Engineering Thymic Selection to Promote Acceptance of Allogeneic Transplantation

Fabio Russo – San Raffaele Telethon Institute for gene therapy (SR-Tiget), IRCCS Ospedale San Raffale, Milan, Italy; Brendan Foley – Department of Plastic and Reconstructive Surgery, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA; Lotte Van Den Goor – Department of Plastic and Reconstructive Surgery, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA; Silvia Gregori – San Raffaele Telethon Institute for gene therapy (SR-Tiget), IRCCS Ospedale San Raffale, Milan, Italy; Giorgio Raimondi – Department of Plastic and Reconstructive Surgery, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA; Andrea Annoni – San Raffaele Telethon Institute for gene therapy (SR-Tiget), IRCCS Ospedale San Raffale, Milan, Italy

Abstract Text: Achieving donor-specific immune tolerance is critical for graft survival without life-long immunosuppression. Thymic epithelial cells (TEC) are pivotal in central tolerance, selecting a self-tolerant T-cell repertoire. We investigated whether mir142-regulated lentiviral vector (LV)-driven expression of donor-related major histocompatibility complexes (MHC) by recipient TEC generates a “hybrid thymus” capable of reshaping T-cell repertoire, promoting tolerance to donor tissue transplantation. We first demonstrated that intra-thymic (it)-LV-driven MHC expression in MHC knock-out (KO) mice restored the T-cell selection and determined responsiveness to allo-antigens(Ags) of the reconstituted T-cell population. Therefore, it-LV injection enables MHC expression on cortical and medullary TEC, restoring positive and negative selection of thymocytes in MHC KO mice. In immunocompetent recipient Balb/c (H-2d), we removed pre-existing T-cell repertoire by applying a lymphodepletion regimen before it-LV injection of a mix of LV encoding for donor H-2b molecules to obtain the hybrid thymus (H-2db). The transgenic H-2b MHC expression was confirmed by immunostaining by TEC of the recipient Balb/c. Proliferative response to allo-H-2b of the reconstituted T cells isolated from it-LV injected mice evaluated by mixed leukocyte reaction (MLR) assay showed an alloreactivity re-shaped by donor- H-2b MHC expressed in the thymus. These findings demonstrate the potential of it-LV-mediated MHC gene transfer to promote donor-specific central tolerance.