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Generation of Hypoimmunogenic Induced Pluripotent Stem Cells for Allogeneic Cell and Tissue Transplantation

Hu, Xiaomeng1,2; Deuse, Tobias1,2,3; Kooremann, Nigel6,7; Malik, Alawi4,5; Wang, Dong1,2; Tediashvili, Grigol1,2; Velden, Joachim8; Schrepfer, Sonja1,2,3

doi: 10.1097/01.tp.0000520292.05650.be
105.6
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1Transplant and Stem Cell Immunobiology (TSI) Laboratory, UCSF, San Francisco, CA, United States; 2Transplant and Stem Cell Immunobiology (TSI) Laboratory, University Heart Center Hamburg, Hamburg, Germany; 3Cardiovascular Surgery, University Heart Center Hamburg, Hamburg, Germany; 4Bioinformatics Service Facility, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; 5Heinrich-Pette Institute, Leibniz Institute for Experimental Virology, Hamburg, Germany; 6Institute of Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, United States; 7Department of Vascular Surgery, Leiden University Medical Center, Leiden, Netherlands; 8Department of Pathology, Marienkrankenhaus, Hamburg, Germany.

Background: The most urgent problem facing transplantation today is the lack of suitable donor organs and tissues. One possible alternative may be cell transplantation with the aim to replace, repair, or enhance the biological function of the diseased organ. Induced pluripotent stem cells are promising candidates for cell-based therapies. Transplanted autologous iPS cell do not cross histocompatibility barriers since such cells are genetically identical to the transplant recipient. However, for acute diseases such as myocardial infarction, treatment options must be readily available, and generating autologous iPSCs for treatment purposes would take too long. Therefore, identifying alternative methods for preventing rejection of iPSCs would be of great interest. In this study, we generated MHC I and MHCII -deficient mouse iPSCs (miPSCs) via disruption of beta-2-microglobulin and C2TA. The antigenicity was investigated in in vivo and in vitro assays.

Methods: IPSCs were generated from C57BL/6 mouse fibroblasts by non-viral minicircle transfection of the transcription factors c-Myc, Sox-2, Klf4 and Oct4. The CRIPSR/Cas9 technology was used to generate β2m and C2TA knockout miPSCs (β2m-C2TA−/− miPSCs). Unmodified and β2m-C2TA −/− miPSCs as well as their derivates were injected in syngeneic C57BL/6 mice and in allogeneic BALB/c mice. Cellular response was quantified and bioluminescence imaging was performed to assess cell survival.

Results: FACS analysis showed a lack of MHC I and MHC II expression on the β2m-C2TA−/− iPSCs but a normal expression in unmodified iPSCs. After allogeneic transplantation into BALB/c mice, β2m-C2TA−/− miPSCs generated a significantly weaker response in both IFNγ (p<0.01) and IL-4 (p<0.01) ELISPOT assays. Teratoma formation occurred in C57BL/6 mice receiving unmodified iPSCs (100%) and BALB/c mice receiving β2m-C2TA−/− iPSCs (100%), whereas BALB/c mice receiving unmodified iPSCs did not form any teratoma (0%). Indeed, also miPSC derivates, such as cardiomyocytes, endothelial and islet cells showed long-term survival after allogeneic transplantation.

Conclusion: Our results clearly demonstrate the hypoimmunogenicity of β2m-C2TA−/− iPSCs after allogeneic transplantation. Thus, MHC I MHC II deficient iPSCs might serve an unlimited cell source for the generation of universally compatible “off-the-shelf” cells grafts or tissues in future clinical applications.

Christiane Pahrmann.

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