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，中位生存时间为15个月。 尽管许多恶性肿瘤的癌症存活率有所提高，但GBM存活率仍然很低， 在过去30年中，这一情况发生了显著变化，强调迫切需要新的治疗方案。之一 癌症的共同特征是剪接干扰，而选择性剪接基因是一种有趣的 潜在的诊断生物标志物和治疗靶点的新来源。目前用于描述 使用mRNA测序的肿瘤中的剪接只能表明选择性外显子被转录，但 需要额外的验证来证实这些剪接形式产生功能蛋白。我们的初步 比较成对的肿瘤/正常样本的结果表明，GBM中的选择性剪接影响更多的 基因比以前预期的，无论是在转录本和蛋白质水平。基于这些数据，我们假设 一组可变剪接基因参与肿瘤的发生和GBM的发病机制。在这一提议中， 我们将使用精确医学方法的组合来检测和量化肿瘤特异性替代物， GBM中的剪接事件。我们将验证这些事件产生不同的proteoforms使用剪接感知 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， 候选人对患者来源的神经胶质瘤干细胞的生长，凋亡和侵袭的影响。如果 成功，这项工作将导致新的GBM蛋白质的生物学影响的新的理解， 可能导致基于独特的肿瘤抗原和分子生物学的治疗GBM的新方法。 途径。

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.