Specific CLK inhibitors from a novel chemotype for regulation of alternative splicing

Oleg Fedorov; Kilian Huber; Andreas Eisenreich; Panagis Filippakopoulos; Oliver King; Alex N Bullock; Damian Szklarczyk; Lars J Jensen; Doriano Fabbro; Jörg Trappe; Ursula Rauch; Franz Bracher; Stefan Knapp

doi:10.1016/j.chembiol.2010.11.009

Specific CLK inhibitors from a novel chemotype for regulation of alternative splicing

Chem Biol. 2011 Jan 28;18(1):67-76. doi: 10.1016/j.chembiol.2010.11.009.

Authors

Oleg Fedorov¹, Kilian Huber, Andreas Eisenreich, Panagis Filippakopoulos, Oliver King, Alex N Bullock, Damian Szklarczyk, Lars J Jensen, Doriano Fabbro, Jörg Trappe, Ursula Rauch, Franz Bracher, Stefan Knapp

Affiliation

¹ University of Oxford, Nuffield Department of Clinical Medicine, Structural Genomics Consortium, Old Road Campus Research Building, Oxford OX37DQ, UK.

Abstract

There is a growing recognition of the importance of protein kinases in the control of alternative splicing. To define the underlying regulatory mechanisms, highly selective inhibitors are needed. Here, we report the discovery and characterization of the dichloroindolyl enaminonitrile KH-CB19, a potent and highly specific inhibitor of the CDC2-like kinase isoforms 1 and 4 (CLK1/CLK4). Cocrystal structures of KH-CB19 with CLK1 and CLK3 revealed a non-ATP mimetic binding mode, conformational changes in helix αC and the phosphate binding loop and halogen bonding to the kinase hinge region. KH-CB19 effectively suppressed phosphorylation of SR (serine/arginine) proteins in cells, consistent with its expected mechanism of action. Chemical inhibition of CLK1/CLK4 generated a unique pattern of splicing factor dephosphorylation and had at low nM concentration a profound effect on splicing of the two tissue factor isoforms flTF (full-length TF) and asHTF (alternatively spliced human TF).

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Alternative Splicing / drug effects*
Catalytic Domain
Endothelial Cells / drug effects
Endothelial Cells / metabolism
Heterocyclic Compounds, 2-Ring / chemical synthesis
Heterocyclic Compounds, 2-Ring / chemistry
Heterocyclic Compounds, 2-Ring / pharmacology
Humans
Models, Molecular
Nitriles / chemical synthesis
Nitriles / chemistry
Nitriles / pharmacology
Phosphorylation / drug effects
Protein Kinase Inhibitors / chemical synthesis
Protein Kinase Inhibitors / chemistry*
Protein Kinase Inhibitors / pharmacology*
Protein Serine-Threonine Kinases / antagonists & inhibitors*
Protein Serine-Threonine Kinases / chemistry
Protein Serine-Threonine Kinases / metabolism
Protein-Tyrosine Kinases / antagonists & inhibitors*
Protein-Tyrosine Kinases / chemistry
Protein-Tyrosine Kinases / metabolism
RNA, Messenger / genetics
RNA-Binding Proteins / metabolism
Substrate Specificity
Thromboplastin / genetics

Substances

Heterocyclic Compounds, 2-Ring
Nitriles
Protein Kinase Inhibitors
RNA, Messenger
RNA-Binding Proteins
enaminonitrile
Thromboplastin
Clk dual-specificity kinases
Protein-Tyrosine Kinases
Protein Serine-Threonine Kinases