Recent reports suggest that Natural Killer (NK) cells may modulate pathogenesis of primary HIV-1 infection. However, HIV dysregulates NK-cell responses. We dissected this bi-directional relationship to understand how HIV impacts NK-cell responses during primary HIV-1 infection.
Paired samples from 41 high-risk, initially HIV-uninfected CAPRISA004 participants were analysed prior to HIV acquisition, and during viraemic primary HIV-1 infection. At the time of sampling post-infection five women were seronegative, 11 women were serodiscordant, and 25 women were seropositive by HIV-1 rapid immunoassay. Flow cytometry was used to measure NK and T-cell activation, NK-cell receptor expression, cytotoxic and cytokine-secretory functions, and trafficking marker expression (CCR7, α4β7). Non-parametric statistical tests were used. Both NK cells and T-cells were significantly activated following HIV acquisition (p = 0.03 and p<0.0001, respectively), but correlation between NK-cell and T-cell activation was uncoupled following infection (pre-infection r = 0.68;p<0.0001; post-infection, during primary infection r = 0.074;p = 0.09). Nonetheless, during primary infection NK-cell and T-cell activation correlated with HIV viral load (r = 0.32'p = 0.04 and r = 0.35;p = 0.02, respectively). The frequency of Killer Immunoglobulin-like Receptor-expressing (KIRpos) NK cells increased following HIV acquisition (p = 0.006), and KIRpos NK cells were less activated than KIRneg NK cells amongst individuals sampled while seronegative or serodiscordant (p = 0.001;p<0.0001 respectively). During HIV-1 infection, cytotoxic NK cell responses evaluated after IL-2 stimulation alone, or after co-culture with 721 cells, were impaired (p = 0.006 and p = 0.002, respectively). However, NK-cell IFN-y secretory function was not significantly altered. The frequency of CCR7+ NK cells was elevated during primary infection, particularly at early time-points (p<0.0001).
Analyses of immune cells before and after HIV infection revealed an increase in both NK-cell activation and KIR expression, but reduced cytotoxicity during acute infection. The increase in frequency of NK cells able to traffic to lymph nodes following HIV infection suggests that these cells may play a role in events in secondary lymphoid tissue.
Concerns about immune reconstitution inflammatory syndrome (IRIS) remain a barrier to antiretroviral therapy (ART) initiation during anti-tuberculosis treatment in co-infected patients.
We assessed IRIS incidence, severity, and outcomes relative to timing of ART initiation in patients with HIV-related tuberculosis (HIV-TB).
An outpatient clinic in Durban, South Africa
642 HIV-TB co-infected patients
In a secondary analysis of the SAPiT trial, IRIS was assessed in patients randomized to initiate ART either within four weeks of tuberculosis treatment initiation (early integrated-treatment arm), within four weeks of completion of the intensive phase of tuberculosis treatment (late integrated-treatment arm) or within four weeks after tuberculosis therapy completion (sequential-treatment arm). IRIS was defined as new onset or worsening symptoms, signs or radiographic manifestations temporally related to treatment initiation accompanied by a treatment response. IRIS severity, hospitalization and time to resolution were monitored.
IRIS incidence was 19.5 (n=43), 7.5 (n=18) and 8.1 (n=19) per 100 person-years in the early integrated-, late integrated-, and sequential-treatment arms, respectively; P < 0.001, and 45.5, 9.7 and 19.7 per 100 person-years in patients with baseline CD4+ counts <50 cells/mm3, P = 0.004. IRIS incidence was higher in the early integrated- compared to the late integrated- (incidence rate ratio (IRR) = 2.6, 95%confidence interval (CI): 1.5 to 4.8; P < 0.001) or sequential-treatment arm (IRR=2.4, 95%CI: 1.4 to 4.4; P < 0.001). IRIS cases in the early integrated-treatment arm were more severe (34.9% vs. 18.9%, P = 0.18); had significantly higher hospitalization rates (18/43 vs. 5/37; P = 0.01), and longer time to resolution (70.5 vs. 29.0 days; P = 0.001) compared to IRIS cases in the other two arms.
IRIS could not be assessed, due to LTFU, withdrawal or death within 6 months of scheduled ART initiation, in more patients from the sequential treatment arm (n=74) than in the late integrated treatment arm (n=50) and in the early integrated treatment arm (n=32). This study did not assess IRIS risk in non-ambulant patients and in patients with extra-pulmonary and smear negative pulmonary tuberculosis.
Initiation of ART early during tuberculosis treatment resulted in significantly higher IRIS rates, with longer time to resolution, and more severe cases of IRIS requiring hospitalization. These findings, particularly relevant to patients initiating ART with CD4+ counts < 50 cells/mm3, need to be considered together with the increased survival benefit of early ART initiation in this group. Clintrials.gov: NCT00398996
To determine the safety and effectiveness of BufferGel and 0.5% PRO2000 microbicide gels for the prevention of male to female HIV transmission
Phase II/IIb, randomized, placebo-controlled trial with three double-blinded gel arms and an open label no gel arm.
Study participants from Malawi, South Africa, Zambia, Zimbabwe and USA were instructed to apply study gel ≤1 hour before each sex act and safety, sexual behavior, pregnancy, gel adherence, acceptability, and HIV serostatus were assessed during follow-up.
The 3101 enrolled women were followed for an average of 20.4 months with 93.6% retention and 81.1% self-reported gel adherence. Adverse event rates were similar in all study arms. HIV incidence rates in the 0.5% PRO2000 Gel, BufferGel, Placebo Gel and No Gel arms were 2.70, 4.14, 3.91 and 4.02 per 100 women-years, respectively. HIV incidence in the 0.5% PRO2000 Gel arm was lower than the Placebo Gel arm (Hazard Ratio (HR)=0.7; p=0.10) and the No Gel arm (HR=0.67; p=0.06). HIV incidence rates were similar in the BufferGel and both Placebo Gel (HR=1.10; p=0.63) and No Gel control arms (HR=1.05; p=0.78). HIV incidence was similar in the Placebo Gel and No Gel arms (HR=0.97; p=0.89).
0.5% PRO2000 Gel demonstrated a modest 30% reduction in HIV acquisition in women. However, these results were not statistically significant and subsequent findings from the MDP 301 trial have confirmed that 0.5% PRO2000 has little or no protective effect. BufferGel did not alter the risk of HIV infection. Both products were safe.
Microbicide; PRO 2000 Gel; BufferGel; HIV Prevention; Women
Frequency of drug changes in combination antiretroviral therapy among patients starting both tuberculosis (TB) and human immunodeficiency virus (HIV) therapy, as a result of treatment-limiting toxicity or virological failure, is not well established.
Patients in the Starting Antiretroviral Therapy at Three Points in Tuberculosis (SAPiT) trial were randomized to initiate antiretroviral therapy either early or late during TB treatment or after completion of TB treatment. Drug changes due to toxicity (defined as due to grade 3 or 4 adverse events) or virological failure (defined as viral load > 1000 copies/ml on two occasions, taken at least 4 weeks apart) were assessed in these patients.
A total of 501 TB-HIV co-infected patients were followed for a mean of 16.0 (95% confidence interval (CI): 15.5 to 16.6) months after antiretroviral therapy (ART) initiation. The standard first-line ARVs used, were efavirenz, lamivudine and didanosine. Individual drug switches for toxicity occurred in 14 patients (incidence rate: 2.1 per 100 person-years; 95% (CI): 1.1 to 3.5), and complete regimen changes due to virological failure in 25 patients (incidence rate: 3.7 per 100 person-years; CI: 2.4 to 5.5). The most common treatment limiting toxicities were neuropsychiatric effects (n=4; 0.8%), elevated transaminase levels and hyperlactatemia (n= 3; 0.6%), and peripheral neuropathy (n=2; 0.4%). Complete regimen change due to treatment failure was more common in patients with CD4+ cell count <50cells/mm3 (p<0.001) at ART initiation and body mass index greater than 25 kg/m2 (p=0.01) at entry into the study.
Both drug switches and complete regimen change were uncommon in patients co-treated for TB-HIV with the chosen regimen. Patients with severe immunosuppression need to be monitored carefully, as they were most at risk for treatment failure requiring regimen change.
To estimate the effectiveness of candidate microbicides BufferGel and 0.5% PRO 2000 Gel (P) (PRO 2000) for prevention of non-ulcerative sexually transmitted infections (STIs).
Between 2005 and 2007, 3099 women were enrolled in HIV Prevention Trials Network (HPTN) protocol 035, a phase II/IIb evaluation of the safety and effectiveness of BufferGel and PRO 2000 for prevention of sexually transmitted infections, including Neisseria gonorrhoeae (GC), Chlamydia trachomatis (CT), and Trichomonas vaginalis (TV). Incidences of STIs were determined by study arm, and hazard ratios (HRs) of BufferGel and PRO 2000 versus placebo gel or no gel control groups were computed using discrete time Andersen-Gill proportional hazards model.
The overall incidence rates were 1.6/100 person-years at risk (PYAR) for GC, 3.9/100 PYAR for CT, and 15.3/100 PYAR for TV. For BufferGel versus placebo gel, HRs were 0.99 (95% CI 0.49–2.00), 1.00 (95% CI 0.64–1.57), and 0.95 (95% CI 0.71–1.25) for prevention of GC, CT, and TV respectively. For PRO 2000, HRs were 1.66 (95% CI 0.90–3.06), 1.16 (95% CI 0.76–1.79), and 1.18 (95% CI 0.90–1.53) for prevention of GC, CT, and TV respectively.
The incidence of STIs was high during HPTN 035 despite provision of free condoms and comprehensive risk-reduction counselling, highlighting the need for effective STI prevention programmes in this population. Unfortunately, candidate microbicides BufferGel and PRO2000 had no protective effect against gonorrhoea, Chlamydia, or trichomoniasis.
Neisseria gonorrhoeae; Chlamydia trachomatis; Trichomonas; microbicides; prevention
Tenofovir disoproxil fumarate, a licensed oral treatment for both HIV and Hepatitis B virus (HBV) infections, has been associated with severe rebound hepatic flares when treatment is interrupted. A gel formulation of tenofovir is currently being assessed as a microbicide against HIV. If licensed, it is possible that tenofovir gel could be used either intentionally or unintentionally by HBV carriers. The purpose of this study was to establish the safety of tenofovir gel use in this patient group participating in the CAPRISA 004 tenofovir gel trial. HBV infection status was assessed at enrolment and study exit. Liver function testing was performed at enrolment, study months 3, 12, 24, study exit, and two months after exiting the study. At enrolment, 34 women were identified as being HBV carriers and 22 women acquired HBV infections during follow-up; 14 and 8 in the tenofovir and placebo gel arms respectively (p=0.21). Intermittent tenofovir gel use did not cause an increase in hepatic flares or impact on viral load suppression in women with HBV infection. There were 2 hepatic flares in each gel arm during follow-up and none two months after cessation of gel at study exit. The mean HBV DNA levels were similar at enrolment and exit in both study arms. Tenofovir gel, when used intermittently, was safe to use in women with HBV infection.
microbicides; Hepatitis B virus; tenofovir gel; HIV prevention; hepatic flares
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.
Methods and Findings
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.
To compare the two control arms of HPTN 035 (a hydroxyethylcellulose (HEC) gel control arm and a no gel control arm) to assess behavioral effects associated with gel use and direct causal effects of the HEC gel on STIs, pregnancy, and genital safety.
Randomized trial with one blinded (HEC gel) and one open label (no gel) control arms.
HIV-uninfected, sexually active women were randomized into the HEC gel arm (n=771) and into the no gel arm (n=772) in five countries. Participants in the HEC gel arm were instructed to insert the study gel intravaginally <1 hour before each vaginal sex act. Data on sexual behavior, adherence, safety, pregnancy, and STIs were collected quarterly for 12 to 30 months of follow-up.
During follow-up, mean reported condom use in the past week was significantly higher in the no gel arm (81% versus 70%, p<0.001). There were no significant differences, after adjusting for this differential condom use, between the two arms in rates of genital safety events, pregnancy outcomes, or STIs, including HIV-1.
In this large randomized trial, we found no significant differences between the no gel and HEC gel arms in rates of genital safety events, pregnancy outcomes, or STIs. These results aid interpretation of the results of previous vaginal microbicide trials that used the HEC gel as a control. The HEC gel is suitable as a control for ongoing and future vaginal microbicide studies.
HIV prevention; vaginal microbicide; placebo; women; sexually transmitted infections
Adherence undeniably impacts product effectiveness in microbicide trials, but the connection has proven challenging to quantify using routinely collected behavioral data. We explored this relationship using a nested case–control study in the CAPRISA 004 Tenofovir (TFV) gel HIV prevention trial. Detailed 3-month recall data on sex events, condom and gel use were collected from 72 incident cases and 205 uninfected controls. We then assessed how the relationship between self-reported adherence and HIV acquisition differed between the TFV and placebo gel groups, an interaction effect that should exist if effectiveness increases with adherence. The CAPRISA 004 trial determined that randomization to TFV gel was associated with a significant reduction in risk of HIV acquisition. In our nested case–control study, however, we did not observe a meaningful decrease in the relative odds of infection—TFV versus placebo—as self-reported adherence increased. To the contrary, exploratory sub-group analysis of the case–control data identified greater evidence for a protective effect of TFV gel among participants reporting less than 80 % adherence to the protocol-defined regimen (odds ratio (OR) 0.30; 95 % CI 0.11–0.78) than among those reporting ≥80 % adherence (Odds Ratio 0.81; 95 % CI 0.34–1.92). The small number of cases may have inhibited our ability to detect the hypothesized interaction between adherence and effectiveness. Nonetheless, our results re-emphasize the challenges faced by investigators when adherence may be miss-measured, miss-reported, or confounded with the risk of HIV.
Microbicides; Tenofovir gel; Adherence; HIV prevention; Self-report
There is limited information on full-length genome sequences and the
early evolution of transmitted HIV-1 subtype C viruses, which constitute the
majority of viruses spread in Africa. The purpose of this study was to
characterize the earliest changes across the genome of subtype C viruses
following transmission, to better understand early control of viremia.
We derived the near full-length genome sequence responsible for
clinical infection from five HIV subtype C-infected individuals with
different disease progression profiles and tracked adaption to immune
responses in the first six months of infection.
Near full-length genomes were generated by single genome
amplification and direct sequencing. Sequences were analyzed for amino acid
mutations associated with cytotoxic T-lymphocyte (CTL) or antibody
(Ab)-mediated immune pressure, and for reversion.
Fifty-five sequence changes associated with adaptation to the new
host were identified with 38% attributed to CTL pressure;
35% to antibody pressure; 16% to reversions and the
remainder were unclassified. Mutations in CTL epitopes were most frequent in
the first 5 weeks of infection, with the frequency declining over time with
the decline in viral load. CTL escape predominantly occurred in
nef, followed by pol and
env. Shuffling/toggling of mutations was identified in
81% of CTL epitopes with only 7% reaching fixation within
the six month period.
There was rapid virus adaptation following transmission,
predominantly driven by CTL pressure, with most changes occurring during
high viremia. Rapid escape and complex escape pathways provide further
challenges for vaccine protection.
HIV-1; Africa; genome; acute infection; cytotoxic T-lymphocytes; progression
Understanding human immunodeficiency virus type 1 (HIV-1) transmission is central to developing effective prevention strategies, including a vaccine. We compared phenotypic and genetic variation in HIV-1 env genes from subjects in acute/early infection and subjects with chronic infections in the context of subtype C heterosexual transmission. We found that the transmitted viruses all used CCR5 and required high levels of CD4 to infect target cells, suggesting selection for replication in T cells and not macrophages after transmission. In addition, the transmitted viruses were more likely to use a maraviroc-sensitive conformation of CCR5, perhaps identifying a feature of the target T cell. We confirmed an earlier observation that the transmitted viruses were, on average, modestly underglycosylated relative to the viruses from chronically infected subjects. This difference was most pronounced in comparing the viruses in acutely infected men to those in chronically infected women. These features of the transmitted virus point to selective pressures during the transmission event. We did not observe a consistent difference either in heterologous neutralization sensitivity or in sensitivity to soluble CD4 between the two groups, suggesting similar conformations between viruses from acute and chronic infection. However, the presence or absence of glycosylation sites had differential effects on neutralization sensitivity for different antibodies. We suggest that the occasional absence of glycosylation sites encoded in the conserved regions of env, further reduced in transmitted viruses, could expose specific surface structures on the protein as antibody targets.
Identification of the epitopes targeted by antibodies that can neutralize diverse HIV-1 strains can provide important clues for the design of a preventative vaccine.
We have developed a computational approach that can identify key amino acids within the HIV-1 envelope glycoprotein that influence sensitivity to broadly cross-neutralizing antibodies. Given a sequence alignment and neutralization titers for a panel of viruses, the method works by fitting a phylogenetic model that allows the amino acid frequencies at each site to depend on neutralization sensitivities. Sites at which viral evolution influences neutralization sensitivity were identified using Bayes factors (BFs) to compare the fit of this model to that of a null model in which sequences evolved independently of antibody sensitivity. Conformational epitopes were identified with a Metropolis algorithm that searched for a cluster of sites with large Bayes factors on the tertiary structure of the viral envelope.
We applied our method to ID50 neutralization data generated from seven HIV-1 subtype C serum samples with neutralization breadth that had been tested against a multi-clade panel of 225 pseudoviruses for which envelope sequences were also available. For each sample, between two and four sites were identified that were strongly associated with neutralization sensitivity (2ln(BF) > 6), a subset of which were experimentally confirmed using site-directed mutagenesis.
Our results provide strong support for the use of evolutionary models applied to cross-sectional viral neutralization data to identify the epitopes of serum antibodies that confer neutralization breadth.
HIV; Antibodies; Neutralization sensitivity; Epitope prediction; Evolutionary model
Young women are particularly vulnerable for acquiring HIV yet they are often excluded from clinical trials testing new biomedical intervention. We assessed the HIV incidence and feasibility of enrolling a cohort of young women for potential participation in future clinical trials. Between March 2004 and May 2007, 594 HIV uninfected 14–30 year old women were enrolled into a longitudinal HIV risk reduction study in KwaZulu-Natal, South Africa. The overall HIV prevalence at screening in young girls below the age of 18 years of age was 27.6% compared to 52.0% in the women above 18 years, p<0.001. HIV incidence was 4.7 [95% Confidence interval (CI) 1.5–10.9) and 6.9 (95% CI 4.8–9.6)/100 women years (wy), p=0.42 and pregnancy rates were 23.7 (95% CI 14.9–35.9) and 16.4 (95% CI 12.9–20.6)/100wy, p=0.29, in the women below and above 18 years respectively. Retention was similar in both groups (71.0% versus 71.5%, p=0.90). This study demonstrates that the inclusion of young girls between the ages of 14 and 17 years in longitudinal studies is feasible and their inclusion in clinical trials would maintain scientific integrity and power of the study.
young girls; biomedical HIV prevention research; South Africa
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.
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.
Alterations of the genital mucosal barrier may influence the number of viruses transmitted from a human immunodeficiency virus–infected source host to the newly infected individual. We used heteroduplex tracking assay and single-genome sequencing to investigate the effect of a tenofovir-based microbicide gel on the transmission bottleneck in women who seroconverted during the CAPRISA 004 microbicide trial. Seventy-seven percent (17 of 22; 95% confidence interval [CI], 56%–90%) of women in the tenofovir gel arm were infected with a single virus compared with 92% (13 of 14; 95% CI, 67%–>99%) in the placebo arm (P = .37). Tenofovir gel had no discernable impact on the transmission bottleneck.
The CAPRISA004 trial demonstrated reduction of sexual HIV-1 acquisition in women using a vaginal microbicide containing tenofovir. A better understanding of the consequences of antiretroviral-containing microbicides for immune responses in individuals with intercurrent HIV-1 infection is needed for future trials combining the use of microbicides with HIV-1 vaccines. Investigation of immune responses in women who acquired HIV-1 whilst using tenofovir gel showed significantly higher (p=0.01) Gag-specific IFNγ+ CD4+ T-cell responses. The use of tenofovir containing gel around the time of infection can modulate HIV-1 immunity, and these immunological changes need to be considered in future trials combining vaccines and microbicides.
HIV-1; vaginal microbicide; tenofovir; HIV-1-specific CD4+ T cell help
Certain immune-driven mutations in HIV-1, such as those arising in p24Gag, decrease viral replicative capacity. However, the intersubtype differences in the replicative consequences of such mutations have not been explored. In HIV-1 subtype B, the p24Gag M250I mutation is a rare variant (0.6%) that is enriched among elite controllers (7.2%) (P = 0.0005) and appears to be a rare escape variant selected by HLA-B58 supertype alleles (P < 0.01). In contrast, in subtype C, it is a relatively common minor polymorphic variant (10 to 15%) whose appearance is not associated with a particular HLA allele. Using site-directed mutant viruses, we demonstrate that M250I reduces in vitro viral replicative capacity in both subtype B and subtype C sequences. However, whereas in subtype C downstream compensatory mutations at p24Gag codons 252 and 260 reduce the adverse effects of M250I, fitness costs in subtype B appear difficult to restore. Indeed, patient-derived subtype B sequences harboring M250I exhibited in vitro replicative defects, while those from subtype C did not. The structural implications of M250I were predicted by protein modeling to be greater in subtype B versus C, providing a potential explanation for its lower frequency and enhanced replicative defects in subtype B. In addition to accounting for genetic differences between HIV-1 subtypes, the design of cytotoxic-T-lymphocyte-based vaccines may need to account for differential effects of host-driven viral evolution on viral fitness.
More than a million people acquire HIV infection annually. Pre-exposure prophylaxis (PrEP) using antiretrovirals is currently being investigated for HIV prevention. Oral and topical formulations of tenofovir have undergone pre-clinical and clinical testing to assess acceptability, safety and effectiveness in preventing HIV infection.
The tenofovir drug development pathway from compound discovery; pre-clinical animal model testing and human testing were reviewed for safety, tolerability, and efficacy. Tenofovir is well tolerated and safe when used both systemically or applied topically for HIV prevention. High drug concentrations at the site of HIV transmission and concomitant low systemic drug concentrations are achieved with vaginal application. Coitally applied gel may be the favoured prevention option for women compared to the tablets which may be more suitable for prevention in men and sero-discordant couples. However, recent contradictory effectiveness outcomes in women need to be better understood.
Emerging evidence has brought new hope that antiretrovirals can potentially change the course of the HIV epidemic when used as early treatment for prevention, as topical or oral PrEP. Although some trial results appear conflicting, behavioral factors, adherence to dosing, and pharmacokinetic properties of the different tenofovir formulations and dosing approaches offer plausible explanations for most of the variations in effectiveness observed in different trials.
ART; NtRTI; tenofovir; tenofovir disoproxil fumarate; HIV prevention; microbicide; PMPA – [9-R-(2-Phosphonomethoxypropyl) adenine]
The generation of polyfunctional CD8+ T cells, in response to vaccination or natural infection, has been associated with improved protective immunity. However, it is unclear whether the maintenance of polyfunctionality is related to particular cellular phenotypic characteristics. To determine whether the cytokine expression profile is linked to the memory differentiation stage, we analyzed the degree of polyfunctionality of HIV-specific CD8+ T cells within different memory subpopulations in 20 ART-naïve HIV-1 infected individuals at approximately 34 weeks post infection. These profiles were compared with CMV-specific CD8+ T cell responses in HIV-uninfected controls and in individuals chronically infected with HIV. Our results showed that the polyfunctional abilities of HIV-specific CD8+ T cells differed according to their memory phenotype. Early-differentiated cells (CD45RO+CD27+) exhibited a higher proportion of cells positive for three or four functions (p<0.001), and a lower proportion of mono-functional cells (p<0.001) compared to terminally-differentiated (CD45RO−CD27−) HIV-specific CD8+ T cells. The majority of terminally-differentiated HIV-specific CD8+ T cells were mono-functional (median 69% [IQR: 57–83]), producing predominantly CD107a or MIP1β. Moreover, proportions of HIV-specific mono-functional CD8+ T cells positively associated with proportions of terminally-differentiated HIV-specific CD8+ T cells (p=0.019, r=0.54). In contrast, CMV-specific CD8+ T cell polyfunctional capacities were similar across all memory subpopulations, with terminally- and early-differentiated cells endowed with comparable polyfunctionality. Overall, these data show that the polyfunctional abilities of HIV-specific CD8+ T cells are influenced by the stage of memory differentiation, which is not the case for CMV-specific responses.
In settings where multiple HIV prevention trials are conducted in close proximity, trial participants may attempt to enroll in more than one trial simultaneously. Co-enrollment impacts on participant’s safety and validity of trial results. We describe our experience, remedial action taken, inter-organizational collaboration and lessons learnt following the identification of co-enrolled participants.
Between February and April 2008, we identified 185 of the 398 enrolled participants as ineligible. In violation of the study protocol exclusion criteria, there was simultaneous enrollment in another HIV prevention trial (ineligible co-enrolled, n=135), and enrollment of women who had participated in a microbicide trial within the past 12 months (ineligible not co-enrolled, n=50). Following a complete audit of all enrolled participants, ineligible participants were discontinued via study exit visits from trial follow-up. Custom-designed education program on co-enrollment impacting on participants’ safety and validity of the trial results were implemented. Shared electronic database between research units were established to enable verification of each volunteer’s trial participation and to prevent future co-enrollments.
Interviews with ineligible enrolled women revealed that high-quality care; financial incentives; altruistic motives; preference for sex with gel; wanting to increase their likelihood of receiving active gel; perceived low risk of discovery and peer pressure as the reasons for their enrolment in the CAPRISA 004 trial.
Instituting education programs based on the reasons reported by women for seeking enrolment in more than one trial and using a shared central database system to identify co-enrollments have effectively prevented further co-enrollments.
HIV prevention trials; co-enrollment; young women
We have entered a new era in HIV prevention whereby priorities have expanded from biomedical discovery to include implementation, effectiveness, and the effect of combination prevention at the population level. However, gaps in knowledge and implementation challenges remain. In this Review we analyse trends in the rapidly changing landscape of HIV prevention, and chart a new path for HIV prevention research that focuses on the implementation of effective and efficient combination prevention strategies to turn the tide on the HIV pandemic.
Recent successes of antiretroviral pre-exposure prophylaxis (PrEP) in preventing HIV infection have raised questions whether further placebo controlled trials of new HIV-prevention technologies are ethically justifiable. A trial with active agent(s) in the comparator group can be designed either as a superiority or non-inferiority trial. In a non-inferiority trial the hypothesis tested is that the intervention is not inferior to, by a predefined clinically relevant amount, or at least as effective as, the comparator. Non-inferiority trials pose challenges in data interpretation.
Firstly it is possible to show equivalence of two non-effective interventions. If the active comparator intervention is ineffective, the new intervention would be shown to be non-inferior to this inactive intervention, while neither intervention is superior to placebo or no intervention. The second challenge is that any effect that dilutes the true efficacy of an intervention in a trial, such as non-adherence, loss to follow-up or protocol violations, makes it easier for the two interventions to be declared equivalent. Non-differential low adherence is unlikely to lead to the conclusion that an inferior intervention is non-inferior. However, differential adherence between study arms, which is more likely in non-blinded trials, is likely to bias the results and lead to incorrect conclusions.
Investigators conducting non-inferiority trials will have to pay special attention to supporting, measuring and maintaining high adherence. The goal in future non-inferiority trials should be to maintain similar levels of high adherence in all study arms, but at a minimum to reduce the likelihood of differential adherence across study arms.
HIV-1 accumulates mutations in and around reactive epitopes to escape recognition and killing by CD8+ T cells. Measurements of HIV-1 time to escape should therefore provide information on which parameters are most important for T cell–mediated in vivo control of HIV-1. Primary HIV-1–specific T cell responses were fully mapped in 17 individuals, and the time to virus escape, which ranged from days to years, was measured for each epitope. While higher magnitude of an individual T cell response was associated with more rapid escape, the most significant T cell measure was its relative immunodominance measured in acute infection. This identified subject-level or “vertical” immunodominance as the primary determinant of in vivo CD8+ T cell pressure in HIV-1 infection. Conversely, escape was slowed significantly by lower population variability, or entropy, of the epitope targeted. Immunodominance and epitope entropy combined to explain half of all the variability in time to escape. These data explain how CD8+ T cells can exert significant and sustained HIV-1 pressure even when escape is very slow and that within an individual, the impacts of other T cell factors on HIV-1 escape should be considered in the context of immunodominance.
Neutralizing antibodies are likely to play a crucial part in a preventative HIV-1 vaccine. Although efforts to elicit broadly cross-neutralizing (BCN) antibodies by vaccination have been unsuccessful1-3, a minority of individuals naturally develop these antibodies after many years of infection4-7. How such antibodies arise, and the role of viral evolution in shaping these responses, is unknown. Here we show, in two HIV-1–infected individuals who developed BCN antibodies targeting the glycan at Asn332 on the gp120 envelope, that this glycan was absent on the initial infecting virus. However, this BCN epitope evolved within 6 months, through immune escape from earlier strain-specific antibodies that resulted in a shift of a glycan to position 332. Both viruses that lacked the glycan at amino acid 332 were resistant to the Asn332-dependent BCN monoclonal antibody PGT128 (ref. 8), whereas escaped variants that acquired this glycan were sensitive. Analysis of large sequence and neutralization data sets showed the 332 glycan to be significantly underrepresented in transmitted subtype C viruses compared to chronic viruses, with the absence of this glycan corresponding with resistance to PGT128. These findings highlight the dynamic interplay between early antibodies and viral escape in driving the evolution of conserved BCN antibody epitopes.