A single fraction of 2 Gy irradiation was sufficient to enhance the in vitro cytotoxicity of NKG2D CAR-T cells toward ESCC cells, while 8 Gy irradiation induced higher susceptibility of ESCC cells to NKG2D CAR-T cells attack compared to 2–4 Gy irradiation. Here, KLRK1 is linked to esophageal squamous cell carcinoma.