Targeting CD20⁺ aggressive B-cell non-hodgkin lymphoma by Anti-CD20 CAR mRNA-modified expanded natural killer cells in vitro and in NSG Mice

The prognosis is very dismal for patients with relapsed CD20⁺ B-cell non-Hodgkin lymphoma (B-NHL). Facilitating the development of alternative novel therapeutic strategies is required to improve outcomes in patients with recurrent/refractory CD20⁺ B-NHL. In this study, we investigated functional activities of anti-CD20 CAR-modified, expanded peripheral blood NK cells (exPBNK) following mRNA nucleofection against CD20⁺ B-NHL in vitro and in vivo. CAR⁺ exPBNK had significantly enhanced in vitro cytotoxicity, compared with CAR^- exPBNK against CD20⁺ Ramos (P < 0.05), Daudi, Raji, and two rituximab-resistant cell lines, Raji-2R and Raji-4RH (P< 0.001). As expected, there was no significant difference against CD20^- RS4;11 and Jurkat cells. CD107a degranulation and intracellular IFNγ production were also enhanced in CAR⁺ exPBNK in response to CD20⁺ B-NHL-specific stimulation. In Raji-Luc and Raji-2R-Luc xenografted NOD/SCID/g-chain^-/-(NSG) mice, the luciferase signals measured in the CAR⁺ exPBNK-treated group were significantly reduced, compared with the signals measured in the untreated mice and in mice treated with the CAR^- exPBNK. Furthermore, the CAR exPBNK-treated mice had significantly extended survival time (P < 0.001) and reduced tumor size, compared with those of the untreated and the CAR^- exPBNK-treated mice (P < 0.05). These preclinical data suggest that ex vivo-exPBNK modified with anti-CD20 CAR may have therapeutic potential for treating patients with poor-risk CD20⁺ hematologic malignancies.