靶向抑制DNA损伤修复激酶ATM增强溶瘤病毒M1的抗结直肠癌作用及其机制研究

The applicant reported that oncolytic virus M1 kills colon cancer cells in vitro and in vivo at varying degrees, suggesting that M1 virus has clinical potential in the treatment of colon cancer. Clinical researches have shown that oncolytic viruses are generally safe, but monotherapy is not good enough. Based on the results of the previous enhancers screening for M1 virus, this project initially identified DNA damage repair kinase ATM inhibitors significantly enhance the anti-cancer effect of M1. On basis of this hypothesis: Targeted inhibition of ATM attenuates DSBs repair by hindering homologous recombination (HR) and non-homologous end repair (NHEJ) pathways, amplifying M1-induced DNA damage, leading to pyroptosis. This project intends to verify ATM inhibitors enhancing oncolytic activity of M1 virus in animal (subcutaneous tumor of immunocompetent and immunodeficient mouse) and PDX clinical models, while we plan to explore the detailed mechanism that ATM inhibitors promoting oncolytic effects of M1. It is the key factors of synergy that elucidating the dual inhibition of DSBs repair and triggering pyroptosis This study can provide a theoretical basis for the exploring DSBs targets for the enhancement of oncolytic activity, and develop new ideas for the combination therapy of colon cancer.

申请者报道在多种结直肠癌体内外模型中，溶瘤病毒M1对癌细胞有不同程度的杀伤，提示M1有应用于临床治疗结直肠癌潜能。临床研究显示溶瘤病毒普遍安全性高，但作为单一疗法疗效欠佳。基于前期M1增效剂筛选结果，本项目初步鉴定出DNA损伤修复激酶ATM抑制剂显著增强M1的抗结直肠癌效应。据此提出假说：靶向抑制ATM通过阻碍同源重组（HR）、非同源末端 (NHEJ) 修复通路抑制DSBs修复，放大M1诱导的DNA损伤，导致细胞焦亡。本项目拟通过细胞、动物（免疫健全、缺陷小鼠肿瘤模型）和临床（PDX）模型验证抑制ATM提高M1的溶瘤效应，挖掘ATM抑制剂增效病毒的机制；阐明双重抑制DSBs修复、诱导细胞焦亡是增效关键原因。本研究可为DSBs修复作为溶瘤病毒增效靶点的开发提供理论依据，为结直肠癌的联合治疗拓展新思路。

甲病毒作为载体已在病毒性疾病的肿瘤疫苗、核酸疫苗、基因治疗等方面得到了大量应用，这提示M1病毒有成为新型抗肿瘤药物的潜力。本课题聚焦于溶瘤病毒M1开展了两方面的研究：（1）拓展肿瘤治疗谱。我们的前期研究显示溶瘤病毒不同程度的杀伤恶性胶质瘤、结肠癌、肝癌细胞等。本项目探究了溶瘤病毒M1诱导宫颈癌细胞凋亡的作用及机制。（2）探究溶瘤病毒M1的联合治疗方案及其机制。通过小分子化合物库筛选和鉴定出增效剂：ATM抑制剂，然后完成了ATM抑制剂作为M1病毒的增效剂增效效应和体内安全性研究；探索了靶向ATM（抑制剂或CRISPR-Cas9敲除）增效M1病毒溶瘤效应的机制。总而言之，本研究在细胞、小鼠皮下肿瘤、临床原代肿瘤模型上确证抑制ATM加强M1病毒的抗肿瘤作用；阐明ATM抑制剂联合M1病毒会加剧肿瘤细胞DNA损伤，诱导大面积细胞凋亡。本项目的完成，为溶瘤病毒M1的联合治疗拓展新的思路。