Molecular Mechanisms in Gastrointestinal Stromal Tumor

摘要 胃肠道间质瘤(GIST)是最常见的人类肉瘤，通常发生于 胃或小肠。癌症通常有一个单一突变，要么是KIT，要么是PDGFRA。伊马替尼 以及其他攻击与这些突变基因相关的癌蛋白的酪氨酸激酶抑制剂是 有效，但耐药性在18个月左右形成，治愈很少见。没有其他有效的方法 因此需要新的治疗方法。在过去的13年里，我们广泛地 在基因工程“敲门”小鼠模型和300多只小鼠中研究了GIST的免疫反应 新鲜的人类GIST手术标本。为了更深入地研究抗肿瘤免疫反应， 我们刚刚对50,000多个肿瘤内免疫细胞进行了单细胞RNA测序 使用赋形剂或伊马替尼治疗的小鼠。我们发现GDT细胞在肿瘤中富含 与血和脾进行比较。我们有初步数据表明GDT细胞在体内抑制了GIST的生长， 至少部分是由于IL-17的分泌。此外，伊马替尼治疗增加了IL-17的产生。我们 还发现GDT细胞渗透到人类GIST中，并产生IL-17，类似于小鼠模型。这个 GDT细胞在人类GIST或任何肉瘤中的重要性尚不清楚。我们有初步的 数据表明，GDT细胞的存在与GIST患者的总生存期呈正相关。我们 假设GDT细胞在GIST的抗肿瘤免疫反应中发挥关键作用，并且可以 为了治疗效果而操控。在这项提案中，我们将研究GDT细胞在 首先明确了小鼠和人GIST细胞抑制小鼠GIST的机制。接下来，我们 将确定伊马替尼如何影响小鼠GIST中的GDT细胞。有趣的是，无人监督 单细胞RNA表达的聚集性揭示了小鼠肿瘤内GDT细胞的两个主要亚群 模特。我们将研究它们的不同功能。我们将建立GDT细胞的翻译相关性 基特。我们将在我们的小鼠模型中测试几种GDT细胞激活剂与伊马替尼的结合。 此外，我们还将研究从新鲜的人类GIST手术标本中分离的GDT细胞的功能。 并将它们的存在与临床病理变量和免疫浸润相关联。最后，我们建议 对新鲜的人类GIST瘤内免疫细胞进行scRNAseq。我们的单细胞RNA 测序数据给了我们一个前所未有的机会来了解免疫反应的复杂性 并构成了这一提议中许多假设的基础。在我们关注GIST的同时， 我们希望我们的发现将与其他人类癌症中的GDT细胞相关。

ABSTRACT Gastrointestinal stromal tumor (GIST) is the most common human sarcoma and typically arises from the stomach or small intestine. The cancer generally has a single mutation, either in KIT or PDGFRA. Imatinib and other tyrosine kinase inhibitors that attack the oncoproteins associated with these mutated genes are effective, but resistance develops around 18 months and cure is rare. There are no other effective treatments so new therapeutic approaches are needed. Over the last 13 years, we have extensively studied the immune response to GIST in a genetically engineered “knockin” mouse model and over 300 fresh human GIST surgical specimens. To investigate the antitumor immune response in greater depth, we have just performed single-cell RNA sequencing on over 50,000 intratumoral immune cells from tumors of mice that were treated with vehicle or imatinib. We discovered that gdT cells are enriched in the tumor compared with blood and spleen. We have preliminary data that gdT cells suppress GIST growth in vivo, at least in part due to IL-17 secretion. Furthermore, imatinib therapy enhanced their IL-17 production. We also found that gdT cells infiltrate human GIST and they produce IL-17, similar to the mouse model. The importance of gdT cells in human GIST, or any sarcoma for that matter, is unknown. We have preliminary data that the presence of gdT cells correlates positively with overall survival in GIST patients. We hypothesize that gdT cells play a critical role in the antitumor immune response to GIST and can be manipulated for therapeutic efficacy. In this proposal, we will investigate the importance of gdT cells in mouse and human GIST by first defining the mechanism of gdT cell suppression of murine GIST. Next, we will determine how imatinib affects intratumoral gdT cells in murine GIST. Interestingly, unsupervised clustering of the single-cell RNA expression revealed 2 main subsets of intratumoral gdT cells in our mouse model. We will study their different function. We will establish the translational relevance of gdT cells in GIST. We will test several gdT cell activating agents in combination with imatinib in our mouse model. Furthermore, we will study the function of gdT cells isolated from fresh human GIST surgical specimens and correlate their presence to clinicopathologic variables and immune infiltrate. Lastly, we propose to perform scRNAseq on intratumoral immune cells from fresh human GISTs. Our single-cell RNA sequencing data have given us an unprecedented look into the complexity of the immune response to GIST and have formed the basis for numerous hypotheses in this proposal. While we are focused on GIST, we expect that our findings will have relevance to gdT cells in other human cancers.