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Sample records for hiv-1 gp120 c4-v3

  1. Prediction of the secondary structure of HIV-1 gp120

    DEFF Research Database (Denmark)

    Hansen, J E; Lund, O; Nielsen, Jens Ole

    1996-01-01

    strain BH10 gp120, as well as in 27 other HIV-1 strains examined. Two helical segments were predicted in regions displaying profound sequence variation, one in a region suggested to be critical for CD4 binding. The predicted content of helix, beta-strand, and coil was consistent with estimates from......The secondary structure of HIV-1 gp120 was predicted using multiple alignment and a combination of two independent methods based on neural network and nearest-neighbor algorithms. The methods agreed on the secondary structure for 80% of the residues in BH10 gp120. Six helices were predicted in HIV...... Fourier transform infrared spectroscopy. The predicted secondary structure of gp120 compared well with data from NMR analysis of synthetic peptides from the V3 loop and the C4 region. As a first step towards modeling the tertiary structure of gp120, the predicted secondary structure may guide the design...

  2. Prediction of the secondary structure of HIV-1 gp120

    DEFF Research Database (Denmark)

    Hansen, J E; Lund, O; Nielsen, Jens Ole

    1996-01-01

    The secondary structure of HIV-1 gp120 was predicted using multiple alignment and a combination of two independent methods based on neural network and nearest-neighbor algorithms. The methods agreed on the secondary structure for 80% of the residues in BH10 gp120. Six helices were predicted in HIV...... strain BH10 gp120, as well as in 27 other HIV-1 strains examined. Two helical segments were predicted in regions displaying profound sequence variation, one in a region suggested to be critical for CD4 binding. The predicted content of helix, beta-strand, and coil was consistent with estimates from...... of future HIV subunit vaccine candidates....

  3. Prediction of the Secondary Structure of HIV-1 gp120

    DEFF Research Database (Denmark)

    Hansen, Jan; Lund, Ole; Nielsen, Jens O.

    1996-01-01

    The secondary structure of HIV-1 gp120 was predicted using multiple alignment and a combination of two independent methods based on neural network and nearest-neighbor algorithms. The methods agreed on the secondary structure for 80% of the residues in BH10 gp120. Six helices were predicted in HIV...... strain BH10 gp120, as well as in 27 other HIV-1 strains examined. Two helical seqments were predicted in regions displaying profound sequence variation, one in a region suggested to be critical for CD4 biding. The predicted content of helix, beta-strand, and coil was consistent with estimates from...... of future HIV sub-unit vaccine candidates....

  4. Antioxidant protection from HIV-1 gp120-induced neuroglial toxicity

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    Walsh Kimberley A

    2004-05-01

    Full Text Available Abstract Background The pathogenesis of HIV-1 glycoprotein 120 (gp120 associated neuroglial toxicity remains unresolved, but oxidative injury has been widely implicated as a contributing factor. In previous studies, exposure of primary human central nervous system tissue cultures to gp120 led to a simplification of neuronal dendritic elements as well as astrocytic hypertrophy and hyperplasia; neuropathological features of HIV-1-associated dementia. Gp120 and proinflammatory cytokines upregulate inducible nitric oxide synthase (iNOS, an important source of nitric oxide (NO and nitrosative stress. Because ascorbate scavenges reactive nitrogen and oxygen species, we studied the effect of ascorbate supplementation on iNOS expression as well as the neuronal and glial structural changes associated with gp120 exposure. Methods Human CNS cultures were derived from 16–18 week gestation post-mortem fetal brain. Cultures were incubated with 400 μM ascorbate-2-O-phosphate (Asc-p or vehicle for 18 hours then exposed to 1 nM gp120 for 24 hours. The expression of iNOS and neuronal (MAP2 and astrocytic (GFAP structural proteins was examined by immunohistochemistry and immunofluorescence using confocal scanning laser microscopy (CSLM. Results Following gp120 exposure iNOS was markedly upregulated from undetectable levels at baseline. Double label CSLM studies revealed astrocytes to be the prime source of iNOS with rare neurons expressing iNOS. This upregulation was attenuated by the preincubation with Asc-p, which raised the intracellular concentration of ascorbate. Astrocytic hypertrophy and neuronal injury caused by gp120 were also prevented by preincubation with ascorbate. Conclusions Ascorbate supplementation prevents the deleterious upregulation of iNOS and associated neuronal and astrocytic protein expression and structural changes caused by gp120 in human brain cell cultures.

  5. Binding kinetics of aptamers to gp120 derived from HIV-1 subtype C

    CSIR Research Space (South Africa)

    Millroy, L

    2011-02-01

    Full Text Available aptamers with specific and strong affinity to the HIV-1 envelope glycoprotein gp120 and act as novel HIV-1 entry inhibitor drugs or as targeted drug delivery systems to HIV-1 infected cells. Prior to any downstream applications, novel gp120 aptamers need...

  6. HIV-1 gp120 mannoses induce immunosuppressive responses from dendritic cells.

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    Meimei Shan

    2007-11-01

    Full Text Available The human immunodeficiency virus type 1 (HIV-1 envelope glycoprotein gp120 is a vaccine immunogen that can signal via several cell surface receptors. To investigate whether receptor biology could influence immune responses to gp120, we studied its interaction with human, monocyte-derived dendritic cells (MDDCs in vitro. Gp120 from the HIV-1 strain JR-FL induced IL-10 expression in MDDCs from 62% of donors, via a mannose C-type lectin receptor(s (MCLR. Gp120 from the strain LAI was also an IL-10 inducer, but gp120 from the strain KNH1144 was not. The mannose-binding protein cyanovirin-N, the 2G12 mAb to a mannose-dependent gp120 epitope, and MCLR-specific mAbs inhibited IL-10 expression, as did enzymatic removal of gp120 mannose moieties, whereas inhibitors of signaling via CD4, CCR5, or CXCR4 were ineffective. Gp120-stimulated IL-10 production correlated with DC-SIGN expression on the cells, and involved the ERK signaling pathway. Gp120-treated MDDCs also responded poorly to maturation stimuli by up-regulating activation markers inefficiently and stimulating allogeneic T cell proliferation only weakly. These adverse reactions to gp120 were MCLR-dependent but independent of IL-10 production. Since such mechanisms might suppress immune responses to Env-containing vaccines, demannosylation may be a way to improve the immunogenicity of gp120 or gp140 proteins.

  7. Molecular recognition of CCR5 by an HIV-1 gp120 V3 loop.

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    Phanourios Tamamis

    Full Text Available The binding of protein HIV-1 gp120 to coreceptors CCR5 or CXCR4 is a key step of the HIV-1 entry to the host cell, and is predominantly mediated through the V3 loop fragment of HIV-1 gp120. In the present work, we delineate the molecular recognition of chemokine receptor CCR5 by a dual tropic HIV-1 gp120 V3 loop, using a comprehensive set of computational tools predominantly based on molecular dynamics simulations and free energy calculations. We report, what is to our knowledge, the first complete HIV-1 gp120 V3 loop : CCR5 complex structure, which includes the whole V3 loop and the N-terminus of CCR5, and exhibits exceptional agreement with previous experimental findings. The computationally derived structure sheds light into the functional role of HIV-1 gp120 V3 loop and CCR5 residues associated with the HIV-1 coreceptor activity, and provides insights into the HIV-1 coreceptor selectivity and the blocking mechanism of HIV-1 gp120 by maraviroc. By comparing the binding of the specific dual tropic HIV-1 gp120 V3 loop with CCR5 and CXCR4, we observe that the HIV-1 gp120 V3 loop residues 13-21, which include the tip, share nearly identical structural and energetic properties in complex with both coreceptors. This result paves the way for the design of dual CCR5/CXCR4 targeted peptides as novel potential anti-AIDS therapeutics.

  8. Structural Basis for Species Selectivity in the HIV-1 gp120-CD4 Interaction: Restoring Affinity to gp120 in Murine CD4 Mimetic Peptides

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    Kristin Kassler

    2011-01-01

    Full Text Available The first step of HIV-1 infection involves interaction between the viral glycoprotein gp120 and the human cellular receptor CD4. Inhibition of the gp120-CD4 interaction represents an attractive strategy to block HIV-1 infection. In an attempt to explore the known lack of affinity of murine CD4 to gp120, we have investigated peptides presenting the putative gp120-binding site of murine CD4 (mCD4. Molecular modeling indicates that mCD4 protein cannot bind gp120 due to steric clashes, while the larger conformational flexibility of mCD4 peptides allows an interaction. This finding is confirmed by experimental binding assays, which also evidenced specificity of the peptide-gp120 interaction. Molecular dynamics simulations indicate that the mCD4-peptide stably interacts with gp120 via an intermolecular β-sheet, while an important salt-bridge formed by a C-terminal lysine is lost. Fixation of the C-terminus by introducing a disulfide bridge between the N- and C-termini of the peptide significantly enhanced the affinity to gp120.

  9. Molecular mechanism of HIV-1 gp120 mutations that reduce CD4 binding affinity.

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    Kassler, Kristin; Sticht, Heinrich

    2014-01-01

    The interaction of the HIV-1 fusion protein gp120 with its cellular receptor CD4 represents a crucial step of the viral infection process, thus rendering gp120 a promising target for the intervention with anti-HIV drugs. Naturally occurring mutations of gp120, however, can decrease its affinity for anti-infective ligands like therapeutic antibodies or soluble CD4. To understand this phenomenon on a structural level, we performed molecular dynamics simulations of two gp120 variants (termed gp1203-2 and gp1202-1), which exhibit a significantly decreased binding of soluble CD4. In both variants, the exchange of a nonpolar residue by glutamate was identified as an important determinant for reduced binding. However, those glutamates are located at different sequence positions and affect different steps of the recognition process: E471 in gp1203-2 predominantly affects the CD4-bound conformation, whereas E372 in gp1202-1 mainly modulates the conformational sampling of free gp120. Despite these differences, there exists an interesting similarity between the two variants: both glutamates exert their function by modulating the conformation and interactions of glycine-rich motifs (G366-G367, G471-G473) resulting in an accumulation of binding incompetent gp120 conformations or a loss of intermolecular gp120-CD4 hydrogen bonds. Thus, the present data suggests that interference with the structure and dynamics of glycine-rich stretches might represent a more widespread mechanism, by which gp120 mutations reduce binding affinity. This knowledge should be helpful to predict the resistance of novel gp120 mutations or to design gp120-ligands with improved binding properties. An animated interactive 3D complement (I3DC) is available in Proteopedia at http://proteopedia.org/w/Journal:JBSD:41.

  10. Extracellular Matrix Proteins Mediate HIV-1 gp120 Interactions with α4β7.

    Science.gov (United States)

    Plotnik, David; Guo, Wenjin; Cleveland, Brad; von Haller, Priska; Eng, Jimmy K; Guttman, Miklos; Lee, Kelly K; Arthos, James; Hu, Shiu-Lok

    2017-11-01

    Gut-homing α4β7high CD4+ T lymphocytes have been shown to be preferentially targeted by human immunodeficiency virus type 1 (HIV-1) and are implicated in HIV-1 pathogenesis. Previous studies demonstrated that HIV-1 envelope protein gp120 binds and signals through α4β7 and that this likely contributes to the infection of α4β7high T cells and promotes cell-to-cell virus transmission. Structures within the second variable loop (V2) of gp120, including the tripeptide motif LDV/I, are thought to mediate gp120-α4β7 binding. However, lack of α4β7 binding has been reported in gp120 proteins containing LDV/I, and the precise determinants of gp120-α4β7 binding are not fully defined. In this work, we report the novel finding that fibronectins mediate indirect gp120-α4β7 interactions. We show that Chinese hamster ovary (CHO) cells used to express recombinant gp120 produced fibronectins and other extracellular matrix proteins that copurified with gp120. CHO cell fibronectins were able to mediate the binding of a diverse panel of gp120 proteins to α4β7 in an in vitro cell binding assay. The V2 loop was not required for fibronectin-mediated binding of gp120 to α4β7, nor did V2-specific antibodies block this interaction. Removal of fibronectin through anion-exchange chromatography abrogated V2-independent gp120-α4β7 binding. Additionally, we showed a recombinant human fibronectin fragment mediated gp120-α4β7 interactions similarly to CHO cell fibronectin. These findings provide an explanation for the apparently contradictory observations regarding the gp120-α4β7 interaction and offer new insights into the potential role of fibronectin and other extracellular matrix proteins in HIV-1 biology.IMPORTANCE Immune tissues within the gut are severely damaged by HIV-1, and this plays an important role in the development of AIDS. Integrin α4β7 plays a major role in the trafficking of lymphocytes, including CD4+ T cells, into gut lymphoid tissues. Previous reports

  11. Transmission routes of HIV-1 gp120 from brain to lymphoid tissues.

    Science.gov (United States)

    Cashion, M F; Banks, W A; Bost, K L; Kastin, A J

    1999-03-20

    The blood-brain barrier (BBB) restricts the entry of antiviral agents into the CNS thereby facilitating the creation of a reservoir of HIV that could potentially reinfect peripheral tissues. We characterized the efflux from brain of radioactively labeled viral coat HIV-1 gp120 (I-gp120) after intracerebroventricular (i.c.v.) injection. The half-time disappearance rate of I-gp120 from brain was 12.6 min, which was faster than could be explained by the reabsorption of cerebrospinal fluid into blood but could not be explained by a saturable transporter. After i.c.v. injection, I-gp120 appeared in the serum and was sequestered by spleen and the cervical nodes, demonstrating a potential for virus within the CNS to reinfect peripheral tissues. However, the amount of I-gp120 appearing in serum was less than that expected based on the efflux rate, whereas uptake by the cervical nodes was much greater after i. c.v. than after i.v. injection of I-gp120. These findings were explained by drainage from the brain directly to the cervical lymph nodes through the brain's primitive lymphatic system. These lymphatics potentially provide a pathway through which CNS reservoirs of HIV-1 could directly reinfect lymphoid tissue without being exposed to circulating antiviral agents. Copyright 1999 Elsevier Science B.V.

  12. CB2 receptor agonists protect human dopaminergic neurons against damage from HIV-1 gp120.

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    Shuxian Hu

    Full Text Available Despite the therapeutic impact of anti-retroviral therapy, HIV-1-associated neurocognitive disorder (HAND remains a serious threat to AIDS patients, and there currently remains no specific therapy for the neurological manifestations of HIV-1. Recent work suggests that the nigrostriatal dopaminergic area is a critical brain region for the neuronal dysfunction and death seen in HAND and that human dopaminergic neurons have a particular sensitivity to gp120-induced damage, manifested as reduced function (decreased dopamine uptake, morphological changes, and reduced viability. Synthetic cannabinoids inhibit HIV-1 expression in human microglia, suppress production of inflammatory mediators in human astrocytes, and there is substantial literature demonstrating the neuroprotective properties of cannabinoids in other neuropathogenic processes. Based on these data, experiments were designed to test the hypothesis that synthetic cannabinoids will protect dopaminergic neurons against the toxic effects of the HIV-1 protein gp120. Using a human mesencephalic neuronal/glial culture model, which contains dopaminergic neurons, microglia, and astrocytes, we were able to show that the CB1/CB2 agonist WIN55,212-2 blunts gp120-induced neuronal damage as measured by dopamine transporter function, apoptosis and lipid peroxidation; these actions were mediated principally by the CB2 receptor. Adding supplementary human microglia to our cultures enhances gp120-induced damage; WIN55,212-2 is able to alleviate this enhanced damage. Additionally, WIN55,212-2 inhibits gp120-induced superoxide production by purified human microglial cells, inhibits migration of human microglia towards supernatants generated from gp120-stimulated human mesencephalic neuronal/glial cultures and reduces chemokine and cytokine production from the human mesencephalic neuronal/glial cultures. These data suggest that synthetic cannabinoids are capable of protecting human dopaminergic neurons from

  13. Curcumin improves synaptic plasticity impairment induced by HIV-1gp120 V3 loop

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    Ling-ling Shen

    2015-01-01

    Full Text Available Curcumin has been shown to significantly improve spatial memory impairment induced by HIV-1 gp120 V3 in rats, but the electrophysiological mechanism remains unknown. Using extracellular microelectrode recording techniques, this study confirmed that the gp120 V3 loop could suppress long-term potentiation in the rat hippocampal CA1 region and synaptic plasticity, and that curcumin could antagonize these inhibitory effects. Using a Fura-2/AM calcium ion probe, we found that curcumin resisted the effects of the gp120 V3 loop on hippocampal synaptosomes and decreased Ca 2+ concentration in synaptosomes. This effect of curcumin was identical to nimodipine, suggesting that curcumin improved the inhibitory effects of gp120 on synaptic plasticity, ameliorated damage caused to the central nervous system, and might be a potential neuroprotective drug.

  14. A Wnt5a signaling pathway in the pathogenesis of HIV-1 gp120-induced pain.

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    Yuan, Su-Bo; Ji, Guangchen; Li, Bei; Andersson, Tommy; Neugebauer, Volker; Tang, Shao-Jun

    2015-07-01

    Pathological pain is one of the most common neurological complications in patients with HIV-1/AIDS. However, the pathogenic process is unclear. Our recent studies show that Wnt5a is upregulated in the spinal cord dorsal horn (SDH) of the patients with HIV who develop pain and that HIV-1 gp120, a potential causal factor of the HIV-associated pain, rapidly upregulates Wnt5a in the mouse SDH. Using a mouse model, we show here that a specific Wnt5a antagonist, Box-5, attenuated gp120-induced mechanical allodynia. Conversely, a Wnt5a agonist, Foxy5, facilitated the allodynia. To elucidate the molecular mechanism by which Wnt5a regulates gp120-induced allodynia, we tested the role of the JNK/TNF-α pathway. We observed that the JNK-specific inhibitor SP600125 blocked either gp120- or Foxy5-induced allodynia. Similarly, the TNF-α-specific antagonist Enbrel also reversed either gp120- or Foxy5-induced allodynia. These data suggest that JNK and TNF-α mediate the biological effects of Wnt5a in regulating gp120-induced allodynia. To investigate the cellular mechanism, we performed extracellular single-unit recording from SDH neurons in anesthetized mice. Both Box-5 and SP600125 negated gp120-induced potentiation of SDH neuron spiking evoked by mechanical stimulation of the hind paw. Furthermore, while Foxy5 potentiated spike frequency of SDH neurons, either SP600125 or Enbrel blocked the potentiation. The data indicate that Wnt5a potentiates the activity of SDH neurons through the JNK-TNF-α pathway. Collectively, our findings suggest that Wnt5a regulates the pathogenesis of gp120-induced pain, likely by sensitizing pain-processing SDH neurons through JNK/TNF-α signaling.

  15. Gp120 on HIV-1 Virions Lacks O-Linked Carbohydrate.

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    Elizabeth Stansell

    Full Text Available As HIV-1-encoded envelope protein traverses the secretory pathway, it may be modified with N- and O-linked carbohydrate. When the gp120s of HIV-1 NL4-3, HIV-1 YU2, HIV-1 Bal, HIV-1 JRFL, and HIV-1 JRCSF were expressed as secreted proteins, the threonine at consensus position 499 was found to be O-glycosylated. For SIVmac239, the corresponding threonine was also glycosylated when gp120 was recombinantly expressed. Similarly-positioned, highly-conserved threonines in the influenza A virus H1N1 HA1 and H5N1 HA1 envelope proteins were also found to carry O-glycans when expressed as secreted proteins. In all cases, the threonines were modified predominantly with disialylated core 1 glycans, together with related core 1 and core 2 structures. Secreted HIV-1 gp140 was modified to a lesser extent with mainly monosialylated core 1 O-glycans, suggesting that the ectodomain of the gp41 transmembrane component may limit the accessibility of Thr499 to glycosyltransferases. In striking contrast to these findings, gp120 on purified virions of HIV-1 Bal and SIV CP-MAC lacked any detectable O-glycosylation of the C-terminal threonine. Our results indicate the absence of O-linked carbohydrates on Thr499 as it exists on the surface of virions and suggest caution in the interpretation of analyses of post-translational modifications that utilize recombinant forms of envelope protein.

  16. Antibodies to the CD4-binding site of HIV-1 gp120 suppress gp120-specific CD4 T cell response while enhancing antibody response

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    Hioe Catarina E

    2008-07-01

    Full Text Available Abstract Background The binding of Abs to the CD4-binding site (CD4bs of HIV-1 envelope gp120 has been shown to obstruct the processing and generation of helper epitopes from this antigen, resulting in poor presentation of various gp120 epitopes by MHC class II to CD4 T cells. However, the physiologic significance of these inhibitory anti-CD4bs Abs in vivo has remained unclear. In this study, we evaluated the immunologic effects of anti-CD4bs Abs in vivo using a murine model. Results Animals were immunized with recombinant envelope proteins with or without CD4-binding activity (designated CD4bs+ Env and CD4bs– Env, respectively. As expected, anti-CD4bs Abs were generated only after immunization with CD4bs+ Env and not with CD4bs– Env. The presence of anti-CD4bs Abs was associated with lower levels of envelope-specific lymphoproliferation in animals immunized with CD4bs+ Env. To further determine the specific role of the anti-CD4bs Abs, we immunized mice with gp120 in the presence of an inhibitory anti-CD4bs mAb or a non-inhibitory anti-gp120 mAb. The data show that the presence of anti-CD4bs mAb reduced CD4 T cell responses to gp120. However, we also detected significantly higher titers of anti-gp120 Abs following immunization with gp120 and the anti-CD4bs mAb. Conclusion Anti-CD4bs Abs can exert discordant effects on the gp120-specific CD4 T cell and Ab responses in vivo, indicating the importance of these particular Abs in influencing both the cellular and the humoral immune responses against HIV-1.

  17. Engineering and exploitation of a fluorescent HIV-1 gp120 for live cell CD4 binding assays

    Energy Technology Data Exchange (ETDEWEB)

    Costantini, Lindsey M. [Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461 (United States); Irvin, Susan C. [Department of Pediatrics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461 (United States); Department of Microbiology and Immunology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461 (United States); Kennedy, Steven C. [Department of Pediatrics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461 (United States); Guo, Feng [Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461 (United States); Goldstein, Harris; Herold, Betsy C. [Department of Pediatrics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461 (United States); Department of Microbiology and Immunology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461 (United States); Snapp, Erik L., E-mail: erik-lee.snapp@einstein.yu.edu [Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461 (United States)

    2015-02-15

    The HIV-1 envelope glycoprotein, gp120, binds the host cell receptor, CD4, in the initial step of HIV viral entry and infection. This process is an appealing target for the development of inhibitory drugs and neutralizing antibodies. To study gp120 binding and intracellular trafficking, we engineered a fluorescent fusion of the humanized gp120 JRFL HIV-1 variant and GFP. Gp120-sfGFP is glycosylated with human sugars, robustly expressed, and secreted from cultured human cells. Protein dynamics, quality control, and trafficking can be visualized in live cells. The fusion protein can be readily modified with different gp120 variants or fluorescent proteins. Finally, secreted gp120-sfGFP enables a sensitive and easy binding assay that can quantitatively screen potential inhibitors of gp120-CD4 binding on live cells via fluorescence imaging or laser scanning cytometry. This adaptable research tool should aid in studies of gp120 cell biology and the development of novel anti-HIV drugs. - Highlights: • Development of fluorescent protein labeled HIV-1 envelope gp120. • Imaging of gp120 dynamics and trafficking in live cells. • Quantitative visual assay of antibody-mediated inhibition of gp120 binding to CD4 on live cells.

  18. Molecular characterization of HIV-1 subtype C gp-120 regions potentially involved in virus adaptive mechanisms.

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    Alessandra Cenci

    Full Text Available The role of variable regions of HIV-1 gp120 in immune escape of HIV has been investigated. However, there is scant information on how conserved gp120 regions contribute to virus escaping. Here we have studied how molecular sequence characteristics of conserved C3, C4 and V3 regions of clade C HIV-1 gp120 that are involved in HIV entry and are target of the immune response, are modulated during the disease course. We found an increase of "shifting" putative N-glycosylation sites (PNGSs in the α2 helix (in C3 and in C4 and an increase of sites under positive selection pressure in the α2 helix during the chronic stage of disease. These sites are close to CD4 and to co-receptor binding sites. We also found a negative correlation between electric charges of C3 and V4 during the late stage of disease counteracted by a positive correlation of electric charges of α2 helix and V5 during the same stage. These data allow us to hypothesize possible mechanisms of virus escape involving constant and variable regions of gp120. In particular, new mutations, including new PNGSs occurring near the CD4 and CCR5 binding sites could potentially affect receptor binding affinity and shield the virus from the immune response.

  19. HLA-C increases HIV-1 infectivity and is associated with gp120

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    Beretta Alberto

    2008-08-01

    Full Text Available Abstract Background A recently identified genetic polymorphism located in the 5' region of the HLA-C gene is associated with individual variations in HIV-1 viral load and with differences in HLA-C expression levels. HLA-C has the potential to restrict HIV-1 by presenting epitopes to cytotoxic T cells but it is also a potent inhibitor of NK cells. In addition, HLA-C molecules incorporated within the HIV-1 envelope have been shown to bind to the envelope glycoprotein gp120 and enhance viral infectivity. We investigated this last property in cell fusion assays where the expression of HLA-C was silenced by small interfering RNA sequences. Syncytia formation was analyzed by co-cultivating cell lines expressing HIV-1 gp120/gp41 from different laboratory and primary isolates with target cells expressing different HIV-1 co-receptors. Virus infectivity was analyzed using pseudoviruses. Molecular complexes generated during cell fusion (fusion complexes were purified and analyzed for their HLA-C content. Results HLA-C positive cells co-expressing HIV-1 gp120/gp41 fused more rapidly and produced larger syncytia than HLA-C negative cells. Transient transfection of gp120/gp41 from different primary isolates in HLA-C positive cells resulted in a significant cell fusion increase. Fusion efficiency was reduced in HLA-C silenced cells compared to non-silenced cells when co-cultivated with different target cell lines expressing HIV-1 co-receptors. Similarly, pseudoviruses produced from HLA-C silenced cells were significantly less infectious. HLA-C was co-purified with gp120 from cells before and after fusion and was associated with the fusion complex. Conclusion Virionic HLA-C molecules associate to Env and increase the infectivity of both R5 and X4 viruses. Genetic polymorphisms associated to variations in HLA-C expression levels may therefore influence the individual viral set point not only by means of a regulation of the virus-specific immune response but also

  20. Insight derived from molecular dynamics simulations into molecular motions, thermodynamics and kinetics of HIV-1 gp120.

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    Peng Sang

    Full Text Available Although the crystal structures of the HIV-1 gp120 core bound and pre-bound by CD4 are known, the details of dynamics involved in conformational equilibrium and transition in relation to gp120 function have remained elusive. The homology models of gp120 comprising the N- and C-termini and loops V3 and V4 in the CD4-bound and CD4-unbound states were built and subjected to molecular dynamics (MD simulations to investigate the differences in dynamic properties and molecular motions between them. The results indicate that the CD4-bound gp120 adopted a more compact and stable conformation than the unbound form during simulations. For both the unbound and bound gp120, the large concerted motions derived from essential dynamics (ED analyses can influence the size/shape of the ligand-binding channel/cavity of gp120 and, therefore, were related to its functional properties. The differences in motion direction between certain structural components of these two forms of gp120 were related to the conformational interconversion between them. The free energy calculations based on the metadynamics simulations reveal a more rugged and complex free energy landscape (FEL for the unbound than for the bound gp120, implying that gp120 has a richer conformational diversity in the unbound form. The estimated free energy difference of ∼-6.0 kJ/mol between the global minimum free energy states of the unbound and bound gp120 indicates that gp120 can transform spontaneously from the unbound to bound states, revealing that the bound state represents a high-probability "ground state" for gp120 and explaining why the unbound state resists crystallization. Our results provide insight into the dynamics-and-function relationship of gp120, and facilitate understandings of the thermodynamics, kinetics and conformational control mechanism of HIV-1 gp120.

  1. Structure-based Design of Cyclically Permuted HIV-1 gp120 Trimers That Elicit Neutralizing Antibodies.

    Science.gov (United States)

    Kesavardhana, Sannula; Das, Raksha; Citron, Michael; Datta, Rohini; Ecto, Linda; Srilatha, Nonavinakere Seetharam; DiStefano, Daniel; Swoyer, Ryan; Joyce, Joseph G; Dutta, Somnath; LaBranche, Celia C; Montefiori, David C; Flynn, Jessica A; Varadarajan, Raghavan

    2017-01-06

    A major goal for HIV-1 vaccine development is an ability to elicit strong and durable broadly neutralizing antibody (bNAb) responses. The trimeric envelope glycoprotein (Env) spikes on HIV-1 are known to contain multiple epitopes that are susceptible to bNAbs isolated from infected individuals. Nonetheless, all trimeric and monomeric Env immunogens designed to date have failed to elicit such antibodies. We report the structure-guided design of HIV-1 cyclically permuted gp120 that forms homogeneous, stable trimers, and displays enhanced binding to multiple bNAbs, including VRC01, VRC03, VRC-PG04, PGT128, and the quaternary epitope-specific bNAbs PGT145 and PGDM1400. Constructs that were cyclically permuted in the V1 loop region and contained an N-terminal trimerization domain to stabilize V1V2-mediated quaternary interactions, showed the highest homogeneity and the best antigenic characteristics. In guinea pigs, a DNA prime-protein boost regimen with these new gp120 trimer immunogens elicited potent neutralizing antibody responses against highly sensitive Tier 1A isolates and weaker neutralizing antibody responses with an average titer of about 115 against a panel of heterologous Tier 2 isolates. A modest fraction of the Tier 2 virus neutralizing activity appeared to target the CD4 binding site on gp120. These results suggest that cyclically permuted HIV-1 gp120 trimers represent a viable platform in which further modifications may be made to eventually achieve protective bNAb responses. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  2. HIV-1 gp120 and Morphine Induced Oxidative Stress: Role in Cell Cycle Regulation

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    Samikkannu eThangavel

    2015-06-01

    Full Text Available HIV infection and illicit drugs are known to induce oxidative stress and linked with severity of viral replication, disease progression, impaired cell cycle regulation and neurodegeneration. Studies have shown that morphine accelerates HIV infection and disease progression mediated by Reactive oxygen species (ROS. Oxidative stress impact redox balance and ROS production affect cell cycle regulation. However, the role of morphine in HIV associated acceleration of oxidative stress and its link to cell cycle regulation and neurodegeneration has not been elucidated. The aim of present study is to elucidate the mechanism of oxidative stress induced glutathione synthases (GSS, super oxide dismutase (SOD, and glutathione peroxidase (GPx impact cell cycle regulated protein cyclin-dependent kinase 1, cell division cycle 2 (CDK-1/CDC-2, cyclin B, and cell division cycle 25C (CDC-25C influencing neuronal dysfunction by morphine co-morbidity with HIV-1 gp120. It was observed that redox imbalance inhibited the GSS, GPx and increased SOD which, subsequently inhibited CDK-1/CDC-2 whereas cyclin B and CDC-25C significantly up regulated in HIV-1 gp120 with morphine compared to either HIV-1 gp120 or morphine treated alone in human microglial cell line. These results suggest that HIV positive morphine users have increased levels of oxidative stress and effect of cell cycle machinery, which may cause the HIV infection and disease progression.

  3. Human antibody response to a strain-specific HIV-1 gp120 epitope associated with cell fusion inhibition

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    Goudsmit, J.; Boucher, C. A.; Meloen, R. H.; Epstein, L. G.; Smit, L.; van der Hoek, L.; Bakker, M.

    1988-01-01

    PEPSCAN analysis, performed using 536 overlapping nonapeptides derived from the HTLV-III B nucleotide sequence of the region encoding the external envelope protein of 120 kDa (gp120), identified in the V3 region of gp120 a major binding site for antibodies of HIV-1-infected humans. The minimal amino

  4. Are blockers of gp120/CD4 interaction effective inhibitors of HIV-1 immunopathogenesis?

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    Herbeuval, Jean-Philippe; Shearer, Gene M

    2006-01-01

    The immunopathogenic mechanisms that result in the depletion of CD4+ T-cells after HIV-1 infection remain controversial. We consider here mechanisms that have been suggested, and propose a data-supported model in which CD4+ T-cells undergo apoptosis that is signaled by the binding of viral gp120 to cellular CD4. Blood leucocytes from HIV-1-uninfected donors, including CD4+ and CD8+ T-cells, monocytes, myeloid and plasmacytoid dendritic cells (pDC) were cultured with either infectious or noninfectious HIV-1. The cultures were tested for expression of interferon-alpha, TRAIL, DR5 and apoptosis. Inhibitors of IFNalpha, TRAIL, DR5 and gp120/CD4 binding were added to the cultures. Ex vivo studies were performed using peripheral blood mononuclear cells (PBMC) from HIV-1-infected patients to test the validity of our in vitro findings. Both infectious and noninfectious HIV-1 induced pDC to produce IFNalpha, which induced expression of TRAIL by CD4+ but not CD8+ T-cells. CD4+ T-cells expressed the TRAIL death receptor 5 (DR5), upon HIV-1 binding to CD4. Antibodies against TRAIL and DR5 partly inhibited apoptosis. However, soluble CD4 (sCD4-IgG) efficiently blocked IFNalpha production, TRAIL and DR5 expression and apoptosis of T helper cells. Studies of HIV-1-infected patients' PBMC indicated increased plasma TRAIL production and CD4+ T-cell DR5 expression, which correlated directly with viral load and inversely with CD4 count. Noninfectious interactions between HIV-1 and CD4 are major contributors to CD4+ T-cell death via IFNalpha-induced TRAIL expression and HIV-1-induced DR5 expression on CD4+ T-cells. Since noninfectious as well as infectious HIV-1 induces the death cascade resulting in selective apoptosis of CD4+ T-cells, these HIV-1/CD4-dependent binding events would not necessarily be reflected in HIV-1 RNA and DNA expression by the CD4+ target T-cells. Because each step of this model leading to apoptosis requires the binding of gp120 to CD4, we suggest that molecules

  5. Curcumin protects microglia and primary rat cortical neurons against HIV-1 gp120-mediated inflammation and apoptosis.

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    Luyan Guo

    Full Text Available Curcumin is a molecule found in turmeric root that has anti-inflammatory, antioxidant, and anti-tumor properties and has been widely used as both an herbal drug and a food additive to treat or prevent neurodegenerative diseases. To explore whether curcumin is able to ameliorate HIV-1-associated neurotoxicity, we treated a murine microglial cell line (N9 and primary rat cortical neurons with curcumin in the presence or absence of neurotoxic HIV-1 gp120 (V3 loop protein. We found that HIV-1 gp120 profoundly induced N9 cells to produce reactive oxygen species (ROS, tumor necrosis factor-α (TNF-α and monocyte chemoattractant protein-1 (MCP-1. HIV-1 gp120 also induced apoptosis of primary rat cortical neurons. Curcumin exerted a powerful inhibitory effect against HIV-1 gp120-induced neuronal damage, reducing the production of ROS, TNF-α and MCP-1 by N9 cells and inhibiting apoptosis of primary rat cortical neurons. Curcumin may exert its biological activities through inhibition of the delayed rectification and transient outward potassium (K(+ current, as curcumin effectively reduced HIV-1 gp120-mediated elevation of the delayed rectification and transient outward K(+ channel current in neurons. We conclude that HIV-1 gp120 increases ROS, TNF-α and MCP-1 production in microglia, and induces cortical neuron apoptosis by affecting the delayed rectification and transient outward K(+ channel current. Curcumin reduces production of ROS and inflammatory mediators in HIV-1-gp120-stimulated microglia, and protects cortical neurons against HIV-1-mediated apoptosis, most likely through inhibition of HIV-1 gp120-induced elevation of the delayed rectification and transient outward K(+ current.

  6. Cocaine enhances HIV-1 gp120-induced lymphatic endothelial dysfunction in the lung.

    Science.gov (United States)

    Zhang, Xuefeng; Jiang, Susan; Yu, Jinlong; Kuzontkoski, Paula M; Groopman, Jerome E

    2015-08-01

    Pulmonary complications are common in both AIDS patients and cocaine users. We addressed the cellular and molecular mechanisms by which HIV and cocaine may partner to induce their deleterious effects. Using primary lung lymphatic endothelial cells (L-LECs), we examined how cocaine and HIV-1 gp120, alone and together, modulate signaling and functional properties of L-LECs. We found that brief cocaine exposure activated paxillin and induced cytoskeletal rearrangement, while sustained exposure increased fibronectin (FN) expression, decreased Robo4 expression, and enhanced the permeability of L-LEC monolayers. Moreover, incubating L-LECs with both cocaine and HIV-1 gp120 exacerbated hyperpermeability, significantly enhanced apoptosis, and further impaired in vitro wound healing as compared with cocaine alone. Our studies also suggested that the sigma-1 receptor (Sigma-1R) and the dopamine-4 receptor (D4R) are involved in cocaine-induced pathology in L-LECs. Seeking clinical correlation, we found that FN levels in sera and lung tissue of HIV(+) donors were significantly elevated as compared to HIV(-) donors. Our in vitro data demonstrate that cocaine and HIV-1 gp120 induce dysfunction and damage of lung lymphatics, and suggest that cocaine use may exacerbate pulmonary edema and fibrosis associated with HIV infection. Continued exploration of the interplay between cocaine and HIV should assist the design of therapeutics to ameliorate HIV-induced pulmonary disorders within the drug using population. © 2015 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.

  7. Immunogenicity of a polyvalent HIV-1 candidate vaccine based on fourteen wild type gp120 proteins in golden hamsters

    Science.gov (United States)

    Azizi, Ali; Anderson, David E; Ghorbani, Masoud; Gee, Katrina; Diaz-Mitoma, Francisco

    2006-01-01

    Background One of the major obstacles in the design of an effective vaccine against HIV-1 is the hypervariability of the HIV-1 envelope glycoprotein. Most HIV-1 vaccine candidates have utilized envelope glycoprotein from a single virus isolate, but to date, none of them elicited broadly reactive humoral immunity. Herein, we hypothesised that a cocktail of HIV-1 gp120 proteins containing multiple epitopes may increase the breadth of immune responses against HIV-1. We compared and evaluated the immunogenicity of HIV-1 vaccines containing either gp120 protein alone or in combinations of four or fourteen gp120s from different primary HIV-1 isolates in immunized hamsters. Results We amplified and characterized 14 different gp120s from primary subtype B isolates with both syncytium and non-syncytium inducing properties, and expressed the proteins in Chinese Hamster Ovary (CHO) cell lines. Purified proteins were used either alone or in combinations of four or fourteen different gp120s to vaccinate golden hamsters. The polyvalent vaccine showed higher antibody titers to HIV-1 subtype B isolates MN and SF162 compared to the groups that received one or four gp120 proteins. However, the polyvalent vaccine was not able to show higher neutralizing antibody responses against HIV-1 primary isolates. Interestingly, the polyvalent vaccine group had the highest proliferative immune responses and showed a substantial proportion of cross-subtype CD4 reactivity to HIV-1 subtypes B, C, and A/E Conclusion Although the polyvalent approach achieved only a modest increase in the breadth of humoral and cellular immunity, the qualitative change in the vaccine (14 vs. 1 gp120) resulted in a quantitative improvement in vaccine-induced immunity. PMID:17076905

  8. Immunogenicity of a polyvalent HIV-1 candidate vaccine based on fourteen wild type gp120 proteins in golden hamsters

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    Ghorbani Masoud

    2006-10-01

    Full Text Available Abstract Background One of the major obstacles in the design of an effective vaccine against HIV-1 is the hypervariability of the HIV-1 envelope glycoprotein. Most HIV-1 vaccine candidates have utilized envelope glycoprotein from a single virus isolate, but to date, none of them elicited broadly reactive humoral immunity. Herein, we hypothesised that a cocktail of HIV-1 gp120 proteins containing multiple epitopes may increase the breadth of immune responses against HIV-1. We compared and evaluated the immunogenicity of HIV-1 vaccines containing either gp120 protein alone or in combinations of four or fourteen gp120s from different primary HIV-1 isolates in immunized hamsters. Results We amplified and characterized 14 different gp120s from primary subtype B isolates with both syncytium and non-syncytium inducing properties, and expressed the proteins in Chinese Hamster Ovary (CHO cell lines. Purified proteins were used either alone or in combinations of four or fourteen different gp120s to vaccinate golden hamsters. The polyvalent vaccine showed higher antibody titers to HIV-1 subtype B isolates MN and SF162 compared to the groups that received one or four gp120 proteins. However, the polyvalent vaccine was not able to show higher neutralizing antibody responses against HIV-1 primary isolates. Interestingly, the polyvalent vaccine group had the highest proliferative immune responses and showed a substantial proportion of cross-subtype CD4 reactivity to HIV-1 subtypes B, C, and A/E Conclusion Although the polyvalent approach achieved only a modest increase in the breadth of humoral and cellular immunity, the qualitative change in the vaccine (14 vs. 1 gp120 resulted in a quantitative improvement in vaccine-induced immunity.

  9. Intrathecal HIV-1 envelope glycoprotein gp120 induces enhanced pain states mediated by spinal cord proinflammatory cytokines.

    Science.gov (United States)

    Milligan, E D; O'Connor, K A; Nguyen, K T; Armstrong, C B; Twining, C; Gaykema, R P; Holguin, A; Martin, D; Maier, S F; Watkins, L R

    2001-04-15

    Perispinal (intrathecal) injection of the human immunodeficiency virus-1 (HIV-1) envelope glycoprotein gp120 creates exaggerated pain states. Decreases in response thresholds to both heat stimuli (thermal hyperalgesia) and light tactile stimuli (mechanical allodynia) are rapidly induced after gp120 administration. gp120 is the portion of HIV-1 that binds to and activates microglia and astrocytes. These glial cells have been proposed to be key mediators of gp120-induced hyperalgesia and allodynia because these pain changes are blocked by drugs thought to affect glial function preferentially. The aim of the present series of studies was to determine whether gp120-induced pain changes involve proinflammatory cytokines [interleukin-1beta (IL-1) and tumor necrosis factor-alpha (TNF-alpha)], substances released from activated glia. IL-1 and TNF antagonists each prevented gp120-induced pain changes. Intrathecal gp120 produced time-dependent, site-specific increases in TNF and IL-1 protein release into lumbosacral CSF; parallel cytokine increases in lumbar dorsal spinal cord were also observed. Intrathecal administration of fluorocitrate (a glial metabolic inhibitor), TNF antagonist, and IL-1 antagonist each blocked gp120-induced increases in spinal IL-1 protein. These results support the concept that activated glia in dorsal spinal cord can create exaggerated pain states via the release of proinflammatory cytokines.

  10. Engineering and exploitation of a fluorescent HIV-1 gp120 for live cell CD4 binding assays.

    Science.gov (United States)

    Costantini, Lindsey M; Irvin, Susan C; Kennedy, Steven C; Guo, Feng; Goldstein, Harris; Herold, Betsy C; Snapp, Erik L

    2015-02-01

    The HIV-1 envelope glycoprotein, gp120, binds the host cell receptor, CD4, in the initial step of HIV viral entry and infection. This process is an appealing target for the development of inhibitory drugs and neutralizing antibodies. To study gp120 binding and intracellular trafficking, we engineered a fluorescent fusion of the humanized gp120 JRFL HIV-1 variant and GFP. Gp120-sfGFP is glycosylated with human sugars, robustly expressed, and secreted from cultured human cells. Protein dynamics, quality control, and trafficking can be visualized in live cells. The fusion protein can be readily modified with different gp120 variants or fluorescent proteins. Finally, secreted gp120-sfGFP enables a sensitive and easy binding assay that can quantitatively screen potential inhibitors of gp120-CD4 binding on live cells via fluorescence imaging or laser scanning cytometry. This adaptable research tool should aid in studies of gp120 cell biology and the development of novel anti-HIV drugs. Copyright © 2014 Elsevier Inc. All rights reserved.

  11. Topological Analysis of HIV-1 Glycoproteins Expressed In Situ on Virus Surfaces Reveals Tighter Packing but Greater Conformational Flexibility than for Soluble gp120

    Science.gov (United States)

    Tong, Tommy; Osawa, Keiko; Robinson, James E.; Crooks, Ema T.

    2013-01-01

    In natural infection, antibodies interact with HIV-1 primarily through nonfunctional forms of envelope glycoproteins (Env), including uncleaved (UNC) gp160 and gp41 stumps. These antigens are important to fully characterize, as they may be decoys that promote nonneutralizing responses and may also be targets for nonneutralizing effector responses. In this study, we compared the antigenic properties of Env expressed in situ on pseudovirion virus-like particle (VLP) surfaces and soluble gp120 using harmonized enzyme-linked immunosorbent assays (ELISAs) and a panel of 51 monoclonal antibodies (MAbs). Only 32 of 46 soluble gp120-reactive MAbs recognized the primary UNC gp160 antigen of VLPs. Indeed, many epitopes were poorly exposed (C1, V2, C1-C4, C4, C4-V3, CD4 induced [CD4i], and PGT group 3) or obscured (C2, C5, and C1-C5) on VLPs. In further studies, VLP Env exhibited an increased degree of inter-MAb competition, the epicenter of which was the base of the V3 loop, where PGT, 2G12, V3, and CD4 binding site specificities competed. UNC gp160 also underwent more drastic soluble CD4 (sCD4)-induced conformational changes than soluble gp120, exposing CD4i, C1-C4, and V2 epitopes. A greater propensity of UNC gp160 to undergo conformational changes was also suggested by the induction of CD4i MAb binding to VLPs by a V3 MAb as well as by soluble CD4. The same effect was not observed for soluble gp120. Taken together, our data suggest that membrane-expressed UNC gp160 exists in a less “triggered” conformational state than soluble gp120 and that MAb binding to UNC gp160 tends to have greater conformational consequences. PMID:23740975

  12. Natural mannosylation of HIV-1 gp120 imposes no immunoregulatory effects in primary human plasmacytoid dendritic cells

    NARCIS (Netherlands)

    Sondergaard, J.N.; Vinner, L.; Brix, S.

    2014-01-01

    Plasmacytoid dendritic cells (pDCs) play a vital role in activation of anti-HIV-1 immunity, and suppression of pDCs might mitigate immune responses against HIV-1. HIV-1 gp120 high-mannose has been attributed immunosuppressive roles in human myeloid DCs, but no receptors for high-mannose have so far

  13. HIV-1 Gp120 clade B/C induces a GRP78 driven cytoprotective mechanism in astrocytoma

    Science.gov (United States)

    López, Sheila N.; Rodríguez-Valentín, Madeline; Rivera, Mariela; Rodríguez, Maridaliz; Babu, Mohan; Cubano, Luis A.; Xiong, Huangui; Wang, Guangdi; Kucheryavykh, Lilia; Boukli, Nawal M.

    2017-01-01

    HIV-1 clades are known to be one of the key factors implicated in modulating HIV-associated neurocognitive disorders. HIV-1 B and C clades account for the majority of HIV-1 infections, clade B being the most neuropathogenic. The mechanisms behind HIV-mediated neuropathogenesis remain the subject of active research. We hypothesized that HIV-1 gp120 clade B and C proteins may exert differential proliferation, cell survival and NeuroAIDS effects in human astrocytoma cells via the Unfolded Protein Response, an endoplasmic reticulum- based cytoprotective mechanism. The differential effect of gp120 clade B and C was evaluated using for the first time a Tandem Mass Tag isobaric labeling quantitative proteomic approach. Flow cytometry analyses were performed for cell cycle and cell death identification. Among the proteins differentiated by HIV-1 gp120 proteins figure cytoskeleton, oxidative stress, UPR markers and numerous glycolytic metabolism enzymes. Our results demonstrate that HIV-1 gp120 B induced migration, proliferative and protective responses granted by the expression of GRP78, while HIV-1 gp120 C induced the expression of key inflammatory and pro-apoptotic markers. These novel findings put forward the first evidence that GRP78 is a key player in HIV-1 clade B and C neuropathogenic discrepancies and can be used as a novel target for immunotherapies. PMID:28978127

  14. Crystal structure of the neutralizing Llama V(HH D7 and its mode of HIV-1 gp120 interaction.

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    Andreas Hinz

    2010-05-01

    Full Text Available HIV-1 entry into host cells is mediated by the sequential binding of the envelope glycoprotein gp120 to CD4 and a chemokine receptor. Antibodies binding to epitopes overlapping the CD4-binding site on gp120 are potent inhibitors of HIV entry, such as the llama heavy chain antibody fragment V(HH D7, which has cross-clade neutralizing properties and competes with CD4 and mAb b12 for high affinity binding to gp120. We report the crystal structure of the D7 V(HH at 1.5 A resolution, which reveals the molecular details of the complementarity determining regions (CDR and substantial flexibility of CDR3 that could facilitate an induced fit interaction with gp120. Structural comparison of CDRs from other CD4 binding site antibodies suggests diverse modes of interaction. Mutational analysis identified CDR3 as a key component of gp120 interaction as determined by surface plasmon resonance. A decrease in affinity is directly coupled to the neutralization efficiency since mutations that decrease gp120 interaction increase the IC50 required for HIV-1 IIIB neutralization. Thus the structural study identifies the long CDR3 of D7 as the key determinant of interaction and HIV-1 neutralization. Furthermore, our data confirm that the structural plasticity of gp120 can accommodate multiple modes of antibody binding within the CD4 binding site.

  15. Mimicking protein-protein interactions through peptide-peptide interactions: HIV-1 gp120 and CXCR4

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    Andrea eGross

    2013-09-01

    Full Text Available We have recently designed a soluble synthetic peptide that functionally mimics the HIV-1 coreceptor CXCR4, which is a chemokine receptor that belongs to the family of seven-transmembrane GPCRs. This CXCR4 mimetic peptide, termed CX4-M1, presents the three extracellular loops (ECLs of the receptor. In binding assays involving recombinant proteins, as well as in cellular infection assays, CX4-M1 was found to selectively recognize gp120 from HIV-1 strains that use CXCR4 for cell entry (X4 tropic HIV-1. Furthermore, anti-HIV-1 antibodies modulate this interaction in a molecular mechanism related to that of their impact on the gp120-CXCR4 interaction. We could now show that the selectivity of CX4-M1 pertains not only to gp120 from X4 tropic HIV-1, but also to synthetic peptides presenting the V3 loops of these gp120 proteins. The V3 loop is thought to be an essential part of the coreceptor binding site of gp120 that contacts the second ECL of the coreceptor. We were able to experimentally confirm this notion in binding assays using substitution analogs of CX4-M1 and the V3 loop peptides, respectively, as well as in cellular infection assays. These results indicate that interactions of the HIV-1 Env with coreceptors can be mimicked by synthetic peptides, which may be useful to explore these interactions at the molecular level in more detail.

  16. Impaired neurogenesis by HIV-1-Gp120 is rescued by genetic deletion of fatty acid amide hydrolase enzyme.

    Science.gov (United States)

    Avraham, H K; Jiang, S; Fu, Y; Rockenstein, E; Makriyannis, A; Wood, J; Wang, L; Masliah, E; Avraham, S

    2015-10-01

    The HIV-envelope glycoprotein Gp120 is involved in neuronal injury and is associated with neuro-AIDS pathogenesis in the brain. Endocannabinoids are important lipid ligands in the CNS regulating neural functions, and their degeneration is controlled by hydrolysing enzymes such as the fatty acid amide hydrolase (FAAH). Here, we examined whether in vivo genetic deletion of Faah gene prevents HIV-1 Gp120-mediated effects on neurogenesis. We generated new GFAP/Gp120 transgenic (Tg) mice that have genetic deletion of Faah gene by mating glial fribillary acidic protein (GFAP)/Gp120 Tg mice with Faah-/- mice. Neurogenesis and cell death were assessed by immunocytochemical analysis. Endocannabinoid levels in the brain of the double GFAP/Gp120//Faah-/- mice were similar to those observed in Faah-/- mice. However, unlike the impaired neurogenesis observed in GFAP/Gp120 Tg mice and Faah-/- mice, these GFAP/Gp120//Faah-/ mice showed significantly improved neurogenesis in the hippocampus, indicated by a significant increase in neuroblasts and neuronal cells, an increase in BrdU(+) cells and doublecortin positive cells (DCX(+) ), and an increase in the number of PCNA. Furthermore, a significant decrease in astrogliosis and gliogenesis was observed in GFAP/Gp120//Faah-/-mice and neurogenesis was stimulated by neural progenitor cells (NPCs) and/or the newly formed NPC niches characterized by increased COX-2 expression and elevated levels of PGE2 . In vivo genetic ablation of Faah, resulted in enhanced neurogenesis through modulation of the newly generated NPC niches in GFAP/Gp120//Faah-/- mice. This suggests a novel approach of using FAAH inhibitors to enhance neurogenesis in HIV-1 infected brain. © 2015 The Authors. British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of The British Pharmacological Society.

  17. Conformational alterations in the CD4 binding cavity of HIV-1 gp120 influencing gp120-CD4 interactions and fusogenicity of HIV-1 envelopes derived from brain and other tissues

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    Ramsland Paul A

    2011-06-01

    Full Text Available Abstract Background CD4-binding site (CD4bs alterations in gp120 contribute to HIV-1 envelope (Env mediated fusogenicity and the ability of gp120 to utilize low levels of cell-surface CD4. In a recent study, we constructed three-dimensional models of gp120 to illustrate CD4bs conformations associated with enhanced fusogenicity and enhanced CD4-usage of a modestly-sized panel of blood-derived HIV-1 Envs (n = 16. These conformations were characterized by a wider aperture of the CD4bs cavity, as constrained by the inner-most atoms at the gp120 V1V2 stem and the V5 loop. Here, we sought to provide further validation of the utility of these models for understanding mechanisms that influence Env function, by characterizing the structure-function relationships of a larger panel of Envs derived from brain and other tissues (n = 81. Findings Three-dimensional models of gp120 were generated by our recently validated homology modelling protocol. Analysis of predicted CD4bs structures showed correlations between the aperture width of the CD4bs cavity and ability of the Envs to mediate cell-cell fusion, scavenge low-levels of cell-surface CD4, bind directly to soluble CD4, and bind to the Env mAb IgG1b12 whose epitope overlaps the gp120 CD4bs. These structural alterations in the CD4bs cavity were associated with repositioning of the V5 loop. Conclusions Using a large, independent panel of Envs, we can confirm the utility of three-dimensional gp120 structural models for illustrating CD4bs alterations that can affect Env function. Furthermore, we now provide new evidence that these CD4bs alterations augment the ability of gp120 to interact with CD4 by increasing the exposure of the CD4bs.

  18. Design and Characterization of a Peptide Mimotope of the HIV-1 gp120 Bridging Sheet

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    Guido Poli

    2012-05-01

    Full Text Available The Bridging Sheet domain of HIV-1 gp120 is highly conserved among the HIV-1 strains and allows HIV-1 binding to host cells via the HIV-1 coreceptors. Further, the bridging sheet domain is a major target to neutralize HIV-1 infection. We rationally designed four linear peptide epitopes that mimic the three-dimensional structure of bridging sheet by using molecular modeling. Chemically synthesized peptides BS3 and BS4 showed a fair degree of antigenicity when tested in ELISA with IgG purified from HIV+ broadly neutralizing sera while the production of synthetic peptides BS1 and BS2 failed due to their high degree of hydrophobicity. To overcome this limitation, we linked all four BS peptides to the COOH-terminus of GST protein to test both their antigenicity and immunogenicity. Only the BS1 peptide showed good antigenicity; however, no envelope specific antibodies were elicited upon mice immunization. Therefore we performed further analyses by linking BS1 peptide to the NH2-terminus of the E2 scaffold from the Geobacillus Stearothermophylus PDH complex. The E2-BS1 fusion peptide showed good antigenic results, however only one immunized rabbit elicited good antibody titers towards both the monomeric and oligomeric viral envelope glycoprotein (Env. In addition, moderate neutralizing antibodies response was elicited against two HIV-1 clade B and one clade C primary isolates. These preliminary data validate the peptide mimotope approach as a promising tool to obtain an effective HIV-1 vaccine.

  19. Small-molecule HIV-1 entry inhibitors targeting gp120 and gp41: a patent review (2010-2015).

    Science.gov (United States)

    Li, Wen; Lu, Lu; Li, Weihua; Jiang, Shibo

    2017-06-01

    It is essential to discover and develop small-molecule HIV-1 entry inhibitors with suitable pharmaceutical properties. Areas covered: We review the development of small-molecule HIV-1 entry inhibitors as evidenced in patents, patent applications, and related research articles published between 2010 and 2015. Expert opinion: HIV-1 Env gp120 and gp41 are important targets for development of HIV-1 entry inhibitors. The Phe43 pocket in gp120 and the highly conserved hydrophobic pocket on gp41 NHR-trimer are important targets for identification of HIV-1 attachment and fusion inhibitors, respectively. Compounds that bind to Phe43 pocket can block viral gp120 binding to CD4 on T cells, thus inhibiting HIV-1 attachment. However, most compounds targeting Phe43 pocket identified so far are HIV-1 entry agonists with the ability to enhance infectivity of HIV-1 in CD4-negative cells. Therefore, it is essential to identify HIV-1 entry antagonist-based HIV-1 attachment/entry inhibitors. Compounds binding to the gp41 hydrophobic pocket may inhibit CHR binding to the gp41 NHR trimer, thus blocking six-helix bundle formation and gp41-mediated virus-cell fusion. However, most lead compounds targeting this pocket have low potency, possibly because the pocket is too big or too deep. Therefore, it is necessary to identify other pockets in gp41 for developing HIV-1 fusion/entry inhibitors.

  20. Pharmacological descriptors related to the binding of Gp120 to CD4 corresponding to 60 representative HIV-1 strains.

    Science.gov (United States)

    Calborean, Octavian; Mernea, Maria; Avram, Speranta; Mihailescu, Dan Florin

    2013-10-01

    The standardization of HIV-1 strains for vaccine tests include the use of viral reference panels. We determined a series of pharmacological descriptors (molecular surfaces, volumes, electrostatic energies, solvation energies, number of atoms, number of hydrogen donors or acceptors and number of rigid bonds) for the gp120 CD4-binding sites structures in the unliganded state from a reference panel of 60 diverse strains of HIV-1. We identified the descriptors that varied significantly between the strains, the outliner strains for each descriptor set and the possible correlations between the descriptors. Our results improve the knowledge about gp120, its molecular and possible neutralization properties.

  1. Mapping the complete glycoproteome of virion-derived HIV-1 gp120 provides insights into broadly neutralizing antibody binding.

    Science.gov (United States)

    Panico, Maria; Bouché, Laura; Binet, Daniel; O'Connor, Michael-John; Rahman, Dinah; Pang, Poh-Choo; Canis, Kevin; North, Simon J; Desrosiers, Ronald C; Chertova, Elena; Keele, Brandon F; Bess, Julian W; Lifson, Jeffrey D; Haslam, Stuart M; Dell, Anne; Morris, Howard R

    2016-09-08

    The surface envelope glycoprotein (SU) of Human immunodeficiency virus type 1 (HIV-1), gp120(SU) plays an essential role in virus binding to target CD4+ T-cells and is a major vaccine target. Gp120 has remarkably high levels of N-linked glycosylation and there is considerable evidence that this "glycan shield" can help protect the virus from antibody-mediated neutralization. In recent years, however, it has become clear that gp120 glycosylation can also be included in the targets of recognition by some of the most potent broadly neutralizing antibodies. Knowing the site-specific glycosylation of gp120 can facilitate the rational design of glycopeptide antigens for HIV vaccine development. While most prior studies have focused on glycan analysis of recombinant forms of gp120, here we report the first systematic glycosylation site analysis of gp120 derived from virions produced by infected T lymphoid cells and show that a single site is exclusively substituted with complex glycans. These results should help guide the design of vaccine immunogens.

  2. Binding of the mannose-specific lectin, Griffithsin, to HIV-1 gp120 exposes the CD4-binding site

    CSIR Research Space (South Africa)

    Alexandre, Kabamba B

    2011-09-01

    Full Text Available The glycans on HIV-1 gp120 play an important role in shielding neutralization-sensitive epitopes from antibody recognition. They also serve as targets for lectins that bind mannose-rich glycans. In this study, the authors investigated...

  3. Fusion proteins of HIV-1 envelope glycoprotein gp120 with CD4-induced antibodies showed enhanced binding to CD4 and CD4 binding site antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Weizao, E-mail: chenw3@mail.nih.gov [Protein Interactions Group, Frederick National Laboratory for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, MD 21702 (United States); Feng, Yang [Protein Interactions Group, Frederick National Laboratory for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, MD 21702 (United States); Wang, Yanping [Protein Interactions Group, Frederick National Laboratory for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, MD 21702 (United States); The Basic Research Program, Science Applications International Corporation-Frederick, Inc., National Cancer Institute, National Institutes of Health, Frederick, MD 21702 (United States); Zhu, Zhongyu; Dimitrov, Dimiter S. [Protein Interactions Group, Frederick National Laboratory for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, MD 21702 (United States)

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer Some recombinant HIV-1 gp120s do not preserve their conformations on gp140s. Black-Right-Pointing-Pointer We hypothesize that CD4i antibodies could induce conformational changes in gp120. Black-Right-Pointing-Pointer CD4i antibodies enhance binding of CD4 and CD4bs antibodies to gp120. Black-Right-Pointing-Pointer CD4i antibody-gp120 fusion proteins could have potential as vaccine immunogens. -- Abstract: Development of successful AIDS vaccine immunogens continues to be a major challenge. One of the mechanisms by which HIV-1 evades antibody-mediated neutralizing responses is the remarkable conformational flexibility of its envelope glycoprotein (Env) gp120. Some recombinant gp120s do not preserve their conformations on gp140s and functional viral spikes, and exhibit decreased recognition by CD4 and neutralizing antibodies. CD4 binding induces conformational changes in gp120 leading to exposure of the coreceptor-binding site (CoRbs). In this study, we test our hypothesis that CD4-induced (CD4i) antibodies, which target the CoRbs, could also induce conformational changes in gp120 leading to better exposed conserved neutralizing antibody epitopes including the CD4-binding site (CD4bs). We found that a mixture of CD4i antibodies with gp120 only weakly enhanced CD4 binding. However, such interactions in single-chain fusion proteins resulted in gp120 conformations which bound to CD4 and CD4bs antibodies better than the original or mutagenically stabilized gp120s. Moreover, the two molecules in the fusion proteins synergized with each other in neutralizing HIV-1. Therefore, fusion proteins of gp120 with CD4i antibodies could have potential as components of HIV-1 vaccines and inhibitors of HIV-1 entry, and could be used as reagents to explore the conformational flexibility of gp120 and mechanisms of entry and immune evasion.

  4. Residues in the gp41 Ectodomain Regulate HIV-1 Envelope Glycoprotein Conformational Transitions Induced by gp120-Directed Inhibitors.

    Science.gov (United States)

    Pacheco, Beatriz; Alsahafi, Nirmin; Debbeche, Olfa; Prévost, Jérémie; Ding, Shilei; Chapleau, Jean-Philippe; Herschhorn, Alon; Madani, Navid; Princiotto, Amy; Melillo, Bruno; Gu, Christopher; Zeng, Xin; Mao, Youdong; Smith, Amos B; Sodroski, Joseph; Finzi, Andrés

    2017-03-01

    Interactions between the gp120 and gp41 subunits of the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env) trimer maintain the metastable unliganded form of the viral spike. Binding of gp120 to the receptor, CD4, changes the Env conformation to promote gp120 interaction with the second receptor, CCR5 or CXCR4. CD4 binding also induces the transformation of Env into the prehairpin intermediate, in which the gp41 heptad repeat 1 (HR1) coiled coil is assembled at the trimer axis. In nature, HIV-1 Envs must balance the requirements to maintain the noncovalent association of gp120 with gp41 and to evade the host antibody response with the need to respond to CD4 binding. Here we show that the gp41 HR1 region contributes to gp120 association with the unliganded Env trimer. Changes in particular amino acid residues in the gp41 HR1 region decreased the efficiency with which Env moved from the unliganded state. Thus, these gp41 changes decreased the sensitivity of HIV-1 to cold inactivation and ligands that require Env conformational changes to bind efficiently. Conversely, these gp41 changes increased HIV-1 sensitivity to small-molecule entry inhibitors that block Env conformational changes induced by CD4. Changes in particular gp41 HR1 amino acid residues can apparently affect the relative stability of the unliganded state and CD4-induced conformations. Thus, the gp41 HR1 region contributes to the association with gp120 and regulates Env transitions from the unliganded state to downstream conformations.IMPORTANCE The development of an efficient vaccine able to prevent HIV infection is a worldwide priority. Knowledge of the envelope glycoprotein structure and the conformational changes that occur after receptor engagement will help researchers to develop an immunogen able to elicit antibodies that block HIV-1 transmission. Here we identify residues in the HIV-1 transmembrane envelope glycoprotein that stabilize the unliganded state by modulating the

  5. Clustering of HIV-1 Subtypes Based on gp120 V3 Loop electrostatic properties

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    López de Victoria Aliana

    2012-02-01

    Full Text Available Abstract Background The V3 loop of the glycoprotein gp120 of HIV-1 plays an important role in viral entry into cells by utilizing as coreceptor CCR5 or CXCR4, and is implicated in the phenotypic tropisms of HIV viruses. It has been hypothesized that the interaction between the V3 loop and CCR5 or CXCR4 is mediated by electrostatics. We have performed hierarchical clustering analysis of the spatial distributions of electrostatic potentials and charges of V3 loop structures containing consensus sequences of HIV-1 subtypes. Results Although the majority of consensus sequences have a net charge of +3, the spatial distribution of their electrostatic potentials and charges may be a discriminating factor for binding and infectivity. This is demonstrated by the formation of several small subclusters, within major clusters, which indicates common origin but distinct spatial details of electrostatic properties. Some of this information may be present, in a coarse manner, in clustering of sequences, but the spatial details are largely lost. We show the effect of ionic strength on clustering of electrostatic potentials, information that is not present in clustering of charges or sequences. We also make correlations between clustering of electrostatic potentials and net charge, coreceptor selectivity, global prevalence, and geographic distribution. Finally, we interpret coreceptor selectivity based on the N6X7T8|S8X9 sequence glycosylation motif, the specific positive charge location according to the 11/24/25 rule, and the overall charge and electrostatic potential distribution. Conclusions We propose that in addition to the sequence and the net charge of the V3 loop of each subtype, the spatial distributions of electrostatic potentials and charges may also be important factors for receptor recognition and binding and subsequent viral entry into cells. This implies that the overall electrostatic potential is responsible for long-range recognition of the V3

  6. HIV-1 and recombinant gp120 affect the survival and differentiation of human vessel wall-derived mesenchymal stem cells

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    Pasquinelli Gianandrea

    2011-05-01

    Full Text Available Abstract Background HIV infection elicits the onset of a progressive immunodeficiency and also damages several other organs and tissues such as the CNS, kidney, heart, blood vessels, adipose tissue and bone. In particular, HIV infection has been related to an increased incidence of cardiovascular diseases and derangement in the structure of blood vessels in the absence of classical risk factors. The recent characterization of multipotent mesenchymal cells in the vascular wall, involved in regulating cellular homeostasis, suggests that these cells may be considered a target of HIV pathogenesis. This paper investigated the interaction between HIV-1 and vascular wall resident human mesenchymal stem cells (MSCs. Results MSCs were challenged with classical R5 and X4 HIV-1 laboratory strains demonstrating that these strains are able to enter and integrate their retro-transcribed proviral DNA in the host cell genome. Subsequent experiments indicated that HIV-1 strains and recombinant gp120 elicited a reliable increase in apoptosis in sub-confluent MSCs. Since vascular wall MSCs are multipotent cells that may be differentiated towards several cell lineages, we challenged HIV-1 strains and gp120 on MSCs differentiated to adipogenesis and endotheliogenesis. Our experiments showed that the adipogenesis is increased especially by upregulated PPARγ activity whereas the endothelial differentiation induced by VEGF treatment was impaired with a downregulation of endothelial markers such as vWF, Flt-1 and KDR expression. These viral effects in MSC survival and adipogenic or endothelial differentiation were tackled by CD4 blockade suggesting an important role of CD4/gp120 interaction in this context. Conclusions The HIV-related derangement of MSC survival and differentiation may suggest a direct role of HIV infection and gp120 in impaired vessel homeostasis and in genesis of vessel damage observed in HIV-infected patients.

  7. Cocaine potentiates astrocyte toxicity mediated by human immunodeficiency virus (HIV-1 protein gp120.

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    Yanjing Yang

    2010-10-01

    Full Text Available It is becoming widely accepted that psychoactive drugs, often abused by HIV-I infected individuals, can significantly alter the progression of neuropathological changes observed in HIV-associated neurodegenerative diseases (HAND. The underlying mechanisms mediating these effects however, remain poorly understood. In the current study, we explored whether the psychostimulant drug cocaine could exacerbate toxicity mediated by gp120 in rat primary astrocytes. Exposure to both cocaine and gp120 resulted in increased cell toxicity compared to cells treated with either factor alone. The combinatorial toxicity of cocaine and gp120 was accompanied by an increase in caspase-3 activation. In addition, increased apoptosis of astrocytes in the presence of both the agents was associated with a concomitant increase in the production of intracellular reactive oxygen species and loss of mitochondrial membrane potential. Signaling pathways including c-jun N-teminal kinase (JNK, p38, extracellular signal-regulated kinase (ERK/mitogen-activated protein kinases (MAPK, and nuclear factor (NF-κB were identified to be major players in cocaine and gp120-mediated apoptosis of astrocytes. Our results demonstrated that cocaine-mediated potentiation of gp120 toxicity involved regulation of oxidative stress, mitochondrial membrane potential and MAPK signaling pathways.

  8. Toll-like receptor expression and activation in astroglia: differential regulation by HIV-1 Tat, gp120, and morphine

    Science.gov (United States)

    El-Hage, Nazira; Podhaizer, Elizabeth M.; Sturgill, Jamie; Hauser, Kurt F.

    2012-01-01

    In this study we determine whether morphine alone or in combination with HIV-1 Tat or gp120 affects the expression of Toll-like receptors (TLRs) by astrocytes and to assess whether TLRs expressed by astrocytes function in the release of inflammatory mediators in vitro. TLR profiling by immunofluorescence microscopy, flow cytometry, in-cell westerns, and RT-PCR showed that subpopulations of astrocytes possessed TLR 2, TLR3, TLR4, and TLR9 antigenicity. Exposure to HIV-1 Tat, gp120, and/or morphine significantly altered the proportion of TLR-immunopositive and/or TLR expression by astroglia in a TLR-specific manner. Subsets of astroglia displayed significant increases in TLR2 with reciprocal decreases in TLR9 expression in response to Tat or gp120 ± morphine treatment. TLR9 expression was also significantly decreased by morphine alone. Exposing astrocytes to the TLR agonists LTA (TLR2), poly I:C (TLR3), LPS (TLR4) and unmethylated CpG ODN (TLR9) resulted in increased secretion of MCP-1/CCL2 and elevations in reactive oxygen species. TLR3 and TLR4 stimulation increased the secretion of TNF-α, IL-6, and RANTES/CCL5, while activation of TLR2 caused a significant increase in nitric oxide levels. The results suggest that HIV-1 proteins and/or opioid abuse disrupt the innate immune response of the central nervous system (CNS) which may lead to increased pathogenicity. PMID:21425908

  9. CD4-gp120 interaction interface - a gateway for HIV-1 infection in human: molecular network, modeling and docking studies.

    Science.gov (United States)

    Pandey, Deeksha; Podder, Avijit; Pandit, Mansi; Latha, Narayanan

    2017-09-01

    The major causative agent for Acquired Immune Deficiency Syndrome (AIDS) is Human Immunodeficiency Virus-1 (HIV-1). HIV-1 is a predominant subtype of HIV which counts on human cellular mechanism virtually in every aspect of its life cycle. Binding of viral envelope glycoprotein-gp120 with human cell surface CD4 receptor triggers the early infection stage of HIV-1. This study focuses on the interaction interface between these two proteins that play a crucial role for viral infectivity. The CD4-gp120 interaction interface has been studied through a comprehensive protein-protein interaction network (PPIN) analysis and highlighted as a useful step towards identifying potential therapeutic drug targets against HIV-1 infection. We prioritized gp41, Nef and Tat proteins of HIV-1 as valuable drug targets at early stage of viral infection. Lack of crystal structure has made it difficult to understand the biological implication of these proteins during disease progression. Here, computational protein modeling techniques and molecular dynamics simulations were performed to generate three-dimensional models of these targets. Besides, molecular docking was initiated to determine the desirability of these target proteins for already available HIV-1 specific drugs which indicates the usefulness of these protein structures to identify an effective drug combination therapy against AIDS.

  10. HIV-1 induced AIDS is an allergy and the allergen is the Shed gp120--a review, hypothesis, and implications.

    Science.gov (United States)

    Becker, Yechiel

    2004-04-01

    The possibility that the induction of T helper 2 (Th2) cytokine synthesis and the gradual increase in interleukin 4 (IL-4) and IgE levels during HIV-1 infection are an allergic response to HIV-1 proteins was raised in the author's previous article [Becker, Virus Genes 28, 1-4, 2004]. The present review extends this hypothesis by citing experimental reports which indicate that HIV-1 shed gp120 virions share a striking resemblance with the allergens that bind to IgE molecules bound to Fc epsilon receptor I-positive (Fc epsilon RI) cells (mast cells, basophils, monocytes, and dendritic cells, DC) and then induce them to release and synthesize the IL-4 cytokine. In the earlier review, it was established that the IL-4 cytokine is responsible for the following processes: IgE synthesis by B cells and the inhibition of antiviral IgG synthesis; the inactivation of T helper 1 (Th1) cells; and the inhibition of the antiviral cytotoxic T cell (CTL) response. The binding of the shed gp120 to B cell-bound immunoglobulin (Ig) depletes these cells. Moreover, it was reported that the viral regulatory proteins Tat and Vpr also stimulate IL-4 release from basophils. The mode of action used by HIV-1 gp120 in the induction of IL-4 [Becker, Virus Genes 28, 1-14, 2004] revealed that the viral protein contains a superantigen (SAg) domain that is capable of binding to the V(H)3 domain of IgE and Ig; namely, the shed gp120 protein act as endogenous and environmental allergens that bind to IgE molecules, which are bound to the Fc epsilon RI receptors on hematopoietic cells. Based on these findings, it is hypothesized that gp120 is an allergen. Consequently, it can be inferred that the active replication of HIV-1 in infected individuals constantly exposes the immune system to an increase in the allergen content until the host immunity is eventually compromised. These finding suggest that HIV-1 engage in a two-pronged attack of the human immune system: it infects Th2 cells, macrophages, and

  11. HIV-1 gp41-targeting fusion inhibitory peptides enhance the gp120-targeting protein-mediated inactivation of HIV-1 virions

    OpenAIRE

    Qi, Qianqian; Wang, Qian; Chen, Weizao; Du, Lanying; Dimitrov, Dimiter S; Lu, Lu; Jiang, Shibo

    2017-01-01

    Protein- or peptide-based viral inactivators are being developed as novel antiviral drugs with improved efficacy, pharmacokinetics and toxicity profiles because they actively inactivate cell-free human immunodeficiency virus type 1 (HIV-1) virions before attachment to host cells. By contrast, most clinically used antiviral drugs must penetrate host cells to inhibit viral replication. In this study, we pre-treated HIV-1 particles with a gp120-targeting bispecific multivalent protein, 2Dm2m or ...

  12. Generation and Characterization of a Bivalent HIV-1 Subtype C gp120 Protein Boost for Proof-of-Concept HIV Vaccine Efficacy Trials in Southern Africa.

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    Carlo Zambonelli

    Full Text Available The viral envelope glycoprotein (Env is the major target for antibody (Ab-mediated vaccine development against the Human Immunodeficiency Virus type 1 (HIV-1. Although several recombinant Env antigens have been evaluated in clinical trials, only the surface glycoprotein, gp120, (from HIV-1 subtype B, MN, and subtype CRF_01AE, A244 used in the ALVAC prime-AIDSVAX gp120 boost RV144 Phase III HIV vaccine trial was shown to contribute to protective efficacy, although modest and short-lived. Hence, for clinical trials in southern Africa, a bivalent protein boost of HIV-1 subtype C gp120 antigens composed of two complementary gp120s, from the TV1.C (chronic and 1086.C (transmitted founder HIV-1 strains, was selected. Stable Chinese Hamster Cell (CHO cell lines expressing these gp120s were generated, scalable purification methods were developed, and a detailed analytical analysis of the purified proteins was conducted that showed differences and complementarity in the antigenicity, glycan occupancy, and glycan content of the two gp120 molecules. Moreover, mass spectrometry revealed some disulfide heterogeneity in the expressed proteins, particularly in V1V2-C1 region and most prominently in the TV1 gp120 dimers. These dimers not only lacked binding to certain key CD4 binding site (CD4bs and V1V2 epitope-directed ligands but also elicited reduced Ab responses directed to those epitopes, in contrast to monomeric gp120, following immunization of rabbits. Both monomeric and dimeric gp120s elicited similarly high titer Tier 1 neutralizing Abs as measured in standard virus neutralization assays. These results provide support for clinical evaluations of bivalent preparations of purified monomeric TV1.C and 1086.C gp120 proteins.

  13. Anti-gp120 minibody gene transfer to female genital epithelial cells protects against HIV-1 virus challenge in vitro.

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    Ussama M Abdel-Motal

    Full Text Available Although cervico-vaginal epithelial cells of the female lower genital tract provide the initial defense system against HIV-1 infection, the protection is sometimes incomplete. Thus, enhancing anti-HIV-1 humoral immunity at the mucosal cell surface by local expression of anti-HIV-1 broadly neutralizing antibodies (BnAb that block HIV-1 entry would provide an important new intervention that could slow the spread of HIV/AIDS.This study tested the hypothesis that adeno-associated virus (AAV-BnAb gene transfer to cervico-vaginal epithelial cells will lead to protection against HIV-1. Accordingly, a recombinant AAV vector that encodes human b12 anti-HIV gp120 BnAb as a single-chain variable fragment Fc fusion (scFvFc, or "minibody" was constructed. The secreted b12 minibody was shown to be biologically functional in binding to virus envelope protein, neutralizing HIV-1 and importantly, blocking transfer and infectivity of HIV-1(bal in an organotypic human vaginal epithelial cell (VEC model. Furthermore, cervico-vaginal epithelial stem cells were found to be efficiently transduced by the optimal AAV serotype mediated expression of GFP.This study provides the foundation for a novel microbicide strategy to protect against sexual transmission of HIV-1 by AAV transfer of broadly neutralizing antibody genes to cervico-vaginal epithelial stem cells that could replenish b12 BnAb secreting cells through multiple menstrual cycles.

  14. Deletion of the highly conserved N-glycan at Asn260 of HIV-1 gp120 affects folding and lysosomal degradation of gp120, and results in loss of viral infectivity.

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    Leen Mathys

    Full Text Available N-linked glycans covering the surface of the HIV-1 glycoprotein gp120 are of major importance for the correct folding of this glycoprotein. Of the, on average, 24 N-linked glycans present on gp120, the glycan at Asn260 was reported to be essential for the correct expression of gp120 and gp41 in the virus particle and deletion of the N260 glycan in gp120 heavily compromised virus infectivity. We show here that gp160 containing the N260Q mutation reaches the Golgi apparatus during biosynthesis. Using pulse-chase experiments with [35S] methionine/cysteine, we show that oxidative folding was slightly delayed in case of mutant N260Q gp160 and that CD4 binding was markedly compromised compared to wild-type gp160. In the search of compensatory mutations, we found a mutation in the V1/V2 loop of gp120 (S128N that could partially restore the infectivity of mutant N260Q gp120 virus. However, the mutation S128N did not enhance any of the above-mentioned processes so its underlying compensatory mechanism must be a conformational effect that does not affect CD4 binding per se. Finally, we show that mutant N260Q gp160 was cleaved to gp120 and gp41 to a much lower extent than wild-type gp160, and that it was subject of lysosomal degradation to a higher extent than wild-type gp160 showing a prominent role of this process in the breakdown of N260-glycan-deleted gp160, which could not be counteracted by the S128N mutation. Moreover, at least part of the wild-type or mutant gp160 that is normally targeted for lysosomal degradation reached a conformation that enabled CD4 binding.

  15. HIV-1 gp41-targeting fusion inhibitory peptides enhance the gp120-targeting protein-mediated inactivation of HIV-1 virions.

    Science.gov (United States)

    Qi, Qianqian; Wang, Qian; Chen, Weizao; Du, Lanying; Dimitrov, Dimiter S; Lu, Lu; Jiang, Shibo

    2017-06-21

    Protein- or peptide-based viral inactivators are being developed as novel antiviral drugs with improved efficacy, pharmacokinetics and toxicity profiles because they actively inactivate cell-free human immunodeficiency virus type 1 (HIV-1) virions before attachment to host cells. By contrast, most clinically used antiviral drugs must penetrate host cells to inhibit viral replication. In this study, we pre-treated HIV-1 particles with a gp120-targeting bispecific multivalent protein, 2Dm2m or 4Dm2m, in the presence or absence of the gp41-targeting HIV-1 fusion inhibitory peptides enfuvirtide (T20), T2635, or sifuvirtide (SFT). HIV-1 virions were separated from the inhibitors using PEG-6000, followed by testing of the residual infectivity of the HIV-1 virions. 2Dm2m and 4Dm2m exhibited significant inactivation activity against all HIV-1 strains tested with EC50 values at the low nanomolar level, whereas none of the gp41-targeting peptides showed inactivation activity at concentrations up to 250 nM. Notably, these three peptides significantly enhanced protein-mediated inactivation against cell-free HIV-1 virions, including HIV-1 laboratory-adapted and primary HIV-1 strains, as well as those resistant to T20 or T2635 and virions released from reactivated latently HIV-1-infected cells. These results indicate that the gp120-targeting bispecific multivalent proteins 2Dm2m and 4Dm2m have potential for further development as HIV-1 inactivator-based antiviral drugs for use in the clinic, either alone or in combination with a gp41-targeting HIV-1 fusion inhibitor such as T20, to treat patients with HIV-1 infection and AIDS.

  16. Indirect detection of an epitope-specific response to HIV-1 gp120 immunization in human subjects.

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    Evgeny Shmelkov

    Full Text Available A specific response of human serum neutralizing antibodies (nAb to a conformational epitope as a result of vaccination of human subjects with the surface envelope glycoprotein (gp120 of HIV-1 has not previously been documented. Here, we used computational analysis to assess the epitope-specific responses of human subjects, which were immunized with recombinant gp120 immunogens in the VAX003 and VAX004 clinical trials. Our computational methodology--a variation of sieve analysis--compares the occurrence of specific nAb targeted conformational 3D epitopes on viruses from infected individuals who received vaccination to the occurrence of matched epitopes in the viruses infecting placebo subjects. We specifically studied seven crystallographically defined nAb targeted conformational epitopes in the V3 loop, an immunogenic region of gp120. Of the six epitopes present in the immunogens and targeted by known monoclonal neutralizing antibodies, only the one targeted by the anti-V3 nAb 2219 exhibited a significant reduction in occurrence in vaccinated subjects compared to the placebo group. This difference occurred only in the VAX003 Thailand cohort. No difference was seen between vaccinated and placebo groups for the occurrence of an epitope that was not present in the immunogen. Thus, it can be theorized that a specific 2219-like human neutralizing antibody immune response to AIDSVAX immunization occurred in the VAX003 cohort, and that this response protected subjects from a narrow subset of HIV-1 viruses circulating in Thailand in the 1990s and bearing the conformational epitope targeted by the neutralizing antibody 2219.

  17. The HIV-1 envelope protein gp120 is captured and displayed for B cell recognition by SIGN-R1(+) lymph node macrophages.

    Science.gov (United States)

    Park, Chung; Arthos, James; Cicala, Claudia; Kehrl, John H

    2015-08-10

    The HIV-1 envelope protein gp120 is both the target of neutralizing antibodies and a major focus of vaccine efforts; however how it is delivered to B cells to elicit an antibody response is unknown. Here, we show that following local gp120 injection lymph node (LN) SIGN-R1(+) sinus macrophages located in interfollicular pockets and underlying SIGN-R1(+) macrophages form a cellular network that rapidly captures gp120 from the afferent lymph. In contrast, two other antigens, phycoerythrin and hen egg lysozyme, were not captured by these cells. Intravital imaging of mouse LNs revealed persistent, but transient interactions between gp120 bearing interfollicular network cells and both trafficking and LN follicle resident gp120 specific B cells. The gp120 specific, but not the control B cells repetitively extracted gp120 from the network cells. Our findings reveal a specialized LN antigen delivery system poised to deliver gp120 and likely other pathogen derived glycoproteins to B cells.

  18. The HIV-1 envelope protein gp120 is captured and displayed for B cell recognition by SIGN-R1+ lymph node macrophages

    Science.gov (United States)

    Park, Chung; Arthos, James; Cicala, Claudia; Kehrl, John H

    2015-01-01

    The HIV-1 envelope protein gp120 is both the target of neutralizing antibodies and a major focus of vaccine efforts; however how it is delivered to B cells to elicit an antibody response is unknown. Here, we show that following local gp120 injection lymph node (LN) SIGN-R1+ sinus macrophages located in interfollicular pockets and underlying SIGN-R1+ macrophages form a cellular network that rapidly captures gp120 from the afferent lymph. In contrast, two other antigens, phycoerythrin and hen egg lysozyme, were not captured by these cells. Intravital imaging of mouse LNs revealed persistent, but transient interactions between gp120 bearing interfollicular network cells and both trafficking and LN follicle resident gp120 specific B cells. The gp120 specific, but not the control B cells repetitively extracted gp120 from the network cells. Our findings reveal a specialized LN antigen delivery system poised to deliver gp120 and likely other pathogen derived glycoproteins to B cells. DOI: http://dx.doi.org/10.7554/eLife.06467.001 PMID:26258881

  19. Binding of HIV-1 gp120 to DC-SIGN Promotes ASK-1-Dependent Activation-Induced Apoptosis of Human Dendritic Cells

    Science.gov (United States)

    Chan, Vera S. F.; Chung, Nancy P. Y.; Wang, Shu-Rong; Li, Zhongye; Ma, Jing; Lin, Chia-Wei; Hsieh, Ya-Ju; Chang, Kao-Ping; Kung, Sui-Sum; Wu, Yi-Chia; Chu, Cheng-Wei; Tai, Hsiao-Ting; Gao, George F.; Zheng, Bojian; Yokoyama, Kazunari K.; Austyn, Jonathan M.; Lin, Chen-Lung S.

    2013-01-01

    During disease progression to AIDS, HIV-1 infected individuals become increasingly immunosuppressed and susceptible to opportunistic infections. It has also been demonstrated that multiple subsets of dendritic cells (DC), including DC-SIGN(+) cells, become significantly depleted in the blood and lymphoid tissues of AIDS patients, which may contribute to the failure in initiating effective host immune responses. The mechanism for DC depletion, however, is unclear. It is also known that vast quantities of viral envelope protein gp120 are shed from maturing HIV-1 virions and form circulating immune complexes in the serum of HIV-1-infected individuals, but the pathological role of gp120 in HIV-1 pathogenesis remains elusive. Here we describe a previously unrecognized mechanism of DC death in chronic HIV-1 infection, in which ligation of DC-SIGN by gp120 sensitizes DC to undergo accelerated apoptosis in response to a variety of activation stimuli. The cultured monocyte-derived DC and also freshly-isolated DC-SIGN(+) blood DC that were exposed to either cross-linked recombinant gp120 or immune-complex gp120 in HIV(+) serum underwent considerable apoptosis after CD40 ligation or exposure to bacterial lipopolysaccharide (LPS) or pro-inflammatory cytokines such as TNFα and IL-1β. Furthermore, circulating DC-SIGN(+) DC that were isolated directly from HIV-1(+) individuals had actually been pre-sensitized by serum gp120 for activation-induced exorbitant apoptosis. In all cases the DC apoptosis was substantially inhibited by DC-SIGN blockade. Finally, we showed that accelerated DC apoptosis was a direct consequence of excessive activation of the pro-apoptotic molecule ASK-1 and transfection of siRNA against ASK-1 significantly prevented the activation-induced excessive DC death. Our study discloses a previously unknown mechanism of immune modulation by envelope protein gp120, provides new insights into HIV immunopathogenesis, and suggests potential therapeutic approaches to

  20. Natural mannosylation of HIV-1 gp120 imposes no immunoregulatory effects in primary human plasmacytoid dendritic cells

    DEFF Research Database (Denmark)

    Søndergaard, Jonas Nørskov; Vinner, Lasse; Pedersen, Susanne Brix

    2014-01-01

    or viable HIV-1 particles with various degrees of mannosylation were cultured with pDCs. Activation of pDCs was determined by assaying secretion of IFN-alpha, viability, and upregulation of several pDC-activation markers: CD40, CD86, HLA-DR, CCR7, and PD-L1. The level of activation negatively correlated......Plasmacytoid dendritic cells (pDCs) play a vital role in activation of anti-HIV-1 immunity, and suppression of pDCs might mitigate immune responses against HIV-1. HIV-1 gp120 high-mannose has been attributed immunosuppressive roles in human myeloid DCs, but no receptors for high-mannose have so far...... been reported on human pDCs. Here we show that upon activation with HIV-1 or by a synthetic compound triggering the same receptor in human pDCs as single-stranded RNA, human pDCs upregulate the mannose receptor (MR, CD206). To examine the functional outcome of this upregulation, inactivated intact...

  1. A single gp120 residue can affect HIV-1 tropism in macaques.

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    Gregory Q Del Prete

    2017-09-01

    Full Text Available Species-dependent variation in proteins that aid or limit virus replication determines the ability of lentiviruses to jump between host species. Identifying and overcoming these differences facilitates the development of animal models for HIV-1, including models based on chimeric SIVs that express HIV-1 envelope (Env glycoproteins, (SHIVs and simian-tropic HIV-1 (stHIV strains. Here, we demonstrate that the inherently poor ability of most HIV-1 Env proteins to use macaque CD4 as a receptor is improved during adaptation by virus passage in macaques. We identify a single amino acid, A281, in HIV-1 Env that consistently changes during adaptation in macaques and affects the ability of HIV-1 Env to use macaque CD4. Importantly, mutations at A281 do not markedly affect HIV-1 Env neutralization properties. Our findings should facilitate the design of HIV-1 Env proteins for use in non-human primate models and thus expedite the development of clinically relevant reagents for testing interventions against HIV-1.

  2. Molecular Characterization of Heterologous HIV-1gp120 Gene Expression Disruption in Mycobacterium bovis BCG Host Strain: A Critical Issue for Engineering Mycobacterial Based-Vaccine Vectors

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    Joan Joseph

    2010-01-01

    Full Text Available Mycobacterium bovis Bacillus Calmette-Guérin (BCG as a live vector of recombinant bacterial vaccine is a promising system to be used. In this study, we evaluate the disrupted expression of heterologous HIV-1gp120 gene in BCG Pasteur host strain using replicative vectors pMV261 and pJH222. pJH222 carries a lysine complementing gene in BCG lysine auxotrophs. The HIV-1 gp120 gene expression was regulated by BCG hsp60 promoter (in plasmid pMV261 and Mycobacteria spp. α-antigen promoter (in plasmid pJH222. Among 14 rBCG:HIV-1gp120 (pMV261 colonies screened, 12 showed a partial deletion and two showed a complete deletion. However, deletion was not observed in all 10 rBCG:HIV-1gp120 (pJH222 colonies screened. In this study, we demonstrated that E. coli/Mycobacterial expression vectors bearing a weak promoter and lysine complementing gene in a recombinant lysine auxotroph of BCG could prevent genetic rearrangements and disruption of HIV 1gp120 gene expression, a key issue for engineering Mycobacterial based vaccine vectors.

  3. Computer-aided design of novel HIV-1 entry inhibitors blocking the virus envelope gp120 V3 loop

    Directory of Open Access Journals (Sweden)

    Tuzikov A. V.

    2012-12-01

    Full Text Available Aim. The object of this study was to implement computer-aided design of the water-soluble analog of glycolipid β -galactosylceramide (β-GalCer, which presents a potential HIV-1 entry inhibitor, by the analysis of intermolecular interactions of β-GalCer with the central region of the virus envelope gp120 V3 loop followed by synthesis of this glycolipid derivative and testing for antiviral activity. Methods. To reach the object of view, computer modeling procedures, such as quantum chemical calculations, molecular docking, molecular dynamics and free energy simulations, were involved in the studies in conjunction with chemical synthesis and anti-HIV-1 assay methods. Results. As a result, the high probability of exhibiting of antiviral activity was predicted for the designed β-GalCer analog. The data of molecular modeling were confirmed by those of primary medical trials of the synthesized compound. Conclusions. In the light of the findings obtained, the designed analog of β-GalCer may be considered as the basic structure for simulation of its more potent structural forms and for posterior selection of drug candidates most promising for synthesis and anti-HIV-1 assays.

  4. HIV-1 gp120 neurotoxicity proximally and at a distance from the point of exposure: protection by rSV40 delivery of antioxidant enzymes.

    Science.gov (United States)

    Louboutin, Jean-Pierre; Agrawal, Lokesh; Reyes, Beverly A S; Van Bockstaele, Elisabeth J; Strayer, David S

    2009-06-01

    Toxicity of HIV-1 envelope glycoprotein (gp120) for substantia nigra (SN) neurons may contribute to the Parkinsonian manifestations often seen in HIV-1-associated dementia (HAD). We studied the neurotoxicity of gp120 for dopaminergic neurons and potential neuroprotection by antioxidant gene delivery. Rats were injected stereotaxically into their caudate-putamen (CP); CP and (substantia nigra) SN neuron loss was quantified. The area of neuron loss extended several millimeters from the injection site, approximately 35% of the CP area. SN neurons, outside of this area of direct neurotoxicity, were also severely affected. Dopaminergic SN neurons (expressing tyrosine hydroxylase, TH, in the SN and dopamine transporter, DAT, in the CP) were mostly affected: intra-CP gp120 caused approximately 50% DAT+ SN neuron loss. Prior intra-CP gene delivery of Cu/Zn superoxide dismutase (SOD1) or glutathione peroxidase (GPx1) protected SN neurons from intra-CP gp120. Thus, SN dopaminergic neurons are highly sensitive to HIV-1 gp120-induced neurotoxicity, and antioxidant gene delivery, even at a distance, is protective.

  5. Reduced expression of glutamate transporter EAAT2 and impaired glutamate transport in human primary astrocytes exposed to HIV-1 or gp120.

    Science.gov (United States)

    Wang, Zhuying; Pekarskaya, Olga; Bencheikh, Meryem; Chao, Wei; Gelbard, Harris A; Ghorpade, Anuja; Rothstein, Jeffrey D; Volsky, David J

    2003-07-20

    L-Glutamate is the major excitatory neurotransmitter in the brain. Astrocytes maintain low levels of synaptic glutamate by high-affinity uptake and defects in this function may lead to neuronal cell death by excitotoxicity. We tested the effects of HIV-1 and its envelope glycoprotein gp120 upon glutamate uptake and expression of glutamate transporters EAAT1 and EAAT2 in fetal human astrocytes in vitro. Astrocytes isolated from fetal tissues between 16 and 19 weeks of gestation expressed EAAT1 and EAAT2 RNA and proteins as detected by Northern blot analysis and immunoblotting, respectively, and the cells were capable of specific glutamate uptake. Exposure of astrocytes to HIV-1 or gp120 significantly impaired glutamate uptake by the cells, with maximum inhibition within 6 h, followed by gradual decline during 3 days of observation. HIV-1-infected cells showed a 59% reduction in V(max) for glutamate transport, indicating a reduction in the number of active transporter sites on the cell surface. Impaired glutamate transport after HIV-1 infection or gp120 exposure correlated with a 40-70% decline in steady-state levels of EAAT2 RNA and protein. EAAT1 RNA and protein levels were less affected. Treatment of astrocytes with tumor necrosis factor-alpha (TNF-alpha) decreased the expression of both EAAT1 and EAAT2, but neither HIV-1 nor gp120 were found to induce TNF-alpha production by astrocytes. These findings demonstrate that HIV-1 and gp120 induce transcriptional downmodulation of the EAAT2 transporter gene in human astrocytes and coordinately attenuate glutamate transport by the cells. Reduction of the ability of HIV-1-infected astrocytes to take up glutamate may contribute to the development of neurological disease.

  6. Structure-based stabilization of HIV-1 gp120 enhances humoral immune responses to the induced co-receptor binding site.

    Directory of Open Access Journals (Sweden)

    Barna Dey

    2009-05-01

    Full Text Available The human immunodeficiency virus type 1 (HIV-1 exterior envelope glycoprotein, gp120, possesses conserved binding sites for interaction with the primary virus receptor, CD4, and also for the co-receptor, generally CCR5. Although gp120 is a major target for virus-specific neutralizing antibodies, the gp120 variable elements and its malleable nature contribute to evasion of effective host-neutralizing antibodies. To understand the conformational character and immunogenicity of the gp120 receptor binding sites as potential vaccine targets, we introduced structure-based modifications to stabilize gp120 core proteins (deleted of the gp120 major variable regions into the conformation recognized by both receptors. Thermodynamic analysis of the re-engineered core with selected ligands revealed significant stabilization of the receptor-binding regions. Stabilization of the co-receptor-binding region was associated with a marked increase in on-rate of ligand binding to this site as determined by surface plasmon resonance. Rabbit immunization studies showed that the conformational stabilization of core proteins, along with increased ligand affinity, was associated with strikingly enhanced humoral immune responses against the co-receptor-binding site. These results demonstrate that structure-based approaches can be exploited to stabilize a conformational site in a large functional protein to enhance immunogenic responses specific for that region.

  7. Broad and potent HIV-1 neutralization by a human antibody that binds the gp41-gp120 interface

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Jinghe; Kang, Byong H.; Pancera, Marie; Lee, Jeong Hyun; Tong, Tommy; Feng, Yu; Imamichi, Hiromi; Georgiev, Ivelin S.; Chuang, Gwo-Yu; Druz, Aliaksandr; Doria-Rose, Nicole A.; Laub, Leo; Sliepen, Kwinten; van Gils, Marit J.; de la Peña, Alba Torrents; Derking, Ronald; Klasse, Per-Johan; Migueles, Stephen A.; Bailer, Robert T.; Alam, Munir; Pugach, Pavel; Haynes, Barton F.; Wyatt, Richard T.; Sanders, Rogier W.; Binley, James M.; Ward, Andrew B.; Mascola, John R.; Kwong, Peter D.; Connors, Mark [NIH

    2015-10-15

    The isolation of human monoclonal antibodies is providing important insights into the specificities that underlie broad neutralization of HIV-1 (reviewed in ref. 1). Here we report a broad and extremely potent HIV-specific monoclonal antibody, termed 35O22, which binds a novel HIV-1 envelope glycoprotein (Env) epitope. 35O22 neutralized 62% of 181 pseudoviruses with a half-maximum inhibitory concentration (IC50) <50 μg ml-1. The median IC50 of neutralized viruses was 0.033 μg ml-1, among the most potent thus far described. 35O22 did not bind monomeric forms of Env tested, but did bind the trimeric BG505 SOSIP.664. Mutagenesis and a reconstruction by negative-stain electron microscopy of the Fab in complex with trimer revealed that it bound to a conserved epitope, which stretched across gp120 and gp41. The specificity of 35O22 represents a novel site of vulnerability on HIV Env, which serum analysis indicates to be commonly elicited by natural infection. Binding to this new site of vulnerability may thus be an important complement to current monoclonal-antibody-based approaches to immunotherapies, prophylaxis and vaccine design.

  8. Structure of HIV-1 gp120 V1/V2 domain with broadly neutralizing antibody PG9

    Energy Technology Data Exchange (ETDEWEB)

    McLellan, Jason S.; Pancera, Marie; Carrico, Chris; Gorman, Jason; Julien, Jean-Philippe; Khayat, Reza; Louder, Robert; Pejchal, Robert; Sastry, Mallika; Dai, Kaifan; O’Dell, Sijy; Patel, Nikita; Shahzad-ul-Hussan, Syed; Yang, Yongping; Zhang, Baoshan; Zhou, Tongqing; Zhu, Jiang; Boyington, Jeffrey C.; Chuang, Gwo-Yu; Diwanji, Devan; Georgiev, Ivelin; Kwon, Young Do; Lee, Doyung; Louder, Mark K.; Moquin, Stephanie; Schmidt, Stephen D.; Yang, Zhi-Yong; Bonsignori, Mattia; Crump, John A.; Kapiga, Saidi H.; Sam, Noel E.; Haynes, Barton F.; Burton, Dennis R.; Koff, Wayne C.; Walker, Laura M.; Phogat, Sanjay; Wyatt, Richard; Orwenyo, Jared; Wang, Lai-Xi; Arthos, James; Bewley, Carole A.; Mascola, John R.; Nabel, Gary J.; Schief, William R.; Ward, Andrew B.; Wilson, Ian A.; Kwong, Peter D. (UWASH); (NIH); (Scripps); (Duke); (IAVI); (Maryland-MED)

    2012-12-13

    Variable regions 1 and 2 (V1/V2) of human immunodeficiency virus-1 (HIV-1) gp120 envelope glycoprotein are critical for viral evasion of antibody neutralization, and are themselves protected by extraordinary sequence diversity and N-linked glycosylation. Human antibodies such as PG9 nonetheless engage V1/V2 and neutralize 80% of HIV-1 isolates. Here we report the structure of V1/V2 in complex with PG9. V1/V2 forms a four-stranded {beta}-sheet domain, in which sequence diversity and glycosylation are largely segregated to strand-connecting loops. PG9 recognition involves electrostatic, sequence-independent and glycan interactions: the latter account for over half the interactive surface but are of sufficiently weak affinity to avoid autoreactivity. The structures of V1/V2-directed antibodies CH04 and PGT145 indicate that they share a common mode of glycan penetration by extended anionic loops. In addition to structurally defining V1/V2, the results thus identify a paradigm of antibody recognition for highly glycosylated antigens, which - with PG9 - involves a site of vulnerability comprising just two glycans and a strand.

  9. Identification and characterization of a broadly cross-reactive HIV-1 human monoclonal antibody that binds to both gp120 and gp41.

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    Mei-Yun Zhang

    Full Text Available Identification of broadly cross-reactive HIV-1-neutralizing antibodies (bnAbs may assist vaccine immunogen design. Here we report a novel human monoclonal antibody (mAb, designated m43, which co-targets the gp120 and gp41 subunits of the HIV-1 envelope glycoprotein (Env. M43 bound to recombinant gp140 s from various primary isolates, to membrane-associated Envs on transfected cells and HIV-1 infected cells, as well as to recombinant gp120 s and gp41 fusion intermediate structures containing N-trimer structure, but did not bind to denatured recombinant gp140 s and the CD4 binding site (CD4bs mutant, gp120 D368R, suggesting that the m43 epitope is conformational and overlaps the CD4bs on gp120 and the N-trimer structure on gp41. M43 neutralized 34% of the HIV-1 primary isolates from different clades and all the SHIVs tested in assays based on infection of peripheral blood mononuclear cells (PBMCs by replication-competent virus, but was less potent in cell line-based pseudovirus assays. In contrast to CD4, m43 did not induce Env conformational changes upon binding leading to exposure of the coreceptor binding site, enhanced binding of mAbs 2F5 and 4E10 specific for the membrane proximal external region (MPER of gp41 Envs, or increased gp120 shedding. The overall modest neutralization activity of m43 is likely due to the limited binding of m43 to functional Envs which could be increased by antibody engineering if needed. M43 may represent a new class of bnAbs targeting conformational epitopes overlapping structures on both gp120 and gp41. Its novel epitope and possibly new mechanism(s of neutralization could helpdesign improved vaccine immunogens and candidate therapeutics.

  10. Thermal hyperalgesia and mechanical allodynia produced by intrathecal administration of the human immunodeficiency virus-1 (HIV-1) envelope glycoprotein, gp120.

    Science.gov (United States)

    Milligan, E D; Mehmert, K K; Hinde, J L; Harvey, L O; Martin, D; Tracey, K J; Maier, S F; Watkins, L R

    2000-04-07

    Astrocytes and microglia in the spinal cord have recently been reported to contribute to the development of peripheral inflammation-induced exaggerated pain states. Both lowering of thermal pain threshold (thermal hyperalgesia) and lowering of response threshold to light tactile stimuli (mechanical allodynia) have been reported. The notion that spinal cord glia are potential mediators of such effects is based on the disruption of these exaggerated pain states by drugs thought to preferentially affect glial function. Activation of astrocytes and microglia can release many of the same substances that are known to mediate thermal hyperalgesia and mechanical allodynia. The aim of the present series of studies was to determine whether exaggerated pain states could also be created in rats by direct, intraspinal immune activation of astrocytes and microglia. The immune stimulus used was peri-spinal (intrathecal, i.t.) application of the Human Immunodeficiency Virus type 1 (HIV-1) envelope glycoprotein, gp120. This portion of HIV-1 is known to bind to and activate microglia and astrocytes. Robust thermal hyperalgesia (tail-flick, TF, and Hargreaves tests) and mechanical allodynia (von Frey and touch-evoked agitation tests) were observed in response to i.t. gp120. Heat denaturing of the complex protein structure of gp120 blocked gp120-induced thermal hyperalgesia. Lastly, both thermal hyperalgesia and mechanical allodynia to i.t. gp120 were blocked by spinal pretreatment with drugs (fluorocitrate and CNI-1493) thought to preferentially disrupt glial function.

  11. Correlation between carbohydrate structures on the envelope glycoprotein gp120 of HIV-1 and HIV-2 and syncytium inhibition with lectins

    DEFF Research Database (Denmark)

    Hansen, J E; Nielsen, C M; Nielsen, C

    1989-01-01

    The binding of 13 different lectins to gp120 partially purified from two HIV-1 isolates and one HIV-2 isolate was studied by in situ staining on electrophoretically separated and electroblotted HIV antigens. The lectins concanavalin A, wheat germ agglutinin, Lens culinaris agglutinin, Vicia faba...

  12. Heterologous protection elicited by candidate monomeric recombinant HIV-1 gp120 vaccine in the absence of cross neutralising antibodies in a macaque model

    Directory of Open Access Journals (Sweden)

    Page Mark

    2012-07-01

    Full Text Available Abstract Background Current data suggest that an efficacious human immunodeficiency virus type 1 (HIV-1 vaccine should elicit both adaptive humoral and cell mediated immune responses. Such a vaccine will also need to protect against infection from a range of heterologous viral variants. Here we have developed a simian-human immunodeficiency virus (SHIV based model in cynomolgus macaques to investigate the breadth of protection conferred by HIV-1W61D recombinant gp120 vaccination against SHIVsbg and SHIVSF33 challenge, and to identify correlates of protection. Results High titres of anti-envelope antibodies were detected in all vaccinees. The antibodies reacted with both the homologous HIV-1W61D and heterologous HIV-1IIIB envelope rgp120 which has an identical sequence to the SHIVsbg challenge virus. Significant titres of virus neutralising antibodies were detected against SHIVW61D expressing an envelope homologous with the vaccine, but only limited cross neutralisation against SHIVsbg, SHIV-4 and SHIVSF33 was observed. Protection against SHIVsbg infection was observed in vaccinated animals but none was observed against SHIVSF33 challenge. Transfer of immune sera from vaccinated macaques to naive recipients did not confer protection against SHIVsbg challenge. In a follow-up study, T cell proliferative responses detected after immunisation with the same vaccine against a single peptide present in the second conserved region 2 of HIV-1 W61D and HIV-1 IIIB gp120, but not SF33 gp120. Conclusions Following extended vaccination with a HIV-1 rgp120 vaccine, protection was observed against heterologous virus challenge with SHIVsbg, but not SHIVSF33. Protection did not correlate with serological responses generated by vaccination, but might be associated with T cell proliferative responses against an epitope in the second constant region of HIV-1 gp120. Broader protection may be obtained with recombinant HIV-1 envelope based vaccines formulated with

  13. Correlation between carbohydrate structures on the envelope glycoprotein gp120 of HIV-1 and HIV-2 and syncytium inhibition with lectins

    DEFF Research Database (Denmark)

    Hansen, J E; Nielsen, C M; Nielsen, C

    1989-01-01

    agglutinin, Pisum sativum agglutinin and phytohaem(erythro)agglutinin bound to gp120 of all three isolates. The carbohydrate of gp120 recognized by lectins was thus arranged in at least four types of glycans: a high mannose type glycan, a bisected hybrid or complex type glycan, a biantennary fucosylated...... complex type glycan and a triantennary bisected complex type glycan. Only lectins which bound at least one of the four types of glycans were capable of inhibiting fusion of HIV-infected cells with CD4 cells by a carbohydrate-specific interaction with the HIV-infected cells. Thus, several different glycan......The binding of 13 different lectins to gp120 partially purified from two HIV-1 isolates and one HIV-2 isolate was studied by in situ staining on electrophoretically separated and electroblotted HIV antigens. The lectins concanavalin A, wheat germ agglutinin, Lens culinaris agglutinin, Vicia faba...

  14. Crystal structures of HIV-1 gp120 envelope glycoprotein in complex with NBD analogues that target the CD4-binding site.

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    Young Do Kwon

    Full Text Available Efforts to develop therapeutic agents that inhibit HIV-1 entry have led to the identification of several small molecule leads. One of the most promising is the NBD series, which binds within a conserved gp120 cavity and possesses para-halogen substituted aromatic rings, a central oxalamide linker, and a tetramethylpiperidine moiety. In this study, we characterized structurally the interactions of four NBD analogues containing meta-fluoro substitution on the aromatic ring and various heterocyclic ring replacements of the tetramethylpiperidine group. The addition of a meta-fluorine to the aromatic ring improved surface complementarity and did not alter the position of the analogue relative to gp120. By contrast, heterocyclic ring replacements of the tetramethylpiperidine moiety exhibited diverse positioning and interactions with the vestibule of the gp120 cavity. Overall, the biological profile of NBD-congeners was modulated by ligand interactions with the gp120-cavity vestibule. Herein, six co-crystal structures of NBD-analogues with gp120 provide a structural framework for continued small molecule-entry inhibitor optimization.

  15. Probability analysis of variational crystallization and its application to gp120, the exterior envelope glycoprotein of type 1 human immunodeficiency virus (HIV-1).

    Science.gov (United States)

    Kwong, P D; Wyatt, R; Desjardins, E; Robinson, J; Culp, J S; Hellmig, B D; Sweet, R W; Sodroski, J; Hendrickson, W A

    1999-02-12

    The extensive glycosylation and conformational mobility of gp120, the envelope glycoprotein of type 1 human immunodeficiency virus (HIV-1), pose formidable barriers for crystallization. To surmount these difficulties, we used probability analysis to determine the most effective crystallization approach and derive equations which show that a strategy, which we term variational crystallization, substantially enhances the overall probability of crystallization for gp120. Variational crystallization focuses on protein modification as opposed to crystallization screening. Multiple variants of gp120 were analyzed with an iterative cycle involving a limited set of crystallization conditions and biochemical feedback on protease sensitivity, glycosylation status, and monoclonal antibody binding. Sources of likely conformational heterogeneity such as N-linked carbohydrates, flexible or mobile N and C termini, and variable internal loops were reduced or eliminated, and ligands such as CD4 and antigen-binding fragments (Fabs) of monoclonal antibodies were used to restrict conformational mobility as well as to alter the crystallization surface. Through successive cycles of manipulation involving 18 different variants, we succeeded in growing six different types of gp120 crystals. One of these, a ternary complex composed of gp120, its receptor CD4, and the Fab of the human neutralizing monoclonal antibody 17b, diffracts to a minimum Bragg spacing of at least 2.2 A and is suitable for structural analysis.

  16. In silico analysis of HIV-1 Env-gp120 reveals structural bases for viral adaptation in growth-restrictive cells

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    Masaru eYokoyama

    2016-02-01

    Full Text Available Variable V1/V2 and V3 loops on human immunodeficiency virus type 1 (HIV-1 envelope-gp120 core play key roles in modulating viral competence to recognize two infection receptors, CD4 and chemokine-receptors. However, molecular bases for the modulation largely remain unclear. To address these issues, we constructed structural models for a full-length gp120 in CD4-free and -bound states. The models showed topologies of gp120 surface loop that agree with those in reported structural data. Molecular dynamics simulation showed that in the unliganded state, V1/V2 loop settled into a thermodynamically stable arrangement near V3 loop for conformational masking of V3 tip, a potent neutralization epitope. In the CD4-bound state, however, V1/V2 loop was rearranged near the bound CD4 to support CD4 binding. In parallel, cell-based adaptation in the absence of anti-viral antibody pressures led to the identification of amino acid substitutions that individually enhance viral entry and growth efficiencies in association with reduced sensitivity to CCR5 antagonist TAK-779. Notably, all these substitutions were positioned on the receptors binding surfaces in V1/V2 or V3 loop. In silico structural studies predicted some physical changes of gp120 by substitutions with alterations in viral replication phenotypes. These data suggest that V1/V2 loop is critical for creating a gp120 structure that masks co-receptor binding site compatible with maintenance of viral infectivity, and for tuning a functional balance of gp120 between immune escape ability and infectivity to optimize HIV-1 replication fitness.

  17. A Peptide Derived from the HIV-1 gp120 Coreceptor-Binding Region Promotes Formation of PAP248-286 Amyloid Fibrils to Enhance HIV-1 Infection.

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    Jinquan Chen

    Full Text Available Semen is a major vehicle for HIV transmission. Prostatic acid phosphatase (PAP fragments, such as PAP248-286, in human semen can form amyloid fibrils to enhance HIV infection. Other endogenous or exogenous factors present during sexual intercourse have also been reported to promote the formation of seminal amyloid fibrils.Here, we demonstrated that a synthetic 15-residue peptide derived from the HIV-1 gp120 coreceptor-binding region, designated enhancing peptide 2 (EP2, can rapidly self-assemble into nanofibers. These EP2-derivated nanofibers promptly accelerated the formation of semen amyloid fibrils by PAP248-286, as shown by Thioflavin T (ThT and Congo red assays. The amyloid fibrils presented similar morphology, assessed via transmission electron microscopy (TEM, in the presence or absence of EP2. Circular dichroism (CD spectroscopy revealed that EP2 accelerates PAP248-286 amyloid fibril formation by promoting the structural transition of PAP248-286 from a random coil into a cross-β-sheet. Newly formed semen amyloid fibrils effectively enhanced HIV-1 infection in TZM-bl cells and U87 cells by promoting the binding of HIV-1 virions to target cells.Nanofibers composed of EP2 promote the formation of PAP248-286 amyloid fibrils and enhance HIV-1 infection.

  18. A human antibody to the CD4 binding site of gp120 capable of highly potent but sporadic cross clade neutralization of primary HIV-1.

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    Johannes S Gach

    Full Text Available Primary isolates of HIV-1 resist neutralization by most antibodies to the CD4 binding site (CD4bs on gp120 due to occlusion of this site on the trimeric spike. We describe 1F7, a human CD4bs monoclonal antibody that was found to be exceptionally potent against the HIV-1 primary isolate JR-FL. However, 1F7 failed to neutralize a patient-matched primary isolate, JR-CSF even though the two isolates differ by <10% in gp120 at the protein level. In an HIV-1 cross clade panel (n = 157, 1F7 exhibited moderate breadth, but occasionally achieved considerable potency. In binding experiments using monomeric gp120s of select resistant isolates and domain-swap chimeras between JR-FL and JR-CSF, recognition by 1F7 was limited by sequence polymorphisms involving at least the C2 region of Env. Putative N-linked glycosylation site (PNGS mutations, notably at position 197, allowed 1F7 to neutralize JR-CSF potently without improving binding to the cognate, monomeric gp120. In contrast, flow cytometry experiments using the same PNGS mutants revealed that 1F7 binding is enhanced on cognate trimeric Env. BN-PAGE mobility shift experiments revealed that 1F7 is sensitive to the diagnostic mutation D368R in the CD4 binding loop of gp120. Our data on 1F7 reinforce how exquisitely targeted CD4bs antibodies must be to achieve cross neutralization of two closely related primary isolates. High-resolution analyses of trimeric Env that show the orientation of glycans and polymorphic elements of the CD4bs that affect binding to antibodies like 1F7 are desirable to understand how to promote immunogenicity of more conserved elements of the CD4bs.

  19. Effect of Glycosylation on an Immunodominant Region in the V1V2 Variable Domain of the HIV-1 Envelope gp120 Protein.

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    Jianhui Tian

    2016-10-01

    Full Text Available Heavy glycosylation of the envelope (Env surface subunit, gp120, is a key adaptation of HIV-1; however, the precise effects of glycosylation on the folding, conformation and dynamics of this protein are poorly understood. Here we explore the patterns of HIV-1 Env gp120 glycosylation, and particularly the enrichment in glycosylation sites proximal to the disulfide linkages at the base of the surface-exposed variable domains. To dissect the influence of glycans on the conformation these regions, we focused on an antigenic peptide fragment from a disulfide bridge-bounded region spanning the V1 and V2 hyper-variable domains of HIV-1 gp120. We used replica exchange molecular dynamics (MD simulations to investigate how glycosylation influences its conformation and stability. Simulations were performed with and without N-linked glycosylation at two sites that are highly conserved across HIV-1 isolates (N156 and N160; both are contacts for recognition by V1V2-targeted broadly neutralizing antibodies against HIV-1. Glycosylation stabilized the pre-existing conformations of this peptide construct, reduced its propensity to adopt other secondary structures, and provided resistance against thermal unfolding. Simulations performed in the context of the Env trimer also indicated that glycosylation reduces flexibility of the V1V2 region, and provided insight into glycan-glycan interactions in this region. These stabilizing effects were influenced by a combination of factors, including the presence of a disulfide bond between the Cysteines at 131 and 157, which increased the formation of beta-strands. Together, these results provide a mechanism for conservation of disulfide linkage proximal glycosylation adjacent to the variable domains of gp120 and begin to explain how this could be exploited to enhance the immunogenicity of those regions. These studies suggest that glycopeptide immunogens can be designed to stabilize the most relevant Env conformations to

  20. Chemokine co-receptor CCR5/CXCR4-dependent modulation of Kv2.1 channel confers acute neuroprotection to HIV-1 glycoprotein gp120 exposure.

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    Andrew J Shepherd

    Full Text Available Infection with human immunodeficiency virus-1 (HIV-1 within the brain has long been known to be associated with neurodegeneration and neurocognitive disorder (referred as HAND, a condition characterized in its early stages by declining cognitive function and behavioral disturbances. Mechanistically, the HIV-1 coat glycoprotein 120 (gp120 has been suggested to be a critical factor inducing apoptotic cell death in neurons via the activation of p38 mitogen-activated protein kinase (MAPK, upon chronic exposure to the virus. Here we show that acute exposure of neurons to HIV-1 gp120 elicits a homeostatic response, which provides protection against non-apoptotic cell death, involving the major somatodendritic voltage-gated K⁺ (Kv channel Kv2.1 as the key mediator. The Kv2.1 channel has recently been shown to provide homeostatic control of neuronal excitability under conditions of seizures, ischemia and neuromodulation/neuroinflammation. Following acute exposure to gp120, cultured rat hippocampal neurons show rapid dephosphorylation of the Kv2.1 protein, which ultimately leads to changes in specific sub-cellular localization and voltage-dependent channel activation properties of Kv2.1. Such modifications in Kv2.1 are dependent on the activation of the chemokine co-receptors CCR5 and CXCR4, and subsequent activation of the protein phosphatase calcineurin. This leads to the overall suppression of neuronal excitability and provides neurons with a homeostatic protective mechanism. Specific blockade of calcineurin and Kv2.1 channel activity led to significant enhancement of non-apoptotic neuronal death upon acute gp120 treatment. These observations shed new light on the intrinsic homeostatic mechanisms of neuronal resilience during the acute stages of neuro-HIV infections.

  1. Structure and Recognition of a Novel HIV-1 gp120-gp41 Interface Antibody that Caused MPER Exposure through Viral Escape

    Energy Technology Data Exchange (ETDEWEB)

    Wibmer, Constantinos Kurt; Gorman, Jason; Ozorowski, Gabriel; Bhiman, Jinal N.; Sheward, Daniel J.; Elliott, Debra H.; Rouelle, Julie; Smira, Ashley; Joyce, M. Gordon; Ndabambi, Nonkululeko; Druz, Aliaksandr; Asokan, Mangai; Burton, Dennis R.; Connors, Mark; Abdool Karim, Salim S.; Mascola, John R.; Robinson, James E.; Ward, Andrew B.; Williamson, Carolyn; Kwong, Peter D.; Morris, Lynn; Moore, Penny L.; Desrosiers, Ronald C.

    2017-01-11

    A comprehensive understanding of the regions on HIV-1 envelope trimers targeted by broadly neutralizing antibodies may contribute to rational design of an HIV-1 vaccine. We previously identified a participant in the CAPRISA cohort, CAP248, who developed trimer-specific antibodies capable of neutralizing 60% of heterologous viruses at three years post-infection. Here, we report the isolation by B cell culture of monoclonal antibody CAP248-2B, which targets a novel membrane proximal epitope including elements of gp120 and gp41. Despite low maximum inhibition plateaus, often below 50% inhibitory concentrations, the breadth of CAP248-2B significantly correlated with donor plasma. Site-directed mutagenesis, X-ray crystallography, and negative-stain electron microscopy 3D reconstructions revealed how CAP248-2B recognizes a cleavage-dependent epitope that includes the gp120 C terminus. While this epitope is distinct, it overlapped in parts of gp41 with the epitopes of broadly neutralizing antibodies PGT151, VRC34, 35O22, 3BC315, and 10E8. CAP248-2B has a conformationally variable paratope with an unusually long 19 amino acid light chain third complementarity determining region. Two phenylalanines at the loop apex were predicted by docking and mutagenesis data to interact with the viral membrane. Neutralization by CAP248-2B is not dependent on any single glycan proximal to its epitope, and low neutralization plateaus could not be completely explained by N- or O-linked glycosylation pathway inhibitors, furin co-transfection, or pre-incubation with soluble CD4. Viral escape from CAP248-2B involved a cluster of rare mutations in the gp120-gp41 cleavage sites. Simultaneous introduction of these mutations into heterologous viruses abrogated neutralization by CAP248-2B, but enhanced neutralization sensitivity to 35O22, 4E10, and 10E8 by 10-100-fold. Altogether, this study expands the region of the HIV-1 gp120-gp41 quaternary interface that is a target for broadly neutralizing

  2. Structure-guided Design and Immunological Characterization of Immunogens Presenting the HIV-1 gp120 V3 Loop on a CTB Scaffold

    Energy Technology Data Exchange (ETDEWEB)

    M Totrov; X Jiang; X Kong; S Cohen; C Krachmarov; A Salomon; C Williams; M Seaman; R Abagyan; et al.

    2011-12-31

    V3 loop is a major neutralizing determinant of the HIV-1 gp120. Using 3D structures of cholera toxin B subunit (CTB), complete V3 in the gp120 context, and V3 bound to a monoclonal antibody (mAb), we designed two V3-scaffold immunogen constructs (V3-CTB). The full-length V3-CTB presenting the complete V3 in a structural context mimicking gp120 was recognized by the large majority of our panel of 24 mAbs. The short V3-CTB presenting a V3 fragment in the conformation observed in the complex with the 447-52D Fab, exhibited high-affinity binding to this mAb. The immunogens were evaluated in rabbits using DNA-prime/protein-boost protocol. Boosting with the full-length V3-CTB induced high anti-V3 titers in sera that potently neutralize multiple HIV virus strains. The short V3-CTB was ineffective. The results suggest that very narrow antigenic profile of an immunogen is associated with poor Ab response. An immunogen with broader antigenic activity elicits robust Ab response.

  3. Influence of the Envelope gp120 Phe 43 Cavity on HIV-1 Sensitivity to Antibody-Dependent Cell-Mediated Cytotoxicity Responses.

    Science.gov (United States)

    Prévost, Jérémie; Zoubchenok, Daria; Richard, Jonathan; Veillette, Maxime; Pacheco, Beatriz; Coutu, Mathieu; Brassard, Nathalie; Parsons, Matthew S; Ruxrungtham, Kiat; Bunupuradah, Torsak; Tovanabutra, Sodsai; Hwang, Kwan-Ki; Moody, M Anthony; Haynes, Barton F; Bonsignori, Mattia; Sodroski, Joseph; Kaufmann, Daniel E; Shaw, George M; Chenine, Agnès L; Finzi, Andrés

    2017-04-01

    HIV-1-infected cells presenting envelope glycoproteins (Env) in the CD4-bound conformation on their surface are preferentially targeted by antibody-dependent cellular-mediated cytotoxicity (ADCC). HIV-1 has evolved sophisticated mechanisms to avoid the exposure of Env ADCC epitopes by downregulating CD4 and by limiting the overall amount of Env on the cell surface. In HIV-1, substitution of large residues such as histidine or tryptophan for serine 375 (S375H/W) in the gp120 Phe 43 cavity, where Phe 43 of CD4 contacts gp120, results in the spontaneous sampling of an Env conformation closer to the CD4-bound state. While residue S375 is well conserved in the majority of group M HIV-1 isolates, CRF01_AE strains have a naturally occurring histidine at this position (H375). Interestingly, CRF01_AE is the predominant circulating strain in Thailand, where the RV144 trial took place. In this trial, which resulted in a modest degree of protection, ADCC responses were identified as being part of the correlate of protection. Here we investigate the influence of the Phe 43 cavity on ADCC responses. Filling this cavity with a histidine or tryptophan residue in Env with a natural serine residue at this position (S375H/W) increased the susceptibility of HIV-1-infected cells to ADCC. Conversely, the replacement of His 375 by a serine residue (H375S) within HIV-1 CRF01_AE decreased the efficiency of the ADCC response. Our results raise the intriguing possibility that the presence of His 375 in the circulating strain where the RV144 trial was held contributed to the observed vaccine efficacy.IMPORTANCE HIV-1-infected cells presenting Env in the CD4-bound conformation on their surface are preferentially targeted by ADCC mediated by HIV-positive (HIV(+)) sera. Here we show that the gp120 Phe 43 cavity modulates the propensity of Env to sample this conformation and therefore affects the susceptibility of infected cells to ADCC. CRF01_AE HIV-1 strains have an unusual Phe 43 cavity

  4. The alpha helix 1 from the first conserved region of HIV1 gp120 is reconstructed in the short NQ21 peptide.

    Science.gov (United States)

    Khrustalev, Vladislav Victorovich; Khrustaleva, Tatyana Aleksandrovna; Kahanouskaya, Ekaterina Yurievna; Rudnichenko, Yulia Anatolyevna; Bandarenka, Hanna Vitalyevna; Arutyunyan, Alexander Migranovich; Girel, Kseniya Victorovna; Khinevich, Nadia Vladimirovna; Ksenofontov, Alexander Leonidovich; Kordyukova, Larisa Valentinovna

    2018-01-15

    Investigations of short peptides that can be used in the next phase of synthetic HIV1 vaccine development are an urgent goal, as well as investigations of peptides that can be used in immunological tests with the aim to check the titer of antibodies against the alpha helix 1 from the first conserved region of HIV1 gp120 that are known to cause antibody-dependent cellular cytotoxicity (ADCC). The aim of this work was to study the structure of the NQ21 peptide corresponding to the less mutable part of the first conserved region of HIV1 gp120 (residues 94-114). The NQ21 peptide and its conjugate with biotin (biotin-NQ21) are absolutely alpha-helical in phosphate buffer solutions at pH = 6.8, 7.4 and 8.0, as well as in the dried form, according to the results of surface-enhanced Raman scattering (SERS) spectroscopy. Results of the native gel electrophoresis and thermal analysis under the control of spectrofluorometer and near UV circular dichroism (CD) showed that the peptide exists in form of octamers and tetramers at pH = 7.4, that is important information for further vaccine development. Strong signal of interacting Trp residues in oligomers in the far UV CD obscures the signal from secondary structure, but becomes less intensive during the heating. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. Broad-spectrum inhibition of HIV-1 by a monoclonal antibody directed against a gp120-induced epitope of CD4.

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    Samuele E Burastero

    Full Text Available To penetrate susceptible cells, HIV-1 sequentially interacts with two highly conserved cellular receptors, CD4 and a chemokine receptor like CCR5 or CXCR4. Monoclonal antibodies (MAbs directed against such receptors are currently under clinical investigation as potential preventive or therapeutic agents. We immunized Balb/c mice with molecular complexes of the native, trimeric HIV-1 envelope (Env bound to a soluble form of the human CD4 receptor. Sera from immunized mice were found to contain gp120-CD4 complex-enhanced antibodies and showed broad-spectrum HIV-1-inhibitory activity. A proportion of MAbs derived from these mice preferentially recognized complex-enhanced epitopes. In particular, a CD4-specific MAb designated DB81 (IgG1Κ was found to preferentially bind to a complex-enhanced epitope on the D2 domain of human CD4. MAb DB81 also recognized chimpanzee CD4, but not baboon or macaque CD4, which exhibit sequence divergence in the D2 domain. Functionally, MAb DB81 displayed broad HIV-1-inhibitory activity, but it did not exert suppressive effects on T-cell activation in vitro. The variable regions of the heavy and light chains of MAb DB81 were sequenced. Due to its broad-spectrum anti-HIV-1 activity and lack of immunosuppressive effects, a humanized derivative of MAb DB81 could provide a useful complement to current preventive or therapeutic strategies against HIV-1.

  6. HIV-1 and its gp120 inhibits the influenza A(H1N1pdm09 life cycle in an IFITM3-dependent fashion.

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    Milene Mesquita

    Full Text Available HIV-1-infected patients co-infected with A(H1N1pdm09 surprisingly presented benign clinical outcome. The knowledge that HIV-1 changes the host homeostatic equilibrium, which may favor the patient resistance to some co-pathogens, prompted us to investigate whether HIV-1 infection could influence A(H1N1pdm09 life cycle in vitro. We show here that exposure of A(H1N1pdm09-infected epithelial cells to HIV-1 viral particles or its gp120 enhanced by 25% the IFITM3 content, resulting in a decrease in influenza replication. This event was dependent on toll-like receptor 2 and 4. Moreover, knockdown of IFITM3 prevented HIV-1 ability to inhibit A(H1N1pdm09 replication. HIV-1 infection also increased IFITM3 levels in human primary macrophages by almost 100%. Consequently, the arrival of influenza ribonucleoproteins (RNPs to nucleus of macrophages was inhibited, as evaluated by different approaches. Reduction of influenza RNPs entry into the nucleus tolled A(H1N1pdm09 life cycle in macrophages earlier than usual, limiting influenza's ability to induce TNF-α. As judged by analysis of the influenza hemagglutin (HA gene from in vitro experiments and from samples of HIV-1/A(H1N1pdm09 co-infected individuals, the HIV-1-induced reduction of influenza replication resulted in delayed viral evolution. Our results may provide insights on the mechanisms that may have attenuated the clinical course of Influenza in HIV-1/A(H1N1pdm09 co-infected patients during the recent influenza form 2009/2010.

  7. HIV-1 and its gp120 inhibits the influenza A(H1N1)pdm09 life cycle in an IFITM3-dependent fashion.

    Science.gov (United States)

    Mesquita, Milene; Fintelman-Rodrigues, Natalia; Sacramento, Carolina Q; Abrantes, Juliana L; Costa, Eduardo; Temerozo, Jairo R; Siqueira, Marilda M; Bou-Habib, Dumith Chequer; Souza, Thiago Moreno L

    2014-01-01

    HIV-1-infected patients co-infected with A(H1N1)pdm09 surprisingly presented benign clinical outcome. The knowledge that HIV-1 changes the host homeostatic equilibrium, which may favor the patient resistance to some co-pathogens, prompted us to investigate whether HIV-1 infection could influence A(H1N1)pdm09 life cycle in vitro. We show here that exposure of A(H1N1)pdm09-infected epithelial cells to HIV-1 viral particles or its gp120 enhanced by 25% the IFITM3 content, resulting in a decrease in influenza replication. This event was dependent on toll-like receptor 2 and 4. Moreover, knockdown of IFITM3 prevented HIV-1 ability to inhibit A(H1N1)pdm09 replication. HIV-1 infection also increased IFITM3 levels in human primary macrophages by almost 100%. Consequently, the arrival of influenza ribonucleoproteins (RNPs) to nucleus of macrophages was inhibited, as evaluated by different approaches. Reduction of influenza RNPs entry into the nucleus tolled A(H1N1)pdm09 life cycle in macrophages earlier than usual, limiting influenza's ability to induce TNF-α. As judged by analysis of the influenza hemagglutin (HA) gene from in vitro experiments and from samples of HIV-1/A(H1N1)pdm09 co-infected individuals, the HIV-1-induced reduction of influenza replication resulted in delayed viral evolution. Our results may provide insights on the mechanisms that may have attenuated the clinical course of Influenza in HIV-1/A(H1N1)pdm09 co-infected patients during the recent influenza form 2009/2010.

  8. Accurate and efficient gp120 V3 loop structure based models for the determination of HIV-1 co-receptor usage

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    Vaisman Iosif I

    2010-10-01

    Full Text Available Abstract Background HIV-1 targets human cells expressing both the CD4 receptor, which binds the viral envelope glycoprotein gp120, as well as either the CCR5 (R5 or CXCR4 (X4 co-receptors, which interact primarily with the third hypervariable loop (V3 loop of gp120. Determination of HIV-1 affinity for either the R5 or X4 co-receptor on host cells facilitates the inclusion of co-receptor antagonists as a part of patient treatment strategies. A dataset of 1193 distinct gp120 V3 loop peptide sequences (989 R5-utilizing, 204 X4-capable is utilized to train predictive classifiers based on implementations of random forest, support vector machine, boosted decision tree, and neural network machine learning algorithms. An in silico mutagenesis procedure employing multibody statistical potentials, computational geometry, and threading of variant V3 sequences onto an experimental structure, is used to generate a feature vector representation for each variant whose components measure environmental perturbations at corresponding structural positions. Results Classifier performance is evaluated based on stratified 10-fold cross-validation, stratified dataset splits (2/3 training, 1/3 validation, and leave-one-out cross-validation. Best reported values of sensitivity (85%, specificity (100%, and precision (98% for predicting X4-capable HIV-1 virus, overall accuracy (97%, Matthew's correlation coefficient (89%, balanced error rate (0.08, and ROC area (0.97 all reach critical thresholds, suggesting that the models outperform six other state-of-the-art methods and come closer to competing with phenotype assays. Conclusions The trained classifiers provide instantaneous and reliable predictions regarding HIV-1 co-receptor usage, requiring only translated V3 loop genotypes as input. Furthermore, the novelty of these computational mutagenesis based predictor attributes distinguishes the models as orthogonal and complementary to previous methods that utilize sequence

  9. HIV-1 subtype C primary isolates exhibit high sensitivity to an anti-gp120 RNA aptamer

    CSIR Research Space (South Africa)

    Mufhandu, Hazel T

    2012-09-01

    Full Text Available Globally, HIV-1 subtype C is the most prevalent subtype, yet most antiretroviral drugs are developed against subtype B. UCLA1 RNA aptamer, which we previously showed neutralizes HIV-1 subtype C Env-pseudotyped viruses was examined for neutralization...

  10. Entropy calculation of HIV-1 Env gp120, its receptor CD4, and their complex: an analysis of configurational entropy changes upon complexation.

    Science.gov (United States)

    Hsu, Shang-Te D; Peter, Christine; van Gunsteren, Wilfred F; Bonvin, Alexandre M J J

    2005-01-01

    The HIV-1 gp120/CD4 interaction shows a large, unprecedented entropy/enthalpy compensation, with the capacity to fine-tune recognition over a broad range of affinity. The intermolecular interaction involves stable hydrophobic contacts with a unique protruding CD4-Phe43 structure surrounded by an intermolecular hydrogen-bond network that covers the hemisphere of the CD4 D1 domain. We have applied a heuristic formula based on the covariance matrix of atom-positional fluctuations to assess the configurational entropy of the gp120/CD4 complex at different levels. The system was dissected into various subsets of atoms to evaluate the entropic contributions of different functional elements. By combining the trajectories of the free and complex forms, further insight into the conformational sampling was extracted. Despite the limited sampling time of 10 ns, the theoretically derived changes in configurational entropy are in fair agreement with the experimentally determined data. The simultaneous evaluation of different interaction modes through a decomposition approach is only feasible with the knowledge of the atomic trajectory of the system. The configurational entropy analysis in terms of combined trajectories presented here shall potentially provide accurate estimations of thermodynamic properties of biomolecules given sufficient sampling of conformational space.

  11. Asn 362 in gp120 contributes to enhanced fusogenicity by CCR5-restricted HIV-1 envelope glycoprotein variants from patients with AIDS

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    Wang Bin

    2007-12-01

    Full Text Available Abstract Background CCR5-restricted (R5 human immunodeficiency virus type 1 (HIV-1 variants cause CD4+ T-cell loss in the majority of individuals who progress to AIDS, but mechanisms underlying the pathogenicity of R5 strains are poorly understood. To better understand envelope glycoprotein (Env determinants contributing to pathogenicity of R5 viruses, we characterized 37 full-length R5 Envs from cross-sectional and longitudinal R5 viruses isolated from blood of patients with asymptomatic infection or AIDS, referred to as pre-AIDS (PA and AIDS (A R5 Envs, respectively. Results Compared to PA-R5 Envs, A-R5 Envs had enhanced fusogenicity in quantitative cell-cell fusion assays, and reduced sensitivity to inhibition by the fusion inhibitor T-20. Sequence analysis identified the presence of Asn 362 (N362, a potential N-linked glycosylation site immediately N-terminal to CD4-binding site (CD4bs residues in the C3 region of gp120, more frequently in A-R5 Envs than PA-R5 Envs. N362 was associated with enhanced fusogenicity, faster entry kinetics, and increased sensitivity of Env-pseudotyped reporter viruses to neutralization by the CD4bs-directed Env mAb IgG1b12. Mutagenesis studies showed N362 contributes to enhanced fusogenicity of most A-R5 Envs. Molecular models indicate N362 is located adjacent to the CD4 binding loop of gp120, and suggest N362 may enhance fusogenicity by promoting greater exposure of the CD4bs and/or stabilizing the CD4-bound Env structure. Conclusion Enhanced fusogenicity is a phenotype of the A-R5 Envs studied, which was associated with the presence of N362, enhanced HIV-1 entry kinetics and increased CD4bs exposure in gp120. N362 contributes to fusogenicity of R5 Envs in a strain dependent manner. Our studies suggest enhanced fusogenicity of A-R5 Envs may contribute to CD4+ T-cell loss in subjects who progress to AIDS whilst harbouring R5 HIV-1 variants. N362 may contribute to this effect in some individuals.

  12. Energetics of dendrimer binding to HIV-1 gp120-CD4 complex and mechanismic aspects of its role as an entry-inhibitor

    Science.gov (United States)

    Saurabh, Suman; Sahoo, Anil Kumar; Maiti, Prabal K.

    2016-10-01

    Experiments and computational studies have established that de-protonated dendrimers (SPL7013 and PAMAM) act as entry-inhibitors of HIV. SPL7013 based Vivagel is currently under clinical development. The dendrimer binds to gp120 in the gp120-CD4 complex, destabilizes it by breaking key contacts between gp120 and CD4 and prevents viral entry into target cells. In this work, we provide molecular details and energetics of the formation of the SPL7013-gp120-CD4 ternary complex and decipher modes of action of the dendrimer in preventing viral entry. It is also known from experiments that the dendrimer binds weakly to gp120 that is not bound to CD4. It binds even more weakly to the CD4-binding region of gp120 and thus cannot directly block gp120-CD4 complexation. In this work, we examine the feasibility of dendrimer binding to the gp120-binding region of CD4 and directly blocking gp120-CD4 complex formation. We find that the process of the dendrimer binding to CD4 can compete with gp120-CD4 binding due to comparable free energy change for the two processes, thus creating a possibility for the dendrimer to directly block gp120-CD4 complexation by binding to the gp120-binding region of CD4.

  13. Identification of an N-linked glycan in the V1-loop of HIV-1 gp120 influencing neutralization by anti-V3 antibodies and soluble CD4

    DEFF Research Database (Denmark)

    Gram, G J; Hemming, A; Bolmstedt, A

    1994-01-01

    affecting viral infectivity in cell culture. We found that the mutated virus lacking an N-linked glycan in the V1-loop of gp120 was more resistant to neutralization by monoclonal antibodies to the V3-loop and neutralization by soluble recombinant CD4 (sCD4). Both viruses were equally well neutralized by Con...... in the V1-loop of HIV-1 gp120. Lack of an N-linked glycan was verified by a mobility enhancement of mutant gp120 in SDS-gel electrophoresis. The mutated virus showed no differences in either gp120 content per infectious unit or infectivity, indicating that the N-linked glycan was neither essential nor...

  14. Molecular interaction of gp120 and B40 aptamer: A potential new HIV-1 entry inhibitor drug

    CSIR Research Space (South Africa)

    Baron, MK

    2008-11-01

    Full Text Available HIV-1 infection and its concomitant disease – Aids, remain major public health problems in southern Africa. While current antiretroviral drugs have prolonged the quality of life for many HIV-positive individuals, they do not eliminate the virus (2...

  15. Binding of the mannose-specific lectin, griffithsin, to HIV-1 gp120 exposes the CD4-binding site

    CSIR Research Space (South Africa)

    Alexandre, Kabamba B

    2011-09-01

    Full Text Available :359-69. 37 8. Burrer, R., S. Haessig-Einius, A. M. Aubertin, and C. Moog. 2005. 38 Neutralizing as well as non-neutralizing polyclonal immunoglobulin (Ig)G 39 from infected patients capture HIV-1 via antibodies directed against the 40 principal.... Moore, G. D. Tomaras, C. K. 5 Wibmer, A. Puren, A. DeCamp, P. B. Gilbert, B. Wood, D. C. Montefiori, 6 J. M. Binley, G. M. Shaw, B. F. Haynes, J. R. Mascola, and L. Morris. 7 2009. Antibody specificities associated with neutralization breadth...

  16. The HIV-1 gp120/V3 modifies the response of uninfected CD4 T cells to antigen presentation: mapping of the specific transcriptional signature

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    Spandidos Demetrios A

    2011-09-01

    Full Text Available Abstract Background The asymptomatic phase of HIV-1 infection is characterized by a progressive depletion of uninfected peripheral effector/memory CD4+ T cells that subsequently leads to immune dysfunction and AIDS symptoms. We have previously demonstrated that the presence of specific gp120/V3 peptides during antigen presentation can modify the activation of normal T-cells leading to altered immune function. The aim of the present study was to map the specific transcriptional profile invoked by an HIV-1/V3 epitope in uninfected T cells during antigen presentation. Methods We exposed primary human peripheral blood monocytes to V3 lipopeptides using a liposome delivery system followed by a superantigen-mediated antigen presentation system. We then evaluated the changes in the T-cell transcriptional profile using oligonucleotide microarrays and performed Ingenuity Pathway Analysis (IPA and DAVID analysis. The results were validated using realtime PCR, FACS, Western blotting and immunofluorescence. Results Our results revealed that the most highly modulated transcripts could almost entirely be categorized as related to the cell cycle or transcriptional regulation. The most statistically significant enriched categories and networks identified by IPA were associated with cell cycle, gene expression, immune response, infection mechanisms, cellular growth, proliferation and antigen presentation. Canonical pathways involved in energy and cell cycle regulation, and in the co-activation of T cells were also enriched. Conclusions Taken together, these results document a distinct transcriptional profile invoked by the HIV-1/V3 epitope. These data could be invaluable to determine the underlying mechanism by which HIV-1 epitopes interfere with uninfected CD4+ T-cell function causing hyper proliferation and AICD.

  17. Drift of the HIV-1 envelope glycoprotein gp120 toward increased neutralization resistance over the course of the epidemic: a comprehensive study using the most potent and broadly neutralizing monoclonal antibodies.

    Science.gov (United States)

    Bouvin-Pley, M; Morgand, M; Meyer, L; Goujard, C; Moreau, A; Mouquet, H; Nussenzweig, M; Pace, C; Ho, D; Bjorkman, P J; Baty, D; Chames, P; Pancera, M; Kwong, P D; Poignard, P; Barin, F; Braibant, M

    2014-12-01

    Extending our previous analyses to the most recently described monoclonal broadly neutralizing antibodies (bNAbs), we confirmed a drift of HIV-1 clade B variants over 2 decades toward higher resistance to bNAbs targeting almost all the identified gp120-neutralizing epitopes. In contrast, the sensitivity to bNAbs targeting the gp41 membrane-proximal external region remained stable, suggesting a selective pressure on gp120 preferentially. Despite this evolution, selected combinations of bNAbs remain capable of neutralizing efficiently most of the circulating variants. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  18. Characterization of a Large Panel of Rabbit Monoclonal Antibodies against HIV-1 gp120 and Isolation of Novel Neutralizing Antibodies against the V3 Loop.

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    Yali Qin

    Full Text Available We recently reported the induction of potent, cross-clade neutralizing antibodies (nAbs against Human Immunodeficiency Virus type-1 (HIV-1 in rabbits using gp120 based on an M-group consensus sequence. To better characterize these antibodies, 93 hybridomas were generated, which represent the largest panel of monoclonal antibodies (mAbs ever generated from a vaccinated rabbit. The single most frequently recognized epitope of the isolated mAbs was at the very C-terminal end of the protein (APTKAKRRVVEREKR, followed by the V3 loop. A total of seven anti-V3 loop mAbs were isolated, two of which (10A3 and 10A37 exhibited neutralizing activity. In contrast to 10A3 and most other anti-V3 loop nAbs, 10A37 was atypical with its epitope positioned more towards the C-terminal half of the loop. To our knowledge, 10A37 is the most potent and broadly neutralizing anti-V3 loop mAb induced by vaccination. Interestingly, all seven anti-V3 loop mAbs competed with PGT121, suggesting a possibility that early induction of potent anti-V3 loop antibodies could prevent induction of more broadly neutralizing PGT121-like antibodies that target the conserved base of the V3 loop stem.

  19. Prolonged expression of an anti-HIV-1 gp120 minibody to the female rhesus macaque lower genital tract by AAV gene transfer.

    Science.gov (United States)

    Abdel-Motal, U M; Harbison, C; Han, T; Pudney, J; Anderson, D J; Zhu, Q; Westmoreland, S; Marasco, W A

    2014-09-01

    Topical microbicides are a leading strategy for prevention of HIV mucosal infection to women; however, numerous pharmacokinetic limitations associated with coitally related dosing strategy have contributed to their limited success. Here we test the hypothesis that adeno-associated virus (AAV) mediated delivery of the b12 human anti-HIV-1 gp120 minibody gene to the lower genital tract of female rhesus macaques (Rh) can provide prolonged expression of b12 minibodies in the cervical-vaginal secretions. Gene transfer studies demonstrated that, of various green fluorescent protein (GFP)-expressing AAV serotypes, AAV-6 most efficiently transduced freshly immortalized and primary genital epithelial cells (PGECs) of female Rh in vitro. In addition, AAV-6-b12 minibody transduction of Rh PGECs led to inhibition of SHIV162p4 transmigration and virus infectivity in vitro. AAV-6-GFP could also successfully transduce vaginal epithelial cells of Rh when applied intravaginally, including p63+ epithelial stem cells. Moreover, intravaginal application of AAV-6-b12 to female Rh resulted in prolonged minibody detection in their vaginal secretions throughout the 79-day study period. These data provide proof of principle that AAV-6-mediated delivery of anti-HIV broadly neutralizing antibody (BnAb) genes to the lower genital tract of female Rh results in persistent minibody detection for several months. This strategy offers promise that an anti-HIV-1 genetic microbicide strategy may be possible in which topical application of AAV vector, with periodic reapplication as needed, may provide sustained local BnAb expression and protection.

  20. Antibodies raised to short synthetic peptides with sequences derived from HIV-1 SF2 gp120 can both neutralize and enhance HIV-1 SF13: a later variant isolated from the same host.

    Science.gov (United States)

    Davis, D; Trischmann, H; Stephens, D M; Lachmann, P J

    2001-07-01

    HIV-1 SF13 emerged in a patient with immunity to HIV-1 SF2. This study determined the effect of antibodies raised to HIV-1 SF2 on the replication of the later variant. Antisera in rats were raised previously to a complete set of overlapping, synthetic 15mer peptides following the sequence of HIV-1 SF2 gp120. These sera have now been used in neutralization and enhancement assays against viruses derived from molecular clones of both variants. The sets of peptides inducing neutralizing antibodies to the two variants overlap. Antibodies to the third variable region of HIV-1 SF2 only neutralize the homologous virus whereas those to the second and fourth variable regions neutralize both variants. In contrast, the sets of major epitopes involved in enhancement do not overlap. Epitopes for both variants form two clusters when superimposed on the conformation of the conserved regions. To determine if antibodies with the potential to enhance or neutralize HIV-1 SF2 change over time in infected individuals sera from chimpanzees were used because no material was still available from the original patient. Antibodies to HIV-1 SF2 neutralizing epitopes and HIV-1 SF13 enhancing epitopes were present in the circulation of chimpanzees infected with HIV-1 SF2. Once antibodies to the neutralizing epitopes were induced they persisted whereas antibodies to the enhancing epitopes varied with time after infection. Conditions may therefore exist within individual hosts where not only neutralizing but also enhancing antibodies have the potential to contribute to the selection pressure operating on the circulating population of polymorphic variants. Copyright 2001 Wiley-Liss, Inc.

  1. Methamphetamine Exposure Combined with HIV-1 Disease or gp120 Expression: Comparison of Learning and Executive Functions in Humans and Mice

    Science.gov (United States)

    Kesby, James P; Heaton, Robert K; Young, Jared W; Umlauf, Anya; Woods, Steven P; Letendre, Scott L; Markou, Athina; Grant, Igor; Semenova, Svetlana

    2015-01-01

    Methamphetamine dependence is a common comorbid condition among people living with HIV, and may exacerbate HIV-associated neurocognitive disorders. Animal models of neuroAIDS suggest that the gp120 protein may also cause cognitive impairment. The present work evaluated the separate and combined effects of HIV/gp120 and methamphetamine on learning and executive functions in both humans and transgenic mice. Human participants were grouped by HIV serostatus (HIV+ or HIV−) and lifetime methamphetamine dependence (METH+ or METH−). A neurocognitive test battery included domain-specific assessments of learning and executive functions. Mice (gp120+ and gp120−) were exposed to either a methamphetamine binge (METH+) or saline (METH−), then tested in the attentional-set-shifting task to assess learning and executive functions. In humans, HIV status was associated with significant impairments in learning, but less so for executive functions. The frequency of learning impairments varied between groups, with the greatest impairment observed in the HIV+/METH+ group. In mice, gp120 expression was associated with impairments in learning but not reversal learning (executive component). The greatest proportion of mice that failed to complete the task was observed in the gp120+/METH+ group, suggesting greater learning impairments. Our cross-species study demonstrated that HIV in humans and gp120 in mice impaired learning, and that a history of methamphetamine exposure increased the susceptibility to HIV-associated neurocognitive deficits in both species. Finally, the similar pattern of results in both species suggest that the gp120 protein may contribute to HIV-associated learning deficits in humans. PMID:25652249

  2. HIV-1 specific IgA detected in vaginal secretions of HIV uninfected women participating in a microbicide trial in Southern Africa are primarily directed toward gp120 and gp140 specificities.

    Directory of Open Access Journals (Sweden)

    Kelly E Seaton

    Full Text Available Many participants in microbicide trials remain uninfected despite ongoing exposure to HIV-1. Determining the emergence and nature of mucosal HIV-specific immune responses in such women is important, since these responses may contribute to protection and could provide insight for the rational design of HIV-1 vaccines.We first conducted a pilot study to compare three sampling devices (Dacron swabs, flocked nylon swabs and Merocel sponges for detection of HIV-1-specific IgG and IgA antibodies in vaginal secretions. IgG antibodies from HIV-1-positive women reacted broadly across the full panel of eight HIV-1 envelope (Env antigens tested, whereas IgA antibodies only reacted to the gp41 subunit. No Env-reactive antibodies were detected in the HIV-negative women. The three sampling devices yielded equal HIV-1-specific antibody titers, as well as total IgG and IgA concentrations. We then tested vaginal Dacron swabs archived from 57 HIV seronegative women who participated in a microbicide efficacy trial in Southern Africa (HPTN 035. We detected vaginal IgA antibodies directed at HIV-1 Env gp120/gp140 in six of these women, and at gp41 in another three women, but did not detect Env-specific IgG antibodies in any women.Vaginal secretions of HIV-1 infected women contained IgG reactivity to a broad range of Env antigens and IgA reactivity to gp41. In contrast, Env-binding antibodies in the vaginal secretions of HIV-1 uninfected women participating in the microbicide trial were restricted to the IgA subtype and were mostly directed at HIV-1 gp120/gp140.

  3. Development of dengue virus replicons expressing HIV-1 gp120 and other heterologous genes: a potential future tool for dual vaccination against dengue virus and HIV

    Directory of Open Access Journals (Sweden)

    Dayton Andrew I

    2001-11-01

    Full Text Available Abstract Background Toward the goals of providing an additional vector to add to the armamentarium available to HIV vaccinologists and of creating a bivalent vaccine effective against dengue virus and HIV, we have attempted to create vectors which express dengue virus non-structural proteins and HIV immunogens. Previously we reported the successful construction of dengue virus replicons which lack structural genes necessary for virion release and spreading infection in culture but which can replicate intracellularly and abundantly produce dengue non-structural proteins. Here we attempted to express heterologous genetic material from these replicons. Results We cloned into a Δpre-M/E dengue virus replicon genes for either green fluorescent protein (GFP, HIV gp160 or HIV gp120 and tested the ability of these constructs to express dengue virus proteins as well as the heterologous proteins in tissue culture after transfection of replicon RNA. Conclusions Heterologous proteins were readily expressed from these constructs. GFP and gp120 demonstrated minimal or no toxicity. Gp160 expressing replicons were found to express proteins abundantly at 36 hours post transfection, but after 50 hrs of transfection, few replicon positive cells could be found despite the presence of cellular debris positive for replicon proteins. This suggested that gp160 expressed from dengue virus replicons is considerably more toxic than either GFP or gp120. The successful expression of heterologous proteins, including HIV gp120 for long periods in culture suggests this vector system may be useful as a vaccine vector, given appropriate delivery methods.

  4. A Cinnamon-Derived Procyanidin Compound Displays Anti-HIV-1 Activity by Blocking Heparan Sulfate- and Co-Receptor- Binding Sites on gp120 and Reverses T Cell Exhaustion via Impeding Tim-3 and PD-1 Upregulation.

    Directory of Open Access Journals (Sweden)

    Bridgette Janine Connell

    Full Text Available Amongst the many strategies aiming at inhibiting HIV-1 infection, blocking viral entry has been recently recognized as a very promising approach. Using diverse in vitro models and a broad range of HIV-1 primary patient isolates, we report here that IND02, a type A procyanidin polyphenol extracted from cinnamon, that features trimeric and pentameric forms displays an anti-HIV-1 activity against CXCR4 and CCR5 viruses with 1-7 μM ED50 for the trimer. Competition experiments, using a surface plasmon resonance-based binding assay, revealed that IND02 inhibited envelope binding to CD4 and heparan sulphate (HS as well as to an antibody (mAb 17b directed against the gp120 co-receptor binding site with an IC50 in the low μM range. IND02 has thus the remarkable property of simultaneously blocking gp120 binding to its major host cell surface counterparts. Additionally, the IND02-trimer impeded up-regulation of the inhibitory receptors Tim-3 and PD-1 on CD4+ and CD8+ cells, thereby demonstrating its beneficial effect by limiting T cell exhaustion. Among naturally derived products significantly inhibiting HIV-1, the IND02-trimer is the first component demonstrating an entry inhibition property through binding to the viral envelope glycoprotein. These data suggest that cinnamon, a widely consumed spice, could represent a novel and promising candidate for a cost-effective, natural entry inhibitor for HIV-1 which can also down-modulate T cell exhaustion markers Tim-3 and PD-1.

  5. Improved humoral and cellular immune responses against the gp120 V3 loop of HIV-1 following genetic immunization with a chimeric DNA vaccine encoding the V3 inserted into the hepatitis B surface antigen

    DEFF Research Database (Denmark)

    Fomsgaard, A; Nielsen, H V; Bryder, K

    1998-01-01

    -2d-restricted cytotoxic T lymphocyte (CTL) epitope. In an attempt to improve the immunogenicity of V3 in DNA vaccines, a plasmid expressing MN V3 as a fusion protein with the highly immunogenic middle (pre-S2 + S) surface antigen of hepatitis B virus (HBsAg) was constructed. Epidermal inoculation......The gp120-derived V3 loop of HIV-1 is involved in co-receptor interaction, it guides cell tropism, and contains an epitope for antibody neutralization. Thus, HIV-1 V3 is an attractive vaccine candidate. The V3 of the MN strain (MN V3) contains both B- and T-cell epitopes, including a known mouse H...... by gene gun was used for genetic immunization in a mouse model. Antibody and CTL responses to MN V3 and HBsAg were measured and compared with the immune responses obtained after vaccination with plasmids encoding the complete HIV-1 MN gp160 and HBsAg (pre-S2 + S), respectively. DNA vaccination...

  6. Improved humoral and cellular immune response against the gp120 V3 loop of HIV-1 following genetic immunization with a chimeric DNA vaccine encoding the V3 inserted into the hepatites B surface antigen

    DEFF Research Database (Denmark)

    Fomsgaard, A.; Nielsen, H.V.; Bryder, K.

    1998-01-01

    -2d-restricted cytotoxic T lymphocyte (CTL) epitope. In an attempt to improve the immunogenicity of V3 in DNA vaccines, a plasmid expressing MN V3 as a fusion protein with the highly immunogenic middle (pre-S2+S) surface antigen of hepatitis B virus (HBsAg) was constructed. Epidermal inoculation......The gp120-derived V3 loop of HIV-1 is involved in co-receptor interaction, it guides cell tropism, and contains an epitope for antibody neutralization. Thus, HIV-1 V3 is an attractive vaccine candidate. The V3 of the MN strain (MN V3) contains both B- and T-cell epitopes, including a known mouse H...... by gene gun was used for genetic immunization in a mouse model. Antibody and CTL responses to MN V3 and HBsAg were measured and compared with the immune responses obtained after vaccination with plasmids encoding the complete HIV-1 MN gp160 and HBsAg (pre-S2+S), respectively. DNA vaccination with the HIV...

  7. Neutralization of several adult and paediatric HIV-1 subtype C isolates using a shortened synthetic derivative of gp120 binding aptamer called UCLA1.

    CSIR Research Space (South Africa)

    Mufhandu, Hazel T

    2009-07-01

    Full Text Available This paper present a chemically synthesised derivative of the B40 parental aptamer, called UCLA1 (Cohen et al., 2008), was used for neutralization of endemic subtype C clinical isolates of HIV-1 from adult and paediatric patients and subtype B lab...

  8. HIV-1 subtype C unproductively infects human cardiomyocytes in vitro and induces apoptosis mitigated by an anti-Gp120 aptamer

    CSIR Research Space (South Africa)

    Rangel Lopes de Campos, W

    2014-10-01

    Full Text Available the direct and indirect effects of HIV-1 subtype C infection of cultured human cardiomyocytes and the mechanisms leading to cardiomyocytes damage; as well as a way to mitigate the damage. We evaluated a novel approach to mitigate HIVCM using a previously...

  9. Vaccine-induced Human Antibodies Specific for the Third Variable Region of HIV-1 gp120 Impose Immune Pressure on Infecting Viruses

    Directory of Open Access Journals (Sweden)

    Susan Zolla-Pazner

    2014-11-01

    Full Text Available To evaluate the role of V3-specific IgG antibodies (Abs in the RV144 clinical HIV vaccine trial, which reduced HIV-1 infection by 31.2%, the anti-V3 Ab response was assessed. Vaccinees' V3 Abs were highly cross-reactive with cyclic V3 peptides (cV3s from diverse virus subtypes. Sieve analysis of CRF01_AE breakthrough viruses from 43 vaccine- and 66 placebo-recipients demonstrated an estimated vaccine efficacy of 85% against viruses with amino acids mismatching the vaccine at V3 site 317 (p = 0.004 and 52% against viruses matching the vaccine at V3 site 307 (p = 0.004. This analysis was supported by data showing that vaccinees' plasma Abs were less reactive with I307 when replaced with residues found more often in vaccinees' breakthrough viruses. Simultaneously, viruses with mutations at F317 were less infectious, possibly due to the contribution of F317 to optimal formation of the V3 hydrophobic core. These data suggest that RV144-induced V3-specific Abs imposed immune pressure on infecting viruses and inform efforts to design an HIV vaccine.

  10. The convertases furin and PC1 can both cleave the human immunodeficiency virus (HIV)-1 envelope glycoprotein gp160 into gp120 (HIV-1 SU) and gp41 (HIV-I TM).

    Science.gov (United States)

    Decroly, E; Vandenbranden, M; Ruysschaert, J M; Cogniaux, J; Jacob, G S; Howard, S C; Marshall, G; Kompelli, A; Basak, A; Jean, F

    1994-04-22

    Intracellular proteolytic processing of human immunodeficiency virus envelope glycoprotein precursor (gp160) is an essential step for virus infectivity. Northern blot analysis provided evidence that furin and PC1, but not PC2, are expressed in the CD4+ human lymphoblastoid H9 cell line, suggesting the possible participation of these convertases in human immunodeficiency virus (HIV) gp160 proteolytic processing. Purified PC1 and furin cleaved specifically in vitro gp160 into gp120 (HIV-I SU) and gp41 (HIV-I TM). NH2-terminal sequence analysis of the produced gp41 (HIV-I TM) demonstrated that the cleavage occurred within the sequence Arg-Glu-Lys-Arg decreases Ala-Val-Gly-Ile, which is identical to the bond cleaved in vivo. Transition state analog peptides were designed and tested in vitro for their ability to inhibit the PC1- or furin-mediated gp160 cleavage. The best inhibitor was decanoyl-Arg-Lys-Arg-Arg-psi [CH2NH]-Phe-Leu-Gly-Phe-NH2.

  11. HIV Subtypes B and C gp120 and Methamphetamine Interaction: Dopaminergic System Implicates Differential Neuronal Toxicity

    Science.gov (United States)

    Samikkannu, Thangavel; Rao, Kurapati V. K.; Salam, Abdul Ajees Abdul; Atluri, Venkata S. R.; Kaftanovskaya, Elena M.; Agudelo, Marisela; Perez, Suray; Yoo, Changwon; Raymond, Andrea D.; Ding, Hong; Nair, Madhavan P. N.

    2015-01-01

    HIV subtypes or clades differentially induce HIV-associated neurocognitive disorders (HAND) and substance abuse is known to accelerate HIV disease progression. The HIV-1 envelope protein gp120 plays a major role in binding and budding in the central nervous system (CNS) and impacts dopaminergic functions. However, the mechanisms utilized by HIV-1 clades to exert differential effects and the methamphetamine (METH)-associated dopaminergic dysfunction are poorly understood. We hypothesized that clade B and C gp120 structural sequences, modeling based analysis, dopaminergic effect, and METH potentiate neuronal toxicity in astrocytes. We evaluated the effect of clade B and C gp120 and/or METH on the DRD-2, DAT, CaMKs and CREBP transcription. Both the structural sequence and modeling studies demonstrated that clade B gp120 in V1-V4, α -2 and N-glycosylated sites are distinct from clade C gp120. The distinct structure and sequence variation of clade B gp120 differentially impact DRD-2, DAT, CaMK II and CaMK IV mRNA, protein and intracellular expression compared to clade C gp120. However, CREB transcription is upregulated by both clade B and C gp120, and METH co-treatment potentiated these effects. In conclusion, distinct structural sequences of HIV-1 clade B and C gp120 differentially regulate the dopaminergic pathway and METH potentiates neurotoxicity. PMID:26057350

  12. Characterization of the Outer Domain of the gp120 Glycoprotein from Human Immunodeficiency Virus Type 1

    Science.gov (United States)

    Yang, Xinzhen; Tomov, Vesko; Kurteva, Svetla; Wang, Liping; Ren, Xinping; Gorny, Miroslaw K.; Zolla-Pazner, Susan; Sodroski, Joseph

    2004-01-01

    The core of the gp120 glycoprotein from human immunodeficiency virus type 1 (HIV-1) is comprised of three major structural domains: the outer domain, the inner domain, and the bridging sheet. The outer domain is exposed on the HIV-1 envelope glycoprotein trimer and contains binding surfaces for neutralizing antibodies such as 2G12, immunoglobulin G1b12, and anti-V3 antibodies. We expressed the outer domain of HIV-1YU2 gp120 as an independent protein, termed OD1. OD1 efficiently bound 2G12 and a large number of anti-V3 antibodies, indicating its structural integrity. Immunochemical studies with OD1 indicated that antibody responses against the outer domain of the HIV-1 gp120 envelope glycoprotein are rare in HIV-1-infected human sera that potently neutralize the virus. Surprisingly, such outer-domain-directed antibody responses are commonly elicited by immunization with recombinant monomeric gp120. Immunization with soluble, stabilized HIV-1 envelope glycoprotein trimers elicited antibody responses that more closely resembled those in the sera of HIV-1-infected individuals. These results underscore the qualitatively different humoral immune responses elicited during natural infection and after gp120 vaccination and help to explain the failure of gp120 as an effective vaccine. PMID:15542649

  13. Conserved molecular signatures in gp120 are associated with the genetic bottleneck during simian immunodeficiency virus (SIV), SIV-human immunodeficiency virus (SHIV), and HIV type 1 (HIV-1) transmission.

    Science.gov (United States)

    Gonzalez, Mileidy W; DeVico, Anthony L; Lewis, George K; Spouge, John L

    2015-04-01

    Human immunodeficiency virus (HIV) transmission typically results from infection by a single transmitted/founder (T/F) variant. Are T/F variants chosen uniformly at random from the donor pool, or are they selected based on advantageous traits facilitating transmission? Finding evidence for selection during transmission is of particular interest, because it would indicate that phenotypic and/or genetic properties of the viruses might be harnessed as potential vaccine targets or immunotherapies. Here, we systematically evaluated the differences between the Env proteins of simian immunodeficiency virus/simian HIV (SIV/SHIV) stock and T/F variants in search of "signature" sites of transmission. We also surveyed residue preferences in HIV at the SIV/SHIV signature sites. Four sites of gp120 showed significant selection, and an additional two sites showed a similar trend. Therefore, the six sites clearly differentiate T/F viruses from the majority of circulating variants in the stocks. The selection of SIV/SHIV could be inferred reasonably across both vaccinated and unvaccinated subjects, with infections resulting from vaginal, rectal, and intravenous routes of transmission and regardless of viral dosage. The evidence for selection in SIV and SHIV T/F variants is strong and plentiful, and in HIV the evidence is suggestive though commensurate with the availability of suitable data for analysis. Two of the signature residues are completely conserved across the SIV, SHIV, and HIV variants we examined. Five of the signature residues map to the C1 region of gp120 and one to the signal peptide. Our data raise the possibility that C1, while governing the association between gp120 and gp41, modulates transmission efficiency, replicative fitness, and/or host cell tropism at the level of virus-cell attachment and entry. The present study finds significant evidence of selection on gp120 molecules of SIV/SHIV T/F viruses. The data provide ancillary evidence suggesting the same sites

  14. Internalization and Axonal Transport of the HIV Glycoprotein gp120

    Science.gov (United States)

    Berth, Sarah; Caicedo, Hector Hugo; Sarma, Tulika; Morfini, Gerardo

    2015-01-01

    The HIV glycoprotein gp120, a neurotoxic HIV glycoprotein that is overproduced and shed by HIV-infected macrophages, is associated with neurological complications of HIV such as distal sensory polyneuropathy, but interactions of gp120 in the peripheral nervous system remain to be characterized. Here, we demonstrate internalization of extracellular gp120 in a manner partially independent of binding to its coreceptor CXCR4 by F11 neuroblastoma cells and cultured dorsal root ganglion neurons. Immunocytochemical and pharmacological experiments indicate that gp120 does not undergo trafficking through the endolysosomal pathway. Instead, gp120 is mainly internalized through lipid rafts in a cholesterol-dependent manner, with a minor fraction being internalized by fluid phase pinocytosis. Experiments using compartmentalized microfluidic chambers further indicate that, after internalization, endocytosed gp120 selectively undergoes retrograde but not anterograde axonal transport from axons to neuronal cell bodies. Collectively, these studies illuminate mechanisms of gp120 internalization and axonal transport in peripheral nervous system neurons, providing a novel framework for mechanisms for gp120 neurotoxicity. PMID:25636314

  15. CXCR4-gp120-IIIB interactions induce caspase-mediated apoptosis of prostate cancer cells and inhibit tumor growth.

    Science.gov (United States)

    Singh, Shailesh; Bond, Vincent C; Powell, Michael; Singh, Udai P; Bumpers, Harvey L; Grizzle, William E; Lillard, James W

    2009-01-01

    CXC chemokine receptor 4 (CXCR4) has been implicated in prostate cancer metastasis and this receptor also acts as a coreceptor for HIV-1 120-kDa glycoprotein variant IIIB (gp120-IIIB). The interaction between CXCR4 and gp120-IIIB has been shown to mediate apoptosis of both immune and endothelial cells. In this study, we have examined the effects of gp120-IIIB on hormone-refractory prostate cancer cells (PC3 and DU145) in vitro and tumor growth in vivo. Normal prostatic epithelial (PrEC) and prostate cancer cell lines were treated with gp120-IIIB with or without anti-CXCR4 antibody. Caspase expression was evaluated by real-time PCR and active caspase assays. Apoptosis was determined by flow cytometry. gp120-IIIB treatment correlated with active caspase-3 and -9 expression and apoptosis of prostate cancer cells but not PrEC cells. This effect was significantly inhibited after CXCR4 blockade. PC3 and DU145 tumor-bearing mice received intraperitoneal injections of gp120-IIIB and controls received bovine serum albumin in PBS. PC3 and DU145 tumor sizes were measured over time and excised tumors were evaluated for CD44, CD34, lymphatic endothelial cell marker LYVE-1, active caspase-3, and active caspase-9 expression by immunohistochemistry. The tumor size in mice receiving gp120-IIIB was significantly smaller than compared with tumors in control mice. This regression was associated with significant decreases in CD44, CD34, and LYVE-1 and increases in active caspase-3 and -9 expression. These results suggest that gp120-IIIB induced apoptosis in prostate cancer cells and reduced tumor-associated lymphoendothelial cells.

  16. Human immunodeficiency virus type 1 gp120 induces abnormal maturation and functional alterations of dendritic cells: a novel mechanism for AIDS pathogenesis.

    Science.gov (United States)

    Fantuzzi, Laura; Purificato, Cristina; Donato, Karim; Belardelli, Filippo; Gessani, Sandra

    2004-09-01

    Dendritic cells (DCs) play a crucial role in bridging innate and acquired immune responses to pathogens. In human immunodeficiency virus type 1 (HIV-1) infection, immature DCs (iDCs) are also main targets for HIV-1 at the mucosal level. In this study, we evaluated the effects of HIV-1-DC interactions on the maturation and functional activity of these cells. Exposure of human monocyte-derived iDCs to either aldrithiol-2-inactivated HIV-1 or gp120 led to an upmodulation of activation markers indicative of functional maturation. Despite their phenotype, these cells retained antigen uptake capacity and showed an impaired ability to secrete cytokines or chemokines and to induce T-cell proliferation. Although gp120 did not interfere with DC differentiation, the capacity of these cells to produce interleukin-12 (IL-12) upon maturation was markedly reduced. Likewise, iDCs stimulated by classical maturation factors in the presence of gp120 lacked allostimulatory capacity and did not produce IL-12, in spite of their phenotype typical of activated DCs. Exogenous addition of IL-12 restores the allostimulatory capacity of gp120-exposed DCs. The finding that gp120 induces abnormal maturation of DCs linked to profound suppression of their activities unravels a novel mechanism by which HIV can lead to immune dysfunction in AIDS patients. Copyright 2004 American Society for Microbiology

  17. UCLA1, a synthetic derivative of a gp120 RNA aptamer, inhibits entry of human immunodeficiency virus type 1 subtype C

    CSIR Research Space (South Africa)

    Mufhandu, Hazel T

    2012-05-01

    Full Text Available Entry of human immunodeficiency virus type 1 (HIV-1) into cells is mediated by the virion surface envelope (Env) glycoproteins, making it a desirable target for antiretroviral entry inhibitors. We previously isolated a family of gp120 binding RNA...

  18. Occluding the Mannose Moieties on Human Immunodeficiency Virus Type 1 gp120 with Griffithsin Improves the Antibody Responses to Both Proteins in Mice

    NARCIS (Netherlands)

    Banerjee, Kaustuv; Michael, Elizabeth; Eggink, Dirk; van Montfort, Thijs; Lasnik, Amanda B.; Palmer, Kenneth E.; Sanders, Rogier W.; Moore, John P.; Klasse, Per Johan

    2012-01-01

    To assess the influence of mannosylated glycans on the immunogenicity of human immunodeficiency virus type 1 (HIV-1) Env proteins, we immunized mice with monomeric gp120 in the presence and absence of the mannose-binding protein, griffithsin (GRFT). For comparison, other groups of mice received the

  19. C-type lectin Mermaid inhibits dendritic cell mediated HIV-1 transmission to CD4+ T cells

    NARCIS (Netherlands)

    Nabatov, Alexey A.; de Jong, Marein A. W. P.; de Witte, Lot; Bulgheresi, Silvia; Geijtenbeek, Teunis B. H.

    2008-01-01

    Dendritic cells (DCs) are important in HIV-1 transmission; DCs capture invading HIV-1 through the interaction of the gp120 oligosaccharides with the C-type lectin DC-SIGN and migrate to the lymphoid tissues where HIV-1 is transmitted to T cells. Thus, the HIV-1 envelope glycoprotein gp120 is an

  20. A glycoconjugate antigen based on the recognition motif of a broadly neutralizing human immunodeficiency virus antibody, 2G12, is immunogenic but elicits antibodies unable to bind to the self glycans of gp120

    DEFF Research Database (Denmark)

    Astronomo, Rena D; Lee, Hing-Ken; Scanlan, Christopher N

    2008-01-01

    The glycan shield of human immunodeficiency virus type 1 (HIV-1) gp120 contributes to viral evasion from humoral immune responses. However, the shield is recognized by the HIV-1 broadly neutralizing antibody (Ab), 2G12, at a relatively conserved cluster of oligomannose glycans. The discovery of 2G...... serum Ab titers to Man(4). However, these Abs are unable to bind gp120. Further analysis reveals that the elicited Abs bind a variety of unbranched and, to a lesser extent, branched Man(9) derivatives but not natural N-linked oligomannose containing the chitobiose core. These results suggest that Abs...

  1. Induction of Potent CD8+ T-Cell Responses by Novel Biodegradable Nanoparticles Carrying Human Immunodeficiency Virus Type 1 gp120

    Science.gov (United States)

    Wang, Xin; Uto, Tomofumi; Akagi, Takami; Akashi, Mitsuru; Baba, Masanori

    2007-01-01

    The mainstream of recent anti-AIDS vaccines is a prime/boost approach with multiple doses of the target DNA of human immunodeficiency virus type 1 (HIV-1) and recombinant viral vectors. In this study, we have attempted to construct an efficient protein-based vaccine using biodegradable poly(γ-glutamic acid) (γ-PGA) nanoparticles (NPs), which are capable of inducing potent cellular immunity. A significant expansion of CD8+ T cells specific to the major histocompatibility complex class I-restricted gp120 epitope was observed in mice intranasally immunized once with gp120-carrying NPs but not with gp120 alone or gp120 together with the B-subunit of cholera toxin. Both the gp120-encapsulating and -immobilizing forms of NPs could induce antigen-specific spleen CD8+ T cells having a functional profile of cytotoxic T lymphocytes. Long-lived memory CD8+ T cells could also be elicited. Although a substantial decay in the effector memory T cells was observed over time in the immunized mice, the central memory T cells remained relatively constant from day 30 to day 238 after immunization. Furthermore, the memory CD8+ T cells rapidly expanded with boosting with the same immunogen. In addition, γ-PGA NPs were found to be a much stronger inducer of antigen-specific CD8+ T-cell responses than nonbiodegradable polystyrene NPs. Thus, γ-PGA NPs carrying various HIV-1 antigens may have great potential as a novel priming and/or boosting tool in current vaccination regimens for the induction of cellular immune responses. PMID:17609261

  2. Induction of potent CD8+ T-cell responses by novel biodegradable nanoparticles carrying human immunodeficiency virus type 1 gp120.

    Science.gov (United States)

    Wang, Xin; Uto, Tomofumi; Akagi, Takami; Akashi, Mitsuru; Baba, Masanori

    2007-09-01

    The mainstream of recent anti-AIDS vaccines is a prime/boost approach with multiple doses of the target DNA of human immunodeficiency virus type 1 (HIV-1) and recombinant viral vectors. In this study, we have attempted to construct an efficient protein-based vaccine using biodegradable poly(gamma-glutamic acid) (gamma-PGA) nanoparticles (NPs), which are capable of inducing potent cellular immunity. A significant expansion of CD8+ T cells specific to the major histocompatibility complex class I-restricted gp120 epitope was observed in mice intranasally immunized once with gp120-carrying NPs but not with gp120 alone or gp120 together with the B-subunit of cholera toxin. Both the gp120-encapsulating and -immobilizing forms of NPs could induce antigen-specific spleen CD8+ T cells having a functional profile of cytotoxic T lymphocytes. Long-lived memory CD8+ T cells could also be elicited. Although a substantial decay in the effector memory T cells was observed over time in the immunized mice, the central memory T cells remained relatively constant from day 30 to day 238 after immunization. Furthermore, the memory CD8+ T cells rapidly expanded with boosting with the same immunogen. In addition, gamma-PGA NPs were found to be a much stronger inducer of antigen-specific CD8+ T-cell responses than nonbiodegradable polystyrene NPs. Thus, gamma-PGA NPs carrying various HIV-1 antigens may have great potential as a novel priming and/or boosting tool in current vaccination regimens for the induction of cellular immune responses.

  3. Antigenic properties of the human immunodeficiency virus envelope glycoprotein gp120 on virions bound to target cells.

    Directory of Open Access Journals (Sweden)

    Meron Mengistu

    2015-03-01

    Full Text Available The HIV-1 envelope glycoprotein, gp120, undergoes multiple molecular interactions and structural rearrangements during the course of host cell attachment and viral entry, which are being increasingly defined at the atomic level using isolated proteins. In comparison, antigenic markers of these dynamic changes are essentially unknown for single HIV-1 particles bound to target cells. Such markers should indicate how neutralizing and/or non-neutralizing antibodies might interdict infection by either blocking infection or sensitizing host cells for elimination by Fc-mediated effector function. Here we address this deficit by imaging fluorescently labeled CCR5-tropic HIV-1 pseudoviruses using confocal and superresolution microscopy to track the exposure of neutralizing and non-neutralizing epitopes as they appear on single HIV-1 particles bound to target cells. Epitope exposure was followed under conditions permissive or non-permissive for viral entry to delimit changes associated with virion binding from those associated with post-attachment events. We find that a previously unexpected array of gp120 epitopes is exposed rapidly upon target cell binding. This array comprises both neutralizing and non-neutralizing epitopes, the latter being hidden on free virions yet capable of serving as potent targets for Fc-mediated effector function. Under non-permissive conditions for viral entry, both neutralizing and non-neutralizing epitope exposures were relatively static over time for the majority of bound virions. Under entry-permissive conditions, epitope exposure patterns changed over time on subsets of virions that exhibited concurrent variations in virion contents. These studies reveal that bound virions are distinguished by a broad array of both neutralizing and non-neutralizing gp120 epitopes that potentially sensitize a freshly engaged target cell for destruction by Fc-mediated effector function and/or for direct neutralization at a post-binding step

  4. Expression of HIV gp120 protein increases sensitivity to the rewarding properties of methamphetamine in mice

    Science.gov (United States)

    Kesby, James P.; Hubbard, David T.; Markou, Athina; Semenova, Svetlana

    2012-01-01

    Methamphetamine abuse and human immunodeficiency virus (HIV) infection induce neuropathological changes in corticolimbic brain areas involved in reward and cognitive function. Little is known about the combined effects of methamphetamine and HIV infection on cognitive and reward processes. The HIV/gp120 protein induces neurodegeneration in mice, similar to HIV-induced pathology in humans. We investigated the effects of gp120 expression on associative learning, preference for methamphetamine and non-drug reinforcers, and sensitivity to the conditioned rewarding properties of methamphetamine in transgenic (tg) mice expressing HIV/gp120 protein (gp120-tg). gp120-tg mice learned the operant response for food at the same rate as non-tg mice. In the two-bottle choice procedure with restricted access to drugs, gp120-tg mice exhibited greater preference for methamphetamine and saccharin than non-tg mice, whereas preference for quinine was similar between genotypes. Under conditions of unrestricted access to methamphetamine, the mice exhibited a decreased preference for increasing methamphetamine concentrations. However, male gp120-tg mice showed a decreased preference for methamphetamine at lower concentrations than non-tg male mice. gp120-tg mice developed methamphetamine-induced conditioned place preference at lower methamphetamine doses compared with non-tg mice. No differences in methamphetamine pharmacokinetics were found between genotypes. These results indicate that gp120-tg mice exhibit no deficits in associative learning or reward/motivational function for a natural reinforcer. Interestingly, gp120 expression resulted in increased preference for methamphetamine and a highly palatable non-drug reinforcer (saccharin) and increased sensitivity to methamphetamine-induced conditioned reward. These data suggest that HIV-positive individuals may have increased sensitivity to methamphetamine, leading to high methamphetamine abuse potential in this population. PMID

  5. HIV gp120 vaccine - VaxGen: AIDSVAX, AIDSVAX B/B, AIDSVAX B/E, HIV gp120 vaccine - Genentech, HIV gp120 vaccine AIDSVAX - VaxGen, HIV vaccine AIDSVAX - VaxGen.

    Science.gov (United States)

    2003-01-01

    VaxGen is developing prophylactic vaccines against HIV-1 consisting of two recombinant gp120 surface proteins from different HIV-1 strains.This profile has been selected from R&D Insight, a pharmaceutical intelligence database produced by Adis International Ltd. The bivalent vaccines [AIDSVAX B/B and AIDSVAX B/E] are being evaluated in two phase III trials. The first multicentre phase III trial of AIDSVAX B/B, was conducted principally in Canada and the US but also at some sites in the Netherlands and Puerto Rico. The trial was completed at the end of 2002. The second phase III trial is being conducted in Thailand with the AIDSVAX B/E vaccine. VaxGen announced in July 2002 that it would be delaying its Biologics License Application (BLA) for AIDSVAX until 2004 to enable the company to fulfil pre-approval manufacturing requirements. AIDSVAX is based on an earlier monovalent gp120 vaccine developed by Genentech that was shown to be safe in humans. VaxGen (formerly Genenvax) was formed as a spin-off company from Genentech with the sole purpose of developing the gp120 vaccine. VaxGen announced in July 2002 that the original License and Supply agreement with Genentech, signed in May 1997, had been amended. Under the revised agreement, Genentech maintains its right to market and sell AIDSVAX in North America, but has relinquished its options to commercialise the vaccine candidate in the rest of the world. Genentech's earlier decision to waive its option to manufacture AIDSVAX has also been formalised in this agreement. Additionally, VaxGen's royalty payments to Genentech for sales to the WHO or UN for underdeveloped nations have also been reduced by up to 50% and Genentech has extended the milestone date associated with VaxGen submitting an NDA. A $US120 million joint venture (Celltrion) has been formed between VaxGen and South Korean investors to manufacture more than 200 million doses of AIDSVAX a year. Celltrion will build and operate two biotechnology manufacturing

  6. Cognitive deficits associated with combined HIV gp120 expression and chronic methamphetamine exposure in mice

    Science.gov (United States)

    Kesby, James P.; Markou, Athina; Semenova, Svetlana

    2014-01-01

    Methamphetamine abuse is common among individuals infected by human immunodeficiency virus (HIV). Neurocognitive outcomes tend to be worse in methamphetamine users with HIV. However, it is unclear whether discrete cognitive domains are susceptible to impairment after combined HIV infection and methamphetamine abuse. The expression of HIV/gp120 protein induces neuropathology in mice similar to HIV-induced pathology in humans. We investigated the separate and combined effects of methamphetamine exposure and gp120 expression on cognitive function in transgenic (gp120-tg) and control mice. The mice underwent an escalating methamphetamine binge regimen and were tested in novel object/location recognition, object-in-place recognition, and Barnes maze tests. gp120 expression disrupted performance in the object-in-place test (i.e., similar time spent with all objects, regardless of location), indicating deficits in associative recognition memory. gp120 expression also altered reversal learning in the Barnes maze, suggesting impairments in executive function. Methamphetamine exposure impaired spatial strategy in the Barnes maze, indicating deficits in spatial learning. Methamphetamine-exposed gp120-tg mice had the lowest spatial strategy scores in the final acquisition trials in the Barnes maze, suggesting greater deficits in spatial learning than all of the other groups. Although HIV infection involves interactions between multiple proteins and processes, in addition to gp120, our findings in gp120-tg mice suggest that humans with the dual insult of HIV infection and methamphetamine abuse may exhibit a broader spectrum of cognitive deficits than those with either factor alone. Depending on the cognitive domain, the combination of both insults may exacerbate deficits in cognitive performance compared with each individual insult. PMID:25476577

  7. Structural plasticity and conformational transitions of HIV envelope glycoprotein gp120.

    Directory of Open Access Journals (Sweden)

    Anil Korkut

    Full Text Available HIV envelope glycoproteins undergo large-scale conformational changes as they interact with cellular receptors to cause the fusion of viral and cellular membranes that permits viral entry to infect targeted cells. Conformational dynamics in HIV gp120 are also important in masking conserved receptor epitopes from being detected for effective neutralization by the human immune system. Crystal structures of HIV gp120 and its complexes with receptors and antibody fragments provide high-resolution pictures of selected conformational states accessible to gp120. Here we describe systematic computational analyses of HIV gp120 plasticity in such complexes with CD4 binding fragments, CD4 mimetic proteins, and various antibody fragments. We used three computational approaches: an isotropic elastic network analysis of conformational plasticity, a full atomic normal mode analysis, and simulation of conformational transitions with our coarse-grained virtual atom molecular mechanics (VAMM potential function. We observe collective sub-domain motions about hinge points that coordinate those motions, correlated local fluctuations at the interfacial cavity formed when gp120 binds to CD4, and concerted changes in structural elements that form at the CD4 interface during large-scale conformational transitions to the CD4-bound state from the deformed states of gp120 in certain antibody complexes.

  8. HIV Glycoprotein Gp120 Impairs Fast Axonal Transport by Activating Tak1 Signaling Pathways

    Directory of Open Access Journals (Sweden)

    Sarah H. Berth

    2016-11-01

    Full Text Available Sensory neuropathies are the most common neurological complication of HIV. Of these, distal sensory polyneuropathy (DSP is directly caused by HIV infection and characterized by length-dependent axonal degeneration of dorsal root ganglion (DRG neurons. Mechanisms for axonal degeneration in DSP remain unclear, but recent experiments revealed that the HIV glycoprotein gp120 is internalized and localized within axons of DRG neurons. Based on these findings, we investigated whether intra-axonal gp120 might impair fast axonal transport (FAT, a cellular process critical for appropriate maintenance of the axonal compartment. Significantly, we found that gp120 severely impaired both anterograde and retrograde FAT. Providing a mechanistic basis for these effects, pharmacological experiments revealed an involvement of various phosphotransferases in this toxic effect, including members of mitogen-activated protein kinase pathways (Tak-1, p38, and c-Jun N-terminal Kinase (JNK, inhibitor of kappa-B-kinase 2 (IKK2, and PP1. Biochemical experiments and axonal outgrowth assays in cell lines and primary cultures extended these findings. Impairments in neurite outgrowth in DRG neurons by gp120 were rescued using a Tak-1 inhibitor, implicating a Tak-1 mitogen-activated protein kinase pathway in gp120 neurotoxicity. Taken together, these observations indicate that kinase-based impairments in FAT represent a novel mechanism underlying gp120 neurotoxicity consistent with the dying-back degeneration seen in DSP. Targeting gp120-based impairments in FAT with specific kinase inhibitors might provide a novel therapeutic strategy to prevent axonal degeneration in DSP.

  9. HIV Glycoprotein Gp120 Impairs Fast Axonal Transport by Activating Tak1 Signaling Pathways.

    Science.gov (United States)

    Berth, Sarah H; Mesnard-Hoaglin, Nichole; Wang, Bin; Kim, Hajwa; Song, Yuyu; Sapar, Maria; Morfini, Gerardo; Brady, Scott T

    2016-12-01

    Sensory neuropathies are the most common neurological complication of HIV. Of these, distal sensory polyneuropathy (DSP) is directly caused by HIV infection and characterized by length-dependent axonal degeneration of dorsal root ganglion (DRG) neurons. Mechanisms for axonal degeneration in DSP remain unclear, but recent experiments revealed that the HIV glycoprotein gp120 is internalized and localized within axons of DRG neurons. Based on these findings, we investigated whether intra-axonal gp120 might impair fast axonal transport (FAT), a cellular process critical for appropriate maintenance of the axonal compartment. Significantly, we found that gp120 severely impaired both anterograde and retrograde FAT. Providing a mechanistic basis for these effects, pharmacological experiments revealed an involvement of various phosphotransferases in this toxic effect, including members of mitogen-activated protein kinase pathways (Tak-1, p38, and c-Jun N-terminal Kinase (JNK)), inhibitor of kappa-B-kinase 2 (IKK2), and PP1. Biochemical experiments and axonal outgrowth assays in cell lines and primary cultures extended these findings. Impairments in neurite outgrowth in DRG neurons by gp120 were rescued using a Tak-1 inhibitor, implicating a Tak-1 mitogen-activated protein kinase pathway in gp120 neurotoxicity. Taken together, these observations indicate that kinase-based impairments in FAT represent a novel mechanism underlying gp120 neurotoxicity consistent with the dying-back degeneration seen in DSP. Targeting gp120-based impairments in FAT with specific kinase inhibitors might provide a novel therapeutic strategy to prevent axonal degeneration in DSP. © The Author(s) 2016.

  10. Serological responses in chimpanzees inoculated with human immunodeficiency virus glycoprotein (gp120) subunit vaccine

    Energy Technology Data Exchange (ETDEWEB)

    Arthur, L.O.; Pyle, S.W.; Nara, P.L.; Bess, J.W. Jr.; Gonda, M.A.; Kelliher, J.C.; Gilden, R.V.; Robey, W.G.; Bolognesi, D.P.; Gallo, R.C.

    1987-12-01

    The major envelope glycoprotein of a human immunodeficiency virus (HIV) has been purified and was utilized as a prototype vaccine in chimpanzees. The 120,000-dalton glycoprotein (gp120) was purified from membranes of human T-lymphotropic virus (HTLV)-IIIB-infected cells and the final preparation contained low levels to no detectable HTLV-IIIB core antigen (p24) and low levels of endotoxin. Chimpanzees inoculated with gp120 responded by developing antibodies that precipitated radiolabeled gp120 and neutralized in vitro infection of HTLV-IIIB. Antibodies to HTLV-IIIB p24 were not detected in the gp120-immunized chimpanzees. Peripheral blood leukocytes from the vaccinated animals were examined for T4/sup +/ and T8/sup +/ cells, and no decrease in the T4/T8 ratio was found, indicating that immunization with a ligand (gp120) that binds to T4 has not detectable adverse effect on the population of T4/sup +/ cells. The only current animal model that can be reproducibly infected with HIV is the chimpanzee. Immunization of chimpanzees with HIV proteins will provide an experimental system for testing the effectiveness of prototype vaccines for preventing HIV infection in vivo.

  11. Characterization of DC-SIGN/R interaction with human immunodeficiency virus type 1 gp120 and ICAM molecules favors the receptor's role as an antigen-capturing rather than an adhesion receptor.

    Science.gov (United States)

    Snyder, Greg A; Ford, Jennifer; Torabi-Parizi, Parizad; Arthos, James A; Schuck, Peter; Colonna, Marco; Sun, Peter D

    2005-04-01

    The dendritic cell (DC)-specific intercellular adhesion molecule 3 (ICAM-3)-grabbing nonintegrin binding receptor (DC-SIGN) was shown to bind human immunodeficiency virus type 1 (HIV-1) viral envelope protein gp120 and proposed to function as a Trojan horse to enhance trans-virus infection to host T cells. To better understand the mechanism by which DC-SIGN and DC-SIGNR selectively bind HIV-1 gp120, we constructed a series of deletion mutations in the repeat regions of both receptors. Different truncated receptors exist in different oligomeric forms. The carbohydrate binding domain without any repeats was monomeric, whereas the full extracellular receptors existed as tetramers. All reconstituted receptors retained their ability to bind gp120. The dissociation constant, however, differed drastically from micromolar values for the monomeric receptors to nanomolar values for the tetrameric receptors, suggesting that the repeat region of these receptors contributes to the avidity of gp120 binding. Such oligomerization may provide a mechanism for the receptor to selectively recognize pathogens containing multiple high-mannose-concentration carbohydrates. In contrast, the receptors bound to ICAMs with submicromolar affinities that are similar to those of two nonspecific cell surface glycoproteins, FcgammaRIIb and FcgammaRIII, and the oligomerization of DC-SIGNR resulted in no increase in binding affinity to ICAM-3. These findings suggest that DC-SIGN may not discriminate other cell surface glycoproteins from ICAM-3 binding. The pH dependence in DC-SIGN binding to gp120 showed that the receptor retained high-affinity gp120 binding at neutral pH but lost gp120 binding at pH 5, suggesting a release mechanism of HIV in the acidic endosomal compartment by DC-SIGN. Our work contradicts the function of DC-SIGN as a Trojan horse to facilitate HIV-1 infection; rather, it supports the function of DC-SIGN/R (a designation referring to both DC-SIGN and DC-SIGNR) as an antigen

  12. Peptide Paratope Mimics of the Broadly Neutralizing HIV-1 Antibody b12.

    Science.gov (United States)

    Haußner, Christina; Damm, Dominik; Nirschl, Sandra; Rohrhofer, Anette; Schmidt, Barbara; Eichler, Jutta

    2017-04-04

    The broadly neutralizing HIV-1 antibody b12 recognizes the CD4 binding site of the HIV-1 envelope glycoprotein gp120 and efficiently neutralizes HIV-1 infections in vitro and in vivo. Based on the 3D structure of a b12⋅gp120 complex, we have designed an assembled peptide (b12-M) that presents the parts of the three heavy-chain complementarity-determining regions (CDRs) of b12, which contain the contact sites of the antibody for gp120. This b12-mimetic peptide, as well as a truncated peptide presenting only two of the three heavy-chain CDRs of b12, were shown to recognize gp120 in a similar manner to b12, as well as to inhibit HIV-1 infection, demonstrating functional mimicry of b12 by the paratope mimetic peptides. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Computational Design of Hypothetical New Peptides Based on a Cyclotide Scaffold as HIV gp120 Inhibitor.

    Directory of Open Access Journals (Sweden)

    Apiwat Sangphukieo

    Full Text Available Cyclotides are a family of triple disulfide cyclic peptides with exceptional resistance to thermal/chemical denaturation and enzymatic degradation. Several cyclotides have been shown to possess anti-HIV activity, including kalata B1 (KB1. However, the use of cyclotides as anti-HIV therapies remains limited due to the high toxicity in normal cells. Therefore, grafting anti-HIV epitopes onto a cyclotide might be a promising approach for reducing toxicity and simultaneously improving anti-HIV activity. Viral envelope glycoprotein gp120 is required for entry of HIV into CD4+ T cells. However, due to a high degree of variability and physical shielding, the design of drugs targeting gp120 remains challenging. We created a computational protocol in which molecular modeling techniques were combined with a genetic algorithm (GA to automate the design of new cyclotides with improved binding to HIV gp120. We found that the group of modified cyclotides has better binding scores (23.1% compared to the KB1. By using molecular dynamic (MD simulation as a post filter for the final candidates, we identified two novel cyclotides, GA763 and GA190, which exhibited better interaction energies (36.6% and 22.8%, respectively when binding to gp120 compared to KB1. This computational design represents an alternative tool for modifying peptides, including cyclotides and other stable peptides, as therapeutic agents before the synthesis process.

  14. DMPD: Macrophage activation through CCR5- and CXCR4-mediated gp120-elicited signalingpathways. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 12960231 Macrophage activation through CCR5- and CXCR4-mediated gp120-elicited sign...82. Epub 2003 Jul 22. (.png) (.svg) (.html) (.csml) Show Macrophage activation through CCR5- and CXCR4-media...on through CCR5- and CXCR4-mediated gp120-elicited signalingpathways. Authors Lee C, Liu QH, Tomkowicz B, Yi

  15. Stoichiometric parameters of HIV-1 entry.

    Science.gov (United States)

    Zarr, Melissa; Siliciano, Robert

    2015-01-01

    During HIV type 1 (HIV-1) entry, trimers of gp120 bind to CD4 and either the CCR5 or CXCR4 coreceptor on the target cell. The stoichiometric parameters associated with HIV-1 entry remain unclear. Important unanswered questions include: how many trimers must attach to CD4 molecules, how many must bind coreceptors, and how many functional gp120 subunits per trimer are required for entry? We performed single round infectivity assays with chimeric viruses and compared the experimental relative infectivity curves with curves generated by mathematical models. Our results indicate that HIV-1 entry requires only a small number of functional spikes (one or two), that Env trimers may function with fewer than three active subunits, and that there is no major difference in the stoichiometric requirements for CCR5 vs. CXCR4 mediated HIV-1 entry into host cells. Copyright © 2014 Elsevier Inc. All rights reserved.

  16. An HIV gp120-CD4 Immunogen Does Not Elicit Autoimmune Antibody Responses in Cynomolgus Macaques.

    Science.gov (United States)

    Schwartz, Jennifer A; Prado, Ilia; Misamore, Johnathan; Weiss, Deborah; Francis, Jesse; Pal, Ranajit; Huaman, Maria; Cristillo, Anthony; Lewis, George K; Gallo, Robert C; DeVico, Anthony L; Fouts, Timothy R

    2016-07-01

    A promising concept for human immunodeficiency virus (HIV) vaccines focuses immunity on the highly conserved transition state structures and epitopes that appear when the HIV glycoprotein gp120 binds to its receptor, CD4. We are developing chimeric antigens (full-length single chain, or FLSC) in which gp120 and CD4 sequences are flexibly linked to allow stable intrachain complex formation between the two moieties (A. DeVico et al., Proc Natl Acad Sci U S A 104:17477-17482, 2007, doi:10.1073/pnas.0707399104; T. R. Fouts et al., J Virol 74:11427-11436, 2000, doi:10.1128/JVI.74.24.11427-11436.2000). Proof of concept studies with nonhuman primates show that FLSC elicited heterologous protection against simian-human immunodeficiency virus (SHIV)/simian immunodeficiency virus (SIV) (T. R. Fouts et al., Proc Natl Acad Sci U S A 112:E992-E999, 2016, doi:10.1073/pnas.1423669112), which correlated with antibodies against transition state gp120 epitopes. Nevertheless, advancement of any vaccine that comprises gp120-CD4 complexes must consider whether the CD4 component breaks tolerance and becomes immunogenic in the autologous host. To address this, we performed an immunotoxicology study with cynomolgus macaques vaccinated with either FLSC or a rhesus variant of FLSC containing macaque CD4 sequences (rhFLSC). Enzyme-linked immunosorbent assay (ELISA) binding titers, primary CD3(+) T cell staining, and temporal trends in T cell subset frequencies served to assess whether anti-CD4 autoantibody responses were elicited by vaccination. We find that immunization with multiple high doses of rhFLSC did not elicit detectable antibody titers despite robust responses to rhFLSC. In accordance with these findings, immunized animals had no changes in circulating CD4(+) T cell counts or evidence of autoantibody reactivity with cell surface CD4 on primary naive macaque T cells. Collectively, these studies show that antigens using CD4 sequences to stabilize transition state gp120 structures

  17. HIV gp120 induces, NF-kappaB dependent, HIV replication that requires procaspase 8.

    Directory of Open Access Journals (Sweden)

    Gary D Bren

    Full Text Available HIV envelope glycoprotein gp120 causes cellular activation resulting in anergy, apoptosis, proinflammatory cytokine production, and through an unknown mechanism, enhanced HIV replication.We describe that the signals which promote apoptosis are also responsible for the enhanced HIV replication. Specifically, we demonstrate that the caspase 8 cleavage fragment Caspase8p43, activates p50/p65 Nuclear Factor kappaB (NF-kappaB, in a manner which is inhibited by dominant negative IkappaBalpha. This caspase 8 dependent NF-kappaB activation occurs following stimulation with gp120, TNF, or CD3/CD28 crosslinking, but these treatments do not activate NF-kappaB in cells deficient in caspase 8. The Casp8p43 cleavage fragment also transactivates the HIV LTR through NF-kappaB, and the absence of caspase 8 following HIV infection greatly inhibits HIV replication.Gp120 induced caspase 8 dependent NF-kappaB activation is a novel pathway of HIV replication which increases understanding of the biology of T-cell death, as well as having implications for understanding treatment and prevention of HIV infection.

  18. Prime-Boost Vaccination Using Chemokine-Fused gp120 DNA and HIV Envelope Peptides Activates Both Immediate and Long-Term Memory Cellular Responses in Rhesus Macaques

    Directory of Open Access Journals (Sweden)

    Hong Qin

    2010-01-01

    Full Text Available HIV vaccine candidates with improved immunogenicity and induction of mucosal T-cell immunity are needed. A prime-boost strategy using a novel HIV glycoprotein 120 DNA vaccine was employed to immunize rhesus macaques. The DNA vaccine encoded a chimeric gp120 protein in fusion with monocyte chemoattractant protein-3, which was hypothesized to improve the ability of antigen-presenting cells to capture viral antigen through chemokine receptor-mediated endocytosis. DNA vaccination induced virus-reactive T cells in peripheral blood, detectable by T cell proliferation, INFγ ELISPOT and sustained IL-6 production, without humoral responses. With a peptide-cocktail vaccine containing a set of conserved polypeptides of HIV-1 envelope protein, given by nasogastric administration, primed T-cell immunity was significantly boosted. Surprisingly, long-term and peptide-specific mucosal memory T-cell immunity was detected in both vaccinated macaques after one year. Therefore, data from this investigation offer proof-of-principle for potential effectiveness of the prime-boost strategy with a chemokine-fused gp120 DNA and warrant further testing in the nonhuman primate models for developing as a potential HIV vaccine candidate in humans.

  19. Structural insights into the specific anti-HIV property of actinohivin: structure of its complex with the α(1–2)mannobiose moiety of gp120

    Energy Technology Data Exchange (ETDEWEB)

    Hoque, M. Mominul [Iwaki Meisei University, 5-5-1 Chuodai-Iino, Iwaki, Fukushima 970-8551 (Japan); Rajshahi University, Rajshahi (Bangladesh); Suzuki, Kaoru [Iwaki Meisei University, Iwaki, Fukushima 970-8551 (Japan); Tsunoda, Masaru; Jiang, Jiandong; Zhang, Fang; Takahashi, Atsushi; Ohbayashi, Naomi; Zhang, Xiaoxue [Iwaki Meisei University, 5-5-1 Chuodai-Iino, Iwaki, Fukushima 970-8551 (Japan); Tanaka, Haruo [Iwaki Meisei University, 5-5-1 Chuodai-Iino, Iwaki, Fukushima 970-8551 (Japan); Iwaki Meisei University, 5-5-1 Chuodai-Iino, Iwaki, Fukushima 970-8551 (Japan); KIIM Pharmaceutical Laboratories Inc., Fukushima 970-8551 (Japan); Ōmura, Satoshi [Kitasato University, Tokyo 108-8641 (Japan); Takénaka, Akio, E-mail: atakenak@sakura.email.ne.jp [Iwaki Meisei University, 5-5-1 Chuodai-Iino, Iwaki, Fukushima 970-8551 (Japan); Iwaki Meisei University, 5-5-1 Chuodai-Iino, Iwaki, Fukushima 970-8551 (Japan); Tokyo Institute of Technology, Yokohama 226-8501 (Japan)

    2012-12-01

    X-ray analysis of anti-HIV actinohivin in complex with the target α(1-2)mannobiose moiety of high-mannose type glycans attached to HIV-1 gp120 reveals that the three rotamers generated with 120 rotations around the molecular pseudo-rotation axis are packed randomly in the unit cell according to the P2{sub 1}2{sub 1}2{sub 1} symmetry to exhibit an apparent space group P2{sub 1}3 as the statistical structure. However, the high-resolution X-ray structure shows the detailed interaction geometry for specific binding. Actinohivin (AH) is an actinomycete lectin with a potent specific anti-HIV activity. In order to clarify the structural evidence for its specific binding to the α(1–2)mannobiose (MB) moiety of the D1 chains of high-mannose-type glycans (HMTGs) attached to HIV-1 gp120, the crystal structure of AH in complex with MB has been determined. The AH molecule is composed of three identical structural modules, each of which has a pocket in which an MB molecule is bound adopting a bracket-shaped conformation. This conformation is stabilized through two weak C—H⋯O hydrogen bonds facilitated by the α(1–2) linkage. The binding features in the three pockets are quite similar to each other, in accordance with the molecular pseudo-threefold symmetry generated from the three tandem repeats in the amino-acid sequence. The shape of the pocket can accept two neighbouring hydroxyl groups of the O{sup 3} and O{sup 4} atoms of the equatorial configuration of the second mannose residue. To recognize these atoms through hydrogen bonds, an Asp residue is located at the bottom of each pocket. Tyr and Leu residues seem to block the movement of the MB molecules. Furthermore, the O{sup 1} atom of the axial configuration of the second mannose residue protrudes from each pocket into an open space surrounded by the conserved hydrophobic residues, suggesting an additional binding site for the third mannose residue of the branched D1 chain of HMTGs. These structural features

  20. Surfactant protein D binds to human immunodeficiency virus (HIV) envelope protein gp120 and inhibits HIV replication

    DEFF Research Database (Denmark)

    Meschi, Joseph; Crouch, Erika C; Skolnik, Paul

    2005-01-01

    The envelope protein (gp120) of human immunodeficiency virus (HIV) contains highly conserved mannosylated oligosaccharides. These glycoconjugates contribute to resistance to antibody neutralization, and binding to cell surface lectins on macrophages and dendritic cells. Mannose-binding lectin (MBL......) binds to gp120 and plays a role in defence against the virus. In this study it is demonstrated that surfactant protein D (SP-D) binds to gp120 and inhibits HIV infectivity at significantly lower concentrations than MBL. The binding of SP-D was mediated by its calcium-dependent carbohydrate...... defence against HIV. A chimeric protein containing the N-terminal and collagen domains of SP-D linked to the neck and carbohydrate-recognition domains of MBL (called SP-D/MBL(neck+CRD)) had greater ability to bind to gp120 and inhibit virus replication than either SP-D or MBL. The enhanced binding of SP...

  1. N-acetylcysteine prevents HIV gp 120-related damage of human cultured astrocytes: correlation with glutamine synthase dysfunction

    Directory of Open Access Journals (Sweden)

    Costa Nicola

    2007-12-01

    Full Text Available Abstract Background HIV envelope gp 120 glycoprotein is released during active HIV infection of brain macrophages thereby generating inflammation and oxidative stress which contribute to the development of the AIDS-Dementia Complex (ADC. Gp120 has also been found capable to generate excitotoxic effect on brain tissue via enhancement of glutamatergic neurotransmission, leading to neuronal and astroglial damage, though the mechanism is still to be better understood. Here we investigated on the effect of N-acetylcysteine (NAC, on gp120-induced damage in human cultured astroglial cells and the possible contribution of gp120-related reacting oxygen species (ROS in the imbalanced activity of glutamine synthase (GS, the enzyme that metabolizes glutamate into glutamine within astroglial cells playing a neuroprotective role in brain disorders. Results Incubation of Lipari human cultured astroglial cells with gp 120 (0.1–10 nM produced a significant reduction of astroglial cell viability and apoptosis as evaluated by TUNEL reaction and flow cytometric analysis (FACS. This effect was accompanied by lipid peroxidation as detected by means of malondialdehyde assay (MDA. In addition, gp 120 reduced both glutamine concentration in astroglial cell supernatants and GS expression as detected by immunocytochemistry and western blotting analysis. Pre-treatment of cells with NAC (0.5–5 mM, dose-dependently antagonised astroglial apoptotic cell death induced by gp 120, an effect accompanied by significant attenuation of MDA accumulation. Furthermore, both effects were closely associated with a significant recovery of glutamine levels in cell supernatants and by GS expression, thus suggesting that overproduction of free radicals might contribute in gp 120-related dysfunction of GS in astroglial cells. Conclusion In conclusion, the present experiments demonstrate that gp 120 is toxic to astroglial cells, an effect accompanied by lipid peroxidation and by altered

  2. Scaffold optimization in discontinuous epitope containing protein mimics of gp120 using smart libraries.

    Science.gov (United States)

    Mulder, Gwenn E; Quarles van Ufford, H Linda C; van Ameijde, Jeroen; Brouwer, Arwin J; Kruijtzer, John A W; Liskamp, Rob M J

    2013-04-28

    A diversity of protein surface discontinuous epitope mimics is now rapidly and efficiently accessible. Despite the important role of protein-protein interactions involving discontinuous epitopes in a wide range of diseases, mimicry of discontinuous epitopes using peptide-based molecules remains a major challenge. Using copper(I) catalyzed azide-alkyne cycloaddition (CuAAC), we have developed a general and efficient method for the synthesis of collections of discontinuous epitope mimics. Up to three different cyclic peptides, representing discontinuous epitopes in HIV-gp120, were conjugated to a selection of scaffold molecules. Variation of the scaffold molecule, optimization of the ring size of the cyclic peptides and screening of the resulting libraries for successful protein mimics led to an HIV gp120 mimic with an IC50 value of 1.7 μM. The approach described here provides rapid and highly reproducible access to clean, smart libraries of very complex bio-molecular constructs representing protein mimics for use as synthetic vaccines and beyond.

  3. Human immunodeficiency virus envelope protein Gp120 induces proliferation but not apoptosis in osteoblasts at physiologic concentrations.

    Directory of Open Access Journals (Sweden)

    Nathan W Cummins

    Full Text Available Patients with HIV infection have decreased numbers of osteoblasts, decreased bone mineral density and increased risk of fracture compared to uninfected patients; however, the molecular mechanisms behind these associations remain unclear. We questioned whether Gp120, a component of the envelope protein of HIV capable of inducing apoptosis in many cell types, is able to induce cell death in bone-forming osteoblasts. We show that treatment of immortalized osteoblast-like cells and primary human osteoblasts with exogenous Gp120 in vitro at physiologic concentrations does not result in apoptosis. Instead, in the osteoblast-like U2OS cell line, cells expressing CXCR4, a receptor for Gp120, had increased proliferation when treated with Gp120 compared to control (P<0.05, which was inhibited by pretreatment with a CXCR4 inhibitor and a G-protein inhibitor. This suggests that Gp120 is not an inducer of apoptosis in human osteoblasts and likely does not directly contribute to osteoporosis in infected patients by this mechanism.

  4. HIV-1 vaccines

    Science.gov (United States)

    Excler, Jean-Louis; Robb, Merlin L; Kim, Jerome H

    2014-01-01

    The development of a safe and effective preventive HIV-1 vaccine remains a public health priority. Despite scientific difficulties and disappointing results, HIV-1 vaccine clinical development has, for the first time, established proof-of-concept efficacy against HIV-1 acquisition and identified vaccine-associated immune correlates of risk. The correlate of risk analysis showed that IgG antibodies against the gp120 V2 loop correlated with decreased risk of HIV infection, while Env-specific IgA directly correlated with increased risk. The development of vaccine strategies such as improved envelope proteins formulated with potent adjuvants and DNA and vectors expressing mosaics, or conserved sequences, capable of eliciting greater breadth and depth of potentially relevant immune responses including neutralizing and non-neutralizing antibodies, CD4+ and CD8+ cell-mediated immune responses, mucosal immune responses, and immunological memory, is now proceeding quickly. Additional human efficacy trials combined with other prevention modalities along with sustained funding and international collaboration remain key to bring an HIV-1 vaccine to licensure. PMID:24637946

  5. Mechanisms of HIV-1 subtype C resistance to GRFT, CV-N and SVN

    CSIR Research Space (South Africa)

    Alexandre, Kabamba B

    2013-11-01

    Full Text Available We examined the ability of HIV-1 subtype C to develop resistance to the inhibitory lectins, griffithsin (GRFT), cyanovirin-N (CV-N) and scytovirin (SVN), which bind multiple mannose-rich glycans on gp120. Four primary HIV-1 strains cultured under...

  6. HIV-1 predisposed to acquiring resistance to maraviroc (MVC) and other CCR5 antagonists in vitro has an inherent, low-level ability to utilize MVC-bound CCR5 for entry

    National Research Council Canada - National Science Library

    Roche, Michael; Jakobsen, Martin R; Ellett, Anne; Salimiseyedabad, Hamid; Jubb, Becky; Westby, Mike; Lee, Benhur; Lewin, Sharon R; Churchill, Melissa J; Gorry, Paul R

    2011-01-01

    Maraviroc (MVC) and other CCR5 antagonists are HIV-1 entry inhibitors that bind to- and alter the conformation of CCR5, such that CCR5 is no longer recognized by the viral gp120 envelope (Env) glycoproteins...

  7. Deficient synthesis of class-switched, HIV-neutralizing antibodies to the CD4 binding site and correction by electrophilic gp120 immunogen

    Science.gov (United States)

    Planque, Stephanie A.; Mitsuda, Yukie; Chitsazzadeh, Vida; Gorantla, Santhi; Poluektova, Larisa; Nishiyama, Yasuhiro; Ochsenbauer, Christina; Morris, Mary-Kate; Sapparapu, Gopal; Hanson, Carl V.; Massey, Richard J.; Paul, Sudhir

    2015-01-01

    Objective HIV is vulnerable to antibodies that recognize a linear CD4 binding site epitope of gp120 (CLIN), but inducing CLIN-directed antibody synthesis by traditional vaccine principles is difficult. We wished to understand the basis for deficient CLIN-directed antibody synthesis and validate correction of the deficiency by an electrophilic gp120 analog (E-gp120) immunogen that binds B-cell receptors covalently. Methods Serum antibody responses to a CLIN peptide and full-length gp120 epitopes induced by HIV infection in humans and immunization of mice with gp120 or E-gp120 were monitored. HIV neutralization by monoclonal and variable domain-swapped antibodies was determined from tissue culture and humanized mouse infection assays. Results We describe deficient CLIN-directed IgG but not IgM antibodies in HIV-infected patients and mice immunized with gp120 accompanied by robust synthesis of IgGs to the immunodominant gp120 epitopes. Immunization with the E-gp120 corrected the deficient CLIN-directed IgG synthesis without overall increased immunogenicity of the CLIN or other gp120 epitopes. E-gp120-induced monoclonal IgGs neutralized diverse HIV strains heterologous to the immunogen. A CLIN-directed IgG neutralized HIV more potently compared to its larger IgM counterpart containing the same variable domains, suggesting obstructed access to HIV surface-expressed CLIN. An E-gp120-induced IgG suppressed HIV infection in humanized mice, validating the tissue culture neutralizing activity. Conclusion A CLIN-selective physiological defect of IgM→IgG class-switch recombination (CSR) or restricted post-CSR B-cell development limits the functional utility of the humoral immune response to gp120. The E-gp120 immunogen is useful to bypass the restriction and induce broadly neutralizing CLIN-directed IgGs (see Supplemental Video Abstract, http://links.lww.com/QAD/A551). PMID:25022597

  8. Synergistic cooperation between methamphetamine and HIV-1 gsp120 through the P13K/Akt pathway induces IL-6 but not IL-8 expression in astrocytes.

    Directory of Open Access Journals (Sweden)

    Ankit Shah

    Full Text Available HIV-1 envelope protein gp120 has been extensively studied for neurotoxic effects that have been attributed to the increased expression of various proinflammatory cytokines in the CNS. Recently we have shown that methamphetamine (MA also increases expression of proinflammatory cytokines in astrocytes. However, combined effect of gp120 and MA is not known. The present study was undertaken to determine cumulative effect and the mechanism(s/pathways involved in the functional interaction between gp120 and MA in SVGA astrocytes. Our results clearly suggest that gp120 and MA affect IL-6 but not IL-8 in a synergistic manner and this synergy was mediated by PI3K/Akt and NF-κB pathways. Inhibition of either of these pathways could abrogate the increased expression of IL-6 due to MA or gp120 alone, as well as the increased expression of IL-6 when the astrocytes were treated with both gp120 and MA. These results were confirmed by both, using chemical inhibitors/siRNA as well as western blotting. This study therefore provides novel information regarding the interaction between MA and gp120 in terms of the expression of IL-6 and the mechanisms underlying potential synergy between MA and gp120 in astrocytes.

  9. Epigallocatechin gallate, the main polyphenol in green tea, binds to the T-cell receptor, CD4: Potential for HIV-1 therapy.

    Science.gov (United States)

    Williamson, Mike P; McCormick, Theron G; Nance, Christina L; Shearer, William T

    2006-12-01

    The green tea flavonoid, epigallocatechin gallate (EGCG), has been proposed to have an anti-HIV-1 effect by preventing the binding of HIV-1 glycoprotein (gp) 120 to the CD4 molecule on T cells. To demonstrate that EGCG binds to the CD4 molecule at the gp120 attachment site and inhibits gp120 binding at physiologically relevant levels, thus establishing EGCG as a potential therapeutic treatment for HIV-1 infection. Nuclear magnetic resonance spectroscopy was used to examine the binding of EGCG and control, (-)-catechin, to CD4-IgG2 (PRO 542). Gp120 binding to human CD4+ T cells was analyzed by flow cytometry. Addition of CD4 to EGCG produced a linear decrease in nuclear magnetic resonance signal intensity from EGCG but not from the control, (-)-catechin. In saturation transfer difference experiments, addition of 5.8 micromol/L CD4 to 310 micromol/L EGCG produced strong saturation at the aromatic rings of EGCG, but identical concentrations of (-)-catechin produced much smaller effects, implying EGCG/CD4 binding strong enough to reduce gp120/CD4 binding substantially. Molecular modeling studies suggested a binding site for EGCG in the D1 domain of CD4, the pocket that binds gp120. Physiologically relevant concentrations of EGCG (0.2 micromol/L) inhibited binding of gp120 to isolated human CD4+ T cells. We have demonstrated clear evidence of high-affinity binding of EGCG to the CD4 molecule with a Kd of approximately 10 nmol/L and inhibition of gp120 binding to human CD4+ T cells. Epigallocatechin gallate has potential use as adjunctive therapy in HIV-1 infection.

  10. Secondary structure of gp160 and gp120 envelope glycoproteins of human immunodeficiency virus type 1: a Fourier transform infrared spectroscopic study.

    Science.gov (United States)

    Decroly, E; Cornet, B; Martin, I; Ruysschaert, J M; Vandenbranden, M

    1993-06-01

    The secondary structure of the precursor (gp160) of the envelope protein of human immunodeficiency virus type 1 (BH10) and its receptor-binding subunit (gp120) was studied by Fourier-transformed attenuated total reflection spectroscopy. A higher alpha-helix/beta-sheet ratio in the gp120 subunit than in the precursor indicates a structural heterogeneity between the two subunits (gp120 and gp41), in agreement with classical secondary-structure predictions. The secondary structure of gp41 was estimated and compared with existing models. The high alpha-helical content in gp41 and the dominant beta-sheet content in gp120 resemble the distribution in influenza virus hemagglutinin subunits.

  11. Towards HIV detection: Novel Poly(propylene imine) Dendrimer-Streptavidin platform for electrochemical DNA and gp120 aptamer biosensors

    CSIR Research Space (South Africa)

    John, SV

    2014-07-01

    Full Text Available (2014) 5425 - 5437 International Journal of ELECTROCHEMICAL SCIENCE www.electrochemsci.org Towards HIV Detection: Novel Poly(propylene imine) Dendrimer-Streptavidin Platform for Electrochemical DNA and gp120 Aptamer Biosensors Suru V.... Gray, M. C. Madiga, N. Tumba, K. B. Alexandre, T. Khoza, C. K. Wibmer, P. L. Moore, L. Morris and M. M. Khati, Journal of Virology, 86 (2012) 1 15. G. Li, X. Li, J. Wan and S. Zhang, Biosensors and Bioelectronics, 24 (2009) 3281 16. A. K. H. Cheng...

  12. HIV-1 envelope sequence-based diversity measures for identifying recent infections.

    Directory of Open Access Journals (Sweden)

    Alexis Kafando

    Full Text Available Identifying recent HIV-1 infections is crucial for monitoring HIV-1 incidence and optimizing public health prevention efforts. To identify recent HIV-1 infections, we evaluated and compared the performance of 4 sequence-based diversity measures including percent diversity, percent complexity, Shannon entropy and number of haplotypes targeting 13 genetic segments within the env gene of HIV-1. A total of 597 diagnostic samples obtained in 2013 and 2015 from recently and chronically HIV-1 infected individuals were selected. From the selected samples, 249 (134 from recent versus 115 from chronic infections env coding regions, including V1-C5 of gp120 and the gp41 ectodomain of HIV-1, were successfully amplified and sequenced by next generation sequencing (NGS using the Illumina MiSeq platform. The ability of the four sequence-based diversity measures to correctly identify recent HIV infections was evaluated using the frequency distribution curves, median and interquartile range and area under the curve (AUC of the receiver operating characteristic (ROC. Comparing the median and interquartile range and evaluating the frequency distribution curves associated with the 4 sequence-based diversity measures, we observed that the percent diversity, number of haplotypes and Shannon entropy demonstrated significant potential to discriminate recent from chronic infections (p<0.0001. Using the AUC of ROC analysis, only the Shannon entropy measure within three HIV-1 env segments could accurately identify recent infections at a satisfactory level. The env segments were gp120 C2_1 (AUC = 0.806, gp120 C2_3 (AUC = 0.805 and gp120 V3 (AUC = 0.812. Our results clearly indicate that the Shannon entropy measure represents a useful tool for predicting HIV-1 infection recency.

  13. Oxidative stress and protein oxidation in the brain of water drinking and alcohol drinking rats administered the HIV envelope protein, gp120.

    Science.gov (United States)

    Singh, Ashok K; Gupta, Shveta; Jiang, Yin

    2008-03-01

    Possible roles of oxidative stress and protein oxidation on alcohol-induced augmentation of cerebral neuropathy in gp120 administered alcohol preferring rats drinking either pure water (W rats) or a free-choice ethanol and water (E rats) for 90 days. This study showed that peripherally administered gp120 accumulated into the brain, liver, and RBCs samples from water drinking - gp120 administered rats (Wg rats) and ethanol drinking - gp120 administered rats (Eg rats), although gp120 levels in samples from Eg rats were significantly greater than the levels in samples from Wg rats. The brain samples from ethanol drinking-saline administered (EC) and Wg rats exhibited comparable levels of free radicals that were significantly lower than the levels in Eg rats. Peroxiredoxin-I (PrxI) activity in the brain samples exhibited the following pattern: Wg > > WC > EC > Eg. Total protein-carbonyl and carbonylated hippocampal cholinergic neurostimulating peptide precursor protein levels, but not N-acetylaspartate or N-acetyl aspartylglutamate or total protein-thiol levels, paralleled the free radical levels in the brain of all four groups. This suggests PrxI inhibition may be more sensitive indicator of oxidative stress than measuring free radicals or metabolites. As PrxI oxidation in WC, Wg, and EC rats was reversible, while PrxI oxidation in Eg rats was not, we suggest that alcohol drinking and gp120 together hyperoxidized and inactivated PrxI that suppressed free radical neutralization in the brain of Eg rats. In conclusion, chronic alcohol drinking, by carbonylating and hyperoxidizing free radical neutralization proteins, augmented the gp120-induced oxidative stress that may be associated with an increase in severity of the brain neuropathy.

  14. Trimeric gp120-specific bovine monoclonal antibodies require cysteine and aromatic residues in CDRH3 for high affinity binding to HIV Env.

    Science.gov (United States)

    Heydarchi, Behnaz; Center, Rob J; Bebbington, Jonathan; Cuthbertson, Jack; Gonelli, Christopher; Khoury, Georges; Mackenzie, Charlene; Lichtfuss, Marit; Rawlin, Grant; Muller, Brian; Purcell, Damian

    2017-04-01

    We isolated HIV-1 Envelope (Env)-specific memory B cells from a cow that had developed high titer polyclonal immunoglobulin G (IgG) with broad neutralizing activity after a long duration vaccination with HIV-1AD8 Env gp140 trimers. We cloned the bovine IgG matched heavy (H) and light (L) chain variable (V) genes from these memory B cells and constructed IgG monoclonal antibodies (mAbs) with either a human constant (C)-region/bovine V-region chimeric or fully bovine C and V regions. Among 42 selected Ig+ memory B cells, two mAbs (6A and 8C) showed high affinity binding to gp140 Env. Characterization of both the fully bovine and human chimeric isoforms of these two mAbs revealed them as highly type-specific and capable of binding only to soluble AD8 uncleaved gp140 trimers and covalently stabilized AD8 SOSIP gp140 cleaved trimers, but not monomeric gp120. Genomic sequence analysis of the V genes showed the third heavy complementarity-determining region (CDRH3) of 6A mAb was 21 amino acids in length while 8C CDRH3 was 14 amino acids long. The entire V heavy (VH) region was 27% and 25% diverged for 6A and 8C, respectively, from the best matched germline V genes available, and the CDRH3 regions of 6A and 8C were 47.62% and 78.57% somatically mutated, respectively, suggesting a high level of somatic hypermutation compared with CDRH3 of other species. Alanine mutagenesis of the VH genes of 6A and 8C, showed that CDRH3 cysteine and tryptophan amino acids were crucial for antigen binding. Therefore, these bovine vaccine-induced anti-HIV antibodies shared some of the notable structural features of elite human broadly neutralizing antibodies, such as CDRH3 size and somatic mutation during affinity-maturation. However, while the 6A and 8C mAbs inhibited soluble CD4 binding to gp140 Env, they did not recapitulate the neutralizing activity of the polyclonal antibodies against HIV infection.

  15. Origin and dynamics of HIV-1 subtype C infection in India.

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    Chengli Shen

    Full Text Available To investigate the geographical origin and evolution dynamics of HIV-1 subtype C infection in India.Ninety HIV-1 subtype C env gp120 subtype C sequences from India were compared with 312 env gp120 reference subtype C sequences from 27 different countries obtained from Los Alamos HIV database. All the HIV-1 subtype C env gp120 sequences from India were used for the geographical origin analysis and 61 subtype C env gp120 sequences with known sampling year (from 1991 to 2008 were employed to determine the origin of HIV infection in India.Phylogenetic analysis of HIV-1 env sequences was used to investigate the geographical origin and tMRCA of Indian HIV-1 subtype C. Evolutionary parameters including origin date and demographic growth patterns of Indian subtype C were estimated using a Bayesian coalescent-based approach under relaxed molecular clock models.The majority of the analyzed Indian and South African HIV-1 subtype C sequences formed a single monophyletic cluster. The most recent common ancestor date was calculated to be 1975.56 (95% HPD, 1968.78-1981.52. Reconstruction of the effective population size revealed three phases of epidemic growth: an initial slow growth, followed by exponential growth, and then a plateau phase approaching present time. Stabilization of the epidemic growth phase correlated with the foundation of National AIDS Control Organization in India.Indian subtype C originated from a single South African lineage in the middle of 1970s. The current study emphasizes not only the utility of HIV-1 sequence data for epidemiological studies but more notably highlights the effectiveness of community or government intervention strategies in controlling the trend of the epidemic.

  16. Anti-HIV double variable domain immunoglobulins binding both gp41 and gp120 for targeted delivery of immunoconjugates.

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    Ryan B Craig

    Full Text Available BACKGROUND: Anti-HIV immunoconjugates targeted to the HIV envelope protein may be used to eradicate the latent reservoir of HIV infection using activate-and-purge protocols. Previous studies have identified the two target epitopes most effective for the delivery of cytotoxic immunoconjugates the CD4-binding site of gp120, and the hairpin loop of gp41. Here we construct and test tetravalent double variable domain immunoglobulin molecules (DVD-Igs that bind to both epitopes. METHODS: Synthetic genes that encode DVD-Igs utilizing V-domains derived from human anti-gp120 and anti-gp41 Abs were designed and expressed in 293F cells. A series of constructs tested different inter-V-linker domains and orientations of the two V domains. Antibodies were tested for binding to recombinant Ag and native Env expressed on infected cells, for neutralization of infectious HIV, and for their ability to deliver cytotoxic immunoconjugates to infected cells. FINDINGS: The outer V-domain was the major determinant of binding and functional activity of the DVD-Ig. Function of the inner V-domain and bifunctional binding required at least 15 AA in the inter-V-domain linker. A molecular model showing the spatial orientation of the two epitopes is consistent with this observation. Linkers that incorporated helical domains (A[EAAAK](nA resulted in more effective DVD-Igs than those based solely on flexible domains ([GGGGS](n. In general, the DVD-Igs outperformed the less effective parental antibody and equaled the activity of the more effective. The ability of the DVD-Igs to deliver cytotoxic immunoconjugates in the absence of soluble CD4 was improved over that of either parent. CONCLUSIONS: DVD-Igs can be designed that bind to both gp120 and gp41 on the HIV envelope. DVD-Igs are effective in delivering cytotoxic immunoconjugates. The optimal design of these DVD-Igs, in which both domains are fully functional, has not yet been achieved.

  17. HIV-1 Fusion Is Blocked through Binding of GB Virus C E2D Peptides to the HIV-1 gp41 Disulfide Loop

    Science.gov (United States)

    Eissmann, Kristin; Mueller, Sebastian; Sticht, Heinrich; Jung, Susan; Zou, Peng; Jiang, Shibo; Gross, Andrea; Eichler, Jutta; Fleckenstein, Bernhard; Reil, Heide

    2013-01-01

    A strategy for antiviral drug discovery is the elucidation and imitation of viral interference mechanisms. HIV-1 patients benefit from a coinfection with GB Virus C (GBV-C), since HIV-positive individuals with long-term GBV-C viraemia show better survival rates than HIV-1 patients without persisting GBV-C. A direct influence of GBV-C on HIV-1 replication has been shown in coinfection experiments. GBV-C is a human non-pathogenic member of the flaviviridae family that can replicate in T and B cells. Therefore, GBV-C shares partly the same ecological niche with HIV-1. In earlier work we have demonstrated that recombinant glycoprotein E2 of GBV-C and peptides derived from the E2 N-terminus interfere with HIV entry. In this study we investigated the underlying mechanism. Performing a virus-cell fusion assay and temperature-arrested HIV-infection kinetics, we provide evidence that the HIV-inhibitory E2 peptides interfere with late HIV-1 entry steps after the engagement of gp120 with CD4 receptor and coreceptor. Binding and competition experiments revealed that the N-terminal E2 peptides bind to the disulfide loop region of HIV-1 transmembrane protein gp41. In conjunction with computational analyses, we identified sequence similarities between the N-termini of GBV-C E2 and the HIV-1 glycoprotein gp120. This similarity appears to enable the GBV-C E2 N-terminus to interact with the HIV-1 gp41 disulfide loop, a crucial domain involved in the gp120-gp41 interface. Furthermore, the results of the present study provide initial proof of concept that peptides targeted to the gp41 disulfide loop are able to inhibit HIV fusion and should inspire the development of this new class of HIV-1 entry inhibitors. PMID:23349893

  18. HIV-1 fusion is blocked through binding of GB Virus C E2-derived peptides to the HIV-1 gp41 disulfide loop [corrected].

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    Kristin Eissmann

    Full Text Available A strategy for antiviral drug discovery is the elucidation and imitation of viral interference mechanisms. HIV-1 patients benefit from a coinfection with GB Virus C (GBV-C, since HIV-positive individuals with long-term GBV-C viraemia show better survival rates than HIV-1 patients without persisting GBV-C. A direct influence of GBV-C on HIV-1 replication has been shown in coinfection experiments. GBV-C is a human non-pathogenic member of the flaviviridae family that can replicate in T and B cells. Therefore, GBV-C shares partly the same ecological niche with HIV-1. In earlier work we have demonstrated that recombinant glycoprotein E2 of GBV-C and peptides derived from the E2 N-terminus interfere with HIV entry. In this study we investigated the underlying mechanism. Performing a virus-cell fusion assay and temperature-arrested HIV-infection kinetics, we provide evidence that the HIV-inhibitory E2 peptides interfere with late HIV-1 entry steps after the engagement of gp120 with CD4 receptor and coreceptor. Binding and competition experiments revealed that the N-terminal E2 peptides bind to the disulfide loop region of HIV-1 transmembrane protein gp41. In conjunction with computational analyses, we identified sequence similarities between the N-termini of GBV-C E2 and the HIV-1 glycoprotein gp120. This similarity appears to enable the GBV-C E2 N-terminus to interact with the HIV-1 gp41 disulfide loop, a crucial domain involved in the gp120-gp41 interface. Furthermore, the results of the present study provide initial proof of concept that peptides targeted to the gp41 disulfide loop are able to inhibit HIV fusion and should inspire the development of this new class of HIV-1 entry inhibitors.

  19. Bioinformatic analysis of neurotropic HIV envelope sequences identifies polymorphisms in the gp120 bridging sheet that increase macrophage-tropism through enhanced interactions with CCR5

    Energy Technology Data Exchange (ETDEWEB)

    Mefford, Megan E., E-mail: megan_mefford@hms.harvard.edu [Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Boston, MA (United States); Kunstman, Kevin, E-mail: kunstman@northwestern.edu [Northwestern University Medical School, Chicago, IL (United States); Wolinsky, Steven M., E-mail: s-wolinsky@northwestern.edu [Northwestern University Medical School, Chicago, IL (United States); Gabuzda, Dana, E-mail: dana_gabuzda@dfci.harvard.edu [Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Boston, MA (United States); Department of Neurology (Microbiology and Immunobiology), Harvard Medical School, Boston, MA (United States)

    2015-07-15

    Macrophages express low levels of the CD4 receptor compared to T-cells. Macrophage-tropic HIV strains replicating in brain of untreated patients with HIV-associated dementia (HAD) express Envs that are adapted to overcome this restriction through mechanisms that are poorly understood. Here, bioinformatic analysis of env sequence datasets together with functional studies identified polymorphisms in the β3 strand of the HIV gp120 bridging sheet that increase M-tropism. D197, which results in loss of an N-glycan located near the HIV Env trimer apex, was detected in brain in some HAD patients, while position 200 was estimated to be under positive selection. D197 and T/V200 increased fusion and infection of cells expressing low CD4 by enhancing gp120 binding to CCR5. These results identify polymorphisms in the HIV gp120 bridging sheet that overcome the restriction to macrophage infection imposed by low CD4 through enhanced gp120–CCR5 interactions, thereby promoting infection of brain and other macrophage-rich tissues. - Highlights: • We analyze HIV Env sequences and identify amino acids in beta 3 of the gp120 bridging sheet that enhance macrophage tropism. • These amino acids at positions 197 and 200 are present in brain of some patients with HIV-associated dementia. • D197 results in loss of a glycan near the HIV Env trimer apex, which may increase exposure of V3. • These variants may promote infection of macrophages in the brain by enhancing gp120–CCR5 interactions.

  20. Escherichia coli surface display of single-chain antibody VRC01 against HIV-1 infection

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Lin-Xu [Nebraska Center for Virology, Lincoln, NE (United States); School of Biological Sciences, University of Nebraska—Lincoln, Lincoln, NE 68583 (United States); Mellon, Michael [Nebraska Center for Virology, Lincoln, NE (United States); School of Veterinary Medicine and Biomedical Sciences, Lincoln, NE (United States); Bowder, Dane [Nebraska Center for Virology, Lincoln, NE (United States); School of Biological Sciences, University of Nebraska—Lincoln, Lincoln, NE 68583 (United States); Quinn, Meghan [Nebraska Center for Virology, Lincoln, NE (United States); School of Veterinary Medicine and Biomedical Sciences, Lincoln, NE (United States); Shea, Danielle; Wood, Charles [Nebraska Center for Virology, Lincoln, NE (United States); School of Biological Sciences, University of Nebraska—Lincoln, Lincoln, NE 68583 (United States); Xiang, Shi-Hua, E-mail: sxiang2@unl.edu [Nebraska Center for Virology, Lincoln, NE (United States); School of Veterinary Medicine and Biomedical Sciences, Lincoln, NE (United States)

    2015-01-15

    Human immunodeficiency virus type 1 (HIV-1) transmission and infection occur mainly via the mucosal surfaces. The commensal bacteria residing in these surfaces can potentially be employed as a vehicle for delivering inhibitors to prevent HIV-1 infection. In this study, we have employed a bacteria-based strategy to display a broadly neutralizing antibody VRC01, which could potentially be used to prevent HIV-1 infection. The VRC01 antibody mimics CD4-binding to gp120 and has broadly neutralization activities against HIV-1. We have designed a construct that can express the fusion peptide of the scFv-VRC01 antibody together with the autotransporter β-barrel domain of IgAP gene from Neisseria gonorrhoeae, which enabled surface display of the antibody molecule. Our results indicate that the scFv-VRC01 antibody molecule was displayed on the surface of the bacteria as demonstrated by flow cytometry and immunofluorescence microscopy. The engineered bacteria can capture HIV-1 particles via surface-binding and inhibit HIV-1 infection in cell culture. - Highlights: • Designed single-chain VRC01 antibody was demonstrated to bind HIV-1 envelope gp120. • Single-chain VRC01 antibody was successfully displayed on the surface of E. coli. • Engineered bacteria can absorb HIV-1 particles and prevent HIV-1 infection in cell culture.

  1. Effects of the I559P gp41 change on the conformation and function of the human immunodeficiency virus (HIV-1 membrane envelope glycoprotein trimer.

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    Nirmin Alsahafi

    Full Text Available The mature human immunodeficiency virus (HIV-1 envelope glycoprotein (Env trimer is produced by proteolytic cleavage of a precursor and consists of three gp120 exterior and three gp41 transmembrane subunits. The metastable Env complex is induced to undergo conformational changes required for virus entry by the binding of gp120 to the receptors, CD4 and CCR5/CXCR4. An isoleucine-to-proline change (I559P in the gp41 ectodomain has been used to stabilize soluble forms of HIV-1 Env trimers for structural characterization and for use as immunogens. In the native membrane-anchored HIV-1BG505 Env, the I559P change modestly decreased proteolytic maturation, increased the non-covalent association of gp120 with the Env trimer, and resulted in an Env conformation distinctly different from that of the wild-type HIV-1BG505 Env. Compared with the wild-type Env, the I559P Env was recognized inefficiently by polyclonal sera from HIV-1-infected individuals, by several gp41-directed antibodies, by some antibodies against the CD4-binding site of gp120, and by antibodies that preferentially recognize the CD4-bound Env. Some of the gp120-associated antigenic differences between the wild-type HIV-1BG505 Env and the I559P mutant were compensated by the SOS disulfide bond between gp120 and gp41, which has been used to stabilize cleaved soluble Env trimers. Nonetheless, regardless of the presence of the SOS changes, Envs with proline 559 were recognized less efficiently than Envs with isoleucine 559 by the VRC01 neutralizing antibody, which binds the CD4-binding site of gp120, and the PGT151 neutralizing antibody, which binds a hybrid gp120-gp41 epitope. The I559P change completely eliminated the ability of the HIV-1BG505 Env to mediate cell-cell fusion and virus entry, and abolished the capacity of the SOS Env to support virus infection in the presence of a reducing agent. These results suggest that differences exist between the quaternary structures of functional Env

  2. Vaccine induced antibodies to the first variable loop of human immunodeficiency virus type 1 gp120, mediate antibody-dependent virus inhibition in macaques.

    Science.gov (United States)

    Bialuk, Izabela; Whitney, Stephen; Andresen, Vibeke; Florese, Ruth H; Nacsa, Janos; Cecchinato, Valentina; Valeri, Valerio W; Heraud, Jean-Michel; Gordon, Shari; Parks, Robyn Washington; Montefiori, David C; Venzon, David; Demberg, Thorsten; Guroff, Marjorie Robert-; Landucci, Gary; Forthal, Donald N; Franchini, Genoveffa

    2011-12-09

    The role of antibodies directed against the hyper variable envelope region V1 of human immunodeficiency virus type 1 (HIV-1), has not been thoroughly studied. We show that a vaccine able to elicit strain-specific non-neutralizing antibodies to this region of gp120 is associated with control of highly pathogenic chimeric SHIV(89.6P) replication in rhesus macaques. The vaccinated animal that had the highest titers of antibodies to the amino terminus portion of V1, prior to challenge, had secondary antibody responses that mediated cell killing by antibody-dependent cellular cytotoxicity (ADCC), as early as 2 weeks after infection and inhibited viral replication by antibody-dependent cell-mediated virus inhibition (ADCVI), by 4 weeks after infection. There was a significant inverse correlation between virus level and binding antibody titers to the envelope protein, (R=-0.83, p=0.015), and ADCVI (R=-0.84 p=0.044). Genotyping of plasma virus demonstrated in vivo selection of three SHIV(89.6P) variants with changes in potential N-linked glycosylation sites in V1. We found a significant inverse correlation between virus levels and titers of antibodies that mediated ADCVI against all the identified V1 virus variants. A significant inverse correlation was also found between neutralizing antibody titers to SHIV(89.6) and virus levels (R=-0.72 p=0.0050). However, passive inoculation of purified immunoglobulin from animal M316, the macaque that best controlled virus, to a naïve macaque, resulted in a low serum neutralizing antibodies and low ADCVI activity that failed to protect from SHIV(89.6P) challenge. Collectively, while our data suggest that anti-envelope antibodies with neutralizing and non-neutralizing Fc(R-dependent activities may be important in the control of SHIV replication, they also demonstrate that low levels of these antibodies alone are not sufficient to protect from infection. Published by Elsevier Ltd.

  3. HIV-1 vaccine induced immune responses in newborns of HIV-1 infected mothers.

    Science.gov (United States)

    McFarland, Elizabeth J; Johnson, Daniel C; Muresan, Petronella; Fenton, Terence; Tomaras, Georgia D; McNamara, James; Read, Jennifer S; Douglas, Steven D; Deville, Jaime; Gurwith, Marc; Gurunathan, Sanjay; Lambert, John S

    2006-07-13

    Breast milk transmission continues to account for a large proportion of cases of mother-to-child transmission of HIV-1 worldwide. An effective HIV-1 vaccine coupled with either passive immunization or short-term antiretroviral prophylaxis represents a potential strategy to prevent breast milk transmission. This study evaluated the safety and immunogenicity of ALVAC HIV-1 vaccine with and without a subunit envelope boost in infants born to HIV-1-infected women. : Placebo-controlled, double-blinded study. Infants born to HIV-1-infected mothers in the US were immunized with a prime-boost regimen using a canarypox virus HIV-1 vaccine (vCP1452) and a recombinant glycoprotein subunit vaccine (rgp120). Infants (n = 30) were randomized to receive: vCP1452 alone, vCP1452 + rgp120, or corresponding placebos. Local reactions were mild or moderate and no significant systemic toxicities occurred. Subjects receiving both vaccines had gp120-specific binding serum antibodies that were distinguishable from maternal antibody. Repeated gp160-specific lymphoproliferative responses were observed in 75%. Neutralizing activity to HIV-1 homologous to the vaccine strain was observed in 50% of the vCP1452 + rgp120 subjects who had lost maternal antibody by week 24. In some infants HIV-1-specific proliferative and antibody responses persisted until week 104. HIV-1-specific cytotoxic T lymphocyte responses were detected in two subjects in each treatment group; the frequency of HIV-1 specific cytotoxic T lymphocyte responses did not differ between vaccine and placebo recipients. The demonstration of vaccine-induced immune responses in early infancy supports further study of HIV-1 vaccination as a strategy to reduce breast milk transmission.

  4. Increased chemokine signaling in a model of HIV1-associated peripheral neuropathy

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    Buchanan David J

    2009-08-01

    Full Text Available Abstract Painful distal sensory polyneuropathy (DSP is the most common neurological complication of HIV1 infection. Although infection with the virus itself is associated with an incidence of DSP, patients are more likely to become symptomatic following initiation of nucleoside reverse transcriptase inhibitor (NRTI treatment. The chemokines monocyte chemoattractant protein-1 (MCP1/CCL2 and stromal derived factor-1 (SDF1/CXCL12 and their respective receptors, CCR2 and CXCR4, have been implicated in HIV1 related neuropathic pain mechanisms including NRTI treatment in rodents. Utilizing a rodent model that incorporates the viral coat protein, gp120, and the NRTI, 2'3'-dideoxycytidine (ddC, we examined the degree to which chemokine receptor signaling via CCR2 and CXCR4 potentially influences the resultant chronic hypernociceptive behavior. We observed that following unilateral gp120 sciatic nerve administration, rats developed profound tactile hypernociception in the hindpaw ipsilateral to gp120 treatment. Behavioral changes were also present in the hindpaw contralateral to the injury, albeit delayed and less robust. Using immunohistochemical studies, we demonstrated that MCP1 and CCR2 were upregulated by primary sensory neurons in lumbar ganglia by post-operative day (POD 14. The functional nature of these observations was confirmed using calcium imaging in acutely dissociated lumbar dorsal root ganglion (DRG derived from gp120 injured rats at POD 14. Tactile hypernociception in gp120 treated animals was reversed following treatment with a CCR2 receptor antagonist at POD 14. Some groups of animals were subjected to gp120 sciatic nerve injury in combination with an injection of ddC at POD 14. This injury paradigm produced pronounced bilateral tactile hypernociception from POD 14–48. More importantly, functional MCP1/CCR2 and SDF1/CXCR4 signaling was present in sensory neurons. In contrast to gp120 treatment alone, the hypernociceptive behavior

  5. Absence of cytotoxic antibody to human immunodeficiency virus-infected cells in humans and its induction in animals after infection or immunization with purified envelope glycoprotein gp120

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    Nara, P.L.; Robey, W.G.; Gonda, M.A.; Carter, S.G.; Fischinger, P.J.

    1987-06-01

    The presence of antibody-dependent complement-mediated cytotoxicity (ACC) was assessed in humans and chimpanzees, which are capable of infection with human immunodeficiency virus isolate HTLV-IIIb, and examined in the goat after immunization with the major viral glycoprotein (gp120) of HTLV-IIIb. In infected humans no antibody mediating ACC was observed regardless of the status of disease. Even healthy individuals with high-titer, broadly reactive, neutralizing antibodies has no ACC. In contrast, chimpanzees infected with HTLV-IIIb, from whom virus could be isolated, not only had neutralizing antibody but also antibodies broadly reactive in ACC, even against distantly related human immunodeficiency virus isolates, as well as against their own reisolated virus. In the goat, the gp120 of HTLV-IIIb induced a highly type-specific response as measured by both ACC and flow cytofluorometry of live infected H9 cells. Normal human cells were not subject to ACC by animal anti-HTLV-III gp120-specific sera. Induction of ACC and neutralizing antibody were closely correlated in the animal experimental models but not in humans. The presence of ACC in gp120-inoculated goats and HTLV-III-infected chimpanzees represent a qualitative difference that may be important in the quest for the elicitation of a protective immunity in humans.

  6. Paring Down HIV Env: Design and Crystal Structure of a Stabilized Inner Domain of HIV-1 gp120 Displaying a Major ADCC Target of the A32 Region

    National Research Council Canada - National Science Library

    Tolbert, William D; Gohain, Neelakshi; Veillette, Maxime; Chapleau, Jean-Philippe; Orlandi, Chiara; Visciano, Maria L; Ebadi, Maryam; DeVico, Anthony L; Fouts, Timothy R; Finzi, Andrés; Lewis, George K; Pazgier, Marzena

    2016-01-01

    .... Through structure-based design, we developed ID2, which consists of the ID expressed independently of the outer domain and stabilized in the CD4-bound conformation by an inter-layer disulfide bond...

  7. Human Ubc9 is involved in intracellular HIV-1 Env stability after trafficking out of the trans-Golgi network in a Gag dependent manner.

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    Christopher R Bohl

    Full Text Available The cellular E2 Sumo conjugase, Ubc9 interacts with HIV-1 Gag, and is important for the assembly of infectious HIV-1 virions. In the previous study we demonstrated that in the absence of Ubc9, a defect in virion assembly was associated with decreased levels of mature intracellular Envelope (Env that affected Env incorporation into virions and virion infectivity. We have further characterized the effect of Ubc9 knockdown on HIV Env processing and assembly. We found that gp160 stability in the endoplasmic reticulum (ER and its trafficking to the trans-Golgi network (TGN were unaffected, indicating that the decreased intracellular mature Env levels in Ubc9-depleted cells were due to a selective degradation of mature Env gp120 after cleavage from gp160 and trafficked out of the TGN. Decreased levels of Gag and mature Env were found to be associated with the plasma membrane and lipid rafts, which suggest that these viral proteins were not trafficked correctly to the assembly site. Intracellular gp120 were partially rescued when treated with a combination of lysosome inhibitors. Taken together our results suggest that in the absence of Ubc9, gp120 is preferentially degraded in the lysosomes likely before trafficking to assembly sites leading to the production of defective virions. This study provides further insight in the processing and packaging of the HIV-1 gp120 into mature HIV-1 virions.

  8. Human Ubc9 is involved in intracellular HIV-1 Env stability after trafficking out of the trans-Golgi network in a Gag dependent manner.

    Science.gov (United States)

    Bohl, Christopher R; Abrahamyan, Levon G; Wood, Charles

    2013-01-01

    The cellular E2 Sumo conjugase, Ubc9 interacts with HIV-1 Gag, and is important for the assembly of infectious HIV-1 virions. In the previous study we demonstrated that in the absence of Ubc9, a defect in virion assembly was associated with decreased levels of mature intracellular Envelope (Env) that affected Env incorporation into virions and virion infectivity. We have further characterized the effect of Ubc9 knockdown on HIV Env processing and assembly. We found that gp160 stability in the endoplasmic reticulum (ER) and its trafficking to the trans-Golgi network (TGN) were unaffected, indicating that the decreased intracellular mature Env levels in Ubc9-depleted cells were due to a selective degradation of mature Env gp120 after cleavage from gp160 and trafficked out of the TGN. Decreased levels of Gag and mature Env were found to be associated with the plasma membrane and lipid rafts, which suggest that these viral proteins were not trafficked correctly to the assembly site. Intracellular gp120 were partially rescued when treated with a combination of lysosome inhibitors. Taken together our results suggest that in the absence of Ubc9, gp120 is preferentially degraded in the lysosomes likely before trafficking to assembly sites leading to the production of defective virions. This study provides further insight in the processing and packaging of the HIV-1 gp120 into mature HIV-1 virions.

  9. Transplanting supersites of HIV-1 vulnerability.

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    Tongqing Zhou

    Full Text Available One strategy for isolating or eliciting antibodies against a specific target region on the envelope glycoprotein trimer (Env of the human immunodeficiency virus type 1 (HIV-1 involves the creation of site transplants, which present the target region on a heterologous protein scaffold with preserved antibody-binding properties. If the target region is a supersite of HIV-1 vulnerability, recognized by a collection of broadly neutralizing antibodies, this strategy affords the creation of "supersite transplants", capable of binding (and potentially eliciting antibodies similar to the template collection of effective antibodies. Here we transplant three supersites of HIV-1 vulnerability, each targeted by effective neutralizing antibodies from multiple donors. To implement our strategy, we chose a single representative antibody against each of the target supersites: antibody 10E8, which recognizes the membrane-proximal external region (MPER on the HIV-1 gp41 glycoprotein; antibody PG9, which recognizes variable regions one and two (V1V2 on the HIV-1 gp120 glycoprotein; and antibody PGT128 which recognizes a glycopeptide supersite in variable region 3 (glycan V3 on gp120. We used a structural alignment algorithm to identify suitable acceptor proteins, and then designed, expressed, and tested antigenically over 100-supersite transplants in a 96-well microtiter-plate format. The majority of the supersite transplants failed to maintain the antigenic properties of their respective template supersite. However, seven of the glycan V3-supersite transplants exhibited nanomolar affinity to effective neutralizing antibodies from at least three donors and recapitulated the mannose9-N-linked glycan requirement of the template supersite. The binding of these transplants could be further enhanced by placement into self-assembling nanoparticles. Essential elements of the glycan V3 supersite, embodied by as few as 3 N-linked glycans and ∼ 25 Env residues, can be

  10. Kinetic studies of HIV-1 and HIV-2 envelope glycoprotein-mediated fusion

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    Doms Robert W

    2006-12-01

    Full Text Available Abstract Background HIV envelope glycoprotein (Env-mediated fusion is driven by the concerted coalescence of the HIV gp41 N-helical and C-helical regions, which results in the formation of 6 helix bundles. Kinetics of HIV Env-mediated fusion is an important determinant of sensitivity to entry inhibitors and antibodies. However, the parameters that govern the HIV Env fusion cascade have yet to be fully elucidated. We address this issue by comparing the kinetics HIV-1IIIB Env with those mediated by HIV-2 from two strains with different affinities for CD4 and CXCR4. Results HIV-1 and HIV-2 Env-mediated cell fusion occurred with half times of about 60 and 30 min, respectively. Binding experiments of soluble HIV gp120 proteins to CD4 and co-receptor did not correlate with the differences in kinetics of fusion mediated by the three different HIV Envs. However, escape from inhibition by reagents that block gp120-CD4 binding, CD4-induced CXCR4 binding and 6-helix bundle formation, respectively, indicated large difference between HIV-1 and HIV-2 envelope glycoproteins in their CD4-induced rates of engagement with CXCR4. Conclusion The HIV-2 Env proteins studied here exhibited a significantly reduced window of time between the engagement of gp120 with CD4 and exposure of the CXCR4 binding site on gp120 as compared with HIV-1IIIB Env. The efficiency with which HIV-2 Env undergoes this CD4-induced conformational change is the major cause of the relatively rapid rate of HIV-2 Env mediated-fusion.

  11. Catechins containing a galloyl moiety as potential anti-HIV-1 compounds.

    Science.gov (United States)

    Zhao, Yali; Jiang, Fan; Liu, Ping; Chen, Wei; Yi, Kejia

    2012-06-01

    Catechins containing galloyl moieties are important natural antioxidant compounds. In this paper, we review the multiple mechanisms whereby catechins containing a galloyl moiety can target key proteins to inhibit sexual transmission of HIV-1, as well as HIV-1 fusion, HIV-1 reverse transcriptase, HIV-1 integrase and HIV-1 protease. Furthermore, catechins with a galloyl moiety can mediate host cell factors such as nitric oxide synthase, nuclear factor-κB and casein kinase II to inhibit HIV-1 infection. The most significant inhibitory effect is blocking gp120 binding to isolated human CD4+ T cells. The multiple mechanisms underlying the anti-HIV activity of galloyl-containing catechins predict that these catechins could be used as alternative therapies in the treatment of HIV infection. Crown Copyright © 2012. Published by Elsevier Ltd. All rights reserved.

  12. Glycosaminoglycans are interactants of Langerin: comparison with gp120 highlights an unexpected calcium-independent binding mode.

    Directory of Open Access Journals (Sweden)

    Eric Chabrol

    Full Text Available Langerin is a C-type lectin specifically expressed in Langerhans cells. As recently shown for HIV, Langerin is thought to capture pathogens and mediate their internalisation into Birbeck Granules for elimination. However, the precise functions of Langerin remain elusive, mostly because of the lack of information on its binding properties and physiological ligands. Based on recent reports that Langerin binds to sulfated sugars, we conducted here a comparative analysis of Langerin interaction with mannose-rich HIV glycoprotein gp120 and glycosaminoglycan (GAGs, a family of sulfated polysaccharides expressed at the surface of most mammalian cells. Our results first revealed that Langerin bound to these different glycans through very distinct mechanisms and led to the identification of a novel, GAG-specific binding mode within Langerin. In contrast to the canonical lectin domain, this new binding site showed no Ca(2+-dependency, and could only be detected in entire, trimeric extracellular domains of Langerin. Interestingly binding to GAGs, did not simply rely on a net charge effect, but rather on more discrete saccharide features, such as 6-O-sulfation, or iduronic acid content. Using molecular modelling simulations, we proposed a model of Langerin/heparin complex, which located the GAG binding site at the interface of two of the three Carbohydrate-recognition domains of the protein, at the edge of the a-helix coiled-coil. To our knowledge, the binding properties that we have highlighted here for Langerin, have never been reported for C-type lectins before. These findings provide new insights towards the understanding of Langerin biological functions.

  13. Glycosylation in HIV-1 envelope glycoprotein and its biological implications

    KAUST Repository

    Ho, Yung Shwen

    2013-08-01

    Glycosylation of HIV-1 envelope proteins (Env gp120/gp41) plays a vital role in viral evasion from the host immune response, which occurs through the masking of key neutralization epitopes and the presentation of the Env glycosylation as \\'self\\' to the host immune system. Env glycosylation is generally conserved, yet its continual evolution plays an important role in modulating viral infectivity and Env immunogenicity. Thus, it is believed that Env glycosylation, which is a vital part of the HIV-1 architecture, also controls intra- and inter-clade genetic variations. Discerning intra- and inter-clade glycosylation variations could therefore yield important information for understanding the molecular and biological differences between HIV clades and may assist in effectively designing Env-based immunogens and in clearly understanding HIV vaccines. This review provides an in-depth perspective of various aspects of Env glycosylation in the context of HIV-1 pathogenesis. © 2013 Future Medicine Ltd.

  14. HIV-1 envelope, integrins and co-receptor use in mucosal transmission of HIV

    Directory of Open Access Journals (Sweden)

    Cicala Claudia

    2010-01-01

    Full Text Available Abstract It is well established that HIV-1 infection typically involves an interaction between the viral envelope protein gp120/41 and the CD4 molecule followed by a second interaction with a chemokine receptor, usually CCR5 or CXCR4. In the early stages of an HIV-1 infection CCR5 using viruses (R5 viruses predominate. In some viral subtypes there is a propensity to switch to CXCR4 usage (X4 viruses. The receptor switch occurs in ~ 40% of the infected individuals and is associated with faster disease progression. This holds for subtypes B and D, but occurs less frequently in subtypes A and C. There are several hypotheses to explain the preferential transmission of R5 viruses and the mechanisms that lead to switching of co-receptor usage; however, there is no definitive explanation for either. One important consideration regarding transmission is that signaling by R5 gp120 may facilitate transmission of R5 viruses by inducing a permissive environment for HIV replication. In the case of sexual transmission, infection by HIV requires the virus to breach the mucosal barrier to gain access to the immune cell targets that it infects; however, the immediate events that follow HIV exposure at genital mucosal sites are not well understood. Upon transmission, the HIV quasispecies that is replicating in an infected donor contracts through a “genetic bottleneck”, and often infection results from a single infectious event. Many details surrounding this initial infection remain unresolved. In mucosal tissues, CD4+ T cells express high levels of CCR5, and a subset of these CD4+/CCR5high cells express the integrin α4β7, the gut homing receptor. CD4+/CCR5high/ α4β7high T cells are highly susceptible to infection by HIV-1 and are ideal targets for an efficient productive infection at the point of transmission. In this context we have demonstrated that the HIV-1 envelope protein gp120 binds to α4β7 on CD4+ T cells. On CD4+/CCR5high/ α4β7high T cells,

  15. Role of the carbohydrate-binding sites of griffithsin in the prevention of DC-SIGN-mediated capture and transmission of HIV-1.

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    Bart Hoorelbeke

    Full Text Available BACKGROUND: The glycan-targeting C-type DC-SIGN lectin receptor is implicated in the transmission of the human immunodeficiency virus (HIV by binding the virus and transferring the captured HIV-1 to CD4(+ T lymphocytes. Carbohydrate binding agents (CBAs have been reported to block HIV-1 infection. We have now investigated the potent mannose-specific anti-HIV CBA griffithsin (GRFT on its ability to inhibit the capture of HIV-1 to DC-SIGN, its DC-SIGN-directed transmission to CD4(+ T-lymphocytes and the role of the three carbohydrate-binding sites (CBS of GRFT in these processes. FINDINGS: GRFT inhibited HIV-1(IIIB infection of CEM and HIV-1(NL4.3 infection of C8166 CD4(+ T-lymphocytes at an EC50 of 0.059 and 0.444 nM, respectively. The single mutant CBS variants of GRFT (in which a key Asp in one of the CBS was mutated to Ala were about ∼20 to 60-fold less potent to prevent HIV-1 infection and ∼20 to 90-fold less potent to inhibit syncytia formation in co-cultures of persistently HIV-1 infected HuT-78 and uninfected C8166 CD4(+ T-lymphocytes. GRFT prevents DC-SIGN-mediated virus capture and HIV-1 transmission to CD4(+ T-lymphocytes at an EC50 of 1.5 nM and 0.012 nM, respectively. Surface plasmon resonance (SPR studies revealed that wild-type GRFT efficiently blocked the binding between DC-SIGN and immobilized gp120, whereas the point mutant CBS variants of GRFT were ∼10- to 15-fold less efficient. SPR-analysis also demonstrated that wild-type GRFT and its single mutant CBS variants have the capacity to expel bound gp120 from the gp120-DC-SIGN complex in a dose dependent manner, a property that was not observed for HHA, another mannose-specific potent anti-HIV-1 CBA. CONCLUSION: GRFT is inhibitory against HIV gp120 binding to DC-SIGN, efficiently prevents DC-SIGN-mediated transfer of HIV-1 to CD4(+ T-lymphocytes and is able to expel gp120 from the gp120-DC-SIGN complex. Functionally intact CBS of GRFT are important for the optimal action of

  16. HIV-1 Phylogenetic analysis shows HIV-1 transits through the meninges to brain and peripheral tissues

    Science.gov (United States)

    Lamers, Susanna L.; Gray, Rebecca R.; Salemi, Marco; Huysentruyt, Leanne C.; McGrath, Michael

    2010-01-01

    Brain infection by the human immunodeficiency virus type 1 (HIV-1) has been investigated in many reports with a variety of conclusions concerning the time of entry and degree of viral compartmentalization. To address these diverse findings, we sequenced HIV-1 gp120 clones from a wide range of brain, peripheral and meningeal tissues from five patients who died from several HIV-1 associated disease pathologies. High-resolution phylogenetic analysis confirmed previous studies that showed a significant degree of compartmentalization in brain and peripheral tissue subpopulations. Some intermixing between the HIV-1 subpopulations was evident, especially in patients that died from pathologies other than HIV-associated dementia. Interestingly, the major tissue harboring virus from both the brain and peripheral tissues was the meninges. These results show that 1) HIV-1 is clearly capable of migrating out of the brain, 2) the meninges are the most likely primary transport tissues, and 3) infected brain macrophages comprise an important HIV reservoir during highly active antiretroviral therapy. PMID:21055482

  17. Short communication: evaluating the level of expressed HIV type 1 gp120 and gag proteins in the vCP1521 vector by two immunoplaque methods.

    Science.gov (United States)

    Azizi, Ali; Edamura, Kerrie Nichol; Leung, Glenda; Gisonni-Lex, Lucy; Mallet, Laurent

    2013-02-01

    Over the past few years, several recombinant ALVAC constructs have been used as delivery systems in various vaccine research studies and trials. The ALVAC-HIV vCP1521 vector has been used as a vaccine delivery system in the RV144 study, a phase III HIV study that displayed over 31% protective efficacy. One of the important parameters for evaluating the potency of an ALVAC construct is the stable expression of proteins encoded by the inserted genes. Herein, the expression of inserted gp120 and gag genes in two manufactured ALVAC-HIV vCP1521 lots have been determined by two immunoplaque methods (dish and plaque lift). Both methods were specific and robust and demonstrated that the ALVAC-HIV vCP1521 lots were able to express gp120 and gag proteins in over 99% of the infectious plaques.

  18. An HIV-1 envelope immunogen with W427S mutation in CD4 binding site induced more T follicular helper memory cells and reduced non-specific antibody responses.

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    Hao-Tong Yu

    Full Text Available The CD4 binding site (CD4BS of the HIV-1 envelope glycoprotein (Env contains epitopes for broadly neutralizing antibody (nAb and is the target for the vaccine development. However, the CD4BS core including residues 425-430 overlaps the B cell superantigen site and may be related to B cell exhaustion in HIV-1 infection. Furthermore, production of nAb and high-affinity plasma cells needs germinal center reaction and the help of T follicular helper (Tfh cells. We believe that strengthening the ability of Env CD4BS in inducing Tfh response and decreasing the effects of the superantigen are the strategies for eliciting nAb and development of HIV-1 vaccine. We constructed a gp120 mutant W427S of an HIV-1 primary R5 strain and examined its ability in the elicitation of Ab and the production of Tfh by immunization of BALB/c mice. We found that the trimeric wild-type gp120 can induce more non-specific antibody-secreting plasma cells, higher serum IgG secretion, and more Tfh cells by splenocyte. The modified W427S gp120 elicits higher levels of specific binding antibodies as well as nAbs though it produces less Tfh cells. Furthermore, higher Tfh cell frequency does not correlate to the specific binding Abs or nAbs indicating that the wild-type gp120 induced some non-specific Tfh that did not contribute to the production of specific Abs. This gp120 mutant led to more memory Tfh production, especially, the effector memory Tfh cells. Taken together, W427S gp120 could induce higher level of specific binding and neutralizing Ab production that may be associated with the reduction of non-specific Tfh but strengthening of the memory Tfh.

  19. Neutralization of X4- and R5-tropic HIV-1 NL4-3 variants by HOCl-modified serum albumins

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    Schwalbe Birco

    2010-06-01

    Full Text Available Abstract Background Myeloperoxidase (MPO, an important element of the microbicidal activity of neutrophils, generates hypochlorous acid (HOCl from H2O2 and chloride, which is released into body fluids. Besides its direct microbicidal activity, HOCl can react with amino acid residues and HOCl-modified proteins can be detected in vivo. Findings This report is based on binding studies of HOCl-modified serum albumins to HIV-1 gp120 and three different neutralization assays using infectious virus. The binding studies were carried out by surface plasmon resonance spectroscopy and by standard ELISA techniques. Virus neutralization assays were carried out using HIV-1 NL4-3 virus and recombinant strains with CXCR4 and CCR5 coreceptor usage. Viral infection was monitored by a standard p24 or X-gal staining assay. Our data demonstrate that HOCl-modified mouse-, bovine- and human serum albumins all bind to the HIV-1 NL4-3 gp120 (LAV glycoprotein in contrast to non-modified albumin. Binding of HOCl-modified albumin to gp120 correlated to the blockade of CD4 as well as that of V3 loop specific monoclonal antibody binding. In neutralization experiments, HOCl-modified serum albumins inhibited replication and syncytium formation of the X4- and R5-tropic NL4-3 isolates in a dose dependent manner. Conclusions Our data indicate that HOCl-modified serum albumin veils the binding site for CD4 and the V3 loop on gp120. Such masking of the viral gp120/gp41 envelope complex might be a simple but promising strategy to inactivate HIV-1 and therefore prevent infection when HOCl-modified serum albumin is applied, for example, as a topical microbicide.

  20. Targeting cell surface HIV-1 Env protein to suppress infectious virus formation

    Science.gov (United States)

    Bastian, Arangassery Rosemary; Ang, Charles G.; Kamanna, Kantharaju; Shaheen, Farida; Huang, Yu-Hung; McFadden, Karyn; Duffy, Caitlin; Bailey, Lauren D.; Sundaram, Ramalingam Venkat Kalyana; Chaiken, Irwin

    2017-01-01

    HIV-1 Env protein is essential for host cell entry, and targeting Env remains an important antiretroviral strategy. We previously found that a peptide triazole thiol KR13 and its gold nanoparticle conjugate AuNP-KR13 directly and irreversibly inactivate the virus by targeting the Env protein, leading to virus gp120 shedding, membrane disruption and p24 capsid protein release. Here, we examined the consequences of targeting cell-surface Env with the virus inactivators. We found that both agents led to formation of non-infectious virus from transiently transfected 293T cells. The budded non-infectious viruses lacked Env gp120 but contained gp41. Importantly, budded virions also retained the capsid protein p24, in stark contrast to p24 leakage from viruses directly treated by these agents and arguing that the agents led to deformed viruses by transforming the cells at a stage before virus budding. We found that the Env inactivators caused gp120 shedding from the transiently transfected 293T cells as well as non-producer CHO-K1-gp160 cells. Additionally, AuNP-KR13 was cytotoxic against the virus-producing 293T and CHO-K1-gp160 cells, but not untransfected 293T or unmodified CHO-K1 cells. The results obtained reinforce the argument that cell-surface HIV-1 Env is metastable, as on virus particles, and provides a conformationally vulnerable target for virus suppression and infectious cell inactivation. PMID:28390972

  1. HIV-1 protein induced modulation of primary human osteoblast differentiation and function via a Wnt/β-catenin-dependent mechanism.

    LENUS (Irish Health Repository)

    Butler, Joseph S

    2013-02-01

    HIV infection is associated with metabolic bone disease resulting in bone demineralization and reduced bone mass. The molecular mechanisms driving this disease process have yet to be elucidated. Wnt\\/β-catenin signaling plays a key role in bone development and remodeling. We attempted to determine the effects of the HIV-1 protein, gp120, on Wnt\\/β-catenin signaling at an intracellular and transcriptional level in primary human osteoblasts (HOBs). This work, inclusive of experimental controls, was part of a greater project assessing the effects of a variety of different agents on Wnt\\/β-catenin signaling (BMC Musculoskelet Disord 2010;11:210).We examined the phenotypic effects of silencing and overexpressing the Wnt antagonist, Dickkopf-1 (Dkk1) in HOBs treated with gp120. HOBs exposed to gp120 displayed a significant reduction in alkaline phosphatase activity (ALP) activity and cell proliferation and increased cellular apoptosis over a 48 h time course. Immunocytochemistry demonstrated a significant reduction in intracytosolic and intranuclear β-catenin in response to HIV-1 protein exposure. These changes were associated with a reduction of TCF\\/LEF-mediated transcription, the transcriptional outcome of canonical Wnt β-catenin signaling. Silencing Dkk1 expression in HOBs exposed to gp120 resulted in increased ALP activity and cell proliferation, and decreased cellular apoptosis relative to scrambled control. Dkk1 overexpression exacerbated the inhibitory effect of gp120 on HOB function, with decreases in ALP activity and cell proliferation and increased cellular apoptosis relative to vector control. Wnt\\/β-catenin signaling plays a key regulatory role in HIV-associated bone loss, with Dkk1, aputative central mediator in this degenerative process. © 2012 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 31: 218-226, 2013.

  2. Stabilizing exposure of conserved epitopes by structure guided insertion of disulfide bond in HIV-1 envelope glycoprotein.

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    Aemro Kassa

    Full Text Available Entry of HIV-1 into target cells requires binding of the viral envelope glycoprotein (Env to cellular receptors and subsequent conformational changes that culminates in fusion of viral and target cell membranes. Recent structural information has revealed that these conformational transitions are regulated by three conserved but potentially flexible layers stacked between the receptor-binding domain (gp120 and the fusion arm (gp41 of Env. We hypothesized that artificial insertion of a covalent bond will 'snap' Env into a conformation that is less mobile and stably expose conserved sites. Therefore, we analyzed the interface between these gp120 layers (layers 1, 2 and 3 and identified residues that may form disulfide bonds when substituted with cysteines. We subsequently probed the structures of the resultant mutant gp120 proteins by assaying their binding to a variety of ligands using Surface Plasmon Resonance (SPR assay. We found that a single disulfide bond strategically inserted between the highly conserved layers 1 and 2 (C65-C115 is able to 'lock' gp120 in a CD4 receptor bound conformation (in the absence of CD4, as indicated by the lower dissociation constant (Kd for the CD4-induced (CD4i epitope binding 17b antibody. When disulfide-stabilized monomeric (gp120 and trimeric (gp140 Envs were used to immunize rabbits, they were found to elicit a higher proportion of antibodies directed against both CD4i and CD4 binding site epitopes than the wild-type proteins. These results demonstrate that structure-guided stabilization of inter-layer interactions within HIV-1 Env can be used to expose conserved epitopes and potentially overcome the sequence diversity of these molecules.

  3. An autoreactive antibody from an SLE/HIV-1 individual broadly neutralizes HIV-1

    Science.gov (United States)

    Bonsignori, Mattia; Wiehe, Kevin; Grimm, Sebastian K.; Lynch, Rebecca; Yang, Guang; Kozink, Daniel M.; Perrin, Florence; Cooper, Abby J.; Hwang, Kwan-Ki; Chen, Xi; Liu, Mengfei; McKee, Krisha; Parks, Robert J.; Eudailey, Joshua; Wang, Minyue; Clowse, Megan; Criscione-Schreiber, Lisa G.; Moody, M. Anthony; Ackerman, Margaret E.; Boyd, Scott D.; Gao, Feng; Kelsoe, Garnett; Verkoczy, Laurent; Tomaras, Georgia D.; Liao, Hua-Xin; Kepler, Thomas B.; Montefiori, David C.; Mascola, John R.; Haynes, Barton F.

    2014-01-01

    Broadly HIV-1–neutralizing antibodies (BnAbs) display one or more unusual traits, including a long heavy chain complementarity-determining region 3 (HCDR3), polyreactivity, and high levels of somatic mutations. These shared characteristics suggest that BnAb development might be limited by immune tolerance controls. It has been postulated that HIV-1–infected individuals with autoimmune disease and defective immune tolerance mechanisms may produce BnAbs more readily than those without autoimmune diseases. In this study, we identified an HIV-1–infected individual with SLE who exhibited controlled viral load (<5,000 copies/ml) in the absence of controlling HLA phenotypes and developed plasma HIV-1 neutralization breadth. We collected memory B cells from this individual and isolated a BnAb, CH98, that targets the CD4 binding site (CD4bs) of HIV-1 envelope glycoprotein 120 (gp120). CH98 bound to human antigens including dsDNA, which is specifically associated with SLE. Anti-dsDNA reactivity was also present in the patient’s plasma. CH98 had a mutation frequency of 25% and 15% nt somatic mutations in the heavy and light chain variable domains, respectively, a long HCDR3, and a deletion in the light chain CDR1. The occurrence of anti-dsDNA reactivity by a HIV-1 CD4bs BnAb in an individual with SLE raises the possibility that some BnAbs and SLE-associated autoantibodies arise from similar pools of B cells. PMID:24614107

  4. HIV-1 envelope subregion length variation during disease progression.

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    Marcel E Curlin

    2010-12-01

    Full Text Available The V3 loop of the HIV-1 Env protein is the primary determinant of viral coreceptor usage, whereas the V1V2 loop region is thought to influence coreceptor binding and participate in shielding of neutralization-sensitive regions of the Env glycoprotein gp120 from antibody responses. The functional properties and antigenicity of V1V2 are influenced by changes in amino acid sequence, sequence length and patterns of N-linked glycosylation. However, how these polymorphisms relate to HIV pathogenesis is not fully understood. We examined 5185 HIV-1 gp120 nucleotide sequence fragments and clinical data from 154 individuals (152 were infected with HIV-1 Subtype B. Sequences were aligned, translated, manually edited and separated into V1V2, C2, V3, C3, V4, C4 and V5 subregions. V1-V5 and subregion lengths were calculated, and potential N-linked glycosylation sites (PNLGS counted. Loop lengths and PNLGS were examined as a function of time since infection, CD4 count, viral load, and calendar year in cross-sectional and longitudinal analyses. V1V2 length and PNLGS increased significantly through chronic infection before declining in late-stage infection. In cross-sectional analyses, V1V2 length also increased by calendar year between 1984 and 2004 in subjects with early and mid-stage illness. Our observations suggest that there is little selection for loop length at the time of transmission; following infection, HIV-1 adapts to host immune responses through increased V1V2 length and/or addition of carbohydrate moieties at N-linked glycosylation sites. V1V2 shortening during early and late-stage infection may reflect ineffective host immunity. Transmission from donors with chronic illness may have caused the modest increase in V1V2 length observed during the course of the pandemic.

  5. Antibodies Elicited by Multiple Envelope Glycoprotein Immunogens in Primates Neutralize Primary Human Immunodeficiency Viruses (HIV-1) Sensitized by CD4-Mimetic Compounds.

    Science.gov (United States)

    Madani, Navid; Princiotto, Amy M; Easterhoff, David; Bradley, Todd; Luo, Kan; Williams, Wilton B; Liao, Hua-Xin; Moody, M Anthony; Phad, Ganesh E; Vázquez Bernat, Néstor; Melillo, Bruno; Santra, Sampa; Smith, Amos B; Karlsson Hedestam, Gunilla B; Haynes, Barton; Sodroski, Joseph

    2016-05-15

    The human immunodeficiency virus (HIV-1) envelope glycoproteins (Env) mediate virus entry through a series of complex conformational changes triggered by binding to the receptors CD4 and CCR5/CXCR4. Broadly neutralizing antibodies that recognize conserved Env epitopes are thought to be an important component of a protective immune response. However, to date, HIV-1 Env immunogens that elicit broadly neutralizing antibodies have not been identified, creating hurdles for vaccine development. Small-molecule CD4-mimetic compounds engage the CD4-binding pocket on the gp120 exterior Env and induce Env conformations that are highly sensitive to neutralization by antibodies, including antibodies directed against the conserved Env region that interacts with CCR5/CXCR4. Here, we show that CD4-mimetic compounds sensitize primary HIV-1 to neutralization by antibodies that can be elicited in monkeys and humans within 6 months by several Env vaccine candidates, including gp120 monomers. Monoclonal antibodies directed against the gp120 V2 and V3 variable regions were isolated from the immunized monkeys and humans; these monoclonal antibodies neutralized a primary HIV-1 only when the virus was sensitized by a CD4-mimetic compound. Thus, in addition to their direct antiviral effect, CD4-mimetic compounds dramatically enhance the HIV-1-neutralizing activity of antibodies that can be elicited with currently available immunogens. Used as components of microbicides, the CD4-mimetic compounds might increase the protective efficacy of HIV-1 vaccines. Preventing HIV-1 transmission is a high priority for global health. Eliciting antibodies that can neutralize transmitted strains of HIV-1 is difficult, creating problems for the development of an effective vaccine. We found that small-molecule CD4-mimetic compounds sensitize HIV-1 to antibodies that can be elicited in vaccinated humans and monkeys. These results suggest an approach to prevent HIV-1 sexual transmission in which a virus

  6. Functional Antibody Response Against V1V2 and V3 of HIV gp120 in the VAX003 and VAX004 Vaccine Trials.

    Science.gov (United States)

    Balasubramanian, Preetha; Williams, Constance; Shapiro, Mariya B; Sinangil, Faruk; Higgins, Keith; Nádas, Arthur; Totrov, Maxim; Kong, Xiang-Peng; Fiore-Gartland, Andrew J; Haigwood, Nancy L; Zolla-Pazner, Susan; Hioe, Catarina E

    2018-01-11

    Immunization with HIV AIDSVAX gp120 vaccines in the phase III VAX003 and VAX004 trials did not confer protection. To understand the shortcomings in antibody (Ab) responses induced by these vaccines, we evaluated the kinetics of Ab responses to the V1V2 and V3 regions of gp120 and the induction of Ab-mediated antiviral functions during the course of 7 vaccinations over a 30.5-month period. Plasma samples from VAX003 and VAX004 vaccinees and placebo recipients were measured for ELISA-binding Abs and for virus neutralization, Ab-dependent cellular phagocytosis (ADCP), and Ab-dependent cellular cytotoxicity (ADCC). Ab responses to V1V2 and V3 peaked after 3 to 4 immunizations and declined after 5 to 7 immunizations. The deteriorating responses were most evident against epitopes in the underside of the V1V2 β-barrel and in the V3 crown. Correspondingly, vaccinees demonstrated higher neutralization against SF162 pseudovirus sensitive to anti-V1V2 and anti-V3 Abs after 3 or 4 immunizations than after 7 immunizations. Higher levels of ADCP and ADCC were also observed at early or mid-time points as compared with the final time point. Hence, VAX003 and VAX004 vaccinees generated V1V2- and V3-binding Abs and functional Abs after 3 to 4 immunizations, but subsequent boosts did not maintain these responses.

  7. Neutrophils Turn Plasma Proteins into Weapons against HIV-1.

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    Cornelia Speth

    Full Text Available As a consequence of innate immune activation granulocytes and macrophages produce hypochlorite/hypochlorous acid (HOCl via secretion of myeloperoxidase (MPO to the outside of the cells, where HOCl immediately reacts with proteins. Most proteins that become altered by this system do not belong to the invading microorganism but to the host. While there is no doubt that the myeloperoxidase system is capable of directly inactivating HIV-1, we hypothesized that it may have an additional indirect mode of action. We show in this article that HOCl is able to chemically alter proteins and thus turn them into Idea-Ps (Idea-P = immune defence-altered protein, potent amyloid-like and SH-groups capturing antiviral weapons against HIV-1. HOCl-altered plasma proteins (Idea-PP have the capacity to bind efficiently and with high affinity to the HIV-1 envelope protein gp120, and to its receptor CD4 as well as to the protein disulfide isomerase (PDI. Idea-PP was able to inhibit viral infection and replication in a cell culture system as shown by reduced number of infected cells and of syncytia, resulting in reduction of viral capsid protein p24 in the culture supernatant. The unmodified plasma protein fraction had no effect. HOCl-altered isolated proteins antithrombin III and human serum albumin, taken as representative examples of the whole pool of plasma proteins, were both able to exert the same activity of binding to gp120 and inhibition of viral proliferation. These data offer an opportunity to improve the understanding of the intricacies of host-pathogen interactions and allow the generation of the following hypothetical scheme: natural immune defense mechanisms generate by posttranslational modification of plasma proteins a potent virucidal weapon that immobilizes the virus as well as inhibits viral fusion and thus entry into the host cells. Furthermore simulation of this mechanism in vitro might provide an interesting new therapeutic approach against

  8. The current status and challenges in the development of fusion inhibitors as therapeutics for HIV-1 infection.

    Science.gov (United States)

    Tan, Jian Jun; Ma, Xue Ting; Liu, Chang; Zhang, Xiao Yi; Wang, Cun Xin

    2013-01-01

    HIV-1 membrane fusion as a part of the process of viral entry in the target cells is facilitated by gp41 and gp120, which are encoded by Env gene of HIV-1. Based on the structure and the mechanism researches, new treatment options targeting HIV-1 entry process have been proposed. Enfuvirtide, which mimics amino acid sequences of viral envelope glycoprotein gp41, is the first HIV-1 fusion inhibitor approved by FDA. Although it fulfills vital functions by binding to gp41 and abolishing the membrane fusion reaction when used in combination, it could induce drug resistant virus variants. Currently, a number of design and modification schemes have been presented, a large number of prospective fusion peptides have emerged. For these fusion inhibitors, multiple mutations in gp41 have been associated with the loss of susceptibility to agents. This review reported the current developments and innovative designs of HIV-1 membrane fusion inhibitors.

  9. Inhibition of human immunodeficiency virus (HIV) infection in vitro by anticarbohydrate monoclonal antibodies: peripheral glycosylation of HIV envelope glycoprotein gp120 may be a target for virus neutralization

    DEFF Research Database (Denmark)

    Hansen, J E; Clausen, H; Nielsen, C

    1990-01-01

    Carbohydrate structures are often involved in the initial adhesion of pathogens to target cells. In the present study, a panel of anticarbohydrate monoclonal antibodies (MAbs) was tested for their ability to inhibit in vitro human immunodeficiency virus infectivity. MAbs against three different N......- and O-linked carbohydrate epitopes (LeY, A1, and sialyl-Tn) were able to block infection by cell-free virus as well as inhibit syncytium formation. Inhibition of virus infectivity was independent of virus strain (HTLVIIIB or patient isolate SSI-002), the cell line used for virus propagation (H9 or MT4......), and the cell type used as the infection target (MT4, PMC, or selected T4 lymphocytes). Inhibition was observed when viruses were preincubated with MAbs but not when cells were preincubated with MAbs before inoculation, and the MAbs were shown to precipitate 125I-labeled gp120. The MAbs therefore define...

  10. Punica granatum (Pomegranate juice provides an HIV-1 entry inhibitor and candidate topical microbicide

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    Li Yun-Yao

    2004-10-01

    Full Text Available Abstract Background For ≈ 24 years the AIDS pandemic has claimed ≈ 30 million lives, causing ≈ 14,000 new HIV-1 infections daily worldwide in 2003. About 80% of infections occur by heterosexual transmission. In the absence of vaccines, topical microbicides, expected to block virus transmission, offer hope for controlling the pandemic. Antiretroviral chemotherapeutics have decreased AIDS mortality in industrialized countries, but only minimally in developing countries. To prevent an analogous dichotomy, microbicides should be: acceptable; accessible; affordable; and accelerative in transition from development to marketing. Already marketed pharmaceutical excipients or foods, with established safety records and adequate anti-HIV-1 activity, may provide this option. Methods Fruit juices were screened for inhibitory activity against HIV-1 IIIB using CD4 and CXCR4 as cell receptors. The best juice was tested for inhibition of: (1 infection by HIV-1 BaL, utilizing CCR5 as the cellular coreceptor; and (2 binding of gp120 IIIB and gp120 BaL, respectively, to CXCR4 and CCR5. To remove most colored juice components, the adsorption of the effective ingredient(s to dispersible excipients and other foods was investigated. A selected complex was assayed for inhibition of infection by primary HIV-1 isolates. Results HIV-1 entry inhibitors from pomegranate juice adsorb onto corn starch. The resulting complex blocks virus binding to CD4 and CXCR4/CCR5 and inhibits infection by primary virus clades A to G and group O. Conclusion These results suggest the possibility of producing an anti-HIV-1 microbicide from inexpensive, widely available sources, whose safety has been established throughout centuries, provided that its quality is adequately standardized and monitored.

  11. Anti-HIV-1 response elicited in rabbits by anti-idiotype monoclonal antibodies mimicking the CD4-binding site.

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    Roberto Burioni

    Full Text Available Antibodies against conserved epitopes on HIV-1 envelope glycoproteins (Env, such as the gp120 CD4-binding site (CD4bs, could contribute to protection against HIV-1. Env-based immunogens inducing such a response could be a major component of future anti-HIV-1 strategies. In this proof-of-concept study we describe the generation of two anti-idiotype (AI murine antibodies mimicking the CD4bs epitope. Sera were collected from long-term non-progressor patients to obtain CD4bs-directed IgG, through sequential purification steps. The purified IgG were then used as Fab fragments to immunize mice for hybridoma generation. Two hybridomas (P1 and P2, reacting only against the CD4bs-directed IgG, were identified and characterized. The P1 and P2 antibodies were shown to recognize the idiotype of the broadly neutralizing anti-CD4bs human mAb b12. Both P1 and P2 Fabs were able to induce a strong anti-gp120 response in rabbits. Moreover, the rabbits' sera were shown to neutralize two sensitive tier 1 strains of HIV-1 in an Env-pseudotype neutralization assay. In particular, 3/5 rabbits in the P1 group and 1/5 in the P2 group showed greater than 80% neutralizing activity against the HXB2 pseudovirus. Two rabbits also neutralized the pseudovirus HIV-MN. Overall, these data describe the first anti-idiotypic vaccine approach performed to generate antibodies to the CD4bs of the HIV-1 gp120. Although future studies will be necessary to improve strength and breadth of the elicited neutralizing response, this proof-of-concept study documents that immunogens designed on the idiotype of broadly neutralizing Abs are feasible and could help in the design of future anti-HIV strategies.

  12. HIV-1 transgene expression in rats causes oxidant stress and alveolar epithelial barrier dysfunction

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    Jacob Barbara A

    2009-02-01

    Full Text Available Abstract Background HIV-infected individuals are at increased risk for acute and chronic airway disease even though there is no evidence that the virus can infect the lung epithelium. Although HIV-related proteins including gp120 and Tat can directly cause oxidant stress and cellular dysfunction, their effects in the lung are unknown. The goal of this study was to determine the effects of HIV-1 transgene expression in rats on alveolar epithelial barrier function. Alveolar epithelial barrier function was assessed by determining lung liquid clearance in vivo and alveolar epithelial monolayer permeability in vitro. Oxidant stress in the alveolar space was determined by measuring the glutathione redox couple by high performance liquid chromatography, and the expression and membrane localization of key tight junction proteins were assessed. Finally, the direct effects of the HIV-related proteins gp120 and Tat on alveolar epithelial barrier formation and tight junction protein expression were determined. Results HIV-1 transgene expression caused oxidant stress within the alveolar space and impaired epithelial barrier function even though there was no evidence of overt inflammation within the airways. The expression and membrane localization of the tight junction proteins zonula occludens-1 and occludin were decreased in alveolar epithelial cells from HIV-1 transgenic rats. Further, treating alveolar epithelial monolayers from wild type rats in vitro with recombinant gp120 or Tat for 24 hours reproduced many of the effects on zonula occludens-1 and occludin expression and membrane localization. Conclusion Taken together, these data indicate that HIV-related proteins cause oxidant stress and alter the expression of critical tight junction proteins in the alveolar epithelium, resulting in barrier dysfunction.

  13. Effect of HIV-1-related protein expression on cardiac and skeletal muscles from transgenic rats

    Directory of Open Access Journals (Sweden)

    Guidot David M

    2008-04-01

    Full Text Available Abstract Background Human immunodeficiency virus type 1 (HIV-1 infection and the consequent acquired immunodeficiency syndrome (AIDS has protean manifestations, including muscle wasting and cardiomyopathy, which contribute to its high morbidity. The pathogenesis of these myopathies remains partially understood, and may include nutritional deficiencies, biochemical abnormalities, inflammation, and other mechanisms due to viral infection and replication. Growing evidence has suggested that HIV-1-related proteins expressed by the host in response to viral infection, including Tat and gp120, may also be involved in the pathophysiology of AIDS, particularly in cells or tissues that are not directly infected with HIV-1. To explore the potentially independent effects of HIV-1-related proteins on heart and skeletal muscles, we used a transgenic rat model that expresses several HIV-1-related proteins (e.g., Tat, gp120, and Nef. Outcome measures included basic heart and skeletal muscle morphology, glutathione metabolism and oxidative stress, and gene expressions of atrogin-1, muscle ring finger protein-1 (MuRF-1 and Transforming Growth Factor-β1 (TGFβ1, three factors associated with muscle catabolism. Results Consistent with HIV-1 associated myopathies in humans, HIV-1 transgenic rats had increased relative heart masses, decreased relative masses of soleus, plantaris and gastrocnemius muscles, and decreased total and myosin heavy chain type-specific plantaris muscle fiber areas. In both tissues, the levels of cystine (Cyss, the oxidized form of the anti-oxidant cysteine (Cys, and Cyss:Cys ratios were significantly elevated, and cardiac tissue from HIV-1 transgenic rats had altered glutathione metabolism, all reflective of significant oxidative stress. In HIV-1 transgenic rat hearts, MuRF-1 gene expression was increased. Further, HIV-1-related protein expression also increased atrogin-1 (~14- and ~3-fold and TGFβ1 (~5-fold and ~3-fold in heart and

  14. Research on anti-HIV-1 agents. Investigation on the CD4-Suradista binding mode through docking experiments

    Science.gov (United States)

    Manetti, Fabrizio; Corelli, Federico; Mongelli, Nicola; Lombardi Borgia, Andrea; Botta, Maurizio

    2000-05-01

    Sulfonated distamycin (Suradista) derivatives exhibit anti-HIV-1 activity by inhibiting the binding of the viral envelope glycoprotein gp120 to its receptor (CD4). With the aim to propose a possible binding mode between Suradistas and the CD4 macromolecule, molecular docking experiments, followed by energy minimization of the complexes thus obtained, were performed. Computational results show that ligand binding at the CD4 surface involves two or three positively charged regions of the macromolecule, in agreement with the results of X-ray crystallographic analysis of a ternary complex (CD4/gp120/neutralizing antibody) recently reported in the literature. Our findings account well for the structure-activity relationship found for Suradista compounds.

  15. Small CD4 Mimetics Prevent HIV-1 Uninfected Bystander CD4 + T Cell Killing Mediated by Antibody-dependent Cell-mediated Cytotoxicity

    Science.gov (United States)

    Richard, Jonathan; Veillette, Maxime; Ding, Shilei; Zoubchenok, Daria; Alsahafi, Nirmin; Coutu, Mathieu; Brassard, Nathalie; Park, Jongwoo; Courter, Joel R.; Melillo, Bruno; Smith, Amos B.; Shaw, George M.; Hahn, Beatrice H.; Sodroski, Joseph; Kaufmann, Daniel E.; Finzi, Andrés

    2015-01-01

    Human immunodeficiency virus type 1 (HIV-1) infection causes a progressive depletion of CD4 + T cells. Despite its importance for HIV-1 pathogenesis, the precise mechanisms underlying CD4 + T-cell depletion remain incompletely understood. Here we make the surprising observation that antibody-dependent cell-mediated cytotoxicity (ADCC) mediates the death of uninfected bystander CD4 + T cells in cultures of HIV-1-infected cells. While HIV-1-infected cells are protected from ADCC by the action of the viral Vpu and Nef proteins, uninfected bystander CD4 + T cells bind gp120 shed from productively infected cells and are efficiently recognized by ADCC-mediating antibodies. Thus, gp120 shedding represents a viral mechanism to divert ADCC responses towards uninfected bystander CD4 + T cells. Importantly, CD4-mimetic molecules redirect ADCC responses from uninfected bystander cells to HIV-1-infected cells; therefore, CD4-mimetic compounds might have therapeutic utility in new strategies aimed at specifically eliminating HIV-1-infected cells. PMID:26870823

  16. Prediction of exposed domains of envelope glycoprotein in Indian HIV-1 isolates and experimental confirmation of their immunogenicity in humans

    Directory of Open Access Journals (Sweden)

    Mohabatkar H.

    2004-01-01

    Full Text Available We describe the impact of subtype differences on the seroreactivity of linear antigenic epitopes in envelope glycoprotein of HIV-1 isolates from different geographical locations. By computer analysis, we predicted potential antigenic sites of envelope glycoprotein (gp120 and gp4l of this virus. For this purpose, after fetching sequences of proteins of interest from data banks, values of hydrophilicity, flexibility, accessibility, inverted hydrophobicity, and secondary structure were considered. We identified several potential antigenic epitopes in a B subtype strain of envelope glycoprotein of HIV-1 (IIIB. Solid- phase peptide synthesis methods of Merrifield and Fmoc chemistry were used for synthesizing peptides. These synthetic peptides corresponded mainly to the C2, V3 and CD4 binding sites of gp120 and some parts of the ectodomain of gp41. The reactivity of these peptides was tested by ELISA against different HIV-1-positive sera from different locations in India. For two of these predicted epitopes, the corresponding Indian consensus sequences (LAIERYLKQQLLGWG and DIIGDIRQAHCNISEDKWNET (subtype C were also synthesized and their reactivity was tested by ELISA. These peptides also distinguished HIV-1-positive sera of Indians with C subtype infections from sera from HIV-negative subjects.

  17. Analysis of a new mechanism of neutralizing antibodies against HIV-1

    OpenAIRE

    原田, 信志; ハラダ, シンジ; Harada, Shinji

    2009-01-01

    脂質二重膜であるHIV-1エンベロープの流動性を薬物等で低下させると、HIV-1の感染は抑制される。HIV-1gp120に附着しHIV-1を中和する抗体は、エンベロープ流動性を低下させた。また、HIV-1上の非特異的蛋白を認識する抗体では、HIV-1を中和すれば、エンベロープの流動性を低下させた。以上より、HIV-1特異的あるいは非特異的蛋白を認識する抗体は、中和抗体であれば、その附着によりエンベロープの流動性を抑制した。これはウイルスの新しい中和機序であると思われる。...

  18. Frequent intra-subtype recombination among HIV-1 circulating in Tanzania.

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    Ireen E Kiwelu

    Full Text Available The study estimated the prevalence of HIV-1 intra-subtype recombinant variants among female bar and hotel workers in Tanzania. While intra-subtype recombination occurs in HIV-1, it is generally underestimated. HIV-1 env gp120 V1-C5 quasispecies from 45 subjects were generated by single-genome amplification and sequencing (median (IQR of 38 (28-50 sequences per subject. Recombination analysis was performed using seven methods implemented within the recombination detection program version 3, RDP3. HIV-1 sequences were considered recombinant if recombination signals were detected by at least three methods with p-values of ≤0.05 after Bonferroni correction for multiple comparisons. HIV-1 in 38 (84% subjects showed evidence for intra-subtype recombination including 22 with HIV-1 subtype A1, 13 with HIV-1 subtype C, and 3 with HIV-1 subtype D. The distribution of intra-patient recombination breakpoints suggested ongoing recombination and showed selective enrichment of recombinant variants in 23 (60% subjects. The number of subjects with evidence of intra-subtype recombination increased from 29 (69% to 36 (82% over one year of follow-up, although the increase did not reach statistical significance. Adjustment for intra-subtype recombination is important for the analysis of multiplicity of HIV infection. This is the first report of high prevalence of intra-subtype recombination in the HIV/AIDS epidemic in Tanzania, a region where multiple HIV-1 subtypes co-circulate. HIV-1 intra-subtype recombination increases viral diversity and presents additional challenges for HIV-1 vaccine design.

  19. Protein-Protein Interaction between Surfactant Protein D and DC-SIGN via C-Type Lectin Domain Can Suppress HIV-1 Transfer.

    Science.gov (United States)

    Dodagatta-Marri, Eswari; Mitchell, Daniel A; Pandit, Hrishikesh; Sonawani, Archana; Murugaiah, Valarmathy; Idicula-Thomas, Susan; Nal, Béatrice; Al-Mozaini, Maha M; Kaur, Anuvinder; Madan, Taruna; Kishore, Uday

    2017-01-01

    Surfactant protein D (SP-D) is a soluble C-type lectin, belonging to the collectin (collagen-containing calcium-dependent lectin) family, which acts as an innate immune pattern recognition molecule in the lungs at other mucosal surfaces. Immune regulation and surfactant homeostasis are salient functions of SP-D. SP-D can bind to a range of viral, bacterial, and fungal pathogens and trigger clearance mechanisms. SP-D binds to gp120, the envelope protein expressed on HIV-1, through its C-type lectin or carbohydrate recognition domain. This is of importance since SP-D is secreted by human mucosal epithelial cells and is present in the female reproductive tract, including vagina. Another C-type lectin, dendritic cell (DC)-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN), present on the surface of the DCs, also binds to HIV-1 gp120 and facilitates viral transfer to the lymphoid tissues. DCs are also present at the site of HIV-1 entry, embedded in vaginal or rectal mucosa. In the present study, we report a direct protein-protein interaction between recombinant forms of SP-D (rfhSP-D) and DC-SIGN via their C-type lectin domains. Both SP-D and DC-SIGN competed for binding to immobilized HIV-1 gp120. Pre-incubation of human embryonic kidney cells expressing surface DC-SIGN with rfhSP-D significantly inhibited the HIV-1 transfer to activated peripheral blood mononuclear cells. In silico analysis revealed that SP-D and gp120 may occupy same sites on DC-SIGN, which may explain the reduced transfer of HIV-1. In summary, we demonstrate, for the first time, that DC-SIGN is a novel binding partner of SP-D, and this interaction can modulate HIV-1 capture and transfer to CD4+ T cells. In addition, the present study also reveals a novel and distinct mechanism of host defense by SP-D against HIV-1.

  20. A novel trifunctional IgG-like bispecific antibody to inhibit HIV-1 infection and enhance lysis of HIV by targeting activation of complement

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    Tomlinson Stephen

    2010-06-01

    Full Text Available Abstract Background The complement system is not only a key component of innate immunity but also provides a first line of defense against invading pathogens, especially for viral pathogens. Human immunodeficiency virus (HIV, however, possesses several mechanisms to evade complement-mediated lysis (CoML and exploit the complement system to enhance viral infectivity. Responsible for this intrinsic resistance against complement-mediated virolysis are complement regulatory membrane proteins derived from the host cell that inherently downregulates complement activation at several stages of the cascade. In addition, HIV is protected from complement-mediated lysis by binding soluble factor H (fH through the viral envelope proteins, gp120 and gp41. Whereas inhibition of complement activity is the desired outcome in the vast majority of therapeutic approaches, there is a broader potential for complement-mediated inhibition of HIV by complement local stimulation. Presentation of the hypothesis Our previous studies have proven that the complement-mediated antibody-dependent enhancement of HIV infection is mediated by the association of complement receptor type 2 bound to the C3 fragment and deposited on the surface of HIV virions. Thus, we hypothesize that another new activator of complement, consisting of two dsFv (against gp120 and against C3d respectively linked to a complement-activating human IgG1 Fc domain ((anti-gp120 × anti-C3d-Fc, can not only target and amplify complement activation on HIV virions for enhancing the efficiency of HIV lysis, but also reduce the infectivity of HIV through blocking the gp120 and C3d on the surface of HIV. Testing the hypothesis Our hypothesis was tested using cell-free HIV-1 virions cultivated in vitro and assessment of virus opsonization was performed by incubating appropriate dilutions of virus with medium containing normal human serum and purified (anti-gp120 × anti-C3d-Fc proteins. As a control group, viruses

  1. Serotyping and genotyping of HIV-1 infection in residents of Khayelitsha, Cape Town, South Africa.

    Science.gov (United States)

    Jacobs, G B; de Beer, C; Fincham, J E; Adams, V; Dhansay, M A; van Rensburg, E Janse; Engelbrecht, S

    2006-12-01

    It is estimated that between 5.5 and 6.1 million people are infected with HIV/acquired immunodeficiency syndrome (AIDS) in South Africa, with subtype C responsible for the majority of these infections. The Khayelitsha suburb of Cape Town has one of the highest HIV prevalence rates in South Africa. Overcrowding combined with unemployment and crime in parts of the area perpetuates high-risk sexual behavior, which increases exposure to infection by HIV. Against this background, the objective of this study was to characterize HIV-1 in residents confirmed to be seropositive. Serotyping was performed through a competitive enzyme-linked immunosorbent assay (cPEIA). Genotyping methods included RNA isolation followed by RT-PCR and sequencing of the gag p24, env gp41 immunodominant region (IDR), and env gp120 V3 genome regions of HIV-1. With the exception of a possible C/D recombinant strain, all HIV-1 strains were characterized as HIV-1 group M subtype C. One individual was shown to harbor multiple strains of HIV-1 subtype C. In Southern Africa, the focus has been to develop a subtype C candidate vaccine, as this is the major subtype found in this geographical area. Therefore, the spread of HIV-1 and its recombinant strains needs to be monitored closely. (c) 2006 Wiley-Liss, Inc.

  2. Diversion of HIV-1 Vaccine-induced Immunity by gp41-Microbiota Cross-reactive Antibodies

    Science.gov (United States)

    Williams, Wilton B; Liao, Hua-Xin; Moody, M. Anthony; Kepler, Thomas B.; Alam, S Munir; Gao, Feng; Wiehe, Kevin; Trama, Ashley M.; Jones, Kathryn; Zhang, Ruijun; Song, Hongshuo; Marshall, Dawn J; Whitesides, John F; Sawatzki, Kaitlin; Hua, Axin; Liu, Pinghuang; Tay, Matthew Z; Seaton, Kelly; Shen, Xiaoying; Foulger, Andrew; Lloyd, Krissey E.; Parks, Robert; Pollara, Justin; Ferrari, Guido; Yu, Jae-Sung; Vandergrift, Nathan; Montefiori, David C.; Sobieszczyk, Magdalena E; Hammer, Scott; Karuna, Shelly; Gilbert, Peter; Grove, Doug; Grunenberg, Nicole; McElrath, Julie; Mascola, John R.; Koup, Richard A; Corey, Lawrence; Nabel, Gary J.; Morgan, Cecilia; Churchyard, Gavin; Maenza, Janine; Keefer, Michael; Graham, Barney S.; Baden, Lindsey R.; Tomaras, Georgia D.; Haynes, Barton F.

    2015-01-01

    A HIV-1 DNA prime-recombinant Adenovirus Type 5 (rAd5) boost vaccine failed to protect from HIV-1 acquisition. We studied the nature of the vaccine-induced antibody (Ab) response to HIV-1 envelope (Env). HIV-1-reactive plasma Ab titers were higher to Env gp41 than gp120, and repertoire analysis demonstrated that 93% of HIV-1-reactive Abs from memory B cells was to Env gp41. Vaccine-induced gp41-reactive monoclonal antibodies (mAbs) were non-neutralizing, and frequently polyreactive with host and environmental antigens including intestinal microbiota (IM). Next generation sequencing of an IGHV repertoire prior to vaccination revealed an Env-IM cross-reactive Ab that was clonally-related to a subsequent vaccine-induced gp41-reactive Ab. Thus, HIV-1 Env DNA-rAd5 vaccine induced a dominant IM-polyreactive, non-neutralizing gp41-reactive Ab repertoire response that was associated with no vaccine efficacy. PMID:26229114

  3. Six Highly Conserved Targets of RNAi Revealed in HIV-1-Infected Patients from Russia Are Also Present in Many HIV-1 Strains Worldwide

    Directory of Open Access Journals (Sweden)

    Olga V. Kretova

    2017-09-01

    Full Text Available RNAi has been suggested for use in gene therapy of HIV/AIDS, but the main problem is that HIV-1 is highly variable and could escape attack from the small interfering RNAs (siRNAs due to even single nucleotide substitutions in the potential targets. To exhaustively check the variability in selected RNA targets of HIV-1, we used ultra-deep sequencing of six regions of HIV-1 from the plasma of two independent cohorts of patients from Russia. Six RNAi targets were found that are invariable in 82%–97% of viruses in both cohorts and are located inside the domains specifying reverse transcriptase (RT, integrase, vpu, gp120, and p17. The analysis of mutation frequencies and their characteristics inside the targets suggests a likely role for APOBEC3G (apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like 3G, A3G in G-to-A mutations and a predominant effect of RT biases in the detected variability of the virus. The lowest frequency of mutations was detected in the central part of all six targets. We also discovered that the identical RNAi targets are present in many HIV-1 strains from many countries and from all continents. The data are important for both the understanding of the patterns of HIV-1 mutability and properties of RT and for the development of gene therapy approaches using RNAi for the treatment of HIV/AIDS.

  4. Lignosulfonic acid exhibits broadly anti-HIV-1 activity--potential as a microbicide candidate for the prevention of HIV-1 sexual transmission.

    Directory of Open Access Journals (Sweden)

    Min Qiu

    Full Text Available Some secondary metabolites from plants show to have potent inhibitory activities against microbial pathogens, such as human immunodeficiency virus (HIV, herpes simplex virus (HSV, Treponema pallidum, Neisseria gonorrhoeae, etc. Here we report that lignosulfonic acid (LSA, a polymeric lignin derivative, exhibits potent and broad activity against HIV-1 isolates of diverse subtypes including two North America strains and a number of Chinese clinical isolates values ranging from 21.4 to 633 nM. Distinct from other polyanions, LSA functions as an entry inhibitor with multiple targets on viral gp120 as well as on host receptor CD4 and co-receptors CCR5/CXCR4. LSA blocks viral entry as determined by time-of-drug addiction and cell-cell fusion assays. Moreover, LSA inhibits CD4-gp120 interaction by blocking the binding of antibodies specific for CD4-binding sites (CD4bs and for the V3 loop of gp120. Similarly, LSA interacts with CCR5 and CXCR4 via its inhibition of specific anti-CCR5 and anti-CXCR4 antibodies, respectively. Interestingly, the combination of LSA with AZT and Nevirapine exhibits synergism in viral inhibition. For the purpose of microbicide development, LSA displays low in vitro cytotoxicity to human genital tract epithelial cells, does not stimulate NF-κB activation and has no significant up-regulation of IL-1α/β and IL-8 as compared with N-9. Lastly, LSA shows no adverse effect on the epithelial integrity and the junctional protein expression. Taken together, our findings suggest that LSA can be a potential candidate for tropical microbicide.

  5. Interaction between Tat and Drugs of Abuse during HIV-1 Infection and Central Nervous System Disease.

    Science.gov (United States)

    Maubert, Monique E; Pirrone, Vanessa; Rivera, Nina T; Wigdahl, Brian; Nonnemacher, Michael R

    2015-01-01

    In many individuals, drug abuse is intimately linked with HIV-1 infection. In addition to being associated with one-third of all HIV-1 infections in the United States, drug abuse also plays a role in disease progression and severity in HIV-1-infected patients, including adverse effects on the central nervous system (CNS). Specific systems within the brain are known to be damaged in HIV-1-infected individuals and this damage is similar to that observed in drug abuse. Even in the era of anti-retroviral therapy (ART), CNS pathogenesis occurs with HIV-1 infection, with a broad range of cognitive impairment observed, collectively referred to as HIV-1-associated neurocognitive disorders (HAND). A number of HIV-1 proteins (Tat, gp120, Nef, Vpr) have been implicated in the etiology of pathogenesis and disease as a result of the biologic activity of the extracellular form of each of the proteins in a number of tissues, including the CNS, even in ART-suppressed patients. In this review, we have made Tat the center of attention for a number of reasons. First, it has been shown to be synthesized and secreted by HIV-1-infected cells in the CNS, despite the most effective suppression therapies available to date. Second, Tat has been shown to alter the functions of several host factors, disrupting the molecular and biochemical balance of numerous pathways contributing to cellular toxicity, dysfunction, and death. In addition, the advantages and disadvantages of ART suppression with regard to controlling the genesis and progression of neurocognitive impairment are currently under debate in the field and are yet to be fully determined. In this review, we discuss the individual and concerted contributions of HIV-1 Tat, drug abuse, and ART with respect to damage in the CNS, and how these factors contribute to the development of HAND in HIV-1-infected patients.

  6. Anti-HIV-1 ADCC Antibodies following Latency Reversal and Treatment Interruption.

    Science.gov (United States)

    Lee, Wen Shi; Kristensen, Anne B; Rasmussen, Thomas A; Tolstrup, Martin; Østergaard, Lars; Søgaard, Ole S; Wines, Bruce D; Hogarth, P Mark; Reynaldi, Arnold; Davenport, Miles P; Emery, Sean; Amin, Janaki; Cooper, David A; Kan, Virginia L; Fox, Julie; Gruell, Henning; Parsons, Matthew S; Kent, Stephen J

    2017-08-01

    There is growing interest in utilizing antibody-dependent cellular cytotoxicity (ADCC) to eliminate infected cells following reactivation from HIV-1 latency. A potential barrier is that HIV-1-specific ADCC antibodies decline in patients on long-term antiretroviral therapy (ART) and may not be sufficient to eliminate reactivated latently infected cells. It is not known whether reactivation from latency with latency-reversing agents (LRAs) could provide sufficient antigenic stimulus to boost HIV-1-specific ADCC. We found that treatment with the LRA panobinostat or a short analytical treatment interruption (ATI), 21 to 59 days, was not sufficient to stimulate an increase in ADCC-competent antibodies, despite viral rebound in all subjects who underwent the short ATI. In contrast, a longer ATI, 2 to 12 months, among subjects enrolled in the Strategies for Management of Antiretroviral Therapy (SMART) trial robustly boosted HIV-1 gp120-specific Fc receptor-binding antibodies and ADCC against HIV-1-infected cells in vitro These results show that there is a lag between viral recrudescence and the boosting of ADCC antibodies, which has implications for strategies toward eliminating latently infected cells.IMPORTANCE The "shock and kill" HIV-1 cure strategy aims to reactivate HIV-1 expression in latently infected cells and subsequently eliminate the reactivated cells through immune-mediated killing. Several latency reversing agents (LRAs) have been examined in vivo, but LRAs alone have not been able to achieve HIV-1 remission and prevent viral rebound following analytical treatment interruption (ATI). In this study, we examined whether LRA treatment or ATI can provide sufficient antigenic stimulus to boost HIV-1-specific functional antibodies that can eliminate HIV-1-infected cells. Our study has implications for the antigenic stimulus required for antilatency strategies and/or therapeutic vaccines to boost functional antibodies and assist in eliminating the latent reservoir

  7. Interaction between Tat and Drugs of Abuse during HIV-1 Infection and Central Nervous System Disease

    Directory of Open Access Journals (Sweden)

    Monique E Maubert

    2016-01-01

    Full Text Available In many individuals, drug abuse is intimately linked with HIV-1 infection. In addition to being associated with one-third of all HIV-1 infections in the United States, drug abuse also plays a role in disease progression and severity in HIV-1-infected patients, including adverse effects on the central nervous system (CNS. Specific systems within the brain are known to be damaged in HIV-1-infected individuals and this damage is similar to that observed in drug abuse. Even in the era of anti-retroviral therapy (ART, CNS pathogenesis occurs with HIV-1 infection, with a broad range of cognitive impairment observed, collectively referred to as HIV-1-associated neurocognitive disorders (HAND. A number of HIV-1 proteins (Tat, gp120, Nef, Vpr have been implicated in the etiology of pathogenesis and disease as a result of the biologic activity of the extracellular form of each of the proteins in a number of tissues, including the CNS, even in ART-suppressed patients. In this review, we have made Tat the center of attention for a number of reasons. First, it has been shown to be synthesized and secreted by HIV-1-infected cells in the CNS, despite the most effective suppression therapies available to date. Second, Tat has been shown to alter the functions of several host factors, disrupting the molecular and biochemical balance of numerous pathways contributing to cellular toxicity, dysfunction, and death. In addition, the advantages and disadvantages of ART suppression with regard to controlling the genesis and progression of neurocognitive impairment are currently under debate in the field and are yet to be fully determined. In this review, we discuss the individual and concerted contributions of HIV-1 Tat, drug abuse, and ART with respect to damage in the CNS, and how these factors contribute to the development of HAND in HIV-1-infected patients.

  8. HIV-1 Tat Protein Enhances Expression and Function of Breast Cancer Resistance Protein.

    Science.gov (United States)

    Zhou, Yancong; Zhang, Kun; Yin, Xiaojie; Nie, Qichang; Ma, Yonggang

    2016-01-01

    ATP binding cassette (ABC) transporters can transfer a variety of antiviral agents from the cytoplasm to body fluid, which results in a reduced intracellular concentration of the drugs. Proteins of HIV-1, e.g., Tat and gp120, altered some types of ABC transporter expression in brain microvascular endothelial cells and astrocytes. However, the effect of Tat on ABC transporters in T lymphocytes is unclear. In this study the status of breast cancer resistance protein (BCRP) in Tat expressing cell lines was examined with real-time PCR and flow cytometry. It was found that HIV-1 Tat protein upregulated BCRP expression and enhanced efflux mediated by BCRP significantly, which could inhibit antiviral drugs from entering infected cells and interfere with the therapeutic effect of HAART.

  9. Reconstitution and characterization of antibody repertoires of HIV-1-infected "elite neutralizers".

    Science.gov (United States)

    Sun, Zehua; Li, Jingjing; Hu, Xintao; Shao, Yiming; Zhang, Mei-Yun

    2015-06-01

    Around 3-5% HIV-1-infected individuals develop high titers of broadly neutralizing HIV-1 antibodies (bnAbs) during chronic infection. However, monoclonal antibodies (mAbs) isolated from such "elite neutralizers" do not, in most cases, depict the serum IgGs in neutralizing the virus. We hypothesize that HIV-1-specific antibodies in infected subjects may work in a population manner in containing the virus in vivo, and in vitro reconstituted antibody repertoires of "elite neutralizers" may mimic the sera in binding and neutralizing the virus. This study aims to investigate the antibody repertoires of three such "elite neutralizers" by reconstituting the immune antibody repertories in vitro followed by a comparative study of the recombinant library IgGs with the corresponding serum IgGs. We found that the recombinant library IgGs were much weaker than the serum IgGs in binding to envelope glycoproteins (Envs) and in neutralizing the virus and inhibiting Env-mediated cell-cell fusion. However, the sorted libraries composing of HIV-1-specific neutralizing antibodies (nAbs) in the three recombinant libraries exhibited comparable binding and inhibitory activities, as well as antibody-dependent cell-mediated cytotoxicity (ADCC), to the serum IgGs. The sorted library IgGs further showed neutralization profiles which are similar to those of the serum IgGs, but they were overall less potent than the serum IgGs. The sorted library IgGs and the serum IgGs bound weakly to the resurfaced Env gp120, RSC3, and did not bind to the CD4 binding site (CD4bs) knock-out mutant, ΔRSC3. Profiling with VRC01 binding site knock-out site mutants of gp120BaL indicates that, if there are any CD4bs bnAbs in these sera, they are more likely b12-like, but not VRC01-class bnAbs. Our results suggest that HIV-1-specific Ab-expressing B cells, especially potent nAb-expressing B cells may not be rich in the B cell repertoires of "elite neutralizers", but they may be highly active in producing nAbs in

  10. A New Glycan-Dependent CD4-Binding Site Neutralizing Antibody Exerts Pressure on HIV-1 In Vivo

    OpenAIRE

    Freund, Natalia?T.; Horwitz, Joshua A.; Lilian Nogueira; Sievers, Stuart A.; Louise Scharf; Scheid, Johannes F.; Anna Gazumyan; Cassie Liu; Klara Velinzon; Ariel Goldenthal; Sanders, Rogier W.; Moore, John P.; Bjorkman, Pamela J.; Seaman, Michael S.; Walker, Bruce D.

    2015-01-01

    The CD4 binding site (CD4bs) on the envelope glycoprotein is a major site of vulnerability that is conserved among different HIV-1 isolates. Many broadly neutralizing antibodies (bNAbs) to the CD4bs belong to the VRC01 class, sharing highly restricted origins, recognition mechanisms and viral escape pathways. We sought to isolate new anti-CD4bs bNAbs with different origins and mechanisms of action. Using a gp120 2CC core as bait, we isolated antibodies encoded by IGVH3-21 and IGVL3-1 genes wi...

  11. Impact of HIV-1 Membrane Cholesterol on Cell-Independent Lytic Inactivation and Cellular Infectivity.

    Science.gov (United States)

    Kalyana Sundaram, Ramalingam Venkat; Li, Huiyuan; Bailey, Lauren; Rashad, Adel A; Aneja, Rachna; Weiss, Karl; Huynh, James; Bastian, Arangaserry Rosemary; Papazoglou, Elisabeth; Abrams, Cameron; Wrenn, Steven; Chaiken, Irwin

    2016-01-26

    Peptide triazole thiols (PTTs) have been found previously to bind to HIV-1 Env spike gp120 and cause irreversible virus inactivation by shedding gp120 and lytically releasing luminal capsid protein p24. Since the virions remain visually intact, lysis appears to occur via limited membrane destabilization. To better understand the PTT-triggered membrane transformation involved, we investigated the role of envelope cholesterol on p24 release by measuring the effect of cholesterol depletion using methyl beta-cyclodextrin (MβCD). An unexpected bell-shaped response of PTT-induced lysis to [MβCD] was observed, involving lysis enhancement at low [MβCD] vs loss of function at high [MβCD]. The impact of cholesterol depletion on PTT-induced lysis was reversed by adding exogenous cholesterol and other sterols that support membrane rafts, while sterols that do not support rafts induced only limited reversal. Cholesterol depletion appears to cause a reduced energy barrier to lysis as judged by decreased temperature dependence with MβCD. Enhancement/replenishment responses to [MβCD] also were observed for HIV-1 infectivity, consistent with a similar energy barrier effect in the membrane transformation of virus cell fusion. Overall, the results argue that cholesterol in the HIV-1 envelope is important for balancing virus stability and membrane transformation, and that partial depletion, while increasing infectivity, also makes the virus more fragile. The results also reinforce the argument that the lytic inactivation and infectivity processes are mechanistically related and that membrane transformations occurring during lysis can provide an experimental window to investigate membrane and protein factors important for HIV-1 cell entry.

  12. Rhesus macaque and chimpanzee DC-SIGN act as HIV/SIV gp120 trans-receptors, similar to human DC-SIGN.

    NARCIS (Netherlands)

    Geijtenbeek, T.B.; Koopman, G.; Duijnhoven, G.C.F. van; Vliet, S.J. van; Schijndel, J.C.H.W. van; Engering, A.J.; Heeney, J.L.; Kooyk, Y. van

    2001-01-01

    Dendritic cells (DC) have been implicated in the pathogenesis of both human and simian immunodeficiency viruses (HIV and SIV, respectively). The DC-specific HIV-1 trans-receptor DC-SIGN is thought to be essential for viral dissemination by DC. Abundant expression in lymphoid tissues also implies a

  13. Sialoadhesin expressed on IFN-induced monocytes binds HIV-1 and enhances infectivity.

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    Hans Rempel

    Full Text Available BACKGROUND: HIV-1 infection dysregulates the immune system and alters gene expression in circulating monocytes. Differential gene expression analysis of CD14(+ monocytes from subjects infected with HIV-1 revealed increased expression of sialoadhesin (Sn, CD169, Siglec 1, a cell adhesion molecule first described in a subset of macrophages activated in chronic inflammatory diseases. METHODOLOGY/PRINCIPAL FINDINGS: We analyzed sialoadhesin expression on CD14(+ monocytes by flow cytometry and found significantly higher expression in subjects with elevated viral loads compared to subjects with undetectable viral loads. In cultured CD14(+ monocytes isolated from healthy individuals, sialoadhesin expression was induced by interferon-alpha and interferon-gamma but not tumor necrosis factor-alpha. Using a stringent binding assay, sialoadhesin-expressing monocytes adsorbed HIV-1 through interaction with the sialic acid residues on the viral envelope glycoprotein gp120. Furthermore, monocytes expressing sialoadhesin facilitated HIV-1 trans infection of permissive cells, which occurred in the absence of monocyte self-infection. CONCLUSIONS/SIGNIFICANCE: Increased sialoadhesin expression on CD14(+ monocytes occurred in response to HIV-1 infection with maximum expression associated with high viral load. We show that interferons induce sialoadhesin in primary CD14(+ monocytes, which is consistent with an antiviral response during viremia. Our findings suggest that circulating sialoadhesin-expressing monocytes are capable of binding HIV-1 and effectively delivering virus to target cells thereby enhancing the distribution of HIV-1. Sialoadhesin could disseminate HIV-1 to viral reservoirs during monocyte immunosurveillance or migration to sites of inflammation and then facilitate HIV-1 infection of permissive cells.

  14. Anti-HIV-1 activity of anionic polymers: a comparative study of candidate microbicides

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    Li Yun-Yao

    2002-11-01

    Full Text Available Abstract Background Cellulose acetate phthalate (CAP in soluble form blocks coreceptor binding sites on the virus envelope glycoprotein gp120 and elicits gp41 six-helix bundle formation, processes involved in virus inactivation. CAP is not soluble at pH Methods Enzyme linked immunosorbent assays (ELISA were used to (1 study HIV-1 IIIB and BaL binding to micronized CAP; (2 detect virus disintegration; and (3 measure gp41 six-helix bundle formation. Cells containing integrated HIV-1 LTR linked to the β-gal gene and expressing CD4 and coreceptors CXCR4 or CCR5 were used to measure virus infectivity. Results 1 HIV-1 IIIB and BaL, respectively, effectively bound to micronized CAP. 2 The interaction between HIV-1 and micronized CAP led to: (a gp41 six-helix bundle formation; (b virus disintegration and shedding of envelope glycoproteins; and (c rapid loss of infectivity. Polymers other than CAP, except Carbomer 974P, elicited gp41 six-helix bundle formation in HIV-1 IIIB but only poly(napthalene sulfonate, in addition to CAP, had this effect on HIV-1 BaL. These polymers differed with respect to their virucidal activities, the differences being more pronounced for HIV-1 BaL. Conclusions Micronized CAP is the only candidate topical microbicide with the capacity to remove rapidly by adsorption from physiological fluids HIV-1 of both the X4 and R5 biotypes and is likely to prevent virus contact with target cells. The interaction between micronized CAP and HIV-1 leads to rapid virus inactivation. Among other anionic polymers, cellulose sulfate, BufferGel and aryl sulfonates appear most effective in this respect.

  15. Psychoneuroimmunology and HIV-1.

    Science.gov (United States)

    Antoni, Michael H.; And Others

    1990-01-01

    Presents evidence describing benefits of behavioral interventions such as aerobic exercise training on both psychological and immunological functioning among high risk human immunodeficiency virus-Type 1 (HIV-1) seronegative and very early stage seropositive homosexual men. HIV-1 infection is cast as chronic disease for which early…

  16. HIV-1 envelope glycoprotein

    Energy Technology Data Exchange (ETDEWEB)

    Caulfield, Michael; Cupo, Albert; Dean, Hansi; Hoffenberg, Simon; King, C. Richter; Klasse, P. J.; Marozsan, Andre; Moore, John P.; Sanders, Rogier W.; Ward, Andrew; Wilson, Ian; Julien, Jean-Philippe

    2017-08-22

    The present application relates to novel HIV-1 envelope glycoproteins, which may be utilized as HIV-1 vaccine immunogens, and antigens for crystallization, electron microscopy and other biophysical, biochemical and immunological studies for the identification of broad neutralizing antibodies. The present invention encompasses the preparation and purification of immunogenic compositions, which are formulated into the vaccines of the present invention.

  17. Inhibition of HIV-1 entry by extracts derived from traditional Chinese medicinal herbal plants

    Directory of Open Access Journals (Sweden)

    Song Xinming

    2009-08-01

    HIV-1 interaction with target cells, i.e., the interaction between gp120 and CD4/CCR5 or gp120 and CD4/CXCR4 and point to the potential of developing these two extracts to be HIV-1 entry inhibitors.

  18. Co-evolution of a broadly neutralizing HIV-1 antibody and founder virus

    Science.gov (United States)

    Liao, Hua-Xin; Lynch, Rebecca; Zhou, Tongqing; Gao, Feng; Alam, S. Munir; Boyd, Scott D.; Fire, Andrew Z.; Roskin, Krishna M.; Schramm, Chaim A.; Zhang, Zhenhai; Zhu, Jiang; Shapiro, Lawrence; Mullikin, James C.; Gnanakaran, S.; Hraber, Peter; Wiehe, Kevin; Kelsoe, Garnett; Yang, Guang; Xia, Shi-Mao; Montefiori, David C.; Parks, Robert; Lloyd, Krissey E.; Scearce, Richard M.; Soderberg, Kelly A.; Cohen, Myron; Kaminga, Gift; Louder, Mark K.; Tran, Lillan M.; Chen, Yue; Cai, Fangping; Chen, Sheri; Moquin, Stephanie; Du, Xiulian; Joyce, Gordon M.; Srivatsan, Sanjay; Zhang, Baoshan; Zheng, Anqi; Shaw, George M.; Hahn, Beatrice H.; Kepler, Thomas B.; Korber, Bette T.M.; Kwong, Peter D.; Mascola, John R.; Haynes, Barton F.

    2013-01-01

    Current HIV-1 vaccines elicit strain-specific neutralizing antibodies. However, cross-reactive neutralizing antibodies arise in ~20% of HIV-1-infected individuals, and details of their generation could provide a roadmap for effective vaccination. Here we report the isolation, evolution and structure of a broadly neutralizing antibody from an African donor followed from time of infection. The mature antibody, CH103, neutralized ~55% of HIV-1 isolates, and its co-crystal structure with gp120 revealed a novel loop-based mechanism of CD4-binding site recognition. Virus and antibody gene sequencing revealed concomitant virus evolution and antibody maturation. Notably, the CH103-lineage unmutated common ancestor avidly bound the transmitted/founder HIV-1 envelope glycoprotein, and evolution of antibody neutralization breadth was preceded by extensive viral diversification in and near the CH103 epitope. These data elucidate the viral and antibody evolution leading to induction of a lineage of HIV-1 broadly neutralizing antibodies and provide insights into strategies to elicit similar antibodies via vaccination. PMID:23552890

  19. Different pattern of immunoglobulin gene usage by HIV-1 compared to non-HIV-1 antibodies derived from the same infected subject.

    Science.gov (United States)

    Li, Liuzhe; Wang, Xiao-Hong; Banerjee, Sagarika; Volsky, Barbara; Williams, Constance; Virland, Diana; Nadas, Arthur; Seaman, Michael S; Chen, Xuemin; Spearman, Paul; Zolla-Pazner, Susan; Gorny, Miroslaw K

    2012-01-01

    A biased usage of immunoglobulin (Ig) genes is observed in human anti-HIV-1 monoclonal antibodies (mAbs) resulting probably from compensation to reduced usage of the VH3 family genes, while the other alternative suggests that this bias usage is due to antigen requirements. If the antigen structure is responsible for the preferential usage of particular Ig genes, it may have certain implications for HIV vaccine development by the targeting of particular Ig gene-encoded B cell receptors to induce neutralizing anti-HIV-1 antibodies. To address this issue, we have produced HIV-1 specific and non-HIV-1 mAbs from an infected individual and analyzed the Ig gene usage. Green-fluorescence labeled virus-like particles (VLP) expressing HIV-1 envelope (Env) proteins of JRFL and BaL and control VLPs (without Env) were used to select single B cells for the production of 68 recombinant mAbs. Ten of these mAbs were HIV-1 Env specific with neutralizing activity against V3 and the CD4 binding site, as well as non-neutralizing mAbs to gp41. The remaining 58 mAbs were non-HIV-1 Env mAbs with undefined specificities. Analysis revealed that biased usage of Ig genes was restricted only to anti-HIV-1 but not to non-HIV-1 mAbs. The VH1 family genes were dominantly used, followed by VH3, VH4, and VH5 among anti-HIV-1 mAbs, while non-HIV-1 specific mAbs preferentially used VH3 family genes, followed by VH4, VH1 and VH5 families in a pattern identical to Abs derived from healthy individuals. This observation suggests that the biased usage of Ig genes by anti-HIV-1 mAbs is driven by structural requirements of the virus antigens rather than by compensation to any depletion of VH3 B cells due to autoreactive mechanisms, according to the gp120 superantigen hypothesis.

  20. Different pattern of immunoglobulin gene usage by HIV-1 compared to non-HIV-1 antibodies derived from the same infected subject.

    Directory of Open Access Journals (Sweden)

    Liuzhe Li

    Full Text Available A biased usage of immunoglobulin (Ig genes is observed in human anti-HIV-1 monoclonal antibodies (mAbs resulting probably from compensation to reduced usage of the VH3 family genes, while the other alternative suggests that this bias usage is due to antigen requirements. If the antigen structure is responsible for the preferential usage of particular Ig genes, it may have certain implications for HIV vaccine development by the targeting of particular Ig gene-encoded B cell receptors to induce neutralizing anti-HIV-1 antibodies. To address this issue, we have produced HIV-1 specific and non-HIV-1 mAbs from an infected individual and analyzed the Ig gene usage. Green-fluorescence labeled virus-like particles (VLP expressing HIV-1 envelope (Env proteins of JRFL and BaL and control VLPs (without Env were used to select single B cells for the production of 68 recombinant mAbs. Ten of these mAbs were HIV-1 Env specific with neutralizing activity against V3 and the CD4 binding site, as well as non-neutralizing mAbs to gp41. The remaining 58 mAbs were non-HIV-1 Env mAbs with undefined specificities. Analysis revealed that biased usage of Ig genes was restricted only to anti-HIV-1 but not to non-HIV-1 mAbs. The VH1 family genes were dominantly used, followed by VH3, VH4, and VH5 among anti-HIV-1 mAbs, while non-HIV-1 specific mAbs preferentially used VH3 family genes, followed by VH4, VH1 and VH5 families in a pattern identical to Abs derived from healthy individuals. This observation suggests that the biased usage of Ig genes by anti-HIV-1 mAbs is driven by structural requirements of the virus antigens rather than by compensation to any depletion of VH3 B cells due to autoreactive mechanisms, according to the gp120 superantigen hypothesis.

  1. Thermal stability of the human immunodeficiency virus type 1 (HIV-1 receptors, CD4 and CXCR4, reconstituted in proteoliposomes.

    Directory of Open Access Journals (Sweden)

    Mikhail A Zhukovsky

    2010-10-01

    Full Text Available The entry of human immunodeficiency virus (HIV-1 into host cells involves the interaction of the viral exterior envelope glycoprotein, gp120, and receptors on the target cell. The HIV-1 receptors are CD4 and one of two chemokine receptors, CCR5 or CXCR4.We created proteoliposomes that contain CD4, the primary HIV-1 receptor, and one of the coreceptors, CXCR4. Antibodies against CD4 and CXCR4 specifically bound the proteoliposomes. CXCL12, the natural ligand for CXCR4, and the small-molecule CXCR4 antagonist, AMD3100, bound the proteoliposomes with affinities close to those associated with the binding of these molecules to cells expressing CXCR4 and CD4. The HIV-1 gp120 exterior envelope glycoprotein bound tightly to proteoliposomes expressing only CD4 and, in the presence of soluble CD4, bound weakly to proteoliposomes expressing only CXCR4. The thermal stability of CD4 and CXCR4 inserted into liposomes was examined. Thermal denaturation of CXCR4 followed second-order kinetics, with an activation energy (E(a of 269 kJ/mol (64.3 kcal/mol and an inactivation temperature (T(i of 56°C. Thermal inactivation of CD4 exhibited a reaction order of 1.3, an E(a of 278 kJ/mol (66.5 kcal/mol, and a T(i of 52.2°C. The second-order denaturation kinetics of CXCR4 is unusual among G protein-coupled receptors, and may result from dimeric interactions between CXCR4 molecules.Our studies with proteoliposomes containing the native HIV-1 receptors allowed an examination of the binding of biologically important ligands and revealed the higher-order denaturation kinetics of these receptors. CD4/CXCR4-proteoliposomes may be useful for the study of virus-target cell interactions and for the identification of inhibitors.

  2. Antibodies to a conformational epitope on gp41 neutralize HIV-1 by destabilizing the Env spike

    Science.gov (United States)

    Lee, Jeong Hyun; Leaman, Daniel P.; Kim, Arthur S.; Torrents de La Peña, Alba; Sliepen, Kwinten; Yasmeen, Anila; Derking, Ronald; Ramos, Alejandra; de Taeye, Steven W.; Ozorowski, Gabriel; Klein, Florian; Burton, Dennis R.; Nussenzweig, Michel C.; Poignard, Pascal; Moore, John P.; Klasse, Per Johan; Sanders, Rogier W.; Zwick, Michael B.; Wilson, Ian A.; Ward, Andrew B.

    2015-09-01

    The recent identification of three broadly neutralizing antibodies (bnAbs) against gp120-gp41 interface epitopes has expanded the targetable surface on the HIV-1 envelope glycoprotein (Env) trimer. By using biochemical, biophysical and computational methods, we map the previously unknown trimer epitopes of two related antibodies, 3BC315 and 3BC176. A cryo-EM reconstruction of a soluble Env trimer bound to 3BC315 Fab at 9.3 Å resolution reveals that the antibody binds between two gp41 protomers, and neutralizes the virus by accelerating trimer decay. In contrast, bnAb 35O22 binding to a partially overlapping quaternary epitope at the gp120-gp41 interface does not induce decay. A conserved gp41-proximal glycan at N88 was also shown to play a role in the binding kinetics of 3BC176 and 3BC315. Finally, our data suggest that the dynamic structure of the Env trimer influences exposure of bnAb epitopes.

  3. Maturation Pathways of Cross-Reactive HIV-1 Neutralizing Antibodies

    Directory of Open Access Journals (Sweden)

    Dimiter S. Dimitrov

    2009-11-01

    Full Text Available Several human monoclonal antibodies (hmAbs and antibody fragments, including the best characterized in terms of structure-function b12 and Fab X5, exhibit relatively potent and broad HIV-1 neutralizing activity. However, the elicitation of b12 or b12-like antibodies in vivo by vaccine immunogens based on the HIV-1 envelope glycoprotein (Env has not been successful. B12 is highly divergent from the closest corresponding germline antibody while X5 is less divergent. We have hypothesized that the relatively high degree of specific somatic hypermutations may preclude binding of the HIV-1 envelope glycoprotein (Env to closest germline antibodies, and that identifying antibodies that are intermediates in the pathways to maturation could help design novel vaccine immunogens to guide the immune system for their enhanced elicitation. In support of this hypothesis we have previously found that a germline-like b12 (monovalent and bivalent scFv as an Fc fusion protein or IgG lacks measurable binding to an Env as measured by ELISA with a sensitivity in the μM range [1]; here we present evidence confirming and expanding these findings for a panel of Envs. In contrast, a germline-like scFv X5 bound Env with high (nM affinity. To begin to explore the maturation pathways of these antibodies we identified several possible b12 intermediate antibodies and tested their neutralizing activity. These intermediate antibodies neutralized only some HIV-1 isolates and with relatively weak potency. In contrast, germline-like scFv X5 neutralized a subset of the tested HIV-1 isolates with comparable efficiencies to that of the mature X5. These results could help explain the relatively high immunogenicity of the coreceptor binding site on gp120 and the abundance of CD4-induced (CD4i antibodies in HIV-1-infected patients (X5 is a CD4i antibody as well as the maturation pathway of X5. They also can help identify antigens that can bind specifically to b12 germline and

  4. X-ray, Cryo-EM, and computationally predicted protein structures used in integrative modeling of HIV Env glycoprotein gp120 in complex with CD4 and 17b

    Directory of Open Access Journals (Sweden)

    Muhibur Rasheed

    2016-03-01

    Full Text Available We present the data used for an integrative approach to computational modeling of proteins with large variable domains, specifically applied in this context to model HIV Env glycoprotein gp120 in its CD4 and 17b bound state. The initial data involved X-ray structure PDBID:1GC1 and electron microscopy image EMD:5020. Other existing X-ray structures were used as controls to validate and hierarchically refine partial and complete computational models. A summary of the experiment protocol and data was published (Rasheed et al., 2015 [26], along with detailed analysis of the final model (PDBID:3J70 and its implications.

  5. Comparative Analysis of the Glycosylation Profiles of Membrane-Anchored HIV-1 Envelope Glycoprotein Trimers and Soluble gp140

    Science.gov (United States)

    Go, Eden P.; Herschhorn, Alon; Gu, Christopher; Castillo-Menendez, Luis; Zhang, Shijian; Mao, Youdong; Chen, Haiyan; Ding, Haitao; Wakefield, John K.; Hua, David; Liao, Hua-Xin; Kappes, John C.; Sodroski, Joseph

    2015-01-01

    ABSTRACT The human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env) trimer, which consists of the gp120 and gp41 subunits, is the focus of multiple strategies for vaccine development. Extensive Env glycosylation provides HIV-1 with protection from the immune system, yet the glycans are also essential components of binding epitopes for numerous broadly neutralizing antibodies. Recent studies have shown that when Env is isolated from virions, its glycosylation profile differs significantly from that of soluble forms of Env (gp120 or gp140) predominantly used in vaccine discovery research. Here we show that exogenous membrane-anchored Envs, which can be produced in large quantities in mammalian cells, also display a virion-like glycan profile, where the glycoprotein is extensively decorated with high-mannose glycans. Additionally, because we characterized the glycosylation with a high-fidelity profiling method, glycopeptide analysis, an unprecedented level of molecular detail regarding membrane Env glycosylation and its heterogeneity is presented. Each glycosylation site was characterized individually, with about 500 glycoforms characterized per Env protein. While many of the sites contain exclusively high-mannose glycans, others retain complex glycans, resulting in a glycan profile that cannot currently be mimicked on soluble gp120 or gp140 preparations. These site-level studies are important for understanding antibody-glycan interactions on native Env trimers. Additionally, we report a newly observed O-linked glycosylation site, T606, and we show that the full O-linked glycosylation profile of membrane-associated Env is similar to that of soluble gp140. These findings provide new insight into Env glycosylation and clarify key molecular-level differences between membrane-anchored Env and soluble gp140. IMPORTANCE A vaccine that protects against human immunodeficiency virus type 1 (HIV-1) infection should elicit antibodies that bind to the surface

  6. Expression of ATP-binding cassette membrane transporters in a HIV-1 transgenic rat model.

    Science.gov (United States)

    Robillard, Kevin R; Hoque, Md Tozammel; Bendayan, Reina

    2014-02-21

    P-glycoprotein (P-gp, product of Mdr1a and Mdr1b genes), multidrug resistance associated proteins (Mrps), and breast cancer resistance protein (Bcrp), all members of the ATP-binding cassette (ABC) membrane-associated drug transporters superfamily, can significantly restrict the entry of antiretroviral drugs (ARVs) into organs which exhibit a barrier function such as the central nervous system (CNS) and the male genital tract (MGT). In vitro, HIV-1 viral proteins such as glycoprotein-120 (gp120) and transcriptional transactivator (tat) have been shown to alter the expression of these transporters and ARVs permeability. The objective of this study was to compare mRNA expression of these transporters, in vivo, in several tissues obtained from HIV-1 transgenic rats (Tg-rat) (8 and 24 weeks) with those of age-matched wild-type rats. At 24 weeks, significant changes in several drug transporter mRNA expressions were observed, in particular, in brain, kidney, liver and testes. These findings suggest that HIV-1 viral proteins can alter the expression of ABC drug transporters, in vivo, in the context of HIV-1 and further regulate ARVs permeability in several organs including the CNS and MGT, two sites which have been reported to display very low ARVs permeability in the clinic. Copyright © 2014 Elsevier Inc. All rights reserved.

  7. Longitudinal Analysis of CCR5 and CXCR4 Usage in a Cohort of Antiretroviral Therapy-Naive Subjects with Progressive HIV-1 Subtype C Infection.

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    Martin R Jakobsen

    Full Text Available HIV-1 subtype C (C-HIV is responsible for most HIV-1 cases worldwide. Although the pathogenesis of C-HIV is thought to predominantly involve CCR5-restricted (R5 strains, we do not have a firm understanding of how frequently CXCR4-using (X4 and R5X4 variants emerge in subjects with progressive C-HIV infection. Nor do we completely understand the molecular determinants of coreceptor switching by C-HIV variants. Here, we characterized a panel of HIV-1 envelope glycoproteins (Envs (n = 300 cloned sequentially from plasma of 21 antiretroviral therapy (ART-naïve subjects who experienced progression from chronic to advanced stages of C-HIV infection, and show that CXCR4-using C-HIV variants emerged in only one individual. Mutagenesis studies and structural models suggest that the evolution of R5 to X4 variants in this subject principally involved acquisition of an "Ile-Gly" insertion in the gp120 V3 loop and replacement of the V3 "Gly-Pro-Gly" crown with a "Gly-Arg-Gly" motif, but that the accumulation of additional gp120 "scaffold" mutations was required for these V3 loop changes to confer functional effects. In this context, either of the V3 loop changes could confer possible transitional R5X4 phenotypes, but when present together they completely abolished CCR5 usage and conferred the X4 phenotype. Our results show that the emergence of CXCR4-using strains is rare in this cohort of untreated individuals with advanced C-HIV infection. In the subject where X4 variants did emerge, alterations in the gp120 V3 loop were necessary but not sufficient to confer CXCR4 usage.

  8. Heroin-HIV-1 (H2) vaccine: induction of dual immunologic effects with a heroin hapten-conjugate and an HIV-1 envelope V2 peptide with liposomal lipid A as an adjuvant.

    Science.gov (United States)

    Torres, Oscar B; Matyas, Gary R; Rao, Mangala; Peachman, Kristina K; Jalah, Rashmi; Beck, Zoltan; Michael, Nelson L; Rice, Kenner C; Jacobson, Arthur E; Alving, Carl R

    2017-01-01

    A synthetic heroin analog (MorHap) and a synthetic 42 amino acid V2 loop peptide from A/E strain of HIV-1 gp120 envelope protein that was previously used in a successful phase III vaccine trial were constructed as antigens together with liposomes containing monophosphoryl lipid A as an adjuvant, to explore the feasibility of producing a dual use vaccine both for treatment of heroin addiction and prevention of HIV-1 infection among injection drug users. The V2 peptide was tethered by a palmitoyl fatty acyl tail embedded in the liposomal lipid bilayer, and the heroin analog was conjugated to tetanus toxoid as a carrier protein that was mixed with the adjuvant. Upon comparison of a linear V2 peptide with a cyclic peptide, differences were found in the secondary configurations by circular dichroism, with the tethered cyclic peptide (palm-cyclic peptide) entirely in a random coil, and the tethered linear V2 peptide (palm-linear V2 peptide) entirely in a beta-sheet. Upon immunization of mice, palm-cyclic peptide induced anti-cyclic peptide endpoint titers >10 6 and was considered to be a better immunogen overall than palm-linear V2 peptide for inducing antibodies to gp120 and gp70-V1V2. The antibodies also inhibited the binding of V2 peptide to the HIV-1 α 4 β 7 integrin receptor. Antibody titers to MorHap, even with the presence of injected cyclic peptide, were very high, and resulted in inhibition of the hyper-locomotion and antinociception effects of injected heroin. From these initial experiments, we conclude that with a potent adjuvant and mostly synthetic constituents, a vaccine directed to heroin and HIV-1 (H2 vaccine) could be a feasible objective.

  9. Mutations at the CXCR4 interaction sites for AMD3100 influence anti-CXCR4 antibody binding and HIV-1 entry

    DEFF Research Database (Denmark)

    Hatse, Sigrid; Princen, Katrien; Vermeire, Kurt

    2003-01-01

    The interaction of the CXCR4 antagonist AMD3100 with its target is greatly influenced by specific aspartate residues in the receptor protein, including Asp(171) and Asp(262). We have now found that aspartate-to-asparagine substitutions at these positions differentially affect the binding of four...... different anti-CXCR4 monoclonal antibodies as well as the infectivity of diverse human immunodeficiency virus type 1 (HIV-1) strains and clinical isolates. Mutation of Asp(262) strongly decreased the coreceptor efficiency of CXCR4 for wild-type but not for AMD3100-resistant HIV-1 NL4.3. Thus, resistance...... of HIV-1 NL4.3 to AMD3100 is associated with a decreased dependence of the viral gp120 on Asp(262) of CXCR4, pointing to a different mode of interaction of wild-type versus AMD3100-resistant virus with CXCR4....

  10. Complementation of diverse HIV-1 Env defects through cooperative subunit interactions: a general property of the functional trimer

    Directory of Open Access Journals (Sweden)

    Salzwedel Karl

    2009-08-01

    Full Text Available Abstract Background The HIV-1 Env glycoprotein mediates virus entry by catalyzing direct fusion between the virion membrane and the target cell plasma membrane. Env is composed of two subunits: gp120, which binds to CD4 and the coreceptor, and gp41, which is triggered upon coreceptor binding to promote the membrane fusion reaction. Env on the surface of infected cells is a trimer consisting of three gp120/gp41 homo-dimeric protomers. An emerging question concerns cooperative interactions between the protomers in the trimer, and possible implications for Env function. Results We extended studies on cooperative subunit interactions within the HIV-1 Env trimer, using analysis of functional complementation between coexpressed inactive variants harboring different functional deficiencies. In assays of Env-mediated cell fusion, complementation was observed between variants with a wide range of defects in both the gp120 and gp41 subunits. The former included gp120 subunits mutated in the CD4 binding site or incapable of coreceptor interaction due either to mismatched specificity or V3 loop mutation. Defective gp41 variants included point mutations at different residues within the fusion peptide or heptad repeat regions, as well as constructs with modifications or deletions of the membrane proximal tryptophan-rich region or the transmembrane domain. Complementation required the defective variants to be coexpressed in the same cell. The observed complementation activities were highly dependent on the assay system. The most robust activities were obtained with a vaccinia virus-based expression and reporter gene activation assay for cell fusion. In an alternative system involving Env expression from integrated provirus, complementation was detected in cell fusion assays, but not in virus particle entry assays. Conclusion Our results indicate that Env function does not require every subunit in the trimer to be competent for all essential activities. Through

  11. Analysis of a Clonal Lineage of HIV-1 Envelope V2/V3 Conformational Epitope-Specific Broadly Neutralizing Antibodies and Their Inferred Unmutated Common Ancestors ▿ †

    Science.gov (United States)

    Bonsignori, Mattia; Hwang, Kwan-Ki; Chen, Xi; Tsao, Chun-Yen; Morris, Lynn; Gray, Elin; Marshall, Dawn J.; Crump, John A.; Kapiga, Saidi H.; Sam, Noel E.; Sinangil, Faruk; Pancera, Marie; Yongping, Yang; Zhang, Baoshan; Zhu, Jiang; Kwong, Peter D.; O'Dell, Sijy; Mascola, John R.; Wu, Lan; Nabel, Gary J.; Phogat, Sanjay; Seaman, Michael S.; Whitesides, John F.; Moody, M. Anthony; Kelsoe, Garnett; Yang, Xinzhen; Sodroski, Joseph; Shaw, George M.; Montefiori, David C.; Kepler, Thomas B.; Tomaras, Georgia D.; Alam, S. Munir; Liao, Hua-Xin; Haynes, Barton F.

    2011-01-01

    V2/V3 conformational epitope antibodies that broadly neutralize HIV-1 (PG9 and PG16) have been recently described. Since an elicitation of previously known broadly neutralizing antibodies has proven elusive, the induction of antibodies with such specificity is an important goal for HIV-1 vaccine development. A critical question is which immunogens and vaccine formulations might be used to trigger and drive the development of memory B cell precursors with V2/V3 conformational epitope specificity. In this paper we identified a clonal lineage of four V2/V3 conformational epitope broadly neutralizing antibodies (CH01 to CH04) from an African HIV-1-infected broad neutralizer and inferred their common reverted unmutated ancestor (RUA) antibodies. While conformational epitope antibodies rarely bind recombinant Env monomers, a screen of 32 recombinant envelopes for binding to the CH01 to CH04 antibodies showed monoclonal antibody (MAb) binding to the E.A244 gp120 Env and to chronic Env AE.CM243; MAbs CH01 and CH02 also bound to transmitted/founder Env B.9021. CH01 to CH04 neutralized 38% to 49% of a panel of 91 HIV-1 tier 2 pseudoviruses, while the RUAs neutralized only 16% of HIV-1 isolates. Although the reverted unmutated ancestors showed restricted neutralizing activity, they retained the ability to bind to the E.A244 gp120 HIV-1 envelope with an affinity predicted to trigger B cell development. Thus, E.A244, B.9021, and AE.CM243 Envs are three potential immunogen candidates for studies aimed at defining strategies to induce V2/V3 conformational epitope-specific antibodies. PMID:21795340

  12. Analysis of Select Herpes Simplex Virus 1 (HSV-1) Proteins for Restriction of Human Immunodeficiency Virus Type 1 (HIV-1): HSV-1 gM Protein Potently Restricts HIV-1 by Preventing Intracellular Transport and Processing of Env gp160.

    Science.gov (United States)

    Polpitiya Arachchige, Sachith; Henke, Wyatt; Pramanik, Ankita; Kalamvoki, Maria; Stephens, Edward B

    2018-01-15

    Virus-encoded proteins that impair or shut down specific host cell functions during replication can be used as probes to identify potential proteins/pathways used in the replication of viruses from other families. We screened nine proteins from herpes simplex virus 1 (HSV-1) for the ability to enhance or restrict human immunodeficiency virus type 1 (HIV-1) replication. We show that several HSV-1 proteins (glycoprotein M [gM], US3, and UL24) potently restricted the replication of HIV-1. Unlike UL24 and US3, which reduced viral protein synthesis, we observed that gM restriction of HIV-1 occurred through interference with the processing and transport of gp160, resulting in a significantly reduced level of mature gp120/gp41 released from cells. Finally, we show that an HSV-1 gM mutant lacking the majority of the C-terminal domain (HA-gM[Δ345-473]) restricted neither gp160 processing nor the release of infectious virus. These studies identify proteins from heterologous viruses that can restrict viruses through novel pathways.IMPORTANCE HIV-1 infection of humans results in AIDS, characterized by the loss of CD4+ T cells and increased susceptibility to opportunistic infections. Both HIV-1 and HSV-1 can infect astrocytes and microglia of the central nervous system (CNS). Thus, the identification of HSV-1 proteins that directly restrict HIV-1 or interfere with pathways required for HIV-1 replication could lead to novel antiretroviral strategies. The results of this study show that select viral proteins from HSV-1 can potently restrict HIV-1. Further, our results indicate that the gM protein of HSV-1 restricts HIV-1 through a novel pathway by interfering with the processing of gp160 and its incorporation into virus maturing from the cell. Copyright © 2018 American Society for Microbiology.

  13. Antiviral mechanism of polyanionic carbosilane dendrimers against HIV-1.

    Science.gov (United States)

    Vacas-Córdoba, Enrique; Maly, Marek; De la Mata, Francisco J; Gómez, Rafael; Pion, Marjorie; Muñoz-Fernández, M Ángeles

    2016-01-01

    Nanotechnology-derived platforms, such as dendrimers, are very attractive in several biological applications. In the case of human immunodeficiency virus (HIV) infection, polyanionic carbosilane dendrimers have shown great potential as antiviral agents in the development of novel microbicides to prevent the sexual transmission of HIV-1. In this work, we studied the mechanism of two sulfated and naphthylsulfonated functionalized carbosilane dendrimers, G3-S16 and G2-NF16. They are able to inhibit viral infection at fusion and thus at the entry step. Both compounds impede the binding of viral particles to target cell surface and membrane fusion through the blockage of gp120-CD4 interaction. In addition, and for the first time, we demonstrate that dendrimers can inhibit cell-to-cell HIV transmission and difficult infectious synapse formation. Thus, carbosilane dendrimers' mode of action is a multifactorial process targeting several proteins from viral envelope and from host cells that could block HIV infection at different stages during the first step of infection.

  14. Isolation of HIV-1-neutralizing mucosal monoclonal antibodies from human colostrum.

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    James Friedman

    Full Text Available Generation of potent anti-HIV antibody responses in mucosal compartments is a potential requirement of a transmission-blocking HIV vaccine. HIV-specific, functional antibody responses are present in breast milk, and these mucosal antibody responses may play a role in protection of the majority of HIV-exposed, breastfeeding infants. Therefore, characterization of HIV-specific antibodies produced by B cells in milk could guide the development of vaccines that elicit protective mucosal antibody responses.We isolated B cells from colostrum of an HIV-infected lactating woman with a detectable neutralization response in milk and recombinantly produced and characterized the resulting HIV-1 Envelope (Env-specific monoclonal antibodies (mAbs.The identified HIV-1 Env-specific colostrum mAbs, CH07 and CH08, represent two of the first mucosally-derived anti-HIV antibodies yet to be reported. Colostrum mAb CH07 is a highly-autoreactive, weakly-neutralizing gp140-specific mAb that binds to linear epitopes in the gp120 C5 region and gp41 fusion domain. In contrast, colostrum mAb CH08 is a nonpolyreactive CD4-inducible (CD4i gp120-specific mAb with moderate breadth of neutralization.These novel HIV-neutralizing mAbs isolated from a mucosal compartment provide insight into the ability of mucosal B cell populations to produce functional anti-HIV antibodies that may contribute to protection against virus acquisition at mucosal surfaces.

  15. V3 loop truncations in HIV-1 envelope impart resistance to coreceptor inhibitors and enhanced sensitivity to neutralizing antibodies.

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    Meg M Laakso

    2007-08-01

    Full Text Available The V1/V2 region and the V3 loop of the human immunodeficiency virus type I (HIV-1 envelope (Env protein are targets for neutralizing antibodies and also play an important functional role, with the V3 loop largely determining whether a virus uses CCR5 (R5, CXCR4 (X4, or either coreceptor (R5X4 to infect cells. While the sequence of V3 is variable, its length is highly conserved. Structural studies indicate that V3 length may be important for interactions with the extracellular loops of the coreceptor. Consistent with this view, genetic truncation of the V3 loop is typically associated with loss of Env function. We removed approximately one-half of the V3 loop from three different HIV-1 strains, and found that only the Env protein from the R5X4 strain R3A retained some fusion activity. Loss of V1/V2 (DeltaV1/V2 was well tolerated by this virus. Passaging of virus with the truncated V3 loop resulted in the derivation of a virus strain that replicated with wild-type kinetics. This virus, termed TA1, retained the V3 loop truncation and acquired several adaptive changes in gp120 and gp41. TA1 could use CCR5 but not CXCR4 to infect cells, and was extremely sensitive to neutralization by HIV-1 positive human sera, and by antibodies to the CD4 binding site and to CD4-induced epitopes in the bridging sheet region of gp120. In addition, TA1 was completely resistant to CCR5 inhibitors, and was more dependent upon the N-terminal domain of CCR5, a region of the receptor that is thought to contact the bridging sheet of gp120 and the base of the V3 loop, and whose conformation may not be greatly affected by CCR5 inhibitors. These studies suggest that the V3 loop protects HIV from neutralization by antibodies prevalent in infected humans, that CCR5 inhibitors likely act by disrupting interactions between the V3 loop and the coreceptor, and that altered use of CCR5 by HIV-1 associated with increased sensitivity to changes in the N-terminal domain can be linked

  16. Immunogenic Display of Purified Chemically Cross-Linked HIV-1 Spikes

    Science.gov (United States)

    Leaman, Daniel P.; Lee, Jeong Hyun; Ward, Andrew B.

    2015-01-01

    ABSTRACT HIV-1 envelope glycoprotein (Env) spikes are prime vaccine candidates, at least in principle, but suffer from instability, molecular heterogeneity and a low copy number on virions. We anticipated that chemical cross-linking of HIV-1 would allow purification and molecular characterization of trimeric Env spikes, as well as high copy number immunization. Broadly neutralizing antibodies bound tightly to all major quaternary epitopes on cross-linked spikes. Covalent cross-linking of the trimer also stabilized broadly neutralizing epitopes, although surprisingly some individual epitopes were still somewhat sensitive to heat or reducing agent. Immunodepletion using non-neutralizing antibodies to gp120 and gp41 was an effective method for removing non-native-like Env. Cross-linked spikes, purified via an engineered C-terminal tag, were shown by negative stain EM to have well-ordered, trilobed structure. An immunization was performed comparing a boost with Env spikes on virions to spikes cross-linked and captured onto nanoparticles, each following a gp160 DNA prime. Although differences in neutralization did not reach statistical significance, cross-linked Env spikes elicited a more diverse and sporadically neutralizing antibody response against Tier 1b and 2 isolates when displayed on nanoparticles, despite attenuated binding titers to gp120 and V3 crown peptides. Our study demonstrates display of cross-linked trimeric Env spikes on nanoparticles, while showing a level of control over antigenicity, purity and density of virion-associated Env, which may have relevance for Env based vaccine strategies for HIV-1. IMPORTANCE The envelope spike (Env) is the target of HIV-1 neutralizing antibodies, which a successful vaccine will need to elicit. However, native Env on virions is innately labile, as well as heterogeneously and sparsely displayed. We therefore stabilized Env spikes using a chemical cross-linker and removed non-native Env by immunodepletion with non

  17. Photoinduced Reactivity of the HIV-1 Envelope Glycoprotein with a Membrane-Embedded Probe Reveals Insertion of Portions of the HIV-1 Gp41 Cytoplasmic Tail into the Viral Membrane†

    Science.gov (United States)

    Viard, Mathias; Ablan, Sherimay D.; Zhou, Ming; Veenstra, Timothy D.; Freed, Eric O.; Raviv, Yossef; Blumenthal, Robert

    2008-01-01

    The interactions of HIV-1 Env (gp120-gp41) with CD4 and coreceptors trigger a barrage of conformational changes in Env that drive the membrane fusion process. Various regions of gp41 have profound effects on HIV entry and budding. However, the precise interactions between gp41 and the membrane have not been elucidated. To examine portions of membrane proteins that are embedded in membrane lipids, we have studied photoinduced chemical reactions in membranes using the lipid bilayer specific probe iodonaphthyl azide (INA). Here we show that in addition to the transmembrane anchor, amphipatic sequences in the cytoplasmic tail (CT) of HIV-1 gp41 are labeled by INA. INA labeling of the HIV-1 gp41 CT was similar whether wild-type or a mutant HIV-1 was used with uncleaved p55 Gag, which does not allow entry. These results shed light on the disposition of the HIV-1 gp41 CT with respect to the membrane. Moreover, our data have general implications for topology of membrane proteins and their in situ interactions with the lipid bilayer. PMID:18198900

  18. Oral and vaginal epithelial cell lines bind and transfer cell-free infectious HIV-1 to permissive cells but are not productively infected.

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    Arinder Kohli

    Full Text Available The majority of HIV-1 infections worldwide are acquired via mucosal surfaces. However, unlike the vaginal mucosa, the issue of whether the oral mucosa can act as a portal of entry for HIV-1 infection remains controversial. To address potential differences with regard to the fate of HIV-1 after exposure to oral and vaginal epithelium, we utilized two epithelial cell lines representative of buccal (TR146 and pharyngeal (FaDu sites of the oral cavity and compared them with a cell line derived from vaginal epithelium (A431 in order to determine (i HIV-1 receptor gene and protein expression, (ii whether HIV-1 genome integration into epithelial cells occurs, (iii whether productive viral infection ensues, and (iv whether infectious virus can be transferred to permissive cells. Using flow cytometry to measure captured virus by HIV-1 gp120 protein detection and western blot to detect HIV-1 p24 gag protein, we demonstrate that buccal, pharyngeal and vaginal epithelial cells capture CXCR4- and CCR5-utilising virus, probably via non-canonical receptors. Both oral and vaginal epithelial cells are able to transfer infectious virus to permissive cells either directly through cell-cell attachment or via transcytosis of HIV-1 across epithelial cells. However, HIV-1 integration, as measured by real-time PCR and presence of early gene mRNA transcripts and de novo protein production were not detected in either epithelial cell type. Importantly, both oral and vaginal epithelial cells were able to support integration and productive infection if HIV-1 entered via the endocytic pathway driven by VSV-G. Our data demonstrate that under normal conditions productive HIV-1 infection of epithelial cells leading to progeny virion production is unlikely, but that epithelial cells can act as mediators of systemic viral dissemination through attachment and transfer of HIV-1 to permissive cells.

  19. HIV-1 gp41 Fusion Intermediate: A Target for HIV Therapeutics

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    Chungen Pan

    2010-02-01

    Full Text Available Human immunodeficiency virus (HIV-1 infection is initiated by the binding of gp120 envelope glyco-protein to its cell receptor (CD4 and a coreceptor (CXCR4 or CCR5, followed by a series of conformational changes in the gp41 transmembrane subunit. These changes include insertion of fusion peptide into the target cell membrane and association of C-heptad repeat (CHR peptide with the N-heptad repeat (NHR trimer, a pre-hairpin fusion intermediate. A stable six-helix bundle core is then formed, bringing the viral envelope and target cell membrane into close proximity for fusion. Peptides derived from the CHR region, such as T20 and C34, inhibit HIV-1 fusion by interacting with the gp41 fusion intermediate. A number of anti-HIV-1 peptides and small molecule compounds targeting the gp41 NHR-trimer have been identified. By combining HIV fusion/entry inhibitors targeting different sites in the gp41 fusion intermediate, a potent synergistic effect takes place, resulting in a potential new therapeutic strategy for the HIV infection/AIDS. Here, we present an overview of the current development of anti-HIV drugs, particularly those targeting the gp41 fusion intermediate.

  20. Mechanisms of HIV-1 Control.

    Science.gov (United States)

    Soliman, Mary; Srikrishna, Geetha; Balagopal, Ashwin

    2017-06-01

    HIV-1 infection is of global importance, and still incurs substantial morbidity and mortality. Although major pharmacologic advances over the past two decades have resulted in remarkable HIV-1 control, a cure is still forthcoming. One approach to a cure is to exploit natural mechanisms by which the host restricts HIV-1. Herein, we review past and recent discoveries of HIV-1 restriction factors, a diverse set of host proteins that limit HIV-1 replication at multiple levels, including entry, reverse transcription, integration, translation of viral proteins, and packaging and release of virions. Recent studies of intracellular HIV-1 restriction have offered unique molecular insights into HIV-1 replication and biology. Studies have revealed insights of how restriction factors drive HIV-1 evolution. Although HIV-1 restriction factors only partially control the virus, their importance is underscored by their effect on HIV-1 evolution and adaptation. The list of host restriction factors that control HIV-1 infection is likely to expand with future discoveries. A deeper understanding of the molecular mechanisms of regulation by these factors will uncover new targets for therapeutic control of HIV-1 infection.

  1. Neutralization sensitivity of HIV-1 subtype B' clinical isolates from former plasma donors in China.

    Science.gov (United States)

    OuYang, Yabo; Sun, Jianping; Huang, Yang; Lu, Lu; Xu, Weisi; Hu, Xintao; Hong, Kunxue; Jiang, Shibo; Shao, Yiming; Ma, Liying

    2013-01-05

    HIV-1 subtype B' isolates have been predominantly circulating in China. Their intra- and inter-subtype neutralization sensitivity to autologous and heterologous plasmas has not been well studied. Twelve HIV-1 B' clinical isolates obtained from patients were tested for their intra- and inter-subtype neutralization sensitivity to the neutralization antibodies in the plasmas from patients infected by HIV-1 B' and CRF07_BC subtypes, respectively. We found that the plasmas from the HIV-1 B'-infected patients could potently neutralize heterologous viruses of subtype B' with mean ID50 titer (1/x) of about 67, but they were not effective in neutralizing autologous viruses of subtype B' with mean ID50 titer (1/x) of about 8. The plasmas from HIV-1 CRF07_BC-infected patients exhibited weak inter-subtype neutralization activity against subtype B' viruses with ID50 titer (1/x) is about 22. The neutralization sensitivity of HIV-1 B' isolates was inversely correlated with the neutralizing activity of plasmas from HIV-1 B'-infected patients (Spearman's r = -0.657, P = 0.020), and with the number of potential N-glycosylation site (PNGS) in V1-V5 region (Spearman's r = -0.493, P = 0.034), but positively correlated with the viral load (Spearman's r = 0.629, P = 0.028). It had no correlation with the length of V1-V5 regions or the CD4+ T cell count. Virus AH259V has low intra-subtype neutralization sensitivity, it can be neutralized by 17b (IC50: 10μg/ml) and 447-52D (IC50: 1.6μg/ml), and the neutralizing antibodies (nAbs) in plasma AH259P are effective in neutralizing infection by the primary HIV-1 isolates with different subtypes with ID50 titers (1/x) in the range of 32-396. These findings suggest that the HIV-1 subtype B' viruses may mutate under the immune pressure, thus becoming resistant to the autologous nAbs, possibly by changing the number of PNGS in the V1-V5 region of the viral gp120. Some of primary HIV-1 isolates are able to induce both intra- and inter-subtype cross

  2. Comprehensive sieve analysis of breakthrough HIV-1 sequences in the RV144 vaccine efficacy trial.

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    Paul T Edlefsen

    2015-02-01

    Full Text Available The RV144 clinical trial showed the partial efficacy of a vaccine regimen with an estimated vaccine efficacy (VE of 31% for protecting low-risk Thai volunteers against acquisition of HIV-1. The impact of vaccine-induced immune responses can be investigated through sieve analysis of HIV-1 breakthrough infections (infected vaccine and placebo recipients. A V1/V2-targeted comparison of the genomes of HIV-1 breakthrough viruses identified two V2 amino acid sites that differed between the vaccine and placebo groups. Here we extended the V1/V2 analysis to the entire HIV-1 genome using an array of methods based on individual sites, k-mers and genes/proteins. We identified 56 amino acid sites or "signatures" and 119 k-mers that differed between the vaccine and placebo groups. Of those, 19 sites and 38 k-mers were located in the regions comprising the RV144 vaccine (Env-gp120, Gag, and Pro. The nine signature sites in Env-gp120 were significantly enriched for known antibody-associated sites (p = 0.0021. In particular, site 317 in the third variable loop (V3 overlapped with a hotspot of antibody recognition, and sites 369 and 424 were linked to CD4 binding site neutralization. The identified signature sites significantly covaried with other sites across the genome (mean = 32.1 more than did non-signature sites (mean = 0.9 (p < 0.0001, suggesting functional and/or structural relevance of the signature sites. Since signature sites were not preferentially restricted to the vaccine immunogens and because most of the associations were insignificant following correction for multiple testing, we predict that few of the genetic differences are strongly linked to the RV144 vaccine-induced immune pressure. In addition to presenting results of the first complete-genome analysis of the breakthrough infections in the RV144 trial, this work describes a set of statistical methods and tools applicable to analysis of breakthrough infection genomes in general vaccine

  3. Cross-Neutralizing Antibodies in HIV-1 Individuals Infected by Subtypes B, F1, C or the B/Bbr Variant in Relation to the Genetics and Biochemical Characteristics of the env Gene.

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    Dalziza Victalina de Almeida

    Full Text Available Various HIV-1 env genetic and biochemical features impact the elicitation of cross-reactive neutralizing antibodies in natural infections. Thus, we aimed to investigate cross-neutralizing antibodies in individuals infected with HIV-1 env subtypes B, F1, C or the B/Bbr variant as well as env characteristics. Therefore, plasma samples from Brazilian chronically HIV-1 infected individuals were submitted to the TZM-bl neutralization assay. We also analyzed putative N-glycosylation sites (PNGLs and the size of gp120 variable domains in the context of HIV-1 subtypes prevalent in Brazil. We observed a greater breadth and potency of the anti-Env neutralizing response in individuals infected with the F1 or B HIV-1 subtypes compared with the C subtype and the variant B/Bbr. We observed greater V1 B/Bbr and smaller V4 F1 than those of other subtypes (p<0.005, however neither was there a correlation verified between the variable region length and neutralization potency, nor between PNLG and HIV-1 subtypes. The enrichment of W at top of V3 loop in weak neutralizing response viruses and the P in viruses with higher neutralization susceptibility was statistically significant (p = 0.013. Some other signatures sites were associated to HIV-1 subtype-specific F1 and B/Bbr samples might influence in the distinct neutralizing response. These results indicate that a single amino acid substitution may lead to a distinct conformational exposure or load in the association domain of the trimer of gp120 and interfere with the induction power of the neutralizing response, which affects the sensitivity of the neutralizing antibody and has significant implications for vaccine design.

  4. Inhibition of HIV-1 infection in ex vivo cervical tissue model of human vagina by palmitic acid; implications for a microbicide development.

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    Xudong Lin

    Full Text Available BACKGROUND: Approximately 80% of all new HIV-1 infections are acquired through sexual contact. Currently, there is no clinically approved microbicide, indicating a clear and urgent therapeutic need. We recently reported that palmitic acid (PA is a novel and specific inhibitor of HIV-1 fusion and entry. Mechanistically, PA inhibits HIV-1 infection by binding to a novel pocket on the CD4 receptor and blocks efficient gp120-to-CD4 attachment. Here, we wanted to assess the ability of PA to inhibit HIV-1 infection in cervical tissue ex vivo model of human vagina, and determine its effect on Lactobacillus (L species of probiotic vaginal flora. PRINCIPAL FINDINGS: Our results show that treatment with 100-200 µM PA inhibited HIV-1 infection in cervical tissue by up to 50%, and this treatment was not toxic to the tissue or to L. crispatus and jensenii species of vaginal flora. In vitro, in a cell free system that is independent of in vivo cell associated CD4 receptor; we determined inhibition constant (Ki to be ∼2.53 µM. SIGNIFICANCE: These results demonstrate utility of PA as a model molecule for further preclinical development of a safe and potent HIV-1 entry microbicide inhibitor.

  5. Bulk culture levels of specific cytotoxic T-cell activity against HIV-1 proteins are not associated with risk of death

    DEFF Research Database (Denmark)

    Aladdin, H; Ullum, H; Lepri, A Cozzi

    1999-01-01

    The ability of cytotoxic T lymphocytes (CTL) to control and influence the outcome of human immunodeficiency virus (HIV) infection is not fully understood. The association between HIV-CTL activity and disease progression was evaluated prospectively in 36 HIV-1-infected individuals with a median...... follow-up of 3.0 years. HIV-CTL activity was measured in a 4 h Cr* release assay using autologous target cells expressing HIV-1 BRU isolate gene products (gp-120, gag, pol, nef) and a bulk culture of autologous effector cells. The CD4 count was measured at enrolment and plasma HIV RNA was measured...... retrospectively. The present study failed to support the hypothesis that HIV-CTL activity, as measured using the present method, is important in reducing the risk of death in HIV-infected individuals. However, using other approaches and methods could possibly yield other conclusions, and further prospective...

  6. Covariance of charged amino acids at positions 322 and 440 of HIV-1 Env contributes to coreceptor specificity of subtype B viruses, and can be used to improve the performance of V3 sequence-based coreceptor usage prediction algorithms.

    Science.gov (United States)

    Cashin, Kieran; Sterjovski, Jasminka; Harvey, Katherine L; Ramsland, Paul A; Churchill, Melissa J; Gorry, Paul R

    2014-01-01

    The ability to determine coreceptor usage of patient-derived human immunodeficiency virus type 1 (HIV-1) strains is clinically important, particularly for the administration of the CCR5 antagonist maraviroc. The envelope glycoprotein (Env) determinants of coreceptor specificity lie primarily within the gp120 V3 loop region, although other Env determinants have been shown to influence gp120-coreceptor interactions. Here, we determined whether conserved amino acid alterations outside the V3 loop that contribute to coreceptor usage exist, and whether these alterations improve the performance of V3 sequence-based coreceptor usage prediction algorithms. We demonstrate a significant covariant association between charged amino acids at position 322 in V3 and position 440 in the C4 Env region that contributes to the specificity of HIV-1 subtype B strains for CCR5 or CXCR4. Specifically, positively charged Lys/Arg at position 322 and negatively charged Asp/Glu at position 440 occurred more frequently in CXCR4-using viruses, whereas negatively charged Asp/Glu at position 322 and positively charged Arg at position 440 occurred more frequently in R5 strains. In the context of CD4-bound gp120, structural models suggest that covariation of amino acids at Env positions 322 and 440 has the potential to alter electrostatic interactions that are formed between gp120 and charged amino acids in the CCR5 N-terminus. We further demonstrate that inclusion of a "440 rule" can improve the sensitivity of several V3 sequence-based genotypic algorithms for predicting coreceptor usage of subtype B HIV-1 strains, without compromising specificity, and significantly improves the AUROC of the geno2pheno algorithm when set to its recommended false positive rate of 5.75%. Together, our results provide further mechanistic insights into the intra-molecular interactions within Env that contribute to coreceptor specificity of subtype B HIV-1 strains, and demonstrate that incorporation of Env

  7. Covariance of charged amino acids at positions 322 and 440 of HIV-1 Env contributes to coreceptor specificity of subtype B viruses, and can be used to improve the performance of V3 sequence-based coreceptor usage prediction algorithms.

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    Kieran Cashin

    Full Text Available The ability to determine coreceptor usage of patient-derived human immunodeficiency virus type 1 (HIV-1 strains is clinically important, particularly for the administration of the CCR5 antagonist maraviroc. The envelope glycoprotein (Env determinants of coreceptor specificity lie primarily within the gp120 V3 loop region, although other Env determinants have been shown to influence gp120-coreceptor interactions. Here, we determined whether conserved amino acid alterations outside the V3 loop that contribute to coreceptor usage exist, and whether these alterations improve the performance of V3 sequence-based coreceptor usage prediction algorithms. We demonstrate a significant covariant association between charged amino acids at position 322 in V3 and position 440 in the C4 Env region that contributes to the specificity of HIV-1 subtype B strains for CCR5 or CXCR4. Specifically, positively charged Lys/Arg at position 322 and negatively charged Asp/Glu at position 440 occurred more frequently in CXCR4-using viruses, whereas negatively charged Asp/Glu at position 322 and positively charged Arg at position 440 occurred more frequently in R5 strains. In the context of CD4-bound gp120, structural models suggest that covariation of amino acids at Env positions 322 and 440 has the potential to alter electrostatic interactions that are formed between gp120 and charged amino acids in the CCR5 N-terminus. We further demonstrate that inclusion of a "440 rule" can improve the sensitivity of several V3 sequence-based genotypic algorithms for predicting coreceptor usage of subtype B HIV-1 strains, without compromising specificity, and significantly improves the AUROC of the geno2pheno algorithm when set to its recommended false positive rate of 5.75%. Together, our results provide further mechanistic insights into the intra-molecular interactions within Env that contribute to coreceptor specificity of subtype B HIV-1 strains, and demonstrate that incorporation

  8. Sexual transmission of HIV-1.

    Science.gov (United States)

    Fox, Julie; Fidler, Sarah

    2010-01-01

    HIV-1 transmission occurs in a limited number of ways all of which are preventable. Overall, the risk of HIV-1 transmission following a single sexual exposure is low especially in comparison with other sexually transmitted infections (STIs); with estimates of the average probability of male to female HIV-1 transmission only 0.0005-0.0026 per coital act. The risk of acquiring HIV-1 from a single contact varies enormously and is dependant upon the infectiousness of the HIV-1 positive individual and the susceptibility to HIV-1 of their sexual partner. An understanding of the determinants of HIV-1 transmission is important not only to assess the infection risk to an individual when exposed to the virus (e.g. to determine the provision of post exposure prophylaxis), but also to make accurate predictions on the potential spread of HIV-1 infection in a population and to direct appropriate targeted prevention strategies. In this review article we summarise the current literature on the major worldwide source of HIV-1 acquisition, sexual transmission. This article forms part of a special issue of Antiviral Research marking the 25th anniversary of antiretroviral drug discovery and development, Vol 85, issue 1, 2010. Copyright 2009 Elsevier B.V. All rights reserved.

  9. Maintenance of syncytium-inducing phenotype of HIV type 1 is associated with positively charged residues in the HIV type 1 gp120 V2 domain without fixed positions, elongation, or relocated N-linked glycosylation sites

    NARCIS (Netherlands)

    Cornelissen, M.; Hogervorst, E.; Zorgdrager, F.; Hartman, S.; Goudsmit, J.

    1995-01-01

    The prevalence of HIV-1 sequences of the envelope domains V1V2 and V3 was analyzed by RT-PCR amplification. Two distinct biological phenotypes of HIV-1 have been described: the nonsyncytium-inducing (NSI) phenotype, best characterized by the inability to infect MT-2 cells, and the syncytium-inducing

  10. Revisiting HIV-1 uncoating

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    Arhel Nathalie

    2010-11-01

    Full Text Available Abstract HIV uncoating is defined as the loss of viral capsid that occurs within the cytoplasm of infected cells before entry of the viral genome into the nucleus. It is an obligatory step of HIV-1 early infection and accompanies the transition between reverse transcription complexes (RTCs, in which reverse transcription occurs, and pre-integration complexes (PICs, which are competent to integrate into the host genome. The study of the nature and timing of HIV-1 uncoating has been paved with difficulties, particularly as a result of the vulnerability of the capsid assembly to experimental manipulation. Nevertheless, recent studies of capsid structure, retroviral restriction and mechanisms of nuclear import, as well as the recent expansion of technical advances in genome-wide studies and cell imagery approaches, have substantially changed our understanding of HIV uncoating. Although early work suggested that uncoating occurs immediately following viral entry in the cell, thus attributing a trivial role for the capsid in infected cells, recent data suggest that uncoating occurs several hours later and that capsid has an all-important role in the cell that it infects: for transport towards the nucleus, reverse transcription and nuclear import. Knowing that uncoating occurs at a later stage suggests that the viral capsid interacts extensively with the cytoskeleton and other cytoplasmic components during its transport to the nucleus, which leads to a considerable reassessment of our efforts to identify potential therapeutic targets for HIV therapy. This review discusses our current understanding of HIV uncoating, the functional interplay between infectivity and timely uncoating, as well as exposing the appropriate methods to study uncoating and addressing the many questions that remain unanswered.

  11. Morphine enhances HIV-1SF162-mediated neuron death and delays recovery of injured neurites.

    Science.gov (United States)

    Masvekar, Ruturaj R; El-Hage, Nazira; Hauser, Kurt F; Knapp, Pamela E

    2014-01-01

    HIV-1 enters the CNS soon after initial systemic infection; within the CNS parenchyma infected and/or activated perivascular macrophages, microglia and astrocytes release viral and cellular toxins that drive secondary toxicity in neurons and other cell types. Our previous work has largely modeled HIV-neuropathology using the individual viral proteins Tat or gp120, with murine striatal neurons as targets. To model disease processes more closely, the current study uses supernatant from HIV-1-infected cells. Supernatant from HIV-1SF162-infected differentiated-U937 cells (HIV+sup) was collected and p24 level was measured by ELISA to assess the infection. Injection drug abuse is a significant risk factor for HIV-infection, and opiate drug abusers show increased HIV-neuropathology, even with anti-retroviral treatments. We therefore assessed HIV+sup effects on neuronal survival and neurite growth/pruning with or without concurrent exposure to morphine, an opiate that preferentially acts through µ-opioid receptors. Effects of HIV+sup ± morphine were assessed on neuronal populations, and also by time-lapse imaging of individual cells. HIV+sup caused dose-dependent toxicity over a range of p24 levels (10-500 pg/ml). Significant interactions occurred with morphine at lower p24 levels (10 and 25 pg/ml), and GSK3β was implicated as a point of convergence. In the presence of glia, selective neurotoxic measures were significantly enhanced and interactions with morphine were also augmented, perhaps related to a decreased level of BDNF. Importantly, the arrest of neurite growth that occurred with exposure to HIV+sup was reversible unless neurons were continuously exposed to morphine. Thus, while reducing HIV-infection levels may be protective, ongoing exposure to opiates may limit recovery. Opiate interactions observed in this HIV-infective environment were similar, though not entirely concordant, with Tat/gp120 interactions reported previously, suggesting unique interactions

  12. Morphine enhances HIV-1SF162-mediated neuron death and delays recovery of injured neurites.

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    Ruturaj R Masvekar

    Full Text Available HIV-1 enters the CNS soon after initial systemic infection; within the CNS parenchyma infected and/or activated perivascular macrophages, microglia and astrocytes release viral and cellular toxins that drive secondary toxicity in neurons and other cell types. Our previous work has largely modeled HIV-neuropathology using the individual viral proteins Tat or gp120, with murine striatal neurons as targets. To model disease processes more closely, the current study uses supernatant from HIV-1-infected cells. Supernatant from HIV-1SF162-infected differentiated-U937 cells (HIV+sup was collected and p24 level was measured by ELISA to assess the infection. Injection drug abuse is a significant risk factor for HIV-infection, and opiate drug abusers show increased HIV-neuropathology, even with anti-retroviral treatments. We therefore assessed HIV+sup effects on neuronal survival and neurite growth/pruning with or without concurrent exposure to morphine, an opiate that preferentially acts through µ-opioid receptors. Effects of HIV+sup ± morphine were assessed on neuronal populations, and also by time-lapse imaging of individual cells. HIV+sup caused dose-dependent toxicity over a range of p24 levels (10-500 pg/ml. Significant interactions occurred with morphine at lower p24 levels (10 and 25 pg/ml, and GSK3β was implicated as a point of convergence. In the presence of glia, selective neurotoxic measures were significantly enhanced and interactions with morphine were also augmented, perhaps related to a decreased level of BDNF. Importantly, the arrest of neurite growth that occurred with exposure to HIV+sup was reversible unless neurons were continuously exposed to morphine. Thus, while reducing HIV-infection levels may be protective, ongoing exposure to opiates may limit recovery. Opiate interactions observed in this HIV-infective environment were similar, though not entirely concordant, with Tat/gp120 interactions reported previously, suggesting

  13. Inter-laboratory assessment of a prototype multiplex kit for determination of recent HIV-1 infection.

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    Kelly A Curtis

    Full Text Available BACKGROUND: Accurate and reliable laboratory-based assays are needed for estimating HIV-1 incidence from cross-sectional samples. We recently described the development of a customized, HIV-1-specific Bio-Plex assay that allows for the measurement of HIV-specific antibody levels and avidity to multiple analytes for improved HIV-1 incidence estimates. METHODS: To assess intra- and inter-laboratory assay performance, prototype multiplex kits were developed and evaluated by three distinct laboratories. Longitudinal seroconversion specimens were tested in parallel by each laboratory and kit performance was compared to that of an in-house assay. Additionally, the ability of the kit to distinguish recent from long-term HIV-1 infection, as compared to the in-house assay, was determined by comparing the reactivity of known recent (infected 12 months drug naïve specimens. RESULTS: Although the range of reactivity for each analyte varied between the prototype kit and in-house assay, a measurable distinction in reactivity between recent and long-term specimens was observed with both assays in all three laboratories. Additionally, kit performance was consistent between all three laboratories. The intra-assay coefficient of variation (CV, between sample replicates for all laboratories, ranged from 0.5% to 6.1%. The inter-laboratory CVs ranged from 8.5% to 21.3% for gp160-avidity index (a and gp120-normalized mean fluorescent intensity (MFI value (n, respectively. CONCLUSION: We demonstrate the feasibility of producing a multiplex kit for measuring HIV antibody levels and avidity, with the potential for improved incidence estimates based on multi-analyte algorithms. The availability of a commercial kit will facilitate the transfer of technology among diverse laboratories for widespread assay use.

  14. PVP-coated silver nanoparticles block the transmission of cell-free and cell-associated HIV-1 in human cervical culture

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    Rodriguez-Padilla Cristina

    2010-07-01

    Full Text Available Abstract Background Previous in vitro studies have demonstrated that polyvinylpyrrolidone coated silver nanoparticles (PVP-coated AgNPs have antiviral activity against HIV-1 at non-cytotoxic concentrations. These particles also demonstrate broad spectrum virucidal activity by preventing the interaction of HIV-1 gp120 and cellular CD4, thereby inhibiting fusion or entry of the virus into the host cell. In this study, we evaluated the antiviral activity of PVP-coated AgNPs as a potential topical vaginal microbicide to prevent transmission of HIV-1 infection using human cervical culture, an in vitro model that simulates in vivo conditions. Results When formulated into a non-spermicidal gel (Replens at a concentration of 0.15 mg/mL, PVP-coated AgNPs prevented the transmission of cell-associated HIV-1 and cell-free HIV-1 isolates. Importantly, PVP-coated AgNPs were not toxic to the explant, even when the cervical tissues were exposed continuously to 0.15 mg/mL of PVP-coated AgNPs for 48 h. Only 1 min of PVP-coated AgNPs pretreatment to the explant was required to prevent transmission of HIV-1. Pre-treatment of the cervical explant with 0.15 mg/mL PVP-coated AgNPs for 20 min followed by extensive washing prevented the transmission of HIV-1 in this model for 48 h. Conclusions A formulation of PVP-coated AgNPs homogenized in Replens gel acts rapidly to inhibit HIV-1 transmission after 1 min and offers long-lasting protection of the cervical tissue from infection for 48 h, with no evidence of cytotoxicity observed in the explants. Based on this data, PVP-coated AgNPs are a promising microbicidal candidate for use in topical vaginal/cervical agents to prevent HIV-1 transmission, and further research is warranted.

  15. Monoclonal Antibodies Specific for the V2, V3, CD4-Binding Site, and gp41 of HIV-1 Mediate Phagocytosis in a Dose-Dependent Manner.

    Science.gov (United States)

    Musich, Thomas; Li, Liuzhe; Liu, Lily; Zolla-Pazner, Susan; Robert-Guroff, Marjorie; Gorny, Miroslaw K

    2017-04-15

    In light of the weak or absent neutralizing activity mediated by anti-V2 monoclonal antibodies (MAbs), we tested whether they can mediate Ab-dependent cellular phagocytosis (ADCP), which is an important element of anti-HIV-1 immunity. We tested six anti-V2 MAbs and compared them with 21 MAbs specific for V3, the CD4-binding site (CD4bs), and gp41 derived from chronically HIV-1-infected individuals and produced by hybridoma cells. ADCP activity was measured by flow cytometry using uptake by THP-1 monocytic cells of fluorescent beads coated with gp120, gp41, BG505 SOSIP.664, or BG505 DS-SOSIP.664 complexed with MAbs. The measurement of ADCP activity by the area under the curve showed significantly higher activity of anti-gp41 MAbs than of the members of the three other groups of MAbs tested using beads coated with monomeric gp41 or gp120; anti-V2 MAbs were dominant compared to anti-V3 and anti-CD4bs MAbs against clade C gp120 ZM109 ADCP activity mediated by V2 and V3 MAbs was positive against stabilized DS-SOSIP.664 trimer but negligible against SOSIP.664 targets, suggesting that a closed envelope conformation better exposes the variable loops. Two IgG3 MAbs against the V2 and V3 regions displayed dominant ADCP activity compared to a panel of IgG1 MAbs. This superior ADCP activity was confirmed when two of three recombinant IgG3 anti-V2 MAbs were compared to their IgG1 counterparts. The study demonstrated dominant ADCP activity of anti-gp41 against monomers but not trimers, with some higher activity of anti-V2 MAbs than of anti-V3 and anti-CD4bs MAbs. The ability to mediate ADCP suggests a mechanism by which anti-HIV-1 envelope Abs can contribute to protective efficacy. IMPORTANCE Anti-V2 antibodies (Abs) correlated with reduced risk of HIV-1 infection in recipients of the RV144 vaccine, suggesting that they play a protective role, but a mechanism providing such protection remains to be determined. The rare and weak neutralizing activities of anti-V2 MAbs prompted us

  16. Performance of V3-based HIV-1 sero subtyping in HIV endemic areas

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    Lara Tavoschi

    2011-12-01

    Full Text Available HIV-1 serosubtyping based on reactivity to peptides from the V3 region of gp120 is a low-cost and easy to perform procedure often used in geographical areas with high prevalence and incidence of HIV infection. We evaluated the performance of V3-based serotyping on 148 sera from 118 HIV-1-infected individuals living in Uganda, with estimated dates of seroconversion. Of the 148 tested samples, 68 (46.0% specifically reacted with only one of the V3 peptides included in the test (SP, 64 (43.2% did not react with any peptide (NR and 16 (10.8% reacted with two or more peptides (CR. According to the estimated seroconversion date, the large majority of samples collected early after infection belonged to the NR group. These samples had also a low Avidity Index. In contrast, samples collected later after infection belonged mainly to CR and SP groups and had also a higher avidity index. These results indicate that the performance of V3-based assays depends on maturation of HIV-specific immune response and can be significantly lowered when these tests are carried out on specimens collected from recently infected individuals.

  17. HIV-1 replication in macrophages

    NARCIS (Netherlands)

    Kootstra, N.A.

    1999-01-01

    Lentiviruses such as the human immunodeficiency virus type 1 (HIV-1) are considered to be unique amongst the retroviruses due to their ability to replicate in macrophages, which are often referred to as non-dividing cells. The studies described in this thesis focus on the ability of HIV-1 to

  18. Analysis of memory B cell responses and isolation of novel monoclonal antibodies with neutralizing breadth from HIV-1-infected individuals.

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    Davide Corti

    2010-01-01

    Full Text Available The isolation of human monoclonal antibodies (mAbs that neutralize a broad spectrum of primary HIV-1 isolates and the characterization of the human neutralizing antibody B cell response to HIV-1 infection are important goals that are central to the design of an effective antibody-based vaccine.We immortalized IgG(+ memory B cells from individuals infected with diverse clades of HIV-1 and selected on the basis of plasma neutralization profiles that were cross-clade and relatively potent. Culture supernatants were screened using various recombinant forms of the envelope glycoproteins (Env in multiple parallel assays. We isolated 58 mAbs that were mapped to different Env surfaces, most of which showed neutralizing activity. One mAb in particular (HJ16 specific for a novel epitope proximal to the CD4 binding site on gp120 selectively neutralized a multi-clade panel of Tier-2 HIV-1 pseudoviruses, and demonstrated reactivity that was comparable in breadth, but distinct in neutralization specificity, to that of the other CD4 binding site-specific neutralizing mAb b12. A second mAb (HGN194 bound a conserved epitope in the V3 crown and neutralized all Tier-1 and a proportion of Tier-2 pseudoviruses tested, irrespective of clade. A third mAb (HK20 with broad neutralizing activity, particularly as a Fab fragment, recognized a highly conserved epitope in the HR-1 region of gp41, but showed striking assay-dependent selectivity in its activity.This study reveals that by using appropriate screening methods, a large proportion of memory B cells can be isolated that produce mAbs with HIV-1 neutralizing activity. Three of these mAbs show unusual breadth of neutralization and therefore add to the current panel of HIV-1 neutralizing antibodies with potential for passive protection and template-based vaccine design.

  19. Peptide-derivatized SB105-A10 dendrimer inhibits the infectivity of R5 and X4 HIV-1 strains in primary PBMCs and cervicovaginal histocultures.

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    Isabella Bon

    Full Text Available Peptide dendrimers are a class of molecules that exhibit a large array of biological effects including antiviral activity. In this report, we analyzed the antiviral activity of the peptide-derivatized SB105-A10 dendrimer, which is a tetra-branched dendrimer synthetized on a lysine core, in activated peripheral blood mononuclear cells (PBMCs that were challenged with reference and wild-type human immunodeficiency virus type 1 (HIV-1 strains. SB105-A10 inhibited infections by HIV-1 X4 and R5 strains, interfering with the early phases of the viral replication cycle. SB105-A10 targets heparan sulfate proteoglycans (HSPGs and, importantly, the surface plasmon resonance (SPR assay revealed that SB105-A10 strongly binds gp41 and gp120, most likely preventing HIV-1 attachment/entry through multiple mechanisms. Interestingly, the antiviral activity of SB105-A10 was also detectable in an organ-like structure of human cervicovaginal tissue, in which SB105-A10 inhibited the HIV-1ada R5 strain infection without altering the tissue viability. These results demonstrated the strong antiviral activity of SB105-A10 and suggest a potential microbicide use of this dendrimer to prevent the heterosexual transmission of HIV-1.

  20. Vaccine Induction of Antibodies Against a Structurally Heterogeneous Site of Immune Pressure within HIV-1 Envelope Protein Variable Regions 1 and 2

    Science.gov (United States)

    Liao, Hua-Xin; Bonsignori, Mattia; Alam, S. Munir; McLellan, Jason S.; Tomaras, Georgia D.; Moody, M. Anthony; Kozink, Daniel M.; Hwang, Kwan-Ki; Chen, Xi; Tsao, Chun-Yen; Liu, Pinghuang; Lu, Xiaozhi; Parks, Robert J.; Montefiori, David C.; Ferrari, Guido; Pollara, Justin; Rao, Mangala; Peachman, Kristina K.; Santra, Sampa; Letvin, Norman L.; Karasavvas, Nicos; Yang, Zhi-Yong; Dai, Kaifan; Pancera, Marie; Gorman, Jason; Wiehe, Kevin; Nicely, Nathan I.; Rerks-Ngarm, Supachai; Nitayaphan, Sorachai; Kaewkungwal, Jaranit; Pitisuttithum, Punnee; Tartaglia, James; Sinangil, Faruk; Kim, Jerome H.; Michael, Nelson L.; Kepler, Thomas B.; Kwong, Peter D.; Mascola, John R.; Nabel, Gary J.; Pinter, Abraham; Zolla-Pazner, Susan; Haynes, Barton F.

    2013-01-01

    Summary The RV144 HIV-1 trial of the canary pox vector (ALVAC-HIV) plus the gp120 AIDSVAX B/E vaccine demonstrated an estimated efficacy of 31%, that correlated directly with antibodies to HIV-1 envelope variable regions 1 and 2 (V1–V2). Genetic analysis of trial viruses revealed increased vaccine efficacy against viruses matching the vaccine strain at V2 residue 169. Here, we isolated four V2 monoclonal antibodies from RV144 vaccinees that recognize residue 169, neutralize laboratory-adapted HIV-1, and mediate killing of field isolate HIV-1-infected CD4+ T cells. Crystal structures of two of the V2 antibodies demonstrated residue 169 can exist within divergent helical and loop conformations, which contrasted dramatically with the beta strand conformation previously observed with a broadly neutralizing antibody PG9. Thus, RV144 vaccine-induced immune pressure appears to target a region that may be both sequence variable and structurally polymorphic. Variation may signal sites of HIV-1 envelope vulnerability, providing vaccine designers with new options. PMID:23313589

  1. McCoy cell line as a possible model containing CD4+ receptors for the study of HIV-1 replication

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    Nogueira Yeda L.

    2003-01-01

    Full Text Available Several studies have recently shown the use of recombinant rabies virus as potential vector-viral vaccine for HIV-1. The sequence homology between gp 120 and rabies virus glycoprotein has been reported. The McCoy cell line has therefore been used to show CD4+ or CD4+ like receptors. Samples of HIV-1 were isolated, when plasma of HIV-1 positive patients was inoculated in the McCoy cell line. The virus infection was then studied during successive virus passages. The proteins released in the extra cellular medium were checked for protein activity, by exposure to SDS Electrophoresis and blotting to nitro-cellulose filter, then reacting with sera of HIV positive and negative patients. Successive passages were performed, and showed viral replication, membrane permeabilization, the syncytium formation, and the cellular lysis (cytopathic effect. Flow cytometry analysis shows clear evidence that CD4+ receptors are present in this cell line, which enhances the likelihood of easy isolation and replication of HIV. The results observed allow the use of this cell line as a possible model for isolating HIV, as well as for carrying out studies of the dynamics of viral infection in several situations, including exposure to drugs in pharmacological studies, and possibly studies and analyses of the immune response in vaccine therapies.

  2. Protein kinase C-delta regulates HIV-1 replication at an early post-entry step in macrophages

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    Contreras Xavier

    2012-05-01

    Full Text Available Abstract Background Macrophages, which are CD4 and CCR5 positive, can sustain HIV-1 replication for long periods of time. Thus, these cells play critical roles in the transmission, dissemination and persistence of viral infection. Of note, current antiviral therapies do not target macrophages efficiently. Previously, it was demonstrated that interactions between CCR5 and gp120 stimulate PKC. However, the PKC isozymes involved were not identified. Results In this study, we identified PKC-delta as a major cellular cofactor for HIV-1 replication in macrophages. Indeed, PKC-delta was stimulated following the interaction between the virus and its target cell. Moreover, inhibition of PKC-delta blocked the replication of R5-tropic viruses in primary human macrophages. However, this inhibition did not have significant effects on receptor and co-receptor expression or fusion. Additionally, it did not affect the formation of the early reverse transcription product containing R/U5 sequences, but did inhibit the synthesis of subsequent cDNAs. Importantly, the inhibition of PKC-delta altered the redistribution of actin, a cellular cofactor whose requirement for the completion of reverse transcription was previously established. It also prevented the association of the reverse transcription complex with the cytoskeleton. Conclusion This work highlights the importance of PKC-delta during early steps of the replicative cycle of HIV-1 in human macrophages.

  3. α1Proteinase inhibitor regulates CD4+ lymphocyte levels and is rate limiting in HIV-1 disease.

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    Cynthia L Bristow

    Full Text Available BACKGROUND: The regulation of adult stem cell migration through human hematopoietic tissue involves the chemokine CXCL12 (SDF-1 and its receptor CXCR4 (CD184. In addition, human leukocyte elastase (HLE plays a key role. When HLE is located on the cell surface (HLE(CS, it acts not as a proteinase, but as a receptor for α(1proteinase inhibitor (α(1PI, α(1antitrypsin, SerpinA1. Binding of α(1PI to HLE(CS forms a motogenic complex. We previously demonstrated that α(1PI deficiency attends HIV-1 disease and that α(1PI augmentation produces increased numbers of immunocompetent circulating CD4(+ lymphocytes. Herein we investigated the mechanism underlying the α(1PI deficiency that attends HIV-1 infection. METHODS AND FINDINGS: Active α(1PI in HIV-1 subjects (median 17 µM, n = 35 was significantly below normal (median 36 µM, p220 CD4 cells/µl, CD4(+ lymphocytes were correlated solely with active α(1PI (r(2 = 0.93, p<0.0001, n = 26. The monoclonal anti-HIV-1 gp120 antibody 3F5 present in HIV-1 patient blood is shown to bind and inactivate human α(1PI. Chimpanzee α(1PI differs from human α(1PI by a single amino acid within the 3F5-binding epitope. Unlike human α(1PI, chimpanzee α(1PI did not bind 3F5 or become depleted following HIV-1 challenge, consistent with the normal CD4(+ lymphocyte levels and benign syndrome of HIV-1 infected chimpanzees. The presence of IgG-α(1PI immune complexes correlated with decreased CD4(+ lymphocytes in HIV-1 subjects. CONCLUSIONS: This report identifies an autoimmune component of HIV-1 disease that can be overcome therapeutically. Importantly, results identify an achievable vaccine modification with the novel objective to protect against AIDS as opposed to the current objective to protect against HIV-1 infection.

  4. Slaying the Trojan horse: natural killer cells exhibit robust anti-HIV-1 antibody-dependent activation and cytolysis against allogeneic T cells.

    Science.gov (United States)

    Gooneratne, Shayarana L; Richard, Jonathan; Lee, Wen Shi; Finzi, Andrés; Kent, Stephen J; Parsons, Matthew S

    2015-01-01

    with HIV-1 gp120 were capable of activating NK cells and/or trigging cytolysis of the allogeneic target cells. This suggests ADCC may be able to assist in preventing infection with cell-associated HIV-1. In order to fully utilize NK cell-mediated Ab-dependent effector functions, it might also be important that educated NK cells, which hold the highest activation potential, can become activated against targets bearing HIV-1 antigens and expressing the ligands for self-inhibitory receptors. Here, we show that with Ab-dependent stimulation, NK cells expressing inhibitory receptors can mediate robust activation against targets expressing the ligands for those receptors. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  5. Efficient single tobamoviral vector-based bioproduction of broadly neutralizing anti-HIV-1 monoclonal antibody VRC01 in Nicotiana benthamiana plants and utility of VRC01 in combination microbicides.

    Science.gov (United States)

    Hamorsky, Krystal Teasley; Grooms-Williams, Tiffany W; Husk, Adam S; Bennett, Lauren J; Palmer, Kenneth E; Matoba, Nobuyuki

    2013-05-01

    Broadly neutralizing monoclonal antibodies (bnMAbs) may offer powerful tools for HIV-1 preexposure prophylaxis, such as topical microbicides. However, this option is hampered due to expensive MAb biomanufacturing based on mammalian cell culture. To address this issue, we developed a new production system for bnMAb VRC01 in Nicotiana benthamiana plants using a tobamovirus replicon vector. Unlike conventional two-vector-based expression, this system was designed to overexpress full-length IgG1 from a single polypeptide by means of kex2p-like enzyme recognition sites introduced between the heavy and light chains. An enzyme-linked immunosorbent assay (ELISA) revealed that gp120-binding VRC01 IgG1 was maximally accumulated on 5 to 7 days following vector inoculation, yielding ~150 mg of the bnMAb per kg of fresh leaf material. The plant-made VRC01 (VRC01p) was efficiently purified by protein A affinity followed by hydrophobic-interaction chromatography. ELISA, surface plasmon resonance, and an HIV-1 neutralization assay demonstrated that VRC01p has gp120-binding affinity and HIV-1-neutralization capacity virtually identical to the human-cell-produced counterpart. To advance VRC01p's use in topical microbicides, we analyzed combinations of the bnMAb with other microbicide candidates holding distinct antiviral mechanisms in an HIV-1 neutralization assay. VRC01p exhibited clear synergy with the antiviral lectin griffithsin, the CCR5 antagonist maraviroc, and the reverse transcriptase inhibitor tenofovir in multiple CCR5-tropic HIV-1 strains from clades A, B, and C. In summary, VRC01p is amenable to robust, rapid, and large-scale production and may be developed as an active component in combination microbicides with other anti-HIV agents such as antiviral lectins, CCR5 antagonists, and reverse transcriptase inhibitors.

  6. HIV-1 Env DNA vaccine plus protein boost delivered by EP expands B- and T-cell responses and neutralizing phenotype in vivo.

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    Kar Muthumani

    Full Text Available An effective HIV vaccine will most likely require the induction of strong T-cell responses, broadly neutralizing antibodies (bNAbs, and the elicitation of antibody-dependent cellular cytotoxicity (ADCC. Previously, we demonstrated the induction of strong HIV/SIV cellular immune responses in macaques and humans using synthetic consensus DNA immunogens delivered via adaptive electroporation (EP. However, the ability of this improved DNA approach to prime for relevant antibody responses has not been previously studied. Here, we investigate the immunogenicity of consensus DNA constructs encoding gp140 sequences from HIV-1 subtypes A, B, C and D in a DNA prime-protein boost vaccine regimen. Mice and guinea pigs were primed with single- and multi-clade DNA via EP and boosted with recombinant gp120 protein. Sera were analyzed for gp120 binding and induction of neutralizing antibody activity. Immunization with recombinant Env protein alone induced low-titer binding antibodies with limited neutralization breath. In contrast, the synthetic DNA prime-protein boost protocol induced significantly higher antibody binding titers. Furthermore, sera from DNA prime-protein boost groups were able to neutralize a broader range of viruses in a panel of tier 1 clade B viruses as well as multiple tier 1 clade A and clade C viruses. Further investigation of synthetic DNA prime plus adaptive EP plus protein boost appears warranted.

  7. Somatic populations of PGT135-137 HIV-1-neutralizing antibodies identified by 454 pyrosequencing and bioinformatics

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    Jiang eZhu

    2012-09-01

    Full Text Available Select HIV-1-infected individuals develop sera capable of neutralizing diverse viral strains. The molecular basis of this neutralization is currently being deciphered by the isolation of HIV-1-neutralizing antibodies. In one infected donor, three neutralizing antibodies, PGT135-137, were identified by assessment of neutralization from individually sorted B cells and found to recognize an epitope containing an N-linked glycan at residue 332 on HIV-1 gp120. Here we use deep sequencing and bioinformatics methods to interrogate the B cell record of this donor to gain a more complete understanding of the humoral immune response. PGT135-137-gene family-specific primers were used to amplify heavy and light chain-variable domain sequences. 454 pyrosequencing produced 141,298 heavy-chain sequences of IGHV4-39 origin and 87,229 light-chain sequences of IGKV3-15 origin. A number of heavy and light chain sequences of ~90% identity to PGT137, several to PGT136, and none of high identity to PGT135 were identified. After expansion of these sequences to include close phylogenetic relatives, a total of 202 heavy-chain sequences and 72 light-chain sequences were identified. These sequences were clustered into populations of 95% identity comprising 15 for heavy chain and 10 for light chain, and a select sequence from each population was synthesized and reconstituted with a PGT137-partner chain. Reconstituted antibodies showed varied neutralization phenotypes for HIV-1 clade A and D isolates. Sequence diversity of the antibody population represented by these tested sequences was notably higher than observed with a 454 pyrosequencing-control analysis on 10 antibodies of defined sequence, suggesting that this diversity results primarily from somatic maturation. Our results thus provide an example of how pathogens like HIV-1 are opposed by a varied humoral immune response, derived from intrinsic mechanisms of antibody development, and embodied by somatic populations

  8. Selection pressure on HIV-1 envelope by broadly neutralizing antibodies to the conserved CD4-binding site.

    Science.gov (United States)

    Wu, Xueling; Wang, Charlene; O'Dell, Sijy; Li, Yuxing; Keele, Brandon F; Yang, Zhongjia; Imamichi, Hiromi; Doria-Rose, Nicole; Hoxie, James A; Connors, Mark; Shaw, George M; Wyatt, Richard T; Mascola, John R

    2012-05-01

    The monoclonal antibody (MAb) VRC01 was isolated from a slowly progressing HIV-1-infected donor and was shown to neutralize diverse HIV-1 strains by binding to the conserved CD4 binding site (CD4bs) of gp120. To better understand the virologic factors associated with such antibody development, we characterized HIV-1 envelope (Env) variants from this donor and five other donors who developed broadly neutralizing antibodies. A total of 473 env sequences were obtained by single-genome amplification, and 100 representative env clones were expressed and tested for entry and neutralization sensitivity. While VRC01 neutralizes about 90% of the genetically diverse heterologous HIV-1 strains tested, only selective archival Env variants from the VRC01 donor were sensitive to VRC01 and all of the Env variants derived from the donor plasma were resistant, indicating strong antibody-based selection pressure. Despite their resistance to this broadly reactive MAb that partially mimics CD4, all Env variants required CD4 for entry. Three other CD4bs MAbs from the same donor were able to neutralize some VRC01 escape variants, suggesting that CD4bs antibodies continued to evolve in response to viral escape. We also observed a relatively high percentage of VRC01-resistant Env clones in the plasma of four of five additional broadly neutralizing donors, suggesting the presence of CD4bs-directed neutralizing antibodies in these donors. In total, these data indicate that the CD4bs-directed neutralizing antibodies exert ongoing selection pressure on the conserved CD4bs epitope of HIV-1 Env.

  9. Structure of an N276-Dependent HIV-1 Neutralizing Antibody Targeting a Rare V5 Glycan Hole Adjacent to the CD4 Binding Site

    Energy Technology Data Exchange (ETDEWEB)

    Wibmer, Constantinos Kurt; Gorman, Jason; Anthony, Colin S.; Mkhize, Nonhlanhla N.; Druz, Aliaksandr; York, Talita; Schmidt, Stephen D.; Labuschagne, Phillip; Louder, Mark K.; Bailer, Robert T.; Karim, Salim S. Abdool; Mascola, John R.; Williamson, Carolyn; Moore, Penny L.; Kwong, Peter D.; Morris, Lynn (NHLS-South Africa); (NIH); (Witwatersrand); (KwaZulu-Natal)

    2016-08-31

    ABSTRACT

    All HIV-1-infected individuals develop strain-specific neutralizing antibodies to their infecting virus, which in some cases mature into broadly neutralizing antibodies. Defining the epitopes of strain-specific antibodies that overlap conserved sites of vulnerability might provide mechanistic insights into how broadly neutralizing antibodies arise. We previously described an HIV-1 clade C-infected donor, CAP257, who developed broadly neutralizing plasma antibodies targeting an N276 glycan-dependent epitope in the CD4 binding site. The initial CD4 binding site response potently neutralized the heterologous tier 2 clade B viral strain RHPA, which was used to design resurfaced gp120 antigens for single-B-cell sorting. Here we report the isolation and structural characterization of CAP257-RH1, an N276 glycan-dependent CD4 binding site antibody representative of the early CD4 binding site plasma response in donor CAP257. The cocrystal structure of CAP257-RH1 bound to RHPA gp120 revealed critical interactions with the N276 glycan, loop D, and V5, but not with aspartic acid 368, similarly to HJ16 and 179NC75. The CAP257-RH1 monoclonal antibody was derived from the immunoglobulin-variable IGHV3-33 and IGLV3-10 genes and neutralized RHPA but not the transmitted/founder virus from donor CAP257. Its narrow neutralization breadth was attributed to a binding angle that was incompatible with glycosylated V5 loops present in almost all HIV-1 strains, including the CAP257 transmitted/founder virus. Deep sequencing of autologous CAP257 viruses, however, revealed minority variants early in infection that lacked V5 glycans. These glycan-free V5 loops are unusual holes in the glycan shield that may have been necessary for initiating this N276 glycan-dependent CD4 binding site B-cell lineage.

    IMPORTANCEThe conserved CD4 binding site on gp120 is a major target for HIV-1 vaccine design, but key events in the elicitation and maturation of

  10. Prevalence, estimated HIV-1 incidence and viral diversity among people seeking voluntary counseling and testing services in Rio de Janeiro, Brazil

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    Bastos Francisco I

    2010-07-01

    Full Text Available Abstract Background BED-EIA HIV-1 Incidence Test (BED-CEIA has been described as a tool to discriminate recent (RS from long-term (LTS seroconversion of HIV-1 infection, contributing to a better understanding of the dynamics of the HIV/AIDS epidemic over time. This study determined the prevalence, estimated incidence and HIV-1 subtype infection among individuals seeking testing in Voluntary Counseling and Testing centers (VCTs from Rio de Janeiro, Brazil. Methods Demographics and behavioral data were obtained from 434 individuals, diagnosed as HIV-positive among 9,008 volunteers screened from November 2004 to October 2005 in three VCTs located in the Rio de Janeiro Metropolitan area, Brazil. BED-CEIA protocol was performed to identify RS. DNA samples from RS and a subset of LTS (under a proportion of 1:2 were selected for gp120 C2-V3 and pol (protease and reverse transcriptase regions genomic sequencing. Results Overall HIV-1 prevalence was 4.8%. Sixty-one of 434 seropositive individuals were classified as RS, corresponding to an incidence rate of 1.68%/year (95%CI 1.26% -2.10%. Estimated incidence between Men Who Have Sex with Men (MSM was 11 times higher than among heterosexual men and 55% of the new cases were identified in volunteers aged 25-40 years. A similar distribution of different HIV-1 subtypes was found among RS and LTS. Conclusions Our data suggest that prevention for MSM remains a challenge and efforts focusing on prevention targeting this population should be prioritized. No significant changes in HIV-1 subtypes were observed among the RS and LTS subgroups. One case of HIV-1 AUK (pol/A (env recombinant genome was detected for the first time in Brazil.

  11. Prevalence, estimated HIV-1 incidence and viral diversity among people seeking voluntary counseling and testing services in Rio de Janeiro, Brazil.

    Science.gov (United States)

    de Castro, Carlos A Velasco; Grinsztejn, Beatriz; Veloso, Valdiléa G; Bastos, Francisco I; Pilotto, José H; Morgado, Mariza G

    2010-07-28

    BED-EIA HIV-1 Incidence Test (BED-CEIA) has been described as a tool to discriminate recent (RS) from long-term (LTS) seroconversion of HIV-1 infection, contributing to a better understanding of the dynamics of the HIV/AIDS epidemic over time. This study determined the prevalence, estimated incidence and HIV-1 subtype infection among individuals seeking testing in Voluntary Counseling and Testing centers (VCTs) from Rio de Janeiro, Brazil. Demographics and behavioral data were obtained from 434 individuals, diagnosed as HIV-positive among 9,008 volunteers screened from November 2004 to October 2005 in three VCTs located in the Rio de Janeiro Metropolitan area, Brazil. BED-CEIA protocol was performed to identify RS. DNA samples from RS and a subset of LTS (under a proportion of 1:2) were selected for gp120 C2-V3 and pol (protease and reverse transcriptase) regions genomic sequencing. Overall HIV-1 prevalence was 4.8%. Sixty-one of 434 seropositive individuals were classified as RS, corresponding to an incidence rate of 1.68%/year (95%CI 1.26% -2.10%). Estimated incidence between Men Who Have Sex with Men (MSM) was 11 times higher than among heterosexual men and 55% of the new cases were identified in volunteers aged 25-40 years. A similar distribution of different HIV-1 subtypes was found among RS and LTS. Our data suggest that prevention for MSM remains a challenge and efforts focusing on prevention targeting this population should be prioritized. No significant changes in HIV-1 subtypes were observed among the RS and LTS subgroups. One case of HIV-1 AUK (pol)/A (env) recombinant genome was detected for the first time in Brazil.

  12. Escape from autologous neutralizing antibodies in acute/early subtype C HIV-1 infection requires multiple pathways.

    Science.gov (United States)

    Rong, Rong; Li, Bing; Lynch, Rebecca M; Haaland, Richard E; Murphy, Megan K; Mulenga, Joseph; Allen, Susan A; Pinter, Abraham; Shaw, George M; Hunter, Eric; Robinson, James E; Gnanakaran, S; Derdeyn, Cynthia A

    2009-09-01

    One aim for an HIV vaccine is to elicit neutralizing antibodies (Nab) that can limit replication of genetically diverse viruses and prevent establishment of a new infection. Thus, identifying the strengths and weaknesses of Nab during the early stages of natural infection could prove useful in achieving this goal. Here we demonstrate that viral escape readily occurred despite the development of high titer autologous Nab in two subjects with acute/early subtype C infection. To provide a detailed portrayal of the escape pathways, Nab resistant variants identified at multiple time points were used to create a series of envelope (Env) glycoprotein chimeras and mutants within the background of a corresponding newly transmitted Env. In one subject, Nab escape was driven predominantly by changes in the region of gp120 that extends from the beginning of the V3 domain to the end of the V5 domain (V3V5). However, Nab escape pathways in this subject oscillated and at times required cooperation between V1V2 and the gp41 ectodomain. In the second subject, escape was driven by changes in V1V2. This V1V2-dependent escape pathway was retained over time, and its utility was reflected in the virus's ability to escape from two distinct monoclonal antibodies (Mabs) derived from this same patient via introduction of a single potential N-linked glycosylation site in V2. Spatial representation of the sequence changes in gp120 suggested that selective pressure acted upon the same regions of Env in these two subjects, even though the Env domains that drove escape were different. Together the findings argue that a single mutational pathway is not sufficient to confer escape in early subtype C HIV-1 infection, and support a model in which multiple strategies, including potential glycan shifts, direct alteration of an epitope sequence, and cooperative Env domain conformational masking, are used to evade neutralization.

  13. Sensitivity of HIV-1 to neutralization by antibodies against O-linked carbohydrate epitopes despite deletion of O-glycosylation signals in the V3 loop

    DEFF Research Database (Denmark)

    Hansen, J E; Jansson, B; Gram, G J

    1996-01-01

    It has been suggested that threonine or serine residues in the V3 loop of HIV-1 gp120 are glycosylated with the short-chain O-linked oligosaccharides Tn or sialosyl-Tn that function as epitopes for broadly neutralizing carbohydrate specific antibodies. In this study we examined whether mutation....... Additionally, one of these T-A mutants (T308A) also abrogated the signal for N-glycosylation at N306 inside the V3-loop. The mutant clones were compared with the wild type virus as to sensitivity to neutralization with monoclonal and polyclonal antibodies specific for the tip of the V3 loop of BRU or for the O...

  14. Improving the Immunogenicity of Native-like HIV-1 Envelope Trimers by Hyperstabilization

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    Alba Torrents de la Peña

    2017-08-01

    Full Text Available The production of native-like recombinant versions of the HIV-1 envelope glycoprotein (Env trimer requires overcoming the natural flexibility and instability of the complex. The engineered BG505 SOSIP.664 trimer mimics the structure and antigenicity of native Env. Here, we describe how the introduction of new disulfide bonds between the glycoprotein (gp120 and gp41 subunits of SOSIP trimers of the BG505 and other genotypes improves their stability and antigenicity, reduces their conformational flexibility, and helps maintain them in the unliganded conformation. The resulting next-generation SOSIP.v5 trimers induce strong autologous tier-2 neutralizing antibody (NAb responses in rabbits. In addition, the BG505 SOSIP.v6 trimers induced weak heterologous NAb responses against a subset of tier-2 viruses that were not elicited by the prototype BG505 SOSIP.664. These stabilization methods can be applied to trimers from multiple genotypes as components of multivalent vaccines aimed at inducing broadly NAbs (bNAbs.

  15. HIV-1 assembly in macrophages

    Directory of Open Access Journals (Sweden)

    Benaroch Philippe

    2010-04-01

    Full Text Available Abstract The molecular mechanisms involved in the assembly of newly synthesized Human Immunodeficiency Virus (HIV particles are poorly understood. Most of the work on HIV-1 assembly has been performed in T cells in which viral particle budding and assembly take place at the plasma membrane. In contrast, few studies have been performed on macrophages, the other major target of HIV-1. Infected macrophages represent a viral reservoir and probably play a key role in HIV-1 physiopathology. Indeed macrophages retain infectious particles for long periods of time, keeping them protected from anti-viral immune response or drug treatments. Here, we present an overview of what is known about HIV-1 assembly in macrophages as compared to T lymphocytes or cell lines. Early electron microscopy studies suggested that viral assembly takes place at the limiting membrane of an intracellular compartment in macrophages and not at the plasma membrane as in T cells. This was first considered as a late endosomal compartment in which viral budding seems to be similar to the process of vesicle release into multi-vesicular bodies. This view was notably supported by a large body of evidence involving the ESCRT (Endosomal Sorting Complex Required for Transport machinery in HIV-1 budding, the observation of viral budding profiles in such compartments by immuno-electron microscopy, and the presence of late endosomal markers associated with macrophage-derived virions. However, this model needs to be revisited as recent data indicate that the viral compartment has a neutral pH and can be connected to the plasma membrane via very thin micro-channels. To date, the exact nature and biogenesis of the HIV assembly compartment in macrophages remains elusive. Many cellular proteins potentially involved in the late phases of HIV-1 cycle have been identified; and, recently, the list has grown rapidly with the publication of four independent genome-wide screens. However, their respective

  16. A next-generation cleaved, soluble HIV-1 Env trimer, BG505 SOSIP.664 gp140, expresses multiple epitopes for broadly neutralizing but not non-neutralizing antibodies.

    Science.gov (United States)

    Sanders, Rogier W; Derking, Ronald; Cupo, Albert; Julien, Jean-Philippe; Yasmeen, Anila; de Val, Natalia; Kim, Helen J; Blattner, Claudia; de la Peña, Alba Torrents; Korzun, Jacob; Golabek, Michael; de Los Reyes, Kevin; Ketas, Thomas J; van Gils, Marit J; King, C Richter; Wilson, Ian A; Ward, Andrew B; Klasse, P J; Moore, John P

    2013-09-01

    A desirable but as yet unachieved property of a human immunodeficiency virus type 1 (HIV-1) vaccine candidate is the ability to induce broadly neutralizing antibodies (bNAbs). One approach to the problem is to create trimeric mimics of the native envelope glycoprotein (Env) spike that expose as many bNAb epitopes as possible, while occluding those for non-neutralizing antibodies (non-NAbs). Here, we describe the design and properties of soluble, cleaved SOSIP.664 gp140 trimers based on the subtype A transmitted/founder strain, BG505. These trimers are highly stable, more so even than the corresponding gp120 monomer, as judged by differential scanning calorimetry. They are also homogenous and closely resemble native virus spikes when visualized by negative stain electron microscopy (EM). We used several techniques, including ELISA and surface plasmon resonance (SPR), to determine the relationship between the ability of monoclonal antibodies (MAbs) to bind the soluble trimers and neutralize the corresponding virus. In general, the concordance was excellent, in that virtually all bNAbs against multiple neutralizing epitopes on HIV-1 Env were highly reactive with the BG505 SOSIP.664 gp140 trimers, including quaternary epitopes (CH01, PG9, PG16 and PGT145). Conversely, non-NAbs to the CD4-binding site, CD4-induced epitopes or gp41ECTO did not react with the trimers, even when their epitopes were present on simpler forms of Env (e.g. gp120 monomers or dissociated gp41 subunits). Three non-neutralizing MAbs to V3 epitopes did, however, react strongly with the trimers but only by ELISA, and not at all by SPR and to only a limited extent by EM. These new soluble trimers are useful for structural studies and are being assessed for their performance as immunogens.

  17. A next-generation cleaved, soluble HIV-1 Env trimer, BG505 SOSIP.664 gp140, expresses multiple epitopes for broadly neutralizing but not non-neutralizing antibodies.

    Directory of Open Access Journals (Sweden)

    Rogier W Sanders

    2013-09-01

    Full Text Available A desirable but as yet unachieved property of a human immunodeficiency virus type 1 (HIV-1 vaccine candidate is the ability to induce broadly neutralizing antibodies (bNAbs. One approach to the problem is to create trimeric mimics of the native envelope glycoprotein (Env spike that expose as many bNAb epitopes as possible, while occluding those for non-neutralizing antibodies (non-NAbs. Here, we describe the design and properties of soluble, cleaved SOSIP.664 gp140 trimers based on the subtype A transmitted/founder strain, BG505. These trimers are highly stable, more so even than the corresponding gp120 monomer, as judged by differential scanning calorimetry. They are also homogenous and closely resemble native virus spikes when visualized by negative stain electron microscopy (EM. We used several techniques, including ELISA and surface plasmon resonance (SPR, to determine the relationship between the ability of monoclonal antibodies (MAbs to bind the soluble trimers and neutralize the corresponding virus. In general, the concordance was excellent, in that virtually all bNAbs against multiple neutralizing epitopes on HIV-1 Env were highly reactive with the BG505 SOSIP.664 gp140 trimers, including quaternary epitopes (CH01, PG9, PG16 and PGT145. Conversely, non-NAbs to the CD4-binding site, CD4-induced epitopes or gp41ECTO did not react with the trimers, even when their epitopes were present on simpler forms of Env (e.g. gp120 monomers or dissociated gp41 subunits. Three non-neutralizing MAbs to V3 epitopes did, however, react strongly with the trimers but only by ELISA, and not at all by SPR and to only a limited extent by EM. These new soluble trimers are useful for structural studies and are being assessed for their performance as immunogens.

  18. Functional and immunochemical cross-reactivity of V2-specific monoclonal antibodies from HIV-1-infected individuals

    Energy Technology Data Exchange (ETDEWEB)

    Gorny, Miroslaw K.; Pan, Ruimin; Williams, Constance; Wang, Xiao-Hong; Volsky, Barbara; O; Neal, Timothy; Spurrier, Brett; Sampson, Jared M.; Li, Liuzhe; Seaman, Michael S.; Kong, Xiang-Peng; Zolla-Pazner, Susan (Harvard-Med); (VA); (NYUSM)

    2012-05-18

    The recent analysis of the first successful RV144 vaccine trial revealed that a high titer of plasma anti-V2 antibodies (Abs) correlated with a decreased risk of HIV-1 infection in vaccine recipients. To understand the mechanism of immune correlates, we studied seven anti-V2 monoclonal Abs (mAbs) developed from HIV-1 infected individuals. The V2 mAbs target conserved epitopes, including the binding site for {alpha}4{beta}7 integrin, and are broadly cross-reactive with various gp120 proteins. Preferential usage of the VH1-69 gene by V2 mAbs may depend on selection by the same antigenic structure. Six of seven V2 mAbs weakly neutralized four to eight of the 41 pseudoviruses tested and resistance to neutralization was correlated with longer V2 domains. The data suggest the presence of shared, conserved structural elements in the V2 loop, and these can be used in the design of vaccine immunogens inducing broadly reactive Abs with anti-viral activities.

  19. Modeling HIV-1 Induced Neuroinflammation in Mice: Role of Platelets in Mediating Blood-Brain Barrier Dysfunction.

    Directory of Open Access Journals (Sweden)

    Letitia D Jones

    Full Text Available The number of HIV-1 positive individuals developing some form of HIV-associated neurocognitive disorder (HAND is increasing. In these individuals, the integrity of the blood-brain barrier (BBB is compromised due to an increase in exposure to pro-inflammatory mediators, viral proteins, and virus released from infected cells. It has been shown that soluble CD40L (sCD40L is released upon platelet activation and is an important mediator of the pathogenesis of HAND but the underlying mechanisms are unclear, emphasizing the need of an effective animal model. Here, we have utilized a novel animal model in which wild-type (WT mice were infected with EcoHIV; a derivative of HIV-1 that contains a substitution of envelope protein gp120 with that of gp80 derived from murine leukemia virus-1 (MuLV-1. As early as two-weeks post-infection, EcoHIV led to increased permeability of the BBB associated with decreased expression of tight junction protein claudin-5, in CD40L and platelet activation-dependent manner. Treatment with an antiplatelet drug, eptifibatide, in EcoHIV-infected mice normalized BBB function, sCD40L release and platelet activity, thus implicating platelet activation and platelet-derived CD40L in virally induced BBB dysfunction. Our results also validate and underscore the importance of EcoHIV infection mouse model as a tool to explore therapeutic targets for HAND.

  20. Non-random pre-transcriptional evolution in HIV-1: a refutation of the foundational conditions for neutral evolution

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    Carlos Y Valenzuela

    2009-01-01

    Full Text Available The complete base sequence of HIV-1 virus and GP120 ENV gene were analyzed to establish their distance to the expected neutral random sequence. An especial methodology was devised to achieve this aim. Analyses included: a proportion of dinucleotides (signatures; b homogeneity in the distribution of dinucleotides and bases (isochores by dividing both segments in ten and three sub-segments, respectively; c probability of runs of bases and No-bases according to the Bose-Einstein distribution. The analyses showed a huge deviation from the random distribution expected from neutral evolution and neutral-neighbor influence of nucleotide sites. The most significant result is the tremendous lack of CG dinucleotides (p < 10-50, a selective trait of eukaryote and not of single stranded RNA virus genomes. Results not only refute neutral evolution and neutral neighbor influence, but also strongly indicate that any base at any nucleotide site correlates with all the viral genome or sub-segments. These results suggest that evolution of HIV-1 is pan-selective rather than neutral or nearly neutral.

  1. Human Non-neutralizing HIV-1 Envelope Monoclonal Antibodies Limit the Number of Founder Viruses during SHIV Mucosal Infection in Rhesus Macaques.

    Science.gov (United States)

    Santra, Sampa; Tomaras, Georgia D; Warrier, Ranjit; Nicely, Nathan I; Liao, Hua-Xin; Pollara, Justin; Liu, Pinghuang; Alam, S Munir; Zhang, Ruijun; Cocklin, Sarah L; Shen, Xiaoying; Duffy, Ryan; Xia, Shi-Mao; Schutte, Robert J; Pemble Iv, Charles W; Dennison, S Moses; Li, Hui; Chao, Andrew; Vidnovic, Kora; Evans, Abbey; Klein, Katja; Kumar, Amit; Robinson, James; Landucci, Gary; Forthal, Donald N; Montefiori, David C; Kaewkungwal, Jaranit; Nitayaphan, Sorachai; Pitisuttithum, Punnee; Rerks-Ngarm, Supachai; Robb, Merlin L; Michael, Nelson L; Kim, Jerome H; Soderberg, Kelly A; Giorgi, Elena E; Blair, Lily; Korber, Bette T; Moog, Christiane; Shattock, Robin J; Letvin, Norman L; Schmitz, Joern E; Moody, M A; Gao, Feng; Ferrari, Guido; Shaw, George M; Haynes, Barton F

    2015-08-01

    HIV-1 mucosal transmission begins with virus or virus-infected cells moving through mucus across mucosal epithelium to infect CD4+ T cells. Although broadly neutralizing antibodies (bnAbs) are the type of HIV-1 antibodies that are most likely protective, they are not induced with current vaccine candidates. In contrast, antibodies that do not neutralize primary HIV-1 strains in the TZM-bl infection assay are readily induced by current vaccine candidates and have also been implicated as secondary correlates of decreased HIV-1 risk in the RV144 vaccine efficacy trial. Here, we have studied the capacity of anti-Env monoclonal antibodies (mAbs) against either the immunodominant region of gp41 (7B2 IgG1), the first constant region of gp120 (A32 IgG1), or the third variable loop (V3) of gp120 (CH22 IgG1) to modulate in vivo rectal mucosal transmission of a high-dose simian-human immunodeficiency virus (SHIV-BaL) in rhesus macaques. 7B2 IgG1 or A32 IgG1, each containing mutations to enhance Fc function, was administered passively to rhesus macaques but afforded no protection against productive clinical infection while the positive control antibody CH22 IgG1 prevented infection in 4 of 6 animals. Enumeration of transmitted/founder (T/F) viruses revealed that passive infusion of each of the three antibodies significantly reduced the number of T/F genomes. Thus, some antibodies that bind HIV-1 Env but fail to neutralize virus in traditional neutralization assays may limit the number of T/F viruses involved in transmission without leading to enhancement of viral infection. For one of these mAbs, gp41 mAb 7B2, we provide the first co-crystal structure in complex with a common cyclical loop motif demonstrated to be critical for infection by other retroviruses.

  2. Phase 1 safety and immunogenicity evaluation of ADMVA, a multigenic, modified vaccinia Ankara-HIV-1 B'/C candidate vaccine.

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    Sandhya Vasan

    Full Text Available BACKGROUND: We conducted a Phase I dose-escalation trial of ADMVA, a Clade-B'/C-based HIV-1 candidate vaccine expressing env, gag, pol, nef, and tat in a modified vaccinia Ankara viral vector. Sequences were derived from a prevalent circulating HIV-1 recombinant form in Yunnan, China, an area of high HIV incidence. The objective was to evaluate the safety and immunogenicity of ADMVA in human volunteers. METHODOLOGY/PRINCIPAL FINDINGS: ADMVA or placebo was administered intramuscularly at months 0, 1 and 6 to 50 healthy adult volunteers not at high risk for HIV-1. In each dosage group [1x10(7 (low, 5x10(7 (mid, or 2.5x10(8 pfu (high] volunteers were randomized in a 3:1 ratio to receive ADMVA or placebo in a double-blinded design. Subjects were followed for local and systemic reactogenicity, adverse events including cardiac adverse events, and clinical laboratory parameters. Study follow up was 18 months. Humoral immunogenicity was evaluated by anti-gp120 binding ELISA, immunoflourescent staining, and HIV-1 neutralization. Cellular immunogenicity was assessed by a validated IFNgamma ELISpot assay and intracellular cytokine staining. Anti-vaccinia binding titers were measured by ELISA. ADMVA was generally well-tolerated, with no vaccine-related serious adverse events or cardiac adverse events. Local or systemic reactogenicity events were reported by 77% and 78% of volunteers, respectively. The majority of events were of mild intensity. The IFNgamma ELISpot response rate to any HIV antigen was 0/12 (0% in the placebo group, 3/12 (25% in the low dosage group, 6/12 (50% in the mid dosage group, and 8/13 (62% in the high dosage group. Responses were often multigenic and occasionally persisted up to one year post vaccination. Antibodies to gp120 were detected in 0/12 (0%, 8/13 (62%, 6/12 (50% and 10/13 (77% in the placebo, low, mid, and high dosage groups, respectively. Antibodies persisted up to 12 months after vaccination, with a trend toward agreement

  3. New crown motif of an HIV-1 V3 loop sequence from a Ugandan AIDS patient.

    Science.gov (United States)

    von Brunn, A; von Brunn, B; Eberle, J; Biryahwaho, B; Downing, R G; Gürtler, L

    1995-01-01

    HIV-1 V3 loop sequences from Ugandan patients include motifs from subtypes A, B, and D. To characterize further HIV isolates, V3 loop sequences were amplified from HIV-1 isolated in 1987 from peripheral blood mononuclear cells (PBL) of three patients with full-blown AIDS from Kampala, Uganda. The PBL were separated by Ficoll Paque gradients and cocultivated with noninfected donor lymphocytes for two weeks. The HIV was then transferred to HUT-78 cells. From extracted DNA of the permanently-infected HUT-78 cells, nested polymerase chain reaction (PCR) was conducted, with V3 loop sequencing performed directly upon PCR fragments derived from two independent DNA preparations and on cloned fragments. Isolates MVP-9801, -9802, and -9803 show 35.6%, 32.4%, and 29.7% nucleotide sequence divergence from the ELI subtype D sequence; 31.5%, 25.7%, and 18.9% divergence from the Z2Z6 subtype D sequence; and 21.9%, 12.2%, and 12.2% divergence from the subtype D consensus sequence. All three deduced amino acid sequences fit into the subtype D consensus sequence rather than into other V3 loop sequences described for Ugandan subtype A isolates. MVP-9802 and MVP-9803 contain the GSGQA pentapeptide motif at the tip of the V3 loop, while MVP-9801 contains GGRA. This may be explained by a deletion of proline codon between the codons for the two glycine residues. The authors believe that this deletion has not been previously reported. They also note that the deletion does not appear to be associated with a growth difference in vitro or with a difference in pathogenicity in vivo. The immunogenic implications of this altered V3 loop crest remain unclear. The Western blot profiles for the gp160, gp120, and gp41 proteins of the three Ugandan isolates manifest normal molecular weights.

  4. Curcumin derivatives as HIV-1 protease inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Sui, Z.; Li, J.; Craik, C.S.; Ortiz de Montellano, P.R. [Univ. of California, San Francisco, CA (United States)

    1993-12-31

    Curcumin, a non-toxic natural compound from Curcuma longa, has been found to be an HIV-1 protease inhibitor. Some of its derivatives were synthesized and their inhibitory activity against the HIV-1 protease was tested. Curcumin analogues containing boron enhanced the inhibitory activity. At least of the the synthesized compounds irreversibly inhibits the HIV-1 protease.

  5. A macaque model of HIV-1 infection

    OpenAIRE

    Hatziioannou, Theodora; Ambrose, Zandrea; Chung, Nancy P. Y.; Piatak, Michael; Yuan, Fang; Trubey, Charles M.; Coalter, Vicky; Kiser, Rebecca; Schneider, Doug; Smedley, Jeremy; Pung, Rhonda; Gathuka, Mercy; Estes, Jacob D.; Veazey, Ronald S.; KewalRamani, Vineet N.

    2009-01-01

    The lack of a primate model that utilizes HIV-1 as the challenge virus is an impediment to AIDS research; existing models generally employ simian viruses that are divergent from HIV-1, reducing their usefulness in preclinical investigations. Based on an understanding of species-specific variation in primate TRIM5 and APOBEC3 antiretroviral genes, we constructed simian-tropic (st)HIV-1 strains that differ from HIV-1 only in the vif gene. We demonstrate that such minimally modified stHIV-1 stra...

  6. Plasticity and Epitope Exposure of the HIV-1 Envelope Trimer.

    Science.gov (United States)

    Powell, Rebecca L R; Totrov, Maxim; Itri, Vincenza; Liu, Xiaomei; Fox, Alisa; Zolla-Pazner, Susan

    2017-09-01

    We recently showed that mutations in the HIV-1 envelope (Env) destabilize the V3 loop, rendering neutralization-resistant viruses sensitive to V3-directed monoclonal antibodies (MAbs). Here, we investigated the propagation of this effect on other Env epitopes, with special emphasis on V2 loop exposure. Wild-type JR-FL and 19 mutant JR-FL pseudoviruses were tested for neutralization sensitivity to 21 MAbs specific for epitopes in V2, the CD4 binding site (CD4bs), and the CD4-induced (CD4i) region. Certain glycan mutants, mutations in the gp120 hydrophobic core, and mutations in residues involved in intraprotomer interactions exposed epitopes in the V2i region (which overlies the α4β7 integrin binding site) and the V3 crown, suggesting general destabilization of the distal region of the trimer apex. In contrast, other glycan mutants, mutations affecting interprotomer interactions, and mutations affecting the CD4bs exposed V3 but not V2i epitopes. These data indicate for the first time that V3 can move independently of V2, with V3 pivoting out from its "tucked" position in the trimer while apparently leaving the V2 apex intact. Notably, none of the mutations exposed V2 epitopes without also exposing V3, suggesting that movement of V2 releases V3. Most mutations increased sensitivity to CD4bs-directed MAbs without exposure of the CD4i epitope, implying these mutations facilitate the trimers' maintenance of an intermediate energy state between open and closed conformations. Taken together, these data indicate that several transient Env epitopes can be rendered more accessible to antibodies (Abs) via specific mutations, and this may facilitate the design of V1V2-targeting immunogens.IMPORTANCE Many epitopes of the HIV envelope (Env) spike are relatively inaccessible to antibodies (Abs) compared to their exposure in the open Env conformation induced by receptor binding. However, the reduced infection rate that resulted from the vaccine used in the RV144 HIV-1 vaccine

  7. A European multicientre study on the comparison of HIV-1 viral loads between VERIS HIV-1 Assay and Roche COBAS® TAQMAN® HIV-1 test, Abbott RealTime HIV-1 Assay, and Siemens VERSANT HIV-1 Assay.

    Science.gov (United States)

    Braun, Patrick; Delgado, Rafael; Drago, Monica; Fanti, Diana; Fleury, Hervé; Hofmann, Jörg; Izopet, Jacques; Kühn, Sebastian; Lombardi, Alessandra; Mancon, Alessandro; Marcos, Mª Angeles; Mileto, Davide; Sauné, Karine; O'Shea, Siobhan; Pérez-Rivilla, Alfredo; Ramble, John; Trimoulet, Pascale; Vila, Jordi; Whittaker, Duncan; Artus, Alain; Rhodes, Daniel

    2017-07-01

    Viral load monitoring is essential for patients under treatment for HIV. Beckman Coulter has developed the VERIS HIV-1 Assay for use on the novel, automated DxN VERIS Molecular Diagnostics System. ¥ OBJECTIVES: Evaluation of the clinical performance of the new quantitative VERIS HIV-1 Assay at multiple EU laboratories. Method comparison with the VERIS HIV-1 Assay was performed with 415 specimens at 5 sites tested with COBAS ® AmpliPrep/COBAS ® TaqMan ® HIV-1 Test, v2.0, 169 specimens at 3 sites tested with RealTime HIV-1 Assay, and 202 specimens from 2 sites tested with VERSANT HIV-1 Assay. Patient monitoring sample results from 4 sites were also compared. Bland-Altman analysis showed the average bias between VERIS HIV-1 Assay and COBAS HIV-1 Test, RealTime HIV-1 Assay, and VERSANT HIV-1 Assay to be 0.28, 0.39, and 0.61 log 10 cp/mL, respectively. Bias at low end levels below 1000cp/mL showed predicted bias to be HIV-1 Assay versus COBAS HIV-1 Test and RealTime HIV-1 Assay, and HIV-1 Assay. Analysis on 174 specimens tested with the 0.175mL volume VERIS HIV-1 Assay and COBAS HIV-1 Test showed average bias of 0.39 log 10 cp/mL. Patient monitoring results using VERIS HIV-1 Assay demonstrated similar viral load trends over time to all comparators. The VERIS HIV-1 Assay for use on the DxN VERIS System demonstrated comparable clinical performance to COBAS ® HIV-1 Test, RealTime HIV-1 Assay, and VERSANT HIV-1 Assay. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. Variations in the Biological Functions of HIV-1 Clade C Envelope in a SHIV-Infected Rhesus Macaque during Disease Progression.

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    For Yue Tso

    Full Text Available A better understanding of how the biological functions of the HIV-1 envelope (Env changes during disease progression may aid the design of an efficacious anti-HIV-1 vaccine. Although studies from patient had provided some insights on this issue, the differences in the study cohorts and methodology had make it difficult to reach a consensus of the variations in the HIV-1 Env functions during disease progression. To this end, an animal model that can be infected under controlled environment and reflect the disease course of HIV-1 infection in human will be beneficial. Such an animal model was previously demonstrated by the infection of macaque with SHIV, expressing HIV-1 clade C Env V1-V5 region. By using this model, we examined the changes in biological functions of Env in the infected animal over the entire disease course. Our data showed an increase in the neutralization resistance phenotype over time and coincided with the decrease in the net charges of the V1-V5 region. Infection of PBMC with provirus expressing various Env clones, isolated from the infected animal over time, showed a surprisingly better replicative fitness for viruses expressing the Env from early time point. Biotinylation and ELISA data also indicated a decrease of cell-surface-associated Env and virion-associated gp120 content with disease progression. This decrease did not affect the CD4-binding capability of Env, but were positively correlated with the decrease of Env fusion ability. Interestingly, some of these changes in biological functions reverted to the pre-AIDS level during advance AIDS. These data suggested a dynamic relationship between the Env V1-V5 region with the host immune pressure. The observed changes of biological functions in this setting might reflect and predict those occurring during natural disease progression in human.

  9. A New Glycan-Dependent CD4-Binding Site Neutralizing Antibody Exerts Pressure on HIV-1 In Vivo.

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    Natalia T Freund

    2015-10-01

    Full Text Available The CD4 binding site (CD4bs on the envelope glycoprotein is a major site of vulnerability that is conserved among different HIV-1 isolates. Many broadly neutralizing antibodies (bNAbs to the CD4bs belong to the VRC01 class, sharing highly restricted origins, recognition mechanisms and viral escape pathways. We sought to isolate new anti-CD4bs bNAbs with different origins and mechanisms of action. Using a gp120 2CC core as bait, we isolated antibodies encoded by IGVH3-21 and IGVL3-1 genes with long CDRH3s that depend on the presence of the N-linked glycan at position-276 for activity. This binding mode is similar to the previously identified antibody HJ16, however the new antibodies identified herein are more potent and broad. The most potent variant, 179NC75, had a geometric mean IC80 value of 0.42 μg/ml against 120 Tier-2 HIV-1 pseudoviruses in the TZM.bl assay. Although this group of CD4bs glycan-dependent antibodies can be broadly and potently neutralizing in vitro, their in vivo activity has not been tested to date. Here, we report that 179NC75 is highly active when administered to HIV-1-infected humanized mice, where it selects for escape variants that lack a glycan site at position-276. The same glycan was absent from the virus isolated from the 179NC75 donor, implying that the antibody also exerts selection pressure in humans.

  10. Neutralizing antibody escape during HIV-1 mother-to-child transmission involves conformational masking of distal epitopes in envelope.

    Science.gov (United States)

    Goo, Leslie; Milligan, Caitlin; Simonich, Cassandra A; Nduati, Ruth; Overbaugh, Julie

    2012-09-01

    HIV-1 variants transmitted to infants are often resistant to maternal neutralizing antibodies (NAbs), suggesting that they have escaped maternal NAb pressure. To define the molecular basis of NAb escape that contributes to selection of transmitted variants, we analyzed 5 viruses from 2 mother-to-child transmission pairs, in which the infant virus, but not the maternal virus, was resistant to neutralization by maternal plasma near transmission. We generated chimeric viruses between maternal and infant envelope clones obtained near transmission and examined neutralization by maternal plasma. The molecular determinants of NAb escape were distinct, even when comparing two maternal variants to the transmitted infant virus within one pair, in which insertions in V4 of gp120 and substitutions in HR2 of gp41 conferred neutralization resistance. In another pair, deletions and substitutions in V1 to V3 conferred resistance, but neither V1/V2 nor V3 alone was sufficient. Although the sequence determinants of escape were distinct, all of them involved modifications of potential N-linked glycosylation sites. None of the regions that mediated escape were major linear targets of maternal NAbs because corresponding peptides failed to compete for neutralization. Instead, these regions disrupted multiple distal epitopes targeted by HIV-1-specific monoclonal antibodies, suggesting that escape from maternal NAbs occurred through conformational masking of distal epitopes. This strategy likely allows HIV-1 to utilize relatively limited changes in the envelope to preserve the ability to infect a new host while simultaneously evading multiple NAb specificities present in maternal plasma.

  11. A New Glycan-Dependent CD4-Binding Site Neutralizing Antibody Exerts Pressure on HIV-1 In Vivo.

    Science.gov (United States)

    Freund, Natalia T; Horwitz, Joshua A; Nogueira, Lilian; Sievers, Stuart A; Scharf, Louise; Scheid, Johannes F; Gazumyan, Anna; Liu, Cassie; Velinzon, Klara; Goldenthal, Ariel; Sanders, Rogier W; Moore, John P; Bjorkman, Pamela J; Seaman, Michael S; Walker, Bruce D; Klein, Florian; Nussenzweig, Michel C

    2015-10-01

    The CD4 binding site (CD4bs) on the envelope glycoprotein is a major site of vulnerability that is conserved among different HIV-1 isolates. Many broadly neutralizing antibodies (bNAbs) to the CD4bs belong to the VRC01 class, sharing highly restricted origins, recognition mechanisms and viral escape pathways. We sought to isolate new anti-CD4bs bNAbs with different origins and mechanisms of action. Using a gp120 2CC core as bait, we isolated antibodies encoded by IGVH3-21 and IGVL3-1 genes with long CDRH3s that depend on the presence of the N-linked glycan at position-276 for activity. This binding mode is similar to the previously identified antibody HJ16, however the new antibodies identified herein are more potent and broad. The most potent variant, 179NC75, had a geometric mean IC80 value of 0.42 μg/ml against 120 Tier-2 HIV-1 pseudoviruses in the TZM.bl assay. Although this group of CD4bs glycan-dependent antibodies can be broadly and potently neutralizing in vitro, their in vivo activity has not been tested to date. Here, we report that 179NC75 is highly active when administered to HIV-1-infected humanized mice, where it selects for escape variants that lack a glycan site at position-276. The same glycan was absent from the virus isolated from the 179NC75 donor, implying that the antibody also exerts selection pressure in humans.

  12. Viral escape from HIV-1 neutralizing antibodies drives increased plasma neutralization breadth through sequential recognition of multiple epitopes and immunotypes.

    Science.gov (United States)

    Wibmer, Constantinos Kurt; Bhiman, Jinal N; Gray, Elin S; Tumba, Nancy; Abdool Karim, Salim S; Williamson, Carolyn; Morris, Lynn; Moore, Penny L

    2013-10-01

    Identifying the targets of broadly neutralizing antibodies to HIV-1 and understanding how these antibodies develop remain important goals in the quest to rationally develop an HIV-1 vaccine. We previously identified a participant in the CAPRISA Acute Infection Cohort (CAP257) whose plasma neutralized 84% of heterologous viruses. In this study we showed that breadth in CAP257 was largely due to the sequential, transient appearance of three distinct broadly neutralizing antibody specificities spanning the first 4.5 years of infection. The first specificity targeted an epitope in the V2 region of gp120 that was also recognized by strain-specific antibodies 7 weeks earlier. Specificity for the autologous virus was determined largely by a rare N167 antigenic variant of V2, with viral escape to the more common D167 immunotype coinciding with the development of the first wave of broadly neutralizing antibodies. Escape from these broadly neutralizing V2 antibodies through deletion of the glycan at N160 was associated with exposure of an epitope in the CD4 binding site that became the target for a second wave of broadly neutralizing antibodies. Neutralization by these CD4 binding site antibodies was almost entirely dependent on the glycan at position N276. Early viral escape mutations in the CD4 binding site drove an increase in wave two neutralization breadth, as this second wave of heterologous neutralization matured to recognize multiple immunotypes within this site. The third wave targeted a quaternary epitope that did not overlap any of the four known sites of vulnerability on the HIV-1 envelope and remains undefined. Altogether this study showed that the human immune system is capable of generating multiple broadly neutralizing antibodies in response to a constantly evolving viral population that exposes new targets as a consequence of escape from earlier neutralizing antibodies.

  13. Viral escape from HIV-1 neutralizing antibodies drives increased plasma neutralization breadth through sequential recognition of multiple epitopes and immunotypes.

    Directory of Open Access Journals (Sweden)

    Constantinos Kurt Wibmer

    2013-10-01

    Full Text Available Identifying the targets of broadly neutralizing antibodies to HIV-1 and understanding how these antibodies develop remain important goals in the quest to rationally develop an HIV-1 vaccine. We previously identified a participant in the CAPRISA Acute Infection Cohort (CAP257 whose plasma neutralized 84% of heterologous viruses. In this study we showed that breadth in CAP257 was largely due to the sequential, transient appearance of three distinct broadly neutralizing antibody specificities spanning the first 4.5 years of infection. The first specificity targeted an epitope in the V2 region of gp120 that was also recognized by strain-specific antibodies 7 weeks earlier. Specificity for the autologous virus was determined largely by a rare N167 antigenic variant of V2, with viral escape to the more common D167 immunotype coinciding with the development of the first wave of broadly neutralizing antibodies. Escape from these broadly neutralizing V2 antibodies through deletion of the glycan at N160 was associated with exposure of an epitope in the CD4 binding site that became the target for a second wave of broadly neutralizing antibodies. Neutralization by these CD4 binding site antibodies was almost entirely dependent on the glycan at position N276. Early viral escape mutations in the CD4 binding site drove an increase in wave two neutralization breadth, as this second wave of heterologous neutralization matured to recognize multiple immunotypes within this site. The third wave targeted a quaternary epitope that did not overlap any of the four known sites of vulnerability on the HIV-1 envelope and remains undefined. Altogether this study showed that the human immune system is capable of generating multiple broadly neutralizing antibodies in response to a constantly evolving viral population that exposes new targets as a consequence of escape from earlier neutralizing antibodies.

  14. Lack of ADCC Breadth of Human Nonneutralizing Anti-HIV-1 Antibodies.

    Science.gov (United States)

    Bruel, Timothée; Guivel-Benhassine, Florence; Lorin, Valérie; Lortat-Jacob, Hugues; Baleux, Françoise; Bourdic, Katia; Noël, Nicolas; Lambotte, Olivier; Mouquet, Hugo; Schwartz, Olivier

    2017-04-15

    Anti-human immunodeficiency virus type 1 (HIV-1) nonneutralizing antibodies (nnAbs) capable of antibody-dependent cellular cytotoxicity (ADCC) have been identified as a protective immune correlate in the RV144 vaccine efficacy trial. Broadly neutralizing antibodies (bNAbs) also mediate ADCC in cell culture and rely on their Fc region for optimal efficacy in animal models. Here, we selected 9 monoclonal nnAbs and 5 potent bNAbs targeting various epitopes and conformations of the gp120/41 complex and analyzed the potency of the two types of antibodies to bind and eliminate HIV-1-infected cells in culture. Regardless of their neutralizing activity, most of the selected antibodies recognized and killed cells infected with two laboratory-adapted HIV-1 strains. Some nnAbs also bound bystander cells that may have captured viral proteins. However, in contrast to the bNAbs, the nnAbs bound poorly to reactivated infected cells from 8 HIV-positive individuals and did not mediate effective ADCC against these cells. The nnAbs also inefficiently recognize cells infected with 8 different transmitted-founder (T/F) isolates. The addition of a synthetic CD4 mimetic enhanced the binding and killing efficacy of some of the nnAbs in an epitope-dependent manner without reaching the levels achieved by the most potent bNAbs. Overall, our data reveal important qualitative and quantitative differences between nnAbs and bNAbs in their ADCC capacity and strongly suggest that the breadth of recognition of HIV-1 by nnAbs is narrow.IMPORTANCE Most of the anti-HIV antibodies generated by infected individuals do not display potent neutralizing activities. These nonneutralizing antibodies (nnAbs) with antibody-dependent cellular cytotoxicity (ADCC) have been identified as a protective immune correlate in the RV144 vaccine efficacy trial. However, in primate models, the nnAbs do not protect against simian-human immunodeficiency virus (SHIV) acquisition. Thus, the role of nnAbs with ADCC activity in

  15. Diagnostik af HIV-1 infektionen

    DEFF Research Database (Denmark)

    Christiansen, C B; Dickmeiss, E; Bygbjerg, Ib Christian

    1991-01-01

    Different methods have been developed for the diagnosis of HIV infection, i.e. detection of antibodies, antigen and proviral DNA. ELISA methods for detecting HIV-1 antibodies are widely used as screening assays. A sample which is repeatedly positive with ELISA is re-tested with a confirmatory test...... in a proportion of patients. Detection and quantitation of HIV antigen are used as indicators of disease progression and for monitoring the antiviral efficacy of therapeutic interventions. When no antibodies or antigens can be detected in persons suspected of having HIV infection, culture of HIV can be performed....... For research purposes, detection of small amounts of proviral DNA can be made with polymerase chain reaction (PCR). The method is not yet applicable in routine diagnosis of HIV infection....

  16. Features of Recently Transmitted HIV-1 Clade C Viruses that Impact Antibody Recognition: Implications for Active and Passive Immunization.

    Science.gov (United States)

    Rademeyer, Cecilia; Korber, Bette; Seaman, Michael S; Giorgi, Elena E; Thebus, Ruwayhida; Robles, Alexander; Sheward, Daniel J; Wagh, Kshitij; Garrity, Jetta; Carey, Brittany R; Gao, Hongmei; Greene, Kelli M; Tang, Haili; Bandawe, Gama P; Marais, Jinny C; Diphoko, Thabo E; Hraber, Peter; Tumba, Nancy; Moore, Penny L; Gray, Glenda E; Kublin, James; McElrath, M Juliana; Vermeulen, Marion; Middelkoop, Keren; Bekker, Linda-Gail; Hoelscher, Michael; Maboko, Leonard; Makhema, Joseph; Robb, Merlin L; Abdool Karim, Salim; Abdool Karim, Quarraisha; Kim, Jerome H; Hahn, Beatrice H; Gao, Feng; Swanstrom, Ronald; Morris, Lynn; Montefiori, David C; Williamson, Carolyn

    2016-07-01

    The development of biomedical interventions to reduce acquisition of HIV-1 infection remains a global priority, however their potential effectiveness is challenged by very high HIV-1 envelope diversity. Two large prophylactic trials in high incidence, clade C epidemic regions in southern Africa are imminent; passive administration of the monoclonal antibody VRC01, and active immunization with a clade C modified RV144-like vaccines. We have created a large representative panel of C clade viruses to enable assessment of antibody responses to vaccines and natural infection in Southern Africa, and we investigated the genotypic and neutralization properties of recently transmitted clade C viruses to determine how viral diversity impacted antibody recognition. We further explore the implications of these findings for the potential effectiveness of these trials. A panel of 200 HIV-1 Envelope pseudoviruses was constructed from clade C viruses collected within the first 100 days following infection. Viruses collected pre-seroconversion were significantly more resistant to serum neutralization compared to post-seroconversion viruses (p = 0.001). Over 13 years of the study as the epidemic matured, HIV-1 diversified (p = 0.0009) and became more neutralization resistant to monoclonal antibodies VRC01, PG9 and 4E10. When tested at therapeutic levels (10ug/ml), VRC01 only neutralized 80% of viruses in the panel, although it did exhibit potent neutralization activity against sensitive viruses (IC50 titres of 0.42 μg/ml). The Gp120 amino acid similarity between the clade C panel and candidate C-clade vaccine protein boosts (Ce1086 and TV1) was 77%, which is 8% more distant than between CRF01_AE viruses and the RV144 CRF01_AE immunogen. Furthermore, two vaccine signature sites, K169 in V2 and I307 in V3, associated with reduced infection risk in RV144, occurred less frequently in clade C panel viruses than in CRF01_AE viruses from Thailand. Increased resistance of pre

  17. Cross-border sexual transmission of the newly emerging HIV-1 clade CRF51_01B.

    Directory of Open Access Journals (Sweden)

    Hui Ting Cheong

    Full Text Available A novel HIV-1 recombinant clade (CRF51_01B was recently identified among men who have sex with men (MSM in Singapore. As cases of sexually transmitted HIV-1 infection increase concurrently in two socioeconomically intimate countries such as Malaysia and Singapore, cross transmission of HIV-1 between said countries is highly probable. In order to investigate the timeline for the emergence of HIV-1 CRF51_01B in Singapore and its possible introduction into Malaysia, 595 HIV-positive subjects recruited in Kuala Lumpur from 2008 to 2012 were screened. Phylogenetic relationship of 485 amplified polymerase gene sequences was determined through neighbour-joining method. Next, near-full length sequences were amplified for genomic sequences inferred to be CRF51_01B and subjected to further analysis implemented through Bayesian Markov chain Monte Carlo (MCMC sampling and maximum likelihood methods. Based on the near full length genomes, two isolates formed a phylogenetic cluster with CRF51_01B sequences of Singapore origin, sharing identical recombination structure. Spatial and temporal information from Bayesian MCMC coalescent and maximum likelihood analysis of the protease, gp120 and gp41 genes suggest that Singapore is probably the country of origin of CRF51_01B (as early as in the mid-1990s and featured a Malaysian who acquired the infection through heterosexual contact as host for its ancestral lineages. CRF51_01B then spread rapidly among the MSM in Singapore and Malaysia. Although the importation of CRF51_01B from Singapore to Malaysia is supported by coalescence analysis, the narrow timeframe of the transmission event indicates a closely linked epidemic. Discrepancies in the estimated divergence times suggest that CRF51_01B may have arisen through multiple recombination events from more than one parental lineage. We report the cross transmission of a novel CRF51_01B lineage between countries that involved different sexual risk groups. Understanding

  18. Features of Recently Transmitted HIV-1 Clade C Viruses that Impact Antibody Recognition: Implications for Active and Passive Immunization.

    Directory of Open Access Journals (Sweden)

    Cecilia Rademeyer

    2016-07-01

    Full Text Available The development of biomedical interventions to reduce acquisition of HIV-1 infection remains a global priority, however their potential effectiveness is challenged by very high HIV-1 envelope diversity. Two large prophylactic trials in high incidence, clade C epidemic regions in southern Africa are imminent; passive administration of the monoclonal antibody VRC01, and active immunization with a clade C modified RV144-like vaccines. We have created a large representative panel of C clade viruses to enable assessment of antibody responses to vaccines and natural infection in Southern Africa, and we investigated the genotypic and neutralization properties of recently transmitted clade C viruses to determine how viral diversity impacted antibody recognition. We further explore the implications of these findings for the potential effectiveness of these trials. A panel of 200 HIV-1 Envelope pseudoviruses was constructed from clade C viruses collected within the first 100 days following infection. Viruses collected pre-seroconversion were significantly more resistant to serum neutralization compared to post-seroconversion viruses (p = 0.001. Over 13 years of the study as the epidemic matured, HIV-1 diversified (p = 0.0009 and became more neutralization resistant to monoclonal antibodies VRC01, PG9 and 4E10. When tested at therapeutic levels (10ug/ml, VRC01 only neutralized 80% of viruses in the panel, although it did exhibit potent neutralization activity against sensitive viruses (IC50 titres of 0.42 μg/ml. The Gp120 amino acid similarity between the clade C panel and candidate C-clade vaccine protein boosts (Ce1086 and TV1 was 77%, which is 8% more distant than between CRF01_AE viruses and the RV144 CRF01_AE immunogen. Furthermore, two vaccine signature sites, K169 in V2 and I307 in V3, associated with reduced infection risk in RV144, occurred less frequently in clade C panel viruses than in CRF01_AE viruses from Thailand. Increased resistance of

  19. Nef decreases HIV-1 sensitivity to neutralizing antibodies that target the membrane-proximal external region of TMgp41.

    Directory of Open Access Journals (Sweden)

    Rachel P J Lai

    2011-12-01

    Full Text Available Primate lentivirus nef is required for sustained virus replication in vivo and accelerated progression to AIDS. While exploring the mechanism by which Nef increases the infectivity of cell-free virions, we investigated a functional link between Nef and Env. Since we failed to detect an effect of Nef on the quantity of virion-associated Env, we searched for qualitative changes by examining whether Nef alters HIV-1 sensitivity to agents that target distinct features of Env. Nef conferred as much as 50-fold resistance to 2F5 and 4E10, two potent neutralizing monoclonal antibodies (nAbs that target the membrane proximal external region (MPER of TMgp41. In contrast, Nef had no effect on HIV-1 neutralization by MPER-specific nAb Z13e1, by the peptide inhibitor T20, nor by a panel of nAbs and other reagents targeting gp120. Resistance to neutralization by 2F5 and 4E10 was observed with Nef from a diverse range of HIV-1 and SIV isolates, as well as with HIV-1 virions bearing Env from CCR5- and CXCR4-tropic viruses, clade B and C viruses, or primary isolates. Functional analysis of a panel of Nef mutants revealed that this activity requires Nef myristoylation but that it is genetically separable from other Nef functions such as the ability to enhance virus infectivity and to downregulate CD4. Glycosylated-Gag from MoMLV substituted for Nef in conferring resistance to 2F5 and 4E10, indicating that this activity is conserved in a retrovirus that does not encode Nef. Given the reported membrane-dependence of MPER-recognition by 2F5 and 4E10, in contrast to the membrane-independence of Z13e1, the data here is consistent with a model in which Nef alters MPER recognition in the context of the virion membrane. Indeed, Nef and Glycosylated-Gag decreased the efficiency of virion capture by 2F5 and 4E10, but not by other nAbs. These studies demonstrate that Nef protects lentiviruses from one of the most broadly-acting classes of neutralizing antibodies. This newly

  20. Reducing V3 Antigenicity and Immunogenicity on Soluble, Native-Like HIV-1 Env SOSIP Trimers.

    Science.gov (United States)

    Ringe, Rajesh P; Ozorowski, Gabriel; Rantalainen, Kimmo; Struwe, Weston B; Matthews, Katie; Torres, Jonathan L; Yasmeen, Anila; Cottrell, Christopher A; Ketas, Thomas J; LaBranche, Celia C; Montefiori, David C; Cupo, Albert; Crispin, Max; Wilson, Ian A; Ward, Andrew B; Sanders, Rogier W; Klasse, P J; Moore, John P

    2017-08-01

    Native-like trimers of the SOSIP design are being developed as immunogens in human immunodeficiency virus type 1 (HIV-1) vaccine development programs. These trimers display the epitopes for multiple broadly neutralizing antibodies (bNAbs) but can also expose binding sites for some types of nonneutralizing antibodies (non-NAbs). Among the latter are epitopes in the gp120 V3 region that are highly immunogenic when SOSIP trimers are evaluated in animal models. It is presently uncertain whether antibodies against V3 can interfere with the induction of NAbs, but there are good arguments in favor of suppressing such "off-target" immune responses. Accordingly, we have assessed how to minimize the exposure of V3 non-NAb epitopes and thereby reduce their immunogenicity by introducing N-glycans within the V3 region of BG505 SOSIP trimers. We found that inserting glycans at positions 306 and 314 (termed M1 and M7) markedly reduced V3 antigenicity while improving the presentation of trimer apex bNAb epitopes. Both added glycans were shown to be predominantly of the Man6GlcNAc2 form. The additional introduction of the E64K ground-state stabilizing substitution markedly reduced or ablated soluble CD4 (sCD4) induction of non-NAb epitopes in V3 and/or associated with the coreceptor binding site. When a V3 glycan- and E64K-modified trimer variant, BG505 SOSIP.664-E64K.M1M7, was tested in rabbits, V3 immunogenicity was eliminated while the autologous NAb response was unchanged.IMPORTANCE Trimeric proteins are being developed for future HIV-1 vaccine trials in humans, with the goal of eliciting broadly active neutralizing antibodies (NAbs) that are active against a wide variety of circulating strains. In animal models, the present generation of native-like trimer immunogens, exemplified by the BG505 SOSIP.664 construct, induces narrow-specificity antibodies against the neutralization-resistant (tier-2), sequence-matched virus and more broadly active antibodies against sequence

  1. Efaverinz and nano-gold-loaded mannosylated niosomes: a host cell-targeted topical HIV-1 prophylaxis via thermogel system.

    Science.gov (United States)

    Malik, Tanushree; Chauhan, Gaurav; Rath, Goutam; Kesarkar, Rohan N; Chowdhary, Abhay S; Goyal, Amit K

    2017-12-12

    Sexual dissemination of Human Immunodeficiency Virus-1 (HIV-1) is the prime mode of its spread. Topical microbicidal approach has gained much attention, but no real success is observed till date, either due to toxicity or resistance of active moieties and the lack of efficient drug delivery approaches. In this research protocol, a unique combination approach of a standard drug moiety, that is, Efaverinz (EFV) and a nanometal, that is, gold nanoparticles (GNPs) was tried. Both these candidates were delivered through a mannosylated niosomal system, to exploit protein (lectins present on HIV host cells) - carbohydrate (oligosaccharides such as mannan present on HIV gp-120 receptor) interaction. GNPs (10.4 nm average size) were entrapped inside the aqueous core, whereas lipophilic EFV was loaded in the bilayer membrane. Results demonstrated a significant increase in antiviral activity when EFV was fired with GNPs. Delivery of this combination via mannosylated niosomes proved to be a perfect approach with exceedingly well potential compared to non liganded niosomal system. A thermosensitive gel vehicle was prepared and the loaded niosomes were dispersed in it to have a nanogel system. The optimized formulation was evaluated for its prophylactic activity and the results showed completely inhibited viral dissemination at folds dilution levels.

  2. Use of T7 gene 6 exonuclease and phosphorothioated primers for the manipulation of HIV-1 infectious clones.

    Science.gov (United States)

    da Costa, L J; Tanuri, A

    1998-05-01

    A method is described for the efficient substitution, deletion or insertion of any desired DNA sequence into any viral infectious clones without the limitation of naturally occurring restriction sites. The technique employs the polymerase chain reaction combined with the resistance of 2'-deoxynucleotides 5'-O-(1-thiotriphosphate) dNTPs [S] bonds (phosphorothiate bonds) to the 5'-3' double strand specific T7 gene 6 exonuclease (T7 Exo) digestion. Primers used to amplify the DNA target regions being manipulated present three phosphorothioate bonds from the fifteenth base at the 5' end. The enzyme activity was shown to be completely inhibited by the presence of more than one phosphorothioate residue at the 5' end of the DNA molecules. When the amplification products are submitted to the exonuclease digestion the hydrolytic T7 Exo activity generates a short single strand DNA tail which contains the nucleotide integrity of the 3' strand. Since the ends of two independently amplified products overlap they can regenerate a stable recombinant structure when further combined in the same reaction tube in the presence of T4 DNA ligase. This new method can be used for manipulating an HIV-1 full-length clone belonging to subtype D replacing the env (gp120) gene for an F subtype sequence.

  3. Complete dissociation of the HIV-1 gp41 ectodomain and membrane proximal regions upon phospholipid binding

    Energy Technology Data Exchange (ETDEWEB)

    Roche, Julien; Louis, John M.; Aniana, Annie [National Institute of Diabetes and Digestive and Kidney Diseases, Laboratory of Chemical Physics (United States); Ghirlando, Rodolfo [National Institutes of Health, Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases (United States); Bax, Ad, E-mail: bax@nih.gov [National Institute of Diabetes and Digestive and Kidney Diseases, Laboratory of Chemical Physics (United States)

    2015-04-15

    The envelope glycoprotein gp41 mediates the process of membrane fusion that enables entry of the HIV-1 virus into the host cell. Strong lipid affinity of the ectodomain suggests that its heptad repeat regions play an active role in destabilizing membranes by directly binding to the lipid bilayers and thereby lowering the free-energy barrier for membrane fusion. In such a model, immediately following the shedding of gp120, the N-heptad and C-heptad helices dissociate and melt into the host cell and viral membranes, respectively, pulling the destabilized membranes into juxtaposition, ready for fusion. Post-fusion, reaching the final 6-helix bundle (6HB) conformation then involves competition between intermolecular interactions needed for formation of the symmetric 6HB trimer and the membrane affinity of gp41’s ectodomain, including its membrane-proximal regions. Our solution NMR study of the structural and dynamic properties of three constructs containing the ectodomain of gp41 with and without its membrane-proximal regions suggests that these segments do not form inter-helical interactions until the very late steps of the fusion process. Interactions between the polar termini of the heptad regions, which are not associating with the lipid surface, therefore may constitute the main driving force initiating formation of the final post-fusion states. The absence of significant intermolecular ectodomain interactions in the presence of dodecyl phosphocholine highlights the importance of trimerization of gp41’s transmembrane helix to prevent complete dissociation of the trimer during the course of fusion.

  4. Quantifying Ongoing HIV-1 Exposure in HIV-1–Serodiscordant Couples to Identify Individuals With Potential Host Resistance to HIV-1

    Science.gov (United States)

    Mackelprang, Romel D.; Baeten, Jared M.; Donnell, Deborah; Celum, Connie; Farquhar, Carey; de Bruyn, Guy; Essex, Max; McElrath, M. Juliana; Nakku-Joloba, Edith; Lingappa, Jairam R.

    2012-01-01

    Background. Immunogenetic correlates of resistance to HIV-1 in HIV-1–exposed seronegative (HESN) individuals with consistently high exposure may inform HIV-1 prevention strategies. We developed a novel approach for quantifying HIV-1 exposure to identify individuals remaining HIV-1 uninfected despite persistent high exposure. Methods. We used longitudinal predictors of HIV-1 transmission in HIV-1 serodiscordant couples to score HIV-1 exposure and define HESN clusters with persistently high, low, and decreasing risk trajectories. The model was validated in an independent cohort of serodiscordant couples. We describe a statistical tool that can be applied to other HESN cohorts to identify individuals with high exposure to HIV-1. Results. HIV-1 exposure was best quantified by frequency of unprotected sex with, plasma HIV-1 RNA levels among, and presence of genital ulcer disease among HIV-1–infected partners and by age, pregnancy status, herpes simplex virus 2 serostatus, and male circumcision status among HESN participants. Overall, 14% of HESN individuals persistently had high HIV-1 exposure and exhibited a declining incidence of HIV-1 infection over time. Conclusions. A minority of HESN individuals from HIV-1–discordant couples had persistent high HIV-1 exposure over time. Decreasing incidence of infection in this group suggests these individuals were selected for resistance to HIV-1 and may be most appropriate for identifying biological correlates of natural host resistance to HIV-1 infection. PMID:22926009

  5. Cross-reactivity of monoclonal antibodies to a chimeric V3 peptide of HIV-1 with peptide analogues studied by biosensor technology and ELISA.

    Science.gov (United States)

    Richalet-Sécordel, P M; Zeder-Lutz, G; Plaue, S; Sommermeyer-Leroux, G; Van Regenmortel, M H

    1994-12-02

    The reactivity of monoclonal antibodies (Mabs) raised against a cyclic peptide representing a chimeric V3 loop of HIV-1 gp120 with different peptide analogues was studied with a biosensor system (BIAcore) and by ELISA. In both assays, the Mabs cross-reacted extensively with the V3 regions of different HIV-1 strains and recognized the cyclic form of the peptide immunogen better than its linear form. The highest degree of cross-reactivity was observed with peptides that shared a Lys312 with the chimeric sequence. Dissociation rate constants of ten Mabs measured with the BIAcore with respect to different peptides increased with increasing numbers of substitutions in the flanking regions of the V3 tip sequence Gly Pro Gly Arg. Immobilization of the cyclic peptide on the sensor chip via a thiol group added near the end of the loop structure preserved the conformation of the peptide. In view of the good correlation between the BIAcore and ELISA results, biosensor data should be useful for selecting peptides to be used in diagnostic solid phase assays.

  6. A heterologous prime-boosting strategy with replicating Vaccinia virus vectors and plant-produced HIV-1 Gag/dgp41 virus-like particles.

    Science.gov (United States)

    Meador, Lydia R; Kessans, Sarah A; Kilbourne, Jacquelyn; Kibler, Karen V; Pantaleo, Giuseppe; Roderiguez, Mariano Esteban; Blattman, Joseph N; Jacobs, Bertram L; Mor, Tsafrir S

    2017-07-01

    Showing modest efficacy, the RV144 HIV-1 vaccine clinical trial utilized a non-replicating canarypox viral vector and a soluble gp120 protein boost. Here we built upon the RV144 strategy by developing a novel combination of a replicating, but highly-attenuated Vaccinia virus vector, NYVAC-KC, and plant-produced HIV-1 virus-like particles (VLPs). Both components contained the full-length Gag and a membrane anchored truncated gp41 presenting the membrane proximal external region with its conserved broadly neutralizing epitopes in the pre-fusion conformation. We tested different prime/boost combinations of these components in mice and showed that the group primed with NYVAC-KC and boosted with both the viral vectors and plant-produced VLPs have the most robust Gag-specific CD8 T cell responses, at 12.7% of CD8 T cells expressing IFN-γ in response to stimulation with five Gag epitopes. The same immunization group elicited the best systemic and mucosal antibody responses to Gag and dgp41 with a bias towards IgG1. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  7. HIV-1 as RNA evolution machine

    NARCIS (Netherlands)

    Berkhout, Ben

    2011-01-01

    We have over the years studied several sequence or structural elements within the HIV-1 RNA genome. Molecular mechanisms have been proposed for the role of these RNA motifs in virus replication. We have developed HIV-1 evolution as a powerful research method to study different aspects of the viral

  8. T cell dynamics in HIV-1 infection

    NARCIS (Netherlands)

    Clark, D.R.; Boer, R.J. de; Wolthers, K.C.; Miedema, F.

    1999-01-01

    One of the most prominent features of HIV-1 infection is CD4⁺ T cell depletion. This statement is widely used in papers on HIV-1 research; however, while true, it is deceptively simplistic in that it fails to describe what is actually a complex change in the representation of T cell

  9. HIV-1 Latency in Monocytes/Macrophages

    Directory of Open Access Journals (Sweden)

    Amit Kumar

    2014-04-01

    Full Text Available Human immunodeficiency virus type 1 (HIV-1 targets CD4+ T cells and cells of the monocyte/macrophage lineage. HIV pathogenesis is characterized by the depletion of T lymphocytes and by the presence of a population of cells in which latency has been established called the HIV-1 reservoir. Highly active antiretroviral therapy (HAART has significantly improved the life of HIV-1 infected patients. However, complete eradication of HIV-1 from infected individuals is not possible without targeting latent sources of infection. HIV-1 establishes latent infection in resting CD4+ T cells and findings indicate that latency can also be established in the cells of monocyte/macrophage lineage. Monocyte/macrophage lineage includes among others, monocytes, macrophages and brain resident macrophages. These cells are relatively more resistant to apoptosis induced by HIV-1, thus are important stable hideouts of the virus. Much effort has been made in the direction of eliminating HIV-1 resting CD4+ T-cell reservoirs. However, it is impossible to achieve a cure for HIV-1 without considering these neglected latent reservoirs, the cells of monocyte/macrophage lineage. In this review we will describe our current understanding of the mechanism of latency in monocyte/macrophage lineage and how such cells can be specifically eliminated from the infected host.

  10. Infected cell killing by HIV-1 protease promotes NF-kappaB dependent HIV-1 replication.

    Directory of Open Access Journals (Sweden)

    Gary D Bren

    2008-05-01

    Full Text Available Acute HIV-1 infection of CD4 T cells often results in apoptotic death of infected cells, yet it is unclear what evolutionary advantage this offers to HIV-1. Given the independent observations that acute T cell HIV-1 infection results in (1 NF-kappaB activation, (2 caspase 8 dependent apoptosis, and that (3 caspase 8 directly activates NF-kappaB, we questioned whether these three events might be interrelated. We first show that HIV-1 infected T cell apoptosis, NF-kappaB activation, and caspase 8 cleavage by HIV-1 protease are coincident. Next we show that HIV-1 protease not only cleaves procaspase 8, producing Casp8p41, but also independently stimulates NF-kappaB activity. Finally, we demonstrate that the HIV protease cleavage of caspase 8 is necessary for optimal NF-kappaB activation and that the HIV-1 protease specific cleavage fragment Casp8p41 is sufficient to stimulate HIV-1 replication through NF-kappaB dependent HIV-LTR activation both in vitro as well as in cells from HIV infected donors. Consequently, the molecular events which promote death of HIV-1 infected T cells function dually to promote HIV-1 replication, thereby favoring the propagation and survival of HIV-1.

  11. GADD45 proteins inhibit HIV-1 replication through specific suppression of HIV-1 transcription.

    Science.gov (United States)

    Liang, Zhibin; Liu, Ruikang; Zhang, Hui; Zhang, Suzhen; Hu, Xiaomei; Tan, Juan; Liang, Chen; Qiao, Wentao

    2016-06-01

    GADD45 proteins are a group of stress-induced proteins and participate in various cellular pathways including cell cycle regulation, cell survival and death, DNA repair and demethylation. It was recently shown that HIV-1 infection induces the expression of GADD45 proteins. However, the effect of GADD45 on HIV-1 replication has not been studied. Here, we report that overexpression of GADD45 proteins reduces HIV-1 production through suppressing transcription from the HIV-1 LTR promoter. This inhibitory effect is specific to HIV-1, since GADD45 proteins neither inhibit the LTR promoters from other retroviruses nor reduce the production of these viruses. Knockdown of endogenous GADD45 modestly activates HIV-1 in the J-Lat A72 latency cell line, which suggests GADD45 proteins might play a role in maintaining HIV-1 latency. Copyright © 2016 Elsevier Inc. All rights reserved.

  12. Electron tomography of the contact between T cells and SIV/HIV-1: implications for viral entry.

    Directory of Open Access Journals (Sweden)

    Rachid Sougrat

    2007-05-01

    Full Text Available The envelope glycoproteins of primate lentiviruses, including human and simian immunodeficiency viruses (HIV and SIV, are heterodimers of a transmembrane glycoprotein (usually gp41, and a surface glycoprotein (gp120, which binds CD4 on target cells to initiate viral entry. We have used electron tomography to determine the three-dimensional architectures of purified SIV virions in isolation and in contact with CD4+ target cells. The trimeric viral envelope glycoprotein surface spikes are heterogeneous in appearance and typically approximately 120 A long and approximately 120 A wide at the distal end. Docking of SIV or HIV-1 on the T cell surface occurs via a neck-shaped contact region that is approximately 400 A wide and consistently consists of a closely spaced cluster of five to seven rod-shaped features, each approximately 100 A long and approximately 100 A wide. This distinctive structure is not observed when viruses are incubated with T lymphocytes in the presence of anti-CD4 antibodies, the CCR5 antagonist TAK779, or the peptide entry inhibitor SIVmac251 C34. For virions bound to cells, few trimers were observed away from this cluster at the virion-cell interface, even in cases where virus preparations showing as many as 70 envelope glycoprotein trimers per virus particle were used. This contact zone, which we term the "entry claw", provides a spatial context to understand the molecular mechanisms of viral entry. Determination of the molecular composition and structure of the entry claw may facilitate the identification of improved drugs for the inhibition of HIV-1 entry.

  13. Cytoplasmic Dynein Promotes HIV-1 Uncoating

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    Paulina Pawlica

    2014-11-01

    Full Text Available Retroviral capsid (CA cores undergo uncoating during their retrograde transport (toward the nucleus, and/or after reaching the nuclear membrane. However, whether HIV-1 CA core uncoating is dependent upon its transport is not understood. There is some evidence that HIV-1 cores retrograde transport involves cytoplasmic dynein complexes translocating on microtubules. Here we investigate the role of dynein-dependent transport in HIV-1 uncoating. To interfere with dynein function, we depleted dynein heavy chain (DHC using RNA interference, and we over-expressed p50/dynamitin. In immunofluorescence microscopy experiments, DHC depletion caused an accumulation of CA foci in HIV-1 infected cells. Using a biochemical assay to monitor HIV-1 CA core disassembly in infected cells, we observed an increase in amounts of intact (pelletable CA cores upon DHC depletion or p50 over-expression. Results from these two complementary assays suggest that inhibiting dynein-mediated transport interferes with HIV-1 uncoating in infected cells, indicating the existence of a functional link between HIV-1 transport and uncoating.

  14. HIV/AIDS Vaccine Candidates Based on Replication-Competent Recombinant Poxvirus NYVAC-C-KC Expressing Trimeric gp140 and Gag-Derived Virus-Like Particles or Lacking the Viral Molecule B19 That Inhibits Type I Interferon Activate Relevant HIV-1-Specific B and T Cell Immune Functions in Nonhuman Primates

    Science.gov (United States)

    García-Arriaza, Juan; Perdiguero, Beatriz; Heeney, Jonathan L.; Seaman, Michael S.; Montefiori, David C.; Yates, Nicole L.; Tomaras, Georgia D.; Ferrari, Guido; Foulds, Kathryn E.; Roederer, Mario; Self, Steven G.; Borate, Bhavesh; Gottardo, Raphael; Phogat, Sanjay; Tartaglia, Jim; Barnett, Susan W.; Burke, Brian; Cristillo, Anthony D.; Weiss, Deborah E.; Lee, Carter; Kibler, Karen V.; Jacobs, Bertram L.; Wagner, Ralf; Ding, Song; Pantaleo, Giuseppe

    2017-01-01

    ABSTRACT The nonreplicating attenuated poxvirus vector NYVAC expressing clade C(CN54) HIV-1 Env(gp120) and Gag-Pol-Nef antigens (NYVAC-C) showed limited immunogenicity in phase I clinical trials. To enhance the capacity of the NYVAC vector to trigger broad humoral responses and a more balanced activation of CD4+ and CD8+ T cells, here we compared the HIV-1-specific immunogenicity elicited in nonhuman primates immunized with two replicating NYVAC vectors that have been modified by the insertion of the K1L and C7L vaccinia virus host range genes and express the clade C(ZM96) trimeric HIV-1 gp140 protein or a Gag(ZM96)-Pol-Nef(CN54) polyprotein as Gag-derived virus-like particles (termed NYVAC-C-KC). Additionally, one NYVAC-C-KC vector was generated by deleting the viral gene B19R, an inhibitor of the type I interferon response (NYVAC-C-KC-ΔB19R). An immunization protocol mimicking that of the RV144 phase III clinical trial was used. Two groups of macaques received two doses of the corresponding NYVAC-C-KC vectors (weeks 0 and 4) and booster doses with NYVAC-C-KC vectors plus the clade C HIV-1 gp120 protein (weeks 12 and 24). The two replicating NYVAC-C-KC vectors induced enhanced and similar HIV-1-specific CD4+ and CD8+ T cell responses, similar levels of binding IgG antibodies, low levels of IgA antibodies, and high levels of antibody-dependent cellular cytotoxicity responses and HIV-1-neutralizing antibodies. Small differences within the NYVAC-C-KC-ΔB19R group were seen in the magnitude of CD4+ and CD8+ T cells, the induction of some cytokines, and the neutralization of some HIV-1 isolates. Thus, replication-competent NYVAC-C-KC vectors acquired relevant immunological properties as vaccine candidates against HIV/AIDS, and the viral B19 molecule exerts some control of immune functions. IMPORTANCE It is of special importance to find a safe and effective HIV/AIDS vaccine that can induce strong and broad T cell and humoral immune responses correlating with HIV-1

  15. HIV/AIDS Vaccine Candidates Based on Replication-Competent Recombinant Poxvirus NYVAC-C-KC Expressing Trimeric gp140 and Gag-Derived Virus-Like Particles or Lacking the Viral Molecule B19 That Inhibits Type I Interferon Activate Relevant HIV-1-Specific B and T Cell Immune Functions in Nonhuman Primates.

    Science.gov (United States)

    García-Arriaza, Juan; Perdiguero, Beatriz; Heeney, Jonathan L; Seaman, Michael S; Montefiori, David C; Yates, Nicole L; Tomaras, Georgia D; Ferrari, Guido; Foulds, Kathryn E; Roederer, Mario; Self, Steven G; Borate, Bhavesh; Gottardo, Raphael; Phogat, Sanjay; Tartaglia, Jim; Barnett, Susan W; Burke, Brian; Cristillo, Anthony D; Weiss, Deborah E; Lee, Carter; Kibler, Karen V; Jacobs, Bertram L; Wagner, Ralf; Ding, Song; Pantaleo, Giuseppe; Esteban, Mariano

    2017-05-01

    The nonreplicating attenuated poxvirus vector NYVAC expressing clade C(CN54) HIV-1 Env(gp120) and Gag-Pol-Nef antigens (NYVAC-C) showed limited immunogenicity in phase I clinical trials. To enhance the capacity of the NYVAC vector to trigger broad humoral responses and a more balanced activation of CD4+ and CD8+ T cells, here we compared the HIV-1-specific immunogenicity elicited in nonhuman primates immunized with two replicating NYVAC vectors that have been modified by the insertion of the K1L and C7L vaccinia virus host range genes and express the clade C(ZM96) trimeric HIV-1 gp140 protein or a Gag(ZM96)-Pol-Nef(CN54) polyprotein as Gag-derived virus-like particles (termed NYVAC-C-KC). Additionally, one NYVAC-C-KC vector was generated by deleting the viral gene B19R, an inhibitor of the type I interferon response (NYVAC-C-KC-ΔB19R). An immunization protocol mimicking that of the RV144 phase III clinical trial was used. Two groups of macaques received two doses of the corresponding NYVAC-C-KC vectors (weeks 0 and 4) and booster doses with NYVAC-C-KC vectors plus the clade C HIV-1 gp120 protein (weeks 12 and 24). The two replicating NYVAC-C-KC vectors induced enhanced and similar HIV-1-specific CD4+ and CD8+ T cell responses, similar levels of binding IgG antibodies, low levels of IgA antibodies, and high levels of antibody-dependent cellular cytotoxicity responses and HIV-1-neutralizing antibodies. Small differences within the NYVAC-C-KC-ΔB19R group were seen in the magnitude of CD4+ and CD8+ T cells, the induction of some cytokines, and the neutralization of some HIV-1 isolates. Thus, replication-competent NYVAC-C-KC vectors acquired relevant immunological properties as vaccine candidates against HIV/AIDS, and the viral B19 molecule exerts some control of immune functions.IMPORTANCE It is of special importance to find a safe and effective HIV/AIDS vaccine that can induce strong and broad T cell and humoral immune responses correlating with HIV-1 protection

  16. Immunogenicity of a novel Clade B HIV-1 vaccine combination: Results of phase 1 randomized placebo controlled trial of an HIV-1 GM-CSF-expressing DNA prime with a modified vaccinia Ankara vaccine boost in healthy HIV-1 uninfected adults.

    Directory of Open Access Journals (Sweden)

    Susan P Buchbinder

    Full Text Available A phase 1 trial of a clade B HIV vaccine in HIV-uninfected adults evaluated the safety and immunogenicity of a DNA prime co-expressing GM-CSF (Dg followed by different numbers and intervals of modified vaccinia Ankara Boosts (M. Both vaccines produce virus-like particles presenting membrane-bound Env.Four US sites randomized 48 participants to receiving 1/10th the DNA dose as DgDgMMM given at 0, 2, 4, 6 and 8 months, or full dose DgDgM_M or DgDgMM_M regimens, given at 0, 2, 4, and 8 months, and 0, 2, 4, 6, and 10 months, respectively. Peak immunogenicity was measured 2 weeks post-last vaccination.All regimens were well tolerated and safe. Full dose DgDgM_M and DgDgMM_M regimens generated Env-specific IgG to HIV-1 Env in >90%, IgG3 in >80%, and IgA in <20% of participants. Responses to gp140 and gp41 targets were more common and of higher magnitude than to gp120 and V1V2. The gp41 antibody included reactivity to the conserved immunodominant region with specificities known to mediate virus capture and phagocytosis and did not cross-react with a panel of intestinal flora antigens. The 3rd dose of MVA increased the avidity of elicited antibody (7.5% to 39%, the ADCC response to Bal gp120 (14% to 64%, and the one-year durability of the IgG3 responses to gp41 by 4-fold (13% vs. 3.5% retention of peak response. The co-expressed GM-CSF did not enhance responses over those in trials testing this vaccine without GM-CSF.This DNA/MVA prime-boost regimen induced durable, functional humoral responses that included ADCC, high antibody avidity, and Env IgG1 and IgG3 binding responses to the immunodominant region of gp41. The third, spaced MVA boost improved the overall quality of the antibody response. These products without co-expressed GM-CSF but combined with protein boosts will be considered for efficacy evaluation.ClinicalTrials.gov NCT01571960.

  17. Inhibiting sexual transmission of HIV-1 infection.

    Science.gov (United States)

    Shattock, Robin J; Moore, John P

    2003-10-01

    The worldwide infection rate for HIV-1 is estimated to be 14,000 per day, but only now, more than 20 years into the epidemic, are the immediate events between exposure to infectious virus and the establishment of infection becoming clear. Defining the mechanisms of HIV-1 transmission, the target cells involved and how the virus attaches to and fuses with these cells, could reveal ways to block the sexual spread of the virus. In this review, we will discuss how our increasing knowledge of the ways in which HIV-1 is transmitted is shaping the development of new, more sophisticated intervention strategies based on the application of vaginal or rectal microbicides.

  18. Characterization of the virus-cell interactions by HIV-1 subtype C variants from an antiretroviral therapy-naïve subject with baseline resistance to the CCR5 inhibitor maraviroc

    DEFF Research Database (Denmark)

    Jakobsen, Martin Roelsgaard

    resistance to MVC that persisted (as a subset of the viral quasispecies) for approximately 12 months in an antiretroviral therapy-naïve subject infected with HIV-1 subtype C (C-HIV). From a large, longitudinal study of C-HIV Envs (n=323) cloned from 21 subjects experiencing progressive C-HIV infection (see...... in vivo, can use the MVCbound form of CCR5 for HIV-1 entry via adaptive alterations in gp120. Partial baseline resistance to another CCR5 inhibitor through this mechanism, AD101, has been noted recently in one subject (1). Here, we identified and characterized envelope (Env) clones with baseline...... related abstract by Jakobsen et al., “Preferential CCR5-usage by R5X4 subtype C HIV-1 imparts sensitivity to maraviroc and tempers disease progression”), nine subjects persistently harboured CCR5-using (R5) Envs to late stages of infection. Virus inhibition assays in NP2-CD4/CCR5 cells using Env...

  19. Improving the Expression and Purification of Soluble, Recombinant Native-Like HIV-1 Envelope Glycoprotein Trimers by Targeted Sequence Changes.

    Science.gov (United States)

    Ringe, Rajesh P; Ozorowski, Gabriel; Yasmeen, Anila; Cupo, Albert; Cruz Portillo, Victor M; Pugach, Pavel; Golabek, Michael; Rantalainen, Kimmo; Holden, Lauren G; Cottrell, Christopher A; Wilson, Ian A; Sanders, Rogier W; Ward, Andrew B; Klasse, P J; Moore, John P

    2017-06-15

    Soluble, recombinant native-like envelope glycoprotein (Env) trimers of various human immunodeficiency virus type 1 (HIV-1) genotypes are being developed for structural studies and as vaccine candidates aimed at the induction of broadly neutralizing antibodies (bNAbs). The prototypic design is designated SOSIP.664, but many HIV-1 env genes do not yield fully native-like trimers efficiently. One such env gene is CZA97.012 from a neutralization-resistant (tier 2) clade C virus. As appropriately purified, native-like CZA97.012 SOSIP.664 trimers induce autologous neutralizing antibodies (NAbs) efficiently in immunized rabbits, we sought to improve the efficiency with which they can be produced and to better understand the limitations to the original design. By using structure- and antigenicity-guided mutagenesis strategies focused on the V2 and V3 regions and the gp120-gp41 interface, we developed the CZA97 SOSIP.v4.2-M6.IT construct. Fully native-like, stable trimers that display multiple bNAb epitopes could be expressed from this construct in a stable CHO cell line and purified at an acceptable yield using either a PGT145 or a 2G12 bNAb affinity column. We also show that similar mutagenesis strategies can be used to improve the yields and properties of SOSIP.664 trimers of the DU422, 426c, and 92UG037 genotypes.IMPORTANCE Recombinant trimeric proteins based on HIV-1 env genes are being developed for future vaccine trials in humans. A feature of these proteins is their mimicry of the envelope glycoprotein (Env) structure on virus particles that is targeted by neutralizing antibodies, i.e., antibodies that prevent cells from becoming infected. The vaccine concept under exploration is that recombinant trimers may be able to elicit virus-neutralizing antibodies when delivered as immunogens. Because HIV-1 is extremely variable, a practical vaccine may need to incorporate Env trimers derived from multiple different virus sequences. Accordingly, we need to understand how to

  20. Molecular Understanding of HIV-1 Latency

    Directory of Open Access Journals (Sweden)

    W. Abbas

    2012-01-01

    Full Text Available The introduction of highly active antiretroviral therapy (HAART has been an important breakthrough in the treatment of HIV-1 infection and has also a powerful tool to upset the equilibrium of viral production and HIV-1 pathogenesis. Despite the advent of potent combinations of this therapy, the long-lived HIV-1 reservoirs like cells from monocyte-macrophage lineage and resting memory CD4+ T cells which are established early during primary infection constitute a major obstacle to virus eradication. Further HAART interruption leads to immediate rebound viremia from latent reservoirs. This paper focuses on the essentials of the molecular mechanisms for the establishment of HIV-1 latency with special concern to present and future possible treatment strategies to completely purge and target viral persistence in the reservoirs.

  1. Clinical research in HIV-1 infected children

    OpenAIRE

    Fraaij, Pieter

    2005-01-01

    textabstractAcquired immune deficiency syndrome (AIDS) was described for the first time in 1981. Two years later the previously unknown human immunodeficiency virus (HIV) was identified as the causative agent. HIV has been included in the genus Lent/viruses of the Retroviridae family. Two types are recognized: HIV-1 and HIV-2. Of these, HIV-1 is the primary etiologic agent of the current pandemic. HIV probably originates from simian immunodeficiency virus (SIV) which is endemic in African mon...

  2. Prospective Memory in HIV-1 Infection

    OpenAIRE

    CAREY, CATHERINE L.; WOODS, STEVEN PAUL; RIPPETH, JULIE D.; HEATON, ROBERT K.; GRANT, IGOR

    2006-01-01

    The cognitive deficits associated with HIV-1 infection are thought to primarily reflect neuropathophysiology within the fronto-striato-thalamo-cortical circuits. Prospective memory (ProM) is a cognitive function that is largely dependent on prefronto-striatal circuits, but has not previously been examined in an HIV-1 sample. A form of episodic memory, ProM involves the complex processes of forming, monitoring, and executing future intentions vis-à-vis ongoing distractions. The current study e...

  3. Exosomes: Implications in HIV-1 Pathogenesis

    Science.gov (United States)

    Madison, Marisa N.; Okeoma, Chioma M.

    2015-01-01

    Exosomes are membranous nanovesicles of endocytic origin that carry host and pathogen derived genomic, proteomic, and lipid cargos. Exosomes are secreted by most cell types into the extracellular milieu and are subsequently internalized by recipient cells. Upon internalization, exosomes condition recipient cells by donating their cargos and/or activating various signal transduction pathways, consequently regulating physiological and pathophysiological processes. The role of exosomes in viral pathogenesis, especially human immunodeficiency virus type 1 [HIV-1] is beginning to unravel. Recent research reports suggest that exosomes from various sources play important but different roles in the pathogenesis of HIV-1. From these reports, it appears that the source of exosomes is the defining factor for the exosomal effect on HIV-1. In this review, we will describe how HIV-1 infection is modulated by exosomes and in turn how exosomes are targeted by HIV-1 factors. Finally, we will discuss potentially emerging therapeutic options based on exosomal cargos that may have promise in preventing HIV-1 transmission. PMID:26205405

  4. Prospective memory in HIV-1 infection.

    Science.gov (United States)

    Carey, Catherine L; Woods, Steven Paul; Rippeth, Julie D; Heaton, Robert K; Grant, Igor

    2006-05-01

    The cognitive deficits associated with HIV-1 infection are thought to primarily reflect neuropathophysiology within the fronto-striato-thalamo-cortical circuits. Prospective memory (ProM) is a cognitive function that is largely dependent on prefronto-striatal circuits, but has not previously been examined in an HIV-1 sample. A form of episodic memory, ProM involves the complex processes of forming, monitoring, and executing future intentions vis-à-vis ongoing distractions. The current study examined ProM in 42 participants with HIV-1 infection and 29 demographically similar seronegative healthy comparison (HC) subjects. The HIV-1 sample demonstrated deficits in time- and event-based ProM, as well as more frequent 24-hour delay ProM failures and task substitution errors relative to the HC group. In contrast, there were no significant differences in recognition performance, indicating that the HIV-1 group was able to accurately retain and recognize the ProM intention when retrieval demands were minimized. Secondary analyses revealed that ProM performance correlated with validated clinical measures of executive functions, episodic memory (free recall), and verbal working memory, but not with tests of semantic memory, retention, or recognition discrimination. Taken together, these findings indicate that HIV-1 infection is associated with ProM impairment that is primarily driven by a breakdown in the strategic (i.e., executive) aspects of retrieving future intentions, which is consistent with a prefronto-striatal circuit neuropathogenesis.

  5. HIV-1 transmission linkage in an HIV-1 prevention clinical trial

    Energy Technology Data Exchange (ETDEWEB)

    Leitner, Thomas [Los Alamos National Laboratory; Campbell, Mary S [UNIV OF WASHINGTON; Mullins, James I [UNIV OF WASHINGTON; Hughes, James P [UNIV OF WASHINGTON; Wong, Kim G [UNIV OF WASHINGTON; Raugi, Dana N [UNIV OF WASHINGTON; Scrensen, Stefanie [UNIV OF WASHINGTON

    2009-01-01

    HIV-1 sequencing has been used extensively in epidemiologic and forensic studies to investigate patterns of HIV-1 transmission. However, the criteria for establishing genetic linkage between HIV-1 strains in HIV-1 prevention trials have not been formalized. The Partners in Prevention HSV/HIV Transmission Study (ClinicaITrials.gov NCT00194519) enrolled 3408 HIV-1 serodiscordant heterosexual African couples to determine the efficacy of genital herpes suppression with acyclovir in reducing HIV-1 transmission. The trial analysis required laboratory confirmation of HIV-1 linkage between enrolled partners in couples in which seroconversion occurred. Here we describe the process and results from HIV-1 sequencing studies used to perform transmission linkage determination in this clinical trial. Consensus Sanger sequencing of env (C2-V3-C3) and gag (p17-p24) genes was performed on plasma HIV-1 RNA from both partners within 3 months of seroconversion; env single molecule or pyrosequencing was also performed in some cases. For linkage, we required monophyletic clustering between HIV-1 sequences in the transmitting and seroconverting partners, and developed a Bayesian algorithm using genetic distances to evaluate the posterior probability of linkage of participants sequences. Adjudicators classified transmissions as linked, unlinked, or indeterminate. Among 151 seroconversion events, we found 108 (71.5%) linked, 40 (26.5%) unlinked, and 3 (2.0%) to have indeterminate transmissions. Nine (8.3%) were linked by consensus gag sequencing only and 8 (7.4%) required deep sequencing of env. In this first use of HIV-1 sequencing to establish endpoints in a large clinical trial, more than one-fourth of transmissions were unlinked to the enrolled partner, illustrating the relevance of these methods in the design of future HIV-1 prevention trials in serodiscordant couples. A hierarchy of sequencing techniques, analysis methods, and expert adjudication contributed to the linkage

  6. Cyanovirin-N produced in rice endosperm offers effective pre-exposure prophylaxis against HIV-1BaL infection in vitro.

    Science.gov (United States)

    Vamvaka, E; Evans, A; Ramessar, K; Krumpe, L R H; Shattock, R J; O'Keefe, B R; Christou, P; Capell, T

    2016-06-01

    Cyanovirin-N produced in rice endosperm provides efficient pre-exposure prophylaxis against HIV-1 BaL infection in vitro. Cyanovirin-N (CV-N) is a lectin with potent antiviral activity that has been proposed as a component of microbicides for the prevention of infection with Human immunodeficiency virus (HIV). The production of protein-based microbicide components requires a platform that is sufficiently economical and scalable to meet the demands of the large at-risk population, particularly in resource poor developing countries. We, therefore, expressed CV-N in rice endosperm, because the dried seed is ideal for storage and transport and crude extracts could be prepared locally and used as a microbicide component without further purification. We found that crude extracts from rice seeds expressing up to 10 µg CV-N per gram dry seed weight showed dose-dependent gp120 binding activity, confirming that the protein was soluble, correctly folded and active. The recombinant lectin ((OS)CV-N) reduced the infectivity of HIV-1BaL (an R5 virus strain representing the majority of transmitted infections) by ~90 % but showed only weak neutralization activity against HIV-1RF (representative of X4 virus, rarely associated with transmission), suggesting it would be highly effective for pre-exposure prophylaxis against the vast majority of transmitted strains. Crude extracts expressing (OS)CV-N showed no toxicity towards human cells at working dilutions indicating that microbicide components produced in rice endosperm are safe for direct application as topical microbicides in humans.

  7. The Changes of Positive Selection Within env Gene of HIV-1 B', CRF07_BC and CRF08_BC from China Over Time.

    Science.gov (United States)

    Li, Tingting; Sun, Binlian; Jiang, Yanyan; Zeng, Haiyan; Li, Yanpeng; Wang, Yan; Yang, Rongge

    2017-01-01

    It is not clear about the possible evolutionary changes of the three predominant strains of HIV-1 in China over the course of the epidemic. Envelope (env) gene of HIV-1 is a good target for this evolutionary pressure for its enriched epitopes. We collected 159 full or part of env sequences sampled between 1997 and 2010 from database of China, we calculated the genetic distance, detected the positively selected sites by PAML suite and then compared the number using Fisher's exact test between the early period 1997 to 2003 and the late period 2004 to 2010. We found that the diversity of env genes had increased significantly and the positively selected sites were significantly becoming more over time. V2, V4, and C3 regions had suffered an increase positive selection pressure, while V1, V3, V5 and other conserved regions were relatively stable. Several sites were widely selected and compactly located in C3 and V4, five sites were consecutive in V4.There were two common positively selected sites in all groups: 413T in gp120 and 619L in gp41. The common positively selected sites identified in our study implied that they are important in viral survival and adaptation. Based on the role of V3 region in coreceptor determination and disease progression, our results suggested that the virulence of HIV-1 in China might be stable in the short time span. Given the overall increased tendency of positively selection sites in env, we might predict a less virulent state in the long run. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  8. Heterologous prime-boost-boost immunisation of Chinese cynomolgus macaques using DNA and recombinant poxvirus vectors expressing HIV-1 virus-like particles.

    Science.gov (United States)

    Bridge, Simon H; Sharpe, Sally A; Dennis, Mike J; Dowall, Stuart D; Getty, Brian; Anson, Donald S; Skinner, Michael A; Stewart, James P; Blanchard, Tom J

    2011-09-07

    There is renewed interest in the development of poxvirus vector-based HIV vaccines due to the protective effect observed with repeated recombinant canarypox priming with gp120 boosting in the recent Thai placebo-controlled trial. This study sought to investigate whether a heterologous prime-boost-boost vaccine regimen in Chinese cynomolgus macaques with a DNA vaccine and recombinant poxviral vectors expressing HIV virus-like particles bearing envelopes derived from the most prevalent clades circulating in sub-Saharan Africa, focused the antibody response to shared neutralising epitopes. Three Chinese cynomolgus macaques were immunised via intramuscular injections using a regimen composed of a prime with two DNA vaccines expressing clade A Env/clade B Gag followed by boosting with recombinant fowlpox virus expressing HIV-1 clade D Gag, Env and cholera toxin B subunit followed by the final boost with recombinant modified vaccinia virus Ankara expressing HIV-1 clade C Env, Gag and human complement protein C3d. We measured the macaque serum antibody responses by ELISA, enumerated T cell responses by IFN-γ ELISpot and assessed seroneutralisation of HIV-1 using the TZM-bl β-galactosidase assay with primary isolates of HIV-1. This study shows that large and complex synthetic DNA sequences can be successfully cloned in a single step into two poxvirus vectors: MVA and FPV and the recombinant poxviruses could be grown to high titres. The vaccine candidates showed appropriate expression of recombinant proteins with the formation of authentic HIV virus-like particles seen on transmission electron microscopy. In addition the b12 epitope was shown to be held in common by the vaccine candidates using confocal immunofluorescent microscopy. The vaccine candidates were safely administered to Chinese cynomolgus macaques which elicited modest T cell responses at the end of the study but only one out of the three macaques elicited an HIV-specific antibody response. However, the

  9. Heterologous prime-boost-boost immunisation of Chinese cynomolgus macaques using DNA and recombinant poxvirus vectors expressing HIV-1 virus-like particles

    Directory of Open Access Journals (Sweden)

    Anson Donald S

    2011-09-01

    Full Text Available Abstract Background There is renewed interest in the development of poxvirus vector-based HIV vaccines due to the protective effect observed with repeated recombinant canarypox priming with gp120 boosting in the recent Thai placebo-controlled trial. This study sought to investigate whether a heterologous prime-boost-boost vaccine regimen in Chinese cynomolgus macaques with a DNA vaccine and recombinant poxviral vectors expressing HIV virus-like particles bearing envelopes derived from the most prevalent clades circulating in sub-Saharan Africa, focused the antibody response to shared neutralising epitopes. Methods Three Chinese cynomolgus macaques were immunised via intramuscular injections using a regimen composed of a prime with two DNA vaccines expressing clade A Env/clade B Gag followed by boosting with recombinant fowlpox virus expressing HIV-1 clade D Gag, Env and cholera toxin B subunit followed by the final boost with recombinant modified vaccinia virus Ankara expressing HIV-1 clade C Env, Gag and human complement protein C3d. We measured the macaque serum antibody responses by ELISA, enumerated T cell responses by IFN-γ ELISpot and assessed seroneutralisation of HIV-1 using the TZM-bl β-galactosidase assay with primary isolates of HIV-1. Results This study shows that large and complex synthetic DNA sequences can be successfully cloned in a single step into two poxvirus vectors: MVA and FPV and the recombinant poxviruses could be grown to high titres. The vaccine candidates showed appropriate expression of recombinant proteins with the formation of authentic HIV virus-like particles seen on transmission electron microscopy. In addition the b12 epitope was shown to be held in common by the vaccine candidates using confocal immunofluorescent microscopy. The vaccine candidates were safely administered to Chinese cynomolgus macaques which elicited modest T cell responses at the end of the study but only one out of the three macaques

  10. CCR5 inhibitors in HIV-1 therapy.

    Science.gov (United States)

    Dorr, Patrick; Perros, Manos

    2008-11-01

    The human immunodeficiency virus 1 (HIV-1) is the causative pathogen of AIDS, the world's biggest infectious disease killer. About 33 million people are infected worldwide, with 2.1 million deaths a year as a direct consequence. The devastating nature of AIDS has prompted widespread research, which has led to an extensive array of therapies to suppress viral replication and enable recovery of the immune system to prolong and improve patient life substantially. However, the genetic plasticity and replication rate of HIV-1 are considerable, which has lead to rapid drug resistance. This, together with the need for reducing drug side effects and increasing regimen compliance, has led researchers to identify antiretroviral drugs with new modes of action. This review describes the discovery and clinical development of CCR5 antagonists and the recent approval of maraviroc as a breakthrough in anti-HIV-1 therapy. CCR5 inhibitors target a human cofactor to disable HIV-1 entry into the cells, and thereby provide a new hurdle for the virus to overcome. The status and expert opinion of CCR5 antagonists for the treatment of HIV-1 infection are detailed.

  11. Specific Elimination of Latently HIV-1 Infected Cells Using HIV-1 Protease-Sensitive Toxin Nanocapsules.

    Science.gov (United States)

    Wen, Jing; Yan, Ming; Liu, Yang; Li, Jie; Xie, Yiming; Lu, Yunfeng; Kamata, Masakazu; Chen, Irvin S Y

    2016-01-01

    Anti-retroviral drugs suppress HIV-1 plasma viremia to undetectable levels; however, latent HIV-1 persists in reservoirs within HIV-1-infected patients. The silent provirus can be activated through the use of drugs, including protein kinase C activators and histone deacetylase inhibitors. This "shock" approach is then followed by "kill" of the producing cells either through direct HIV-1-induced cell death or natural immune mechanisms. However, these mechanisms are relatively slow and effectiveness is unclear. Here, we develop an approach to specifically target and kill cells that are activated early in the process of virus production. We utilize a novel nanocapsule technology whereby the ricin A chain is encapsulated in an inactive form within a polymer shell. Specificity for release of the ricin A toxin is conferred by peptide crosslinkers that are sensitive to cleavage by HIV-1 protease. By using well-established latent infection models, J-Lat and U1 cells, we demonstrate that only within an HIV-1-producing cell expressing functional HIV-1 protease will the nanocapsule release its ricin A cargo, shutting down viral and cellular protein synthesis, and ultimately leading to rapid death of the producer cell. Thus, we provide proof of principle for a novel technology to kill HIV-1-producing cells without effects on non-target cells.

  12. Identification of HIV-1 Tat-Associated Proteins Contributing to HIV-1 Transcription and Latency

    Science.gov (United States)

    Jean, Maxime Junior; Power, Derek; Kong, Weili; Huang, Huachao; Santoso, Netty; Zhu, Jian

    2017-01-01

    Human immunodeficiency virus type 1 (HIV-1) Tat is a virus-encoded trans-activator that plays a central role in viral transcription. We used our recently developed parallel analysis of in vitro translated open reading frames (ORFs) (PLATO) approach to identify host proteins that associate with HIV-1 Tat. From this proteomic assay, we identify 89 Tat-associated proteins (TAPs). We combine our results with other datasets of Tat or long terminal repeat (LTR)-associated proteins. For some of these proteins (NAT10, TINP1, XRCC5, SIN3A), we confirm their strong association with Tat. These TAPs also suppress Tat-mediated HIV-1 transcription. Removing suppression of HIV-1 transcription benefits the reversal of post-integrated, latent HIV-1 proviruses. We demonstrate that these transcriptionally suppressing TAPs contribute to HIV-1 latency in Jurkat latency (J-LAT) cells. Therefore, our proteomic analysis highlights the previously unappreciated TAPs that play a role in maintaining HIV-1 latency and can be further studied as potential pharmacological targets for the “shock and kill” HIV-1 cure strategy. PMID:28368303

  13. Resistance to V3-directed neutralization caused by an N-linked oligosaccharide depends on the quaternary structure of the HIV-1 envelope oligomer

    DEFF Research Database (Denmark)

    Schønning, Kristian; Jansson, B; Olofsson, S

    1996-01-01

    to wild-type HIVLAI. This difference in sensitivity was not caused by an alteration of the antibody binding site itself, as NEA-9205 had equal affinity for both wild-type and mutant monomeric gp120. In contrast, virion-associated wild-type gp120 was immunoprecipitated less efficiently with NEA-9205 than...

  14. Cross-Reactivity of Anti-HIV-1 T Cell Immune Responses among the Major HIV-1 Clades in HIV-1-Positive Individuals from 4 Continents

    National Research Council Canada - National Science Library

    Paul M. Coplan; Swati B. Gupta; Sheri A. Dubey; Punnee Pitisuttithum; Alex Nikas; Bernard Mbewe; Efthyia Vardas; Mauro Schechter; Esper G. Kallas; Dan C. Freed; Tong-Ming Fu; Christopher T. Mast; Pilaipan Puthavathana; James Kublin; Kelly Brown Collins; John Chisi; Richard Pendame; Scott J. Thaler; Glenda Gray; James Mcintyre; Walter L. Straus; Jon H. Condra; Devan V. Mehrotra; Harry A. Guess; Emilio A. Emini; John W. Shiver

    2005-01-01

    .... Therefore, we quantified the cross-clade reactivity, among unvaccinated individuals, of anti-HIV-1 T cell responses to the infecting HIV-1 clade relative to other major circulating clades. Methods...

  15. HIV-1 Reservoir Association with Immune Activation

    Directory of Open Access Journals (Sweden)

    Alejandro Vallejo

    2015-09-01

    Full Text Available In this issue of EBioMedicine, Ruggiero and colleagues describe immune activation biomarkers associated with the size of the HIV reservoir in a carefully designed cross-sectional study. The cohort consists of a homogeneous sample of HIV-1-infected patients with long-term plasma HIV-1 RNA suppression under antiretroviral treatment (ART. It is crucial to explore the potential utility of biomarkers that are easier (less labor intensive, less expensive to measure than integrated HIV DNA load, in order to quickly and accurately quantify cellular reservoirs of HIV.

  16. Viral linkage in HIV-1 seroconverters and their partners in an HIV-1 prevention clinical trial.

    Directory of Open Access Journals (Sweden)

    Mary S Campbell

    2011-03-01

    Full Text Available Characterization of viruses in HIV-1 transmission pairs will help identify biological determinants of infectiousness and evaluate candidate interventions to reduce transmission. Although HIV-1 sequencing is frequently used to substantiate linkage between newly HIV-1 infected individuals and their sexual partners in epidemiologic and forensic studies, viral sequencing is seldom applied in HIV-1 prevention trials. The Partners in Prevention HSV/HIV Transmission Study (ClinicalTrials.gov #NCT00194519 was a prospective randomized placebo-controlled trial that enrolled serodiscordant heterosexual couples to determine the efficacy of genital herpes suppression in reducing HIV-1 transmission; as part of the study analysis, HIV-1 sequences were examined for genetic linkage between seroconverters and their enrolled partners.We obtained partial consensus HIV-1 env and gag sequences from blood plasma for 151 transmission pairs and performed deep sequencing of env in some cases. We analyzed sequences with phylogenetic techniques and developed a Bayesian algorithm to evaluate the probability of linkage. For linkage, we required monophyletic clustering between enrolled partners' sequences and a Bayesian posterior probability of ≥ 50%. Adjudicators classified each seroconversion, finding 108 (71.5% linked, 40 (26.5% unlinked, and 3 (2.0% indeterminate transmissions, with linkage determined by consensus env sequencing in 91 (84%. Male seroconverters had a higher frequency of unlinked transmissions than female seroconverters. The likelihood of transmission from the enrolled partner was related to time on study, with increasing numbers of unlinked transmissions occurring after longer observation periods. Finally, baseline viral load was found to be significantly higher among linked transmitters.In this first use of HIV-1 sequencing to establish endpoints in a large clinical trial, more than one-fourth of transmissions were unlinked to the enrolled partner

  17. Epidemiology of HIV-1 and emerging problems

    NARCIS (Netherlands)

    Lukashov, V. V.; de Ronde, A.; de Jong, J. J.; Goudsmit, J.

    2000-01-01

    Broad use of antiretroviral drugs is becoming a factor that is important to consider for understanding the HIV-1 epidemiology. Since 1993, we observe that a proportion of new infections within major risk groups in Amsterdam is caused by azidothymidine (AZT)-resistant viruses. After the introduction

  18. HIV-1 transcription and latency: an update.

    Science.gov (United States)

    Van Lint, Carine; Bouchat, Sophie; Marcello, Alessandro

    2013-06-26

    Combination antiretroviral therapy, despite being potent and life-prolonging, is not curative and does not eradicate HIV-1 infection since interruption of treatment inevitably results in a rapid rebound of viremia. Reactivation of latently infected cells harboring transcriptionally silent but replication-competent proviruses is a potential source of persistent residual viremia in cART-treated patients. Although multiple reservoirs may exist, the persistence of resting CD4+ T cells carrying a latent infection represents a major barrier to eradication. In this review, we will discuss the latest reports on the molecular mechanisms that may regulate HIV-1 latency at the transcriptional level, including transcriptional interference, the role of cellular factors, chromatin organization and epigenetic modifications, the viral Tat trans-activator and its cellular cofactors. Since latency mechanisms may also operate at the post-transcriptional level, we will consider inhibition of nuclear RNA export and inhibition of translation by microRNAs as potential barriers to HIV-1 gene expression. Finally, we will review the therapeutic approaches and clinical studies aimed at achieving either a sterilizing cure or a functional cure of HIV-1 infection, with a special emphasis on the most recent pharmacological strategies to reactivate the latent viruses and decrease the pool of viral reservoirs.

  19. Molecular mechanisms of HIV-1 associated neurodegeneration

    Indian Academy of Sciences (India)

    Unknown

    Erichsen D, Lopez A L, Peng H, Niemann D, Williams C,. Bauer M, Morgello S, Cotter R L, Ryan L A, Ghorpade A,. Gendelman H E and Zheng J 2003 Neuronal injury regulates fractalkine: relevance for HIV-1 associated dementia; J. Neu- roimmunol. 138 144–155. Enting R H, Hoetelmans R M, Lange J M, Burger D M and.

  20. Molecular mechanisms of HIV-1 associated neurodegeneration

    Indian Academy of Sciences (India)

    Since identification of the human immunodeficiency virus-1 (HIV-1), numerous studies suggest a link between neurological impairments, in particular dementia, with acquired immunodeficiency syndrome (AIDS) with alarming occurrence worldwide. Approximately, 60% of HIV-infected people show some form of neurological ...

  1. Epigenetic heterogeneity in HIV-1 latency establishment.

    Science.gov (United States)

    Matsuda, Yuka; Kobayashi-Ishihara, Mie; Fujikawa, Dai; Ishida, Takaomi; Watanabe, Toshiki; Yamagishi, Makoto

    2015-01-09

    Despite prolonged antiretroviral therapy, HIV-1 persists as transcriptionally inactive proviruses. The HIV-1 latency remains a principal obstacle in curing AIDS. It is important to understand mechanisms by which HIV-1 latency is established to make the latent reservoir smaller. We present a molecular characterization of distinct populations at an early phase of infection. We developed an original dual-color reporter virus to monitor LTR kinetics from establishment to maintenance stage. We found that there are two ways of latency establishment i.e., by immediate silencing and slow inactivation from active infection. Histone covalent modifications, particularly polycomb repressive complex 2 (PRC2)-mediated H3K27 trimethylation, appeared to dominate viral transcription at the early phase. PRC2 also contributes to time-dependent LTR dormancy in the chronic phase of the infection. Significant differences in sensitivity against several stimuli were observed between these two distinct populations. These results will expand our understanding of heterogeneous establishment of HIV-1 latency populations.

  2. Enteric viruses in HIV-1 seropositive and HIV-1 seronegative children with diarrheal diseases in Brazil

    Science.gov (United States)

    Rocha, Monica Simões; Fumian, Tulio Machado; Maranhão, Adriana Gonçalves; de Assis, Rosane Maria; Xavier, Maria da Penha Trindade Pinheiro; Rocha, Myrna Santos; Miagostovich, Marize Pereira; Leite, José Paulo Gagliardi; Volotão, Eduardo de Mello

    2017-01-01

    Diarrheal diseases (DD) have distinct etiological profiles in immune-deficient and immune-competent patients. This study compares detection rates, genotype distribution and viral loads of different enteric viral agents in HIV-1 seropositive (n = 200) and HIV-1 seronegative (n = 125) children hospitalized with DD in Rio de Janeiro, Brazil. Except for group A rotavirus (RVA), which were detected through enzyme immunoassay, the other enteric viruses (norovirus [NoV], astrovirus [HAstV], adenovirus [HAdV] and bocavirus [HBoV]) were detected through PCR or RT-PCR. A quantitative PCR was performed for RVA, NoV, HAstV, HAdV and HBoV. Infections with NoV (19% vs. 9.6%; p<0.001), HBoV (14% vs. 7.2%; p = 0.042) and HAdV (30.5% vs. 14.4%; p<0.001) were significantly more frequent among HIV-1 seropositive children. RVA was significantly less frequent among HIV-1 seropositive patients (6.5% vs. 20%; p<0.001). Similarly, frequency of infection with HAstV was lower among HIV-1 seropositive children (5.5% vs. 12.8%; p = 0.018). Among HIV-1 seropositive children 33 (16.5%) had co-infections, including three enteric viruses, such as NoV, HBoV and HAdV (n = 2) and NoV, HAstV and HAdV (n = 2). The frequency of infection with more than one virus was 17 (13.6%) in the HIV-1 negative group, triple infection (NoV + HAstV + HBoV) being observed in only one patient. The median viral load of HAstV in feces was significantly higher among HIV-1 positive children compared to HIV-1 negative children. Concerning children infected with RVA, NoV, HBoV and HAdV, no statistically significant differences were observed in the medians of viral loads in feces, comparing HIV-1 seropositive and HIV-1 seronegative children. Similar detection rates were observed for RVA, HAstV and HAdV, whilst NoV and HBoV were significantly more prevalent among children with CD4+ T lymphocyte count below 200 cells/mm3. Enteric viruses should be considered an important cause of DD in HIV-1 seropositive children, along with

  3. Enteric viruses in HIV-1 seropositive and HIV-1 seronegative children with diarrheal diseases in Brazil.

    Directory of Open Access Journals (Sweden)

    Silvana Augusta Rodrigues Portes

    Full Text Available Diarrheal diseases (DD have distinct etiological profiles in immune-deficient and immune-competent patients. This study compares detection rates, genotype distribution and viral loads of different enteric viral agents in HIV-1 seropositive (n = 200 and HIV-1 seronegative (n = 125 children hospitalized with DD in Rio de Janeiro, Brazil. Except for group A rotavirus (RVA, which were detected through enzyme immunoassay, the other enteric viruses (norovirus [NoV], astrovirus [HAstV], adenovirus [HAdV] and bocavirus [HBoV] were detected through PCR or RT-PCR. A quantitative PCR was performed for RVA, NoV, HAstV, HAdV and HBoV. Infections with NoV (19% vs. 9.6%; p<0.001, HBoV (14% vs. 7.2%; p = 0.042 and HAdV (30.5% vs. 14.4%; p<0.001 were significantly more frequent among HIV-1 seropositive children. RVA was significantly less frequent among HIV-1 seropositive patients (6.5% vs. 20%; p<0.001. Similarly, frequency of infection with HAstV was lower among HIV-1 seropositive children (5.5% vs. 12.8%; p = 0.018. Among HIV-1 seropositive children 33 (16.5% had co-infections, including three enteric viruses, such as NoV, HBoV and HAdV (n = 2 and NoV, HAstV and HAdV (n = 2. The frequency of infection with more than one virus was 17 (13.6% in the HIV-1 negative group, triple infection (NoV + HAstV + HBoV being observed in only one patient. The median viral load of HAstV in feces was significantly higher among HIV-1 positive children compared to HIV-1 negative children. Concerning children infected with RVA, NoV, HBoV and HAdV, no statistically significant differences were observed in the medians of viral loads in feces, comparing HIV-1 seropositive and HIV-1 seronegative children. Similar detection rates were observed for RVA, HAstV and HAdV, whilst NoV and HBoV were significantly more prevalent among children with CD4+ T lymphocyte count below 200 cells/mm3. Enteric viruses should be considered an important cause of DD in HIV-1 seropositive children, along

  4. An anti-HIV-1 V3 loop antibody fully protects cross-clade and elicits T-cell immunity in macaques mucosally challenged with an R5 clade C SHIV.

    Directory of Open Access Journals (Sweden)

    Jennifer D Watkins

    2011-03-01

    Full Text Available Neutralizing antibodies have been shown to protect macaques against SHIV challenge. However, genetically diverse HIV-1 clades have evolved, and a key question left unanswered is whether neutralizing antibodies can confer cross-clade protection in vivo. The novel human monoclonal antibody HGN194 was isolated from an individual infected with an HIV-1 clade AG recombinant circulating recombinant form (CRF. HGN194 targets an epitope in the third hypervariable loop (V3 of HIV-1 gp120 and neutralizes a range of relatively neutralization-sensitive and resistant viruses. We evaluated the potential of HGN194 to protect infant rhesus monkeys against a SHIV encoding a primary CCR5-tropic HIV-1 clade C envelope. After high-dose mucosal challenge, all untreated controls became highly viremic while all HGN194-treated animals (50 mg/kg were completely protected. When HGN194 was given at 1 mg/kg, one out of two monkeys remained aviremic, whereas the other had delayed, lower peak viremia. Interestingly, all protected monkeys given high-dose HGN194 developed Gag-specific proliferative responses of both CD4+ and CD8+ T cells. To test whether generation of the latter involved cryptic infection, we ablated CD8+ cells after HGN194 clearance. No viremia was detected in any protected monkeys, thus ruling out virus reservoirs. Thus, induction of CD8 T-cell immunity may have resulted from transient "Hit and Run" infection or cross priming via Ag-Ab-mediated cross-presentation. Together, our data identified the HGN194 epitope as protective and provide proof-of-concept that this anti-V3 loop mAb can prevent infection with sterilizing immunity after challenge with virus of a different clade, implying that V3 is a potential vaccine target.

  5. Modeling Anti-HIV-1 HSPC-Based Gene Therapy in Humanized Mice Previously Infected with HIV-1.

    Science.gov (United States)

    Khamaikawin, Wannisa; Shimizu, Saki; Kamata, Masakazu; Cortado, Ruth; Jung, Yujin; Lam, Jennifer; Wen, Jing; Kim, Patrick; Xie, Yiming; Kim, Sanggu; Arokium, Hubert; Presson, Angela P; Chen, Irvin S Y; An, Dong Sung

    2018-06-15

    Investigations of anti-HIV-1 human hematopoietic stem/progenitor cell (HSPC)-based gene therapy have been performed by HIV-1 challenge after the engraftment of gene-modified HSPCs in humanized mouse models. However, the clinical application of gene therapy is to treat HIV-1-infected patients. Here, we developed a new method to investigate an anti-HIV-1 HSPC-based gene therapy in humanized mice previously infected with HIV-1. First, humanized mice were infected with HIV-1. When plasma viremia reached >107 copies/mL 3 weeks after HIV-1 infection, the mice were myeloablated with busulfan and transplanted with anti-HIV-1 gene-modified CD34+ HSPCs transduced with a lentiviral vector expressing two short hairpin RNAs (shRNAs) against CCR5 and HIV-1 long terminal repeat (LTR), along with human thymus tissue under the kidney capsule. Anti-HIV-1 vector-modified human CD34+ HSPCs successfully repopulated peripheral blood and lymphoid tissues in HIV-1 previously infected humanized mice. Anti-HIV-1 shRNA vector-modified CD4+ T lymphocytes showed selective advantage in HIV-1 previously infected humanized mice. This new method will be useful for investigations of anti-HIV-1 gene therapy when testing in a more clinically relevant experimental setting.

  6. Identification of Acute HIV-1 Infection by Hologic Aptima HIV-1 RNA Qualitative Assay

    Science.gov (United States)

    Eller, Leigh Anne; Malia, Jennifer; Jagodzinski, Linda L.; Trichavaroj, Rapee; Oundo, Joseph; Lueer, Cornelia; Cham, Fatim; de Souza, Mark; Michael, Nelson L.; Robb, Merlin L.; Peel, Sheila A.

    2017-01-01

    ABSTRACT The Hologic Aptima HIV-1 Qualitative RNA assay was used in a rigorous screening approach designed to identify individuals at the earliest stage of HIV-1 infection for enrollment into subsequent studies of cellular and viral events in early infection (RV 217/Early Capture HIV Cohort [ECHO] study). Volunteers at high risk for HIV-1 infection were recruited from study sites in Thailand, Tanzania, Uganda, and Kenya with high HIV-1 prevalence rates among the populations examined. Small-volume blood samples were collected by finger stick at twice-weekly intervals and tested with the Aptima assay. Participants with reactive Aptima test results were contacted immediately for entry into a more comprehensive follow-up schedule with frequent blood draws. Evaluation of the Aptima test prior to use in this study showed a detection sensitivity of 5.5 copies/ml (50%), with all major HIV-1 subtypes detected. A total of 54,306 specimens from 1,112 volunteers were examined during the initial study period (August 2009 to November 2010); 27 individuals were identified as converting from uninfected to infected status. A sporadic reactive Aptima signal was observed in HIV-1-infected individuals under antiretroviral therapy. Occasional false-reactive Aptima results in uninfected individuals, or nonreactive results in HIV-1-infected individuals not on therapy, were observed and used to calculate assay sensitivity and specificity. The sensitivity and specificity of the Aptima assay were 99.03% and 99.23%, respectively; positive and negative predictive values were 92.01% and 99.91%, respectively. Conversion from HIV-1-uninfected to -infected status was rapid, with no evidence of a prolonged period of intermittent low-level viremia. PMID:28424253

  7. Picomolar dichotomous activity of gnidimacrin against HIV-1.

    Directory of Open Access Journals (Sweden)

    Li Huang

    Full Text Available Highly active antiretroviral therapy (HAART has offered a promising approach for controlling HIV-1 replication in infected individuals. However, with HARRT, HIV-1 is suppressed rather than eradicated due to persistence of HIV-1 in latent viral reservoirs. Thus, purging the virus from latent reservoirs is an important strategy toward eradicating HIV-1 infection. In this study, we discovered that the daphnane diterpene gnidimacrin, which was previously reported to have potent anti-cancer cell activity, activated HIV-1 replication and killed persistently-infected cells at picomolar concentrations. In addition to its potential to purge HIV-1 from latently infected cells, gnidimacrin potently inhibited a panel of HIV-1 R5 virus infection of peripheral blood mononuclear cells (PBMCs at an average concentration lower than 10 pM. In contrast, gnidimacrin only partially inhibited HIV-1 ×4 virus infection of PBMCs. The strong anti-HIV-1 R5 virus activity of gnidimacrin was correlated with its effect on down-regulation of the HIV-1 coreceptor CCR5. The anti-R5 virus activity of gnidimacrin was completely abrogated by a selective protein kinase C beta inhibitor enzastaurin, which suggests that protein kinase C beta plays a key role in the potent anti-HIV-1 activity of gnidimacrin in PBMCs. In summary, these results suggest that gnidimacrin could activate latent HIV-1, specifically kill HIV-1 persistently infected cells, and inhibit R5 viruses at picomolar concentrations.

  8. Picomolar dichotomous activity of gnidimacrin against HIV-1.

    Science.gov (United States)

    Huang, Li; Ho, Phong; Yu, Jie; Zhu, Lei; Lee, Kuo-Hsiung; Chen, Chin-Ho

    2011-01-01

    Highly active antiretroviral therapy (HAART) has offered a promising approach for controlling HIV-1 replication in infected individuals. However, with HARRT, HIV-1 is suppressed rather than eradicated due to persistence of HIV-1 in latent viral reservoirs. Thus, purging the virus from latent reservoirs is an important strategy toward eradicating HIV-1 infection. In this study, we discovered that the daphnane diterpene gnidimacrin, which was previously reported to have potent anti-cancer cell activity, activated HIV-1 replication and killed persistently-infected cells at picomolar concentrations. In addition to its potential to purge HIV-1 from latently infected cells, gnidimacrin potently inhibited a panel of HIV-1 R5 virus infection of peripheral blood mononuclear cells (PBMCs) at an average concentration lower than 10 pM. In contrast, gnidimacrin only partially inhibited HIV-1 ×4 virus infection of PBMCs. The strong anti-HIV-1 R5 virus activity of gnidimacrin was correlated with its effect on down-regulation of the HIV-1 coreceptor CCR5. The anti-R5 virus activity of gnidimacrin was completely abrogated by a selective protein kinase C beta inhibitor enzastaurin, which suggests that protein kinase C beta plays a key role in the potent anti-HIV-1 activity of gnidimacrin in PBMCs. In summary, these results suggest that gnidimacrin could activate latent HIV-1, specifically kill HIV-1 persistently infected cells, and inhibit R5 viruses at picomolar concentrations.

  9. Assessment of HIV-1 entry inhibitors by MLV/HIV-1 pseudotyped vectors

    Directory of Open Access Journals (Sweden)

    Thaler Sonja

    2005-09-01

    Full Text Available Abstract Background Murine leukemia virus (MLV vector particles can be pseudotyped with a truncated variant of the human immunodeficiency virus type 1 (HIV-1 envelope protein (Env and selectively target gene transfer to human cells expressing both CD4 and an appropriate co-receptor. Vector transduction mimics the HIV-1 entry process and is therefore a safe tool to study HIV-1 entry. Results Using FLY cells, which express the MLV gag and pol genes, we generated stable producer cell lines that express the HIV-1 envelope gene and a retroviral vector genome encoding the green fluorescent protein (GFP. The BH10 or 89.6 P HIV-1 Env was expressed from a bicistronic vector which allowed the rapid selection of stable cell lines. A codon-usage-optimized synthetic env gene permitted high, Rev-independent Env expression. Vectors generated by these producer cells displayed different sensitivity to entry inhibitors. Conclusion These data illustrate that MLV/HIV-1 vectors are a valuable screening system for entry inhibitors or neutralizing antisera generated by vaccines.

  10. Dual role of autophagy in HIV-1 replication and pathogenesis

    Directory of Open Access Journals (Sweden)

    Killian M

    2012-05-01

    Full Text Available Abstract Autophagy, the major mechanism for degrading long-lived intracellular proteins and organelles, is essential for eukaryotic cell homeostasis. Autophagy also defends the cell against invasion by microorganisms and has important roles in innate and adaptive immunity. Increasingly evident is that HIV-1 replication is dependent on select components of autophagy. Fittingly, HIV-1 proteins are able to modulate autophagy to maximize virus production. At the same time, HIV-1 proteins appear to disrupt autophagy in uninfected cells, thereby contributing to CD4+ cell death and HIV-1 pathogenesis. These observations allow for new approaches for the treatment and possibly the prevention of HIV-1 infection. This review focuses on the relationship between autophagy and HIV-1 infection. Discussed is how autophagy plays dual roles in HIV-1 replication and HIV-1 disease progression.

  11. Intestinal microbiota and HIV-1 infection

    Directory of Open Access Journals (Sweden)

    E. B. S. M. Trindade

    2007-01-01

    Full Text Available The intestinal microbiota consists of a qualitatively and quantitatively diverse range of microorganisms dynamically interacting with the host. It is remarkably stable with regard to the presence of microorganisms and their roles which, however, can be altered due to pathological conditions, diet composition, gastrointestinal disturbances and/or drug ingestion. The present review aimed at contributing to the discussion about changes in the intestinal microbiota due to HIV-1 infection, focusing on the triad infection-microbiota-nutrition as factors that promote intestinal bacterial imbalance. Intestinal microbiota alterations can be due to the HIV-1 infection as a primary factor or the pharmacotherapy employed, or they can be one of the consequences of the disease.

  12. Limits on oral transmission of HIV-1.

    Science.gov (United States)

    Cohen, M S; Shugars, D C; Fiscus, S A

    2000-07-22

    This article discusses the potential of acquiring an HIV-1 infection through an oral route, with a view of offering clues for its prevention. In a study of adult animals given low concentration cell-free simian immunodeficiency virus (SIV) orally, histological examination suggested that SIV infected lymphoid tissue through the antigen-transporting crypt epithelium rather than through dendritic cells. The investigators found no evidence of acquiring SIV via the gastrointestinal tract. For humans, HIV transmission from saliva or intimate family contact seems to be extremely rare. This could be because of the low concentration of HIV-1 in saliva. A study of 40 people found that significantly less HIV was found in salivary secretions than in plasma. Another possible explanation for inefficient oral transmission might be that HIV-1 in the oropharynx is inhibited by components found in salivary secretions. Conversely, studies have noted that risk of oral transmission of HIV from contaminated breast milk and semen is higher than from saliva. Breast-feeding by an HIV-infected woman puts the baby at substantial risk of infection and receptive fellatio cannot be considered a safe sex act.

  13. HIV-1 vaccine design: Learning from natural infection

    NARCIS (Netherlands)

    van den Kerkhof, T.L.G.M.

    2016-01-01

    Het humane immuundeficiëntie virus type 1 (hiv-1) is het virus dat aids veroorzaakt. Er is nog steeds geen bescherming tegen een hiv-1 infectie en de beëindiging van de wereldwijde epidemie kan waarschijnlijk alleen worden bereikt met behulp van een vaccin. Een hiv-1 vaccin zal bescherming moeten

  14. HIV-1 envelope trimer fusion proteins and their applications

    NARCIS (Netherlands)

    Sliepen, K.H.E.W.J.

    2016-01-01

    HIV-1 is a major threat to global health and a vaccine is not yet on the horizon. A successful HIV-1 vaccine should probably induce HIV-1 neutralizing antibodies that target the envelope glycoprotein (Env) trimer on the outside of the virion. A possible starting point for such a vaccine are soluble

  15. HIV-1 and SIV Predominantly Use CCR5 Expressed on a Precursor Population to Establish Infection in T Follicular Helper Cells

    Directory of Open Access Journals (Sweden)

    John Zaunders

    2017-04-01

    Full Text Available BackgroundT follicular helper (Tfh cells are increasingly recognized as a major reservoir of HIV infection that will likely need to be addressed in approaches to curing HIV. However, Tfh express minimal CCR5, the major coreceptor for HIV-1, and the mechanism by which they are infected is unclear. We have previously shown that macaque Tfh lack CCR5, but are infected in vivo with CCR5-using SIV at levels comparable to other memory CD4+ T cells. Similarly, human splenic Tfh cells are highly infected with HIV-1 DNA. Therefore, we set out to examine the mechanism of infection of Tfh cells.MethodologyTfh and other CD4+ T cell subsets from macaque lymph nodes and spleens, splenic Tfh from HIV+ subjects, and tonsillar Tfh from HIV-uninfected subjects were isolated by cell sorting prior to cell surface and molecular characterization. HIV proviral gp120 sequences were submitted to genotypic and phenotypic tropism assays. Entry of CCR5- and CXCR4-using viruses into Tfh from uninfected tonsillar tissue was measured using a fusion assay.ResultsPhylogenetic analysis, genotypic, and phenotypic analysis showed that splenic Tfh cells from chronic HIV+ subjects were predominantly infected with CCR5-using viruses. In macaques, purified CCR5+PD-1intermediate(int+ memory CD4+ T cells were shown to include pre-Tfh cells capable of differentiating in vitro to Tfh by upregulation of PD-1 and Bcl6, confirmed by qRT-PCR and single-cell multiplex PCR. Infected PD-1int cells survive, carry SIV provirus, and differentiate into PD-1hi Tfh after T cell receptor stimulation, suggesting a pathway for SIV infection of Tfh. In addition, a small subset of macaque and human PD-1hi Tfh can express low levels of CCR5, which makes them susceptible to infection. Fusion assays demonstrated CCR5-using HIV-1 entry into CCR5+ Tfh and pre-Tfh cells from human tonsils.ConclusionThe major route of infection of Tfh in macaques and humans appears to be via a CCR5-expressing pre-Tfh population

  16. Quantification of the Epitope Diversity of HIV-1-Specific Binding Antibodies by Peptide Microarrays for Global HIV-1 Vaccine Development

    OpenAIRE

    Stephenson, Kathryn E.; Neubauer, George H.; Reimer, Ulf; Pawlowski, Nikolaus; Knaute, Tobias; Zerweck, Johannes; Korber, Bette T; Barouch, Dan H.

    2014-01-01

    An effective vaccine against human immunodeficiency virus type 1 (HIV-1) will have to provide protection against a vast array of different HIV-1 strains. Current methods to measure HIV-1-specific binding antibodies following immunization typically focus on determining the magnitude of antibody responses, but the epitope diversity of antibody responses has remained largely unexplored. Here we describe the development of a global HIV-1 peptide microarray that contains 6,564 peptides from across...

  17. Evaluation of Xpert HIV-1 Qual assay for resolution of HIV-1 infection in samples with negative or indeterminate Geenius HIV-1/2 results.

    Science.gov (United States)

    Michaeli, Michal; Wax, Marina; Gozlan, Yael; Rakovsky, Aviya; Mendelson, Ella; Mor, Orna

    2016-03-01

    Diagnosis of HIV infection is a multistage algorithm. Following screening with 4(th) generation combination immunoassay, confirmation of HIV infection is performed with an antibody assay that differentiates HIV-1 from HIV-2 infection. In the newly updated algorithm, samples that are nonreactive or indeterminate in the differentiation assay are to be tested with an HIV-1 nucleic acid amplification (NAAT) test for resolution. Xpert HIV-1 Qual is a new NAAT assay approved for the identification of HIV infection in whole and dried blood. To assess the performance of Xpert HIV-1 Qual supplementary assay in resolving the clinical status of serum samples reactive by 4(th) generation immunoassays and indeterminate or negative by Geenius HIV-1/2 confirmatory assay. In a retrospective study, samples from 97 individuals for whom the true HIV-1 status was already known (by follow-up samples) and which were negative or indeterminate by HIV-1/2 Geenius assay were tested with Xpert Qual HIV-1 assay. Xpert Qual assay correctly classified all 97 samples from HIV-1 positive (n=49) and negative (n=48) individuals. The sensitivity and specificity of Xpert Qual when using the true HIV status as a reference were 100% (92.7-100% at 95% confidence interval [CI] and 92.6-100% at 95% CI, respectively). Applying Xpert Qual HIV-1 assay in the new HIV multi-stage diagnostic algorithm correctly classified 100% of HIV-1 infections including 49 from HIV-1 carriers who have not yet seroconverted. With this assay the total time required for acute HIV diagnosis could be significantly reduced. Copyright © 2015 Elsevier B.V. All rights reserved.

  18. Photochemical neutralization of HIV-1 and inhibition of HIV-1 induced syncytium formation

    Energy Technology Data Exchange (ETDEWEB)

    Lewis, D.E.; Utecht, R.E. [South Dakota State Univ., Brookings, SD (United States); Chanh, T.C.; Allan, J.S. [Southwest Foundation for Biomedical Research, San Antonio, TX (United States); Sogandares-Bernal, F.; Judy, M.M.; Matthews J.L. [Baylor Research Foundation, Dallas, TX (United States)

    1993-12-31

    The authors have prepared a new class of photochemically activatable antiviral compounds based on the 1,8-naphthalimide skeleton which are excited by visible (420 nm) light, and which are highly effective in causing neutralization of enveloped viruses including HIV-1, HSV-1, and VSV. One such photoactive compound, 1,14-bis-(N-hexyl-3-bromo-1,8-naphthalimid-4-yl)-1,4,11,14-tetrazatetradecane-5,10-dione (diED66Br) effectively neutralized HIV-1 in vitro at concentrations below .1{mu}M; similar results are obtained for HSV-1 and VSV. DiED66Br also effectively inhibits syncytium formation induced by cells infected with HIV-1 at doses which had no effect on normal human blood peripheral mononuclear cells. The synthesis of the photochemically active compounds and the mode of antiviral action will be discussed.

  19. Defining HIV-1 transmission clusters based on sequence data.

    Science.gov (United States)

    Hassan, Amin S; Pybus, Oliver G; Sanders, Eduard J; Albert, Jan; Esbjörnsson, Joakim

    2017-06-01

    : Understanding HIV-1 transmission dynamics is relevant to both screening and intervention strategies of HIV-1 infection. Commonly, HIV-1 transmission chains are determined based on sequence similarity assessed either directly from a sequence alignment or by inferring a phylogenetic tree. This review is aimed at both nonexperts interested in understanding and interpreting studies of HIV-1 transmission, and experts interested in finding the most appropriate cluster definition for a specific dataset and research question. We start by introducing the concepts and methodologies of how HIV-1 transmission clusters usually have been defined. We then present the results of a systematic review of 105 HIV-1 molecular epidemiology studies summarizing the most common methods and definitions in the literature. Finally, we offer our perspectives on how HIV-1 transmission clusters can be defined and provide some guidance based on examples from real life datasets.

  20. Factors of intermittent HIV-1 excretion in semen and efficiency of sperm processing in obtaining spermatozoa without HIV-1 genomes.

    Science.gov (United States)

    Bujan, Louis; Daudin, Myriam; Matsuda, Tomohiro; Righi, Laurence; Thauvin, Laurence; Berges, Laetitia; Izopet, Jacques; Berrebi, Alain; Massip, Patrice; Pasquier, Christophe

    2004-03-26

    To study the risk factors for HIV-1 in semen according to the localization of HIV-1 in sperm cell fractions and to assess the efficiency of sperm processing in obtaining spermatozoa without HIV-1 genomes. Ninety-four HIV-infected patients provided 281 paired blood and semen samples. Sperm cell separation was performed using two successive methods. HIV-1 RNA was quantified in blood and seminal plasma. HIV-1 RNA and DNA were detected in cell fractions. HIV-1 RNA was found in 14% of seminal plasma samples and up to 8.7% of native semen cells were positive for HIV-1 RNA and DNA. Ten seminal plasma samples had detectable RNA although blood viral load was undetectable. Antiretroviral treatment reduced the likelihood of RNA detection in seminal plasma. For semen with polynuclear cells and HIV-1 RNA in seminal plasma, the likelihood of detecting HIV-1 genomes in semen cells was increased fourfold and sixfold, respectively. In 25% of patients, HIV-1 excretion was intermittent. In the group of patients with systematic negative seminal plasma, HIV-1 genomes were detected in up to 10% of sperm cell samples. Our method of sperm processing always enabled us to obtain spermatozoa without detectable HIV-1 genomes. Polynuclear cells in semen are a risk factor for seminal HIV-1 excretion. Blood viral load was the only predictive factor for the intermittence of HIV-1 excretion in semen over time. Sperm processing using two successive methods was effective in obtaining spermatozoa without detectable HIV-1 genomes regardless of the viral load level in native semen.

  1. Plasmacytoid dendritic cells suppress HIV-1 replication but contribute to HIV-1 induced immunopathogenesis in humanized mice.

    Directory of Open Access Journals (Sweden)

    Guangming Li

    2014-07-01

    Full Text Available The role of plasmacytoid dendritic cells (pDC in human immunodeficiency virus type 1 (HIV-1 infection and pathogenesis remains unclear. HIV-1 infection in the humanized mouse model leads to persistent HIV-1 infection and immunopathogenesis, including type I interferons (IFN-I induction, immune-activation and depletion of human leukocytes, including CD4 T cells. We developed a monoclonal antibody that specifically depletes human pDC in all lymphoid organs in humanized mice. When pDC were depleted prior to HIV-1 infection, the induction of IFN-I and interferon-stimulated genes (ISGs were abolished during acute HIV-1 infection with either a highly pathogenic CCR5/CXCR4-dual tropic HIV-1 or a standard CCR5-tropic HIV-1 isolate. Consistent with the anti-viral role of IFN-I, HIV-1 replication was significantly up-regulated in pDC-depleted mice. Interestingly, the cell death induced by the highly pathogenic HIV-1 isolate was severely reduced in pDC-depleted mice. During chronic HIV-1 infection, depletion of pDC also severely reduced the induction of IFN-I and ISGs, associated with elevated HIV-1 replication. Surprisingly, HIV-1 induced depletion of human immune cells including T cells in lymphoid organs, but not the blood, was reduced in spite of the increased viral replication. The increased cell number in lymphoid organs was associated with a reduced level of HIV-induced cell death in human leukocytes including CD4 T cells. We conclude that pDC play opposing roles in suppressing HIV-1 replication and in promoting HIV-1 induced immunopathogenesis. These findings suggest that pDC-depletion and IFN-I blockade will provide novel strategies for treating those HIV-1 immune non-responsive patients with persistent immune activation despite effective anti-retrovirus treatment.

  2. Increased Risk of HIV-1 Transmission in Pregnancy: A Prospective Study among African HIV-1 Serodiscordant Couples

    Science.gov (United States)

    MUGO, Nelly R.; HEFFRON, Renee; DONNELL, Deborah; WALD, Anna; WERE, Edwin O.; REES, Helen; CELUM, Connie; KIARIE, James N.; COHEN, Craig R.; KAYINTEKORE, Kayitesi; BAETEN, Jared M.

    2011-01-01

    Background Physiologic and behavioral changes during pregnancy may alter HIV-1 susceptibility and infectiousness. Prospective studies exploring pregnancy and HIV-1 acquisition risk in women have found inconsistent results. No study has explored the effect of pregnancy on HIV-1 transmission risk from HIV-1 infected women to male partners. Methods In a prospective study of African HIV-1 serodiscordant couples, we evaluated the relationship between pregnancy and the risk of 1) HIV-1 acquisition among women and 2) HIV-1 transmission from women to men. Results 3321 HIV-1 serodiscordant couples were enrolled, 1085 (32.7%) with HIV-1 susceptible female partners and 2236 (67.3%) with susceptible male partners. HIV-1 incidence in women was 7.35 versus 3.01 per 100 person-years during pregnant and non-pregnant periods (hazard ratio [HR] 2.34, 95% confidence interval [CI] 1.33–4.09). This effect was attenuated and not statistically significant after adjusting for sexual behavior and other confounding factors (adjusted HR 1.71, 95% CI 0.93–3.12). HIV-1 incidence in male partners of infected women was 3.46 versus 1.58 per 100 person-years when their partners were pregnant versus not pregnant (HR 2.31, 95% CI 1.22–4.39). This effect was not attenuated in adjusted analysis (adjusted HR 2.47, 95% CI 1.26–4.85). Conclusions HIV-1 risk increased two-fold during pregnancy. Elevated risk of HIV-1 acquisition in pregnant women appeared in part to be explained by behavioral and other factors. This is the first study to show pregnancy increased the risk of female-to-male HIV-1 transmission, which may reflect biological changes of pregnancy that could increase HIV-1 infectiousness. PMID:21785321

  3. Role of Endolysosomes in HIV-1 Tat-Induced Neurotoxicity

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    Liang Hui

    2012-05-01

    Full Text Available Combined anti-retroviral therapeutic drugs effectively increase the lifespan of HIV-1-infected individuals who then have a higher prevalence of HAND (HIV-1 associated neurocognitive disorder. Soluble factors including HIV-1 proteins released from HIV-1-infected cells have been implicated in the pathogenesis of HAND, and particular attention has been paid to the HIV-1 Tat (transactivator of transcription protein because of its ability to directly excite neurons and cause neuronal cell death. Since HIV-1 Tat enters cells by receptor-mediated endocytosis and since endolysosomes play an important role in neuronal cell life and death, we tested here the hypothesis that HIV-1 Tat neurotoxicity is associated with changes in the endolysosome structure and function and also autophagy. Following the treatment of primary cultured rat hippocampal neurons with HIV-1 Tat or as controls mutant-Tat or PBS, neuronal viability was determined using a triple staining method. Preceding observations of HIV-1 Tat-induced neuronal cell death, we observed statistically significant changes in the structure and membrane integrity of endolysosomes, endolysosome pH and autophagy. As early as 24 h after HIV-1 Tat was applied to neurons, HIV-1 Tat accumulated in endolysosomes, endolysosome morphology was affected and their size increased, endolysosome membrane integrity was disrupted, endolysosome pH increased, specific activities of endolysosome enzymes decreased and autophagy was inhibited, as indicated by the significant changes in three markers for autophagy. In contrast, statistically significant levels of HIV-1 Tat-induced neuronal cell death were observed only after 48 h of HIV-1 Tat treatment. Our findings suggest that endolysosomes are involved in HIV-1 Tat-induced neurotoxicity and may represent a target for therapeutic intervention against HAND.

  4. Human Cytosolic Extracts Stabilize the HIV-1 Core

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    Fricke, Thomas; Brandariz-Nuñez, Alberto; Wang, Xiaozhao; Smith, Amos B.

    2013-01-01

    The stability of the HIV-1 core in the cytoplasm is crucial for productive HIV-1 infection. Mutations that stabilize or destabilize the core showed defects on HIV-1 reverse transcription and infection. We developed a novel and simple assay to measure the stability of in vitro-assembled HIV-1 CA-NC complexes. The assay allowed us to demonstrate that cytosolic extracts strongly stabilize the HIV-1 core. Interestingly, stabilization of in vitro-assembled HIV-1 CA-NC complexes is not due solely to macromolecular crowding, suggesting the presence of specific cellular factors that stabilize the HIV-1 core. By using our novel assay, we measured the abilities of different drugs, such as PF74, CAP-1, IXN-053, cyclosporine, Bi2 (also known as BI-2), and the peptide CAI, to modulate the stability of in vitro-assembled HIV-1 CA-NC complexes. Interestingly, we found that PF74 and Bi2 strongly stabilized HIV-1 CA-NC complexes. On the other hand, the peptide CAI destabilized HIV-1 CA-NC complexes. We also found that purified cyclophilin A destabilizes in vitro-assembled HIV-1 CA-NC complexes in the presence of cellular extracts in a cyclosporine-sensitive manner. In agreement with previous observations using the fate-of-the-capsid assay, we also demonstrated the ability of recombinant CPSF6 to stabilize HIV-1 CA-NC complexes. Overall, our findings suggested that cellular extracts specifically stabilize the HIV-1 core. We believe that our assay can be a powerful tool to assess HIV-1 core stability in vitro. PMID:23885082

  5. Inhibitory effects of Sudanese plant extracts on HIV-1 replication and HIV-1 protease.

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    Hussein, G; Miyashiro, H; Nakamura, N; Hattori, M; Kawahata, T; Otake, T; Kakiuchi, N; Shimotohno, K

    1999-02-01

    Forty-eight methanol and aqueous extracts from Sudanese plants were screened for their inhibitory activity on viral replication. Nineteen extracts showed inhibitory effects on HIV-induced cytopathic effects (CPE) on MT-4 cells. The extracts were further screened against HIV-1 protease (PR) using an HPLC assay method. Of the tested extracts, the methanol extracts of Acacia nilotica (bark and pods), Euphorbia granulata (leaves), Maytenus senegalensis (stem-bark) and aqueous extracts of A. nilotica (pods) and M. senegalensis (stem-bark) showed considerable inhibitory effects against HIV-1 PR. Inhibitory principles were isolated from M. senegalensis and their activities were also discussed.

  6. HIV-1 genetic variants in Kyrgyzstan

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    V Laga

    2012-11-01

    Full Text Available Objectives: During the last two decades, HIV-1 has been spreading rapidly in former Soviet Union republics including Kyrgyzstan. The current molecular monitoring of HIV-infection epidemic is carried out in Russia only with no or limited data from the other FSU countries. The aim of this work was to investigate the prevalence of HIV-1 genetic variants circulating in Kyrgyzstan. Methods: Blood collection from the HIV-infected patients was carried out by local specialists with the informed consent and the questionnaire was answered by each of the patients. The total number of samples was 100. The washed cell pellets were transferred to Moscow following with proviral DNA extraction, PCR amplification and gag, pol and env genes sequencing. The phylogenetic analysis of nucleotide sequences using neighbor-joining method was carried out by MEGA 3 program. The preliminary data were obtained in 22 samples isolated from PBMC of HIV-infected patients from Kyrgyzstan. Results: Among the samples studied 6 (27.3% samples belonged to a subtype CRF02_AG, 16 samples - to subtype A (A1. One of the samples belonging to CRF02_AG, probably, is a recombinant between CRF02_AG and A1. There was no major drug resistance mutations in the samples studied. The minor mutations were presented in small proportions: 1 in PR (L10I, 6 in RT (A62V - in 3 samples, V108G, E138A, Y181F, M184I, L210M - on one sample and 1 in IN (L74M. It was impossible to associate the distribution of mutations with HIV-1 genetic variant. The V3 loop (env gene in 17 samples was analyzed for tropism using geno2pheno program; all samples were found to be R5-viruses. Conclusion: The HIV-1 subtype A seems to dominate in Kyrgyzstan like in other FSU countries. The recombinant CRF02_AG epidemiologically linked to Uzbekistan is quite widespread. The rest of Kyrgyzstan collection is under investigation and the data will be refined soon.

  7. Rigidity analysis of HIV-1 protease

    Science.gov (United States)

    Heal, J. W.; Wells, S. A.; Jimenez-Roldan, E.; Freedman, R. F.; Römer, R. A.

    2011-03-01

    We present a rigidity analysis on a large number of X-ray crystal structures of the enzyme HIV-1 protease using the 'pebble game' algorithm of the software FIRST. We find that although the rigidity profile remains similar across a comprehensive set of high resolution structures, the profile changes significantly in the presence of an inhibitor. Our study shows that the action of the inhibitors is to restrict the flexibility of the β-hairpin flaps which allow access to the active site. The results are discussed in the context of full molecular dynamics simulations as well as data from NMR experiments.

  8. HIV-1 is not a major driver of increased plasma IL-6 levels in chronic HIV-1 disease

    Science.gov (United States)

    Shive, Carey L.; Biancotto, Angélique; Funderburg, Nicholas T.; Pilch-Cooper, Heather A.; Valdez, Hernan; Margolis, Leonid; Sieg, Scott F.; McComsey, Grace A.; Rodriguez, Benigno; Lederman, Michael M.

    2012-01-01

    Objective Increased plasma IL-6 levels have been associated with HIV-1 disease progression risk, yet the drivers of IL-6 production in HIV-1 infection are not known. This study was designed to explore the relationship between HIV-1 replication and IL-6 induction. Design Correlations between plasma levels of IL-6 and HIV-1 RNA were examined in two clinical studies. To more directly assess the induction of IL-6 by HIV-1, several cell and tissue types that support HIV-1 replication in vivo were infected with HIV-1 and expression of IL-6 was measured. Methods Spearman’s rank correlations were used to examine the relationship between plasma levels of IL-6 and HIV-1 RNA. Macrophages, and colonic and lymph node histocultures were infected with HIV-1 or stimulated with bacterial products, LPS or flagellin, and IL-6 levels in supernatant were measured by ELISA or multiplex bead assay. Results In the clinical studies there was weak or no correlation between plasma levels of IL-6 and HIV-1 RNA but IL-6 levels were correlated with plasma levels of the LPS coreceptor CD14. Macrophages stimulated with LPS or flagellin showed robust production of IL-6, but there was no increase in IL-6 production after HIV-1 infection. IL-6 expression was not increased in lymph node histocultures obtained from HIV-1 infected subjects nor after productive HIV-1 infection of colonic or lymph node histocultures ex vivo. Conclusions We find no evidence that HIV-1 replication is an important driver of IL-6 expression in vivo or in in vitro systems. PMID:22659649

  9. Cyclophilin B enhances HIV-1 infection

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    DeBoer, Jason; Madson, Christian J. [Department of Medical Microbiology and Immunology, Creighton University, Omaha, NE (United States); Belshan, Michael, E-mail: michaelbelshan@creighton.edu [Department of Medical Microbiology and Immunology, Creighton University, Omaha, NE (United States); The Nebraska Center for Virology, University of Nebraska, Lincoln, NE (United States)

    2016-02-15

    Cyclophilin B (CypB) is a member of the immunophilin family and intracellular chaperone. It predominantly localizes to the ER, but also contains a nuclear localization signal and is secreted from cells. CypB has been shown to interact with the Gag protein of human immunodeficiency type 1 (HIV-1). Several proteomic and genetic studies identified it as a potential factor involved in HIV replication. Herein, we show that over-expression of CypB enhances HIV infection by increasing nuclear import of viral DNA. This enhancement was unaffected by cyclosporine treatment and requires the N-terminus of the protein. The N-terminus contains an ER leader sequence, putative nuclear localization signal, and is required for secretion. Deletion of the N-terminus resulted in mislocalization from the ER and suppression of HIV infection. Passive transfer experiments showed that secreted CypB did not impact HIV infection. Combined, these experiments show that intracellular CypB modulates a pathway of HIV nuclear import. - Highlights: • CypB has been identified in several proteomic studies of HIV-1 infection. • CypB expression is upregulated in activated and infected T-cells. • Over-expression of CypB enhances HIV nuclear import and infection. • The N-terminus of CypB is necessary for these effects.

  10. Evaluation of three enzyme immunoassays for HIV-1 antigen detection.

    Science.gov (United States)

    Willoughby, P B; Lisker, A; Folds, J D

    1989-01-01

    Three enzyme immunoassay (EIA) methods for the detection of human immunodeficiency virus (HIV-1) were evaluated. Serum or plasma samples from 22 individuals seropositive for HIV-1 antibodies were tested with the Abbott, Coulter, and DuPont kits for presence of HIV-1 p24 antigen. Another 12 samples were tested with two kits only. Discordant results were obtained with 9 of 34 (26%) HIV-1-antibody-positive patient samples tested. Most of these discrepancies were found in samples containing less than 30 pg/ml of HIV-1 p24 core antigen. A sampling of sera from normal blood donors and patients with infectious or autoimmune diseases revealed a low level of false positive reactions, especially with sera containing antinuclear antibodies or rheumatoid factor. Noteworthy is the frequency of false positive reactions seen with the DuPont EIA for HIV-1 p24 antigen. 18/111 sera (16.2%) containing auto-antibodies tested positively with the DuPont HIV-1 p24 antigen EIA. The nonspecific nature of the test reactivity for 9/10 of these samples was confirmed using an HIV-1 p24 antigen inhibition assay. These findings are discussed in light of the need for HIV-1 antigen detection in the clinical laboratory and of other methods for HIV-1 detection: the polymerase chain reaction and measurements of reverse transcriptase activity.

  11. Genetic composition of replication competent clonal HIV-1 variants isolated from peripheral blood mononuclear cells (PBMC), HIV-1 proviral DNA from PBMC and HIV-1 RNA in serum in the course of HIV-1 infection.

    Science.gov (United States)

    Edo-Matas, Diana; van Gils, Marit J; Bowles, Emma J; Navis, Marjon; Rachinger, Andrea; Boeser-Nunnink, Brigitte; Stewart-Jones, Guillaume B; Kootstra, Neeltje A; van 't Wout, Angélique B; Schuitemaker, Hanneke

    2010-09-30

    The HIV-1 quasispecies in peripheral blood mononuclear cells (PBMC) is considered to be a mix of actively replicating, latent, and archived viruses and may be genetically distinct from HIV-1 variants in plasma that are considered to be recently produced. Here we analyzed the genetic relationship between gp160 env sequences from replication competent clonal HIV-1 variants that were isolated from PBMC and from contemporaneous HIV-1 RNA in serum and HIV-1 proviral DNA in PBMC of four longitudinally studied therapy naïve HIV-1 infected individuals. Replication competent clonal HIV-1 variants, HIV-1 RNA from serum, and HIV-1 proviral DNA from PBMC formed a single virus population at most time points analyzed. However, an under-representation in serum of HIV-1 sequences with predicted CXCR4 usage was sometimes observed implying that the analysis of viral sequences from different sources may provide a more complete assessment of the viral quasispecies in peripheral blood in vivo. Copyright 2010 Elsevier Inc. All rights reserved.

  12. Nup153 and Nup98 bind the HIV-1 core and contribute to the early steps of HIV-1 replication.

    Science.gov (United States)

    Di Nunzio, Francesca; Fricke, Thomas; Miccio, Annarita; Valle-Casuso, Jose Carlos; Perez, Patricio; Souque, Philippe; Rizzi, Ermanno; Severgnini, Marco; Mavilio, Fulvio; Charneau, Pierre; Diaz-Griffero, Felipe

    2013-05-25

    The early steps of HIV-1 replication involve the entry of HIV-1 into the nucleus, which is characterized by viral interactions with nuclear pore components. HIV-1 developed an evolutionary strategy to usurp the nuclear pore machinery and chromatin in order to integrate and efficiently express viral genes. In the current work, we studied the role of nucleoporins 153 and 98 (Nup153 and Nup98) in infection of human Jurkat lymphocytes by HIV-1. We showed that Nup153-depleted cells exhibited a defect in nuclear import, while depletion of Nup 98 caused a slight defect in HIV integration. To explore the biochemical viral determinants for the requirement of Nup153 and Nup98 during HIV-1 infection, we tested the ability of these nucleoporins to interact with HIV-1 cores. Our findings showed that both nucleoporins bind HIV-1 cores suggesting that this interaction is important for HIV-1 nuclear import and/or integration. Distribution analysis of integration sites in Nup153-depleted cells revealed a reduced tendency of HIV-1 to integrate in intragenic sites, which in part could account for the large infectivity defect observed in Nup153-depleted cells. Our work strongly supports a role for Nup153 in HIV-1 nuclear import and integration. Copyright © 2013 Elsevier Inc. All rights reserved.

  13. Synergistic Inhibition of R5 HIV-1 by the Fusion Protein (FLSC) IgG1 Fc and Maraviroc in Primary Cells: Implications for Prevention and Treatment.

    Science.gov (United States)

    Latinovic, Olga S; Zhang, Jian; Tagaya, Yutaka; DeVico, Anthony L; Fouts, Timothy R; Schneider, K; Lakowicz, Joseph R; Heredia, Alonso; Redfield, Robert R

    2016-01-01

    Antiretroviral (ARV) drugs targeting retroviral enzymes have been extensively employed to treat HIV-1 infection. Drawbacks of this approach include cost, toxicity, and the eventual emergence of resistant strains that threaten prophylactic and/or therapeutic efficacy. Accordingly, efforts to develop next-generation ARV approaches are warranted, particularly if they can offer a higher threshold of resistance. We have previously shown that FLSC, a fusion protein containing gp120(BAL) and the D1 and D2 domains of human CD4, specifically binds CCR5, an important cellular co-receptor, and inhibits the entry of R5 HIV isolates. (FLSC) IgG1, a fusion of FLSC and the hinge-C(H)2-C(H)3 region of human IgG1, has an increased antiviral activity, likely due to the resultant bivalency. In this study, we show CCR5 reduction upon (FLSC) IgG1 treatment both by standard flow cytometry and visualized using a novel nanoparticle method. A β-lactamase virus-cell fusion assay was used to quantify (FLSC) IgG1 inhibition of HIV-1 entry into both cell lines and primary cells. Synergistic anti-viral activities of (FLSC) IgG1 and MVC in primary cells were evaluated by measuring supernatant p24 levels via ELISA and calculated using the MacSynergy™ II program. We previously reported that treatment with the CCR5 small molecule antagonist Maraviroc (MVC) increased the apparent exposure of the (FLSC) IgG1 binding sites on CCR5, leading us to wonder if the two compounds used in combination might synergize in their anti-viral activity. Here we show that this is indeed the case. We demonstrate that fusion protein (FLSC) IgG1, strongly synergizes with the CCR5 antagonist Maraviroc to successfully inhibit both MVC-sensitive and MVC-resistant R5 HIV-1. Observed synergy between (FLSC) IgG1 and MVC was high in both, cell lines and primary PBMCs. This has relevance for future in vivo studies. In addition, synergy occurred both with MVC-sensitive viruses and MVC-resistant viruses, partially restoring the

  14. CoRSeqV3-C: a novel HIV-1 subtype C specific V3 sequence based coreceptor usage prediction algorithm.

    Science.gov (United States)

    Cashin, Kieran; Gray, Lachlan R; Jakobsen, Martin R; Sterjovski, Jasminka; Churchill, Melissa J; Gorry, Paul R

    2013-02-27

    The majority of HIV-1 subjects worldwide are infected with HIV-1 subtype C (C-HIV). Although C-HIV predominates in developing regions of the world such as Southern Africa and Central Asia, C-HIV is also spreading rapidly in countries with more developed economies and health care systems, whose populations are more likely to have access to wider treatment options, including the CCR5 antagonist maraviroc (MVC). The ability to reliably determine C-HIV coreceptor usage is therefore becoming increasingly more important. In silico V3 sequence based coreceptor usage prediction algorithms are a relatively rapid and cost effective method for determining HIV-1 coreceptor specificity. In this study, we elucidated the V3 sequence determinants of C-HIV coreceptor usage, and used this knowledge to develop and validate a novel, user friendly, and highly sensitive C-HIV specific coreceptor usage prediction algorithm. We characterized every phenotypically-verified C-HIV gp120 V3 sequence available in the Los Alamos HIV Database. Sequence analyses revealed that compared to R5 C-HIV V3 sequences, CXCR4-using C-HIV V3 sequences have significantly greater amino acid variability, increased net charge, increased amino acid length, increased frequency of insertions and substitutions within the GPGQ crown motif, and reduced frequency of glycosylation sites. Based on these findings, we developed a novel C-HIV specific coreceptor usage prediction algorithm (CoRSeqV3-C), which we show has superior sensitivity for determining CXCR4 usage by C-HIV strains compared to all other available algorithms and prediction rules, including Geno2pheno[coreceptor] and WebPSSMSINSI-C, which has been designed specifically for C-HIV. CoRSeqV3-C is now openly available for public use at http://www.burnet.edu.au/coreceptor. Our results show that CoRSeqV3-C is the most sensitive V3 sequence based algorithm presently available for predicting CXCR4 usage of C-HIV strains, without compromising specificity. CoRSeqV3

  15. Species tropism of HIV-1 modulated by viral accessory proteins

    OpenAIRE

    Masako eNomaguchi; Naoya eDoi; Yui eMatsumoto; Yosuke eSakai; Sachi eFujiwara; Akio eAdachi

    2012-01-01

    Human immunodeficiency virus type 1 (HIV-1) is tropic and pathogenic only for humans, and does not replicate in macaque monkeys routinely used for experimental infections. This specially narrow host range (species tropism) has impeded much the progress of HIV-1/acquired immunodeficiency syndrome (AIDS) basic research. Extensive studies on the underlying mechanism have revealed that Vif, one of viral accessory proteins, is critical for the HIV-1 species tropism in addition to Gag-capsid protei...

  16. Reverse transcription of the HIV-1 pandemic.

    Science.gov (United States)

    Basavapathruni, Aravind; Anderson, Karen S

    2007-12-01

    The HIV/AIDS pandemic has existed for >25 years. Extensive work globally has provided avenues to combat viral infection, but the disease continues to rage on in the human population and infected approximately 4 million people in 2006 alone. In this review, we provide a brief history of HIV/AIDS, followed by analysis of one therapeutic target of HIV-1: its reverse transcriptase (RT). We discuss the biochemical characterization of RT in order to place emphasis on possible avenues of inhibition, which now includes both nucleoside and non-nucleoside modalities. Therapies against RT remain a cornerstone of anti-HIV treatment, but the virus eventually resists inhibition through the selection of drug-resistant RT mutations. Current inhibitors and associated resistance are discussed, with the hopes that new therapeutics can be developed against RT.

  17. TIM-family proteins inhibit HIV-1 release.

    Science.gov (United States)

    Li, Minghua; Ablan, Sherimay D; Miao, Chunhui; Zheng, Yi-Min; Fuller, Matthew S; Rennert, Paul D; Maury, Wendy; Johnson, Marc C; Freed, Eric O; Liu, Shan-Lu

    2014-09-02

    Accumulating evidence indicates that T-cell immunoglobulin (Ig) and mucin domain (TIM) proteins play critical roles in viral infections. Herein, we report that the TIM-family proteins strongly inhibit HIV-1 release, resulting in diminished viral production and replication. Expression of TIM-1 causes HIV-1 Gag and mature viral particles to accumulate on the plasma membrane. Mutation of the phosphatidylserine (PS) binding sites of TIM-1 abolishes its ability to block HIV-1 release. TIM-1, but to a much lesser extent PS-binding deficient mutants, induces PS flipping onto the cell surface; TIM-1 is also found to be incorporated into HIV-1 virions. Importantly, TIM-1 inhibits HIV-1 replication in CD4-positive Jurkat cells, despite its capability of up-regulating CD4 and promoting HIV-1 entry. In addition to TIM-1, TIM-3 and TIM-4 also block the release of HIV-1, as well as that of murine leukemia virus (MLV) and Ebola virus (EBOV); knockdown of TIM-3 in differentiated monocyte-derived macrophages (MDMs) enhances HIV-1 production. The inhibitory effects of TIM-family proteins on virus release are extended to other PS receptors, such as Axl and RAGE. Overall, our study uncovers a novel ability of TIM-family proteins to block the release of HIV-1 and other viruses by interaction with virion- and cell-associated PS. Our work provides new insights into a virus-cell interaction that is mediated by TIMs and PS receptors.

  18. RNA Interference Therapies for an HIV-1 Functional Cure.

    Science.gov (United States)

    Scarborough, Robert J; Gatignol, Anne

    2017-12-27

    HIV-1 drug therapies can prevent disease progression but cannot eliminate HIV-1 viruses from an infected individual. While there is hope that elimination of HIV-1 can be achieved, several approaches to reach a functional cure (control of HIV-1 replication in the absence of drug therapy) are also under investigation. One of these approaches is the transplant of HIV-1 resistant cells expressing anti-HIV-1 RNAs, proteins or peptides. Small RNAs that use RNA interference pathways to target HIV-1 replication have emerged as competitive candidates for cell transplant therapy and have been included in all gene combinations that have so far entered clinical trials. Here, we review RNA interference pathways in mammalian cells and the design of therapeutic small RNAs that use these pathways to target pathogenic RNA sequences. Studies that have been performed to identify anti-HIV-1 RNA interference therapeutics are also reviewed and perspectives on their use in combination gene therapy to functionally cure HIV-1 infection are provided.

  19. Chronic HIV-1 infection frequently fails to protect against superinfection.

    Directory of Open Access Journals (Sweden)

    Anne Piantadosi

    2007-11-01

    Full Text Available Reports of HIV-1 superinfection (re-infection have demonstrated that the immune response generated against one strain of HIV-1 does not always protect against other strains. However, studies to determine the incidence of HIV-1 superinfection have yielded conflicting results. Furthermore, few studies have attempted to identify superinfection cases occurring more than a year after initial infection, a time when HIV-1-specific immune responses would be most likely to have developed. We screened a cohort of high-risk Kenyan women for HIV-1 superinfection by comparing partial gag and envelope sequences over a 5-y period beginning at primary infection. Among 36 individuals, we detected seven cases of superinfection, including cases in which both viruses belonged to the same HIV-1 subtype, subtype A. In five of these cases, the superinfecting strain was detected in only one of the two genome regions examined, suggesting that recombination frequently occurs following HIV-1 superinfection. In addition, we found that superinfection occurred throughout the course of the first infection: during acute infection in two cases, between 1-2 y after infection in three cases, and as late as 5 y after infection in two cases. Our results indicate that superinfection commonly occurs after the immune response against the initial infection has had time to develop and mature. Implications from HIV-1 superinfection cases, in which natural re-exposure leads to re-infection, will need to be considered in developing strategies for eliciting protective immunity to HIV-1.

  20. Progress in HIV-1 antibody research using humanized mice.

    Science.gov (United States)

    Gruell, Henning; Klein, Florian

    2017-05-01

    Recent discoveries of highly potent broadly HIV-1 neutralizing antibodies provide new opportunities to successfully prevent, treat, and potentially cure HIV-1 infection. To test their activity in vivo, humanized mice have been shown to be a powerful model and were used to investigate antibody-mediated prevention and therapy approaches. In this review, we will summarize recent findings in humanized mice that have informed on the potential use of broadly neutralizing antibodies targeting HIV-1 in humans. Humanized mouse models have been used to demonstrate the antiviral efficacy of HIV-1 neutralizing antibodies in vivo. It has been shown that a combination of antibodies can suppress viremia below the limit of detection and targets the HIV-1 reservoir. Moreover, passively administered antibodies and vector-mediated antibody production protect humanized mice from HIV-1 infection. Finally, immunization studies in knock-in/transgenic mice carrying human antibody gene segments have informed on potential vaccination strategies to induce broad and potent HIV-1 neutralizing antibodies. Humanized mouse models are of great value for HIV-1 research. They represent a highly versatile in vivo system to investigate novel approaches for HIV-1 prevention and therapy and expedite the critical translation from basic findings to clinical application.

  1. Innate immune sensing of HIV-1 infection.

    Science.gov (United States)

    Jakobsen, Martin R; Olagnier, David; Hiscott, John

    2015-03-01

    The innate immune system plays a critical role in the control of viral infections. Although the mechanisms involved in sensing and response to viral pathogens has progressed tremendously in the last decade, an understanding of the innate antiviral response to human retroviruses lagged behind. Recent studies now demonstrate that human retroviruses such as human immunodeficiency virus type 1 (HIV-1) and human T-lymphotropic virus 1 (HTLV-1) trigger a type I interferon antiviral response through novel cytosolic sensors that detect DNA intermediates of reverse transcription; in addition, these early host-pathogen interactions may trigger cell death pathways depending on the activation state of the target cell. The purpose of this review is to summarize the recent progress in the understanding of innate immune sensing of human retroviruses. Innate immune sensing of HIV-1 and HTLV-1 is influenced by the target cell phenotype, viral replicative intermediates, and host restriction factors that limit retroviral replication. Macrophages and dendritic cells detect HIV-DNA intermediates, whereas CD4 T cells differentially sense HIV DNA depending on the level of T-cell activation. Furthermore, the structure of the viral capsid and interplay between innate DNA sensors and host restriction factors all contribute to the magnitude of the ensuing innate immune response. The interplay between HIV infection and the innate immune system has emerged as an important component of HIV pathogenesis, linked to both induction of innate immunity and stimulation of cell death mechanisms. Ultimately, an in-depth knowledge of the mechanisms of innate immune control of human retrovirus infection may facilitate the development of novel treatment strategies to control retrovirus-induced immunopathology.

  2. Characteristics of HIV-1 Serodiscordant Couples Enrolled in a Clinical Trial of Antiretroviral Pre-Exposure Prophylaxis for HIV-1 Prevention

    OpenAIRE

    Andrew Mujugira; Baeten, Jared M.; Deborah Donnell; Patrick Ndase; Mugo, Nelly R.; Linda Barnes; Campbell, James D.; Jonathan Wangisi; Tappero, Jordan W.; Elizabeth Bukusi; Cohen, Craig R.; Elly Katabira; Allan Ronald; Elioda Tumwesigye; Edwin Were

    2011-01-01

    Introduction Stable heterosexual HIV-1 serodiscordant couples in Africa have high HIV-1 transmission rates and are a critical population for evaluation of new HIV-1 prevention strategies. The Partners PrEP Study is a randomized, double-blind, placebo-controlled trial of tenofovir and emtricitabine-tenofovir pre-exposure prophylaxis to decrease HIV-1 acquisition within heterosexual HIV-1 serodiscordant couples. We describe the trial design and characteristics of the study cohort. Methods HIV-1...

  3. HIV-1 infection of in vitro cultured human monocytes: early events and influence of anti HIV-1 antibodies

    DEFF Research Database (Denmark)

    Arendrup, M; Olofsson, S; Nielsen, Jens Ole

    1994-01-01

    To characterize the role of the humoral immune response on HIV-1 infection of monocytes and macrophages (M phi s) we examined the susceptibility of in vitro cultured monocyte/M phi s to various HIV-1 isolates and the influence of heterologous and particularly autologous anti HIV-1 sera...... on this infection. Depending on the period of in vitro cultivation and the virus isolate used different patterns of susceptibility were detected. One week old monocyte/M phi s were highly susceptible to HIV-1 infection, in contrast to monocyte/M phi s cultured 4 weeks. The infection by virus isolated immediately...

  4. Nup153 and Nup98 bind the HIV-1 core and contribute to the early steps of HIV-1 replication

    OpenAIRE

    Di Nunzio, Francesca; Fricke, Thomas; Miccio, Annarita; Valle-Casuso, Jose Carlos; Perez, Patricio; Souque, Philippe; Rizzi, Ermanno; Severgnini, Marco; Mavilio, Fulvio; Charneau, Pierre; Diaz-Griffero, Felipe

    2013-01-01

    The early steps of HIV-1 replication involve the entry of HIV-1 into the nucleus, which is characterized by viral interactions with nuclear pore components. HIV-1 developed an evolutionary strategy to usurp the nuclear pore machinery and chromatin in order to integrate and efficiently express viral genes. In the current work, we studied the role of nucleoporins 153 and 98 (Nup153 and Nup98) in infection of human Jurkat lymphocytes by HIV-1. We showed that Nup153-depleted cells exhibited a def...

  5. rOv-ASP-1, a recombinant secreted protein of the helminth Onchocercavolvulus, is a potent adjuvant for inducing antibodies to ovalbumin, HIV-1 polypeptide and SARS-CoV peptide antigens.

    Science.gov (United States)

    MacDonald, Angus J; Cao, Long; He, Yuxian; Zhao, Qian; Jiang, Shibo; Lustigman, Sara

    2005-05-16

    We studied the adjuvanticity of recombinant Onchocerca volvulus activation associated protein-1 (rOv-ASP-1) for ovalbumin (OVA) in mice. After a single immunization and one boost, rOv-ASP-1 exceeded the efficacy of alum or MPL + TDM adjuvants in terms of end-point total IgG or IgG1 and IgG2a anti-OVA titres. Using the helminth-derived adjuvant, IgG isotype responses to OVA were of a mixed Th1/Th2 profile and spleen cell cytokines exclusively Th1-type. The potent adjuvanticity of rOv-ASP-1 was confirmed in mice vaccinated with a 37-mer peptide from the S protein of SARS-CoV and an HIV-1 gp120-CD4 chimeric polypeptide antigen. Unusually for a helminth product, the rOv-ASP-1 adjuvant augmented not only Th2 but also Th1 responses, the latter property being of potential utility in stimulating anti-viral immune responses.

  6. Mimetics of beta-galactosylceramide with simple ceramide substitutes: Synthesis and binding togp 120 of HIV-1, and, Enactment of chemistry knowledge by a high school student at a summer program

    Science.gov (United States)

    Augustin, Line A.

    This thesis is the account of two research works. The first part reports the synthesis of O- and C- and aza-C-glycosides of beta-Galactosylceramide (GalCer) that contain simple ceramide substitutes, and the initial results of their binding with gp120 of HIV-1. The O-glycosides were prepared via an established procedure. The C- and aza-C-glycosides originated from a central C1-substituted galactal precursor, and their synthesis is illustrative of a potentially general method for pairs of C- and aza-C-beta-galactosides. They aza-C-glycoside with a simple C-17 hydrocarbon chain exhibited significant higher affinity than GalCer, whereas the corresponding C-glycoside was as active as GalCer. The second part describes the ethnographic study of the enactment of the chemistry knowledge of a high school student at a summer program and the influence of a cultural practice, othermothering, on her ability to perform well on her chemistry Regents Exams. Kelly, an 11th grade student exhibited very good understanding of the chemistry curriculum in the classroom, the laboratory period and the tutoring sessions where she plays a caring role for her peers. The same level of understanding was not reflected on the paper pencil exams taken during the summer program.

  7. Antiviral Therapy by HIV-1 Broadly Neutralizing and Inhibitory Antibodies

    Directory of Open Access Journals (Sweden)

    Zhiqing Zhang

    2016-11-01

    Full Text Available Human immunodeficiency virus type 1 (HIV-1 infection causes acquired immune deficiency syndrome (AIDS, a global epidemic for more than three decades. HIV-1 replication is primarily controlled through antiretroviral therapy (ART but this treatment does not cure HIV-1 infection. Furthermore, there is increasing viral resistance to ART, and side effects associated with long-term therapy. Consequently, there is a need of alternative candidates for HIV-1 prevention and therapy. Recent advances have discovered multiple broadly neutralizing antibodies against HIV-1. In this review, we describe the key epitopes on the HIV-1 Env protein and the reciprocal broadly neutralizing antibodies, and discuss the ongoing clinical trials of broadly neutralizing and inhibitory antibody therapy as well as antibody combinations, bispecific antibodies, and methods that improve therapeutic efficacy by combining broadly neutralizing antibodies (bNAbs with latency reversing agents. Compared with ART, HIV-1 therapeutics that incorporate these broadly neutralizing and inhibitory antibodies offer the advantage of decreasing virus load and clearing infected cells, which is a promising prospect in HIV-1 prevention and treatment.

  8. HIV-1, how llamas help us fight the AIDS pandemic

    NARCIS (Netherlands)

    Strokappe, N.M.|info:eu-repo/dai/nl/314411534

    2013-01-01

    Human Immunodeficiency Virus type 1 (HIV-1) is one of the major health problems worldwide and has been for over thirty years. Most (67%) of the people infected with HIV-1 are living in sub-Saharan Africa. Here, the access to treatments is limited and most women are not in a position to protect

  9. Vaginalmycosis and HIV-1 infection in Kaduna, Nigeria. | Eni ...

    African Journals Online (AJOL)

    Vaginal mycosis and HIV-1 infection are common health problems in females. These infections cause high mortality, morbidity and reproductive health disorders in females. The study is to investigate to what extent these infections are prevalent in this centre. 300 non- pregnant females who tested positive with HIV-1 ...

  10. The origin and emergence of an HIV-1 epidemic:

    DEFF Research Database (Denmark)

    Bruhn, Christian Anders Wathne; Audelin, Anne M.; Helleberg, Marie

    2014-01-01

    To describe, at patient-level detail, the determining events and factors involved in the development of a country's HIV-1 epidemic.......To describe, at patient-level detail, the determining events and factors involved in the development of a country's HIV-1 epidemic....

  11. Telomeres and HIV-1 infection: in search of exhaustion

    NARCIS (Netherlands)

    Wolthers, K. C.; Miedema, F.

    1998-01-01

    Telomere length analysis could be helpful in determining if exhaustion and replicative senescence are involved in HIV-1 pathogenesis. Evidence that CD8+ T cells have shorter telomeres may point towards an increased turnover of CD8+ T cells and exhaustion of the CD8+ T-cell responses in HIV-1

  12. Schistosomiasis and HIV-1 infection in rural Zimbabwe

    DEFF Research Database (Denmark)

    Kallestrup, Per; Zinyama, Rutendo; Gomo, Exnevia

    2005-01-01

    Stunted development and reduced fecundity of Schistosoma parasites in immunodeficient mice and the impaired ability of human immunodeficiency virus 1 (HIV-1)-infected humans to excrete schistosome eggs have been described. This study explores the effect that HIV-1-associated immunodeficiency has ...

  13. Development of aptamer based HIV-1 entry inhibitor prophylactic drugs

    CSIR Research Space (South Africa)

    London, G

    2013-08-01

    Full Text Available AIDS remains a major public health problem globally, especially in Southern Africa where over 6.4 million people are infected by the most prevalent HIV-1 subtype C. To help stop the spread of HIV-1 subtype C, we isolated 2ʹ-F-RNA aptamers against gp...

  14. Dendritic Cell Immune Responses in HIV-1 Controllers.

    Science.gov (United States)

    Martin-Gayo, Enrique; Yu, Xu G

    2017-02-01

    Robust HIV-1-specific CD8 T cell responses are currently regarded as the main correlate of immune defense in rare individuals who achieve natural, drug-free control of HIV-1; however, the mechanisms that support evolution of such powerful immune responses are not well understood. Dendritic cells (DCs) are specialized innate immune cells critical for immune recognition, immune regulation, and immune induction, but their possible contribution to HIV-1 immune defense in controllers remains ill-defined. Recent studies suggest that myeloid DCs from controllers have improved abilities to recognize HIV-1 through cytoplasmic immune sensors, resulting in more potent, cell-intrinsic type I interferon secretion in response to viral infection. This innate immune response may facilitate DC-mediated induction of highly potent antiviral HIV-1-specific T cells. Moreover, protective HLA class I isotypes restricting HIV-1-specific CD8 T cells may influence DC function through specific interactions with innate myelomonocytic MHC class I receptors from the leukocyte immunoglobulin-like receptor family. Bi-directional interactions between dendritic cells and HIV-1-specific T cells may contribute to natural HIV-1 immune control, highlighting the importance of a fine-tuned interplay between innate and adaptive immune activities for effective antiviral immune defense.

  15. Raltegravir with optimized background therapy for resistant HIV-1 infection

    DEFF Research Database (Denmark)

    Steigbigel, Roy T; Cooper, David A; Kumar, Princy N

    2008-01-01

    BACKGROUND: Raltegravir (MK-0518) is an inhibitor of human immunodeficiency virus type 1 (HIV-1) integrase active against HIV-1 susceptible or resistant to older antiretroviral drugs. METHODS: We conducted two identical trials in different geographic regions to evaluate the safety and efficacy...

  16. Antibody function in neutralization and protection against HIV-1

    NARCIS (Netherlands)

    Hessell, A.J.

    2009-01-01

    The ability to induce neutralizing antibodies is generally thought to be of great importance for vaccine efficacy. In HIV-1 research this quality has been elusive as the HIV-1 virus has evolved multiple mechanisms to evade neutralizing antibodies. This thesis traces studies with four broadly

  17. HTLV-1 Tax activates HIV-1 transcription in latency models.

    Science.gov (United States)

    Geddes, Victor Emmanuel Viana; José, Diego Pandeló; Leal, Fabio E; Nixon, Douglas F; Tanuri, Amilcar; Aguiar, Renato Santana

    2017-04-01

    HIV-1 latency is a major obstacle to HIV-1 eradication. Coinfection with HTLV-1 has been associated with faster progression to AIDS. HTLV-1 encodes the transactivator Tax which can activate both HTLV-1 and HIV-1 transcription. Here, we demonstrate that Tax activates HIV transcription in latent CD4(+) T cells. Tax promotes the activation of P-TEFb, releasing CDK9 and Cyclin T1 from inactive forms, promoting transcription elongation and reactivation of latent HIV-1. Tax mutants lacking interaction with the HIV-1-LTR promoter were not able to activate P-TEFb, with no subsequent activation of latent HIV. In HIV-infected primary resting CD4(+) T cells, Tax-1 reactivated HIV-1 transcription up to five fold, confirming these findings in an ex vivo latency model. Finally, our results confirms that HTLV-1/Tax hijacks cellular partners, promoting HIV-1 transcription, and this interaction should be further investigated in HIV-1 latency studies in patients with HIV/HTLV-1 co-infection. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. Molecular Mechanisms in Activation of Latent HIV-1

    NARCIS (Netherlands)

    H. Rafati (Haleh)

    2014-01-01

    markdownabstract__Abstract__ Finding a cure for the human immunodeficiency virus type 1 (HIV-1) is extremely challenging. Development of highly active anti-retroviral therapy (HAART), transformed HIV-1 infection from an acute syndrome into chronic disease. Although using HAART results in

  19. Neutralizing antibodies in slowly progressing HIV-1 infection

    DEFF Research Database (Denmark)

    Schønning, Kristian; Nielsen, C; Iversen, Johan

    1995-01-01

    Ten asymptomatic individuals who had experienced only limited CD4+ cell loss after prolonged infection with HIV-1 were studied. These individuals had a mean CD4+ cell count of 674 x 10(6) cells/L and a mean duration of infection of 8.5 years. Also included were 10 asymptomatic HIV-1-infected...

  20. The role of polymorphonuclear neutrophils during HIV-1 infection.

    Science.gov (United States)

    Yaseen, Mahmoud Mohammad; Abuharfeil, Nizar Mohammad; Yaseen, Mohammad Mahmoud; Shabsoug, Barakat Mohammad

    2018-01-01

    It is well-recognized that human immunodeficiency virus type-1 (HIV-1) mainly targets CD4+ T cells and macrophages. Nonetheless, during the past three decades, a huge number of studies have reported that HIV-1 can directly or indirectly target other cellular components of the immune system including CD8+ T cells, B cells, dendritic cells, natural killer cells, and polymorphonuclear neutrophils (PMNs), among others. PMNs are the most abundant leukocytes in the human circulation, and are known to play principal roles in the elimination of invading pathogens, regulating different immune responses, healing of injured tissues, and maintaining mucosal homeostasis. Until recently, little was known about the impact of HIV-1 infection on PMNs as well as the impact of PMNs on HIV-1 disease progression. This is because early studies focused on neutropenia and recurrent microbial infections, particularly, during advanced disease. However, recent studies have extended the investigation area to cover new aspects of the interactions between HIV-1 and PMNs. This review aims to summarize these advances and address the impact of HIV-1 infection on PMNs as well as the impact of PMNs on HIV-1 disease progression to better understand the pathophysiology of HIV-1 infection.

  1. Probing the sequence space available for HIV-1 evolution

    NARCIS (Netherlands)

    ter Brake, Olivier; Von Eije, Karin J.; Berkhout, Ben

    2008-01-01

    We designed a novel experimental approach to probe the sequence space available for HIV-1 evolution. Selective pressure was put on conserved HIV-1 genomic sequences by means of RNA interference (RNAi). Virus escape was monitored in many parallel cultures, and we scored the mutations selected in the

  2. Daily Acyclovir Delays HIV-1 Disease Progression Among HIV-1/HSV-2 Dually-Infected Persons: A Randomized Trial

    Science.gov (United States)

    Lingappa, Jairam R.; Baeten, Jared M.; Wald, Anna; Hughes, James P.; Thomas, Katherine K.; Mujugira, Andrew; Mugo, Nelly; Bukusi, Elizabeth A.; Cohen, Craig R.; Katabira, Elly; Ronald, Allan; Kiarie, James; Farquhar, Carey; Stewart, Grace John; Makhema, Joseph; Essex, Myron; Were, Edwin; Fife, Kenneth H.; de Bruyn, Guy; Gray, Glenda E.; McIntyre, James; Manongi, Rachel; Kapiga, Saidi; Coetzee, David; Allen, Susan; Inambao, Mubiana; Kayitenkore, Kayitesi; Karita, Etienne; Kanweka, William; Delany, Sinead; Rees, Helen; Vwalika, Bellington; Magaret, Amalia; Wang, Richard S.; Kidoguchi, Lara; Barnes, Linda; Ridzon, Renee; Corey, Lawrence; Celum, Connie

    2010-01-01

    Background Well-tolerated medications that slow HIV-1 disease progression and delay initiation of antiretroviral therapy (ART) are needed. Most HIV-1-infected persons are dually-infected with herpes simplex virus type 2 (HSV-2). Daily HSV-2 suppression reduces plasma HIV-1 levels, but whether HSV-2 suppression delays HIV-1 disease progression is unknown. Methods Within a randomized, placebo-controlled trial of HSV-2 suppressive therapy (acyclovir 400 mg orally bid) to decrease HIV-1 transmission, 3381 HSV-2/HIV-1 dually-infected heterosexual Africans who at enrollment had CD4 counts ≥250 cells/mm3 and were not taking ART were followed for up to 24 months. We evaluated the effect of acyclovir on HIV-1 disease progression, defined by a primary composite endpoint of first occurrence of CD4 count death. As an exploratory analysis, we evaluated the endpoint of CD4 decline to HIV-1 plasma RNA was 4.1 log10 copies/mL. Acyclovir reduced risk of HIV-1 disease progression: 284 participants on acyclovir versus 324 on placebo reached the primary endpoint (hazard ratio [HR] 0.84, 95% confidence interval [CI] 0.71–0.98, p=0.03). Among participants with CD4 counts ≥350 cells/mm3, acyclovir delayed risk of CD4 decline to HIV-1 disease progression by 16% (95% CI 2–29%). The role of HSV-2 suppression in reducing HIV-1 disease progression prior to ART initiation warrants consideration (ClinicalTrials.gov #NCT00194519). PMID:20153888

  3. Astrocytes Resist HIV-1 Fusion but Engulf Infected Macrophage Material

    Directory of Open Access Journals (Sweden)

    Rebecca A. Russell

    2017-02-01

    Full Text Available HIV-1 disseminates to diverse tissues and establishes long-lived viral reservoirs. These reservoirs include the CNS, in which macrophage-lineage cells, and as suggested by many studies, astrocytes, may be infected. Here, we have investigated astrocyte infection by HIV-1. We confirm that astrocytes trap and internalize HIV-1 particles for subsequent release but find no evidence that these particles infect the cell. Astrocyte infection was not observed by cell-free or cell-to-cell routes using diverse approaches, including luciferase and GFP reporter viruses, fixed and live-cell fusion assays, multispectral flow cytometry, and super-resolution imaging. By contrast, we observed intimate interactions between HIV-1-infected macrophages and astrocytes leading to signals that might be mistaken for astrocyte infection using less stringent approaches. These results have implications for HIV-1 infection of the CNS, viral reservoir formation, and antiretroviral therapy.

  4. AIDS in rural Africa: a paradigm for HIV-1 prevention.

    Science.gov (United States)

    Hudson, C P

    1996-07-01

    Networks of concurrent sexual partnerships may be the primary cause of epidemic spread of HIV-1 in parts of sub-Saharan Africa. This pattern of sexual behaviour increases the likelihood that individuals experiencing primary HIV-1 infection transmit the virus to other persons. Networks of concurrent partnerships are likely to be important in both the early ('epidemic') and late ('endemic') phases of HIV-1 transmission. Interventions should aim to break the sexual networks, whatever the stage of the epidemic. However, prevention of transmission in the endemic phase also requires a greater awareness of early clinical manifestations of HIV-1 infection in the general population. Such awareness, coupled with the availability of condoms and access to HIV-1 testing facilities, may reduce transmission in discordant couples.

  5. Sexually transmitted infections among HIV-1-discordant couples.

    Directory of Open Access Journals (Sweden)

    Brandon L Guthrie

    2009-12-01

    Full Text Available More new HIV-1 infections occur within stable HIV-1-discordant couples than in any other group in Africa, and sexually transmitted infections (STIs may increase transmission risk among discordant couples, accounting for a large proportion of new HIV-1 infections. Understanding correlates of STIs among discordant couples will aid in optimizing interventions to prevent HIV-1 transmission in these couples.HIV-1-discordant couples in which HIV-1-infected partners were HSV-2-seropositive were tested for s