Altered RNA Export in SF3B1 Mutants Increases Sensitivity to Nuclear Export Inhibition

SF3B1 mutations are the most frequent spliceosomal alterations across cancers, yet no successful therapy exists to target this pathway. Previous findings from a phase 2 clinical trial of the XPO1 inhibitor selinexor in patients with high-risk myelodysplastic syndrome (MDS) relapsed or refractory to...

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Published in:Blood 2023-11, Vol.142 (Supplement 1), p.44-44
Main Authors: Chaudhry, Sana, Beckedorff, Felipe, Totiger, Tulasigeri M, Affer, Maurizio, Hariramani, Nidhi, Montoya, Skye, Cornista, Alyssa, Bilbao, Daniel, Roberts, Evan R., Afaghani, Jumana, Rodríguez, Jose Antonio, Wang, Eric, Taylor, Justin
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Language:English
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Summary:SF3B1 mutations are the most frequent spliceosomal alterations across cancers, yet no successful therapy exists to target this pathway. Previous findings from a phase 2 clinical trial of the XPO1 inhibitor selinexor in patients with high-risk myelodysplastic syndrome (MDS) relapsed or refractory to hypomethylating agents (HMA) revealed increased activity in patients with SF3B1 mutations. XPO1 (Exportin-1) is responsible for the nuclear export of over 200 proteins, but also plays a role in the transport of multiple RNA species, including small nuclear RNAs (snRNAs), ribosomal RNAs (rRNAs), and select messenger RNAs (mRNAs) out of the nucleus. Given the role of XPO1 in exporting snRNAs, which form the catalytic portion of the spliceosome, we hypothesized that XPO1 inhibition may preferentially affect SF3B1-mutant cells via altered splicing, and that SF3B1-mutant high-risk MDS patients would have a better response to rational targeted drug combinations with next generation XPO1 inhibitors. To evaluate the mechanism underlying the preferential sensitivity of SF3B1-mutant cells to XPO1 inhibition, we performed nuclear and cytoplasmic fractionation followed by RNA sequencing (subcellular RNA-seq) before and after XPO1 inhibition in SF3B1 wildtype and SF3B1 K666N mutant cells. Whole transcriptomic analysis of subcellular RNA-seq data revealed increased global retention of RNA transcripts in the nucleus after XPO1 inhibition in the SF3B1 mutant cell line ( Figure 1). Validation assays using qPCR confirmed nuclear retention of several key transcripts such as Salt Inducible Kinase 1 (SIK1) and Cancer/Testis Antigen Family 45 Member A2 (CT45A2) in SF3B1-mutants (both SF3B1 K666N and K700E). Similarly, we performed subcellular RNA-seq for small RNAs and found snRNAs to be increased in the nucleus after XPO1 inhibition in the SF3B1 mutant cells. We then performed total cellular RNA sequencing to understand the effect of XPO1 inhibition on global RNA expression and RNA splicing. Differential gene expression analysis revealed that XPO1 inhibition had a more pronounced effect on cell cycle in SF3B1 wildtype cells, but differentiation pathways were more significantly affected in SF3B1-mutant cells. Alternative splicing analysis showed increased 3' alternative splicing events in SF3B1 mutant after XPO1 inhibition. These results suggest that the preferential sensitivity of SF3B1 mutant cells to nuclear export inhibition arises through increased nuclear retention of spliceoso
ISSN:0006-4971
1528-0020
DOI:10.1182/blood-2023-182877