Erythropoietin-Directed Erythropoiesis Depends Upon Serpin Inhibition of Erythroblast Lysosomal Cathepsins

Abstract 981

EPO is used to treat anemia associated with chronic kidney disease, and cancer chemotherapy. However, EPO has hypertensive and thrombolytic side-effects and can impact on innate immunity, diabetes, vasculogenesis, and the progression of certain cancers. These observations, together with the clinical emergence of new EPOR agonists provide compelling reasons to better understand key EPO/EPOR actions. This includes effects on erythroid progenitor cells (EPC's) as a prime target in which EPO/EPOR actions remain incompletely understood. To discover novel EPO action modes we profiled the transcriptome of primary EPC's. We report Serpina3g/Spi2A as a major EPO/EPOR target, and a key EPC survival factor. In knockout mice, loss of Spi2A sharply worsened anemia due to hemolysis, radiation or transplantation (p = 0.008; p = 0.0001; p = 0.010, respectively). EPO-induced red cell formation also was limited to 8.6 + 0.7% of wild-type control responses. Unexpectedly, maturing erythroblasts proved to require Spi2A for cytoprotection, with iron and ROS defined as cytotoxic agents. Notably, Spi2A defects were ameliorated by cathepsin-B/L inhibition, and by the genetic co-deletion of lysosomal cathepsin-B. In vivo, pharmacological inhibition of cathepsin B/L enhanced EPO's effects on hematocrits by 5⁺ points (d4 and d6 post- EPO). Overall, we define an unexpected EPO action mode via an EPOR- Spi2A serpin- cathepsin axis in maturing erythroblasts, with oxidants (and iron) as cytotoxic potentiators, and lysosomal cathepsins as novel therapeutic targets.