FGFR4-targeted chimeric antigen receptors (CARs) combined with anti-myeloid polypharmacy effectively treats orthotopic rhabdomyosarcoma

Rhabdomyosarcoma (RMS) is the most common soft tissue cancer in children. Treatment outcomes, particularly for relapsed/refractory or metastatic disease, have not improved in decades. The current lack of novel therapies and low tumor mutational burden suggest that CAR T therapy could be a promising approach to treating RMS. Previous work identified Fibroblast Growth Factor Receptor 4 (FGFR4, CD334) as being specifically upregulated in RMS, making it a candidate target for CAR-T cells. We tested the feasibility of an FGFR4 targeted CAR for treating RMS using an NSG mouse with RH30 orthotopic (intramuscular) tumors. The first barrier we noted was that RMS tumors produce a collagen-rich stroma, replete with immunosuppressive myeloid cells, when T cell therapy is initiated. This stromal response is not seen in tumor-only xenografts. When scFV-based binders were selected from phage display, CARs targeting FGFR4 were not effective until our screening approach was refined to identify binders to the membrane-proximal domain of FGFR4. Having improved the CAR, we devised a pharmacologic strategy to augment CAR-T activity by inhibiting the myeloid component of the T cell-induced tumor stroma. The combined treatment of mice with anti-myeloid polypharmacy (targeting CSF1R, IDO1, iNOS, TGFbeta, PDL1, MIF and myeloid misdifferentiation) allowed FGFR4 CAR-T to successfully clear orthotopic RMS tumors, demonstrating that RMS tumors, even with very low copy number targets, can be targeted by CAR-T upon reversal of an immunosuppressive microenvironment.