Using proteogenomics to assess the functional impact of alternative splicing events in glioblastoma

项目摘要 胶质母细胞瘤(GBM)是最恶性的脑肿瘤类型，中位生存期为15个月。 尽管许多恶性肿瘤的癌症存活率取得了进展，但基底膜存活率仍然很低，而且 在过去30年里发生了显著变化，强调迫切需要新的治疗方案。其中之一 癌症的共同特征是剪接干扰，另一种剪接基因是一个有趣的 潜在的诊断生物标记物和治疗靶点的新来源。当前要表征的方法 在肿瘤中使用mRNA测序进行剪接只能表明选择性外显子被转录，但 还需要进一步的验证来验证这些剪接形式是否能产生功能蛋白质。我们的预赛 结果比较成对的肿瘤和正常样本表明，GBM中的选择性剪接影响更大 在转录本和蛋白质水平上，基因都比之前预期的要好。基于这些数据，我们假设 一组选择性剪接的基因参与了肿瘤的发生和GBM的发病。在这份提案中， 我们将使用精准医学方法的组合来检测和量化肿瘤特异性替代方案 在GBM中拼接事件。我们将使用剪接感知来验证这些事件是否会产生不同的蛋白质形式 蛋白质组学实验。最后，我们将从功能上表征差异表达的剪接异构体 候选药物对患者来源的胶质瘤干细胞的生长、凋亡和侵袭的影响。如果 这项工作的成功将导致对新的GBM蛋白形式的生物学影响的新的理解，以及 基于独特的肿瘤抗原和分子可能导致治疗基底膜的新方法 小路。

PROJECT ABSTRACT Glioblastoma (GBM) is the most malignant type of brain tumor, with a median survival time of 15 months. Despite advances in cancer survival for many malignancies, GBM survival rates still remain low and have not significantly changed over the last 30 years, emphasizing the urgent need for new treatment options. One of the common hallmarks of cancer is splicing perturbations and alternatively spliced genes are an interesting new source for potential diagnostic biomarkers and therapeutic targets. Current methodologies to characterize splicing in tumors using mRNA sequencing can only indicate that alternative exons are transcribed, but additional validation is needed to verify that these spliceforms produce functional proteins. Our preliminary results comparing paired tumor/normal samples indicate that alternative splicing in GBM affects many more genes than previously expected, both at the transcript and protein levels. Based on these data, we hypothesize that a set of alternatively spliced genes are involved in tumor initiation and GBM pathogenesis. In this proposal, we will use a combination of precision medicine approaches to detect and quantify tumor-specific alternative splicing events in GBMs. We will verify that these events produce distinct proteoforms using splicing-aware proteogenomics experiments. Finally, we will functionally characterize differentially expressed splice isoform candidates for their effects on growth, apoptosis, and invasion in patient-derived glioma stem cells. If successful, this work will lead to new understanding of the biological impact of novel GBM proteoforms, and potentially lead to novel approaches to treatment of GBM based on unique tumor antigens and molecular pathways.