Associate Professor The Ohio State University Columbus, Ohio, United States
Introduction: Multiple myeloma (MM) is a bone marrow-resident hematological malignancy of plasma cells and the second most diagnosed blood cancer. Despite the development of myeloma-targeted immunotherapies, MM has remained largely incurable. It is well documented that a suppressive bone marrow tumor microenvironment (TME), including dysfunctional natural killer (NK) cells, can cause the failure of current immunotherapies such as immune checkpoint blockade therapy (ICBT). The objective of this research is to target tissue factor as a new MM target and test corresponding chimeric antigen receptor-modified NK (CAR-NK) cell therapy as a monotherapy and in combination therapy to overcome the suppressive TME to enhance ICBT efficacy in MM.
Methods: To validate TF as a new target in MM, we examined its expression on human MM lines (U266.B1, MM.1S, OPM2 and IM9) using flow cytometry. To target TF for CAR-NK cell immunotherapy, we developed TF-targeted human CAR constructs, which are composed of human fVII light chain followed sequentially by a hinge region of human IgG1, CD28 transmembrane and cytoplasmic domains and then by the cytoplasmic domains of 4-1BB and CD3ΞΆ (TF-CAR dimer). To test the in vivo efficacy of TF CAR-NK cells as a monotherapy, we generated subcutaneous xenograft mouse models of human MM (U266.B1) and treated the animals with TF CAR-NK cells and control NK cells. To test the efficacy of TF CAR-NK cells in a novel combination therapy with ICBT (anti-PD-1 antibody, Ab), we first generated human MM xenografts in immunodeficient NSG mice (to mimic the suppressive TME) and then treated the animals with TF CAR-NK cells and anti-PD-1 Ab, TF CAR-NK cells plus IgG isotype control and control NK cells plus IgG isotype control. Whole body weights were recorded and blood chemistry from terminal blood samples were assayed as part of safety studies.
Results: We showed and verified tissue factor expression in all four examined human MM cell lines, of which U266.B1 has a relatively higher expression level. After verifying TF expression in MM lines, we showed that TF CAR-NK cells alone are effective and safe for the treatment of human MM in preclinical mouse models. Furthermore, we showed that TF CAR-NK cells are more effective in combination therapy with ICBT (anti-PD-1 Ab) than TF CAR-NK cell alone in preclinical mouse models. There was no significant difference in whole body weights and blood chemistry panels between treated and control animals, suggesting that TF CAR-NK cells are safe.
Conclusions: This study established the proof of concept of targeting TF as a new target in CAR-NK immunotherapy for effective and safe treatment of MM as a monotherapy and in a novel combination therapy with ICBT and may warrant further preclinical study and potentially future investigation in MM patients.
