Eradication of brain/CNS HIV reservoirs via anti-HIV gene-modified macrophages

通过抗 HIV 基因修饰的巨噬细胞根除大脑/中枢神经系统 HIV 储存库

• 批准号: 10589937
• 负责人: Masakazu Kamata
• 金额: $ 18.56万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-01 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10589937
• 关键词: Adoptive Immunotherapy; Animal Model; Anti-Retroviral Agents; Antibodies; Antigens; Autologous; Brain; CAR T cell therapy; CCR5 gene; CD19 gene; CRISPR/Cas technology; CXCR4 gene; Cell Lineage; Cells; Cellular Membrane; Central Nervous System; Central Nervous System Lymphoma; Cerebrospinal Fluid; Clinical; Clinical Trials; Combined Modality Therapy; Complex; Cytotoxic T-Lymphocytes; DNA; Data; Drug Carriers; Effector Cell; Ensure; Epitopes; Excretory function; Exhibits; Fumarates; Gene Modified; Genes; HIV; HIV Infections; HIV envelope protein; Immune system; Immunotherapeutic agent; Individual; Infection; Inflammation; Location; Macrophage; Maintenance; Mediating; Mediator; Microglia; Modeling; Myelogenous; Myeloid Cells; Natural Killer Cells; Neurons; Nevirapine; Passive Immunotherapy; Penetration; Performance; Peripheral; Phagocytosis; Pharmaceutical Preparations; Pharmacotherapy; Plasma; Production; Risk; Shock; Source; System; T-Lymphocyte; Technology; Tenofovir; Testing; Tissue Extracts; Toxic effect; Transcript; Variant; Viral Load result; Viremia; Virus; Work; antibody-dependent cell cytotoxicity; antibody-dependent cellular phagocytosis; antiretroviral therapy; blood-brain barrier penetration; cell type; chimeric antigen receptor; cytokine release syndrome; cytotoxicity; digital; efficacy evaluation; genome editing; humanized mouse; in vivo; intravenous injection; migration; monocyte; nanocapsule; neuroinflammation; neurotoxicity; neutralizing antibody; novel; novel strategies; purge; recruit; small hairpin RNA; small molecule inhibitor; tool; vector

PROJECT SUMMARY Combination antiretroviral therapy (cART) is highly effective, but it is weakened in the central nervous system (CNS) due to poor drug penetration. In addition, cells infected by HIV in the CNS - termed “CNS reservoirs” - are mainly constituted by myeloid lineage cells, such as microglia cells and macrophages, thus anti- HIV strategies employed are needed to be adapted to these cell types. Various approaches have been applied for controlling viral load in the CNS - for example, T cells expressing anti-HIV chimeric antigen receptor (CAR- T) or broadly neutralizing antibodies (bNAb) – and these approaches effectively target HIV or HIV-infected cells systemically but are ineffective against the CNS reservoirs since cytotoxic-T cells cannot attack myeloid-lineage cells well and antibody penetration into the CNS is extremely low. As such, there is no practical approach successful enough to eliminate these reservoirs safely. We here intend to purge the CNS reservoirs via using genetically modified macrophages expressing anti- HIV CARs (CAR-M) (Aim 1) and multidirectional bNAb (bNAb-M) (Aim 2). Both will be expressed in freshly isolated monocytes from blood sources as a carrier vehicle to the CNS and an effector cell. Importantly, monocytes will be recruited to the CNS when neuroinflammation occurs; HIV infection itself as well as some antiretroviral drugs such as Tenofovir disoproxil fumarate (TDF) or nevirapine (NVP), are known mediators of neuronal inflammation. To overcome the issue of HIV quasispecies, we will use each tool to target several different epitopes, i.e., express 2 or more anti-HIV CARs in the same cells (Aim 1) or multidirectional bNAbs in one molecule which target 3 independent epitopes on the HIV envelope protein (Aim 2). In addition, to protect the gene-modified monocytes, we will simultaneously express two anti-HIV genes with these immunotherapeutic tools; a) C46 that interferes fusion between HIV envelope and cellular membranes, thus blocking HIV infections and b) shRNA against HIV LTR (sh516) that destructs all HIV transcripts including the sequence derived from HIV LTR. Altogether, our approach will ensure CNS delivery of monocytes following gene modification to express multiple anti-HIV CARs or multidirectional bNAbs whilst protecting from HIV infections and/or productions. The monocytes will then differentiate into macrophages in the CNS and attack the infected cells via anti-HIV CAR (CAR-M) or secreting multidirectional bNAbs (bNAb-M) inducing antibody-dependent cytotoxicity or phagocytosis (ADCC/ADCP) against the infected cells. In case of the bNAb-M approach, secreted bNAbs will also work on adjacent myeloid cells which support both ADCC/ADCP. In addition, the bNAbs will neutralize new HIV infection. As such, our approach is highly unique, and, if successful, will open a new avenue to defeat HIV reservoirs in the CNS.

项目摘要 组合抗逆转录病毒疗法（CART）非常有效，但中枢神经弱 由于药物渗透不良而导致的系统（CNS）。此外，CNS中被HIV感染的细胞称为“ CNS” 水库” - 主要由髓样谱系细胞（例如小胶质细胞和巨噬细胞）组织，因此 艾滋病毒策略需要适应这些细胞类型。已经采用了各种方法 为了控制中枢神经系统的病毒载量 - 例如，表达抗HIV嵌合抗原受体的T细胞（CAR- t）或广泛中和抗体（BNAB） - 这些方法有效地靶向HIV或HIV感染的细胞 在系统上，但对中枢神经系统储层无效，因为细胞毒性-T细胞无法攻击髓样细胞 细胞良好，抗体渗透到中枢神经系统中极低。因此，没有实用的方法 成功地消除了这些水库。 我们在这里打算通过使用表达抗 - HIV汽车（CAR-M）（AIM 1）和多向BNAB（BNAB-M）（AIM 2）。两者都将以新鲜的方式表达 从血液源分离的单核细胞作为CNS的载体和效应细胞。重要的是， 当神经炎症发生时，单核细胞将被招募到中枢神经系统。艾滋病毒感染本身以及一些 抗逆转录病毒药物，例如替诺福韦毒素（TDF）或奈韦拉平（NVP），是已知的介体 神经元炎症。为了克服艾滋病毒准特质问题，我们将使用每个工具来定位几个 不同的表位，即在同一细胞中表达2个或更多的抗HIV汽车（AIM 1）或多向BNABS 一个针对HIV包膜蛋白上的3个独立表位的分子（AIM 2）。另外，保护 基因修饰的单核细胞，我们将仅以这些免疫治疗性表达两个抗HIV基因 工具; A）C46干扰HIV包膜和细胞膜之间的融合，从而阻断HIV感染 b）针对HIV LTR（SH516）的shRNA破坏了所有HIV转录本，包括源自 HIV LTR。总之，我们的方法将确保CNS在基因修饰后的单核细胞递送以表达 在保护HIV感染和/或生产的同时，多种抗HIV汽车或多向BNAB。这 然后，单核细胞将分化为中枢神经系统中的巨噬细胞，并通过抗HIV CAR攻击感染的细胞 （CAR-M）或分泌多向BNAB（BNAB-M）诱导抗体依赖性细胞毒性或 针对感染细胞的吞噬作用（ADCC/ADCP）。如果采用BNAB-M方法，则分泌的BNAB将 还可以在支持ADCC/ADCP的相邻髓样细胞上工作。此外，bnabs将中和新 艾滋病毒感染。因此，我们的方法是高度独特的，如果成功的话，将为击败艾滋病毒的新途径 中枢神经系统中的水库。