No evidence for selection of HIV-1 with enhanced Gag-Protease or Nef function among breakthrough infections in the CAPRISA 004 tenofovir microbicide trial

BACKGROUND: Use of antiretroviral-based microbicides for HIV-1 prophylaxis could introduce a transmission barrier that inadvertently facilitates the selection of fitter viral variants among incident infections. To investigate this, we assessed the in vitro function of gag-protease and nef sequences from participants who acquired HIV-1 during the CAPRISA 004 1% tenofovir microbicide gel trial. Methods and RESULTS: We isolated the earliest available gag-protease and nef gene sequences from 83 individuals and examined their in vitro function using recombinant viral replication capacity assays and surface protein downregulation assays, respectively. No major phylogenetic clustering and no significant differences in gag-protease or nef function were observed in participants who received tenofovir gel versus placebo gel prophylaxis. CONCLUSION: Results indicate that the partial protective effects of 1% tenofovir gel use in the CAPRISA 004 trial were not offset by selection of transmitted/early HIV-1 variants with enhanced Gag-Protease or Nef fitness

Subtle Longitudinal Alterations in Env Sequence Potentiate Differences in Sensitivity to Broadly Neutralizing Antibodies following Acute HIV-1 Subtype C Infection

Broadly neutralizing antibodies (bNAbs) for HIV-1 prevention or cure strategies must inhibit transmitted/founder and reservoir viruses. Establishing sensitivity of circulating viruses to bNAbs and genetic patterns affecting neutralization variability may guide rational bNAbs selection for clinical development. We analyzed 326 single env genomes from nine individuals followed longitudinally following acute HIV-1 infection, with samples collected at ~1 week after the first detection of plasma viremia; 300 to 1,709 days postinfection but prior to initiating antiretroviral therapy (ART) (median = 724 days); and ~1 year post ART initiation. Sequences were assessed for phylogenetic relatedness, potential N- and O-linked glycosylation, and variable loop lengths (V1 to V5). A total of 43 env amplicons (median = 3 per patient per time point) were cloned into an expression vector and the TZM-bl assay was used to assess the neutralization profiles of 15 bNAbs targeting the CD4 binding site, V1/V2 region, V3 supersite, MPER, gp120/gp41 interface, and fusion peptide. At 1 μg/mL, the neutralization breadths were as follows: VRC07-LS and N6.LS (100%), VRC01 (86%), PGT151 (81%), 10-1074 and PGT121 (80%), and less than 70% for 10E8, 3BNC117, CAP256.VRC26, 4E10, PGDM1400, and N123-VRC34.01. Features associated with low sensitivity to V1/V2 and V3 bNAbs were higher potential glycosylation sites and/or relatively longer V1 and V4 domains, including known "signature" mutations. The study shows significant variability in the breadth and potency of bNAbs against circulating HIV-1 subtype C envelopes. VRC07-LS, N6.LS, VRC01, PGT151, 10-1074, and PGT121 display broad activity against subtype C variants, and major determinants of sensitivity to most bNAbs were within the V1/V4 domains. IMPORTANCE Broadly neutralizing antibodies (bNAbs) have potential clinical utility in HIV-1 prevention and cure strategies. However, bNAbs target diverse epitopes on the HIV-1 envelope and the virus may evolve to evade immune responses. It is therefore important to identify antibodies with broad activity in high prevalence settings, as well as the genetic patterns that may lead to neutralization escape. We investigated 15 bNAbs with diverse biophysical properties that target six epitopes of the HIV-1 Env glycoprotein for their ability to inhibit viruses that initiated infection, viruses circulating in plasma at chronic infection before antiretroviral treatment (ART), or viruses that were archived in the reservoir during ART in subtype C infected individuals in South Africa, a high burden country. We identify the antibodies most likely to be effective for clinical use in this setting and describe mutational patterns associated with neutralization escape from these antibodies

No evidence for selection of HIV-1 with enhanced gag-protease or nef function among breakthrough infections in the CAPRISA 004 tenofovir microbicide trial.

CAPRISA, 2013.Background: Use of antiretroviral-based microbicides for HIV-1 prophylaxis could introduce a transmission barrier that inadvertently facilitates the selection of fitter viral variants among incident infections. To investigate this, we assessed the in vitro function of gag-protease and nef sequences from participants who acquired HIV-1 during the CAPRISA 004 1% tenofovir microbicide gel trial. Methods and Results: We isolated the earliest available gag-protease and nef gene sequences from 83 individuals and examined their in vitro function using recombinant viral replication capacity assays and surface protein down regulation assays, respectively. No major phylogenetic clustering and no significant differences in gag-protease or nef function were observed in participants who received tenofovir gel versus placebo gel prophylaxis. Conclusion: Results indicate that the partial protective effects of 1% tenofovir gel use in the CAPRISA 004 trial were not offset by selection of transmitted/early HIV-1 variants with enhanced Gag-Protease or Nef fitness

Early immune adaptation in HIV-1 revealed by population-level approaches

Background: The reproducible nature of HIV-1 escape from HLA-restricted CD8+ T-cell responses allows the identification of HLA-associated viral polymorphisms “at the population level” – that is, via analysis of cross-sectional, linked HLA/HIV-1 genotypes by statistical association. However, elucidating their timing of selection traditionally requires detailed longitudinal studies, which are challenging to undertake on a large scale. We investigate whether the extent and relative timecourse of immune-driven HIV adaptation can be inferred via comparative cross-sectional analysis of independent early and chronic infection cohorts. Results: Similarly-powered datasets of linked HLA/HIV-1 genotypes from individuals with early (median  200/dataset), HLA class I and HIV-1 Gag/Pol/Nef diversity, were established. These datasets were first used to define a list of 162 known HLA-associated polymorphisms detectable at the population level in cohorts of the present size and host/viral genetic composition. Of these 162 known HLA-associated polymorphisms, 15% (occurring at 14 Gag, Pol and Nef codons) were already detectable via statistical association in the early infection dataset at p ≤ 0.01 (q < 0.2) – identifying them as the most consistently rapidly escaping sites in HIV-1. Among these were known rapidly-escaping sites (e.g. B*57-Gag-T242N) and others not previously appreciated to be reproducibly rapidly selected (e.g. A*31:01-associated adaptations at Gag codons 397, 401 and 403). Escape prevalence in early infection correlated strongly with first-year escape rates (Pearson’s R = 0.68, p = 0.0001), supporting cross-sectional parameters as reliable indicators of longitudinally-derived measures. Comparative analysis of early and chronic datasets revealed that, on average, the prevalence of HLA-associated polymorphisms more than doubles between these two infection stages in persons harboring the relevant HLA (p < 0.0001, consistent with frequent and reproducible escape), but remains relatively stable in persons lacking the HLA (p = 0.15, consistent with slow reversion). Published HLA-specific Hazard Ratios for progression to AIDS correlated positively with average escape prevalence in early infection (Pearson’s R = 0.53, p = 0.028), consistent with high early within-host HIV-1 adaptation (via rapid escape and/or frequent polymorphism transmission) as a correlate of progression. Conclusion: Cross-sectional host/viral genotype datasets represent an underutilized resource to identify reproducible early pathways of HIV-1 adaptation and identify correlates of protective immunity.Medicine, Faculty ofNon UBCReviewedFacult

Sequence analyses of HIV-1 Gag and Nef from treatment and placebo arms.

<p>Maximum-likelihood phylogenetic trees of bulk <i>Gag</i> sequences (Panel A) and clonal <i>Nef</i> sequences (panel B) from participants who received 1% tenofovir gel (red) or placebo (black) demonstrated no major clustering between arms of the trial. HIV-1 subtype B reference sequence HXB2 is shown in green.</p

Replication capacity of recombinant viruses expressing <i>Gag-Protease</i> sequences from 1% tenofovir gel and placebo participants.

<p>Panel A: Replication data are shown for recombinant NL4-3 viruses encoding <i>gag-protease</i> from participants in the 1% tenofovir gel (red) and placebo (black) study arms, expressed as fold-increase in GFP expression over 6 days, relative to day 2. WT NL4-3 control is indicated in blue. Panel B: The replication capacities of recombinant viruses encoding participant-derived <i>gag-protease</i> sequences, calculated as the slope of viral spread normalised to WT NL4-3, are shown. No signifant difference in Gag-Protease function was observed between study arms (Mann-Whitney, p = 0.2).</p