Optimized Adaptation of Simian-tropic R5 HIV Clade C to Pig-tailed Macaques

描述（由申请人提供）：HIV进化枝C（HIV-C）导致全世界所有HIV/AIDS病例的56%以上，主要分布在撒哈拉以南非洲和印度。非人灵长类动物（NHP）模型将大大有利于预防策略的临床前开发，包括药物，杀微生物剂和疫苗。由于宿主限制因素，HIV-1仅在人类和黑猩猩中复制，而不在其他NHP物种中复制。编码HIV-1 vpu、达特、rev和env的猴-人免疫缺陷病毒（SHIV）的构建已经在恒河猴（RM）或猪尾猕猴（PMs~ Macaca nemestrina）中产生了致病模型。主要基于HIV-1但在猕猴中具有复制能力的灵长类慢病毒模型将极大地促进新治疗或预防策略的临床前开发。最近，已经认识到不同的宿主限制因子阻断HIV-1在RM细胞中的复制，即1）TRIM 5？，其靶向HIV-1衣壳蛋白并阻断早期进入后复制步骤，和2）APOBEC 3G（A3 G），一种不被HIV-1 Vif识别的胞苷脱氨酶，因此逃避这种病毒体相关蛋白的抑制。与RM不同，PM缺乏TRIM 5亚型，这可能解释了为什么PM在早期研究中支持未修饰的HIV-1的有限急性病毒血症。用SIVmac 239、SIVmne或HIV-2的vif替换HIV-1 vif已经产生了在PM中复制延长时间的嵌合体，然而，这些重组体基于X4-嗜性进化枝B克隆，并且不导致持续的病毒血症和疾病进展的迹象。我们已经产生了一个嗜猴的HIV-C克隆，称为stHIV-C，通过替换HIV vif与SIVmac 239 vif在感染性分子HIV-C克隆从赞比亚。嵌合体在M.奈美丝蒂娜在初步研究中，新的stHIV-C-vif 239嵌合体也诱导了两个PM中的病毒血症，然而，适应是不完全的。具体目标是：1.选择在CD 8+和CD 20+细胞的长期消耗下在PM中具有改进的复制适应性的stHIV-C子代~ 2。通过免疫活性PM连续传代进一步适应子代病毒，从最后一个受体中重新分离病毒，产生大量储备液，并在体外对其进行表征。3.进行直肠内滴定并评估病毒致病性4.评估stHIV-C在缺乏和存在适应性免疫以及疾病进展期间的分子演变。我们还将比较不同隔室（PBMC、淋巴结和脑）中的病毒进化。拟议的实验是重要的，因为他们专注于发展的NHP模式的艾滋病毒-C，这在发展中国家占主导地位，艾滋病流行病继续升级，从而将创建一个新的工具，临床前开发不同的战略，预防或根除。

DESCRIPTION (provided by applicant): HIV clade C (HIV-C) causes >56% of all cases of HIV/AIDS worldwide and predominates in sub-Saharan Africa and India. A non-human primate (NHP) model would greatly benefit the preclinical development of prevention strategies, including drugs, microbicides, and vaccines. Due to host restriction factors, HIV-1 replicates only in humans and chimpanzees, but not in other NHP species. The construction of simian-human immunodeficiency viruses (SHIVs) encoding HIV-1 vpu, tat, rev and env has yielded pathogenic models in rhesus macaques (RMs) or pig-tailed macaques (PMs~ Macaca nemestrina). A primate lentivirus model based mostly upon HIV-1 but replication-competent in macaques would greatly facilitate the preclinical development of new treatment or prevention strategies. Recently, different host restriction factors that block HIV-1 replication in RM cells have been recognized, namely 1) TRIM5?, which targets HIV-1 capsid proteins and blocks early post-entry replication steps, and 2) APOBEC3G (A3G), a cytidine deaminase that is not recognized by HIV-1 Vif and therefore escapes inhibition by this virion-associated protein. Unlike RMs, PMs lack a TRIM5 isoform, which may explain why PMs supported limited acute viremia of unmodified HIV-1 in earlier studies. Replacing HIV-1 vif with that of SIVmac239, SIVmne or HIV-2 has yielded chimeras that replicated for extended periods of time in PMs~ however, these recombinants were based upon the X4-tropic clade B clones, and did not result in persistent viremia and signs of disease progression. We have generated a simian-tropic HIV-C clone, termed stHIV-C, by replacing HIV vif with SIVmac239 vif in an infectious molecular HIV-C clone from Zambia. The chimera is replication-competent in peripheral blood mononuclear cells (PBMC) of M. nemestrina. In a pilot study, the new stHIV-C-vif239 chimera also induced viremia in two PMs~ however, adaptation is incomplete. The Specific Aims are to: 1. Select stHIV-C progeny with improved replication fitness in PMs under prolonged depletion of CD8+ and CD20+ cells~ 2. Further adapt progeny virus by serial passage through immunocompetent PMs, re-isolate virus from the last recipient, generate a large stock, and characterize it in vitro. 3. Perform an intrarectal titration and assess viral pathogenicity 4. Assess the molecular evolution of stHIV-C in the absence and presence of adaptive immunity and during disease progression. We will also compare viral evolution in different compartments (PBMC, lymph nodes, and brain). The proposed experiments are significant because they focus on the development of an NHP model of HIV-C, which predominates in developing countries where the AIDS epidemic continues to escalate and will thus create a new tool for the preclinical development of different strategies for prevention or eradication.