Abstract
Lenalidomide displays erythroid and cytogenetic remitting activity in MDS that is karyotype-dependent, yielding a high response rate in patients with chromosome 5q deletion (del5q; 76%) whereas cytogenetic and pathologic improvement in non-del5q patients is less common despite transfusion response in a significant proportion of patients [
NEJM 2005;352:549–57
]. We postulated that lenalidomide targets regulatory proteins that repress the erythropoietin (EPO) signal in non-del5q MDS while inhibiting a critical del5q-associated target to account for karyotype-specific clonal suppression. To evaluate the mechanism by which lenalidomide enhances erythropoiesis in non-del5q MDS, we investigated its effects in MDS [N=9] and normal [N=7] marrow specimens, and in an EPO-responsive UT7 cell line model. Pre-incubation exposure to lenalidomide augmented EPO-induced BFU-E recovery up to 2.5-fold in non-del5q MDS specimens [N=7] over a concentration range of 0.001–1 mM, whereas change in BFU-E recovery from normals was nominal. In UT7 cells, lenalidomide [1mM] enhanced clonogenic response to EPO in a supra-additive fashion. To characterize effects on receptor signal response, we investigated the interaction between lenalidomide and EPO on STAT5 activation, and corresponding EPO-receptor (R) associated kinases. Lenalidomide enhanced both EPO-induced STAT5 phosphorylation and DNA binding in UT7 and MDS CD71Hi MDS erythroid precursors with corresponding Bcl-XL up-regulation. The magnitude of lenalidomide enhancement of EPO/STAT5 phosphorylation was significantly higher in MDS CD71Hi erythroid precursors compared to normal donors (median log10 geometric MFI pSTAT5 EPO+CC5013:EPO, 0.989 [s.d.,0.25] MDS vs. 0.0195 [0.07] Normal; P=0.0005). Lenalidomide enhancement of the EPO-R/STAT5 signal was associated with increased Janus kinase-2 (Jak-2) and Lyn kinase auto-phosphorylation and was abolished only by dual Jak and Src kinase inhibition, implicating targeted inhibition of a common Jak/Lyn kinase regulatory molecule. Indeed, lenalidomide potentiation of the EPO/STAT5 axis was replicated by inhibition of the dual Jak-2/Lyn PTP, CD45, with the selective antagonist RWJ-60475. In a cell free assay employing rhu-CD45 and a pp60 c-Src reporter substrate, lenalidomide inhibited CD45 PTP activity in a time-dependent fashion with a 50% inhibitory concentration [IC50] of 10nM, whereas thalidomide was largely ineffective. PTP inhibition was CD45 selective, demonstrating no significant inhibition of rhu-SHP-2 PTP activity. Our data suggest that lenalidomide is a PTP inhibitor that enhances EPO receptor signaling in non-del5q erythroid precursors by relieving CD45 repression of ligand-dependent Jak2 and Lyn kinase mediated STAT5 activation. Moreover, these findings suggest that CD45 over-activity may contribute to impaired EPO-R signaling in MDS erythroid precursors.Disclosures: Dr. Alan List has a relationship with the Celgene Corporation.; Dr. Alan List has a relationship with the Celgene Corporation.; Dr. Alan List has a relationship with the Celgene Corporation.
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2006, The American Society of Hematology
2006
