去磷酸化介导的KMT2A相分离调控肺癌奥西替尼耐药的分子机制研究

肺癌靶向治疗耐药极大限制了患者长期获益，是肺癌临床治疗的痛点。既往研究倾向以关键分子及其调控元件的线性关系为基础，而近期研究表明基因三维空间调控网络能更真实地阐释耐药过程。申请者通过Hi-C测序发现肺癌奥西替尼耐药前后染色质三维结构互作改变。染色质三维结构变化和组蛋白H3K4me3修饰密切相关，后者受组蛋白甲基转移酶KMT2A调控，这一修饰过程的本质是相分离。结构生物学分析表明KMT2A和组蛋白H3有高比例内在无序结构域，提示两者均能发生相分离。相分离主要由磷酸化修饰驱动，申请者发现耐药过程中KMT2A磷酸化水平差异改变。拟通过Hi-C测序、相分离模型构建等阐释去磷酸化介导的KMT2A相分离通过调控组蛋白甲基化修饰，介导染色质三维结构重塑，进而诱导肺癌奥西替尼耐药的分子机制；通过人源肿瘤异种移植模型验证KMT2A作为逆转耐药靶点的有效性，为临床克服靶向治疗抵抗提供新思路和实验证据。

Drug resistance to targeted therapy of lung cancer greatly limits the long-term benefit of patients, and this is a pain point of clinical treatment for lung cancer. Previous studies have tended to research the mechanisms of drug resistance based on the linear relationship between key molecules and their regulatory elements, however, recent studies have shown that three-dimensional spatial regulatory networks of gene can explain the drug resistance process more truthfully. The applicant has identified the changes of three-dimensional chromatin structure interactions after osimertinib resistance in lung cancer by Hi-C sequencing. The three-dimensional structural changes of chromatin are closely related to H3K4me3 modification of histone H3, which is regulated by KMT2A. KMT2A is a kind of histone methyltransferase. And this modification process is phase separation. Structural biological analysis showed that KMT2A and histone H3 have a high proportion of internal disordered domains, suggesting that both KMT2A and histone H3 can develop phase separation. Phase separation is mainly driven by phosphorylation modification, while the applicant has found that the phosphorylation level of KMT2A changed during drug resistance. This project aims to explain the molecular mechanism of KMT2A phase separation mediated by dephosphorylation controls chromatin three-dimensional structural remodeling by regulating histone methylation modification, and thus induce osimertinib resistance, based on Hi-C sequencing, phase separation model construction and other techniques. Based on patient-derived xenograft, the effectiveness of KMT2A as a target for reversing osimertinib resistance would be verified, which could provide new insights and experimental evidence for overcoming resistance to targeted therapy in clinical.