高级别肺神经内分泌癌ASCL1–DLL3通路的分子异质性研究

High-grade neuroendocrine carcinomas(HGNEC), including the large cell neuroendocrine carcinoma (LCNEC) and small cell lung cancer(SCLC), harbor the most pathological and genetic heterogeneity, and bear the poorest survival. Targeted therapies for these patients have been confronted with rough rides for decades due to failures of discovering or verifying responsive driver genes. Latest researches have revealed that ASCL1-DLL3 pathway is the potentially actionable target. However, results from a phase I clinical trial indicated that the objective response rate of refractory SCLC to Rova-T which targets the DLL3 was only 23%. Recently, molecular heterogeneity has been playing an increasingly important role in pathological classification of lung cancer, tumor evolutions and resistance to targeted regimens. Our previous studies have confirmed that inter- and intra-tumoral heterogeneity in pulmonary adenocarcinomas are decided by various driver and passenger genes, which indicated potential mechanisms of molecular evolutions in early pulmonary adenocarcinomas and corresponding differences in their responses to targeted therapies. Aided by laser captured sequencing, IHC, FISH and next generation sequencing, this study aim to picture the trunk-branch tree model and explore intratumoral heterogeneity in HGNEC and their evolution principles of gene mutations, protein expressions and signal pathways. Then how the targeted regimen, Rova-T, interferes in ASCL1-DLL3 pathway is also explored based on constructions of HGNEC cell lines. Our study will provide solid evidences to explore the molecular evolution and confirm the driver genes of HGNEC for further targeted therapies.

高级别肺神经内分泌癌（HGNEC），包括LCNEC和SCLC，瘤内病理和基因异质性高、传统化疗有效率低，预后恶劣，靶向治疗举步维艰。研究提示ASCL1-DLL3是HGNEC的潜在关键靶点；而在I期临床试验中，针对DLL3的靶向药物Rova-T在SCLC的客观缓解率仅23%。分子异质性在肺癌病理分型、肿瘤发生演化和靶向耐药中的价值日显重要。我们前期研究发现肺腺癌瘤内、瘤间异质性决定于驱动和旁路基因的差异，揭示了早期腺癌潜在的分子演化机制和靶向治疗反应差异。本课题拟通过激光显微切割多点采样肿瘤标本，运用IHC、FISH、二代测序等方法，绘制基因树干-分支模型，探索HGNEC瘤内异质性及基因、蛋白表达、信号调节通路的演变规律。并构建HGNEC肺癌细胞株，探索ASCL1-DLL3通路在靶向药物Rova-T干预中的关键作用。将为HGNEC肿瘤分子进化机制、制定HGNEC靶向治疗策略提供实验室依据。

根据2015年病理新分类中，肺高级别神经内分泌肿瘤分为大细胞神经内分泌肿瘤（LCNEC）以及小细胞肺癌（SCLC）。在本项目资助下，通过开放网络搭建多媒体数据平台，邀请国内十余家单位参与，构建国内最大型的肺LCNEC数据库，最终纳入381例LCNEC，通过回顾性分析其临床特征、病理及预后相关信息，探索这一少见癌种的独特性及化疗模式优化选择;在预后相关指标上，我们发现CD56，Syn，CgA，Ki-67等免疫组化指标均与LCNEC预后相关；同时我们发现，无论是术后辅助治疗还是一线治疗，小细胞肺癌化疗方案相对于非小细胞化疗方案而言，都是更优的方案（11.5 vs. 7.2 个月, P=0.003）；此多中心回顾数据提供了目前阶段针对LCNEC治疗高级别的循证医学证据。另外，进一步汇总目前SCLC治疗进展并发表前沿综述，我们发现SCLC治疗中免疫治疗联合化疗模式具备前景。除此以外，通过血浆外泌体miRNA，肺部磨玻璃结节高通量测序，围术期改良改良等方面对肺癌综合诊疗形成筛查-诊断-治疗-机制链条化研究输出。以上研究均已总结成文，分别发表在Lung Cancer, J Extracellular Vesicle, J Thorac Oncol, Ann Surg Oncol, Ann Thorac Surg等杂志上(均为2016年国家自然科学基金基金81673031资助下完成)。本研究的结果进一步提高了人们对于肺高级别神经内分泌肿瘤的认识，为进一步探索肺癌个体化治疗提供基础。