Oncolytic virus therapy is a novel and promising approach in tumor immunotherapy. We have developed the oncolytic virus OH2 and completed its preclinical studies. This oncolytic virus is currently undergoing clinical trials in China (NMPA, 2018L02743) and the United States (FDA IND 27137) targeting various solid tumors. One of the main mechanisms by which the oncolytic virus exerts its antitumor effect is through the direct killing of tumor cells. In our previous studies, we found that the structural protein VP5 of oncolytic virus OH2 can induce apoptosis in A549 cells via the TP53I3 molecule. This study further investigates the solid tumor cell lines involved in the clinical trials of oncolytic virus OH2. We found that VP5 protein exhibits different functions depending on the tumor cell type. Stable expression of VP5 protein enhanced the cytotoxicity of UV-OH2 against BGC823, LoVo, and CT26 cells, while inhibiting its cytotoxicity against 4T1 and HeLa cells. To explore the differences in the cytotoxicity induced by VP5 protein in different tumor cells, we performed transcriptome sequencing on 4T1-VP5 cells and OH2-stimulated 4T1-VP5 cells, comparing them with 4T1 cells and OH2-stimulated 4T1 cells, respectively. We also compared the transcriptome sequencing results with those of A549 cells from previous studies. Through this, we identified nectin-3 as a differentially expressed gene associated with tumor cell killing induced by VP5 protein. Further studies on nectin-3 revealed that the structural protein VP5 of oncolytic virus OH2 regulates the expression of nectin-3 in tumor cells, thereby promoting or inhibiting tumor cell apoptosis.