Ruth L. Kirschstein National Research Service Awards for Individual Postdoctoral

描述(申请人提供)：癌症是全球主要的死亡原因，预计其数量将进一步上升，2030年估计将导致1200万人死亡。目前的标准疗法，如化疗和放射治疗，已经在少数癌症中产生了显著的好处，但长期生存增加是罕见的，特别是对晚期疾病。因此，这些治疗本身不足以对抗癌症死亡率不断上升的趋势。免疫疗法为癌症治疗提供了一种有趣的方法，尤其是过继细胞转移(ACT)疗法，在正在进行的临床试验中显示出巨大的前景。ACT利用大量自体肿瘤相关抗原(TAA)特异性T细胞用于患者治疗，并在转移性黑色素瘤中产生了~50%的客观有效率(Dudley等人，2005年；Rosenberg等人，2008年)。然而，临床上的ACT疗法仅限于实体瘤，主要集中在黑色素瘤的治疗上。这在一定程度上是由于许多黑色素瘤相关抗原的鉴定和治疗性MART-1特异性T细胞受体(TCR)的发展(Hughes等人，2005年；Novellino等人，2005年)。相反，对于其他癌症类型，缺乏已知的TAA，可用于临床研究的匹配TCR要少得多。因此，为了扩大ACT治疗的范围，将进行一项大规模的努力，以确定用于卵巢癌免疫治疗的新的治疗靶点(即TAA)和试剂(即互补TCR)。利用蛋白质组学方法，主要组织相容性复合体(MHC)I类呈递的多肽将从HL A-A2+卵巢癌细胞系中提取并测序，以产生一个可能的肿瘤相关抗原的综合文库。在这个文库中，免疫原性抗原将通过使用来自卵巢癌患者样本的肿瘤浸润性淋巴细胞(TIL)来鉴定。然后，免疫原性抗原将被用来从TIL样本中物理地提取抗原特异性T细胞，并从单个细胞中克隆TCR基因，以产生一个肿瘤反应性T细胞受体基因库。这项任务将需要开发和使用一种新的集成微流控平台，用于单细胞捕获、全基因组扩增和顺序TCR特异性聚合酶链式反应。一旦TAA和TCR文库都组装好，将进行文库对文库的筛选，以配对功能TCR基因并鉴定它们的互补TAA。成功匹配的TAA和TCR随后将被鉴定，以获得有助于将TAA-TCR对分层用于潜在治疗用途的信息。在西方世界，卵巢癌是妇科癌症的主要死亡原因，因此，新的治疗干预措施将使卵巢癌受益匪浅。这一总体策略在未来也可以应用于其他癌症。 公共卫生相关性：过继T细胞转移涉及使用肿瘤靶向T细胞进行癌症治疗，并已在晚期黑色素瘤患者的临床试验中展示了真正的前景。我们的目标是通过识别相关的肿瘤抗原和相应的T细胞受体，将这种治疗扩大到包括卵巢癌患者。肿瘤抗原是癌症免疫治疗的潜在靶点，而T细胞受体是实际治疗的基础。

DESCRIPTION (provided by applicant): Cancer is the leading cause of death worldwide and its numbers are projected to rise further, resulting in an estimated 12 million deaths in 2030. Current standard therapies, such as chemotherapy and radiation therapy, have yielded significant benefit in a few cancers, but long-term survival gains are uncommon, especially for advanced disease. Thus, these treatments are insufficient by themselves to combat the escalating trend in cancer mortality. Immune therapy offers an interesting approach to cancer treatment and adoptive cell transfer (ACT) therapy, in particular, has demonstrated great promise in ongoing clinical trials. ACT utilizes large numbers of autologous tumor-associated antigen (TAA)-specific T cells for patient treatment and has yielded objective response rates of ~50% in metastatic melanoma (Dudley et al., 2005; Rosenberg et al., 2008). ACT therapy in the clinic, however, has been limited for solid tumors and predominantly focused on the treatment of melanoma. This is partly due to the identification of many melanoma-associated antigens and the development of a therapeutic MART-1-specific T cell receptor (TCR) (Hughes et al., 2005; Novellino et al., 2005). Conversely, for other cancer types, there is a dearth of known TAAs and far fewer matching TCRs that are viable for clinical investigation. Thus, to expand the scope of ACT therapy, a large-scale effort to identify new therapeutic targets (i.e. TAAs) and reagents (i.e. complementary TCRs) for immune therapy against ovarian cancer will be undertaken. Utilizing a proteomics approach, major histocompatibility complex (MHC) class I- presented peptides will be extracted and sequenced from HLA-A2+ ovarian cancer cell lines to generate a comprehensive library of putative tumor-associated antigens. Within this library, the immunogenic antigens will be identified through the use of tumor-infiltrating lymphocytes (TILs) from ovarian cancer patient samples. Then, in turn, the immunogenic antigens will be used to physically retrieve antigen-specific T cells from TIL samples and TCR genes from individual cells will be cloned to generate a library of tumor-reactive T cell receptor genes. This task will require the development and employment of a new integrated microfluidics platform for single cell capture, whole-genome amplification, and sequential TCR-specific PCR. Once TAA and TCR libraries are both assembled, library-against-library screens will be performed to pair functional TCR genes and identify their complementary TAAs. Successfully matched TAAs and TCRs will subsequently be characterized to acquire information that will help in stratifying TAA-TCR pairs for potential therapeutic use. Ovarian cancer is the leading cause of death due to gynecological cancer in the Western world and, thus, stands to benefit greatly from new therapeutic interventions. This general strategy can be applied to other cancers in the future. PUBLIC HEALTH RELEVANCE: Adoptive T cell transfer involves the use of tumor-targeting T cells for cancer therapy and has demonstrated real promise in clinical trials in patients with advanced melanoma. We aim to expand this treatment to include patients with ovarian cancer by identifying relevant tumor antigens and corresponding T cell receptors. Tumor antigens serve as potential targets for cancer immune therapy, while the T cell receptors form the basis of the actual treatment.