Human CD117 As an Immunotherapeutic Target in preclinical models of Advanced Systemic Mastocytosis

Topic: 24. Gene therapy, cellular immunotherapy and vaccination - Biology & Translational Research Background: Systemic Mastocytosis (SM) is a malignant disease resulting from oncogenically transformed mast cells. Up to 90% of patients harbor the D816V mutation in the KIT-receptor (CD117), leading to constitutive activation of this tyrosine kinase receptor, resulting in abnormal mast cell proliferation and survival. Although the multi tyrosine kinase inhibitors (TKI) Midostaurin and Avapritinib are approved for the treatment of SM with overall response rates (ORR) of 75% for Avapritinib, the median overall survival ranges from 3.5 years (aggressive SM: ASM) to less than six months (mast cell leukemia: MCL). Currently, the only available curative approach is conditioning poly-chemotherapy followed by allogeneic stem cell transplantation (allo-HSCT). Thus, better therapeutic options are needed. Our previous work demonstrated that CD117 (KIT-receptor) positive human AML are efficiently eradicated by anti-CD117 CAR T-cells in vitro and in vivo (Myburgh et al., Leukemia 2020). Aims: This study aims at employing CAR T-cells targeting human CD117 in preclinical models of SM and testing the efficacy of this approach in vitro and in vivo as a novel therapy option for SM. Methods: 1. Mast cell lines (with and without the oncogenic driver mutation KIT D816V), 2. SM patient-specific iPS cell-derived mast cells (KIT D816V positive), and 3. SM patient-derived mast cells were in vitro co-cultured for up to three days with anti-CD117 CAR T-cells. CAR T-cell activation markers, cytokine production and proliferation were assessed. Mast cell line (ROSAKIT D816V) engrafted immunodeficient mice (NSG) were treated with anti-CD117 CAR T-cells or untransduced T-cells (control). In vivo live imaging was performed, and peripheral blood, bone marrow, liver and spleen were terminally analyzed for human mast cell infiltration. Results: After 24h of in vitro co-culturing, the tumor cells were effectively killed by CAR T-cells compared to the control. Activation of CAR T-cells in the presence of CD117-bearing cells led to T-cell proliferation and production of proinflammatory and activation-stimulating cytokines (e.g., IL-2). Tumor cells were efficiently controlled by anti-CD117 CAR T-cells for up to 28 days of follow-up. Sequential injection of anti-CD117 CAR T-cells in ROSAKIT D816V engrafted mice led to a substantial in vivo tumor burden reduction and increased survival of ROSAKIT D816V-engrafted NSG mice. Summary/Conclusion: In conclusion, we demonstrate in preclinical SM models the efficient targeting and killing in vitro and in vivo by CAR T-cells directed against human CD117. Given that CD117 is expressed on healthy hematopoietic stem and progenitor cells (HSPCs) on a substantially lower level, there might be a therapeutic window for anti-CD117 immunotherapy in advanced forms of mastocytosis. However, as CAR T-cells are highly efficient, collateral damage on healthy HSPCs will likely need to be compensated by subsequent HSC transplantation. Our results open up future opportunities for clinical applications of CAR T-cell immunotherapy, broadening the armamentarium of therapeutic options in the so far very limited field of SM. Keywords: Systemic mastocytosis, c-kit, CAR-T, Immunotherapy