Role of Viral Reservoirs in the Pathogenesis of HIV Disease

研究表明，在感染的急性/早期阶段开始抗逆转录病毒治疗的艾滋病毒感染者中，有一小部分人能够在分析治疗中断（ATI）后较长时间内控制血浆病毒血症。更好地了解这些个体的病毒学控制的潜在机制可以为新疗法的开发和应用提供见解。我们对两名hiv感染者进行了一项长期研究，他们之前曾参加过包括ATI在内的随机对照治疗性疫苗试验的安慰剂组。两名研究参与者（04岁和30岁）在感染急性期开始抗逆转录病毒治疗，在开始ATI之前分别接受了6.7年和6.5年的临床有效抗逆转录病毒治疗。在ATI之后，参与者04在第56天经历了第一次实质性的血浆病毒反弹（26,967拷贝HIV RNA/ml），随后持续了500多天的持续自发病毒学控制。此后，他的血浆病毒血症在第581天和第875天分别短暂回升至778和1784拷贝/ml。为了深入了解他的连续血浆病毒反弹的基因组性质，我们使用血浆中3-一半HIV RNA的单基因组扩增（SGA）序列进行了系统发育分析。第56天血浆HIV反弹序列相对均匀。然而，第581天第二次血浆病毒反弹的序列与第56天的序列在很大程度上不同，这表明病毒持续进化和/或其他预先存在的病毒库重新激活。参与者04在1250天之前没有检测到抗逆转录病毒药物，包括紧接着三次主要血浆病毒反弹的时间点。然而，药物检测显示参与者04在大约1250天开始了未公开的次优抗逆转录病毒治疗。相比之下，参与者30在整个随访期间没有检测到抗逆转录病毒药物。尽管在整个随访期间几乎完全抑制病毒学，参与者30在1434天经历了显著水平的血浆病毒反弹。系统发育显示，1434天血浆HIV反弹的HIV环境序列与之前的时间点不同，并且与不同的HIV亚型B株一样存在差异，强烈提示参与者30发生了HIV重复感染。为了研究HIV特异性CD8+ T细胞在没有ART的情况下抑制HIV的潜在作用，我们使用跨越整个HIV Gag蛋白的重叠肽池进行了纵向细胞内细胞因子染色试验。与参与者30相比，参与者04中多功能HIV gag特异性CD8+ T细胞的频率始终较高。最后，我们调查了在没有抗逆转录病毒治疗的情况下，研究参与者血浆中的中和抗体是否可能导致病毒学抑制。对于参与者04，热灭活血浆显示出低血浆50%中和效价（ID50）（<1:30稀释）。与此形成鲜明对比的是，参与者30第117天的血浆显示出强大的igg介导的中和活性（ID50为1:16 07稀释），可以抵抗自身的当代感染分离物，这表明参与者30在重复感染之前在体内实现接近完全病毒学抑制的潜在机制之一可能与中和HIV抗体有关。总的来说，我们的数据为治疗后中断控制的不同机制提供了见解，并强调了在ATI阶段频繁监测未公开使用抗逆转录病毒治疗和重复感染的重要性。

It has been shown that a small percentage of HIV-infected individuals in whom ART was initiated during the acute/early phase of infection can control plasma viremia for extended periods following analytical treatment interruption (ATI). A better understanding of the underlying mechanism(s) of virologic control in these individuals could provide insight into the development and application of novel therapies. We conducted a long-term study of two HIV-infected individuals who had previously participated in the placebo arm of a randomized, controlled therapeutic vaccine trial that included ATI. Two study participants (04 and 30) initiated ART during the acute phase of their infections and had been receiving clinically effective ART for 6.7 and 6.5 years, respectively, prior to initiating ATI. Following ATI, Participant 04 experienced a first substantial plasma viral rebound on day 56 (26,967 copies of HIV RNA/ml), followed by sustained spontaneous virologic control that lasted over 500 days. Thereafter, his plasma viremia rebounded briefly to 778 and 1,784 copies/ml on days 581 and 875, respectively. To gain insight into the genomic nature of his sequential plasma viral rebounds, we conducted phylogenetic analyses using sequences derived from single genome amplification (SGA) of the 3-half of HIV RNA in the plasma. The sequences of rebounding plasma HIV on day 56 were relatively homogeneous. However, the sequences derived from the second plasma viral rebound on day 581 were largely distinct from those on day 56, suggesting continuous viral evolution and/or reactivation of other pre-existing viral reservoirs. There were no antiretroviral drugs detected in Participant 04 prior to day 1,250, including the time points immediately following the three major episodes of plasma viral rebound. However, the drug testing revealed that Participant 04 had initiated undisclosed, suboptimal ART at approximately day 1,250. In contrast, there were no antiretroviral drugs detected in Participant 30 for the entire follow-up period. Despite near complete virologic suppression throughout the entire follow-up period, Participant 30 experienced a significant level of plasma viral rebound on day 1,434. Phylogenetic showed that the HIV env sequences of rebounding plasma HIV on day 1,434 did not resemble those from previous time points and were as divergent as different HIV subtype B strains, strongly suggesting that HIV superinfection had occurred in Participant 30. To investigate the potential role of HIV-specific CD8+ T cells in suppressing HIV in the absence of ART, we performed longitudinal intracellular cytokine staining assays using a pool of overlapping peptides that spanned the entire HIV Gag protein. Frequencies of polyfunctional HIV Gag-specific CD8+ T cells were consistently higher in Participant 04 compared to those of Participant 30. Finally, we investigated whether neutralizing antibodies in the plasma of the study participants potentially contributed to virologic suppression in the absence of ART. For Participant 04, heat-inactivated plasma exhibited low plasma 50% neutralization titer (ID50) (<1:30 dilution). In striking contrast, day 117 plasma of Participant 30 showed strong IgG-mediated neutralization activity (ID50 of 1:1,607 dilution) against autologous contemporary infectious isolates, suggesting that one of the potential mechanisms by which Participant 30 achieved near complete virologic suppression in vivo, prior to superinfection, may have involved neutralizing antibodies against HIV. Collectively, our data provide insight into distinct mechanisms of post-treatment interruption control and highlight the importance of frequent monitoring of undisclosed use of ART and superinfection during the ATI phase. Between September 2018 and January 2021, we conducted a phase 1 clinical trial to assess the safety, tolerability, and efficacy of the combination of bNAbs 3BNC117 and 10-1074 in HIV-infected individuals. This trial consisted of two components: 1) a randomized, double-blind, placebo-controlled study involving 14 participants in whom ART was initiated during the acute/early phase of infection and who subsequently underwent ATI shortly after receiving the first infusion of bNAbs or placebo (Group 1) and 2) an open-label study involving 5 viremic controllers who were ART-nave and had baseline plasma viremia between 200-5,000 copies/ml (Group 2). Study participants received 4-8 (median 8) 3BNC117 (30mg/kg) and 10-1074 (30mg/kg) infusions. After receiving the first infusion of 3BNC117 and 10-1074 or placebo, the study participants in Group 1 underwent ATI and plasma viremia and CD4+ T cell counts were measured every two weeks. Six of the 7 study participants in the placebo arm experienced plasma viral rebound and met criteria to restart ART prior to study week 28 compared with none of the 7 participants in the treatment arm. The median duration off ART was 39.6 weeks and 9.4 weeks for the Group 1 bNAb and placebo participants, respectively (P = 0.001).The median duration of plasma viremia suppression at <200 copies/ml in Group 1 was 33.4 weeks and 3.4 weeks in the bNAb and placebo arms, respectively (P = 0.002). In Group 2, 2 of the 5 study participants whose baseline infectious HIV was sensitive to both antibodies maintained complete suppression of plasma viremia for an average of 41.7 weeks. The levels of CD4+ T cells carrying total HIV DNA, cell-associated HIV RNA, intact proviral DNA, and replication-competent virus were longitudinally monitored in Group 1 bNAb participants. No statistical significance was reached when the levels of HIV reservoirs were compared between the two time points (Weeks 0 and 24), suggesting that the combination bNAbs did not have a significant impact on the persistent HIV reservoir in our study participants. We considered whether immunologic abnormalities (such as inflammation and exhaustion) occurred in study participants who received infusions of combination bNAbs and maintained extended periods of virologic suppression. We used high-dimensional flow cytometry to longitudinally examine a wide array of biomarkers and immune parameters in peripheral blood mononuclear cells (PBMCs) of our study participants. Our data suggest that significant immunologic abnormalities did not arise during prolonged ATI in study participants in whom the bNAb-mediated virologic suppression was achieved. We longitudinally examined HIV-specific CD8+ T cells in our study participants by performing intracellular cytokine staining following stimulation with a pool of overlapping HIV Gag peptides. The level of polyfunctional HIV Gag-specific CD8+ T cells remained unchanged in Group 1 bNAb and Group 2 participants. Collectively, our data demonstrate that the combination therapy with 3BNC117 and 10-1074 is highly effective in suppressing HIV in the absence of ART for extended periods provided that antibody-resistant virus is not present at baseline and offer guidance for future clinical trials involving next-generation antibodies with long half-lives.