Investigating splicing-derived antigens in the context of Rbm10-mutant lung adenocarcinoma

项目概要： 适应性免疫系统可以特异性地识别和杀死癌细胞。恢复活力的疗法 肿瘤特异性CD 8 T细胞治愈了一部分肺癌患者，促使FDA批准和使用 作为护理标准。肿瘤反应性T细胞识别癌细胞抗原-展示的蛋白质片段 是肿瘤细胞所独有的迄今为止，抗肿瘤免疫的研究主要集中在来源于肿瘤细胞的抗原。 癌症基因组中的体细胞突变，这不足以完全解释抗肿瘤免疫 反应此外，突变衍生的新抗原通常对于每个肿瘤是独特的，并且靶向它们是可能的。 可能需要为每个病人量身定制治疗方案。该提案的长期目标是扩大 癌症特异性抗原的范例。最近的研究发现了一类新的癌症限制性抗原 由异常剪接事件产生。虽然几种癌症在剪接中含有复发性驱动突变， 由于这些因素，很少有研究探讨剪接失调与肿瘤免疫原性之间的关系。 值得注意的是，约10%的肺腺癌的特征是RBM 10（一种关键剪接因子）的功能缺失突变。 这项工作将检查Rbm 10缺陷肿瘤中的剪接衍生抗原。核心假设是 Rbm 10缺陷会产生异常剪接的转录本，这将引发适应性免疫， 如果肿瘤细胞将其翻译并呈递为新抗原，则会产生免疫应答。这一假设将在一个良好的测试- 通过追求两个特定的目标来表征肺腺癌的小鼠模型。在目标1中，mRNA 将进行测序以表征在Rbm 10背景下发生的异常剪接事件 缺陷质谱法将用于验证异常剪接的RNA产生抗原 呈现给免疫系统。在目标2中，CRISPR/Cas9将被用于在一个细胞中引入Rbm 10突变。 免疫活性的、本地鼠模型。组织学和流式细胞术分析将在 Rbm 10缺陷型和有效型肿瘤中的浸润淋巴细胞，以评估内源性抗肿瘤免疫。 这项研究的创新之处在于，它提出了经验性地测试剪接衍生抗原在以下方面的作用： 在本地小鼠模型中使用基因组工程的抗肿瘤免疫。这一点很重要，因为它 可以识别新的肿瘤限制性抗原，这些抗原可被癌症免疫疗法靶向。虽然T细胞 靶向免疫疗法为晚期癌症患者提供了持久的治疗，先前的研究强调， 癌症特异性T细胞抗原的有限集合。这项工作的最终目的是揭示小说 增强癌细胞的免疫识别机制，从而开发出新的治疗方法， 肺癌

Project summary: The adaptive immune system can specifically recognize and kill cancer cells. Therapies that reinvigorate tumor-specific CD8 T cells cure a subset of patients with lung cancer, prompting their FDA approval and use as standard of care. Tumor-reactive T cells recognize cancer cell antigens -- displayed fragments of proteins that are unique to tumor cells. To date, study of anti-tumor immunity has primarily focused on antigens derived from somatic mutations in the cancer genome, which are insufficient to fully explain the anti-tumor immune response. Moreover, mutation-derived neoantigens are generally unique to each tumor, and targeting them might require bespoke treatments for each patient. The long-term goal of this proposal is to expand the paradigm of cancer-specific antigens. Recent work has uncovered a novel class of cancer-restricted antigens produced by abnormal splicing events. While several cancers harbor recurrent driver mutations in splicing factors, few studies have explored the relationship between dysregulated splicing and tumor immunogenicity. Notably, ~10% of lung adenocarcinomas feature loss-of-function mutations in RBM10, a critical splicing factor. This work will examine splicing-derived antigens in Rbm10-deficient tumors. The central hypothesis is that Rbm10 deficiency will generate aberrantly spliced transcripts, which will elicit an adaptive immune response if translated and presented as novel antigens by tumor cells. This hypothesis will be tested in a well- characterized murine model of lung adenocarcinoma by pursuing two specific aims. In Aim 1, mRNA sequencing will be performed to characterize aberrant splicing events that occur in the context of Rbm10 deficiency. Mass spectrometry will be used to validate that aberrantly spliced RNAs produce antigens presented to the immune systems. In Aim 2, CRISPR/Cas9 will be used to introduce Rbm10 mutations in an immunocompetent, autochthonous murine model. Histologic and flow cytometric analyses will be performed on infiltrating lymphocytes in Rbm10-deficient and proficient tumors to assess endogenous anti-tumor immunity. This research study is innovative in that it proposes to empirically test the role of splicing-derived antigens in anti-tumor immunity using genome engineering in autochthonous mouse models. It is significant because it may identify new, tumor-restricted antigens that are targetable by cancer immunotherapies. While T-cell targeted immunotherapies provide durable cures for patients with advanced cancer, prior studies emphasize a limited set of cancer-specific T cell antigens. The ultimate objective of this work is to uncover novel mechanisms to potentiate immune recognition of cancer cells, leading to the development of new therapies for lung cancer.