SPECIFIC T CELL IMMUNITY TO MUTATED RAS PROTEIN

Ras原癌基因的体细胞突变发生在大约20%的ALL中 人类的恶性肿瘤。这些突变涉及单核苷酸交换。 在导致p21ras蛋白表达的密码子12或61内 在第12或61位有单一取代氨基酸。初步 在小鼠身上的研究表明，p21ras可以通过以下途径对T细胞产生免疫原性 单一取代氨基酸的优点。因此，p21ras蛋白 代表与转化相关的潜在肿瘤特异性抗原。 活动，并由许多个人分享。 在类似的研究中，我们已经证明了嵌合体的连接区 慢性粒细胞白血病中表达的蛋白(p210bcr-abl)是 同样具有免疫原性。因此，表达的蛋白质的免疫原性 异常癌基因并不局限于p21ras。人们认识到的主要问题 使用p21ras、p21ras、p210bcr-abl或其他类似蛋白作为靶点 对于T细胞的攻击是因为它们是细胞内的蛋白质。T细胞可以 不对全蛋白起反应，而是对经过加工的多肽起反应 与MHC分子结合。初步数据证实，突变的 P21ras蛋白片段可被抗原提呈细胞加工 (APC)，并由II类MHC分子提呈，因此具有免疫原性 CD4+T细胞。CD4+T细胞的体内治疗效果需要 P21ras在肿瘤环境中被APC摄取、加工和 以足够的浓度呈现以刺激近似性免疫 T细胞分泌参与直接和/或间接细胞溶解的细胞因子 机械装置。目前，这些严格的条件已经在两个方面得到了满足 小鼠模型中p21ras特异性T细胞已被证明 有效地防止ras阳性肿瘤的生长。 突变的p21ras是一种肿瘤特异性抗原的观察提供了一种 发起人体研究以确定是否 RAS特异性反应可以产生并在治疗上用于 治疗或预防ras阳性恶性肿瘤的复发。第一 目的是确定携带ras阳性的患者 恶性肿瘤对异常的p21ras有免疫应答。 最终，可能有必要对患者进行免疫以诱导 RAS特异性T细胞。目前的拨款建议完成这些研究。 在着手进行人类免疫研究之前，有必要这样做。 目前提案的具体目标是：(1)确定是否 异常ras阳性肿瘤患者存在CD8+T细胞 启动异常的p21ras蛋白；(2)确定患者 在异常ras阳性的肿瘤中，存在CD8+T细胞 异常的p21ras蛋白；(3)确定T细胞是否来自正常 可以检测到对异常p21ras具有潜在反应性的个体， 在体外激活和扩增；以及(4)确定患者是否有 异常的ras阳性肿瘤对 异常的p21ras蛋白。

Somatic mutations of ras proto-oncogenes occur in approximately 20% of all human malignancies. The mutations involve a single nucleotide exchange within codons 12 or 61 which result in the expression of a p21ras protein with a single substituted amino acid at residues 12 or 61. Preliminary studies in mice have shown that p21ras can become immunogenic to T cells by virtue of the single substituted amino acid. Thus, p21ras protein represents a potential tumor-specific antigen related to the transforming events and shared by many individuals. In similar studies, we have shown that the joining region of a chimeric protein (p210bcr-abl) which is expressed in chronic myelogenous leukemia is likewise immunogenic. Thus, the immunogenicity of proteins expressed by aberrant oncogenes is not limited to p21ras. The major perceived problem of using p21ras, p21ras, p210bcr-abl or other similar proteins as targets for T cell attack is that they are internal cellular proteins. T cells do not respond to whole protein, but rather respond to processed peptides bound to MHC molecules. Preliminary data validated that the mutated segment of p21ras protein can be processed by antigen presenting cells (APC) and presented by class II MHC molecules and thereby is immunogenic to CD4+ T cells. In vivo therapeutic efficacy of CD4+ T cells requires that p21ras be taken up by APC in the environment of tumor, processed and presented in sufficient enough concentration to stimulate proximate immune T cells to secrete cytokines involved in direct and/or indirect cytolytic mechanisms. At present, these stringent conditions have been met in two murine models in which p21ras specific T cells have been shown to be effective at preventing the growth of ras-positive tumors. The observation that mutated p21ras is a tumor-specific antigen provides a strong impetus for initiating human studies to determine whether ras-specific responses can be generated and utilized therapeutically to treat or prevent the recurrence of ras-positive malignancies. The first objective is to determine whether patients bearing ras-positive malignancies have existent immune responses to aberrant p21ras. Ultimately, it might be necessary to immunize patients to elicit ras-specific T cells. The current grant proposes to perfom the studies necessary prior to embarking upon human immunization studies. The specific aims of the current proposal are: (1) to determine whether patients with aberrant ras-positive tumors have existent CD8+ T cells primed to the aberrant p21ras protein; (2) to determine whether patients with aberrant ras-positive tumors have existent CD8+ T cells primed to the aberrant p21ras protein; (3) to determine whether T cells from normal individuals with potential reactivity to aberrant p21ras can be detected, activated and expanded in vitro; and (4) to determine whether patients with aberrant ras-positive tumors develop specific humoral responses to the aberrant p21ras protein.