Dysregulation of TGF-Beta Stimulated Smad Signaling Is Seen in Myelodysplasia and Points to the Potential Therapeutic Efficacy of TGF-Beta Receptor I Kinase Inhibition in Low Grade Disease.

Abstract 737

Myelodysplastic syndromes (MDS) are characterized by ineffective hematopoiesis that leads to peripheral cytopenias. TGF-beta is a hematopoietic inhibitory cytokine that has been indirectly linked to the pathogenesis of some subsets of MDS and acute leukemias. We have shown (Blood, 112(8):3434; 2008) that smad2, a component of the TGF-beta signaling pathway, is constitutively activated and upregulated in MDS progenitors. Since there is conflicting data about upregulation of TGF-beta levels in MDS, we next sought to determine the molecular basis of TGF-beta receptor-I (TBRI) overactivation and subsequent smad2 phosphorylation / activation in this disease. We observed that smad-7, a negative regulator of TBRI kinase, is markedly down regulated in a meta-analysis of gene expression studies from 89 MDS bone marrow derived CD34+ cells when compared to 61 normal controls (Adjusted P<0.001, Benjamini Hochberg). Smad-7 protein was also found to be significantly decreased in MDS BM progenitors (n=14) when compared to anemic controls (n=18) when examined immunohistochemically in a bone marrow tissue microarray (P=0.04, T test). siRNA mediated knockdown of smad-7 levels led to increased TGF mediated gene transcription over baseline, that could be effectively inhibited by a TBRI (ALK5 kinase) inhibitor. TBRI kinase inhibition effectively abrogated TGF-beta mediated activation of smad-2 in hematopoietic cells. Most importantly, treatment with the TBRI inhibitor increased erythroid and myeloid colony formation from primary MDS bone marrow specimens in a concentration-dependent fashion. Furthermore, the hematopoietic potentiating effect of the TBRI kinase inhibition was validated in a TGF-overexpressing transgenic mouse model. This mouse constitutively expresses TGF-beta through an albumin promoter, develops progressive anemia and dysplasia and thus serves as a novel model of human bone marrow failure. Treatment with the TBRI inhibitor by oral lavage at 100 mg/kg/d for two weeks led to significant increases in hematocrit when compared to placebo (Mean increase in hematocrit=48%, P value=0.01, T test). Taken together, these studies demonstrate an important role for reduction in smad-7 in contributing to ineffective hematopoiesis in MDS by activating TGF-beta signaling in hematopoietic cells. These studies also illustrate the promising therapeutic potential of TBRI inhibitors in this disease.