Hematopoietic chimerism achieved by bone marrow transplantation (BMT) induces donor-specific tolerance to solid organ grafts. We previously demonstrated that by targeting recipient immune cells (T, NK cells and macrophages) total body irradiation (TBI) can be reduced to safe and clinically acceptable levels in a mouse BMT model. Therefore, we hypothesized that various immune signalling pathways may be novel targets for mixed chimerism as an alternative to immune cell depletion. B6 (H2^b) mice were nonmyeloablatively conditioned with 300cGy TBI (day 0) and cyclophosphamide (CyP, day +2) in combination with different pathway inhibitors (day 0 to +9). 15 x 10⁶ bone marrow cells from MHC-disparate BALB/c (H2^d) mice were transplanted on day 0. With 300cGy/CyP, 100% engraftment only occurred in recipients treated with JAK3/1/2 inhibitor [tofacitinib (Tofa)], but not with inhibitors of MEK, BTK, PI3K/mTOR, or S1P/S1P1. The JAK/STAT pathway serves a fundamental role in regulation of cell growth and differentiation by transducing signals from extracellular cytokines to their transmembrane receptors, and then to the nucleus. With Tofa, 100% and 58.3% of recipients engrafted with 200 and 100 cGy TBI plus CyP, respectively. Donor chimerism was 57.7 ± 10.8%, 22.7 ± 14.5%, and 3.8 ± 1.2% with 300, 200, 100cGy TBI plus Tofa/CyP at 1 month, respectively. When host T cells were depleted with anti-αβTCR in addition to Tofa/CyP, 100% engraftment regained with 100cGy TBI. The level of donor chimerism with Tofa/anti-αβTCR/100cGy/CyP was significantly higher at 1 month post BMT (8.9 ± 8.7%, P < 0.0001) compared to Tofa/100cGy/CyP conditioning. Engrafted animals exhibited multilineage cell production, including donor T, B, NK cells, macrophages, and granulocytes. Chimerism remained stable up to 6 months. Moreover, specific Vβ5.[frac12] and Vβ11 clonal deletion was detected in host T cells in all chimeric mice, indicating central tolerance to donor alloantigens. Targeting JAK3/1/2 with Tofa can decrease TBI dose to a minimum to achieve mixed chimerism. Tofa and T cell depletion have a synergistic effect to enhance alloengraftment, suggesting that this JAK inhibitor targets cells other than T cells, including innate immune cells. Our study may have a significant impact on the development of immune-based nonmyeloablative conditioning strategies for tolerance induction.

CITATION INFORMATION: Xu H, Merchak A, Xu M, Mort C, Dowling T, Kahn L, Binion M, Chhabra A, Wen Y, Huang Y, Ildstad S. Targeting JAK-STAT Pathways with Tofacitinib Provides a Novel Approach for Conditioning in Allogeneic BMT Recipients. Am J Transplant. 2017;17 (suppl 3).

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