Modified Activin Receptor Type IIb-Fc Fusion Protein (RAP-536) Decreases Anemia In a Murine Model Of Myelodysplastic Syndrome and Improves Overall Survival

Myelodysplastic syndromes (MDS) are caused due to abnormal proliferation and differentiation of pluripotent hematopoietic stem cells leading to peripheral cytopenias including anemia and an increased risk of progression to acute myelogenous leukemia (AML). The mainstay of anemia treatments for majority of non-del (5q) patients are frequent transfusions but often leads to iron overloading and enhanced progression to AML, causing a negative impact on overall survival (OS). Therefore, alternative therapies that promote effective erythropoiesis, decrease anemia, and improve OS, are needed.

Members of the TGF-β superfamily are known regulators of erythropoiesis. ACE-536 is a modified soluble activin receptor type IIB-Fc fusion protein that acts as a ligand trap for certain TGF-β family ligands and prevents Smad 2/3 signaling. ACE-536 has shown a robust increase in RBCs in mice, rats and monkeys. In normal mice, ACE-536 promotes maturation but not proliferation of late stage erythroid precursors. Additionally, we have shown that RAP-536 (murine ortholog of ACE-536) corrected anemia in the NUP98-HOXD13 (NHD13) murine model of MDS. In this study, we evaluated the progression of MDS disease and OS of NHD13 mice administered RAP-536.

NHD13 mice begin to develop anemia, neutropenia and lymphopenia at four months of age. NHD13 mice die by 14 months due to severe pancytopenia or progression to AML. In this study, 4-month old NHD13 mice (N=12-16/group) were dosed with RAP-536 (10 mg/kg) or vehicle (VEH) twice per week for 5 or 10 months. Age matched wild type mice were used as controls. At each time point, blood samples were collected for CBCs. Bone marrow and splenic hematopoietic precursors of various cell lineages were immuno-stained and analyzed by flow cytometry (FCM). Spleen sections, blood and bone marrow smears were also analyzed for histopathological changes.

After 5 months of treatment, VEH treated NHD13 mice had decreased RBC (-19.6%, P<0.001), WBC (-30.8%, P<0.001), lymphocytes (-63.2%, P<0.001) and increased platelet counts (+89.2%, P<0.05) compared to wild type mice. Treatment with RAP-536 increased RBC (+7.2%, P<0.05) and reduced platelet counts compared to VEH control. No significant changes in other blood lineages were observed following RAP-536 treatment, demonstrating that RAP-536 is selective of the erythroid lineage.

After 10 months of treatment, VEH treated NHD13 mice had severely decreased RBC (-32.9%) and hemoglobin (-21.8%) compared to wild-type mice. RAP-536 treatment increased RBC (+21.4%) and hemoglobin (+16.6%) compared to VEH treatment. FCM evaluation of erythroid precursors from bone marrow of NHD13 mice demonstrated increased immature CD71⁺Ter119⁺ erythroblasts (from 13.1% to 18.3%), and decreased mature CD71^-Ter119⁺ erythroblasts (from 13.2% to 3.8%) compared to wild-type mice. Treatment with RAP-536 increased mature erythroblasts (from 3.8% to 9.6%) consistent with improved RBC parameters, indicating the stimulation of erythroid differentiation. Additionally, bone marrow from NHD13 mice had significantly elevated Gr1⁺ & CD11b⁺ (from 33.6% to 62.6%) and CD4⁺ & CD8⁺(from 19.1% to 32.3%) precursors, while peripheral blood displayed a concomitant decreases in granulocytes (-22.5%), WBC (-37.5%) and lymphocytes (-45%) compared to wild type mice, demonstrating ineffective hematopoiesis. Treatment with RAP-536 displayed a non-statistical decrease in these precursors in bone marrow and a similar increase in peripheral blood compared to VEH control. No changes in platelets were observed after ten months of treatment. These data suggests that the effect of RAP-536 on other hematopoietic lineages is likely secondary to its effect on erythropoiesis. Importantly, histopathological findings revealed no indication of increased leukemic progression in RAP-536 treated NHD13 mice compared to VEH treated mice. Furthermore, RAP-536 treated NHD13 mice demonstrated a trend for increased median survival compared to VEH treated mice, from 238 days to 277 days (P=0.08).

Together, these data demonstrate that RAP-536 corrects anemia associated with ineffective erythropoiesis in NHD13 mouse model of MDS. RAP-536 does not enhance progression to AML, and may increase overall survival of NHD13 mice. ACE-536 is currently being evaluated for the treatment of anemia in patients with MDS and β-thalassemia, conditions characterized by ineffective erythropoiesis.