Modeling monocyte and macrophage based gene therapy for neuroAIDS

描述（由申请人提供）：人类免疫缺陷病毒1型（HIV）感染脑巨噬细胞和小胶质细胞并导致HIV相关痴呆（HAD），这是一种中枢神经系统（CNS）的原发性疾病，影响约20%的HIV感染者。目前对HAD的治疗受到许多抗逆转录病毒药物通过血脑屏障（BBB）效率低下的阻碍。因此，需要新的HAD疗法。众所周知，在正常和某些情况下，循环血液中单核细胞来源的巨噬细胞（MDM）都可以穿过血脑屏障进入中枢神经系统；一些随后成熟为长期存在于脑组织的巨噬细胞和小胶质细胞。本研究的假设是，有可能利用血液MDM的自然归巢/迁移特性，以非侵入性和非手术方式将神经保护因子传递到中枢神经系统。我们实验室和其他实验室之前的研究表明，MDM（人类和小鼠）可以在体外使用有缺陷的慢病毒载体（DLV）进行基因修饰，而不会对其生物学特性产生不利影响。此外，我们已经证明，经DLV基因修饰的原代小鼠血液单核细胞和骨髓来源的巨噬细胞可以进入大脑，并且CNS对这些细胞的摄取效率可以通过短暂破坏血脑屏障而提高。我们最近发现，从dlv转导的MDM中分泌的抗hiv - tat单链抗体（scFv）和可溶性肿瘤坏死因子受体（sTNFR）可以有效阻断条件培养基对HIV-1感染细胞或相应重组蛋白的神经毒性作用。在本项目中，我们将建立一种高效（>50%）、稳定的用编码可分泌scFv和sTNFR的载体转导小鼠原代MDM的基因转移方法。然后，我们将进行研究，以确定是否可以通过暂时破坏血脑屏障来增强这些转基因细胞进入小鼠大脑的吸收，我们将评估外周输注转基因MDM后cns内基因表达的时间过程和稳定性。最后，我们将确定稳定表达TNFR或抗hiv - tat scFv的原代小鼠MDM是否可以改善两种经过充分研究的神经艾滋病小鼠模型系统的疾病进展。这项研究的成功完成有望为对抗神经艾滋病和其他神经系统疾病的新方法提供重要的见解。公共卫生相关性：这项研究的成功完成有望为对抗神经艾滋病和其他神经疾病的新方法提供重要的见解。

DESCRIPTION (provided by applicant): Human immunodeficiency virus type 1 (HIV) infects brain macrophages and microglia and causes HIV- associated dementia (HAD), a primary disorder of the central nervous system (CNS) that affects about 20% of HIV-infected individuals. Current treatment of HAD is hampered by the poor efficiency of many antiretroviral drugs to cross the blood-brain barrier (BBB). Hence, new therapies for HAD are needed. Circulating blood monocyte-derived macrophages (MDM) are known to migrate across the BBB and to enter the CNS under both normal and certain circumstances; some subsequently maturing into long-lived tissue-resident brain macrophages and microglia. The hypothesis of this research is that it may be possible to exploit the natural homing/migratory properties of blood MDM in order to deliver neuroprotective factors into the CNS, in a non- invasive and non-surgical manner. Previous studies from our laboratory and others, have shown that MDM (human and mouse) can be genetically modified in vitro using defective lentiviral vectors (DLV) without adversely affecting their biological properties. Furthermore, we have shown that primary mouse blood monocytes and bone marrow-derived macrophages genetically modified by DLV can enter the brain, and that the efficiency of CNS uptake of these cells can be enhanced through transient disruption of the BBB. We have recently shown that secreted anti-HIV-Tat single chain antibodies (scFv) and soluble tumor necrosis factor receptor (sTNFR) from DLV-transduced MDM can effectively block the neurotoxic effects of conditioned medium from HIV-1 infected cells or respective recombinant proteins. In this project, we will establish a gene transfer method for high efficiency (>50%), stable transduction of primary mouse MDM with vectors that encode secretable scFv and sTNFR. We will then conduct studies to determine whether uptake of these genetically modified cells into the mouse brain can be enhanced using temporary disruption of the BBB, and we will evaluate the time course and stability of intra-CNS gene expression following peripheral infusion of the genetically-modified MDM. Finally, we will determine whether primary mouse MDM that stably express TNFR or anti-HIV-Tat scFv can ameliorate disease progression in two well-studied murine model systems for neuroAIDS. Successful completion of this study is expected to provide significant insight into new approaches for combating neuroAIDS and other neurologic disorders. PUBLIC HEALTH RELEVANCE: Successful completion of this study is expected to provide significant insight into new approaches for combating neuroAIDS and other neurologic disorders.