关于HIV-1病毒储存库形成的表观遗传学机制及其清除手段的机理研究

HIV-1-infected individuals have to take the antiviral pills for lifelong because of the existence of HIV-1 reservoir. It is well accepted that the “shock and kill” strategy could be an efficient method to eradicate viral reservoir. A series of our previous works have identified multiple layers of mechanisms contributing to reservoir formation and developed the chimeric antigen receptor T-cell (CAR-T) technique to reactivate and eradicate HIV-1-latently infected cells. For this proposal, our preliminary results demonstrated that chromatin assembly factor-1 (CAF-1) could massively influence the epigenetic regulation and play a key role in the formation of viral reservoir. In addition, tannic acid, which was identified by high throughput screening, potently induced the degradation of CBX4, an E3 SUMO ligase which is a part of CAF-1 complex, and reactivated HIV-1 latency. We have also found that the expression of several inhibitory receptors, as well as CAF-1 and CBX4, significantly enhanced in HIV-1-specific CAR-T cells when the functional exhaustion occurred. Based upon these works, we will further determine the mechanisms of how CAF-1/CBX4 pathway contributes to HIV-1 reservoir formation and the mechanism for tannic acid to affect this pathway and potently reactivate HIV-1 latency, as well as the impact of CAF-1/CBX4 pathway to the function of CAR-T cells. Through these various approaches, we expect to further clarify the epigenetic regulations underlying reservoir formation, reactivation, and clearance, and develop new strategies for functional cure of AIDS.

HIV-1病毒储存库的存在，使得艾滋病人必须终身服用抗病毒药物。“震与杀（shock and kill）”策略是当前普遍认可的清除储存库方法。我们之前一系列工作发现储存库形成涉及多种机制及可被CAR-T细胞有效清除。我们的初步实验显示，染色质组装因子（CAF-1）可广泛影响表观遗传调控而导致储存库形成；成药库筛选出来的单宁酸特异地诱导CAF-1复合物中的E3 SUMO化连接酶CBX4降解并激活储存库；HIV-1特异性CAR-T细胞清除储存库的功能出现衰减时不仅若干抑制性受体，而且CAF-1和CBX4的表达也增加。鉴此，本项目将进一步研究CAF-1/CBX4通路导致病毒储存库的形成和单宁酸作用于此通路并激活潜伏病毒的机制，以及此通路的调控对CAR-T细胞功能出现衰减及清除储存库功能的影响。期望通过进一步阐明储存库形成、激活及清除的表观遗传学机制，为制定功能性治愈艾滋病的新策略提供理论基础。

HIV-1病毒储存库的存在，使得艾滋病人必须终身服用抗病毒药物。探索HIV-1病毒储存库建立和维持的表观遗传学机制、开发新型针对表观遗传学修饰因子的潜伏感染激活药物和CAR-T疗法是本项目的核心任务。我们发现（1）CAF-1通过形成多功能抑制性细胞内液-液相分离（LLPS）小体促进HIV-1的潜伏感染；（2）单宁酸（TA）可以高效激活HIV-1潜伏感染，及其下游靶点CBX4通过SUMO化EZH2帮助维持HIV-1潜伏；（3）TRIM28，USP49，PIWIL4作为被CAF-1调控及招募的宿主因子，也参与维持HIV-1的潜伏感染；（4）滤泡辅助性T细胞是一类新型的HIV-1储藏库，更容易受到HIV-1感染；（5）广谱中和抗体来源、插入三重抑制性受体shRNA的HIV-CAR-T细胞，可有效延后停药病人的病毒反弹时间，并减少体内的病毒储存库。以上成果发现紧密契合“功能性治愈HIV-1”的理念，进一步阐明HIV-1病毒储存库形成及维持的表观遗传学机制，从分子机制探索、激活药物筛选和CAR-T疗法设计等环节连贯性致力于从根本缩小HIV-1感染者病毒储存库。