44: Successful Transplantation of Human Muscle Precursor Cells in Nonhuman Primates Using Tacrolimus Immunosuprpression: A New Model for the Preclinical Test of Other Potentially Myogenic Human Cells

Background Different cells isolated from human tissues were proposed for cell transplantation in the skeletal muscle as a treatment of myopathies, essentially muscular dystrophies. However, the myogenic properties of most of these cells were found only after xenotransplantation in immunodeficient mice. Given that cell transplantation protocols developed in nonhuman primates (NHPs) were better extrapolated to humans than those that were not verified in this model, and considering that NHPs are crucial in preclinical transplantation research, we wanted to test the feasibility of xenotransplant human myogenic cells in NHP muscles. Methods Human CD56+ muscle precursor cells (MPCs) were transduced with the LacZ gene using a replication defective retroviral vector. They were injected into muscle regions of 1 cm3 (around 25 x 106 viable cells per site) in four cynomolgus macaques immunosuppressed with oral tacrolimus (Advagraf) and dexamethasone. Allogeneic LacZ-labeled MPCs were transplanted similarly in other muscles as a positive control for cell engraftment. Cell-grafted regions were sampled 1 month later and analyzed by histology in cryostat serial sections using ß-galactosidase (ß-Gal) histochemical detection, hematoxylin and eosin stain, and CD8 immunodetection. Blood was taken before transplantation and before muscle sampling to detect antibodies against the grafted cells. Tacrolimus blood levels were quantified by liquid chromatography tandem mass spectrometry. Results Tacrolimus blood levels and Advagraf doses in the monkeys of the study are shown in figure 1.{{AbstractFigure.1}} In monkeys #1 and #2 we targeted tacrolimus blood levels over 40 µg/L, while in monkeys #3 and #4 we targeted levels closer to those used in humans (around 20 µg/L). Abundant ß-Gal+ myofibers were found in all regions grafted with human MPCs (an average of around 25/mm2) distributed in bands aligned according to the injection trajectories (figure 2, red arrows indicate the original sense of the cell injections). The histological analyzes showed absence of specific cellular immune responses in three monkeys and minimal focal lymphocytic infiltrates in the monkey that had the lowest tacrolimus blood levels. Similar patterns of ß-Gal+ myofibers were observed in all regions grafted with cynomolgus MPCs (figure 2), in the absence of specific immune responses. Anti-donor antibodies were not detected in the sera. Conclusions We demonstrated that human MPCs can form hybrid myofibers in NHP muscles and that a conventional tacrolimus-based immunosuppression is sufficient to control rejection in this case. This opens the door to NHP studies with other human cells in which myogenic properties were found by xenotransplantation in immunodeficient mice, validating the myogenicity of these cells in a more appropriate model than mice for clinical translation and investigating the administration parameters necessary in humans. This work was supported by a grant of the Jesse’s Journey Foundation for Gene and Cell Therapy of Canada to D.S. and a grant of the Canadian Institutes of Health Research to J.P.T.

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