PD-L1过表达对CLL-1 CAR-T细胞抗急性髓系白血病作用的影响

OBJECTIVE: To investigate the effects of programmed death receptor ligand 1（PD-L1）on CLL-1 CAR-T against acute myeloid leukemia（AML）. METHODS: In this experiment, the PD-L1 expression vector was constructed, and then the lentivirus vector was packaged by three-plasmid packaging system. THP-1 monoclonal cell lines stably expressing PD-L1 were set up. The CLL-1 CAR-T was developed by our team, as the effector cell for co-culture with the THP-1 or THP1-PDL1 cell lines, respectively. Then, the LDH was tested using the kit, the supernatant cytokine was detected by CBA, and the CLL-1 CAR-T cell proliferation was demonstrated by flow cytometry（FCM）with CSFE labeled. RESULTS: ①The PD-L1 lentivirus vector was successfully constructed, and monoclonal cell lines of THP-1 with stable PD-L1 was set up and verified by FCM and PCR. ②The overexpression of PD-L1 inhibited CLL-1 CAR-T's ability to lyse THP-1 cells（E∶F ratio 10∶1）; the killing efficiency of CLL-1 CAR-T on THP1-PDL1 cells was lower than that of THP-1 cells［（15.70±9.90）％ vs（51.95 ± 2.52）％, P<0.05］. ③The overexpression of PD-L1 decrease the release of cytokine［THP1-PDL1 group vs THP-1 group: IFN-γ（115.66±3.13）pg/ml vs（1708.16±26.76）pg/ml, P<0.05; IL-6（17.37±0.72）pg/ml vs（124.92±4.26）pg/ml, P<0.05; IL-10（5.69±0.13）pg/ml vs（124.12±3.02）pg/ml, P<0.05］. Additionally, the proliferation of CLL-1 CAR-T was also inhibited. CONCLUSION: Monoclonal cell lines of THP-1 with stable PD-L1 expression were successfully constructed, and the adverse effect of PD-L1 overexpression on CLL-1 CAR-T anti-AML was confirmed, which provided a theoretical basis for the regulation of CLL-1CAR-T through the PD-1/PD-L1 pathway.