由于肿瘤浸润有限和免疫抑制的微环境，嵌合抗原受体T细胞（CAR-T）对实体瘤的疗效仍然不尽人意。为了克服这些局限性，开发了能够在肿瘤细胞中有条件地复制的基因工程重组溶瘤腺病毒（OAVs），用于改变肿瘤微环境（TME），以促进CAR-T介导的肿瘤消灭。在本研究中，构建了一种新的重组OAV，该OAV携带由Ki67启动子控制的CCL5、IL12和IFN-γ（命名为AdKi67-C3）。通过单独给药或与靶向B7H3的CAR-T细胞相结合，测试了AdKi67-C3的抗肿瘤活性。结果表明，由AdKi67-C3表达的CCL5在体外诱导了更多的CAR-T细胞迁移，并在体内促进了CAR-T细胞在肿瘤块中的浸润。同时，由AdKi67-C3感染的肿瘤细胞分泌的IFN-γ和IL12显著促进了CAR-T细胞在体外和体内的增殖和持久存在。在肾癌、前列腺癌或胰腺癌的移植瘤小鼠模型中，局部预处理AdKi67-C3显著增强了CAR-T细胞的疗效，消除了局部和远处的肿瘤。更重要的是，完全肿瘤消退的小鼠对同一肿瘤细胞的再挑战表现出抵抗，提示建立了长期的抗肿瘤免疫反应。因此，携带细胞因子的OAVs可作为生物增效剂的开发，以克服免疫抑制的微环境，并改善CAR-T对实体瘤的治疗效果。© 2023. 细胞死亡分化协会（ADMC）。

The efficacy of chimeric antigen receptor T (CAR-T) cells for solid tumors remains unsatisfactory due to the limited tumor infiltration and immunosuppressive microenvironment. To overcome these limitations, the genetically engineered recombinant oncolytic adenoviruses (OAVs) that conditionally replicate in tumor cells were developed to modify the tumor microenvironment (TME) to facilitate CAR-T-mediated tumor eradication. Here in the present study, a novel recombinant OAV carrying CCL5, IL12, and IFN-γ controlled by Ki67 promoter was constructed (named AdKi67-C3). The antitumor activity of AdKi67-C3 was tested in vitro and in vivo by using mono administration or combing with CAR-T cells targeting B7H3. It proved that CCL5 expressed by AdKi67-C3 indeed induced more CAR-T migration in vitro and CAR-T infiltration in tumor mass in vivo. Meanwhile, cytokines of IFN-γ and IL12 secreted by AdKi67-C3-infected tumor cells significantly promoted proliferation and persistence of CAR-T cells in vitro and in vivo. In tumor-bearing xenograft mouse models of kidney, prostate or pancreatic cancer, local pretreatment with AdKi67-C3 dramatically enhanced CAR-T cell efficacy and eliminated local and distant tumors. More importantly, mice achieving complete tumor regression resisted to re-challenge with the same tumor cells, suggesting establishment of long-term antitumor immune response. Therefore, OAVs armored with cytokines could be developed as a bioenhancer to defeat the immunosuppressive microenvironment and improve therapeutic efficacy of CAR-T in solid tumors.© 2023. Cell Death Differentiation Association (ADMC).