Lei Guanglin, Li Baofa, Yang Hao, Sun Fang, Li Donghui, Yan Jin, Wang Yonggang, Li Ruisheng, Liu Honghong, Zhang Shaogeng, Li Yufeng, Yang Penghui
Fifth Medical Center of Chinese PLA General Hospital, Beijing, China.
Department of Liver Disease Kaifeng Infectious Disease Hospital, Henan, China.
Hum Gene Ther. 2022 Mar;33(5-6):309-317. doi: 10.1089/hum.2021.167. Epub 2022 Feb 23.
Oncolytic virus therapy is a promising novel immunotherapy. In this report, we engineered a novel oncolytic influenza virus (IV) carrying an antihuman programmed cell death 1 (PD-1) monoclonal antibody utilizing reverse genetics. A reassortant chimeric IV, named rFlu-huPD1, was synthesized as follows: the heavy chain of the PD-1 antibody was encoded on the PB1 fragment, and the light chain of the PD-1 antibody was encoded on the polymerase acid protein fragment. rFlu-huPD1 antibodies were produced in infected ovalantoic eggs and could replicate to high titers. Moreover, selective cytotoxicity of rFlu-huPD1 was upregulated in multiple hepatocellular carcinoma (HCC) cell lines compared with a control, as determined by a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Furthermore, the activation of T cells in the spleen of tumor-bearing BALB/c mice treated with rFlu-huPD1 was observed, especially cytotoxic CD8 T cell activation . In addition, in a patient-derived xenograft liver cancer mouse model, tumor growth was reduced and the overall survival of the mice was increased by intratumoral injections with rFlu-huPD1 compared with wild-type PR8 virus. Taken together, these findings provide evidence for the utility of a combination of oncolytic IVs expressing PD-1 inhibitors for use in HCC virotherapy.
溶瘤病毒疗法是一种很有前景的新型免疫疗法。在本报告中，我们利用反向遗传学技术构建了一种携带抗人程序性细胞死亡蛋白1（PD-1）单克隆抗体的新型溶瘤流感病毒（IV）。一种名为rFlu-huPD1的重配嵌合IV按如下方式合成：PD-1抗体的重链编码于PB1片段上，而PD-1抗体的轻链编码于聚合酶酸性蛋白片段上。rFlu-huPD1抗体在感染的鸡胚卵中产生，并能复制至高滴度。此外，通过3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐（MTT）试验测定，与对照相比，rFlu-huPD1在多种肝癌（HCC）细胞系中的选择性细胞毒性上调。此外，观察到用rFlu-huPD1治疗的荷瘤BALB/c小鼠脾脏中的T细胞被激活，尤其是细胞毒性CD8 T细胞的激活。此外，在患者来源的异种移植肝癌小鼠模型中，与野生型PR8病毒相比，瘤内注射rFlu-huPD1可减少肿瘤生长并提高小鼠的总体生存率。综上所述，这些发现为表达PD-1抑制剂的溶瘤IV联合用于HCC病毒治疗的效用提供了证据。
