吸烟对KRAS突变肺腺癌异质性的作用机制研究

KRAS-mutant lung adenocarcinomas are malignant, poor prognosis and heterogeneous. Due to the structure and location of KRAS protein, there is still no targeted therapy available yet. The major strategies against KRAS-mutant lung adenocarcinomas including 1. Bypassing the KRAS gene and targeting its downstream pathways, however the response rate is relatively low due to the heterogeneity among patients; 2. Screening for small molecules that have synthetically lethal interactions with oncogenic KRAS, which means the small molecule inhibitors specifically "kill" tumor cells bearing KRAS mutations. However, there are rare common hits among these screenings due to the heterogeneity and fruitful results are suffered from poor reproducibility. It is very hard to translate laboratory findings into clinically applicable therapeutics. Therefore, it is critical to find the molecular subtypes of KRAS mutant lung adenocarcinomas and study the underlying mechanism of the heterogeneity of KRAS-mutant lung adenocarcinomas. Our preliminary results found that KRAS mutant lung adenocarcinomas can be classified into two subtypes. These two subtypes are closely linked to smoking-induced somatic mutations and aberrant DNA methylation. However, it is unclear how somatic mutations and abnormal DNA methylation caused by smoking affect the different biological characteristics of KRAS-mutated lung adenocarcinomas and therapeutic strategies. In this study, we aim to study the underlying mechanisms of the heterogeneity of KRAS-mutant lung adenocarcinomas from the aspect of smoking, identify the biological characteristics of KRAS-subtypes, and provide a theoretical basis for the molecular diagnosis, prevention and personalized therapy of this malignant tumor type.

KRAS突变肺腺癌具有恶性程度高和异质性高的特点。由于KRAS蛋白的结构特性及胞内位置等因素，目前临床上尚无有效的靶向药物，而主要应用以下策略：1. 靶向KRAS下游通路，但因病人间存在异质性而响应率低；2. 筛选能特异性“杀死”携带KRAS突变的肿瘤细胞的小分子抑制剂，但因其亚型不明确，使得研究间的可重复性差，临床转化效率低。因此对KRAS突变肺腺癌的精准医疗的关键是理解其异质性的机理。我们的前期工作综合多个平台的高通量数据，发现两个KRAS突变肺腺癌的亚型，且分别与吸烟导致的体细胞突变和异常的DNA甲基化有紧密关联。但目前尚不清楚吸烟导致体细胞突变和异常DNA甲基化是如何影响KRAS突变肺腺癌的生物学特征，进而形成不同的分子亚型。本申请旨在深入研究吸烟导致KRAS突变肺腺癌异质性的分子机制并阐明各亚型的生物学特性，为KRAS突变肺腺癌的分子诊断、风险预警及治疗策略提供依据。

肺腺癌的异质性，特别是KRAS突变型肺腺癌的异质性一直是临床治疗的难点，但其异质性的分子机制却仍不清楚。因此，只有一部分患者可以从靶向治疗或免疫检查点阻断中获益。而关于肺腺癌异质性的研究非常少，在本研究中，我们综合了各组学的数据类型，旨在对肺腺癌的异质性的因素做深入的探讨，并对肺腺癌做更精准的分型。我们发现KRAS突变型肺腺癌的异质性与吸烟引起的DNA甲基化、体细胞突变和肿瘤浸润的免疫细胞组成相关。我们从中提取出了亚型特异性的基因组及表观基因组的特征，这些特征有助于我们发现亚型特异性的药物。我们利用两个亚型的不同的特征进行了药物敏感性预测，发现了几个具有亚型特异性杀伤作用的小分子药物。我们的研究解释了KRAS突变型肺腺癌的异质性潜在的分子机理可能与吸烟相关，为KRAS突变肺腺癌的临床分型和治疗提供了潜在的治疗靶点。此外，我们挖掘并整合了肺腺癌单细胞测序数据来研究吸烟对于肺癌免疫微环境的影响，我们发现了一簇与吸烟密切相关的调节性T细胞（Treg细胞）的亚群，并对这个亚群的时间轨迹、空间相互作用、分子特征以及临床预后做了详细的分析，我们发现这一簇与吸烟相关的Treg亚群, ADAM12+CTLA4+Treg，高度表达细胞粘附途径和脂质代谢的基因，且具有很强的免疫抑制活性，我们通过受体-配体分析和重构肿瘤免疫微环境的空间，发现ADAM12+CTLA4+Tregs与耗竭性T细胞在空间上非常接近且配体受体结合更紧密。我们还利用了TCGA的肺腺癌（LUAD）数据集的生存信息来评估ADAM12+CTLA4+Tregs的预后价值，我们发现ADAM12+CTLA4+Tregs高比例组的总体生存期显著低于ADAM12+ CTLA4+ Tregs低比例组。综上所述，我们从不同的维度（基因组、表观基因组、转录组、单细胞转录组）系统分析了吸烟对肺腺癌的临床分型、靶向治疗和免疫微环境三个方面的影响，为肺腺癌（特别是KRAS突变肺腺癌) 的临床分型和治疗提供了潜在的生物标志物和药物靶点。