Mis-splicing of mitotic regulators sensitizes SF3B1-mutated human HSCs to CHK1 inhibition.

Splicing factor SF3B1 mutations are frequent somatic lesions in myeloid neoplasms that transform hematopoietic stem cells (HSCs) by inducing mis-splicing of target genes. However, the molecular and functional consequences of SF3B1 mutations in human HSCs remain unclear. Here, we identify the mis-splicing program in human HSCs as a targetable vulnerability by precise gene editing of SF3B1 K700E mutations in primary CD34+ cells. Mutant SF3B1 induced pervasive mis-splicing and reduced expression of genes regulating mitosis and genome maintenance leading to altered differentiation, delayed G2/M progression, and profound sensitivity to CHK1 inhibition (CHK1i). Mis-splicing or reduced expression of mitotic regulators BUBR1 and CDC27 delayed G2/M transit and promoted CHK1i sensitivity. Clinical CHK1i prexasertib selectively targeted SF3B1-mutant HSCs and abrogated engraftment in vivo. These findings identify mis-splicing of mitotic regulators in SF3B1-mutant HSCs as a targetable vulnerability engaged by pharmacological CHK1 inhibition.