The development of latency reversing agents that potently reactivate HIV without inducing global T cell activation would benefit the field of HIV reservoir research and could pave the way to a functional cure. Here, we explore the reactivation capacity of a lipid nanoparticle containing Tat mRNA (Tat-LNP) in CD4 T cells from people living with HIV undergoing antiretroviral therapy (ART). When combined with panobinostat, Tat-LNP induces latency reversal in a significantly higher proportion of latently infected cells compared to PMA/ionomycin (≈ 4-fold higher). We demonstrate that Tat-LNP does not alter the transcriptome of CD4 T cells, enabling the characterization of latently infected cells in their near-native state. Upon latency reversal, we identify transcriptomic differences between infected cells carrying an inducible provirus and non-infected cells (e.g. LINC02964, GZMA, CCL5). We confirm the transcriptomic differences at the protein level and provide evidence that the long non-coding RNA LINC02964 plays a role in active HIV infection. Furthermore, p24+ cells exhibit heightened PI3K/Akt signaling, along with downregulation of protein translation, suggesting that HIV-infected cells display distinct signatures facilitating their long-term persistence. Tat-LNP represents a valuable research tool for in vitro reservoir studies as it greatly facilitates the in-depth characterization of HIV reservoir cells’ transcriptome and proteome profiles.

开发潜伏期逆转剂可以有效地重新激活 HIV，而不诱导整体 T 细胞激活，这将有利于 HIV 储存库研究领域，并可能为功能性治愈铺平道路。在这里，我们探索了含有 Tat mRNA (Tat-LNP) 的脂质纳米颗粒对接受抗逆转录病毒治疗 (ART) 的 HIV 感染者的 CD4 T 细胞的再激活能力。当与帕比司他联合使用时，与 PMA/离子霉素相比，Tat-LNP 在显着更高比例的潜伏感染细胞中诱导潜伏期逆转（约高 4 倍）。我们证明 Tat-LNP 不会改变 CD4 T 细胞的转录组，从而能够表征处于接近天然状态的潜伏感染细胞。潜伏期逆转后，我们鉴定了携带可诱导原病毒的感染细胞和未感染细胞（例如LINC02964 、 GZMA 、 CCL5 ）之间的转录组差异。我们确认了蛋白质水平上的转录组差异，并提供了长非编码 RNA LINC02964在活动性 HIV 感染中发挥作用的证据。此外，p24+细胞表现出增强的PI3K/Akt信号传导，以及蛋白质翻译的下调，表明HIV感染的细胞表现出独特的特征，有利于其长期持续存在。 Tat-LNP 是体外病毒库研究的一种有价值的研究工具，因为它极大地促进了 HIV 病毒库细胞转录组和蛋白质组谱的深入表征。