Ludwig Institute for Cancer Research, University of Lausanne, Lausanne, Switzerland.
Department of Oncology, Centre Hospitalier Universitaire Vaudois, Lausanne, Switzerland.
Nat Commun. 2024 Mar 15;15(1):2357. doi: 10.1038/s41467-024-46408-3.
Circular RNAs (circRNAs) are covalently closed non-coding RNAs lacking the 5' cap and the poly-A tail. Nevertheless, it has been demonstrated that certain circRNAs can undergo active translation. Therefore, aberrantly expressed circRNAs in human cancers could be an unexplored source of tumor-specific antigens, potentially mediating anti-tumor T cell responses. This study presents an immunopeptidomics workflow with a specific focus on generating a circRNA-specific protein fasta reference. The main goal of this workflow is to streamline the process of identifying and validating human leukocyte antigen (HLA) bound peptides potentially originating from circRNAs. We increase the analytical stringency of our workflow by retaining peptides identified independently by two mass spectrometry search engines and/or by applying a group-specific FDR for canonical-derived and circRNA-derived peptides. A subset of circRNA-derived peptides specifically encoded by the region spanning the back-splice junction (BSJ) are validated with targeted MS, and with direct Sanger sequencing of the respective source transcripts. Our workflow identifies 54 unique BSJ-spanning circRNA-derived peptides in the immunopeptidome of melanoma and lung cancer samples. Our approach enlarges the catalog of source proteins that can be explored for immunotherapy.
环状 RNA（circRNAs）是共价闭合的非编码 RNA，缺乏 5' 帽和 poly-A 尾。然而，已经证明某些 circRNAs 可以进行主动翻译。因此，人类癌症中异常表达的 circRNAs 可能是肿瘤特异性抗原的一个未被探索的来源，潜在地介导抗肿瘤 T 细胞反应。本研究提出了一种免疫肽组学工作流程，特别侧重于生成 circRNA 特异性蛋白 fasta 参考。该工作流程的主要目标是简化鉴定和验证潜在源自 circRNAs 的人类白细胞抗原（HLA）结合肽的过程。我们通过保留由两个质谱搜索引擎独立鉴定的肽以及对经典衍生肽和 circRNA 衍生肽应用特定的 FDR，增加了我们工作流程的分析严格性。跨越后剪接连接（BSJ）的区域特异性编码的 circRNA 衍生肽的子集通过靶向 MS 以及相应源转录物的直接 Sanger 测序进行验证。我们的工作流程在黑色素瘤和肺癌样本的免疫肽组中鉴定了 54 个独特的 BSJ 跨越 circRNA 衍生肽。我们的方法扩大了可以探索用于免疫治疗的源蛋白目录。
