B-cell-specific transduction for anti-HIV antibody and B-cell receptor expression

用于抗 HIV 抗体和 B 细胞受体表达的 B 细胞特异性转导

• 批准号: 10549760
• 负责人: Koki Morizono
• 金额: $ 39万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-02-15 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10549760
• 关键词: Amino Acid Sequence; Animal Model; Antibodies; Antibody Formation; Antibody Therapy; Antigens; Applications Grants; Autoantigens; Autoimmune Diseases; B cell differentiation; B-Cell Antigen Receptor; B-Cell Development; B-Cell Receptor Binding; B-Lymphocytes; BLT mice; Binding; Cell Differentiation process; Cells; Chromosomes; Communicable Diseases; Development; Differentiation and Growth; Experimental Designs; Factor IX; Fc domain; Gene Transduction Agent; Genes; Genetic Recombination; Genetic Transcription; Growth; HIV Antibodies; HIV therapy; HIV-1; Hemophilia A; Human; Immune; Immune response; Immunocompetent; Immunoglobulin G; Infection; Intravenous; Lentivirus Vector; Ligands; Longevity; Membrane; Memory B-Lymphocyte; Methods; Mus; Mutation; Pathology; Physiological; Plasma Cells; Positioning Attribute; Proliferating; Proteins; Public Health; Reagent; Research; Serum; Signal Transduction; Specificity; Structural Protein; System; Therapeutic; Therapeutic Effect; Therapeutic antibodies; Time; Tissues; Transgenes; Viral Proteins; Viral Vector; Virus; Virus Replication; cell growth; cell type; cellular transduction; cost; env Gene Products; gene product; gene therapy; immunoreaction; in vivo; in vivo imaging; inhibitor; mouse model; natural antibodies; neutralizing antibody; novel; promoter; receptor expression; therapeutic gene; therapeutically effective; tool; transgene expression; transmission process; vector

PROJECT SUMMARY Neutralizing antibodies against HIV-1 are very potent in suppressing HIV-1 replication. However, the serum concentrations of the antibodies must be kept at therapeutic levels for long periods of time to eliminate HIV-1-infected cells, because persistently infected cells reside in the deep tissue. Therefore, multiple administrations of the antibodies are required, which could be expensive and labor-intensive. Gene therapy vectors can be used to produce therapeutic antibodies. However, transgene expression by any gene therapy vectors, including lentiviral vectors, can induce immune reactions against the transgenes, thereby decreasing the therapeutic effects of transgene products and eliminating the transduced cells. Therefore, it is important to express transgenes in the cell types that can develop tolerance to transgene products. B-cells physiologically express wide varieties of valuable antibody regions generated by recombination and mutations in genes. B-cells are known to induce tolerance to the valuable regions. This ability of B-cells was previously used to induce tolerance to self-antigens for therapy of autoimmune diseases and coagulation factor IX for treatment of hemophilia. Therefore, transduction of B-cells is likely to generate long-term anti-HIV-1 antibodies without inducing immune reactions to the antibodies. We have developed lentiviral vectors that can specifically transduce desired target cell types after systemic administration. The vector can selectively transduces splenic B-cells without conjugation of any B-cell-targeting ligand. By exploiting this B-cell-specific transduction by our lentiviral vectors, we will attempt to express anti-HIV-1 proteins specifically in B-cells to develop tolerance to the transgene products. We will further increase specificity of transgene expression with our targeting vector by combining it with transcriptional B-cell targeting by a B-cell-specific promoter. Using this highly B-cell-specific transgene expression system, we will express eCD4-Ig, a highly potent anti-HIV-1 reagent consisting of IgG and soluble CD4, in immunocompetent mice. We will investigate whether eCD4-Ig expression, specifically in B-cells, can induce long-term expression of eCD4-Ig by inducing developing tolerance. We will next express both secretory and membrane-anchored forms of eCD4-Ig in B-cells to generate an anti-HIV-1 B-cell receptor (BCR) on B-cells. We will investigate whether anti-HIV-1 BCR elicits signals upon binding to the HIV-1 envelope protein, inducing differentiation and growth of the transduced cells. The growth of transduced B-cells will increase the serum concentrations of eCD4-Ig. Differentiation of transduced cells to memory B-cells and plasma cells will extend the period of eCD4-Ig expression by prolonging the life span of transduced B-cells. We will then evaluate the therapeutic effects of B-cell-specific expression of eCD4-Ig in HIV-1 infection of humanized BLT mice. These experiments are designed to develop a novel gene therapeutic antibody that can be applied not only to therapy of HIV-1 infection, but also other types of infectious diseases.

项目概要 HIV-1 中和抗体能够非常有效地抑制 HIV-1 复制。然而，血清 抗体浓度必须长期保持在治疗水平以消除 HIV-1 感染的细胞，因为持续感染的细胞驻留在深层组织中。因此，多 需要施用抗体，这可能是昂贵且劳动密集型的。基因治疗 载体可用于产生治疗性抗体。然而，任何基因疗法的转基因表达 载体，包括慢病毒载体，可以诱导针对转基因的免疫反应，从而减少 转基因产品的治疗效果和消除转导的细胞。因此，重要的是 在可以对转基因产品产生耐受性的细胞类型中表达转基因。 B细胞生理学 表达由基因重组和突变产生的多种有价值的抗体区域。 B细胞 已知会诱导对有价值区域的耐受性。 B细胞的这种能力以前被用来诱导 对自身抗原的耐受性用于治疗自身免疫性疾病，对凝血因子 IX 的耐受性用于治疗 血友病。因此，B细胞的转导很可能会产生长期的抗HIV-1抗体，而无需 诱导针对抗体的免疫反应。我们开发了慢病毒载体，可以特异性地 全身给药后转导所需的靶细胞类型。该载体可以选择性转导脾脏 未结合任何 B 细胞靶向配体的 B 细胞。通过我们的研究人员利用这种 B 细胞特异性转导 慢病毒载体，我们将尝试在 B 细胞中特异性表达抗 HIV-1 蛋白，以培养对慢病毒载体的耐受性 转基因产品。我们将通过我们的靶向载体进一步提高转基因表达的特异性 将其与 B 细胞特异性启动子的转录 B 细胞靶向相结合。利用这种高度 B 细胞特异性 转基因表达系统，我们将表达 eCD4-Ig，一种高效抗 HIV-1 试剂，由 IgG 和 可溶性 CD4，在免疫功能正常的小鼠中。我们将研究 eCD4-Ig 表达（特别是在 B 细胞中）是否 可以通过诱导耐受性的产生来诱导 eCD4-Ig 的长期表达。接下来我们将表达两者 B 细胞中分泌型和膜锚定型的 eCD4-Ig 可产生抗 HIV-1 B 细胞受体 (BCR) 在 B 细胞上。我们将研究抗 HIV-1 BCR 是否在与 HIV-1 包膜蛋白结合后引发信号， 诱导转导细胞的分化和生长。转导的 B 细胞的生长会增加 eCD4-Ig 的血清浓度。转导细胞分化为记忆 B 细胞和浆细胞 通过延长转导的 B 细胞的寿命来延长 eCD4-Ig 表达的时间。然后我们将评估 eCD4-Ig B 细胞特异性表达对人源化 BLT 小鼠 HIV-1 感染的治疗作用。这些 实验旨在开发一种新型基因治疗抗体，该抗体不仅可用于治疗 HIV-1 感染，还有其他类型的传染病。