Receptor for gp96 on macrophages and dendritic cells

巨噬细胞和树突状细胞上的 gp96 受体

• 批准号: 6723510
• 负责人: Pramod K. Srivastava
• 金额: $ 32.63万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-12-01 至 2009-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6723510
• 关键词: antigen presenting cell; binding proteins; dendritic cells; differential display technique; electron microscopy; flow cytometry; genetically modified animals; heat shock proteins; laboratory mouse; laboratory rabbit; laboratory rat; macrophage; molecular chaperones; monoclonal antibody; neoplasm /cancer immunotherapy; polymerase chain reaction; protein protein interaction; protein purification; protein sequence; protein structure function; protein transport; receptor expression

DESCRIPTION (provided by applicant): Heat shock proteins (HSPs) (such as hsp70, hsp90, calreticulin families) are abundant soluble intracellular proteins. They possess powerful immunological properties, that include potentiation of innate and adaptive components of T cell immune response, as well as its down-regulation. The immunological activities of HSPs are dependent upon the interaction of HSPs with antigen presenting cells (APCs). Based on this dependence, the presence of an HSP receptor was envisaged in 1994. Such a receptor, CD91, was structurally identified and characterized and its functional activity demonstrated, in 2000. In this application, we aim to (i) assess other reported HSP receptors and identify new HSP receptors on human dendritic cells (DCs), (ii) define HSP-CD91 interaction in structural terms, (iii) test functional activity of CD91 in vivo in mediating cross-presentation antigens, and (iv) test the adjuvanticity of other CD91 ligands for immunotherapy. Specifically, we propose to study: Aim 1: Assessment and identification of HSP receptors on murine and human DCs. (a) Assessment of LOX-1 and other reported HSP receptors with respect to their relative roles vis-a-vis CD91, in cross-presentation of HSP-chaperoned peptides;(b) Generation of monoclonal antibodies to human HSP receptors, characterized by their ability to inhibit cross-presentation of HSP-chaperoned peptides. Aim 2: Structural analysis of HSP-CD91 interaction. (a) Definition of the hsp90/gp96/hsp70 domains involved in interaction with CD91, and if necessary, LOX1 ;(b) Definition of the CD91 domain(s) involved in interaction with hsp90 and gp96. Aim 3: Role of CD91 in cross-presentation and cross-priming. (a) Test influence of anti-CD91 antibodies on cross-presentation in vitro and on cross-priming in vivo, using HSP peptide complexes or whole cells as immunogens; (b) Test the influence of CD91 ligands alpha2 macroglobulin (alpha2M) and RAP on the same; (c) Test the influence of excess peptide-free HSPs on the same; (d) Construct mice expressing dominant negative forms of CD91 and test priming and cross-priming of T cell responses in such mice. Aim 4: Mechanism of immunogenicity of alpha2M and its application to cancer immunotherapy (a) Examine the mechanism of immunogenicity and anti-tumor reactivity of alpha2M-peptide complexes; (b) Test complexes of antigenic peptides with alpha2M in cross-presentation in vitro and for therapy of pre-existing cancers.

描述（由申请人提供）：热休克蛋白（HSP）（例如hsp70、hsp90、钙网蛋白家族）是丰富的可溶性细胞内蛋白。它们具有强大的免疫学特性，包括 T 细胞免疫反应的先天性和适应性成分的增强及其下调。 HSP 的免疫活性取决于 HSP 与抗原呈递细胞 (APC) 的相互作用。基于这种依赖性，1994 年设想了 HSP 受体的存在。2000 年对这种受体 CD91 进行了结构鉴定和表征，并证明了其功能活性。 在此应用中，我们的目标是 (i) 评估其他报道的 HSP 受体并鉴定人类树突状细胞 (DC) 上的新 HSP 受体，(ii) 从结构角度定义 HSP-CD91 相互作用，(iii) 测试 CD91 的体内功能活性介导交叉呈递抗原，以及 (iv) 测试其他 CD91 配体对免疫治疗的佐剂作用。具体来说，我们建议研究： 目标 1：评估和鉴定小鼠和人类 DC 上的 HSP 受体。 (a) 评估 LOX-1 和其他报道的 HSP 受体在 HSP 陪伴肽交叉呈递中相对于 CD91 的相对作用；(b) 人 HSP 受体单克隆抗体的生成，其特征在于它们抑制 HSP 陪伴肽交叉呈递的能力。 目标 2：HSP-CD91 相互作用的结构分析。 (a) 涉及与 CD91 相互作用的 hsp90/gp96/hsp70 结构域的定义，必要时还包括 LOX1；(b) 涉及与 hsp90 和 gp96 相互作用的 CD91 结构域的定义。 目标 3：CD91 在交叉呈递和交叉引发中的作用。 (a) 使用 HSP 肽复合物或全细胞作为免疫原，测试抗 CD91 抗体对体外交叉呈递和体内交叉引发的影响； (b)测试CD91配体α2巨球蛋白(α2M)和RAP对其的影响； (c) 测试过量的无肽热休克蛋白对其的影响； (d)构建表达显性阴性形式的CD91的小鼠并测试这些小鼠中T细胞反应的启动和交叉启动。 目标4：α2M的免疫原性机制及其在癌症免疫治疗中的应用 (a) 检查α2M-肽复合物的免疫原性和抗肿瘤反应性机制； (b) 测试抗原肽与 alpha2M 的复合物在体外交叉呈递以及用于治疗已存在的癌症。