HIV-1 EnvLRS: a scalable, sequence-based assay for in-depth assessment of HIV-1 bNAb resistance

项目总结/摘要 人类免疫缺陷病毒1型（HIV-1）是一种逆转录病毒，感染免疫系统的CD 4 + T细胞， 系统如果不及时治疗，HIV-1感染者（PLWH）将发展为艾滋病，并可能最终死亡。 结果.联合抗逆转录病毒疗法（ART）与小分子药物是非常有效的停止 HIV-1在受感染个体中的复制，但需要每日给药，通常具有相当大的副作用。 重要的是，尽管这种方法成功地将HIV-1复制抑制到临床检测不到的水平， 抗逆转录病毒疗法不能治愈疾病。这是由于HIV-1一直处于沉默或潜伏状态 在长寿命的CD 4 + T细胞中以极低的频率。这些潜伏感染的细胞不是靶细胞， 目前的小分子抗逆转录病毒疗法。因此，艾滋病毒携带者必须终身接受抗逆转录病毒治疗。 靶向HIV-1 Env中关键表位的广泛中和抗体（HIV-1 bNAb）目前正在开发中。 正在开发作为消除潜伏HIV-1的潜在方法和/或作为小分子药物的替代品， 条与传统的小分子ART相比，HIV-1 bNAb提供了几个优势，包括潜在的 长效制剂，减少副作用，并随着时间的推移消除潜伏感染细胞的潜力。 然而，在治疗和治愈中实施HIV-1 bNAb的主要挑战是预先存在的 bNAb靶向表位的变异或抗性。需要可扩展的临床试验来确定（1） 将接受HIV-1 bNAb的人是否具有预先存在的耐药性，以及（2）个性化HIV-1 bNAb 组合到每个人。为了解决这一关键的未满足的需求，AccelevirDx正在开发HIV-1 EnvLRS检测是第一个可扩展的基于序列的检测，用于评估HIV-1 bNAb耐药性并预测 通过深入的env序列分析确定抗体效力。从广义上讲，该提案旨在（1）在分析上限定 AccelevirDx的新型专有HiFi-dePCR方法用于HIV-1 EnvLRS的env扩增，（2） 确定来自PLWH的样品的HIV-1 EnvLRS测定的再现性，和（3）将测定应用于 最近在PLWH中完成了HIV-1 bNAb VRC 01的ACTG A5340临床试验， 性能和实用性。

PROJECT SUMMARY/ABSTRACT Human immunodeficiency virus type-1 (HIV-1) is a retrovirus that infects CD4+ T cells of the immune system. If left untreated, people living with HIV-1 (PLWH) will progress to AIDS and may ultimately die as a result. Combination antiretroviral therapy (ART) with small-molecule drugs is extremely effective at stopping the replication of HIV-1 in infected individuals but requires daily dosing often with considerable side effects. Importantly, despite the success of this approach at suppressing HIV-1 replication to clinically undetectable levels, antiretroviral therapy is not curative. This is due to the persistence of HIV-1 in a silent, or latent, state within long-lived CD4+ T cells at extremely low frequencies. These latently infected cells are not targeted by current small-molecule ART regimens. As a result, PLWH must remain on lifelong antiretroviral therapy. Broadly neutralizing antibodies targeting critical epitopes in HIV-1 Env (HIV-1 bNAbs) are currently being developed as a potential approach to eliminate latent HIV-1 and/or as an alternative to small-molecule ART. HIV-1 bNAbs offer several advantages over traditional small-molecule ART, including the potential for long-acting formulations, reduced side effects, and the potential to eliminate latently infected cells over time. However, a major challenge to the implementation of HIV-1 bNAbs in treatment and cure is pre-existing variation or resistance in the bNAb-targeted epitopes. Scalable clinical tests are needed to determine (1) whether people who will receive HIV-1 bNAbs have pre-existing resistance, and (2) personalize HIV-1 bNAb combinations to each individual. To address this critical unmet need, AccelevirDx is developing the HIV-1 EnvLRS assay as the first scalable sequence-based test to assess HIV-1 bNAb resistance and predict antibody efficacy by in-depth env sequence analysis. Broadly, this proposal aims to (1) analytically qualify AccelevirDx’s novel, proprietary HiFi-dePCR approach for env amplification underlying HIV-1 EnvLRS, (2) determine the reproducibility of the HIV-1 EnvLRS assay of samples from PLWH, and (3) apply the assay to a recently completed ACTG A5340 clinical trial of the HIV-1 bNAb VRC01 in PLWH to assess assay performance and utility.