Abstract
Genomic analyses of cancer have identified recurrent point mutations in the RNA splicing factor–encoding genes SF3B1, U2AF1, and SRSF2 that confer an alteration of function1,2,3,4,5,6. Cancer cells bearing these mutations are preferentially dependent on wild-type (WT) spliceosome function7,8,9,10,11, but clinically relevant means to therapeutically target the spliceosome do not currently exist. Here we describe an orally available modulator of the SF3b complex, H3B-8800, which potently and preferentially kills spliceosome-mutant epithelial and hematologic tumor cells. These killing effects of H3B-8800 are due to its direct interaction with the SF3b complex, as evidenced by loss of H3B-8800 activity in drug-resistant cells bearing mutations in genes encoding SF3b components. Although H3B-8800 modulates WT and mutant spliceosome activity, the preferential killing of spliceosome-mutant cells is due to retention of short, GC-rich introns, which are enriched for genes encoding spliceosome components. These data demonstrate the therapeutic potential of splicing modulation in spliceosome-mutant cancers.
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References
Yoshida, K. et al. Frequent pathway mutations of splicing machinery in myelodysplasia. Nature 478, 64–69 (2011).
Wang, L. et al. SF3B1 and other novel cancer genes in chronic lymphocytic leukemia. N. Engl. J. Med. 365, 2497–2506 (2011).
Harbour, J.W. et al. Recurrent mutations at codon 625 of the splicing factor SF3B1 in uveal melanoma. Nat. Genet. 45, 133–135 (2013).
Imielinski, M. et al. Mapping the hallmarks of lung adenocarcinoma with massively parallel sequencing. Cell 150, 1107–1120 (2012).
Bailey, P. et al. Genomic analyses identify molecular subtypes of pancreatic cancer. Nature 531, 47–52 (2016).
Ciriello, G. et al. Comprehensive molecular portraits of invasive lobular breast cancer. Cell 163, 506–519 (2015).
Zhou, Q. et al. A chemical genetics approach for the functional assessment of novel cancer genes. Cancer Res. 75, 1949–1958 (2015).
Fei, D.L. et al. Wild-Type U2AF1 antagonizes the splicing program characteristic of U2AF1-mutant tumors and is required for cell survival. PLoS Genet. 12, e1006384 (2016).
Obeng, E.A. et al. Physiologic expression of Sf3b1(K700E) causes impaired erythropoiesis, aberrant splicing, and sensitivity to therapeutic spliceosome modulation. Cancer Cell 30, 404–417 (2016).
Shirai, C.L. et al. Mutant U2AF1-expressing cells are sensitive to pharmacological modulation of the spliceosome. Nat. Commun. 8, 14060 (2017).
Lee, S.C. et al. Modulation of splicing catalysis for therapeutic targeting of leukemia with mutations in genes encoding spliceosomal proteins. Nat. Med. 22, 672–678 (2016).
Papaemmanuil, E. et al. Somatic SF3B1 mutation in myelodysplasia with ring sideroblasts. N. Engl. J. Med. 365, 1384–1395 (2011).
Lee, Y. & Rio, D.C. Mechanisms and regulation of alternative pre-mRNA splicing. Annu. Rev. Biochem. 84, 291–323 (2015).
Nguyen, T.H. et al. CryoEM structures of two spliceosomal complexes: starter and dessert at the spliceosome feast. Curr. Opin. Struct. Biol. 36, 48–57 (2016).
Kim, E. et al. SRSF2 mutations contribute to myelodysplasia by mutant-specific effects on exon recognition. Cancer Cell 27, 617–630 (2015).
Ilagan, J.O. et al. U2AF1 mutations alter splice site recognition in hematological malignancies. Genome Res. 25, 14–26 (2015).
Darman, R.B. et al. Cancer-Associated SF3B1 hotspot mutations induce cryptic 3′ splice site selection through use of a different branch point. Cell Rep. 13, 1033–1045 (2015).
Alsafadi, S. et al. Cancer-associated SF3B1 mutations affect alternative splicing by promoting alternative branchpoint usage. Nat. Commun. 7, 10615 (2016).
Zhang, J. et al. Disease-associated mutation in SRSF2 misregulates splicing by altering RNA-binding affinities. Proc. Natl. Acad. Sci. USA 112, E4726–E4734 (2015).
Kotake, Y. et al. Splicing factor SF3b as a target of the antitumor natural product pladienolide. Nat. Chem. Biol. 3, 570–575 (2007).
Teng, T. et al. Splicing modulators act at the branch point adenosine binding pocket defined by the PHF5A-SF3b complex. Nat. Commun. 8, 15522 (2017).
Cretu, C. et al. Molecular Architecture of SF3b and structural consequences of its cancer-related mutations. Mol. Cell 64, 307–319 (2016).
Folco, E.G., Coil, K.E. & Reed, R. The anti-tumor drug E7107 reveals an essential role for SF3b in remodeling U2 snRNP to expose the branch point-binding region. Genes Dev. 25, 440–444 (2011).
Amit, M. et al. Differential GC content between exons and introns establishes distinct strategies of splice-site recognition. Cell Rep. 1, 543–556 (2012).
Jenkins, J.L., Agrawal, A.A., Gupta, A., Green, M.R. & Kielkopf, C.L. U2AF65 adapts to diverse pre-mRNA splice sites through conformational selection of specific and promiscuous RNA recognition motifs. Nucleic Acids Res. 41, 3859–3873 (2013).
Mercer, T.R. et al. Genome-wide discovery of human splicing branchpoints. Genome Res. 25, 290–303 (2015).
Corrionero, A., Miñana, B. & Valcárcel, J. Reduced fidelity of branch point recognition and alternative splicing induced by the anti-tumor drug spliceostatin A. Genes Dev. 25, 445–459 (2011).
Cvitkovic, I. & Jurica, M.S. Spliceosome database: a tool for tracking components of the spliceosome. Nucleic Acids Res. 41, D132–D141 (2013).
Hegele, A. et al. Dynamic protein–protein interaction wiring of the human spliceosome. Mol. Cell 45, 567–580 (2012).
Barbosa-Morais, N.L., Carmo-Fonseca, M. & AparÃcio, S. Systematic genome-wide annotation of spliceosomal proteins reveals differential gene family expansion. Genome Res. 16, 66–77 (2006).
Pellizzoni, L., Kataoka, N., Charroux, B. & Dreyfuss, G. A novel function for SMN, the spinal muscular atrophy disease gene product, in pre-mRNA splicing. Cell 95, 615–624 (1998).
Luo, M.J. & Reed, R. Splicing is required for rapid and efficient mRNA export in metazoans. Proc. Natl. Acad. Sci. USA 96, 14937–14942 (1999).
Yoshimi, A. et al. Robust patient-derived xenografts of MDS/MPN overlap syndromes capture the unique characteristics of CMML and JMML. Blood 130, 397–407 (2017).
Cheng, D.T. et al. Memorial Sloan Kettering-integrated mutation profiling of actionable cancer targets (MSK-IMPACT): a hybridization capture-based next-generation sequencing clinical assay for solid tumor molecular oncology. J. Mol. Diagn. 17, 251–264 (2015).
Dobin, A. et al. STAR: ultrafast universal RNA-seq aligner. Bioinformatics 29, 15–21 (2013).
Bray, N.L., Pimentel, H., Melsted, P. & Pachter, L. Near-optimal probabilistic RNA-seq quantification. Nat. Biotechnol. 34, 525–527 (2016).
Robinson, M.D., McCarthy, D.J. & Smyth, G.K. edgeR: a Bioconductor package for differential expression analysis of digital gene expression data. Bioinformatics 26, 139–140 (2010).
Hunter, J. D. Matplotlib: A 2D Graphics Environment. Computing in Science and Engineering 9, 90–95 (2007).
Sgouropoulos, N., Yao, Q. & Yastremiz, C. Matching a Distribution by Matching Quantiles Estimation. J. Am. Stat. Assoc. 110, 742–759 (2015).
Croft, D. et al. The Reactome pathway knowledgebase. Nucleic Acids Res. 42, D472–D477 (2014).
Thomas, P.D. et al. PANTHER: a browsable database of gene products organized by biological function, using curated protein family and subfamily classification. Nucleic Acids Res. 31, 334–341 (2003).
Acknowledgements
We thank H3 Biomedicine employees for their support in this project. A.Y. was supported by grants from the Aplastic Anemia and Myelodysplastic Syndromes (MDS) International Foundation and the Lauri Strauss Leukemia Foundation. O.A.-W. was supported by grants from the Edward P. Evans Foundation, the Taub Foundation, the Department of Defense Bone Marrow Failure Research Program (BM150092 and W81XWH-12-1-0041), National Institutes of Health National Heart, Lung and Blood Institute (R01 HL128239), the Josie Robertson Investigator Program, an award from the Starr Foundation (I8-A8-075), the Leukemia and Lymphoma Society (2314-17) and the Pershing Square Sohn Cancer Research Alliance.
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G.K., S. Prajapati, J.W., and X.L guided and performed the medicinal chemistry efforts that led to identification of H3B-8800. E. Park, A.A.A., B.C., and P.F. performed the in vitro biochemical assays. A.Y., A.C., B.C., R.D., C.K., L.L., P.K., C. Mackenzie, Y.M., T.T., H.Y., P.Z., P.G.S., and S.B. performed experiments and analyses of H3B-8800 performance in in vitro cellular assays. M.T., E.S., G.L., E. Pazolli, C. Meeske, P.G.S., M.W., and S.B. performed studies and analyzed data from the cell line xenografts. V.K. and E. Padron provided patient materials used for PDX models. A.Y., S.C-W.L., J.T., and O.A.-W. generated PDX models and performed PDX experiments and analyses. M.S., S. Peng, and L.Y. performed RNA-seq analyses. O.A.-W., M.S., A.A.A., P.G.S., and S.B. wrote the manuscript.
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M.T., B.C., M.S., A.A.A., B. Chan, B. Caleb, A.C., R.D., P.F., C.K., G.K., L.L., P.K., X.L., C. Mackenzie, C. Meeske, Y.M., E. Park, S. Peng, S. Prajapati, T.T., J.W., M.W., H.Y., L.Y., P.Z., P.G.S. and S.B. are employees of H3 Biomedicine, Inc., and E.S. and G.L. are employees of Eisai, Inc.
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Seiler, M., Yoshimi, A., Darman, R. et al. H3B-8800, an orally available small-molecule splicing modulator, induces lethality in spliceosome-mutant cancers. Nat Med 24, 497–504 (2018). https://doi.org/10.1038/nm.4493
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DOI: https://doi.org/10.1038/nm.4493
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