Experimental models of CAR-redirected cellular therapy with CIK lymphocytes against ovarian cancer peritoneal carcinomatosis

Background Peritoneal carcinomatosis (PC) from ovarian cancer (OC) has poor prognosis and is in need for optimized therapies. Such a clinical condition displays a composite structure with floating cell aggregates with-in ascites and solid-like masses invading the peritoneum. A deep understanding of the dynamics is thus crucial to optimize the therapeutic strategies and knowledge. Chimeric antigen receptor (CAR)-based immunotherapies are among the most suited treatments for the compartmentalized pathophysiology of PC. The ex vivo expanded cytokineinduced killer lymphocytes (CIK), intrinsically endowed with HLA-independent antitumor activity, are a valid platform to be exploited against solid tumors. However, appropriate preclinical models recapitulating the clinical hurdles are needed to better tailor the clinical translation and optimize currently available therapies. Methods Here, we explored the activity of mesothelin (MSLN) CAR-redirected CIK (MSLN-CAR.CIK) against advanced OC, by developing suitable 3D models resembling the complexity of peritoneal metastasis, where 3D mOC spheroids were co-cultured with MSLN-CAR.CIK, either floating in liquid medium or embedded in a 3D hydrogel. To measure MSLN-CAR.CIK activity we employed imaging-based methods. Results MSLN-CAR.CIK resulted functional in both 3D biological models recapitulating the floating as well as solid component of OC PC. Specifically, CAR.CIK were found to localize faster than NTD.CIK in both 3D liquid and solid settings. Recruitment kinetics of CAR.CIK on mOC aggregates in liquid was significantly faster when coupled to fluid flow in shaking cultures, reminiscent of spontaneous patient movement, than in absence of flow, coherently with the relative increase in cytotoxicity. Even in the 3D solid setting, CAR.CIK were functionally more efficient on mOC spheroids than NTD.CIK. However, the solid setting was characterized by slower kinetics and significantly influenced by the distance traveled by the effectors. CAR.CIK antitumor activity was found to be comparable to the liquid condition. Our data indicate that the kinetics of recruitment should therefore be considered when assessing CAR.CIK killing. Conclusions Our findings provide therapeutically relevant information on intraperitoneal approaches with CAR.CIK, supporting further developments and improvements for clinical studies in the context of locoregional cell-therapy approaches for patients with PC from OC.