title: A triple exon-skipping luciferase reporter assay identifies a new CLK inhibitor pharmacophore.
abstract: The splicing of pre-mRNA is a critical process in normal cells and is deregulated in cancer. Compounds that modulate this process have recently been shown to target a specific vulnerability in tumors. We have developed a novel cell-based assay that specifically activates luciferase in cells exposed to SF3B1 targeted compounds, such as sudemycin D6. This assay was used to screen a combined collection of approved drugs and bioactive compounds. This screening approach identified several active hits, the most potent of which were CGP-74514A and aminopurvalanol A, both have been reported to be cyclin-dependent kinases (CDKs) inhibitors. We found that these compounds, and their analogs, show significant cdc2-like kinase (CLK) inhibition and clear structure-activity relationships (SAR) at CLKs. We prepared a set of analogs and were able to 'dial out' the CDK activity and simultaneously developed CLK inhibitors with low nanomolar activity. Thus, we have demonstrated the utility of our exon-skipping assay and identified new molecules that exhibit potency and selectivity for CLK, as well as some structurally related dual CLK/CDK inhibitors.
authors.affiliation: Division of Biosciences, SRI International, 333 Ravenswood Ave., Menlo Park, CA 94025, USA.
authors.forename: Yihui
authors.initials: Y
authors.lastname: Shi
authors.affiliation: Division of Biosciences, SRI International, 333 Ravenswood Ave., Menlo Park, CA 94025, USA.
authors.forename: Jaehyeon
authors.initials: J
authors.lastname: Park
authors.affiliation: Division of Biosciences, SRI International, 333 Ravenswood Ave., Menlo Park, CA 94025, USA.
authors.forename: Chandraiah
authors.initials: C
authors.lastname: Lagisetti
authors.affiliation: Division of Biosciences, SRI International, 333 Ravenswood Ave., Menlo Park, CA 94025, USA.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhou
authors.affiliation: Division of Biosciences, SRI International, 333 Ravenswood Ave., Menlo Park, CA 94025, USA.
authors.forename: Lidia C
authors.initials: LC
authors.lastname: Sambucetti
authors.affiliation: Division of Biosciences, SRI International, 333 Ravenswood Ave., Menlo Park, CA 94025, USA. Electronic address: thomas.webb@sri.com.
authors.forename: Thomas R
authors.initials: TR
authors.lastname: Webb
mesh_terms: D015075:2-Aminopurine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D000225:Adenine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D001665:Binding Sites; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D005091:Exons; D017930:Genes, Reporter; D057166:High-Throughput Screening Assays; D006801:Humans; D020128:Inhibitory Concentration 50; D008156:Luciferases / Q000235:genetics; D056004:Molecular Dynamics Simulation; D011485:Protein Binding; D047428:Protein Kinase Inhibitors / Q000737:chemistry* / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012326:RNA Splicing; D013329:Structure-Activity Relationship
chemical_list: C469937:N(2)-(2-aminocyclohexyl)-N(6)-(3-chlorophenyl)-9-ethyl-9H-purine-2,6-diamine; D047428:Protein Kinase Inhibitors; C502582:aminopurvalanol A; D015075:2-Aminopurine; D008156:Luciferases; D018844:Cyclin-Dependent Kinases; D000225:Adenine
keywords: Anti-tumor agents; Cdc-like kinase (CLK) inhibitors; High throughput screening; Modulators of pre-mRNA splicing