A Clinical Study Of NKG2D Chimeric Antigen Receptor Engineered Natural Killer Cells In Treating Colorectal Cancer Patients

Objectives:Colorectal cancer is one of the most common malignant tumors in the world,and its morbidity and mortality are among the top three.About 50% of patients have distant metastasis,and the five-year survival rate of patients with stage ? colorectal cancer is less than 10%.Although Chimeric antigen receptor(CAR)T cells have achieved significant efficacy in the treatment of hematological cancer,CAR-T cells are still difficult to overcome solid tumor,and have risks of severe cytokine storm,neurotoxicity and so on.Natural killer(NK)cells are superior to T cells in safety and accessibility,so we intend to use NK cells modified with NKG2 D CAR to evaluate the safety and feasibility of NKG2 D CAR-NK cells in the treatment of patients with advanced colorectal cancer.Methods:We adopted an mRNA electroporation approach to modify NK cells with NKG2 D mRNA CAR,and used Flow cytometric analysis,IFN-? ELISPOT assay and cytotoxicity assay to detect the tumor killing activity of NKG2 D CAR-NK cells in vitro.NSG mice were inoculated via i.p.injection of HCT116-luc cells to generate tumor models.The anti-tumor effect of NKG2 D CAR-NK cells in vivo was detected by whole body bioluminescent imaging and Kaplan-Meier analysis of survival.Furthermore,a single-arm,open label,dose-escalation and single-center pilot study(NCT03415100)was conducted to evaluate the safety and feasibility of NKG2 D CAR-NK cell therapy.Three patients,who were failed to be cured by standard treatment,with peritoneal metastatic colorectal cancer were enrolled.Results:It was found that NKG2D-DAP12 CAR-NK cells showed more robust IFN-? secretion and cytotoxic immune response than NKG2D-CD3? CAR-NK cells.Besides,in the human colorectal cancer-bearing NSG mouse model,the mice receiving NKG2 Dp CAR-modified NK cells were significantly protected from rapid tumor progression,and the median survival time of these mice was longer than that of the control group.Subsequently,3 patients with metastatic colorectal cancer were treated by local infusion of NKG2 Dp CAR-NK.For the first two patients,after intraperitoneal infusion of low-dose CAR-NK cells,it was observed that the production of ascites slowed down and the number of tumor cells in ascites samples decreased sharply.The third case of liver metastases patient was treated with CAR-NK cells by ultrasound-guided percutaneous injection and intraperitoneal infusion.After that,rapid tumor regression in the liver region was observed by Doppler ultrasound imaging and complete metabolic response in the treated liver lesions was confirmed by positron emission tomography-computed tomographic(PET-CT)scanning.No grade 3 or more serious adverse events occurred in all the three patients during the treatment.Conclusion:The results of this study show that local injection of NKG2 D mRNA CAR-NK cells is safe and feasible in the treatment of patients with metastatic colorectal cancer,which provides a new strategy for cancer immunotherapy.