REVERSE PSEUDOTYPE HIV 1 TARGETING OF NEURAL CELLS

HIV-1感染表达CD4的细胞需要包膜、CD4和一个或多个趋化因子受体(七个跨膜G蛋白偶联受体)之间的相互作用。感染需要包膜与CD4和趋化因子受体结合，然后病毒与细胞膜融合。在大脑中，感染发生在巨噬细胞和小胶质细胞中，它们表达CCR5、CXCR4和CCR3趋化因子受体。我们已经证明，当CD4和趋化因子受体在病毒粒子上表达，并且包膜在靶细胞上表达时，包膜与这些受体发生有效的相互作用。我们发现，根据典型的HIV-1毒株的包膜利用模式，这种CD4/趋化因子受体假病毒粒子特异性地针对HIV-1感染的细胞。此外，我们还使用基于HIV的载体假型在人类星形胶质细胞和神经元中表达转基因。我们的假设是，基于HIV-1的载体作为趋化因子受体/CD4伪病毒，可以靶向HIV-1感染的细胞，传递抗病毒和毒性基因，以限制感染并潜在地消除中枢神经系统(CNS)中受感染的细胞库，并且基于HIV-1的载体还允许引入神经保护基因来限制HIV-1诱导的神经毒性。我们的目标是开发基于HIV-1的伪型策略，以消除HIV-1的中枢神经系统储存库，并保护神经元免受HIV-1感染的影响。为此，我们将：1)确定趋化因子受体/CD4假型(反向假型)在转导和杀伤HIV-1感染的巨噬细胞方面的效率。我们将利用表达促凋亡基因的反向伪型来诱导凋亡基因的构建，以诱导具有不同趋化因子受体利用模式的原代CNS HIV-1分离株感染的巨噬细胞的凋亡；2)确定基于HIV-1的伪型转导人类神经元、星形胶质细胞和巨噬细胞的能力，以建立针对HIV-1的神经保护策略。我们将利用原代CNS细胞的假型转导来测试抗凋亡基因作为神经保护剂的作用；以及3)确定从特异性AIMS 1和2中选择的假型在体内转导原代CNS细胞和在体内靶向HIV-1感染的巨噬细胞的能力。我们将与我们的神经病理学核心和克尔博士合作，利用SCID-HU小鼠建立HIV-1脑感染的模型，以研究体内的载体转导。基于HIV的载体在有丝分裂后细胞靶向和表达的能力为治疗和预防艾滋病痴呆提供了新的方法。

HIV-1 infection of CD4-expressing cells requires interaction between envelope, CD4 and one or more chemokine receptors (seven transmembrane-spanning G protein-coupled receptors). Infection requires envelope binding to both CD4 and chemokine receptor, followed by fusion of virus and cell membrane. In the brain, infection occurs in macrophages and microglia, which express CCR5, CXCR4, and CCR3chemokine receptors. We have demonstrated that effective interaction of envelope with such receptors occurs when CD4 and chemokine receptor are expressed on the virion and envelop is expressed on the target cell. We showed that such CD4/chemokine receptor pseudovirions specifically target HIV-1 infected cells, as predicted by envelope utilization patterns by prototypic HIV-1 strains. In addition, we have used HIV-based vector pseudotypes to express transgenes in human astrocytes and neurons. Our hypothesis is that HIV-1 based vectors utilized as chemokine receptor/CD4 pseudovirus can target HIV-1- infected cells to deliver antiviral and toxicity genes to limit infection and potentially eliminate infected cellular reservoirs in the central nervous system (CNS), and that HIV-1 based vectors also allow introduction of neuroprotective genes to limit HIV-1- induced neurotoxicity. Our goal is to develop HIV-1-based pseudotype strategies to eliminate CNS reservoirs for HIV-1 and to protect neurons form effects of HIV-1 infection. To do this we will: 1) Determine the efficiency of chemokine receptor/CD4 pseudotypes (reverse pseudotypes) in transducing and killing HIV-1 infected macrophages. We will utilize reverse pseudotypes expressing a pro-apoptosis genes construct to induce apoptosis gene construct to induce apoptosis in macrophages infected with primary CNS HIV-1 isolates with different chemokine receptor utilization patterns; 2) Define the ability of HIV-1- based pseudotypes to transduce human neurons, astrocytes and macrophages to establish neuroprotective strategies against HIV-1. We will utilize pseudotype transduction of primary CNS cells to test anti-apoptosis genes as neuroprotectants; and 3) Determine the ability of pseudotypes selected from specific aims 1 & 2 to transduce primary CNS cells in vivo and to target HIV-1 infected macrophages in vivo. In collaboration with our Neuropathology core and Dr. Kerr, we will utilize a SCID-hu mouse to establish a model for HIV-1 infection of the brain to study vector transduction in vivo. The ability of HIV-based vectors to target and express in post-mitotic cells suggests new approaches for the treatment and prevention of AIDS dementia.