Clinical development of tacrolimus-resistant regulatory T cells to enable simultaneous immunosuppression and immune regulation.
Zarrinrad G., Burkhardt L-M., Beltran-Mestres C., Romero-Suárez S., Wagner DL., Schlickeiser S., Stein M., Wendering DJ., Carrera Diaz de la Cebosa IJ., Ehlen L., Hamm F., Kurgan GL., Durek P., Heinrich F., Giri A., Pu Y., Ou K., Hoffmann H., Muench S., Stuewe I., Dolling S., McCallion O., Kaeda J., Kath J., Schulenberg S., Wiese N., Nazari A., Peter L., Picht S., Sánchez-Peña A., Usman O., Brommel C., Turk R., Rettig G., Sturgeon M., Osborne TL., Jacobi A., Friedrich R., Kaufmann MM., Klermund J., Fink S., Templin MF., Roemhild A., Kaiser D., Klein O., Cathomen T., Mashreghi M-F., Issa F., Polánsky JK., Volk H-D., Schmueck-Henneresse M., Reinke P., Amini L.
Unwanted alloimmune responses are a central driver of solid organ transplant rejection and currently managed with life-long immunosuppression, which imposes substantial risks and burdens on patients. Adoptive transfer of regulatory T cells (Tregs) offers a strategy to restore immunological balance and reduce long-term adverse effects of generalized immunosuppression. However, although Tregs can potently inhibit incipient immune activation, they struggle to suppress established memory effector T cells, necessitating the continued use of immunosuppression. Calcineurin inhibitors such as Tacrolimus effectively control both, newly activated and pre-existing effector T cells, but unfortunately, they also impair Treg function. Therefore, we hypothesized that gene-editing of Tregs inducing tacrolimus resistance (FKBP12KO) would enable combined therapy that curbs effector T cell responses without compromising Treg efficacy. Here, we developed FKBP12KO-Tregs using a ribonucleoprotein-based CRISPR-Cas9 approach and characterized them extensively in vitro. FKBP12KO-Tregs retained phenotype, high viability, and suppressive function comparable to unedited TregWT, and they remained functionally impervious to Tacrolimus, while preserving sensitivity to alternative CNIs. We additionally established a good-manufacturing practice process for FKBP12KO-Tregs. Comprehensive in vitro phenotypic, functional, and molecular characterization, together with the established manufacturing, provide the rationale for a proof-of-concept clinical trial assessing the feasibility and safety of co-administration of FKBP12KO-Tregs with Tacrolimus in living-donor kidney transplant recipients.