DNA-PKcs Inhibition Extends Allogeneic Skin Graft Survival

Abstract

Background. Organ transplantation is life-saving and continued investigations into immunologic mechanisms that drive organ rejection are needed to improve immunosuppression therapies and prevent graft failure. DNA-dependent protein kinase catalytic subunit, DNA dependent-protein kinase catalytic subunit (DNA-PKcs), is a critical component of both the cellular and humoral immune responses. In this study, we investigate the contribution of DNA-PKcs to allogeneic skin graft rejection to potentially highlight a novel strategy for inhibiting transplant rejection. Methods. Fully MHC mismatched murine allogeneic skin graft studies were performed by transplanting skin from BalbC mice to C57bl6 mice and treating with either vehicle or the DNA-PKcs inhibitor NU7441. Graft rejection, cytokine production, immune cell infiltration, and donor-specific antibody formation were analyzed. Results. DNA-PKcs inhibition significantly reduced necrosis and extended graft survival compared with controls (mean survival 14 d versus 9 d, respectively). Inhibition reduced the production of the cytokines interleukin (IL)-2, IL-4, IL-6, IL-10, TNF-α, and IFN-γ and the infiltration of CD3+ lymphocytes into grafts. Furthermore, DNA-PKcs inhibition reduced the number of CD19+ B cells and CD19+ CD138+ plasma cells coinciding with a significant reduction in donor-specific antibodies. At a molecular level, we determined that the immunosuppressive effects of DNA-PKcs inhibition were mediated, in part, via inhibition of nuclear factor kappa-light-chain-enhancer of activated B cells signaling through reduced expression of the p65 subunit. Conclusions. Our data confirm that DNA-PKcs contributes to allogeneic graft rejection and highlight a novel immunologic function for DNA-PKcs in the regulation of nuclear factor kappa-light-chain-enhancer of activated B cells and concomitant cytokine production.

DNA-PKcs kinase activity stabilizes the transcription factor Egr1 in activated immune cells

“…For instance, our previous study indicates that DNA-PKcs is a critical regulator of the T cell response to allogeneic antigens. 13 Loss of DNA-PKcs activity prevented T cells from producing a host of inflammatory cytokines in response to alloantigen recognition resulting in reduced graft rejection in vivo. This suggests it may be a prime therapeutic target for the prevention of transplant rejection.…”

“…11,12 Our laboratory recently reported that DNA-PKcs also controls expression of the p65 subunit of NF-κB in activated T cells and loss of DNA-PKcs activity significantly reduces expression of NF-κB target genes including Interleukin (IL)-6. 13 Ferguson et al determined that following viral DNA detection, DNA-PKcs drives activation of the innate immune response by directly binding the transcription factor interferon (IFN) regulator factor-3 (Irf-3) and promoting its translocation into the nucleus to induce cytokine gene expression. 14 Similarly, our studies indicate that DNA-PKcs plays a pivotal role in the calcineurinmediated translocation of NFAT to the nucleus.…”

“…In CD4+ T cells, following TCR activation, DNA-PKcs regulates expression of both T-bet and Gata3 highlighting it as a master regulator of Th1 and Th2 differentiation ( 7 , 11 ). Our laboratory recently reported that DNA-PKcs also controls expression of the p65 subunit of NF-κB in activated T cells, and loss of DNA-PKcs activity significantly reduces expression of NF-κB target genes including Interleukin (IL)-6 ( 12 ). Ferguson et al .…”

Cyclic Helix B Peptide Prolongs Skin Allograft Survival via Inhibition of B Cell Immune Responses in a Murine Model

“…The differentiation of B cells into plasma cells to secrete DSAs is a critical component of the development of AMR and a major risk factor for allograft rejection (22)(23)(24). Given that the percentage of plasma cells decreased, the DSA levels were subsequently assessed.…”

DNA-PKcs kinase activity stabilizes the transcription factor Egr1 in activated immune cells

“…11,12 Our laboratory recently reported that DNA-PKcs also controls expression of the p65 subunit of NF-κB in activated T cells and loss of DNA-PKcs activity significantly reduces expression of NF-κB target genes including Interleukin (IL)-6. 13 Ferguson et al determined that following viral DNA detection, DNA-PKcs drives activation of the innate immune response by directly binding the transcription factor interferon (IFN) regulator factor-3 (Irf-3) and promoting its translocation into the nucleus to induce cytokine gene expression. 14 Similarly, our studies indicate that DNA-PKcs plays a pivotal role in the calcineurin-mediated translocation of NFAT to the nucleus.…”