The cryo-EM structure of the SF3b spliceosome complex bound to a splicing modulator reveals a pre-mRNA substrate competitive mechanism of action
- Lorenzo I. Finci1,
- Xiaofeng Zhang1,
- Xiuliang Huang1,
- Qiang Zhou1,
- Jennifer Tsai2,
- Teng Teng2,
- Anant Agrawal2,
- Betty Chan2,
- Sean Irwin2,
- Craig Karr2,
- Andrew Cook2,
- Ping Zhu2,
- Dominic Reynolds2,
- Peter G. Smith2,
- Peter Fekkes2,
- Silvia Buonamici2 and
- Nicholas A. Larsen2
- 1Beijing Advanced Innovation Center for Structural Biology, Tsinghua-Peking Joint Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing 100084, China;
- 2H3 Biomedicine, Inc., Cambridge, Massachusetts 02139, USA
- Corresponding authors: lfinci{at}hotmail.com, nicholas_larsen{at}h3biomedicine.com
Abstract
Somatic mutations in spliceosome proteins lead to dysregulated RNA splicing and are observed in a variety of cancers. These genetic aberrations may offer a potential intervention point for targeted therapeutics. SF3B1, part of the U2 small nuclear RNP (snRNP), is targeted by splicing modulators, including E7107, the first to enter clinical trials, and, more recently, H3B-8800. Modulating splicing represents a first-in-class opportunity in drug discovery, and elucidating the structural basis for the mode of action opens up new possibilities for structure-based drug design. Here, we present the cryogenic electron microscopy (cryo-EM) structure of the SF3b subcomplex (SF3B1, SF3B3, PHF5A, and SF3B5) bound to E7107 at 3.95 Å. This structure shows that E7107 binds in the branch point adenosine-binding pocket, forming close contacts with key residues that confer resistance upon mutation: SF3B1R1074H and PHF5AY36C. The structure suggests a model in which splicing modulators interfere with branch point adenosine recognition and supports a substrate competitive mechanism of action (MOA). Using several related chemical probes, we validate the pose of the compound and support their substrate competitive MOA by comparing their activity against both strong and weak pre-mRNA substrates. Finally, we present functional data and structure–activity relationship (SAR) on the PHF5AR38C mutation that sensitizes cells to some chemical probes but not others. Developing small molecule splicing modulators represents a promising therapeutic approach for a variety of diseases, and this work provides a significant step in enabling structure-based drug design for these elaborate natural products. Importantly, this work also demonstrates that the utilization of cryo-EM in drug discovery is coming of age.
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Footnotes
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Supplemental material is available for this article.
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Article is online at http://www.genesdev.org/cgi/doi/10.1101/gad.311043.117.
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Freely available online through the Genes & Development Open Access option.
- Received December 20, 2017.
- Accepted February 7, 2018.
This article, published in Genes & Development, is available under a Creative Commons License (Attribution 4.0 International), as described at http://creativecommons.org/licenses/by/4.0/.