroy

  5. Hydrodynamic gene delivery in human skin using a hollow microneedle device

    OpenAIRE

    Dul, M.; Stefanidou, M.; Porta, P.; Serve, J.; O'Mahony, Conor; Malissen, B.; Henri, S.; Levin, Y.; Kochba, E.; Wong, F. S.; Dayan, C.; Coulman, S. A.; Birchall, J. C.

    2017-01-01

    Microneedle devices have been proposed as a minimally invasive delivery system for the intradermal administration of nucleic acids, both plasmid DNA (pDNA) and siRNA, to treat localised disease or provide vaccination. Different microneedle types and application methods have been investigated in the laboratory, but limited and irreproducible levels of gene expression have proven to be significant challenges to pre-clinical to clinical progression. This study is the first to explore the potenti...

  6. Short Hairpin RNA (shRNA): Design, Delivery, and Assessment of Gene Knockdown

    Science.gov (United States)

    Moore, Chris B.; Guthrie, Elizabeth H.; Huang, Max Tze-Han; Taxman, Debra J.

    2013-01-01

    Shortly after the cellular mechanism of RNA interference (RNAi) was first described, scientists began using this powerful technique to study gene function. This included designing better methods for the successful delivery of small interfering RNAs (siRNAs) and short hairpin RNAs (shRNAs) into mammalian cells. While the simplest method for RNAi is the cytosolic delivery of siRNA oligonucleotides, this technique is limited to cells capable of transfection and is primarily utilized during transient in vitro studies. The introduction of shRNA into mammalian cells through infection with viral vectors allows for stable integration of shRNA and long-term knockdown of the targeted gene; however, several challenges exist with the implementation of this technology. Here we describe some well-tested protocols which should increase the chances of successful design, delivery, and assessment of gene knockdown by shRNA. We provide suggestions for designing shRNA targets and controls, a protocol for sequencing through the secondary structure of the shRNA hairpin structure, and protocols for packaging and delivery of shRNA lentiviral particles. Using real-time PCR and functional assays we demonstrate the successful knockdown of ASC, an inflammatory adaptor molecule. These studies demonstrate the practicality of including two shRNAs with different efficacies of knockdown to provide an additional level of control and to verify dose dependency of functional effects. Along with the methods described here, as new techniques and algorithms are designed in the future, shRNA is likely to include further promising application and continue to be a critical component of gene discovery. PMID:20387148

  7. Prevention of adverse events of interferon γ gene therapy by gene delivery of interferon γ-heparin-binding domain fusion protein in mice

    Directory of Open Access Journals (Sweden)

    Mitsuru Ando

    2014-01-01

    Full Text Available Sustained gene delivery of interferon (IFN γ can be an effective treatment, but our previous study showed high levels of IFNγ-induced adverse events, including the loss of body weight. These unwanted events could be reduced by target-specific delivery of IFNγ after in vivo gene transfer. To achieve this, we selected the heparin-binding domain (HBD of extracellular superoxide dismutase as a molecule to anchor IFNγ to the cell surface. We designed three IFNγ derivatives, IFNγ-HBD1, IFNγ-HBD2, and IFNγ-HBD3, each of which had 1, 2, or 3 HBDs, respectively. Each plasmid-encoding fusion proteins was delivered to the liver, a model target in this study, by hydrodynamic tail vein injection. The serum concentration of IFNγ-HBD2 and IFNγ-HBD3 after gene delivery was lower than that of IFNγ or IFNγ-HBD1. Gene delivery of IFNγ-HBD2, but not of IFNγ-HBD3, effectively increased the mRNA expression of IFNγ-inducible genes in the liver, suggesting liver-specific distribution of IFNγ-HBD2. Gene delivery of IFNγ-HBD2-suppressed tumor growth in the liver as efficiently as that of IFNγ with much less symptoms of adverse effects. These results indicate that the adverse events of IFNγ gene transfer can be prevented by gene delivery of IFNγ-HBD2, a fusion protein with high cell surface affinity.

  8. Polydnaviruses of Parasitic Wasps: Domestication of Viruses To Act as Gene Delivery Vectors

    Directory of Open Access Journals (Sweden)

    Michael R. Strand

    2012-01-01

    Full Text Available Symbiosis is a common phenomenon in which associated organisms can cooperate in ways that increase their ability to survive, reproduce, or utilize hostile environments. Here, we discuss polydnavirus symbionts of parasitic wasps. These viruses are novel in two ways: (1 they have become non-autonomous domesticated entities that cannot replicate outside of wasps; and (2 they function as a delivery vector of genes that ensure successful parasitism of host insects that wasps parasitize. In this review we discuss how these novelties may have arisen, which genes are potentially involved, and what the consequences have been for genome evolution.

  9. Comparative analysis of DNA nanoparticles and AAVs for ocular gene delivery.

    Directory of Open Access Journals (Sweden)

    Zongchao Han

    Full Text Available Gene therapy is a critical tool for the treatment of monogenic retinal diseases. However, the limited vector capacity of the current benchmark delivery strategy, adeno-associated virus (AAV, makes development of larger capacity alternatives, such as compacted DNA nanoparticles (NPs, critical. Here we conduct a side-by-side comparison of self-complementary AAV and CK30PEG NPs using matched ITR plasmids. We report that although AAVs are more efficient per vector genome (vg than NPs, NPs can drive gene expression on a comparable scale and longevity to AAV. We show that subretinally injected NPs do not leave the eye while some of the AAV-injected animals exhibited vector DNA and GFP expression in the visual pathways of the brain from PI-60 onward. As a result, these NPs have the potential to become a successful alternative for ocular gene therapy, especially for the multitude of genes too large for AAV vectors.

  10. Efficient and safe gene delivery to human corneal endothelium using magnetic nanoparticles.

    Science.gov (United States)

    Czugala, Marta; Mykhaylyk, Olga; Böhler, Philip; Onderka, Jasmine; Stork, Björn; Wesselborg, Sebastian; Kruse, Friedrich E; Plank, Christian; Singer, Bernhard B; Fuchsluger, Thomas A

    2016-07-01

    To develop a safe and efficient method for targeted, anti-apoptotic gene therapy of corneal endothelial cells (CECs). Magnetofection (MF), a combination of lipofection with magnetic nanoparticles (MNPs; PEI-Mag2, SO-Mag5, PalD1-Mag1), was tested in human CECs and in explanted human corneas. Effects on cell viability and function were investigated. Immunocompatibility was assessed in human peripheral blood mononuclear cells. Silica iron-oxide MNPs (SO-Mag5) combined with X-tremeGENE-HP achieved high transfection efficiency in human CECs and explanted human corneas, without altering cell viability or function. Magnetofection caused no immunomodulatory effects in human peripheral blood mononuclear cells. Magnetofection with anti-apoptotic P35 gene effectively blocked apoptosis in CECs. Magnetofection is a promising tool for gene therapy of corneal endothelial cells with potential for targeted on-site delivery.

  11. Transferrin-bearing polypropylenimine dendrimer for targeted gene delivery to the brain.

    Science.gov (United States)

    Somani, Sukrut; Blatchford, David R; Millington, Owain; Stevenson, M Lynn; Dufès, Christine

    2014-08-28

    The possibility of using genes as medicines to treat brain diseases is currently limited by the lack of safe and efficacious delivery systems able to cross the blood-brain barrier, thus resulting in a failure to reach the brain after intravenous administration. On the basis that iron can effectively reach the brain by using transferrin receptors for crossing the blood-brain barrier, we propose to investigate if a transferrin-bearing generation 3-polypropylenimine dendrimer would allow the transport of plasmid DNA to the brain after intravenous administration. In vitro, the conjugation of transferrin to the polypropylenimine dendrimer increased the DNA uptake by bEnd.3 murine brain endothelioma cells overexpressing transferrin receptors, by about 1.4-fold and 2.3-fold compared to that observed with the non-targeted dendriplex and naked DNA. This DNA uptake appeared to be optimal following 2h incubation with the treatment. In vivo, the intravenous injection of transferrin-bearing dendriplex more than doubled the gene expression in the brain compared to the unmodified dendriplex, while decreasing the non-specific gene expression in the lung. Gene expression was at least 3-fold higher in the brain than in any tested peripheral organs and was at its highest 24h following the injection of the treatments. These results suggest that transferrin-bearing polypropylenimine dendrimer is a highly promising gene delivery system to the brain. Copyright © 2014 Elsevier B.V. All rights reserved.

  12. Gene editing in hematopoietic stem cells: a potential therapeutic approach for Fanconi anemia

    International Nuclear Information System (INIS)

    Diez Cabezas, B.

    2015-01-01

    targeting efficiency, due to the toxicity associated with the nucleofection of cells treated with these nanoparticles. In our next step, we moved from healthy donor HSCs to FA hematopoietic cells. Using a therapeutic donor vector carrying the FANCA gene, we demonstrated that gene targeting can correct the phenotype in a FA-A LCL. This was deduced from the restoration of FANCD2 foci formation and the reversion of the sensitivity of FA-A cells to interstrand cross linkers, such as mitomycin C (MMC). To improve the gene targeting efficiency in FA-A hematopoietic cells, we also investigated the effects mediated by the transient inhibition of anti-recombinase PARI. Although the inhibition of PARI increased RAD51 foci, no significant increase of homology directed repair efficiency was observed. In a final set of experiments we demonstrated that our gene targeting approach has also taken place in hematopoietic progenitor cells from FA-A patients, leading to a partial reversion in their hyper-sensitivity to MMC. Our study demonstrates for the first time that gene targeting in the AAVS1 safe harbor locus is feasible in hematopoietic cells from Fanconi anemia-A patients, opening up new perspectives for the future gene therapy of this and other monogenic diseases of the hematopoietic system.(Author)

  13. Gene editing in hematopoietic stem cells: a potential therapeutic approach for Fanconi anemia

    Energy Technology Data Exchange (ETDEWEB)

    Diez Cabezas, B.

    2015-07-01

    targeting efficiency, due to the toxicity associated with the nucleofection of cells treated with these nanoparticles. In our next step, we moved from healthy donor HSCs to FA hematopoietic cells. Using a therapeutic donor vector carrying the FANCA gene, we demonstrated that gene targeting can correct the phenotype in a FA-A LCL. This was deduced from the restoration of FANCD2 foci formation and the reversion of the sensitivity of FA-A cells to interstrand cross linkers, such as mitomycin C (MMC). To improve the gene targeting efficiency in FA-A hematopoietic cells, we also investigated the effects mediated by the transient inhibition of anti-recombinase PARI. Although the inhibition of PARI increased RAD51 foci, no significant increase of homology directed repair efficiency was observed. In a final set of experiments we demonstrated that our gene targeting approach has also taken place in hematopoietic progenitor cells from FA-A patients, leading to a partial reversion in their hyper-sensitivity to MMC. Our study demonstrates for the first time that gene targeting in the AAVS1 safe harbor locus is feasible in hematopoietic cells from Fanconi anemia-A patients, opening up new perspectives for the future gene therapy of this and other monogenic diseases of the hematopoietic system.(Author)

  14. Evaluation of gene delivery strategies to efficiently overexpress functional HLA-G on human bone marrow stromal cells

    Directory of Open Access Journals (Sweden)

    Joana S Boura

    2014-01-01

    Full Text Available Mesenchymal stromal cells (MSC constitutively express low levels of human leukocyte antigen-G (HLA-G, which has been shown to contribute to their immunomodulatory and anti-inflammatory properties. Here, we hypothesized that overexpression of HLA-G on bone marrow-derived MSC would improve their immunomodulatory function, thus increasing their therapeutic potential. Therefore, we investigated which gene transfer system is best suited for delivering this molecule while maintaining its immunomodulatory effects. We performed a side-by-side comparison between three nonviral plasmid-based platforms (pmax-HLA-G1; MC-HLA-G1; pEP-HLA-G1 and a viral system (Lv-HLA-G1 using gene transfer parameters that yielded similar levels of HLA-G1-expressing MSC. Natural killer (NK cell–mediated lysis assays and T cell proliferation assays showed that MSC modified with the HLA-G1 expressing viral vector had significantly lower susceptibility to NK-lysis and significantly reduced T cell proliferation when compared to nonmodified cells or MSC modified with plasmid. We also show that, in plasmid-modified MSC, an increase in Toll-like receptor (TLR9 expression is the mechanism responsible for the abrogation of HLA-G1's immunomodulatory effect. Although MSC can be efficiently modified to overexpress HLA-G1 using viral and nonviral strategies, only viral-based delivery of HLA-G1 is suitable for improvement of MSC's immunomodulatory properties.

  15. QUADrATiC: scalable gene expression connectivity mapping for repurposing FDA-approved therapeutics.

    Science.gov (United States)

    O'Reilly, Paul G; Wen, Qing; Bankhead, Peter; Dunne, Philip D; McArt, Darragh G; McPherson, Suzanne; Hamilton, Peter W; Mills, Ken I; Zhang, Shu-Dong

    2016-05-04

    Gene expression connectivity mapping has proven to be a powerful and flexible tool for research. Its application has been shown in a broad range of research topics, most commonly as a means of identifying potential small molecule compounds, which may be further investigated as candidates for repurposing to treat diseases. The public release of voluminous data from the Library of Integrated Cellular Signatures (LINCS) programme further enhanced the utilities and potentials of gene expression connectivity mapping in biomedicine. We describe QUADrATiC ( http://go.qub.ac.uk/QUADrATiC ), a user-friendly tool for the exploration of gene expression connectivity on the subset of the LINCS data set corresponding to FDA-approved small molecule compounds. It enables the identification of compounds for repurposing therapeutic potentials. The software is designed to cope with the increased volume of data over existing tools, by taking advantage of multicore computing architectures to provide a scalable solution, which may be installed and operated on a range of computers, from laptops to servers. This scalability is provided by the use of the modern concurrent programming paradigm provided by the Akka framework. The QUADrATiC Graphical User Interface (GUI) has been developed using advanced Javascript frameworks, providing novel visualization capabilities for further analysis of connections. There is also a web services interface, allowing integration with other programs or scripts. QUADrATiC has been shown to provide an improvement over existing connectivity map software, in terms of scope (based on the LINCS data set), applicability (using FDA-approved compounds), usability and speed. It offers potential to biological researchers to analyze transcriptional data and generate potential therapeutics for focussed study in the lab. QUADrATiC represents a step change in the process of investigating gene expression connectivity and provides more biologically-relevant results than

  16. Combination therapy and evaluation of therapeutic effect in hepatocellular carcinoma cell using triple reporter genes; containing for NIS, HSV1-sr39tk and GFP

    Energy Technology Data Exchange (ETDEWEB)

    Lee, You La; Lee, Yong Jin; Ahn, Sohn Joo; Ahn, Byeong Cheol; Lee, Sang Woo; Yoo, Jeong Soo; Lee, Jae Tae [Kyungpook National University, Daegu (Korea, Republic of)

    2007-07-01

    To identify therapeutic effect after combine Sodium Iodine Symporter (NIS) and Mutant Herpes-simplex virus type 1 sr39tk (HSV1-sr39tk) expression in hepatocellular carcinoma cell, we transfected triple gene and investigated the properties of these gene ability in hepatocellular carcinoma cell line. After making vector with gene encoding a fusion protein comprised of HSV1-sr39tk and green florescence protein (GFP), to make triple reporter genes NIS gene was further fused to the vector using IRES vector. The vector expressing triple reporter gene was transfected to the Huh-7 cell line using liposome. Functions of hNIS and HSV1-sr39tk expression were confirmed by radio iodine uptake with and without perchlorate and [3H]-penciclovir (3-H PCV) uptake, respectively. To evaluate therapeutic effect in vitro, GCV and I-131 was treated in Huh-7/NTG cell and dual therapy performed. An animal imaging acquired using Optix and microPET in vivo. I-125 uptake was increased up to 100-fold compare to that of non-transfected cells. The transfected cell accumulated H-3 PCV up to 53 times higher at 2 hour than that of non-transfected cells. With fluorescence microscopy, green fluorescence was detected in the transfected cell. In cytotoxic studies, the cell viability of Huh-7/NTG cell was decreased to 41 % of control cell at 10ug/ml GCV concentrations. The survival rate of the Huh-7/NTG cell treated with I-131 decreased up to 16%. In I-131 and GCV dual therapy, Huh-7/NTG cell survival rate decreased up to 4%. In animal studies, Huh-7/NTG tumors showed higher uptake of 18F-FHBG and I-124 than Huh-7 tumors. GFP signal is also higher in Huh-7/NTG tumor than control. We successfully constructed a vector with delivery two therapeutic genes and one reporter gene and transfected the vector to a Huh-7 cell. The hepatocellular carcinoma cell transfected with the vector can be treated with GCV and I-131. The effect of dual gene therapy could be easily assessed by the optical reporter gene imaging.

  17. Mesenchymal stem cell-based gene therapy: A promising therapeutic strategy.

    Science.gov (United States)

    Mohammadian, Mozhdeh; Abasi, Elham; Akbarzadeh, Abolfazl

    2016-08-01

    Mesenchymal stem cells (MSCs) are multipotent stromal cells that exist in bone marrow, fat, and so many other tissues, and can differentiate into a variety of cell types including osteoblasts, chondrocytes, and adipocytes, as well as myocytes and neurons. Moreover, they have great capacity for self-renewal while maintaining their multipotency. Their capacity for proliferation and differentiation, in addition to their immunomodulatory activity, makes them very promising candidates for cell-based regenerative medicine. Moreover, MSCs have the ability of mobilization to the site of damage; therefore, they can automatically migrate to the site of injury via their chemokine receptors following intravenous transplantation. In this respect, they can be applied for MSC-based gene therapy. In this new therapeutic method, genes of interest are introduced into MSCs via viral and non-viral-based methods that lead to transgene expression in them. Although stem cell-based gene therapy is a relatively new strategy, it lights a new hope for the treatment of a variety of genetic disorders. In the near future, MSCs can be of use in a vast number of clinical applications, because of their uncomplicated isolation, culture, and genetic manipulation. However, full consideration is still crucial before they are utilized for clinical trials, because the number of studies that signify the advantageous effects of MSC-based gene therapy are still limited.

  18. Pseudotyped Lentiviral Vectors for Retrograde Gene Delivery into Target Brain Regions

    Directory of Open Access Journals (Sweden)

    Kenta Kobayashi

    2017-08-01

    Full Text Available Gene transfer through retrograde axonal transport of viral vectors offers a substantial advantage for analyzing roles of specific neuronal pathways or cell types forming complex neural networks. This genetic approach may also be useful in gene therapy trials by enabling delivery of transgenes into a target brain region distant from the injection site of the vectors. Pseudotyping of a lentiviral vector based on human immunodeficiency virus type 1 (HIV-1 with various fusion envelope glycoproteins composed of different combinations of rabies virus glycoprotein (RV-G and vesicular stomatitis virus glycoprotein (VSV-G enhances the efficiency of retrograde gene transfer in both rodent and nonhuman primate brains. The most recently developed lentiviral vector is a pseudotype with fusion glycoprotein type E (FuG-E, which demonstrates highly efficient retrograde gene transfer in the brain. The FuG-E–pseudotyped vector permits powerful experimental strategies for more precisely investigating the mechanisms underlying various brain functions. It also contributes to the development of new gene therapy approaches for neurodegenerative disorders, such as Parkinson’s disease, by delivering genes required for survival and protection into specific neuronal populations. In this review article, we report the properties of the FuG-E–pseudotyped vector, and we describe the application of the vector to neural circuit analysis and the potential use of the FuG-E vector in gene therapy for Parkinson’s disease.

  19. Onconase responsive genes in human mesothelioma cells: implications for an RNA damaging therapeutic agent

    International Nuclear Information System (INIS)

    Altomare, Deborah A; Rybak, Susanna M; Pei, Jianming; Maizel, Jacob V; Cheung, Mitchell; Testa, Joseph R; Shogen, Kuslima

    2010-01-01

    Onconase represents a new class of RNA-damaging drugs. Mechanistically, Onconase is thought to internalize, where it degrades intracellular RNAs such as tRNA and double-stranded RNA, and thereby suppresses protein synthesis. However, there may be additional or alternative mechanism(s) of action. In this study, microarray analysis was used to compare gene expression profiles in untreated human malignant mesothelioma (MM) cell lines and cells exposed to 5 μg/ml Onconase for 24 h. A total of 155 genes were found to be regulated by Onconase that were common to both epithelial and biphasic MM cell lines. Some of these genes are known to significantly affect apoptosis (IL-24, TNFAIP3), transcription (ATF3, DDIT3, MAFF, HDAC9, SNAPC1) or inflammation and the immune response (IL-6, COX-2). RT-PCR analysis of selected up- or down-regulated genes treated with varying doses and times of Onconase generally confirmed the expression array findings in four MM cell lines. Onconase treatment consistently resulted in up-regulation of IL-24, previously shown to have tumor suppressive activity, as well as ATF3 and IL-6. Induction of ATF3 and the pro-apoptotic factor IL-24 by Onconase was highest in the two most responsive MM cell lines, as defined by DNA fragmentation analysis. In addition to apoptosis, gene ontology analysis indicated that pathways impacted by Onconase include MAPK signaling, cytokine-cytokine-receptor interactions, and Jak-STAT signaling. These results provide a broad picture of gene activity after treatment with a drug that targets small non-coding RNAs and contribute to our overall understanding of MM cell response to Onconase as a therapeutic strategy. The findings provide insights regarding mechanisms that may contribute to the efficacy of this novel drug in clinical trials of MM patients who have failed first line chemotherapy or radiation treatment

  20. Freeze-dried allograft-mediated gene or protein delivery of growth and differentiation factor 5 reduces reconstructed murine flexor tendon adhesions

    DEFF Research Database (Denmark)

    Svensson, Sys Hasslund; Dadali, Tulin; Ulrich-Vinther, Michael

    2014-01-01

    reverse transcription polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay (ELISA), and in vivo bioluminescent imaging. We then reconstructed flexor digitorum longus (FDL) tendons of the mouse hindlimb with allografts loaded with low and high doses of recombinant GDF-5 protein and r......Advances in allograft processing have opened new horizons for clinical adaptation of flexor tendon allografts as delivery scaffolds for antifibrotic therapeutics. Recombinant adeno-associated-virus (rAAV) gene delivery of the growth and differentiation factor 5 (GDF-5) has been previously...... associated with antifibrotic effects in a mouse model of flexor tendoplasty. In this study, we compared the effects of loading freeze-dried allografts with different doses of GDF-5 protein or rAAV-Gdf5 on flexor tendon healing and adhesions. We first optimized the protein and viral loading parameters using...

  1. CRISPR/Cas9 delivery with one single adenoviral vector devoid of all viral genes.

    Science.gov (United States)

    Ehrke-Schulz, Eric; Schiwon, Maren; Leitner, Theo; Dávid, Stephan; Bergmann, Thorsten; Liu, Jing; Ehrhardt, Anja

    2017-12-07

    The Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9 system revolutionized the field of gene editing but viral delivery of the CRISPR/Cas9 system has not been fully explored. Here we adapted clinically relevant high-capacity adenoviral vectors (HCAdV) devoid of all viral genes for the delivery of the CRISPR/Cas9 machinery using a single viral vector. We present a platform enabling fast transfer of the Cas9 gene and gRNA expression units into the HCAdV genome including the option to choose between constitutive or inducible Cas9 expression and gRNA multiplexing. Efficacy and versatility of this pipeline was exemplified by producing different CRISPR/Cas9-HCAdV targeting the human papillomavirus (HPV) 18 oncogene E6, the dystrophin gene causing Duchenne muscular dystrophy (DMD) and the HIV co-receptor C-C chemokine receptor type 5 (CCR5). All CRISPR/Cas9-HCAdV proved to be efficient to deliver the respective CRISPR/Cas9 expression units and to introduce the desired DNA double strand breaks at their intended target sites in immortalized and primary cells.

  2. Progranulin gene delivery protects dopaminergic neurons in a mouse model of Parkinson's disease.

    Directory of Open Access Journals (Sweden)

    Jackalina M Van Kampen

    Full Text Available Parkinson's disease (PD is a progressive neurodegenerative disorder characterized by tremor, rigidity and akinesia/bradykinesia resulting from the progressive loss of nigrostriatal dopaminergic neurons. To date, only symptomatic treatment is available for PD patients, with no effective means of slowing or stopping the progression of the disease. Progranulin (PGRN is a 593 amino acid multifunction protein that is widely distributed throughout the CNS, localized primarily in neurons and microglia. PGRN has been demonstrated to be a potent regulator of neuroinflammation and also acts as an autocrine neurotrophic factor, important for long-term neuronal survival. Thus, enhancing PGRN expression may strengthen the cells resistance to disease. In the present study, we have used the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP model of PD to investigate the possible use of PGRN gene delivery as a therapy for the prevention or treatment of PD. Viral vector delivery of the PGRN gene was an effective means of elevating PGRN expression in nigrostriatal neurons. When PGRN expression was elevated in the SNC, nigrostriatal neurons were protected from MPTP toxicity in mice, along with a preservation of striatal dopamine content and turnover. Further, protection of nigrostriatal neurons by PGRN gene therapy was accompanied by reductions in markers of MPTP-induced inflammation and apoptosis as well as a complete preservation of locomotor function. We conclude that PGRN gene therapy may have beneficial effects in the treatment of PD.

  3. Enhancing gene delivery of adeno-associated viruses by cell-permeable peptides

    Directory of Open Access Journals (Sweden)

    Yarong Liu

    2014-01-01

    Full Text Available Adeno-associated virus type 2 (AAV2 is considered a promising gene delivery vector and has been extensively applied in several disease models; however, inefficient transduction in various cells and tissues has limited its widespread application in many areas of gene therapy. In this study, we have developed a general, but efficient, strategy to enhance viral transduction, both in vitro and in vivo, by incubating viral particles with cell-permeable peptides (CPPs. We show that CPPs increase internalization of viral particles into cells by facilitating both energy-independent and energy-dependent endocytosis. Moreover, CPPs can significantly enhance the endosomal escape process of viral particles, thus enhancing viral transduction to those cells that have exhibited very low permissiveness to AAV2 infection as a result of impaired intracellular viral processing. We also demonstrated that this approach could be applicable to other AAV serotypes. Thus, the membrane-penetrating ability of CPPs enables us to generate an efficient method for enhanced gene delivery of AAV vectors, potentially facilitating its applicability to human gene therapy.

  4. Design of magnetic gene complexes as effective and serum resistant gene delivery systems for mesenchymal stem cells.

    Science.gov (United States)

    Zhang, Tian-Yuan; Wu, Jia-He; Xu, Qian-Hao; Wang, Xia-Rong; Lu, Jingxiong; Hu, Ying; Jo, Jun-Ichiro; Yamamoto, Masaya; Ling, Daishun; Tabata, Yasuhiko; Gao, Jian-Qing

    2017-03-30

    Gene engineered mesenchymal stem cells (MSCs) have been proposed as promising tools for their various applications in biomedicine. Nevertheless, the lack of an effective and safe way to genetically modify these stem cells is still a major obstacle in the current studies. Herein, we designed novel magnetic complexes by assembling cationized pullulan derivatives with magnetic iron oxide nanoparticles for delivering target genes to MSCs. Results showed that this complexes achieved effective gene expression with the assistance of external magnetic field, and resisted the adverse effect induced by serum proteins on the gene delivery. Moreover, neither significant cytotoxicity nor the interference on the osteogenic differentiation to MSCs were observed after magnetofection. Further studies revealed that this effective and serum resistant gene transfection was partly due to the accelerated and enhanced intracellular uptake process driven by external magnetic field. To conclude, the current study presented a novel option for genetic modification of MSCs in an effective, relatively safe and serum compatible way. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. Efficient in vivo gene transfer to xenotransplanted human skin by lentivirus-mediated, but not by AAV-directed, gene delivery

    DEFF Research Database (Denmark)

    Jakobsen, Maria Vad; Askou, Anne Louise; Dokkedahl, Karin Stenderup

    skin graft, and firefly luciferase expression was observed primarily in neighboring tissue beneath or surrounding the graft. In contrast, gene delivery by intradermally injected lentiviral vectors was efficient and led to extensive and persistent firefly luciferase expression within the human skin...... graft only. The study demonstrates limited capacity of single-stranded AAV vectors of six commonly used serotypes for gene delivery to human skin in vivo....

  6. Comparison of different cationized proteins as biomaterials for nanoparticle-based ocular gene delivery.

    Science.gov (United States)

    Zorzi, Giovanni K; Párraga, Jenny E; Seijo, Begoña; Sanchez, Alejandro

    2015-11-01

    Cationized polymers have been proposed as transfection agents for gene therapy. The present work aims to improve the understanding of the potential use of different cationized proteins (atelocollagen, albumin and gelatin) as nanoparticle components and to investigate the possibility of modulating the physicochemical properties of the resulting nanoparticle carriers by selecting specific protein characteristics in an attempt to improve current ocular gene-delivery approaches. The toxicity profiles, as well as internalization and transfection efficiency, of the developed nanoparticles can be modulated by modifying the molecular weight of the selected protein and the amine used for cationization. The most promising systems are nanoparticles based on intermediate molecular weight gelatin cationized with the endogenous amine spermine, which exhibit an adequate toxicological profile, as well as effective association and protection of pDNA or siRNA molecules, thereby resulting in higher transfection efficiency and gene silencing than the other studied formulations. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. Feasibility of baculovirus-mediated reporter gene delivery for efficient monitoring of islet transplantation in vivo

    International Nuclear Information System (INIS)

    Liu, Shuai; Pan, Yu; Lv, Jing; Wu, Haifei; Tian, Jingyan; Zhang, Yifan

    2014-01-01

    Objective: The objective of this study was to explore the feasibility of baculovirus vector-mediated sodium iodide symporter (NIS) gene delivery to monitor islet transplantation. Methods: Baculovirus vectors expressing green fluorescent protein (GFP) or NIS (Bac-GFP and Bac-NIS) were established using the Bac-to-Bac baculovirus expression system. The GFP expression of Bac-GFP-infected rat islets was observed in vitro by fluorescence microscopy. Iodine uptake and inhibition of iodine uptake by NaClO 4 in Bac-NIS-infected islets were dynamically monitored in vitro. Bac-GFP- or Bac-NIS-infected islets were implanted into the left axillary cavity of NOD-SCID mice, and fluorescence imaging and 125 I NanoSPECT/CT imaging were subsequently performed in vivo. Results: Bac-GFP efficiently infected rat islets (over 95% infected at MOI = 40), and the expression of GFP lasted approximately two weeks. NaClO 4 could inhibit iodine uptake by Bac-NIS-infected islets. In vivo imaging revealed that the fluorescence intensity of the transplant sites in Bac-GFP-infected groups was significantly higher than in the non-infected group. Grafts could be clearly observed by 125 I NanoSPECT/CT imaging for up to 8 h. Conclusion: Baculovirus vectors are powerful vehicles for studying rat islets in gene delivery. It is feasible to use a baculovirus vector to delivery an NIS gene for non-invasive monitoring transplanted islets in vivo by the expression of the target gene

  8. Intracellular Protein Delivery and Gene Transfection by Electroporation Using a Microneedle Electrode Array

    Science.gov (United States)

    Choi, Seong-O; Kim, Yeu-Chun; Lee, Jeong Woo; Park, Jung-Hwan

    2012-01-01

    The impact of many biopharmaceuticals, including protein- and gene-based therapies, has been limited by the need for better methods of delivery into cells within tissues. Here, we present intracellular delivery of molecules and transfection with plasmid DNA by electroporation using a novel microneedle electrode array designed for targeted treatment of skin and other tissue surfaces. The microneedle array is molded out of polylactic acid. Electrodes and circuitry required for electroporation are applied to the microneedle array surface by a new metal-transfer micromolding method. The microneedle array maintains mechanical integrity after insertion into pig cadaver skin and is able to electroporate human prostate cancer cells in vitro. Quantitative measurements show that increasing electroporation pulse voltage increases uptake efficiency of calcein and bovine serum albumin, whereas increasing pulse length has lesser effects over the range studied. Uptake of molecules by up to 50 % of cells and transfection of 12 % of cells with a gene for green fluorescent protein is demonstrated at high cell viability. We conclude that the microneedle electrode array is able to electroporate cells, resulting in intracellular uptake of molecules, and has potential applications to improve intracellular delivery of proteins, DNA and other biopharmaceuticals. PMID:22328093

  9. Polyethylenimine-coated iron oxide magnetic nanoparticles for high efficient gene delivery

    Science.gov (United States)

    Nguyen, Anh H.; Abdelrasoul, Gaser N.; Lin, Donghai; Maadi, Hamid; Tong, Junfeng; Chen, Grace; Wang, Richard; Anwar, Afreen; Shoute, Lian; Fang, Qiang; Wang, Zhixiang; Chen, Jie

    2018-04-01

    Properties of magnetic nanoparticles (MNPs) are of notable interest in many fields of biomedical engineering, especially for gene therapy. In this paper, we report a method for synthesis and delivery of MNPs loaded with DNAs, which overcomes the drawbacks of high cost and cytotoxicity associated with current delivery techniques (chemical- and liposome-based designs). 24-nm MNPs (Fe3O4) were synthesized, functionalized and characterized by analytical techniques to understand the surface properties for DNA binding and cellular uptake. The simple surface functionalization with polyethylenimine (PEI) through glutaraldehyde linker activation gave the complex of PEI-coated MNPs, resulting in high stability with a positive surface charge of about + 31 mV. Under the guidance of an external magnetic field, the functionalized MNPs with a loaded isothiocyanate (FITC) or green fluorescent protein (GFP) will enter the cells, which can be visualized by the fluorescence of FITC or GFP. We also examined the cytotoxicity of our synthesized MNPs by MTT assay. We showed that the IC50s of these MNPs for COS-7 and CHO cells were low and at 0.2 and 0.26 mg/mL, respectively. Moreover, our synthesized MNPs that were loaded with plasmids encoding GFP showed high transfection rate, 38.3% for COS-7cells and 27.6% for CHO cells. In conclusion, we established a promising method with low cost, low toxicity, and high transfection efficiency for siRNA and gene delivery.

  10. Anchoring of self-assembled plasmid DNA/ anti-DNA antibody/cationic lipid micelles on bisphosphonate-modified stent for cardiovascular gene delivery

    Directory of Open Access Journals (Sweden)

    Ma G

    2013-03-01

    -immobilized PAA-BP-modified stents were successful as a gene delivery system. Gene delivery using DAC micelle-tethered stent-based PAA-BP functionalization should be suitable for a wide array of single or multiple therapeutic gene strategies, and could be used on cardiovascular metallic implants for achieving efficient gene therapy. Keywords: stent, polyallylamine bisphosphonate, plasmid DNA, micelles, gene delivery

  11. Utilizing Biomarker Signature Pairs To Develop Gene Therapeutic Viral Delivery Platforms For Treating Prostate Cancer

    Science.gov (United States)

    Dr. Tamaro Hudson is currently an Assistant Professor at Howard University in the Department of Pharmacology and holds an appointment as a Health Research Specialist at the Washington VA Medical Center. Dr. Hudson received his Bachelor of Science from Iowa State University in Biology in 1994 and went on to receive a Master of Science in Preventive Medicine from Ohio State University in 2007. Afterwards, he received a Ph.D. from Ohio State University in 2002 where he focused on evaluating the functional differences among isothiocyanates in the rat esophageal tumor model. Following his Ph.D., Dr. Hudson was selected to complete a prestigious Cancer Prevention Fellowship Program at the National Institute of Health, National Cancer Institute, where he focused on utilizing in vitro and in vivo cancer models to assess the biological activity of bioactive compounds on prostate cancer molecular pathways. Concurrently, he completed a Master of Public Health degree from George Washington University in 2003 where he focused on assessing the degree of agreement between a food frequency questionnaire and a 4-day food record as it related to dietary fiber intake. Upon completion of his MPH and Fellowship, he was recruited by Howard University Cancer Center in 2007 as an Assistant Professor. Since joining the Howard faculty, Dr. Hudson has integrated his research focus by identifying novel signature biomarkers – that could have a significant impact on both the diagnosis and targeted treatment of prostate cancer – with the evaluation of new chemopreventive strategies, which have been evaluated in Phase I and Phase II clinical trials. Dr. Hudson received the first five-year VA-HBCU Research, Scientist, and Training grant that focuses on developing a biomarker-based risk prediction model for prostate cancer. Dr. Hudson serves on several Howard University committees and has many peer-reviewed publications. Dr. Hudson's research interests continue to expand as he tries to build collaborations across broad disciplines to make an impact in translational research.  

  12. Activation of Fetal γ-globin Gene Expression via Direct Protein Delivery of Synthetic Zinc-finger DNA-Binding Domains

    Directory of Open Access Journals (Sweden)

    Mir A Hossain

    2016-01-01

    Full Text Available Reactivation of γ-globin expression has been shown to ameliorate disease phenotypes associated with mutations in the adult β-globin gene, including sickle cell disease. Specific mutations in the promoter of the γ-globin genes are known to prevent repression of the genes in the adult and thus lead to hereditary persistence of fetal hemoglobin. One such hereditary persistence of fetal hemoglobin is associated with a sequence located 567 bp upstream of the Gγ-globin gene which assembles a GATA-containing repressor complex. We generated two synthetic zinc-finger DNA-binding domains (ZF-DBDs targeting this sequence. The -567Gγ ZF-DBDs associated with high affinity and specificity with the target site in the γ-globin gene promoter. We delivered the -567Gγ ZF-DBDs directly to primary erythroid cells. Exposure of these cells to the recombinant -567Gγ ZF-DBDs led to increased expression of the γ-globin gene. Direct protein delivery of ZF-DBDs that compete with transcription regulatory proteins will have broad implications for modulating gene expression in analytical or therapeutic settings.

  13. Design and application of cationic amphiphilic β-cyclodextrin derivatives as gene delivery vectors

    Science.gov (United States)

    Wan, Ning; Huan, Meng-Lei; Ma, Xi-Xi; Jing, Zi-Wei; Zhang, Ya-Xuan; Li, Chen; Zhou, Si-Yuan; Zhang, Bang-Le

    2017-11-01

    The nano self-assembly profiles of amphiphilic gene delivery vectors could improve the density of local cationic head groups to promote their DNA condensation capability and enhance the interaction between cell membrane and hydrophobic tails, thus increasing cellular uptake and gene transfection. In this paper, two series of cationic amphiphilic β-cyclodextrin (β-CD) derivatives were designed and synthesized by using 6-mono-OTs-β-CD (1) as the precursor to construct amphiphilic gene vectors with different building blocks in a selective and controlled manner. The effect of different type and degree of cationic head groups on transfection and the endocytic mechanism of β-CD derivatives/DNA nanocomplexes were also investigated. The results demonstrated that the designed β-cyclodextrin derivatives were able to compact DNA to form stable nanocomplexes and exhibited low cytotoxicity. Among them, PEI-1 with PEI head group showed enhanced transfection activity, significantly higher than commercially available agent PEI25000 especially in the presence of serum, showing potential application prospects in clinical trials. Moreover, the endocytic uptake mechanism involved in the gene transfection of PEI-1 was mainly through caveolae-mediated endocytosis, which could avoid the lysosomal degradation of loaded gene, and had great importance for improving gene transfection activity.

  14. Gene Transfer Corrects Acute GM2 Gangliosidosis—Potential Therapeutic Contribution of Perivascular Enzyme Flow

    Science.gov (United States)

    Cachón-González, M Begoña; Wang, Susan Z; McNair, Rosamund; Bradley, Josephine; Lunn, David; Ziegler, Robin; Cheng, Seng H; Cox, Timothy M

    2012-01-01

    The GM2 gangliosidoses are fatal lysosomal storage diseases principally affecting the brain. Absence of β-hexosaminidase A and B activities in the Sandhoff mouse causes neurological dysfunction and recapitulates the acute Tay–Sachs (TSD) and Sandhoff diseases (SD) in infants. Intracranial coinjection of recombinant adeno-associated viral vectors (rAAV), serotype 2/1, expressing human β-hexosaminidase α (HEXA) and β (HEXB) subunits into 1-month-old Sandhoff mice gave unprecedented survival to 2 years and prevented disease throughout the brain and spinal cord. Classical manifestations of disease, including spasticity—as opposed to tremor-ataxia—were resolved by localized gene transfer to the striatum or cerebellum, respectively. Abundant biosynthesis of β-hexosaminidase isozymes and their global distribution via axonal, perivascular, and cerebrospinal fluid (CSF) spaces, as well as diffusion, account for the sustained phenotypic rescue—long-term protein expression by transduced brain parenchyma, choroid plexus epithelium, and dorsal root ganglia neurons supplies the corrective enzyme. Prolonged survival permitted expression of cryptic disease in organs not accessed by intracranial vector delivery. We contend that infusion of rAAV into CSF space and intraparenchymal administration by convection-enhanced delivery at a few strategic sites will optimally treat neurodegeneration in many diseases affecting the nervous system. PMID:22453766

  15. Restoration of central nervous system alpha-N-acetylglucosaminidase activity and therapeutic benefits in mucopolysaccharidosis IIIB mice by a single intracisternal recombinant adeno-associated viral type 2 vector delivery.

    Science.gov (United States)

    Fu, Haiyan; DiRosario, Julianne; Kang, Lu; Muenzer, Joseph; McCarty, Douglas M

    2010-07-01

    Finding efficient central nervous system (CNS) delivery approaches has been the major challenge facing therapeutic development for treating diseases with global neurological manifestation, such as mucopolysaccharidosis (MPS) IIIB, a lysosomal storage disease, caused by autosomal recessive defect of alpha-N-acetylglucosaminidase (NaGlu). Previously, we developed an approach, intracisternal (i.c.) injection, to deliver recombinant adeno-associated viral (rAAV) vector to the CNS of mice, leading to a widespread periventricular distribution of transduction. In the present study, we delivered rAAV2 vector expressing human NaGlu into the CNS of MPS IIIB mice by an i.c. injection approach, to test its therapeutic efficacy and feasibility for treating the neurological manifestation of the disease. We demonstrated significant functional neurological benefits of a single i.c. vector infusion in adult MPS IIIB mice. The treatment slowed the disease progression by mediating widespread recombinant NaGlu expression in the CNS, resulting in the reduction of brain lysosomal storage pathology, significantly improved cognitive function and prolonged survival. However, persisting motor function deficits suggested that pathology in areas outside the CNS contributes to the MPS IIIB behavioral phenotype. The therapeutic benefit of i.c. rAAV2 delivery was dose-dependent and could be attribute solely to the CNS transduction because the procedure did not lead to detectable transduction in somatic tissues. A single IC rAAV2 gene delivery is functionally beneficial for treating the CNS disease of MPS IIIB in mice. It is immediately clinically translatable, with the potential of improving the quality of life for patients with MPS IIIB.

  16. Optimizing hyaluronidase dose and plasmid DNA delivery greatly improves gene electrotransfer efficiency in rat skeletal muscle

    DEFF Research Database (Denmark)

    Åkerström, Thorbjörn; Vedel, Kenneth; Needham Andersen, Josefine

    2015-01-01

    Transfection of rat skeletal muscle in vivo is a widely used research model. However, gene electrotransfer protocols have been developed for mice and yield variable results in rats. We investigated whether changes in hyaluronidase pre-treatment and plasmid DNA delivery can improve transfection...... with a homogenous distribution. We also show that transfection was stable over five weeks of regular exercise or inactivity. Our findings show that species-specific plasmid DNA delivery and hyaluronidase pre-treatment greatly improves transfection efficiency in rat skeletal muscle....... efficiency in rat skeletal muscle. We found that pre-treating the muscle with a hyaluronidase dose suitable for rats (0.56. U/g b.w.) prior to plasmid DNA injection increased transfection efficiency by >200% whereas timing of the pre-treatment did not affect efficiency. Uniformly distributing plasmid DNA...

  17. In Silico, In Vitro, and In Vivo Studies Indicate the Potential Use of Bolaamphiphiles for Therapeutic siRNAs Delivery

    Directory of Open Access Journals (Sweden)

    Taejin Kim

    2013-01-01

    Full Text Available Specific small interfering RNAs (siRNAs designed to silence different oncogenic pathways can be used for cancer therapy. However, non-modified naked siRNAs have short half-lives in blood serum and encounter difficulties in crossing biological membranes due to their negative charge. These obstacles can be overcome by using siRNAs complexed with bolaamphiphiles, consisting of two positively charged head groups that flank an internal hydrophobic chain. Bolaamphiphiles have relatively low toxicities, long persistence in the blood stream, and most importantly, in aqueous conditions can form poly-cationic micelles thus, becoming amenable to association with siRNAs. Herein, two different bolaamphiphiles with acetylcholine head groups attached to an alkyl chain in two distinct configurations are compared for their abilities to complex with siRNAs and deliver them into cells inducing gene silencing. Our explicit solvent molecular dynamics (MD simulations showed that bolaamphiphiles associate with siRNAs due to electrostatic, hydrogen bonding, and hydrophobic interactions. These in silico studies are supported by various in vitro and in cell culture experimental techniques as well as by some in vivo studies. Results demonstrate that depending on the application, the extent of siRNA chemical protection, delivery efficiency, and further intracellular release can be varied by simply changing the type of bolaamphiphile used.

  18. Functionalized gold nanoparticles for the binding, stabilization, and delivery of therapeutic DNA, RNA, and other biological macromolecules

    Directory of Open Access Journals (Sweden)

    Robert K DeLong

    2010-09-01

    Full Text Available Robert K DeLong1, Christopher M Reynolds1, Yaneika Malcolm1, Ashley Schaeffer1, Tiffany Severs2, Adam Wanekaya21Department of Biomedical Science (Cell and Molecular Biology Program, 2Department of Chemistry, Missouri State University, Springfield, MO, USAAbstract: Nanotechnology has virtually exploded in the last few years with seemingly limitless opportunity across all segments of our society. If gene and RNA therapy are to ever realize their full potential, there is a great need for nanomaterials that can bind, stabilize, and deliver these macromolecular nucleic acids into human cells and tissues. Many researchers have turned to gold nanomaterials, as gold is thought to be relatively well tolerated in humans and provides an inert material upon which nucleic acids can attach. Here, we review the various strategies for associating macromolecular nucleic acids to the surface of gold nanoparticles (GNPs, the characterization chemistries involved, and the potential advantages of GNPs in terms of stabilization and delivery.Keywords: gold, nanoparticles, nanomaterials, RNA, nucleic acid

  19. Delivery of human NKG2D-IL-15 fusion gene by chitosan nanoparticles to enhance antitumor immunity

    International Nuclear Information System (INIS)

    Yan, Chen; Jie, Leng; Yongqi, Wang; Weiming, Xiao; Juqun, Xi; Yanbing, Ding; Li, Qian; Xingyuan, Pan; Mingchun, Ji; Weijuan, Gong

    2015-01-01

    Nanoparticles are becoming promising carriers for gene delivery because of their high capacity in gene loading and low cell cytotoxicity. In this study, a chitosan-based nanoparticle encapsulated within a recombinant pcDNA3.1-dsNKG2D-IL-15 plasmid was generated. The fused dsNKG2D-IL-15 gene fragment consisted of double extracellular domains of NKG2D with IL-15 gene at downstream. The average diameter of the gene nanoparticles ranged from 200 nm to 400 nm, with mean zeta potential value of 53.8 ± 6.56 mV. The nanoparticles which were loaded with the dsNKG2D-IL-15 gene were uptaken by tumor cells with low cytotoxicity. Tumor cells pre-transfected by gene nanopartilces stimulated NK and T cells in vitro. Intramuscular injection of gene nanoparticles suppressed tumor growth and prolonged survival of tumor-bearing mice through activation of NK and CD8 + T cells. Thus, chitosan-based nanoparticle delivery of dsNKG2D-IL-15 gene vaccine can be potentially used for tumor therapy. - Highlights: • Generation of a nanoparticle for delivery of dsNKG2D-IL-15 gene. • Characterization of the gene nanoparticle. • Antitumor activity mediated by the gene nanoparticle

  20. Delivery of human NKG2D-IL-15 fusion gene by chitosan nanoparticles to enhance antitumor immunity

    Energy Technology Data Exchange (ETDEWEB)

    Yan, Chen; Jie, Leng; Yongqi, Wang [Department of Immunology, School of Medicine, Yangzhou University, Yangzhou, 225009 (China); Weiming, Xiao [Department of Gastroenterology, The Second Clinical Medical College, Yangzhou University, Yangzhou, 225009 (China); Juqun, Xi [Jiangsu Key Laboratory of Integrated Traditional Chinese and Western Medicine for Prevention and Treatment of Senile Diseases, Yangzhou, 225009 (China); Yanbing, Ding [Department of Gastroenterology, The Second Clinical Medical College, Yangzhou University, Yangzhou, 225009 (China); Li, Qian [Department of Immunology, School of Medicine, Yangzhou University, Yangzhou, 225009 (China); Xingyuan, Pan [Jiangsu Key Laboratory of Zoonosis, Yangzhou University, Yangzhou, 225009 (China); Mingchun, Ji [Department of Immunology, School of Medicine, Yangzhou University, Yangzhou, 225009 (China); Weijuan, Gong, E-mail: wjgong@yzu.edu.cn [Department of Immunology, School of Medicine, Yangzhou University, Yangzhou, 225009 (China); Department of Gastroenterology, The Second Clinical Medical College, Yangzhou University, Yangzhou, 225009 (China); Jiangsu Key Laboratory of Integrated Traditional Chinese and Western Medicine for Prevention and Treatment of Senile Diseases, Yangzhou, 225009 (China); Jiangsu Key Laboratory of Zoonosis, Yangzhou University, Yangzhou, 225009 (China); Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, 225009 (China)

    2015-07-31

    Nanoparticles are becoming promising carriers for gene delivery because of their high capacity in gene loading and low cell cytotoxicity. In this study, a chitosan-based nanoparticle encapsulated within a recombinant pcDNA3.1-dsNKG2D-IL-15 plasmid was generated. The fused dsNKG2D-IL-15 gene fragment consisted of double extracellular domains of NKG2D with IL-15 gene at downstream. The average diameter of the gene nanoparticles ranged from 200 nm to 400 nm, with mean zeta potential value of 53.8 ± 6.56 mV. The nanoparticles which were loaded with the dsNKG2D-IL-15 gene were uptaken by tumor cells with low cytotoxicity. Tumor cells pre-transfected by gene nanopartilces stimulated NK and T cells in vitro. Intramuscular injection of gene nanoparticles suppressed tumor growth and prolonged survival of tumor-bearing mice through activation of NK and CD8{sup +} T cells. Thus, chitosan-based nanoparticle delivery of dsNKG2D-IL-15 gene vaccine can be potentially used for tumor therapy. - Highlights: • Generation of a nanoparticle for delivery of dsNKG2D-IL-15 gene. • Characterization of the gene nanoparticle. • Antitumor activity mediated by the gene nanoparticle.

  1. Development of Non-Viral, Trophoblast-Specific Gene Delivery for Placental Therapy.

    Directory of Open Access Journals (Sweden)

    Noura Abd Ellah

    Full Text Available Low birth weight is associated with both short term problems and the fetal programming of adult onset diseases, including an increased risk of obesity, diabetes and cardiovascular disease. Placental insufficiency leading to intrauterine growth restriction (IUGR contributes to the prevalence of diseases with developmental origins. Currently there are no therapies for IUGR or placental insufficiency. To address this and move towards development of an in utero therapy, we employ a nanostructure delivery system complexed with the IGF-1 gene to treat the placenta. IGF-1 is a growth factor critical to achieving appropriate placental and fetal growth. Delivery of genes to a model of human trophoblast and mouse placenta was achieved using a diblock copolymer (pHPMA-b-pDMAEMA complexed to hIGF-1 plasmid DNA under the control of trophoblast-specific promoters (Cyp19a or PLAC1. Transfection efficiency of pEGFP-C1-containing nanocarriers in BeWo cells and non-trophoblast cells was visually assessed via fluorescence microscopy. In vivo transfection and functionality was assessed by direct placental-injection into a mouse model of IUGR. Complexes formed using pHPMA-b-pDMAEMA and CYP19a-923 or PLAC1-modified plasmids induce trophoblast-selective transgene expression in vitro, and placental injection of PLAC1-hIGF-1 produces measurable RNA expression and alleviates IUGR in our mouse model, consequently representing innovative building blocks towards human placental gene therapies.

  2. Systemic gene delivery transduces the enteric nervous system of guinea pigs and cynomolgus macaques.

    Science.gov (United States)

    Gombash, S E; Cowley, C J; Fitzgerald, J A; Lepak, C A; Neides, M G; Hook, K; Todd, L J; Wang, G-D; Mueller, C; Kaspar, B K; Bielefeld, E C; Fischer, A J; Wood, J D; Foust, K D

    2017-10-01

    Characterization of adeno-associated viral vector (AAV) mediated gene delivery to the enteric nervous system (ENS) was recently described in mice and rats. In these proof-of-concept experiments, we show that intravenous injections of clinically relevant AAVs can transduce the ENS in guinea pigs and non-human primates. Neonatal guinea pigs were given intravenous injections of either AAV8 or AAV9 vectors that contained a green fluorescent protein (GFP) expression cassette or phosphate-buffered saline. Piglets were euthanized three weeks post injection and tissues were harvested for immunofluorescent analysis. GFP expression was detected in myenteric and submucosal neurons along the length of the gastrointestinal tract in AAV8 injected guinea pigs. GFP-positive neurons were found in dorsal motor nucleus of the vagus and dorsal root ganglia. Less transduction occurred in AAV9-treated tissues. Gastrointestinal tissues were analyzed from young cynomolgus macaques that received systemic injection of AAV9 GFP. GFP expression was detected in myenteric neurons of the stomach, small and large intestine. These data demonstrate that ENS gene delivery translates to larger species. This work develops tools for the field of neurogastroenterology to explore gut physiology and anatomy using emerging technologies such as optogenetics and gene editing. It also provides a basis to develop novel therapies for chronic gut disorders.

  3. Water soluble cationic dextran derivatives containing poly(amidoamine) dendrons for efficient gene delivery.

    Science.gov (United States)

    Mai, Kaijin; Zhang, Shanshan; Liang, Bing; Gao, Cong; Du, Wenjun; Zhang, Li-Ming

    2015-06-05

    To develop new dextran derivatives for efficient gene delivery, the conjugation of poly(amidoamine) dendrons onto biocompatible dextran was carried out by a Cu(I)-catalyzed azide-alkyne cycloaddition, as confirmed by FTIR and (1)H NMR analyses. For resultant dextran conjugates with various generations of poly(amidoamine) dendrons, their buffering capacity and in vitro cytotoxicity were evaluated by acid-base titration and MTT tests, respectively. In particular, their physicochemical characteristics for the complexation with plasmid DNA were investigated by the combined analyses from agarose gel electrophoresis, zeta potential, particle size, transmission electron microscopy and fluorescence emission spectra. Moreover, their complexes with plasmid DNA were studied with respect to their transfection efficiency in human embryonic kidney (HEK293) cell lines. In the case of a higher generation of poly(amidoamine) dendrons, such a dextran conjugate was found to have much lower cytotoxicity and better gene delivery capability when compared to branched polyethylenimine (bPEI, 25kDa), a commonly used gene vector. Copyright © 2015 Elsevier Ltd. All rights reserved.

  4. Cell-type-specific gene delivery into neuronal cells in vitro and in vivo

    International Nuclear Information System (INIS)

    Parveen, Zahida; Mukhtar, Muhammad; Rafi, Mohammed; Wenger, David A.; Siddiqui, Khwaja M.; Siler, Catherine A.; Dietzschold, Bernhard; Pomerantz, Roger J.; Schnell, Matthias J.; Dornburg, Ralph

    2003-01-01

    The avian retroviruses reticuloendotheliosis virus strain A (REV-A) and spleen necrosis virus (SNV) are not naturally infectious in human cells. However, REV-A-derived viral vectors efficiently infect human cells when they are pseudotyped with envelope proteins displaying targeting ligands specific for human cell-surface receptors. Here we report that vectors containing the gag region of REV-A and pol of SNV can be pseudotyped with the envelope protein of vesicular stomatitis virus (VSV) and the glycoproteins of different rabies virus (RV) strains. Vectors pseudotyped with the envelope protein of the highly neurotropic RV strain CVS-N2c facilitated cell type-specific gene delivery into mouse and human neurons, but did not infect other human cell types. Moreover, when such vector particles were injected into the brain of newborn mice, only neuronal cells were infected in vivo. Cell-type-specific gene delivery into neurons may present quite specific gene therapy approaches for many degenerative diseases of the brain

  5. p53, SKP2, and DKK3 as MYCN Target Genes and Their Potential Therapeutic Significance

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Lindi; Tweddle, Deborah A., E-mail: deborah.tweddle@ncl.ac.uk [Newcastle Cancer Centre, Northern Institute for Cancer Research, Newcastle University, Newcastle (United Kingdom)

    2012-11-28

    Neuroblastoma is the most common extra-cranial solid tumor of childhood. Despite significant advances, it currently still remains one of the most difficult childhood cancers to cure, with less than 40% of patients with high-risk disease being long-term survivors. MYCN is a proto-oncogene implicated to be directly involved in neuroblastoma development. Amplification of MYCN is associated with rapid tumor progression and poor prognosis. Novel therapeutic strategies which can improve the survival rates whilst reducing the toxicity in these patients are therefore required. Here we discuss genes regulated by MYCN in neuroblastoma, with particular reference to p53, SKP2, and DKK3 and strategies that may be employed to target them.

  6. Genome-wide gene expression dataset used to identify potential therapeutic targets in androgenetic alopecia

    Directory of Open Access Journals (Sweden)

    R. Dey-Rao

    2017-08-01

    Full Text Available The microarray dataset attached to this report is related to the research article with the title: “A genomic approach to susceptibility and pathogenesis leads to identifying potential novel therapeutic targets in androgenetic alopecia” (Dey-Rao and Sinha, 2017 [1]. Male-pattern hair loss that is induced by androgens (testosterone in genetically predisposed individuals is known as androgenetic alopecia (AGA. The raw dataset is being made publicly available to enable critical and/or extended analyses. Our related research paper utilizes the attached raw dataset, for genome-wide gene-expression associated investigations. Combined with several in silico bioinformatics-based analyses we were able to delineate five strategic molecular elements as potential novel targets towards future AGA-therapy.

  7. Dual responsive promoters to target therapeutic gene expression to radiation-resistant hypoxic tumor cells

    International Nuclear Information System (INIS)

    Chadderton, Naomi; Cowen, Rachel L.; Sheppard, Freda C.D.; Robinson, Suzanne; Greco, Olga; Scott, Simon D.; Stratford, Ian J.; Patterson, Adam V.; Williams, Kaye J.

    2005-01-01

    Purpose: Tumor hypoxia is unequivocally linked to poor radiotherapy outcome. This study aimed to identify enhancer sequences that respond maximally to a combination of radiation and hypoxia for use in genetic radiotherapy approaches. Methods and materials: The influence of radiation (5 Gy) and hypoxia (1% O 2 ) on reporter-gene expression driven by hypoxia (HRE) and radiation (Egr-1) responsive elements was evaluated in tumor cells grown as monolayers or multicellular spheroids. Hypoxia-inducible factor-1α (HIF-1α) and HIF-2α protein expression was monitored in parallel. Results: Of the sequences tested, an HRE from the phosphoglycerate kinase-1 gene (PGK-18[5+]) was maximally induced in response to hypoxia plus radiation in all 5 cell lines tested. The additional radiation treatment afforded a significant increase in the induction of PGK-18[5+] compared with hypoxia alone in 3 cell lines. HIF-1α/2α were induced by radiation but combined hypoxia/radiation treatment did not yield a further increase. The dual responsive nature of HREs was maintained when spheroids were irradiated after delivery of HRE constructs in a replication-deficient adenovirus. Conclusions: Hypoxia-responsive enhancer element sequences are dually responsive to combined radiation and hypoxic treatment. Their use in genetic radiotherapy in vivo could maximize expression in the most radio-resistant population at the time of radiation and also exploit microenvironmental changes after radiotherapy to yield additional switch-on

  8. Adeno-associated viral vectors as agents for gene delivery : application in disorders and trauma of the central nervous system

    NARCIS (Netherlands)

    Ruitenberg, Marc J; Eggers, Ruben; Boer, Gerard J; Verhaagen, J.

    2002-01-01

    The use of viral vectors as agents for gene delivery provides a direct approach to manipulate gene expression in the mammalian central nervous system (CNS). The present article describes in detail the methodology for the injection of viral vectors, in particular adeno-associated virus (AAV) vectors,

  9. Generation of a Vero-Based Packaging Cell Line to Produce SV40 Gene Delivery Vectors for Use in Clinical Gene Therapy Studies

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    Miguel G. Toscano

    2017-09-01

    Full Text Available Replication-defective (RD recombinant simian virus 40 (SV40-based gene delivery vectors hold a great potential for clinical applications because of their presumed non-immunogenicity and capacity to induce immune tolerance to the transgene products in humans. However, the clinical use of SV40 vectors has been hampered by the lack of a packaging cell line that produces replication-competent (RC free SV40 particles in the vector production process. To solve this problem, we have adapted the current SV40 vector genome used for the production of vector particles and generated a novel Vero-based packaging cell line named SuperVero that exclusively expresses the SV40 large T antigen. SuperVero cells produce similar numbers of SV40 vector particles compared to the currently used packaging cell lines, albeit in the absence of contaminating RC SV40 particles. Our unique SV40 vector platform named SVac paves the way to clinically test a whole new generation of SV40-based therapeutics for a broad range of important diseases.

  10. Different Influences of Lipofection and Electrotransfection on In Vitro Gene Delivery to Primary Cultured Cortex Neurons.

    Science.gov (United States)

    Zhang, Xui-Si; Huang, Jing; Zhan, Cong-Qing; Chen, Jing; Li, Tao; Kaye, Alan D; Wu, Sheng-Xi; Xiao, Lan

    2016-03-01

    Many pain states are linked to central nervous system (CNS) diseases involving the dysfunction of dendritic arborization, making restoration a promising therapeutic strategy. Transfection of primary cortex neurons offers the possibility to study mechanisms which are important for the restoration of proper arborization. Its progress is, however, limited at present due to the lack of suitable gene transfer techniques. To obtain better insight into the transfection potential of currently used techniques, 2 non-viral transfection methods, lipofection and gene electrotransfer (GET), were compared. This is a comparison study performed on cultured cells. The transfection efficiency and neuronal viability, as well as the neuronal dendritic arborization after lipofection or GET, were compared. Primary cultured cortex neurons were transfected with the pEGFP-N1 plasmid, either using Lipofectamine 2000 (2, 3, or 4µL) or with electroporation, with our previously optimized protocol (200V/25 ms). Transfection efficiency and cell viability were inversely proportional for lipofection. The appropriate ratio of Lipofectamine and plasmid DNA provides optimal conditions for lipofection. Although GET offered higher transfection efficiency, it could not induce complex dendritic arborization, which made it unsuitable for in vitro gene transfer into cortex neurons. Limitations include species variability and translational applicability for CNS diseases and pain states related to potential toxicity. Based on these findings, lipofection might be advantageous for in vitro application to primary cultured cortex neurons. Pain states, stress mediated pathogenesis, and certain CNS diseases might potentially utilize this important technique in the future as a therapeutic modality.

  11. Transferrin receptor molecular imaging: targeting for diagnosis and monitoring of gene delivery

    International Nuclear Information System (INIS)

    Eun-Mi Kim; Hwan-Jeong Jeong; Jin-Hee Kim; Chang-Guhn Kim

    2004-01-01

    Objective: In this study, we investigated the targetability of Tf conjugated compounds to Tf-R expressed on cancer cells for detection and diagnosis and the usefulness of gamma probe-targeting delivery system on monitoring whether the gene complex bind to the cells specifically. Methods: For the detection and diagnosis of Tf-R positive cancer cells, Tf-chitosan conjugates were synthesized as previously described by Kircheis et al with some modifications. Succinimidyl 6-hydrazino nicotinate hydrochloride (HYNIC) was bound to Tf-chitosan conjugates. HYNIC-Tf-chitosan conjugates were labelled with 99mTc. In the monitoring of Tf-R specific gene delivery system, we used the HYNIC-Tf conjugated dendrimer. For tumor model, 5- to 6-week-old female BALB/c nude mice were injected subcutaneously in the left thigh with Ramos cells (human Burkitt's lymphoma). The gamma imagings were acquired after administration of 99mTc HYNIC-Tf conjugates and 99mTc HYNIC-Tf-DNA polyplexes via the tail vein of tumor bearing nude mice at 10, 30, 60, 90, and 120 min. To compare the image acquired with HYNIC-Tf conjugate, Ga-67 study was performed. To certify the expression of delivered gene via DNA polyplexes, 2 days after gene complex injection we inspected the expression of GFP in dissected tumor tissue. Results: Radiolabeling yields of both HYNIC-Tf conjugate and HYNIC-Tf-dendrimer gene complex were above 90% until 12hr. Uptake in the Ramos model of 99mTc HYNIC-Tf conjugate showed higher than those of Ga-67. A few minutes after injection 99mTc HYNIC-Tf conjugate localized mainly in the circulation (heart), kidneys, and tumor. At later times, radioactivity in tumor increased until 90 min. Pharmacokinetics of Ga-67 were different from those of 99mTc HYNIC-Tf conjugate. Tumor to nontumor ratio of Ga-67 was approximately 2 but in case of 99mTc HYNIC-Tf conjugate showed until 5. In Ramos lymphoma model, 99mTc HYNIC-Tf-DNA polyplexes accumulated the tumor site, and the gene expression of 99m

  12. AAV9-mediated central nervous system–targeted gene delivery via cisterna magna route in mice

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    Vera Lukashchuk

    2016-01-01

    Full Text Available Current barriers to the use of adeno-associated virus serotype 9 (AAV9 in clinical trials for treating neurological disorders are its high expression in many off-target tissues such as liver and heart, and lack of cell specificity within the central nervous system (CNS when using ubiquitous promoters such as human cytomegalovirus (CMV or chicken-β-actin hybrid (CAG. To enhance targeting the transgene expression in CNS cells, self-complementary (sc AAV9 vectors, scAAV9-GFP vectors carrying neuronal Hb9 and synapsin 1, and nonspecific CMV and CAG promoters were constructed. We demonstrate that synapsin 1 and Hb9 promoters exclusively targeted neurons in vitro, although their strengths were up to 10-fold lower than that of CMV. In vivo analyses of mouse tissue after scAAV9-GFP vector delivery via the cisterna magna revealed a significant advantage of synapsin 1 promoter over both Hb9 variants in targeting neurons throughout the brain, since Hb9 promoters were driving gene expression mainly within the motor-related areas of the brain stem. In summary, this study demonstrates that cisterna magna administration is a safe alternative to intracranial or intracerebroventricular vector delivery route using scAAV9, and introduces a novel utility of the Hb9 promoter for the targeted gene expression for both in vivo and in vitro applications.

  13. Biofunctionalized nanoparticles with pH-responsive and cell penetrating blocks for gene delivery

    International Nuclear Information System (INIS)

    Gaspar, V M; Marques, J G; Sousa, F; Queiroz, J A; Correia, I J; Louro, R O

    2013-01-01

    Bridging the gap between nanoparticulate delivery systems and translational gene therapy is a long sought after requirement in nanomedicine-based applications. However, recent developments regarding nanoparticle functionalization have brought forward the ability to synthesize materials with biofunctional moieties that mimic the evolved features of viral particles. Herein we report the versatile conjugation of both cell penetrating arginine and pH-responsive histidine moieties into the chitosan polymeric backbone, to improve the physicochemical characteristics of the native material. Amino acid coupling was confirmed by 2D TOCSY NMR and Fourier transform infrared spectroscopy. The synthesized chitosan–histidine–arginine (CH–H–R) polymer complexed plasmid DNA biopharmaceuticals, and spontaneously assembled into stable 105 nm nanoparticles with spherical morphology and positive surface charge. The functionalized delivery systems were efficiently internalized into the intracellular compartment, and exhibited remarkably higher transfection efficiency than unmodified chitosan without causing any cytotoxic effect. Additional findings regarding intracellular trafficking events reveal their preferential escape from degradative lysosomal pathways and nuclear localization. Overall, this assembly of nanocarriers with bioinspired moieties provides the foundations for the design of efficient and customizable materials for cancer gene therapy. (paper)

  14. Novel PVA-DNA nanoparticles prepared by ultra high pressure technology for gene delivery

    International Nuclear Information System (INIS)

    Kimura, Tsuyoshi; Okuno, Akira; Miyazaki, Kozo; Furuzono, Tsutomu; Ohya, Yuichi; Ouchi, Tatsuro; Mutsuo, Shingo; Yoshizawa, Hidekazu; Kitamura, Yoshiro; Fujisato, Toshiyta; Kishida, Akio

    2004-01-01

    Polyvinyl alcohol (PVA)-DNA nanoparticles have been developed by ultra high pressure (UHP) technology. Mixture solutions of DNA and PVA having various molecular weights (Mw) and degree of saponifications (DS) were treated under 10,000 atmospheres (981 MPa) condition at 40 deg. C for 10 min. Agarose gel electrophoresis and scanning electron microscope observation revealed that the PVA-DNA nanoparticles with average diameter of about 200 nm were formed. Using PVA of higher Mw and degree of saponifications, the amount of nanoparticles formed increased. The driving force of nanoparticle formation was the hydrogen bonding between DNA and PVA. In order to apply the PVA-DNA nanoparticles for gene delivery, the cytotoxicity and the cellular uptake of them were investigated using Raw264 cell lines. The cell viability was not influenced whether the presence of the PVA-DNA nanoparticles. Further, the nanoparticles internalized into cells were observed by fluorescent microscope. These results indicates that the PVA-DNA nanoparticles prepared by UHP technology showed be useful as drug carrier, especially for gene delivery

  15. Patient-controlled analgesia: therapeutic interventions using transdermal electro-activated and electro-modulated drug delivery.

    Science.gov (United States)

    Indermun, Sunaina; Choonara, Yahya E; Kumar, Pradeep; Du Toit, Lisa C; Modi, Girish; Luttge, Regina; Pillay, Viness

    2014-02-01

    Chronic pain poses a major concern to modern medicine and is frequently undertreated, causing suffering and disability. Patient-controlled analgesia, although successful, does have limitations. Transdermal delivery is the pivot to which analgesic research in drug delivery has centralized, especially with the confines of needle phobias and associated pain related to traditional injections, and the existing limitations associated with oral drug delivery. Highlighted within is the possibility of further developing transdermal drug delivery for chronic pain treatment using iontophoresis-based microneedle array patches. A concerted effort was made to review critically all available therapies designed for the treatment of chronic pain. The drug delivery systems developed for this purpose and nondrug routes are elaborated on, in a systematic manner. Recent developments and future goals in transdermal delivery as a means to overcome the individual limitations of the aforementioned delivery routes are represented as well. The approval of patch-like devices that contain both the microelectronic-processing mechanism and the active medicament in a small portable device is still awaited by the pharmaceutical industry. This anticipated platform may provide transdermal electro-activated and electro-modulated drug delivery systems a feasible attempt in chronic pain treatment. Iontophoresis has been proven an effective mode used to administer ionized drugs in physiotherapeutic, diagnostic, and dermatological applications and may be an encouraging probability for the development of devices and aids in the treatment of chronic pain. © 2013 Wiley Periodicals, Inc. and the American Pharmacists Association.

  16. Patient-controlled analgesia : therapeutic interventions using transdermal electro-activated and electro-modulated drug delivery

    NARCIS (Netherlands)

    Indermun, S.; Choonara, Y.E.; Kumar, P.; Du Toit, L.C.; Modi, G.; Luttge, R.; Pillay, V.

    2014-01-01

    Chronic pain poses a major concern to modern medicine and is frequently undertreated, causing suffering and disability. Patient-controlled analgesia, although successful, does have limitations. Transdermal delivery is the pivot to which analgesic research in drug delivery has centralized, especially

  17. Endocrine aspects of cancer gene therapy.

    Science.gov (United States)

    Barzon, Luisa; Boscaro, Marco; Palù, Giorgio

    2004-02-01

    The field of cancer gene therapy is in continuous expansion, and technology is quickly moving ahead as far as gene targeting and regulation of gene expression are concerned. This review focuses on the endocrine aspects of gene therapy, including the possibility to exploit hormone and hormone receptor functions for regulating therapeutic gene expression, the use of endocrine-specific genes as new therapeutic tools, the effects of viral vector delivery and transgene expression on the endocrine system, and the endocrine response to viral vector delivery. Present ethical concerns of gene therapy and the risk of germ cell transduction are also discussed, along with potential lines of innovation to improve cell and gene targeting.

  18. Cationic lipid-coated PEI/DNA polyplexes with improved efficiency and reduced cytotoxicity for gene delivery into mesenchymal stem cells

    Directory of Open Access Journals (Sweden)

    Song HM

    2012-08-01

    Full Text Available Hongmei Song, Gang Wang, Bin He, Li Li, Caixia Li, Yusi Lai, Xianghui Xu, Zhongwei GuNational Engineering Research Center for Biomaterials, Sichuan University, Chengdu, Sichuan, People's Republic of ChinaBackground: Effective gene transfection without serum deprivation is a prerequisite for successful stem cell-based gene therapy. Polyethylenimine (PEI is an efficient nonviral gene vector, but its application has been hindered by serum sensitivity and severe cytotoxicity.Methods: To solve this problem, a new family of lipopolyplexes was developed by coating PEI/DNA polyplexes with three serum-resistant cationic lipids, namely, lysinylated, histidylated, and arginylated cholesterol. The physical properties, transfection efficiency, cellular uptake, subcellular distribution, and cytotoxicity of the lipopolyplexes was investigated.Results: The outer coat composed of lysinylated or histidylated cholesterol remarkably improved the transfection efficiency of the polyplex with a low PEI/DNA ratio of 2 in the presence of serum. The resulting lysinylated and histidylated cholesterol lipopolyplexes were even more efficient than the best performing polyplex with a high PEI/DNA ratio of 10. Results from cellular uptake and subcellular distribution studies suggest that their higher transfection efficiency may result from accelerated DNA nuclear localization. The superiority of the lipopolyplexes over the best performing polyplex was also confirmed by delivering the therapeutic gene, hVEGF165. Equally importantly, the lipid coating removed the necessity of introducing excess free PEI chains into the transfection solution for higher efficiency, generating lipopolyplexes with no signs of cytotoxicity.Conclusion: Noncovalent modification of polyplexes with lysinylated and histidylated cholesterol lipids can simultaneously improve efficiency and reduce the toxicity of gene delivery under serum conditions, showing great promise for genetic modification of bone

  19. Safe and stable noninvasive focal gene delivery to the mammalian brain following focused ultrasound.

    Science.gov (United States)

    Stavarache, Mihaela A; Petersen, Nicholas; Jurgens, Eric M; Milstein, Elizabeth R; Rosenfeld, Zachary B; Ballon, Douglas J; Kaplitt, Michael G

    2018-04-27

    OBJECTIVE Surgical infusion of gene therapy vectors has provided opportunities for biological manipulation of specific brain circuits in both animal models and human patients. Transient focal opening of the blood-brain barrier (BBB) by MR-guided focused ultrasound (MRgFUS) raises the possibility of noninvasive CNS gene therapy to target precise brain regions. However, variable efficiency and short follow-up of studies to date, along with recent suggestions of the potential for immune reactions following MRgFUS BBB disruption, all raise questions regarding the viability of this approach for clinical translation. The objective of the current study was to evaluate the efficiency, safety, and long-term stability of MRgFUS-mediated noninvasive gene therapy in the mammalian brain. METHODS Focused ultrasound under the control of MRI, in combination with microbubbles consisting of albumin-coated gas microspheres, was applied to rat striatum, followed by intravenous infusion of an adeno-associated virus serotype 1/2 (AAV1/2) vector expressing green fluorescent protein (GFP) as a marker. Following recovery, animals were followed from several hours up to 15 months. Immunostaining for GFP quantified transduction efficiency and stability of expression. Quantification of neuronal markers was used to determine histological safety over time, while inflammatory markers were examined for evidence of immune responses. RESULTS Transitory disruption of the BBB by MRgFUS resulted in efficient delivery of the AAV1/2 vector to the targeted rodent striatum, with 50%-75% of striatal neurons transduced on average. GFP transgene expression appeared to be stable over extended periods of time, from 2 weeks to 6 months, with evidence of ongoing stable expression as long as 16 months in a smaller cohort of animals. No evidence of substantial toxicity, tissue injury, or neuronal loss was observed. While transient inflammation from BBB disruption alone was noted for the first few days, consistent

  20. Direct Head-To-Head Comparison of Cationic Liposome-Mediated Gene Delivery to Mesenchymal Stem/Stromal Cells of Different Human Sources: A Comprehensive Study

    Science.gov (United States)

    Boura, Joana S.; dos Santos, Francisco; Gimble, Jeffrey M.; Cardoso, Carla M.P.; Madeira, Catarina; Cabral, Joaquim M.S.

    2013-01-01

    Abstract Nonviral gene delivery to human mesenchymal stem/stromal cells (MSC) can be considered a very promising strategy to improve their intrinsic features, amplifying the therapeutic potential of these cells for clinical applications. In this work, we performed a comprehensive comparison of liposome-mediated gene transfer efficiencies to MSC derived from different human sources—bone marrow (BM MSC), adipose tissue-derived cells (ASC), and umbilical cord matrix (UCM MSC). The results obtained using a green fluorescent protein (GFP)-encoding plasmid indicated that MSC isolated from BM and UCM are more amenable to genetic modification when compared to ASC as they exhibited superior levels of viable, GFP+ cells 48 hr post-transfection, 58±7.1% and 54±3.8%, respectively, versus 33±4.7%. For all cell sources, high cell recoveries (≈50%) and viabilities (>85%) were achieved, and the transgene expression was maintained for 10 days. Levels of plasmid DNA uptake, as well as kinetics of transgene expression and cellular division, were also determined. Importantly, modified cells were found to retain their characteristic immunophenotypic profile and multilineage differentiation capacity. By using the lipofection protocol optimized herein, we were able to maximize transfection efficiencies to human MSC (maximum of 74% total GFP+ cells) and show that lipofection is a promising transfection strategy for MSC genetic modification, especially when a transient expression of a therapeutic gene is required. Importantly, we also clearly demonstrated that intrinsic features of MSC from different sources should be taken into consideration when developing and optimizing strategies for MSC engineering with a therapeutic gene. PMID:23360350

  1. Combined anti-tumor therapeutic effect of targeted gene, hyperthermia, radionuclide brachytherapy in breast carcinoma

    International Nuclear Information System (INIS)

    Chen Daozhen; Tang Qiusha; Xiang Jingying; Xu Fei; Zhang Li; Wang Junfeng

    2011-01-01

    Objective: To investigate the antitumor therapeutic effect of combined therapy of magnetic induction heating by nano-magnetic particles, herpes simplex virus thymidine kinase gene (HSV-tk suicide gene) and internal radiation in mice bearing MCF-7 breast carcinoma. Methods: The transfection reagents, plasmids heat shock protein-HSV-tk (pHSP-HSV-tk), ferroso-ferric oxide nano-magnetic fluid flow and 188 Re-ganciclovir-bovine serum albumin-nanopaticles (GCV-BSA-NP) were prepared. The heating experiments in vivo were carried out using ferroso-ferric oxide nano-magnetic fluid flow. Sixty mice tumor models bearing MCF-7 breast carcinoma were established and randomly divided into six groups. Group A was the control group, B was gene transfection therapy group, C was hyperthermia group, D was gene transfection therapy combined with radionuclide brachytherapy group, E was gene therapy combined with hyperthermia group, and F was gene therapy, hyperthermia combined with radionuclide brachytherapy group. The tumor growth, tumor mass and histopathological changes were evaluated. The expression of HSV-tk in the groups of B, D, E and F was detected by RT-PCR. Poisson distribution and one-way analysis of variance (ANOVA) were used for statistical analysis by SPSS 10.0 software. Results: In the animal heating experiments, the temperature of tumor increased up to 39.6 degree C, 43.2 degree C, and 48.1 degree C quickly with different injected doses (2, 4 and 6 mg respectively) of nano-magnetic particles and maintained for 40 min. The temperature of tumor tissue reduced to 36.8 degree C, 37.5 degree C and 37.8 degree C in 10 min when alternating magnetic field (AMF) stopped. The tumor mass in Groups C ((452.50±30.29) mg), D ((240.98±35.32)mg), E((231.87±27.41) mg) and F ((141.55±23.78) mg) were much lower than that in Group A ((719.12±22.65) mg) (F=800.07, P<0.01), with the most significant treatment effect in Group F.The tumor mass in Group B((684.05±24.02) mg) was higher than

  2. Myostatin propeptide gene delivery by gene gun ameliorates muscle atrophy in a rat model of botulinum toxin-induced nerve denervation.

    Science.gov (United States)

    Tsai, Sen-Wei; Tung, Yu-Tang; Chen, Hsiao-Ling; Yang, Shang-Hsun; Liu, Chia-Yi; Lu, Michelle; Pai, Hui-Jing; Lin, Chi-Chen; Chen, Chuan-Mu

    2016-02-01

    Muscle atrophy is a common symptom after nerve denervation. Myostatin propeptide, a precursor of myostatin, has been documented to improve muscle growth. However, the mechanism underlying the muscle atrophy attenuation effects of myostatin propeptide in muscles and the changes in gene expression are not well established. We investigated the possible underlying mechanisms associated with myostatin propeptide gene delivery by gene gun in a rat denervation muscle atrophy model, and evaluated gene expression patterns. In a rat botulinum toxin-induced nerve denervation muscle atrophy model, we evaluated the effects of wild-type (MSPP) and mutant-type (MSPPD75A) of myostatin propeptide gene delivery, and observed changes in gene activation associated with the neuromuscular junction, muscle and nerve. Muscle mass and muscle fiber size was moderately increased in myostatin propeptide treated muscles (pmyostatin propeptide gene delivery, especially the mutant-type of MSPPD75A, attenuates muscle atrophy through myogenic regulatory factors and acetylcholine receptor regulation. Our data concluded that myostatin propeptide gene therapy may be a promising treatment for nerve denervation induced muscle atrophy. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. Selective Gene Delivery for Integrating Exogenous DNA into Plastid and Mitochondrial Genomes Using Peptide-DNA Complexes.

    Science.gov (United States)

    Yoshizumi, Takeshi; Oikawa, Kazusato; Chuah, Jo-Ann; Kodama, Yutaka; Numata, Keiji

    2018-05-14

    Selective gene delivery into organellar genomes (mitochondrial and plastid genomes) has been limited because of a lack of appropriate platform technology, even though these organelles are essential for metabolite and energy production. Techniques for selective organellar modification are needed to functionally improve organelles and produce transplastomic/transmitochondrial plants. However, no method for mitochondrial genome modification has yet been established for multicellular organisms including plants. Likewise, modification of plastid genomes has been limited to a few plant species and algae. In the present study, we developed ionic complexes of fusion peptides containing organellar targeting signal and plasmid DNA for selective delivery of exogenous DNA into the plastid and mitochondrial genomes of intact plants. This is the first report of exogenous DNA being integrated into the mitochondrial genomes of not only plants, but also multicellular organisms in general. This fusion peptide-mediated gene delivery system is a breakthrough platform for both plant organellar biotechnology and gene therapy for mitochondrial diseases in animals.

  4. Chitosan-Graft-Polyethylenimine/DNA Nanoparticles as Novel Non-Viral Gene Delivery Vectors Targeting Osteoarthritis

    Science.gov (United States)

    Lv, Lulu; Zhao, Huiqing

    2014-01-01

    The development of safe and efficient gene carriers is the key to the clinical success of gene therapy. The present study was designed to develop and evaluate the chitosan-graft-polyethylenimine (CP)/DNA nanoparticles as novel non-viral gene vectors for gene therapy of osteoarthritis. The CP/DNA nanoparticles were produced through a complex coacervation of the cationic polymers with pEGFP after grafting chitosan (CS) with a low molecular weight (Mw) PEI (Mw = 1.8 kDa). Particle size and zeta potential were related to the weight ratio of CP:DNA, where decreases in nanoparticle size and increases in surface charge were observed as CP content increased. The buffering capacity of CP was significantly greater than that of CS. The transfection efficiency of CP/DNA nanoparticles was similar with that of the Lipofectamine™ 2000, and significantly higher than that of CS/DNA and PEI (25 kDa)/DNA nanoparticles. The transfection efficiency of the CP/DNA nanoparticles was dependent on the weight ratio of CP:DNA (w/w). The average cell viability after the treatment with CP/DNA nanoparticles was over 90% in both chondrocytes and synoviocytes, which was much higher than that of PEI (25 kDa)/DNA nanoparticles. The CP copolymers efficiently carried the pDNA inside chondrocytes and synoviocytes, and the pDNA was detected entering into nucleus. These results suggest that CP/DNA nanoparticles with improved transfection efficiency and low cytotoxicity might be a safe and efficient non-viral vector for gene delivery to both chondrocytes and synoviocytes. PMID:24392152

  5. Bioreducible liposomes for gene delivery: from the formulation to the mechanism of action.

    Directory of Open Access Journals (Sweden)

    Gabriele Candiani

    Full Text Available BACKGROUND: A promising strategy to create stimuli-responsive gene delivery systems is to exploit the redox gradient between the oxidizing extracellular milieu and the reducing cytoplasm in order to disassemble DNA/cationic lipid complexes (lipoplexes. On these premises, we previously described the synthesis of SS14 redox-sensitive gemini surfactant for gene delivery. Although others have attributed the beneficial effects of intracellular reducing environment to reduced glutathione (GSH, these observations cannot rule out the possible implication of the redox milieu in its whole on transfection efficiency of bioreducible transfectants leaving the determinants of DNA release largely undefined. METHODOLOGY/PRINCIPAL FINDINGS: With the aim of addressing this issue, SS14 was here formulated into binary and ternary 100 nm-extruded liposomes and the effects of the helper lipid composition and of the SS14/helper lipids molar ratio on chemical-physical and structural parameters defining transfection effectiveness were investigated. Among all formulations tested, DOPC/DOPE/SS14 at 25:50:25 molar ratio was the most effective in transfection studies owing to the presence of dioleoyl chains and phosphatidylethanolamine head groups in co-lipids. The increase in SS14 content up to 50% along DOPC/DOPE/SS14 liposome series yielded enhanced transfection, up to 2.7-fold higher than that of the benchmark Lipofectamine 2000, without altering cytotoxicity of the corresponding lipoplexes at charge ratio 5. Secondly, we specifically investigated the redox-dependent mechanisms of gene delivery into cells through tailored protocols of transfection in GSH-depleted and repleted vs. increased oxidative stress conditions. Importantly, GSH specifically induced DNA release in batch and in vitro. CONCLUSIONS/SIGNIFICANCE: The presence of helper lipids carrying unsaturated dioleoyl chains and phosphatidylethanolamine head groups significantly improved transfection efficiencies

  6. A theranostic prodrug delivery system based on Pt(IV) conjugated nano-graphene oxide with synergistic effect to enhance the therapeutic efficacy of Pt drug.

    Science.gov (United States)

    Li, Jingwen; Lyv, Zhonglin; Li, Yanli; Liu, Huan; Wang, Jinkui; Zhan, Wenjun; Chen, Hong; Chen, Huabing; Li, Xinming

    2015-05-01

    Due to their high NIR-optical absorption and high specific surface area, graphene oxide and graphene oxide-based nanocomposites have great potential in both drug delivery and photothermal therapy. In the work reported herein we successfully integrate a Pt(IV) complex (c,c,t-[Pt(NH3)2Cl2(OH)2]), PEGylated nano-graphene oxide (PEG-NGO), and a cell apoptosis sensor into a single platform to generate a multifunctional nanocomposite (PEG-NGO-Pt) which shows potential for targeted drug delivery and combined photothermal-chemotherapy under near infrared laser irradiation (NIR), and real-time monitoring of its therapeutic efficacy. Non-invasive imaging using a fluorescent probe immobilized on the GO shows an enhanced therapeutic effect of PEG-NGO-Pt in cancer treatment via apoptosis and cell death. Due to the enhanced cytotoxicity of cisplatin and the highly specific tumor targeting of PEG-NGO-Pt at elevated temperatures, this nanocomposite displays a synergistic effect in improving the therapeutic efficacy of the Pt drug with complete destruction of tumors, no tumor recurrence and minimal systemic toxicity in comparison with chemotherapy or photothermal treatment alone, highlighting the advantageous effects of integrating Pt(IV) with GO for anticancer treatment. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. Efficient gene delivery to human umbilical cord mesenchymal stem cells by cationized Porphyra yezoensis polysaccharide nanoparticles.

    Science.gov (United States)

    Yu, Qingtong; Cao, Jin; Chen, Baoding; Deng, Wenwen; Cao, Xia; Chen, Jingjing; Wang, Yan; Wang, Shicheng; Yu, Jiangnan; Xu, Ximing; Gao, Xiangdong

    2015-01-01

    This study centered on an innovative application of Porphyra yezoensis polysaccharide (PPS) with cationic modification as a safe and efficient nonviral gene vector to deliver a plasmid encoding human Wnt3a (pWnt3a) into human umbilical cord mesenchymal stem cells (HUMSCs). After modification with branched low-molecular-weight (1,200 Da) polyethylenimine, the cationized PPS (CPPS) was combined with pWnt3a to form spherical nanoscale particles (CPPS-pWnt3a nanoparticles). Particle size and distribution indicated that the CPPS-pWnt3a nanoparticles at a CPPS:pWnt3a weight ratio of 40:1 might be a potential candidate for DNA plasmid transfection. A cytotoxicity assay demonstrated that the nanoparticles prepared at a CPPS:pWnt3a weight ratio of 40:1 were nontoxic to HUMSCs compared to those of Lipofectamine 2000 and polyethylenimine (25 kDa). These nanoparticles were further transfected to HUMSCs. Western blotting demonstrated that the nanoparticles (CPPS:pWnt3a weight ratio 40:1) had the greatest transfection efficiency in HUMSCs, which was significantly higher than that of Lipofectamine 2000; however, when the CPPS:pWnt3a weight ratio was increased to 80:1, the nanoparticle-treated group showed no obvious improvement in translation efficiency over Lipofectamine 2000. Therefore, CPPS, a novel cationic polysaccharide derived from P. yezoensis, could be developed into a safe, efficient, nonviral gene vector in a gene-delivery system.

  8. One-Pot Parallel Synthesis of Lipid Library via Thiolactone Ring Opening and Screening for Gene Delivery.

    Science.gov (United States)

    Molla, Mijanur R; Böser, Alexander; Rana, Akshita; Schwarz, Karina; Levkin, Pavel A

    2018-04-18

    Efficient delivery of nucleic acids into cells is of great interest in the field of cell biology and gene therapy. Despite a lot of research, transfection efficiency and structural diversity of gene-delivery vectors are still limited. A better understanding of the structure-function relationship of gene delivery vectors is also essential for the design of novel and intelligent delivery vectors, efficient in "difficult-to-transfect" cells and in vivo clinical applications. Most of the existing strategies for the synthesis of gene-delivery vectors require multiple steps and lengthy procedures. Here, we demonstrate a facile, three-component one-pot synthesis of a combinatorial library of 288 structurally diverse lipid-like molecules termed "lipidoids" via a thiolactone ring opening reaction. This strategy introduces the possibility to synthesize lipidoids with hydrophobic tails containing both unsaturated bonds and reducible disulfide groups. The whole synthesis and purification are convenient, extremely fast, and can be accomplished within a few hours. Screening of the produced lipidoids using HEK293T cells without addition of helper lipids resulted in identification of highly stable liposomes demonstrating ∼95% transfection efficiency with low toxicity.

  9. A new optimized formulation of cationic solid lipid nanoparticles intended for gene delivery: development, characterization and DNA binding efficiency of TCERG1 expression plasmid.

    Science.gov (United States)

    Fàbregas, Anna; Sánchez-Hernández, Noemí; Ticó, Josep Ramon; García-Montoya, Encarna; Pérez-Lozano, Pilar; Suñé-Negre, Josep M; Hernández-Munain, Cristina; Suñé, Carlos; Miñarro, Montserrat

    2014-10-01

    Solid lipid nanoparticles (SLNs) are being considered as a new approach for therapeutics for many known diseases. In addition to drug delivery, their use as non-viral vectors for gene delivery can be achieved by the inclusion of cationic lipids, which provide a positive surface potential that favours binding to the DNA backbone. This work is based on the idea that the optimization of the components is required as the first step in simplifying the qualitative and quantitative composition of SLNs as much as possible without affecting the essential properties that define SLNs as optimal non-viral vectors for gene delivery. We selected the best lipids and surfactants in terms of particle size and zeta potential and characterized the properties of the resulting nanoparticles using X-ray photoelectron spectroscopy (XPS) and atomic force microscopy (AFM). The SLNs had a particle size of approximately 120 nm and a positive surface charge of 42 mV. In addition, we analysed the main physicochemical characteristics of the bulk components of the nanoparticles using X-ray diffraction (XRD), differential scanning calorimetry (DSC) and mass spectrometry (MS). The suitability of the optimized SLNs for DNA binding was evaluated after the lyophilisation process using a carboxyl-terminal region of the TCERG1 gene, a human factor that has been implicated in several diseases. We show that the SLNs presented high efficiency in the binding of DNA, and importantly, they presented no toxicity when assayed in an in vivo system. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. Gene delivery by microfluidic flow-through electroporation based on constant DC and AC field.

    Science.gov (United States)

    Geng, Tao; Zhan, Yihong; Lu, Chang

    2012-01-01

    Electroporation is one of the most widely used physical methods to deliver exogenous nucleic acids into cells with high efficiency and low toxicity. Conventional electroporation systems typically require expensive pulse generators to provide short electrical pulses at high voltage. In this work, we demonstrate a flow-through electroporation method for continuous transfection of cells based on disposable chips, a syringe pump, and a low-cost power supply that provides a constant voltage. We successfully transfect cells using either DC or AC voltage with high flow rates (ranging from 40 µl/min to 20 ml/min) and high efficiency (up to 75%). We also enable the entire cell membrane to be uniformly permeabilized and dramatically improve gene delivery by inducing complex migrations of cells during the flow.

  11. Peptide-conjugated micelles as a targeting nanocarrier for gene delivery

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Wen Jen, E-mail: wjlin@ntu.edu.tw; Chien, Wei Hsuan [National Taiwan University, School of Pharmacy, Graduate Institute of Pharmaceutical Sciences (China)

    2015-09-15

    The aim of this study was to develop peptide-conjugated micelles possessing epidermal growth factor receptor (EGFR) targeting ability for gene delivery. A sequence-modified dodecylpeptide, GE11(2R), with enhancing EGF receptor binding affinity, was applied in this study as a targeting ligand. The active targeting micelles were composed of poly(d,l-lactide-co-glycolide)-poly(ethylene glycol) (PLGA-PEG) copolymer conjugated with GE11(2R)-peptide. The particle sizes of peptide-free and peptide-conjugated micelles were 277.0 ± 5.1 and 308.7 ± 14.5 nm, respectively. The peptide-conjugated micelles demonstrated the cellular uptake significantly higher than peptide-free micelles in EGFR high-expressed MDA-MB-231 and MDA-MB-468 cells due to GE11(2R)-peptide specificity. Furthermore, the peptide-conjugated micelles were able to encapsulate plasmid DNA and expressed cellular transfection higher than peptide-free micelles in EGFR high-expressed cells. The EGFR-targeting delivery micelles enhanced DNA internalized into cells and achieved higher cellular transfection in EGFR high-expressed cells.

  12. The Increasing Complexity of the Oncofetal H19 Gene Locus: Functional Dissection and Therapeutic Intervention

    Directory of Open Access Journals (Sweden)

    Abraham Hochberg

    2013-02-01

    Full Text Available The field of the long non-coding RNA (lncRNA is advancing rapidly. Currently, it is one of the most popular fields in the biological and medical sciences. It is becoming increasingly obvious that the majority of the human transcriptome has little or no-protein coding capacity. Historically, H19 was the first imprinted non-coding RNA (ncRNA transcript identified, and the H19/IGF2 locus has served as a paradigm for the study of genomic imprinting since its discovery. In recent years, we have extensively investigated the expression of the H19 gene in a number of human cancers and explored the role of H19 RNA in tumor development. Here, we discuss recently published data from our group and others that provide further support for a central role of H19 RNA in the process of tumorigenesis. Furthermore, we focus on major transcriptional modulators of the H19 gene and discuss them in the context of the tumor-promoting activity of the H19 RNA. Based on the pivotal role of the H19 gene in human cancers, we have developed a DNA-based therapeutic approach for the treatment of cancers that have upregulated levels of H19 expression. This approach uses a diphtheria toxin A (DTA protein expressed under the regulation of the H19 promoter to treat tumors with significant expression of H19 RNA. In this review, we discuss the treatment of four cancer indications in human subjects using this approach, which is currently under development. This represents perhaps one of the very few examples of an existing DNA-based therapy centered on an lncRNA system. Apart from cancer, H19 expression has been reported also in other conditions, syndromes and diseases, where deregulated imprinting at the H19 locus was obvious in some cases and will be summarized below. Moreover, the H19 locus proved to be much more complicated than initially thought. It houses a genomic sequence that can transcribe, yielding various transcriptional outputs, both in sense and antisense directions. The

  13. Nonviral gene-delivery by highly fluorinated gemini bispyridinium surfactant-based DNA nanoparticles.

    Science.gov (United States)

    Fisicaro, Emilia; Compari, Carlotta; Bacciottini, Franco; Contardi, Laura; Pongiluppi, Erika; Barbero, Nadia; Viscardi, Guido; Quagliotto, Pierluigi; Donofrio, Gaetano; Krafft, Marie Pierre

    2017-02-01

    Biological and thermodynamic properties of a new homologous series of highly fluorinated bispyridinium cationic gemini surfactants, differing in the length of the spacer bridging the pyridinium polar heads in 1,1' position, are reported for the first time. Interestingly, gene delivery ability is closely associated with the spacer length due to a structural change of the molecule in solution. This conformation change is allowed when the spacer reaches the right length, and it is suggested by the trends of the apparent and partial molar enthalpies vs molality. To assess the compounds' biological activity, they were tested with an agarose gel electrophoresis mobility shift assay (EMSA), MTT proliferation assay and Transient Transfection assays on a human rhabdomyosarcoma cell line. Data from atomic force microscopy (AFM) allow for morphological characterization of DNA nanoparticles. Dilution enthalpies, measured at 298K, enabled the determination of apparent and partial molar enthalpies vs molality. All tested compounds (except that with the longest spacer), at different levels, can deliver the plasmid when co-formulated with 1,2-dioleyl-sn-glycero-3-phosphoethanolamine (DOPE). The compound with a spacer formed by eight carbon atoms gives rise to a gene delivery ability that is comparable to that of the commercial reagent. The compound with the longest spacer compacts DNA in loosely condensed structures by forming bows, which are not suitable for transfection. Regarding the compounds' hydrogenated counterparts, the tight relationship between the solution thermodynamics data and their biological performance is amazing, making "old" methods the foundation to deeply understanding "new" applications. Copyright © 2016 Elsevier Inc. All rights reserved.

  14. Mesoporous Silica Nanomaterials for Applications in Catalysis, Sensing, Drug Delivery and Gene Transfection

    Energy Technology Data Exchange (ETDEWEB)

    Radu, Daniela Rodica [Iowa State Univ., Ames, IA (United States)

    2004-01-01

    The central theme of this dissertation is represented by the versatility of mesoporous silica nanomaterials in various applications such as catalysis and bio-applications, with main focus on biological applications of Mesoporous Silica Nanospheres (MSN). The metamorphosis that we impose to these materials from catalysis to sensing and to drug and gene delivery is detailed in this dissertation. First, we developed a synthetic method that can fine tune the amount of chemically accessible organic functional groups on the pores surface of MSN by exploiting electrostatic and size matching between the cationic alkylammonium head group of the cetyltrimethylammonium bromide (CTAB) surfactant and various anionic organoalkoxysilane precursors at the micelle-water interface in a base-catalyzed condensation reaction of silicate. Aiming nature imitation, we demonstrated the catalytic abilities of the MSNs, We utilized an ethylenediamine functional group for chelating Cu2+ as a catalytic functional group anchored inside the mesopores. Thus, a polyalkynylene-based conducting polymer (molecular wire) was synthesized within the Cu-functionalized MSNs silica catalyst. For sensing applications, we have synthesized a poly(lactic acid) coated mesoporous silica nanosphere (PLA-MSN) material that serves as a fluorescence sensor system for detection of amino-containing neurotransmitters in neutral aqueous buffer. We exploited the mesoporosity of MSNs for encapsulating pharmaceutical drugs. We examined bio-friendly capping molecules such as polyamidoamine dendrimers of generations G2 to G4, to prevent the drug leaching. Next, the drug delivery system employed MSNs loaded with Doxorubicin, an anticancer drug. The results demonstrated that these nano-Trojan horses have ability to deliver Doxorubicin to cancer cells and induce their death. Finally, to demonstrate the potential of MSN as an universal cellular transmembrane nanovehicle, we anchored positively charged dendrimers on

  15. Kinase Gene Expression Profiling of Metastatic Clear Cell Renal Cell Carcinoma Tissue Identifies Potential New Therapeutic Targets.

    Directory of Open Access Journals (Sweden)

    Pooja Ghatalia

    Full Text Available Kinases are therapeutically actionable targets. Kinase inhibitors targeting vascular endothelial growth factor receptors (VEGFR and mammalian target of rapamycin (mTOR improve outcomes in metastatic clear cell renal cell carcinoma (ccRCC, but are not curative. Metastatic tumor tissue has not been comprehensively studied for kinase gene expression. Paired intra-patient kinase gene expression analysis in primary tumor (T, matched normal kidney (N and metastatic tumor tissue (M may assist in identifying drivers of metastasis and prioritizing therapeutic targets. We compared the expression of 519 kinase genes using NanoString in T, N and M in 35 patients to discover genes over-expressed in M compared to T and N tissue. RNA-seq data derived from ccRCC tumors in The Cancer Genome Atlas (TCGA were used to demonstrate differential expression of genes in primary tumor tissue from patients that had metastasis at baseline (n = 79 compared to those that did not develop metastasis for at least 2 years (n = 187. Functional analysis was conducted to identify key signaling pathways by using Ingenuity Pathway Analysis. Of 10 kinase genes overexpressed in metastases compared to primary tumor in the discovery cohort, 9 genes were also differentially expressed in TCGA primary tumors with metastasis at baseline compared to primary tumors without metastasis for at least 2 years: EPHB2, AURKA, GSG2, IKBKE, MELK, CSK, CHEK2, CDC7 and MAP3K8; p<0.001. The top pathways overexpressed in M tissue were pyridoxal 5'-phosphate salvage, salvage pathways of pyrimidine ribonucleotides, NF-kB signaling, NGF signaling and cell cycle control of chromosomal replication. The 9 kinase genes validated to be over-expressed in metastatic ccRCC may represent currently unrecognized but potentially actionable therapeutic targets that warrant functional validation.

  16. Mannosylated Chitosan Nanoparticles Based Macrophage-Targeting Gene Delivery System Enhanced Cellular Uptake and Improved Transfection Efficiency.

    Science.gov (United States)

    Peng, Yixing; Yao, Wenjun; Wang, Bo; Zong, Li

    2015-04-01

    Gene transfer mediated by mannosylated chitosan (MCS) is a safe and promising approach for gene and vaccine delivery. MCS nanoparticles based gene delivery system showed high in vivo delivery efficiency and elicited strong immune responses in mice. However, little knowledge about the cell binding, transfection efficiency and intracellular trafficking of MCS nanoparticles had been acquired. In this study, using gastrin-releasing peptide as a model plasmid (pGRP), the binding of MCS/pGRP nanoparticles to macrophages and the intracellular trafficking of MCS/pGRP nanoparticles in macrophages were investigated. MCS-mediated transfection efficiency in macrophages was also evaluated using pGL-3 as a reporter gene. The results showed that the binding and transfection efficiency of MCS nanoparticles in macrophages was higher than that of CS, which was attributed to the interaction between mannose ligands in MCS and mannose receptors on the surface of macrophages. Observation with a confocal laser scanning microscope indicated the cellular uptake of MCS/pGRP nanoparticles were more than that of CS/pGRP nanoparticles in macrophages. MCS/pGRP nanoparticles were taken up by macrophages and most of them were entrapped in endosomal/lysosomal compartments. After the nanoparticles escaping from endosomal/lysosomal compartments, naked pGRP entered the nucleus, and a few MCS might enter the nucleus in terms of nanoparticles. Overall, MCS has the potential to be an excellent macrophage-targeting gene delivery carrier.

  17. Virus Delivery of CRISPR Guides to the Murine Prostate for Gene Alteration.

    Science.gov (United States)

    Riedel, Maria; Berthelsen, Martin F; Bakiri, Latifa; Wagner, Erwin F; Thomsen, Martin K

    2018-04-27

    With an increasing incidence of prostate cancer, identification of new tumor drivers or modulators is crucial. Genetically engineered mouse models (GEMM) for prostate cancer are hampered by tumor heterogeneity and its complex microevolution dynamics. Traditional prostate cancer mouse models include, amongst others, germline and conditional knockouts, transgenic expression of oncogenes, and xenograft models. Generation of de novo mutations in these models is complex, time-consuming, and costly. In addition, most of traditional models target the majority of the prostate epithelium, whereas human prostate cancer is well known to evolve as an isolated event in only a small subset of cells. Valuable models need to simulate not only prostate cancer initiation, but also progression to advanced disease. Here we describe a method to target a few cells in the prostate epithelium by transducing cells by viral particles. The delivery of an engineered virus to the murine prostate allows alteration of gene expression in the prostate epithelia. Virus type and quantity will hereby define the number of targeted cells for gene alteration by transducing a few cells for cancer initiation and many cells for gene therapy. Through surgery-based injection in the anterior lobe, distal from the urinary track, the tumor in this model can expand without impairing the urinary function of the animal. Furthermore, by targeting only a subset of prostate epithelial cells the technique enables clonal expansion of the tumor, and therefore mimics human tumor initiation, progression, as well as invasion through the basal membrane. This novel technique provides a powerful prostate cancer model with improved physiological relevance. Animal suffering is limited, and since no additional breeding is required, overall animal count is reduced. At the same time, analysis of new candidate genes and pathways is accelerated, which in turn is more cost efficient.

  18. Structure-function correlation of chloroquine and analogues as transgene expression enhancers in nonviral gene delivery.

    Science.gov (United States)

    Cheng, Jianjun; Zeidan, Ryan; Mishra, Swaroop; Liu, Aijie; Pun, Suzie H; Kulkarni, Rajan P; Jensen, Gregory S; Bellocq, Nathalie C; Davis, Mark E

    2006-11-02

    To understand how chloroquine (CQ) enhances transgene expression in polycation-based, nonviral gene delivery systems, a number of CQ analogues with variations in the aliphatic amino side chain or in the aromatic ring are synthesized and investigated. Our studies indicate that the aliphatic amino moiety of CQ is essential to provide increased gene expression. Further, the enhancements are more dramatically affected by changes to the aromatic ring and are positively correlated to the strength of intercalation between DNA and the CQ analogues. Quinacrine (QC), a CQ analogue with a fused acridinyl structure that can strongly intercalate DNA, enhances transfection similarly to CQ at a concentration 10 times lower, while N(4)-(4-pyridinyl)-N(1),N(1)-diethyl-1,4-pentanediamine (CP), a CQ analogue that has a weakly intercalating pyridinyl ring, shows no effect on gene expression. Subtle change on the 7-substituent of the chloroquine aromatic structure can also greatly affect the ability of the CQ analogues to enhance transgene expression. Transfection in the presence of N(4)-(7-trifluoromethyl-4-quinolinyl)-N(1),N(1)-diethyl-1,4-pentanediamin e (CQ7a) shows expression efficiency 10 times higher than in the presence of CQ at same concentration, while transfection in the presence of N(4)-(4-quinolinyl)-N(1),N(1)-diethyl-1,4-pentanediamine (CQ7b) does not reveal any enhancing effects on expression. Through a number of comparative studies with CQ and its analogues, we conclude that there are at least three mechanistic features of CQ that lead to the enhancement in gene expression: (i) pH buffering in endocytic vesicles, (ii) displacement of polycations from the nucleic acids in polyplexes, and (iii) alteration of the biophysical properties of the released nucleic acid.

  19. Identification of genes highly downregulated in pancreatic cancer through a meta-analysis of microarray datasets: implications for discovery of novel tumor-suppressor genes and therapeutic targets.

    Science.gov (United States)

    Goonesekere, Nalin C W; Andersen, Wyatt; Smith, Alex; Wang, Xiaosheng

    2018-02-01

    The lack of specific symptoms at early tumor stages, together with a high biological aggressiveness of the tumor contribute to the high mortality rate for pancreatic cancer (PC), which has a 5-year survival rate of about 7%. Recent failures of targeted therapies inhibiting kinase activity in clinical trials have highlighted the need for new approaches towards combating this deadly disease. In this study, we have identified genes that are significantly downregulated in PC, through a meta-analysis of large number of microarray datasets. We have used qRT-PCR to confirm the downregulation of selected genes in a panel of PC cell lines. This study has yielded several novel candidate tumor-suppressor genes (TSGs) including GNMT, CEL, PLA2G1B and SERPINI2. We highlight the role of GNMT, a methyl transferase associated with the methylation potential of the cell, and CEL, a lipase, as potential therapeutic targets. We have uncovered genetic links to risk factors associated with PC such as smoking and obesity. Genes important for patient survival and prognosis are also discussed, and we confirm the dysregulation of metabolic pathways previously observed in PC. While many of the genes downregulated in our dataset are associated with protein products normally produced by the pancreas for excretion, we have uncovered some genes whose downregulation appear to play a more causal role in PC. These genes will assist in providing a better understanding of the disease etiology of PC, and in the search for new therapeutic targets and biomarkers.

  20. Id-1 gene and gene products as therapeutic targets for treatment of breast cancer and other types of carcinoma

    Science.gov (United States)

    Desprez, Pierre-Yves; Campisi, Judith

    2014-08-19

    A method for treatment of breast cancer and other types of cancer. The method comprises targeting and modulating Id-1 gene expression, if any, for the Id-1 gene, or gene products in breast or other epithelial cancers in a patient by delivering products that modulate Id-1 gene expression. When expressed, Id-1 gene is a prognostic indicator that cancer cells are invasive and metastatic.

  1. Poly[N-(2-aminoethyl)ethyleneimine] as a New Non-Viral Gene Delivery Carrier : The Effect of Two Protonatable Nitrogens in the Monomer Unit on Gene Delivery Efficiency

    NARCIS (Netherlands)

    Khazaie, Yahya; Novo, Luis; van Gaal, Ethlinn; Fassihi, Afshin; Mirahmadi-Zareh, Seyedeh Zohreh; Esfahani, Mohammad Hossein Nasr; van Nostrum, Cornelus F.; Hennink, Wim E.; Dorkoosh, Farid

    2014-01-01

    Purpose. The aim of this study was to investigate the in vitro gene delivery efficiency of poly[N-(2-aminoethyl)ethylene-imine](PAEEI), a polymer with a linear Polyethyleneimine (LPEI) backbone and with aminoethyl side groups that has two protonatable nitrogen atoms per monomer unit instead of one

  2. AAV serotype 2/1-mediated gene delivery of anti-inflammatory interleukin-10 enhances neurogenesis and cognitive function in APP+PS1 mice.

    Science.gov (United States)

    Kiyota, T; Ingraham, K L; Swan, R J; Jacobsen, M T; Andrews, S J; Ikezu, T

    2012-07-01

    Brain inflammation is a double-edged sword. It is required for brain repair in acute damage, whereas chronic inflammation and autoimmune disorders are neuropathogenic. Certain proinflammatory cytokines and chemokines are closely related to cognitive dysfunction and neurodegeneration. Representative anti-inflammatory cytokines, such as interleukin (IL)-10, can suppress neuroinflammation and have significant therapeutic potentials in ameliorating neurodegenerative disorders such as Alzheimer's disease (AD). Here, we show that adeno-associated virus (AAV) serotype 2/1 hybrid-mediated neuronal expression of the mouse IL-10 gene ameliorates cognitive dysfunction in amyloid precursor protein+ presenilin-1 bigenic mice. AAV2/1 infection of hippocampal neurons resulted in sustained expression of IL-10 without its leakage into the blood, reduced astro/microgliosis, enhanced plasma amyloid-β peptide (Aβ) levels and enhanced neurogenesis. Moreover, increased levels of IL-10 improved spatial learning, as determined by the radial arm water maze. Finally, IL-10-stimulated microglia enhanced proliferation but not differentiation of primary neural stem cells in the co-culture system, whereas IL-10 itself had no effect. Our data suggest that IL-10 gene delivery has a therapeutic potential for a non-Aβ-targeted treatment of AD.

  3. A rapid pathway toward a superb gene delivery system: programming structural and functional diversity into a supramolecular nanoparticle library.

    Science.gov (United States)

    Wang, Hao; Liu, Kan; Chen, Kuan-Ju; Lu, Yujie; Wang, Shutao; Lin, Wei-Yu; Guo, Feng; Kamei, Ken-ichiro; Chen, Yi-Chun; Ohashi, Minori; Wang, Mingwei; Garcia, Mitch André; Zhao, Xing-Zhong; Shen, Clifton K-F; Tseng, Hsian-Rong

    2010-10-26

    Nanoparticles are regarded as promising transfection reagents for effective and safe delivery of nucleic acids into a specific type of cells or tissues providing an alternative manipulation/therapy strategy to viral gene delivery. However, the current process of searching novel delivery materials is limited due to conventional low-throughput and time-consuming multistep synthetic approaches. Additionally, conventional approaches are frequently accompanied with unpredictability and continual optimization refinements, impeding flexible generation of material diversity creating a major obstacle to achieving high transfection performance. Here we have demonstrated a rapid developmental pathway toward highly efficient gene delivery systems by leveraging the powers of a supramolecular synthetic approach and a custom-designed digital microreactor. Using the digital microreactor, broad structural/functional diversity can be programmed into a library of DNA-encapsulated supramolecular nanoparticles (DNA⊂SNPs) by systematically altering the mixing ratios of molecular building blocks and a DNA plasmid. In vitro transfection studies with DNA⊂SNPs library identified the DNA⊂SNPs with the highest gene transfection efficiency, which can be attributed to cooperative effects of structures and surface chemistry of DNA⊂SNPs. We envision such a rapid developmental pathway can be adopted for generating nanoparticle-based vectors for delivery of a variety of loads.

  4. Ultrasound-mediated gene delivery of naked plasmid DNA in skeletal muscles: a case for bolus injections.

    Science.gov (United States)

    Sanches, Pedro Gomes; Mühlmeister, Mareike; Seip, Ralf; Kaijzel, Eric; Löwik, Clemens; Böhmer, Marcel; Tiemann, Klaus; Grüll, Holger

    2014-12-10

    Localized gene delivery has many potential clinical applications. However, the nucleic acids (e.g. pDNA and siRNA) are incapable of passively crossing the endothelium, cell membranes and other biological barriers which must be crossed to reach their intracellular targets. A possible solution is the use of ultrasound to burst circulating microbubbles inducing transient permeabilization of surrounding tissues which mediates nucleic acid extravasation and cellular uptake. In this study we report on an optimization of the ultrasound gene delivery technique. Naked pDNA (200 μg) encoding luciferase and SonoVue® microbubbles were co-injected intravenously in mice. The hindlimb skeletal muscles were exposed to ultrasound from a non-focused transducer (1 MHz, 1.25 MPa, PRI 30s) and injection protocols and total amounts as well as ultrasound parameters were systemically varied. Gene expression was quantified relative to a control using a bioluminescence camera system at day 7 after sonication. Bioluminescence ratios in sonicated/control muscles of up to 101× were obtained. In conclusion, we were able to specifically deliver genetic material to the selected skeletal muscles and overall, the use of bolus injections and high microbubble numbers resulted in increased gene expression reflected by stronger bioluminescence signals. Based on our data, bolus injections seem to be required in order to achieve transient highly concentrated levels of nucleic acids and microbubbles at the tissue of interest which upon ultrasound exposure should lead to increased levels of gene delivery. Thus, ultrasound mediated gene delivery is a promising technique for the clinical translation of localized drug delivery. Copyright © 2014 Elsevier B.V. All rights reserved.

  5. Polyglycerol-functionalized nanodiamond as a platform for gene delivery: Derivatization, characterization, and hybridization with DNA

    Directory of Open Access Journals (Sweden)

    Li Zhao

    2014-03-01

    Full Text Available A gene vector consisting of nanodiamond, polyglycerol, and basic polypeptide (ND-PG-BPP has been designed, synthesized, and characterized. The ND-PG-BPP was synthesized by PG functionalization of ND through ring-opening polymerization of glycidol on the ND surface, multistep organic transformations (–OH → –OTs (tosylate → –N3 in the PG layer, and click conjugation of the basic polypeptides (Arg8, Lys8 or His8 terminated with propargyl glycine. The ND-PG-BPP exhibited good dispersibility in water (>1.0 mg/mL and positive zeta potential ranging from +14.2 mV to +44.1 mV at neutral pH in Milli-Q water. It was confirmed by gel retardation assay that ND-PG-Arg8 and ND-PG-Lys8 with higher zeta potential hybridized with plasmid DNA (pDNA through electrostatic attraction, making them promising as nonviral vectors for gene delivery.

  6. Polyglycerol-functionalized nanodiamond as a platform for gene delivery: Derivatization, characterization, and hybridization with DNA

    Science.gov (United States)

    Zhao, Li; Nakae, Yuki; Qin, Hongmei; Ito, Tadamasa; Kimura, Takahide; Kojima, Hideto; Chan, Lawrence

    2014-01-01

    Summary A gene vector consisting of nanodiamond, polyglycerol, and basic polypeptide (ND-PG-BPP) has been designed, synthesized, and characterized. The ND-PG-BPP was synthesized by PG functionalization of ND through ring-opening polymerization of glycidol on the ND surface, multistep organic transformations (–OH → –OTs (tosylate) → –N3) in the PG layer, and click conjugation of the basic polypeptides (Arg8, Lys8 or His8) terminated with propargyl glycine. The ND-PG-BPP exhibited good dispersibility in water (>1.0 mg/mL) and positive zeta potential ranging from +14.2 mV to +44.1 mV at neutral pH in Milli-Q water. It was confirmed by gel retardation assay that ND-PG-Arg8 and ND-PG-Lys8 with higher zeta potential hybridized with plasmid DNA (pDNA) through electrostatic attraction, making them promising as nonviral vectors for gene delivery. PMID:24778723

  7. Development of a DNA-liposome complex for gene delivery applications

    Energy Technology Data Exchange (ETDEWEB)

    Rasoulianboroujeni, M. [Marquette University School of Dentistry, Milwaukee, WI 53233 (United States); Kupgan, G. [Department of Chemical Engineering, Oklahoma State University, 423 Engineering North, Stillwater, OK 74078 (United States); Moghadam, F. [School of Biological and Health Systems Engineering, Arizona State University, Tempe, AZ (United States); Tahriri, M. [Marquette University School of Dentistry, Milwaukee, WI 53233 (United States); Boughdachi, A. [Polymer Engineering Department, Amirkabir University of Technology, Tehran (Iran, Islamic Republic of); Khoshkenar, P. [Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, MA 01605 (United States); Ambrose, J.J. [Biomedical Engineering Department, Louisiana Tech University, Ruston, LA 71272 (United States); Kiaie, N. [Tissue Engineering Department, Faculty of Biomedical Engineering, Amirkabir University of Technology, Tehran (Iran, Islamic Republic of); Vashaee, D. [Electrical and Computer Engineering Department, North Carolina State University, Raleigh, NC 27606 (United States); Ramsey, J.D. [Department of Chemical Engineering, Oklahoma State University, 423 Engineering North, Stillwater, OK 74078 (United States); Tayebi, L., E-mail: lobat.tayebi@marquette.edu [Marquette University School of Dentistry, Milwaukee, WI 53233 (United States)

    2017-06-01

    The association structures formed by cationic liposomes and DNA (Deoxyribonucleic acid)-liposome have been effectively utilized as gene carriers in transfection assays. In this research study, cationic liposomes were prepared using a modified lipid film hydration method consisting of a lyophilization step for gene delivery applications. The obtained results demonstrated that the mean particle size had no significant change while the polydispersity (PDI) increased after lyophilization. The mean particle size slightly reduced after lyophilization (520 ± 12 nm to 464 ± 25 nm) while the PDI increased after lyophilization (0.094 ± 0.017 to 0.220 ± 0.004). In addition. The mean particle size of vesicles increases when DNA is incorporated to the liposomes (673 ± 27 nm). According to the Scanning Electron Microscopy (SEM) and transmission electron microscopy (TEM) images, the spherical shape of liposomes confirmed their successful preservation and reconstitution from the powder. It was found that liposomal formulation has enhanced transfection considerably compared to the naked DNA as negative control. Finally, liposomal formulation in this research had a better function than Lipofectamine® 2000 as a commercialized product because the cellular activity (cellular protein) was higher in the prepared lipoplex than Lipofectamine® 2000. - Highlights: • Liposomal formulation in this research had a better function than Lipofectamine® 2000. • The average particle size had no significant change while the PDI increased after lyophilization. • LacZ expression of the developed cationic liposomes is approximately equal to the Lipofectamine® 2000.

  8. Nano and Microtechnologies for the Delivery of Oligonucleotides with Gene Silencing Properties

    Directory of Open Access Journals (Sweden)

    Giuseppe De Rosa

    2009-07-01

    Full Text Available Oligonucleotides (ONs are synthetic fragments of nucleic acid designed to modulate the expression of target proteins. DNA-based ONs (antisense, antigene, aptamer or decoy and more recently a new class of RNA-based ONs, the small interfering RNAs (siRNAs, have gained great attention for the treatment of different disease states, such as viral infections, inflammation, diabetes, and cancer. However, the development of therapeutic strategies based on ONs is hampered by their low bioavailability, poor intracellular uptake and rapid degradation in biological fluids. The use of a non-viral carrier can be a powerful tool to overcome these drawbacks. Lipid or polymer-based nanotechnologies can improve biological stability and cellular uptake of ONs, with possibility of tissue and/or cellular targeting. The use of polymeric devices can also produce a prolonged release of the ON, thus reducing the need of frequent administrations. This review summarizes advantages and issues related to the main non-viral vectors used for ON delivery.

  9. An efficient nonviral gene-delivery vector based on hyperbranched cationic glycogen derivatives

    Directory of Open Access Journals (Sweden)

    Liang X

    2014-01-01

    Full Text Available Xuan Liang,1,* Xianyue Ren,2,* Zhenzhen Liu,1 Yingliang Liu,1 Jue Wang,2 Jingnan Wang,2 Li-Ming Zhang,1 David YB Deng,2 Daping Quan,1 Liqun Yang1 1Institute of Polymer Science, School of Chemistry and Chemical Engineering, Key Laboratory of Designed Synthesis and Application of Polymer Material, Key Laboratory for Polymeric Composite and Functional Materials of Ministry of Education, Sun Yat-Sen University, Guangzhou, People's Republic of China; 2Research Center of Translational Medicine, The First Affiliated Hospital, Sun Yat-Sen University, Guangzhou, People's Republic of China *Both these authors contributed equally to this work Background: The purpose of this study was to synthesize and evaluate hyperbranched cationic glycogen derivatives as an efficient nonviral gene-delivery vector. Methods: A series of hyperbranched cationic glycogen derivatives conjugated with 3-(dimethylamino-1-propylamine (DMAPA-Glyp and 1-(2-aminoethyl piperazine (AEPZ-Glyp residues were synthesized and characterized by Fourier-transform infrared and hydrogen-1 nuclear magnetic resonance spectroscopy. Their buffer capacity was assessed by acid–base titration in aqueous NaCl solution. Plasmid deoxyribonucleic acid (pDNA condensation ability and protection against DNase I degradation of the glycogen derivatives were assessed using agarose gel electrophoresis. The zeta potentials and particle sizes of the glycogen derivative/pDNA complexes were measured, and the images of the complexes were observed using atomic force microscopy. Blood compatibility and cytotoxicity were evaluated by hemolysis assay and MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay, respectively. pDNA transfection efficiency mediated by the cationic glycogen derivatives was evaluated by flow cytometry and fluorescence microscopy in the 293T (human embryonic kidney and the CNE2 (human nasopharyngeal carcinoma cell lines. In vivo delivery of pDNA in model animals (Sprague Dawley

  10. Exploring advantages/disadvantages and improvements in overcoming gene delivery barriers of amino acid modified trimethylated chitosan.

    Science.gov (United States)

    Zheng, Hao; Tang, Cui; Yin, Chunhua

    2015-06-01

    Present study aimed at exploring advantages/disadvantages of amino acid modified trimethylated chitosan in conquering multiple gene delivery obstacles and thus providing comprehensive understandings for improved transfection efficiency. Arginine, cysteine, and histidine modified trimethyl chitosan were synthesized and employed to self-assemble with plasmid DNA (pDNA) to form nanocomplexes, namely TRNC, TCNC, and THNC, respectively. They were assessed by structural stability, cellular uptake, endosomal escape, release behavior, nuclear localization, and in vitro and in vivo transfection efficiencies. Besides, sodium tripolyphosphate (TPP) was added into TRNC to compromise certain disadvantageous attributes for pDNA delivery. Optimal endosomal escape ability failed to bring in satisfactory transfection efficiency of THNC due to drawbacks in structural stability, cellular uptake, pDNA liberation, and nuclear distribution. TCNC evoked the most potent gene expression owing to multiple advantages including sufficient stability, preferable uptake, efficient pDNA release, and high nucleic accumulation. Undesirable stability and insufficient pDNA release adversely affected TRNC-mediated gene transfer. However, incorporation of TPP could improve such disadvantages and consequently resulted in enhanced transfection efficiencies. Coordination of multiple contributing effects to conquer all delivery obstacles was necessitated for improved transfection efficiency, which would provide insights into rational design of gene delivery vehicles.

  11. Ultrasound-mediated gene delivery of naked plasmid DNA in skeletal muscles : a case for bolus injections

    NARCIS (Netherlands)

    Gomes Sanches, P.; Muehlmeister, M.; Seip, R.; Kaijzel, E.L.; Loewik, C.; Boehmer, M.; Tiemann, K.; Grüll, H.

    2014-01-01

    Localized gene delivery has many potential clinical applications. However, the nucleic acids (e.g. pDNA and siRNA) are incapable of passively crossing the endothelium, cell membranes and other biological barriers which must be crossed to reach their intracellular targets. A possible solution is the

  12. Smart and Controllable rAAV Gene Delivery Carriers in Progenitor Cells for Human Musculoskeletal Regenerative Medicine with a Focus on the Articular Cartilage.

    Science.gov (United States)

    Rey-Rico, Ana; Cucchiarini, Magali

    2017-01-01

    Cell therapy using mesenchymal stem cells (MSCs) is a powerful tool for the treatment of various diseases and injuries. Still, important limitations including the large amounts of cells required for application in vivo and the age-related decline in lifespan, proliferation, and potency may hinder the use of MSCs in patients. In this regard, gene therapy may offer strong approaches to optimize the use of MSCs for regenerative medicine. Diverse nonviral and viral gene vehicles have been manipulated to genetically modify MSCs, among which the highly effective and relatively safe recombinant adeno-associated viral (rAAV) vectors that emerged as the preferred gene delivery system to treat human disorders. Yet, clinical adaptation of such gene vehicles may be limited by several hurdles, including the possibility of dissemination to nontarget sites and the presence of immune and toxic responses in the host organism that may impair their therapeutic actions. The use of smart biomaterials acting as interfaces to enhance the temporal and spatial presentation of therapeutic agents in the target place and/or acting as scaffolding for MSC growth is an innovative, valuable approach to overcome these shortcomings that else restrain the efficacy of such potent cell populations. Here, we provide an overview on the most recent tissue engineering approaches based on the use of biomaterials acting as vehicles for rAAV vectors to target MSCs directly in the recipient (in vivo strategy) or as supportive matrices for rAAV-modified MSCs for indirect cell reimplantation (ex vivo strategy) as means to activate the reparative processes in tissues of the musculoskeletal system. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  13. Gene delivery to pancreatic exocrine cells in vivo and in vitro

    Directory of Open Access Journals (Sweden)

    Houbracken Isabelle

    2012-10-01

    Full Text Available Abstract Background Effective gene transfer to the pancreas or to pancreatic cells has remained elusive although it is essential for studies of genetic lineage tracing and modulation of gene expression. Different transduction methods and viral vectors were tested in vitro and in vivo, in rat and mouse pancreas. Results For in vitro transfection/transduction of rat exocrine cells lipofection reagents, adenoviral vectors, and Mokola- and VSV-G pseudotyped lentiviral vectors were used. For in vivo transduction of mouse and rat pancreas adenoviral vectors and VSV-G lentiviral vectors were injected into the parenchymal tissue. Both lipofection of rat exocrine cell cultures and transduction with Mokola pseudotyped lentiviral vectors were inefficient and resulted in less than 4% EGFP expressing cells. Adenoviral transduction was highly efficient but its usefulness for gene delivery to rat exocrine cells in vitro was hampered by a drastic increase in cell death. In vitro transduction of rat exocrine cells was most optimal with VSV-G pseudotyped lentiviral vectors, with stable transgene expression, no significant effect on cell survival and about 40% transduced cells. In vivo, pancreatic cells could not be transduced by intra-parenchymal administration of lentiviral vectors in mouse and rat pancreas. However, a high efficiency could be obtained by adenoviral vectors, resulting in transient transduction of mainly exocrine acinar cells. Injection in immune-deficient animals diminished leukocyte infiltration and prolonged transgene expression. Conclusions In summary, our study remarkably demonstrates that transduction of pancreatic exocrine cells requires lentiviral vectors in vitro but adenoviral vectors in vivo.

  14. Photodegradable neutral-cationic brush block copolymers for nonviral gene delivery.

    Science.gov (United States)

    Hu, Xianglong; Li, Yang; Liu, Tao; Zhang, Guoying; Liu, Shiyong

    2014-08-01

    We report on the fabrication of a photodegradable gene-delivery vector based on PEO-b-P(GMA-g-PDMAEMA) neutral-cationic brush block copolymers that possess cationic poly(N,N-dimethylaminoethyl methacrylate) (PDMAEMA) brushes for DNA compaction, poly(ethylene oxide) (PEO) as a hydrophilic block, and poly(glycidyl methacrylate) (PGMA) as the backbone. The PEO-b-P(GMA-g-PDMAEMA) copolymers were synthesized through the combination of reversible addition-fragmentation transfer (RAFT) polymerization and postmodification. A photocleavable PEO-based macroRAFT agent was first synthesized; next, the PEO-b-PGMA block copolymer was prepared by RAFT polymerization of GMA; this was followed by a click reaction to introduce the RAFT initiators on the side chains of the PGMA block; then, RAFT polymerization of DMAEMA afforded the PEO-b-P(GMA-g-PDMAEMA) copolymer. The obtained neutral-cationic brush block copolymer could effectively complex plasmid DNA (pDNA) into nanoparticles at an N/P ratio (i.e., the number of nitrogen residues per DNA phosphate) of 4. Upon UV irradiation, pDNA could be released owing to cleavage of the pDNA-binding cationic PDMAEMA side chains as well as the nitrobenzyl ester linkages at the diblock junction point. In addition, in vitro gene transfection results demonstrated that the polyplexes could be effectively internalized by cells with good transfection efficiency, and the UV irradiation protocol could considerably enhance the efficiency of gene transfection. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Recombinant human parvovirus B19 vectors: erythroid cell-specific delivery and expression of transduced genes.

    Science.gov (United States)

    Ponnazhagan, S; Weigel, K A; Raikwar, S P; Mukherjee, P; Yoder, M C; Srivastava, A

    1998-06-01

    A novel packaging strategy combining the salient features of two human parvoviruses, namely the pathogenic parvovirus B19 and the nonpathogenic adeno-associated virus type 2 (AAV), was developed to achieve erythroid cell-specific delivery as well as expression of the transduced gene. The development of such a chimeric vector system was accomplished by packaging heterologous DNA sequences cloned within the inverted terminal repeats of AAV and subsequently packaging the DNA inside the capsid structure of B19 virus. Recombinant B19 virus particles were assembled, as evidenced by electron microscopy as well as DNA slot blot analyses. The hybrid vector failed to transduce nonerythroid human cells, such as 293 cells, as expected. However, MB-02 cells, a human megakaryocytic leukemia cell line which can be infected by B19 virus following erythroid differentiation with erythropoietin (N. C. Munshi, S. Z. Zhou, M. J. Woody, D. A. Morgan, and A. Srivastava, J. Virol. 67:562-566, 1993) but lacks the putative receptor for AAV (S. Ponnazhagan, X.-S. Wang, M. J. Woody, F. Luo, L. Y. Kang, M. L. Nallari, N. C. Munshi, S. Z. Zhou, and A. Srivastava, J. Gen. Virol. 77:1111-1122, 1996), were readily transduced by this vector. The hybrid vector was also found to specifically target the erythroid population in primary human bone marrow cells as well as more immature hematopoietic progenitor cells following erythroid differentiation, as evidenced by selective expression of the transduced gene in these target cells. Preincubation with anticapsid antibodies against B19 virus, but not anticapsid antibodies against AAV, inhibited transduction of primary human erythroid cells. The efficiency of transduction of primary human erythroid cells by the recombinant B19 virus vector was significantly higher than that by the recombinant AAV vector. Further development of the AAV-B19 virus hybrid vector system should prove beneficial in gene therapy protocols aimed at the correction of inherited and

  16. Recombinant Human Parvovirus B19 Vectors: Erythroid Cell-Specific Delivery and Expression of Transduced Genes

    Science.gov (United States)

    Ponnazhagan, Selvarangan; Weigel, Kirsten A.; Raikwar, Sudhanshu P.; Mukherjee, Pinku; Yoder, Mervin C.; Srivastava, Arun

    1998-01-01

    A novel packaging strategy combining the salient features of two human parvoviruses, namely the pathogenic parvovirus B19 and the nonpathogenic adeno-associated virus type 2 (AAV), was developed to achieve erythroid cell-specific delivery as well as expression of the transduced gene. The development of such a chimeric vector system was accomplished by packaging heterologous DNA sequences cloned within the inverted terminal repeats of AAV and subsequently packaging the DNA inside the capsid structure of B19 virus. Recombinant B19 virus particles were assembled, as evidenced by electron microscopy as well as DNA slot blot analyses. The hybrid vector failed to transduce nonerythroid human cells, such as 293 cells, as expected. However, MB-02 cells, a human megakaryocytic leukemia cell line which can be infected by B19 virus following erythroid differentiation with erythropoietin (N. C. Munshi, S. Z. Zhou, M. J. Woody, D. A. Morgan, and A. Srivastava, J. Virol. 67:562–566, 1993) but lacks the putative receptor for AAV (S. Ponnazhagan, X.-S. Wang, M. J. Woody, F. Luo, L. Y. Kang, M. L. Nallari, N. C. Munshi, S. Z. Zhou, and A. Srivastava, J. Gen. Virol. 77:1111–1122, 1996), were readily transduced by this vector. The hybrid vector was also found to specifically target the erythroid population in primary human bone marrow cells as well as more immature hematopoietic progenitor cells following erythroid differentiation, as evidenced by selective expression of the transduced gene in these target cells. Preincubation with anticapsid antibodies against B19 virus, but not anticapsid antibodies against AAV, inhibited transduction of primary human erythroid cells. The efficiency of transduction of primary human erythroid cells by the recombinant B19 virus vector was significantly higher than that by the recombinant AAV vector. Further development of the AAV-B19 virus hybrid vector system should prove beneficial in gene therapy protocols aimed at the correction of inherited

  17. Tumor cell-targeted delivery of CRISPR/Cas9 by aptamer-functionalized lipopolymer for therapeutic genome editing of VEGFA in osteosarcoma.

    Science.gov (United States)

    Liang, Chao; Li, Fangfei; Wang, Luyao; Zhang, Zong-Kang; Wang, Chao; He, Bing; Li, Jie; Chen, Zhihao; Shaikh, Atik Badshah; Liu, Jin; Wu, Xiaohao; Peng, Songlin; Dang, Lei; Guo, Baosheng; He, Xiaojuan; Au, D W T; Lu, Cheng; Zhu, Hailong; Zhang, Bao-Ting; Lu, Aiping; Zhang, Ge

    2017-12-01

    Osteosarcoma (OS) is a highly aggressive pediatric cancer, characterized by frequent lung metastasis and pathologic bone destruction. Vascular endothelial growth factor A (VEGFA), highly expressed in OS, not only contributes to angiogenesis within the tumor microenvironment via paracrine stimulation of vascular endothelial cells, but also acts as an autocrine survival factor for tumor cell themselves, thus making it a promising therapeutic target for OS. CRISPR/Cas9 is a versatile genome editing technology and holds tremendous promise for cancer treatment. However, a major bottleneck to achieve the therapeutic potential of the CRISPR/Cas9 is the lack of in vivo tumor-targeted delivery systems. Here, we screened an OS cell-specific aptamer (LC09) and developed a LC09-functionalized PEG-PEI-Cholesterol (PPC) lipopolymer encapsulating CRISPR/Cas9 plasmids encoding VEGFA gRNA and Cas9. Our results demonstrated that LC09 facilitated selective distribution of CRISPR/Cas9 in both orthotopic OS and lung metastasis, leading to effective VEGFA genome editing in tumor, decreased VEGFA expression and secretion, inhibited orthotopic OS malignancy and lung metastasis, as well as reduced angiogenesis and bone lesion with no detectable toxicity. The delivery system simultaneously restrained autocrine and paracrine VEGFA signaling in tumor cells and could facilitate translating CRISPR-Cas9 into clinical cancer treatment. Copyright © 2017 Elsevier Ltd. All rights reserved.

  18. Tumor-directed gene therapy in mice using a composite nonviral gene delivery system consisting of the piggyBac transposon and polyethylenimine

    International Nuclear Information System (INIS)

    Kang, Yu; Zhang, Xiaoyan; Jiang, Wei; Wu, Chaoqun; Chen, Chunmei; Zheng, Yufang; Gu, Jianren; Xu, Congjian

    2009-01-01

    Compared with viral vectors, nonviral vectors are less immunogenic, more stable, safer and easier to replication for application in cancer gene therapy. However, nonviral gene delivery system has not been extensively used because of the low transfection efficiency and the short transgene expression, especially in vivo. It is desirable to develop a nonviral gene delivery system that can support stable genomic integration and persistent gene expression in vivo. Here, we used a composite nonviral gene delivery system consisting of the piggyBac (PB) transposon and polyethylenimine (PEI) for long-term transgene expression in mouse ovarian tumors. A recombinant plasmid PB [Act-RFP, HSV-tk] encoding both the herpes simplex thymidine kinase (HSV-tk) and the monomeric red fluorescent protein (mRFP1) under PB transposon elements was constructed. This plasmid and the PBase plasmid were injected into ovarian cancer tumor xenografts in mice by in vivo PEI system. The antitumor effects of HSV-tk/ganciclovir (GCV) system were observed after intraperitoneal injection of GCV. Histological analysis and TUNEL assay were performed on the cryostat sections of the tumor tissue. Plasmid construction was confirmed by PCR analysis combined with restrictive enzyme digestion. mRFP1 expression could be visualized three weeks after the last transfection of pPB/TK under fluorescence microscopy. After GCV admission, the tumor volume of PB/TK group was significantly reduced and the tumor inhibitory rate was 81.96% contrasted against the 43.07% in the TK group. Histological analysis showed that there were extensive necrosis and lymphocytes infiltration in the tumor tissue of the PB/TK group but limited in the tissue of control group. TUNEL assays suggested that the transfected cells were undergoing apoptosis after GCV admission in vivo. Our results show that the nonviral gene delivery system coupling PB transposon with PEI can be used as an efficient tool for gene therapy in ovarian cancer

  19. 3D Porous Chitosan-Alginate Scaffolds as an In Vitro Model for Evaluating Nanoparticle-Mediated Tumor Targeting and Gene Delivery to Prostate Cancer.

    Science.gov (United States)

    Wang, Kui; Kievit, Forrest M; Florczyk, Stephen J; Stephen, Zachary R; Zhang, Miqin

    2015-10-12

    Cationic nanoparticles (NPs) for targeted gene delivery are conventionally evaluated using 2D in vitro cultures. However, this does not translate well to corresponding in vivo studies because of the marked difference in NP behavior in the presence of the tumor microenvironment. In this study, we investigated whether prostate cancer (PCa) cells cultured in three-dimensional (3D) chitosan-alginate (CA) porous scaffolds could model cationic NP-mediated gene targeted delivery to tumors in vitro. We assessed in vitro tumor cell proliferation, formation of tumor spheroids, and expression of marker genes that promote tumor malignancy in CA scaffolds. The efficacy of NP-targeted gene delivery was evaluated in PCa cells in 2D cultures, PCa tumor spheroids grown in CA scaffolds, and PCa tumors in a mouse TRAMP-C2 flank tumor model. PCa cells cultured in CA scaffolds grew into tumor spheroids and displayed characteristics of higher malignancy as compared to those in 2D cultures. Significantly, targeted gene delivery was only observed in cells cultured in CA scaffolds, whereas cells cultured on 2D plates showed no difference in gene delivery between targeted and nontarget control NPs. In vivo NP evaluation confirmed targeted gene delivery, indicating that only CA scaffolds correctly modeled NP-mediated targeted delivery in vivo. These findings suggest that CA scaffolds serve as a better in vitro platform than 2D cultures for evaluation of NP-mediated targeted gene delivery to PCa.

  20. Oligopeptide complex for targeted non-viral gene delivery to adipocytes

    Science.gov (United States)

    Won, Young-Wook; Adhikary, Partho Protim; Lim, Kwang Suk; Kim, Hyung Jin; Kim, Jang Kyoung; Kim, Yong-Hee

    2014-12-01

    Commercial anti-obesity drugs acting in the gastrointestinal tract or the central nervous system have been shown to have limited efficacy and severe side effects. Anti-obesity drug development is thus focusing on targeting adipocytes that store excess fat. Here, we show that an adipocyte-targeting fusion-oligopeptide gene carrier consisting of an adipocyte-targeting sequence and 9-arginine (ATS-9R) selectively transfects mature adipocytes by binding to prohibitin. Injection of ATS-9R into obese mice confirmed specific binding of ATS-9R to fat vasculature, internalization and gene expression in adipocytes. We also constructed a short-hairpin RNA (shRNA) for silencing fatty-acid-binding protein 4 (shFABP4), a key lipid chaperone in fatty-acid uptake and lipid storage in adipocytes. Treatment of obese mice with ATS-9R/shFABP4 led to metabolic recovery and body-weight reduction (>20%). The ATS-9R/shFABP4 oligopeptide complex could prove to be a safe therapeutic approach to regress and treat obesity as well as obesity-induced metabolic syndromes.