Effects and mechanism of recombinant oncolytic adenovirus expressing soluble programmed cell death protein 1 on proliferation and apoptosis of B16/F10 cell

Objective To construct a recombinant oncolytic adenovirus expressing soluble programmed cell death protein 1 (sPD1) and investigate its effects and mechanism on proliferation and inhibition of B16/F10 cells. Methods The recombinant oncolytic adenovirus Ad5-sPD1 was constructed, and infected into B16/f10 cells. The quantitative fluorescent polymerase chain reaction was used to detect the viral replication. MTT assay was applied to examine the inhibition of B16/F10 cells. The viruses were injected subcutaneously into the mouse B16/F10 tumor model to study the antitumor effect of Ad5-sPD1 in vivo. EliSpot technique was exployed to test the number of tumor proliferative lymph nodes. Results After viral treatment of B16/F10 cells for 6, 24, 36, 48, 60 and 72 h, the copies fold increases in the Ad5 treatment group were respectively 0.99±0.10, 1.12±0.05, 1.57±0.15, 3.59±1.03, 6.26±0.74, 9.69±1.46, and those in the Ad5-sPD1 treatment group were respectively 1.00±0.104, 0.99±0.08, 1.05±0.03, 3.56±1.04, 6.04±0.38, 11.05±1.12. There was no significant difference between the two groups (P=0.754). After treating B16/F10 cells for 72 h with multiplicity of infection (MOI) of 0, 5, 10, 15 and 20, the cell viability of Ad5 treatment group was respectively (100.00±6.39)%, (101.60±5.21)%, (98.66±7.31)%, (93.50±4.54)%, (73.16±2.45)%, and that in the Ad5-sPD1 treatment group was respectively (100.00±6.38)%, (97.10±6.37)%, (96.96±7.44)%, (95.67±7.46)%, (74.49±4.14)%. There was no significant difference between the two groups (P=0.779). After treating mice model of B16/F10 subcutaneous tumor with Ad5 and Ad5-sPD1 viruses, tumor volume was respectively 3 638±1.823, 2 603±336 and 1 097±532 at the termination of the experiments with the difference being significant (Mock and Ad5: P=0.044; Mock and Ad5-sPD1: P=0.000; Ad5 and Ad5-sPD1: P=0.000). After respectively injecting mice model of B16/F10 subcutaneous tumor with Mock, Ad5 and Ad5 -sPD1 viruses, the number of IFN-γ spots was respectively 19.4±7.75, 149.8±25.18 and 314±32.34 with the the difference being significant (Mock and Ad5: P=0.000; Mock and Ad5-sPD1: P=0.000; Ad5 and Ad5-sPD1: P=0.000). Conclusion Recombinant oncolytic adenovirus expressing PD1 gene can significantly improve the anti-tumor effect of the oncolytic adenovirus by blocking PD-1/PD-L1 inhibitory pathway. Key words: Oncolytic adenovirus; Tumor; Immune response; Programmed cell death protein 1