Conformal external beam radiation increases B7-H3 expression in a murine model of pancreatic cancer liver metastases and promotes CAR T cell function

Background: Pancreatic ductal adenocarcinoma (PDAC) is a lethal disease characterized by early liver metastases. Chimeric antigen receptor (CAR) modified cells (T and NK) have been largely unsuccessful in PDAC and other solid tumors, despite success in hematologic malignancies. Limitations to CAR therapy success in solid tumors include trafficking, engraftment, and antigen loss. Liver-directed radiation (RT) may improve CAR therapy through altering the tumor and tumor microenvironment. We investigated the impact of liver-directed radiation therapy (RT) to augment B7-H3 CAR T cell homing and function in a xenogeneic model of PDAC liver metastases. Methods: We evaluated dose-dependent expression of B7-H3 and NKG2D activating ligands by flow cytometry in human PANC-1 and AsPC-1 cells in vitro and in vivo after RT, including a liver-only metastatic model using a hemi-spleen implantation technique to seed the liver. B7-H3 CAR T cells were generated and assessed by flow cytometry. B7-H3 CAR T cells were then co-cultured with RT-treated target cells, and cytotoxicity was analyzed using Incucyte live cell imaging. Results: Following RT in vitro , PANC-1 and AsPC-1 cells exhibited significant increase in expression of both B7-H3 and NKG2D ligands MICA/B by either median fluorescent intensity or percent expression. Liver-directed RT led to significantly increased expression of B7-H3 and MICA/B on PANC-1 liver metastases compared to untreated mice. Percent killing by B7-H3 CAR T cells against PANC-1 and AsPC-1 cells was significantly increased when target cells were pre-treated with 4 Gy RT prior to killing assay. Conclusions: RT increases the expression of both B7-H3 and the NKG2D ligands MICA/MICB on pancreas cancer cell lines and this is recapitulated in vivo following liver-directed RT to metastasis-bearing mice. Pre-treating target cells with RT results in greater B7-H3 CAR T cell killing in vitro . Liver-directed RT may be a strategy to better target PDAC liver metastases with CAR cell therapy.