Thrombopoietin-Increased DNA-PK-Dependent DNA Repair Limits Hematopoietic STEM and Progenitor CELL Mutagenesis in Response to Irradiation

Abstract 3447

Radiation-induced double-strand breaks (DSBs) represent a serious threat to the preservation of genetic information when it reaches hematopoietic stem cells (HSCs). Residual loss of HSC functions and increased risk of developing hematopoietic malignancies are two concerning complications of anti-cancer radiotherapy. Management of acute myelosuppression following radio- or chemotherapy has been significantly improved in recent years by the use of growth factors. However, how cytokine/environmental signals integrate the DNA damage responses in HSCs and regulate the long-term residual HSC defects following radio-or chemotherapy is unknown. Notably, the contribution of cytokines regulating HSC functions to HSC intrinsic DNA damage repair processes remains to be delineated. Thrombopoietin (TPO) and its receptor, Mpl, are critical factors supporting HSC self-renewal, survival and expansion posttransplantation. In this study, we uncover an unknown and unique function for TPO/Mpl in the regulation the DNA damage response. We show that DSB repair, measured by both γH2Ax foci resolution and neutral comet assays, following γ-irradiation (IR) or topoisomerase II inhibitor treatments, is defective in Mpl^−/− and Mpl^+/− HS and progenitor cells (HSPCs). Similar defects were found in wild-type cells treated in the absence of TPO. This indicates that the impaired DNA repair of Mpl^−/− and Mpl^+/− cells results from a specific loss of TPO-mediated DNA damage response signaling at the time of IR rather than from intrinsic constitutive differences. TPO stimulates DNA repair by increasing IR-induced DNA-PK phosphorylation at Ser2056 and Thr2609 and non-homologous end joining (NHEJ) efficiency in both HSPCs and the human UT7-Mpl cell line. This is to our knowledge the first demonstration that a cytokine involved in the homeostatic maintenance of HSCs may also regulate their response to external DNA damaging insults by controlling the DSB repair machinery. Short TPO treatment in vitro or single TPO injection to TPO/Mpl proficient mice prior to sublethal total body IR reduced IR-induced HSC genomic instability and loss of long-term reconstitution ability. This may open new avenues for administration of TPO agonists before radiotherapy to minimize radiation-induced HSC injury and mutagenesis. In addition, since Mpl is haploinsufficient in the regulation of DNA damage repair, these data suggest that Mpl might also act as a tumor suppressor in response to radiotherapy.