title: Identification of a novel function of CX-4945 as a splicing regulator.
abstract: Alternative splicing is a nearly ubiquitous versatile process that controls gene expression and creates numerous protein isoforms with different functions from a single gene. The significance of alternative splicing has been confirmed by the increasing number of human diseases that are caused by misregulation of splicing events. Very few compounds, however, have been reported to act as inhibitors of alternative splicing, and their potential clinical use needs to be evaluated. Here, we report that CX-4945, a previously well-characterized inhibitor of casein kinase 2 (CK2) and a molecule currently in clinical trials (Phase II) for cancer treatment, regulates splicing in mammalian cells in a CK2-independent manner. Transcriptome-wide analysis using exon array also showed a widespread alteration in alternative splicing of numerous genes. We found that CX-4945 potently inhibits the Cdc2-like kinases (Clks) in vitro and in turn, leads to suppression of the phosphorylation of serine/arginine-rich (SR) proteins in mammalian cells. Surprisingly, the overall efficacy of CX-4945 on Clks (IC50 = 3-90 nM) was stronger than that of TG-003, the strongest inhibitor reported to date. Of the Clks, Clk2 was most strongly inhibited by CX-4945 in an ATP-competitive manner. Our research revealed an unexpected activity of the drug candidate CX-4945 as a potent splicing modulator and also suggested a potential application for therapy of diseases caused by abnormal splicing.
authors.affiliation: Targeted Medicine Research Center, Korea Research Institute of Bioscience and Biotechnology, Cheongwon, Chungbuk, Korea; Department of Biomolecular Science, University of Science and Technology, Daejeon, Korea.
authors.forename: Hyeongki
authors.initials: H
authors.lastname: Kim
authors.affiliation: Targeted Medicine Research Center, Korea Research Institute of Bioscience and Biotechnology, Cheongwon, Chungbuk, Korea.
authors.forename: Kwangman
authors.initials: K
authors.lastname: Choi
authors.affiliation: Targeted Medicine Research Center, Korea Research Institute of Bioscience and Biotechnology, Cheongwon, Chungbuk, Korea.
authors.forename: Hyunju
authors.initials: H
authors.lastname: Kang
authors.affiliation: International Cooperation Office, Ministry of Food & Drug Safety, Cheongwon, Chungbuk, Korea.
authors.forename: So-Young
authors.initials: SY
authors.lastname: Lee
authors.affiliation: BioMedical Proteomics Research Center, Korea Research Institute of Bioscience and Biotechnology, Cheongwon, Chungbuk, Korea.
authors.forename: Seung-Wook
authors.initials: SW
authors.lastname: Chi
authors.affiliation: BioMedical Proteomics Research Center, Korea Research Institute of Bioscience and Biotechnology, Cheongwon, Chungbuk, Korea.
authors.forename: Min-Sung
authors.initials: MS
authors.lastname: Lee
authors.affiliation: Targeted Medicine Research Center, Korea Research Institute of Bioscience and Biotechnology, Cheongwon, Chungbuk, Korea.
authors.forename: Jaehyoung
authors.initials: J
authors.lastname: Song
authors.affiliation: Targeted Medicine Research Center, Korea Research Institute of Bioscience and Biotechnology, Cheongwon, Chungbuk, Korea.
authors.forename: Donghwa
authors.initials: D
authors.lastname: Im
authors.affiliation: Targeted Medicine Research Center, Korea Research Institute of Bioscience and Biotechnology, Cheongwon, Chungbuk, Korea.
authors.forename: Yura
authors.initials: Y
authors.lastname: Choi
authors.affiliation: Targeted Medicine Research Center, Korea Research Institute of Bioscience and Biotechnology, Cheongwon, Chungbuk, Korea; Department of Biomolecular Science, University of Science and Technology, Daejeon, Korea.
authors.forename: Sungchan
authors.initials: S
authors.lastname: Cho
mesh_terms: D017398:Alternative Splicing / Q000187:drug effects*; D016203:CDC2 Protein Kinase; D047390:Casein Kinase II / Q000037:antagonists & inhibitors / Q000378:metabolism; D018844:Cyclin-Dependent Kinases / Q000378:metabolism; D056945:Hep G2 Cells; D006801:Humans; D009287:Naphthyridines / Q000494:pharmacology*; D010619:Phenazines; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*
chemical_list: D009287:Naphthyridines; D010619:Phenazines; D047428:Protein Kinase Inhibitors; C555142:silmitasertib; D047390:Casein Kinase II; D016203:CDC2 Protein Kinase; C528899:CDK1 protein, human; D018844:Cyclin-Dependent Kinases
references.citation: Genes Dev. 1998 Jan 1;12(1):55-66
references.pmid: 9420331
references.citation: J Mol Graph Model. 2003 Jan;21(4):289-307
references.pmid: 12479928
references.citation: J Allergy Clin Immunol. 2004 Dec;114(6):1463-70
references.pmid: 15577853
references.citation: Biochem Soc Trans. 2008 Jun;36(Pt 3):483-90
references.pmid: 18481986
references.citation: Hepatology. 2001 Jul;34(1):55-61
references.pmid: 11431734
references.citation: Trends Genet. 2008 Aug;24(8):416-25
references.pmid: 18597886
references.citation: Biochem J. 2003 Jan 1;369(Pt 1):1-15
references.pmid: 12396231
references.citation: Adv Exp Med Biol. 2007;623:123-47
references.pmid: 18380344
references.citation: Cell Death Differ. 2005 Jun;12(6):668-77
references.pmid: 15818404
references.citation: Cell Mol Life Sci. 2009 Jun;66(11-12):1858-67
references.pmid: 19387548
references.citation: Cancer Res. 2004 Aug 1;64(15):5154-61
references.pmid: 15289319
references.citation: Nat Genet. 2008 Dec;40(12):1413-5
references.pmid: 18978789
references.citation: Cancer Res. 2010 Dec 15;70(24):10288-98
references.pmid: 21159648
references.citation: Chembiochem. 2007 Jan 2;8(1):129-39
references.pmid: 17133643
references.citation: Genes Dev. 1997 Feb 1;11(3):334-44
references.pmid: 9030686
references.citation: Cell. 2006 Jul 14;126(1):37-47
references.pmid: 16839875
references.citation: Trends Biochem Sci. 1996 Aug;21(8):296-301
references.pmid: 8772383
references.citation: Proc Natl Acad Sci U S A. 2008 Aug 12;105(32):11218-23
references.pmid: 18678901
references.citation: Cell Metab. 2010 Jan;11(1):23-34
references.pmid: 20074525
references.citation: Nat Rev Neurosci. 2001 Jan;2(1):43-50
references.pmid: 11253358
references.citation: Annu Rev Biochem. 2003;72:291-336
references.pmid: 12626338
references.citation: Proc Natl Acad Sci U S A. 2005 Jun 14;102(24):8764-9
references.pmid: 15939885
references.citation: Neuron. 2006 Oct 5;52(1):93-101
references.pmid: 17015229
references.citation: J Biol Chem. 2004 Jun 4;279(23):24246-54
references.pmid: 15010457
references.citation: RNA. 2006 May;12(5):925-30
references.pmid: 16556940
references.citation: Histol Histopathol. 2001 Apr;16(2):573-82
references.pmid: 11332713
references.citation: Proc Natl Acad Sci U S A. 1994 Nov 8;91(23):10824-8
references.pmid: 7526381
references.citation: Nature. 2008 Nov 27;456(7221):470-6
references.pmid: 18978772
references.citation: Genes Dev. 1992 May;6(5):837-47
references.pmid: 1577277
references.citation: Trends Biochem Sci. 2005 Mar;30(3):115-8
references.pmid: 15752982
references.citation: FEBS Lett. 2001 May 4;496(1):44-8
references.pmid: 11343704
references.citation: FEBS Lett. 2011 Jan 3;585(1):104-10
references.pmid: 21093442
references.citation: EMBO J. 1996 Jan 15;15(2):265-75
references.pmid: 8617202
references.citation: Methods Enzymol. 2006;410:3-28
references.pmid: 16938544
references.citation: Nat Chem Biol. 2007 Sep;3(9):570-5
references.pmid: 17643112
references.citation: Nat Chem Biol. 2007 Sep;3(9):576-83
references.pmid: 17643111
references.citation: Mol Cell Biol. 2010 Apr;30(7):1718-28
references.pmid: 20123975
references.citation: Biochem Biophys Res Commun. 1998 Jan 14;242(2):357-64
references.pmid: 9446799
references.citation: RNA Biol. 2008 Jan-Mar;5(1):17-9
references.pmid: 18388487
references.citation: Trends Genet. 1996 Nov;12(11):472-8
references.pmid: 8973158
references.citation: Curr Med Chem. 2008;15(19):1870-86
references.pmid: 18691045
references.citation: J Cell Biol. 1998 Oct 19;143(2):297-307
references.pmid: 9786943
references.citation: Mol Cell Biol. 1997 Oct;17(10):5996-6001
references.pmid: 9315658
references.citation: Biochem J. 1997 Sep 15;326 ( Pt 3):693-700
references.pmid: 9307018
references.citation: J Biol Chem. 1981 Apr 10;256(7):3319-25
references.pmid: 6937462
references.citation: Genes Dev. 2003 Feb 15;17(4):419-37
references.pmid: 12600935
references.citation: Cancer Res. 2001 Sep 15;61(18):6876-84
references.pmid: 11559564
references.citation: Prog Mol Subcell Biol. 2003;31:33-58
references.pmid: 12494762
references.citation: RNA. 1995 Sep;1(7):663-80
references.pmid: 7585252
references.citation: Bioorg Med Chem Lett. 2013 Jun 15;23(12):3654-61
references.pmid: 23642479
references.citation: J Cell Biochem. 2004 Sep 1;93(1):2-10
references.pmid: 15352156
references.citation: Nat Rev Mol Cell Biol. 2005 May;6(5):386-98
references.pmid: 15956978
references.citation: Adv Enzyme Regul. 2008;48:179-87
references.pmid: 18492491