Regulation of Class I Gene Expressions in Tumor Cells

描述：(改编自申请者的摘要)大多数恶性肿瘤 目前肿瘤是无法治愈的，除非在早期阶段被发现；免疫学 治疗方法在改善治疗方面往往很有希望。这样做的总体目标是 拟议的项目是通过以下方式对基本机制有新的了解 人类主要组织相容性抗原基因在肿瘤细胞中的表达是什么 受干扰素调节。预计这样的知识最终将 促进开发更有效的治疗目前不治之症的尝试 通过细胞免疫治疗和疫苗策略治疗癌症。中环 这一提议的假设是干扰素刺激 人类白细胞抗原(人类白细胞抗原)Ia类基因，即人类白细胞抗原A2和人类白细胞抗原1b类基因， 通过不同和新颖的机制。因为这两个人类白细胞抗原I类 基因在细胞毒性T细胞和自然杀伤细胞中都起着关键作用 淋巴细胞介导的肿瘤细胞杀伤，了解其选择性 监管应该增加关于肿瘤免疫学的基本知识。少校 这项建议的具体目标是：(1)确定 干扰素-γ介导的人类白细胞抗原-A2基因转录后激活 通过3‘’干扰素反应元件，以及(2)表征不同的 人类白细胞抗原E基因启动子中的干扰素-γ转录反应元件。 为了实现这些特定的目的，一组顺序的分子生物学基因 将进行转移和体外诱变试验，以确定 这两者的不同调控所需的确切DNA和RNA序列 肿瘤细胞系和新鲜白血病细胞中的基因。细胞和分子 生物分析，包括共聚焦显微镜和体外RNA剪接 分析，将进行，以定义新的机制 人类白细胞抗原A2基因的转录后调控。然后用生化方法 将被用来鉴定、表征和纯化可能的蛋白质 影响人类白细胞抗原A2和E基因干扰素应答的因素。 在鉴定顺式作用DNA序列和反式作用之后 调节人类白细胞抗原-E基因转录反应的因素， 将进行生物化学和遗传学研究，以确定蛋白质-蛋白质 相互作用与介导干扰素的信号转导事件 回应。从这个项目中获得的知识将增加基本的 理解人类白细胞抗原Ia类和Lb类基因的差异 在肿瘤细胞中被调节，并最终可能导致新的治疗方法 优化肿瘤和异基因骨髓的免疫治疗 跨越组织相容抗原屏障的移植。

DESCRIPTION: (Adapted from applicant's abstract) The majority of malignant tumors are currently incurable unless detected at an early stage; immunologic approaches often promise in improving therapy. The overall objective of this proposed project is to obtain new understanding of the basic mechanisms by which human major histocompatibility antigen gene expression in tumor cells is regulated by interferon. It is anticipated that such knowledge will ultimately facilitate attempts to develop more effective treatment of currently incurable cancers through cellular immune therapy and vaccine strategies. The central hypothesis of this proposal is that interferon stimulates expression of the human leukocyte antigen (HLA) Class Ia gene, HLA-A2, and the HLA Class lb gene, HLA-E, through differing and novel mechanisms. Since these two HLA Class I genes play critical roles in both cytotoxic T-cell and natural killer cell lymphocyte mediated tumor cell killing, understanding their selective regulation should increase basic knowledge about tumor immunology. The major specific aims of this proposal are (1) to determine the mechanism by which interferon-gamma mediates post-transcriptional stimulation of the HLA-A2 gene through a 3' interferon response element, and (2) to characterize the distinct interferon-gamma transcriptional response elements in the HLA-E gene promoter. To achieve these specific aims, a sequential set of molecular biological gene transfer and in vitro mutagenesis assays will be carried out to define the exact DNA and RNA sequences required for the differing regulation of these two genes in tumor cell lines and fresh leukemia cells. Cell and molecular biological assays, including confocal microscopy and in vitro RNA splicing analysis, will be carried out to define the novel mechanism of post-transcriptional regulation of the HLA-A2 gene. Biochemical methods then will be utilized to identify, characterize and purify the putative protein factors that mediate the interferon response of the HLA-A2 and HLA-E genes. Following the identification of the cis-acting DNA sequences and trans-acting factors that mediate the transcriptional response of the HLA-E gene, biochemical and genetic studies will be carried out to define protein-protein interactions and the signal transduction events that mediate the interferon response. The knowledge gained from this project will increase the basic understanding of how the HLA Class Ia and Class lb genes are differentially regulated in tumor cells, and may ultimately lead to new therapeutic approaches to optimize immunotherapy of cancer as well as allogeneic bone marrow transplantation across histocompatibility antigen barriers.