Oncostatin M Enhances Hematopoietic Stem Cell Homing and Engraftment

Hematopoietic stem cell (HSC) transplant is widely used to treat hematologic, metabolic, and malignant diseases. HSCs reside within a microenvironmental stem cell niche; the cellular makeup of which is complex, with contributions from endothelial, mesenchymal, and mature hematopoietic cells. Despite the longstanding use of HSC transplantation, there are remarkably few, if any, therapeutics that are available today that can be administered to the recipient of a transplant to enhance hematopoietic engraftment. We believe that revealing the molecular toolkit that naturally exists within the niche provides a rich source of potential novel therapies. Recently, three independent groups (Winkler et al., 2010; Christopher et al., 2011; Chow et al., 2011) have implicated macrophages as a key regulator in G-CSF-mediated mobilization of HSCs. These groups demonstrated that macrophages express the G-CSF receptor, and that depletion of macrophages leads to niche attenuation, reduced levels of stromal derived factor-1 (SDF-1), and mobilization of HSCs. This data implied that macrophages produce a positive supporting factor(s) in the niche to support HSC retention, though none of these prior reports were able to identify the key molecule(s). We now report that Oncostatin M (OSM) is a macrophage-produced protein that regulates HSC retention within the niche. Initial identification was achieved utilizing an in vitro co-culture screening system with macrophage-conditioned media and response of MS-5 stromal cells as measured by SDF-1 production. As previously demonstrated, macrophages enhanced SDF-1 production; however, when OSM production was reduced via shRNA knockdown, or the OSM receptor was knocked down in MS-5 cells or blocked with antibody, the enhanced SDF-1 production was abrogated. Intriguingly, this effect was specific to OSM, as changes in IL-6 or LIF signaling, both members of a similar gp130 signaling family along with OSM, did not have any effects. When mice were treated G-CSF, significant reductions in bone marrow SDF-1 levels and mobilization resulted. However, when mice were co-treated with OSM, levels of bone marrow SDF-1 remained the same as untreated mice (n=10 mice per group, 2 independent expts, P<0.01). Similarly, when macrophages were depleted in vivo via clodronate-loaded liposome treatment, significant HSC mobilization occurred, which was blocked with co-treatment of OSM (n=10 mice per group, 2 independent expts, P<0.01). These results now elucidate a previously unknown key regulatory mechanism governing G-CSF and macrophage mediated mobilization of HSCs. We hypothesized that this new biologic insight could be leveraged as a novel therapeutic strategy to enhance HSC homing and engraftment. To test this, we conducted a series of experiments in which we conditioned mice with myeloablative irradiation and treated with either vehicle control or recombinant OSM (0.5ug per injection) every 6 hours for 48 hours at which point mice received a transplant of bone marrow cells and were subsequently analyzed post-transplant. Mice pre-treated with OSM prior to transplantation exhibited a 2-fold increase in HSC homing compared to vehicle control groups (n=10 mice per group, 3 independent expts, P<0.05). Furthermore, these homed progenitors demonstrated remarkably enhanced hematopoietic expansion in OSM treated mice, as demonstrated by increased numbers of colony forming units (CFUs) from bone marrow assessed days 4, 7, 11, and 14 post-transplant (n=10 mice per group, P<0.01). Excitingly, using a limited cell number transplant, mimicking settings of single cord blood unit transplantation in adults or other settings of limited HSC number, we demonstrate enhanced hematopoietic engraftment and survival in OSM pre-treated mice, with 100 percent survival compared to 50 percent survival of mice treated with vehicle control (n=20 per group, P<0.001). Collectively, our results demonstrate how illumination of endogenous regulatory mechanisms within the hematopoietic niche can reveal molecular agents and pathways with potential to serve as new therapeutic agents in the clinic. Specifically, we identify OSM as the key regulatory molecule governing the G-CSF-macrophage mediated mobilization of HSCs, and describe for the first time a novel therapeutic approach using recombinant OSM as a therapeutic to meet a currently unmet clinical need for HSC transplantation. Disclosures Hoggatt: Harvard University: Patents & Royalties.