Prime shock and kill for HIV erradication

描述：HIV的主要储存库由潜伏感染的静止记忆CD4 T细胞组成。新出现的信息表明，这些细胞本质上抵抗细胞凋亡有两个不同的原因：(i)慢性HIV感染的T细胞通过HIV蛋白引起多种细胞凋亡调节蛋白的表达改变而诱导细胞凋亡抵抗表型，以及（ii）静息记忆T细胞，由于作为先前免疫反应的历史档案而发展为静止和细胞凋亡抵抗状态，以保存记忆反应。目前“治愈”艾滋病毒的方法广泛涉及基因治疗、免疫治疗和病毒再激活。后一种策略涉及在药理学上重新激活HIV，期望重新激活病毒的CD4 T细胞将死于病毒蛋白表达的细胞毒性作用。迄今为止的研究已经证实，病毒再激活是可能的（例如，使用亚酰苯胺羟肟酸，SAHA）并且是安全的，但考虑到这些细胞对凋亡的内在抗性，重新激活病毒的细胞既不会在再激活后死亡，也不会被细胞毒性T淋巴细胞有效杀死，这并不奇怪。我们在含有潜伏HIV的静息记忆CD4 T细胞中表征了一些凋亡调节蛋白的表达，发现这些细胞具有低水平的促凋亡蛋白procaspase 8和高水平的抗凋亡蛋白Bcl2。我们认为，这种不平衡是潜伏HIV感染的CD4 T细胞在HIV再激活后不会死亡的原因，尽管它们在病毒再激活后在细胞内表达了有效的凋亡诱导蛋白——HIV Tat、nef、Vpr和蛋白酶。因此，被潜伏感染的细胞可以

DESCRIPTION: The primary reservoir of HIV consists of latently infected resting memory CD4 T cells. Emerging information indicates that these cells are intrinsically resistant to apoptosis fr two distinct reasons: (i) chronic HIV infection of T cells induces an apoptosis resistant phenotype by virtue of HIV proteins causing altered expression of a wide variety of apoptosis regulatory proteins, and (ii) resting memory T cells, by virtue of being an historical archive of prior immune responses developing a quiescent and apoptosis resistant state in order to preserve the memory responses. Current approaches to "cure" HIV broadly involve gene therapy, immune based therapy, and viral reactivation. The latter strategy involves reactivating HIV pharmacologically, with the expectation that CD4 T cells which reactivate virus will die from the cytotoxic effects of viral protein expression. Work to date has established that viral reactivation is possible (e.g., with suberoylanilide hydroxamic acid, SAHA) and safe, but given the intrinsic resistance of these cells to apoptosis, it is not surprising that the cells that reacivate virus neither die after reactivation, nor are they efficiently killed by cytotoxic T lymphocytes. e have characterized the expression of select apoptosis regulatory proteins in resting memory CD4 T cells which contain latent HIV, and found the cells to have low levels of the proapoptotic protein procaspase 8 and high levels of the antiapoptotic protein Bcl2. We propose that this imbalance is the reason why latently HIV infected CD4 T cells do not die after HIV reactivation, despite the fact that they express potent apoptosis-inducing proteins intracellularly - HIV Tat, nef, Vpr and protease after viral reactivation. Therefore, the cells that were latently infected do not die even after they are induced to express proapoptotic HIV proteins such as HIV protease. The overarching goal of the proposed study is to identify ways to alter latently infected HIV T cells such that they die in response to viral reactivation. In this application, we present three independent lines of evidence that this approach is justified and these cells can be altered in such a way that when HIV is reactivated, the cells will die. First using the Lewin model of HIV latency in primary CD4 T cells, we show that pharmacologically up-regulating the host protein procaspase 8, in resting memory CD4 T cells, allows these cells to be killed after viral reactivation, resulting in lower HIV replication (because infected cells are killed) and less integrated HIV copies. Next we summarize our previously published work that treatment of resting memory CD4 T cells from HIV infected patients with TRAIL agonists reduces that amount of replication competent HIV and the amount of HIV provirus, without deleterious effects on uninfected bystander cells. Finally, we present preliminary evidence that the first in class Bcl2 inhibitor, ABT-737, primes latently infected cells to undergo death upon HIV reactivation. These approaches specifically target HIV infected cells to die because, using this tactic, all cell will be primed to become apoptosis susceptible, however, only those cells which contain intracellular HIV proteins (the HIV infected cells) contain the apoptosis inducing stimulus. Having shown proof of concept for our "Prime Shock and Kill" model of HIV eradication, we now propose to adopt a high throughput screening approach to identify optimum pharmacologic methods of i) inducing apoptosis sensitivity, and then, ii) test these treatments in combination with stimuli that induce viral reactivation. This approach will then be tested for their ability to cause latently HIV infected T cell death using in vitro models of HIV latency and ex vivo testing of primary resting CD4 T cells from HIV-infected patients. Ultimately successful approaches will be fully vetted using the BLT mouse model of HIV infection.