Sudemycin Alters Alternative Splicing in Primary Murine Hematopoietic Cells Expressing Mutant U2AF1

Myelodysplastic syndromes (MDS) are a heterogeneous group of clonal hematopoietic stem cell disorders characterized by ineffective hematopoiesis and low blood counts, often progressing to secondary acute myeloid leukemia (sAML). Recent whole genome sequencing of sAML samples has revealed a novel recurrent missense mutation within the coding region of U2AF1, a gene encoding a splicing factor involved in 3’-splice site recognition. The U2AF1 (S34F or S34Y) mutation is an acquired heterozygous mutation in the first zinc finger of U2AF1 present in 13/150 (8.7%) MDS bone marrow samples. The mechanism by which mutations in the spliceosome contribute to MDS pathogenesis is unknown. Sudemycins, modulate the activity of the spliceosome and bind SF3B1, a member of the U2 snRNP. Treatment of cell lines with sudemycins causes accumulation of aberrantly spliced gene products. In low nanomolar concentrations, sudemycins induce cell cycle arrest and apoptosis in cell line in vitro. Sudemycins induce a significant decrease in cell growth and an increase in apoptosis in murine bone marrow cells expressing mutant U2AF1. We hypothesize that splicing gene mutations cause alterations in splice isoforms that provide a selective advantage to tumor cells and that perturbation of this environment could cause a selective disadvantage for cells with these mutations. To test this, we have generated c-kit+ primary murine bone marrow cells expressing wildtype or mutant U2AF1. We are investigating alternative splicing of candidate genes in treated and untreated cells by RT-PCR.