Suppression of TGF-ß Signaling by ALK5 Inhibition As Treatment for MPN-associated Myelofibrosis in Mice

Myelofibrosis (MF) is a common feature of the Philadelphia-Chromosome negative (Ph-) myeloproliferative neoplasms (MPN) that does not respond to current treatment with Jak2 inhibitors. TGF-ß1 is elevated in PMF patient plasma, progenitors and megakaryocytes (MK) and is known to promote fibrosis through stimulation of fibroblasts and their progenitors, mesenchymal stroma cells (MSC). We tested the efficacy of the orally available ALK5 inhibitor Galunisertib in counteracting fibrosis in two MF mouse models of common MPN oncogenes Jak2 and Mpn. Transgenic Vav1-Jak2V617F mice show a Polycythemia Vera-like phenotype at 6 weeks old, progressing to fibrosis of bone marrow (BM) and spleen around 25-30 weeks. Thirty week old mice were treated with Galunisertib for 4 weeks and showed a significant decrease in fibrosis compared to control mice. Similar results were observed in 50 week old mice treated with a murine antibody against TGF-ß1 (IMC-TR1,LY3022859) for 4 weeks. Mice transplanted with BM cells transduced with MPLW515L rapidly develop MF, myeloproliferation and splenomegaly. Flow cytometry showed an expansion of immature and mature megakaryocytes with overexpression of TGF-ß1 in MPLW515L cells. Treatment from day 12 to day 26 after transplantation significantly reduced fibrosis as measured by reticulin staining and hydroxyproline quantification. In vitro, Galunisertib counteracted the pro-fibrotic effects of TGF-ß1 on both murine and human MSC by antagonizing its stimulatory effects on collagen I and III production. Analysis of MSCs from Galunisertib-treated MPLW515L mice also showed reduced collagen I and III production compared to vehicle controls. In conclusion, we offer further evidence that aberrant megakaryopoiesis drives TGF-ß1 overproduction in MPN by increasing megakaryocyte numbers and overexpressing TGF-ß1 in mutant MKs. Galunisertib counteracts TGF-ß1 induced fibrosis in two MPN mouse-models and is a promising candidate for symptomatic treatment of myelofibrosis.