Si Z, Gorry P, Babcock G, Owens CM, Cayabyab M, Phan N, Sodroski JEnvelope glycoprotein determinants of increased entry in a pathogenic simian-human immunodeficiency virus (SHIV-HXBc2P 3.2) passaged in monkeys. AIDS Res. Hum. Retroviruses 2004;20(2):163-73
Passage of a nonpathogenic simian-human immunodeficiency virus (SHIV-HXBc2) in monkeys resulted in changes in the viral envelope glycoproteins that are responsible for a dramatic increase in replication and pathogenicity in vivo. Here, we show that the envelope glycoproteins of the pathogenic SHIV-HXBc2P 3.2 mediate virus entry into rhesus monkey peripheral blood mononuclear cells (PBMC) more efficiently than the parental SHIV-HXBc2 envelope glycoproteins, and study the basis for this increase. Both parental and pathogenic SHIVs exclusively use CXCR4 as a coreceptor. The determinants of the increased entry associated with the SHIV-HXBc2P 3.2 envelope glycoproteins are located in both the gp120 and gp41 subunits. Changes in the gp120 V3 variable loop specify a decreased sensitivity to SDF-1, consistent with an increase in the affinity of the HXBc2P 3.2 gp120 glycoprotein for CXCR4. Thus, multiple changes in the gp120 variable loops and the gp41 ectodomain of a pathogenic SHIV cooperate to allow enhanced replicative capacity, which in part results from increased chemokine receptor binding.
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Desrosiers RCProspects for an AIDS vaccine. Nat. Med. 2004;10(3):221-3
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Friedrich TC, Dodds EJ, Yant LJ, Vojnov L, Rudersdorf R, Cullen C, Evans DT, Desrosiers RC, Mothé BR, Sidney J, Sette A, Kunstman K, Wolinsky S, Piatak M, Lifson J, Hughes AL, Wilson N, O'Connor DH, Watkins DIReversion of CTL escape-variant immunodeficiency viruses in vivo. Nat. Med. 2004;10(3):275-81
Engendering cytotoxic T-lymphocyte (CTL) responses is likely to be an important goal of HIV vaccines. However, CTLs select for viral variants that escape immune detection. Maintenance of such escape variants in human populations could pose an obstacle to HIV vaccine development. We first observed that escape mutations in a heterogeneous simian immunodeficiency virus (SIV) isolate were lost upon passage to new animals. We therefore infected macaques with a cloned SIV bearing escape mutations in three immunodominant CTL epitopes, and followed viral evolution after infection. Here we show that each mutant epitope sequence continued to evolve in vivo, often re-establishing the original, CTL-susceptible sequence. We conclude that escape from CTL responses may exact a cost to viral fitness. In the absence of selective pressure upon transmission to new hosts, these original escape mutations can be lost. This suggests that some HIV CTL epitopes will be maintained in human populations.
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Mwau M, Cebere I, Sutton J, Chikoti P, Winstone N, Wee EG, Beattie T, Chen YH, Dorrell L, McShane H, Schmidt C, Brooks M, Patel S, Roberts J, Conlon C, Rowland-Jones SL, Bwayo JJ, McMichael AJ, Hanke T
A human immunodeficiency virus 1 (HIV-1) clade A vaccine in clinical trials: stimulation of HIV-specific T-cell responses by DNA and recombinant modified vaccinia virus Ankara (MVA) vaccines in humans. J. Gen. Virol. 2004;85(Pt 4):911-9
The immunogenicities of candidate DNA- and modified vaccinia virus Ankara (MVA)-vectored human immunodeficiency virus (HIV) vaccines were evaluated on their own and in a prime-boost regimen in phase I clinical trials in healthy uninfected individuals in the United Kingdom. Given the current lack of approaches capable of inducing broad HIV-neutralizing antibodies, the pTHr.HIVA DNA and MVA.HIVA vaccines focus solely on the induction of cell-mediated immunity. The vaccines expressed a common immunogen, HIVA, which consists of consensus HIV-1 clade A Gag p24/p17 proteins fused to a string of clade A-derived epitopes recognized by cytotoxic T lymphocytes (CTLs). Volunteers' fresh peripheral blood mononuclear cells were tested for HIV-specific responses in a validated gamma interferon enzyme-linked immunospot (ELISPOT) assay using four overlapping peptide pools across the Gag domain and three pools of known CTL epitopes present in all of the HIVA protein. Both the DNA and the MVA vaccines alone and in a DNA prime-MVA boost combination were safe and induced HIV-specific responses in 14 out of 18, seven out of eight and eight out of nine volunteers, respectively. These results are very encouraging and justify further vaccine development.
Topics: IAVI Clinical Trial
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Rybarczyk BJ, Montefiori D, Johnson PR, West A, Johnston RE, Swanstrom RCorrelation between env V1/V2 region diversification and neutralizing antibodies during primary infection by simian immunodeficiency virus sm in rhesus macaques. J. Virol. 2004;78(7):3561-71
Evolution of the domain encoding the V1/V2 variable region of the simian immunodeficiency virus sm (SIVsm) envelope (env) gene was analyzed in relation to route of virus challenge, virus load, and neutralizing antibody (NAb) titers during primary infection of rhesus macaques with the pathogenic SIVsmE660 isolate. In this model system animals are initially infected with multiple viruses as evidenced by the presence of multiple V1/V2 genotypic variants that could be resolved by using a heteroduplex tracking assay (HTA). Overlapping subsets of the multiple variants were established in each animal. There was no selection for the establishment of specific variants in comparing intravenous- and intrarectal-challenged macaques at week 2 postinfection, suggesting that no genotypic selection occurred at the mucosal surface. There was an initial period of significant stability of the V1/V2 variants. Macaques challenged intravenously displayed subsequent V1/V2 diversification significantly earlier than macaques challenged intrarectally and well past the initial resolution of viremia. The time when SIVsmE660-specific NAbs reached a threshold titer of 100 was significantly correlated with the timing of V1/V2 diversification, even though antibodies to the Env protein could be detected much earlier. The time when NAbs reached a titer of 400 was significantly correlated with virus load late in infection. These results show that the route of infection affects the timing of V1/V2 diversification and that this diversification is correlated with the maturation of a specific NAb response. However, prior immunization capable of priming an anamnestic Env antibody response did not accelerate V1/V2 diversification. This result suggests that diversification of the SIV env V1/V2 region is the result of a type-specific antibody response.
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Kuhmann SE, Pugach P, Kunstman KJ, Taylor J, Stanfield RL, Snyder A, Strizki JM, Riley J, Baroudy BM, Wilson IA, Korber BT, Wolinsky SM, Moore JPGenetic and phenotypic analyses of human immunodeficiency virus type 1 escape from a small-molecule CCR5 inhibitor. J. Virol. 2004;78(6):2790-807
We have described previously the generation of an escape variant of human immunodeficiency virus type 1 (HIV-1), under the selection pressure of AD101, a small molecule inhibitor that binds the CCR5 coreceptor (A. Trkola, S. E. Kuhmann, J. M. Strizki, E. Maxwell, T. Ketas, T. Morgan, P. Pugach, S. X. L. Wojcik, J. Tagat, A. Palani, S. Shapiro, J. W. Clader, S. McCombie, G. R. Reyes, B. M. Baroudy, and J. P. Moore, Proc. Natl. Acad. Sci. USA 99:395-400, 2002). The escape mutant, CC101.19, continued to use CCR5 for entry, but it was at least 20,000-fold more resistant to AD101 than the parental virus, CC1/85. We have now cloned the env genes from the the parental and escape mutant isolates and made chimeric infectious molecular clones that fully recapitulate the phenotypes of the corresponding isolates. Sequence analysis of the evolution of the escape mutants suggested that the most relevant changes were likely to be in the V3 loop of the gp120 glycoprotein. We therefore made a series of mutant viruses and found that full AD101 resistance was conferred by four amino acid changes in V3. Each change individually caused partial resistance when they were introduced into the V3 loop of a CC1/85 clone, but their impact was dependent on the gp120 context in which they were made. We assume that these amino acid changes alter how the HIV-1 Env complex interacts with CCR5. Perhaps unexpectedly, given the complete dependence of the escape mutant on CCR5 for entry, monomeric gp120 proteins expressed from clones of the fully resistant isolate failed to bind to CCR5 on the surface of L1.2-CCR5 cells under conditions where gp120 proteins from the parental virus and a partially AD101-resistant virus bound strongly. Hence, the full impact of the V3 substitutions may only be apparent at the level of the native Env complex.
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Madani N, Perdigoto AL, Srinivasan K, Cox JM, Chruma JJ, LaLonde J, Head M, Smith AB, Sodroski JGLocalized changes in the gp120 envelope glycoprotein confer resistance to human immunodeficiency virus entry inhibitors BMS-806 and #155. J. Virol. 2004;78(7):3742-52
BMS-806 and the related compound, #155, are novel inhibitors of human immunodeficiency virus type 1 (HIV-1) entry that bind the gp120 exterior envelope glycoprotein. BMS-806 and #155 block conformational changes in the HIV-1 envelope glycoproteins that are induced by binding to the host cell receptor, CD4. We tested a panel of HIV-1 envelope glycoprotein mutants and identified several that were resistant to the antiviral effects of BMS-806 and #155. In the CD4-bound conformation of gp120, the amino acid residues implicated in BMS-806 and #155 resistance line the 'phenylalanine 43 cavity' and a water-filled channel that extends from this cavity to the inner domain. Structural considerations suggest a model in which BMS-806 and #155 bind gp120 prior to receptor binding and, upon CD4 binding, are accommodated in the Phe-43 cavity and adjacent channel. The integrity of the nearby V1/V2 variable loops and N-linked carbohydrates on the V1/V2 stem indirectly influences sensitivity to the drugs. A putative binding site for BMS-806 and #155 between the gp120 receptor-binding regions and the inner domain, which is thought to interact with the gp41 transmembrane envelope glycoprotein, helps to explain the mode of action of these drugs.
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McDermott AB, Mitchen J, Piaskowski S, De Souza I, Yant LJ, Stephany J, Furlott J, Watkins DIRepeated low-dose mucosal simian immunodeficiency virus SIVmac239 challenge results in the same viral and immunological kinetics as high-dose challenge: a model for the evaluation of vaccine efficacy in nonhuman primates. J. Virol. 2004;78(6):3140-4
Simian immunodeficiency virus (SIV) challenge of rhesus macaques provides a relevant model for the assessment of human immunodeficiency virus (HIV) vaccine strategies. To ensure that all macaques become infected, the vaccinees and controls are exposed to large doses of pathogenic SIV. These nonphysiological high-dose challenges may adversely affect vaccine evaluation by overwhelming potentially efficacious vaccine responses. To determine whether a more physiologically relevant low-dose challenge can initiate infection and cause disease in Indian rhesus macaques, we used a repeated low-dose challenge strategy designed to reduce the viral inoculum to more physiologically relevant doses. In an attempt to more closely mimic challenge with HIV, we administered repeated mucosal challenges with 30, 300, and 3,000 50% tissue culture infective doses (TCID(50)) of pathogenic SIVmac239 to six animals in three groups. Infection was assessed by sensitive quantitative reverse transcription-PCR and was achieved following a mean of 8, 5.5, and 1 challenge(s) in the 30, 300, and 3,000 TCID(50) groups, respectively. Mortality, humoral immune responses, and peak plasma viral kinetics were similar in five of six animals, regardless of challenge dose. Interestingly, macaques challenged with lower doses of SIVmac239 developed broad T-cell immune responses as assessed by ELISPOT assay. This low-dose repeated challenge may be a valuable tool in the evaluation of potential vaccine regimes and offers a more physiologically relevant regimen for pathogenic SIVmac239 challenge experiments.
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Stremlau M, Owens CM, Perron MJ, Kiessling M, Autissier P, Sodroski JThe cytoplasmic body component TRIM5alpha restricts HIV-1 infection in Old World monkeys. Nature 2004;427(6977):848-53
Host cell barriers to the early phase of immunodeficiency virus replication explain the current distribution of these viruses among human and non-human primate species. Human immunodeficiency virus type 1 (HIV-1), the cause of acquired immunodeficiency syndrome (AIDS) in humans, efficiently enters the cells of Old World monkeys but encounters a block before reverse transcription. This species-specific restriction acts on the incoming HIV-1 capsid and is mediated by a dominant repressive factor. Here we identify TRIM5alpha, a component of cytoplasmic bodies, as the blocking factor. HIV-1 infection is restricted more efficiently by rhesus monkey TRIM5alpha than by human TRIM5alpha. The simian immunodeficiency virus, which naturally infects Old World monkeys, is less susceptible to the TRIM5alpha-mediated block than is HIV-1, and this difference in susceptibility is due to the viral capsid. The early block to HIV-1 infection in monkey cells is relieved by interference with TRIM5alpha expression. Our studies identify TRIM5alpha as a species-specific mediator of innate cellular resistance to HIV-1 and reveal host cell components that modulate the uncoating of a retroviral capsid.
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Cole KS, Steckbeck JD, Rowles JL, Desrosiers RC, Montelaro RCRemoval of N-linked glycosylation sites in the V1 region of simian immunodeficiency virus gp120 results in redirection of B-cell responses to V3. J. Virol. 2004;78(3):1525-39
One mechanism of immune evasion utilized by human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) envelope glycoproteins is the presence of a dense carbohydrate shield. Accumulating evidence from in vitro and in vivo experiments suggests that alterations in N-linked glycosylation of SIV gp120 can enhance host humoral immune responses that may be involved in immune control. The present study was designed to determine the ability of glycosylation mutant viruses to redirect antibody responses to shielded envelope epitopes. The influence of glycosylation on the maturation and specificity of antibody responses elicited by glycosylation mutant viruses containing mutations of specific N-linked sites in and near the V1 and V2 regions of SIVmac239 gp120 was determined. Results from these studies demonstrated a remarkably similar maturation of antibody responses to native, fully glycosylated envelope proteins. However, analyses of antibodies to defined envelope domains revealed that mutation of glycosylation sites in V1 resulted in increased antibody recognition to epitopes in V1. In addition, we demonstrated for the first time that mutation of glycosylation sites in V1 resulted in a redirection of antibody responses to the V3 loop. Taken together, these results demonstrate that N-linked glycosylation is a determinant of SIV envelope B-cell immunogenicity in addition to in vitro antigenicity. In addition, our results demonstrate that the absence of N-linked carbohydrates at specific sites can influence the exposure of epitopes quite distant in the linear sequence.
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