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

项目总结： 适应性免疫系统可以特异性地识别和杀死癌细胞。重振旗鼓的疗法 肿瘤特异性CD8T细胞治愈了一部分肺癌患者，促使FDA批准和使用 作为护理的标准。肿瘤反应性T细胞识别癌细胞抗原--显示的蛋白质片段 是肿瘤细胞所特有的。到目前为止，抗肿瘤免疫的研究主要集中在来源于 来自癌症基因组中的体细胞突变，这不足以充分解释抗肿瘤免疫 回应。此外，突变衍生的新抗原通常对每个肿瘤都是独一无二的，并针对它们 可能需要为每个病人量身定做治疗。这项提议的长期目标是扩大 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存在反复发生的驱动突变 因素方面，很少有研究探讨剪接异常与肿瘤免疫原性的关系。 值得注意的是，大约10%的肺腺癌具有RBM10功能缺失突变，RBM10是一个关键的剪接因子。 这项工作将检查Rbm10缺陷肿瘤中的剪接衍生抗原。中心假设是 Rbm10缺陷会产生异常的剪接转录本，这将引发适应性免疫 如果被肿瘤细胞翻译并呈现为新的抗原，则会产生应答。这一假设将在一口井中进行检验- 通过追求两个特定的目标来表征小鼠肺腺癌模型。在目标1中，信使核糖核酸 将进行测序以表征在Rbm10的背景下发生的异常剪接事件 缺乏症。质谱学将被用来验证异常拼接的RNA产生抗原 呈现给免疫系统。在Aim 2中，CRISPR/Cas9将被用于在An中引入Rbm10突变 具有免疫功能的自体小鼠模型。组织学和流式细胞术分析将在 Rbm10缺陷和成熟肿瘤中淋巴细胞的浸润性以评估内源性抗肿瘤免疫。 这项研究的创新之处在于，它建议经验性地测试剪接衍生抗原在 利用基因组工程技术在自体小鼠模型中进行抗肿瘤免疫。它意义重大，因为它 可能识别新的肿瘤限制性抗原，这些抗原是癌症免疫疗法的靶点。而T细胞 靶向免疫疗法为晚期癌症患者提供持久的治疗，先前的研究强调 一组有限的癌症特异性T细胞抗原。这项工作的最终目的是揭示小说 增强癌细胞免疫识别的机制，导致开发新的治疗方法 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.