Inhibitors of casein kinase 1 block the growth of Leishmania major promastigotes in vitro.
Casein kinase 1 (CK1) is a family of multifunctional Ser/Thr protein kinases that are ubiquitous in eukaryotic cells. Recent studies have demonstrated the existence of, and role for, CK1 in protozoan parasites such as Leishmania, Plasmodium and Trypanosoma. The value of protein kinases as potential drug targets in protozoa is evidenced by the successful exploitation of cyclic guanosine monophosphate-dependent protein kinase (PKG) with selective tri-substituted pyrrole and imidazopyridine inhibitors. These compounds exhibit in vivo efficacy against Eimeria tenella in chickens and Toxoplasma gondii in mice. We now report that both of these protein kinase inhibitor classes inhibit the growth of Leishmania major promastigotes and Trypanosoma brucei bloodstream forms in vitro. Genome informatics predicts that neither of these trypanosomatids codes for a PKG orthologue. Biochemical studies have led to the unexpected discovery that an isoform of CK1 represents the primary target of the pyrrole and imidazopyridine kinase inhibitors in these organisms. CK1 from extracts of L. major promastigotes co-fractionated with [(3)H]imidazopyridine binding activity. Further purification of CK1 activity from L. major and characterization via liquid chromatography coupled tandem mass spectrometry identified CK1 isoform 2 as the specific parasite protein inhibited by imidazopyridines. L. major CK1 isoform 2 expressed as a recombinant protein in Escherichia coli displayed biochemical and inhibition characteristics similar to those of the purified native enzyme. The results described here warrant further evaluation of the activity of these kinase inhibitors against mammalian stage Leishmania parasites in vitro and in animal models of infection, as well as studies to genetically validate CK1 as a therapeutic target in trypanosomatid parasites.
Allocco
John J
JJ
Department of Infectious Disease Research, Merck Research Laboratories, Merck and Co., Inc., P.O. Box 2000 Rahway, NJ 07065-0900, USA. john_allocco@merck.com
Donald
Robert
R
Zhong
Tanya
T
Lee
Anita
A
Tang
Yui Sing
YS
Hendrickson
Ronald C
RC
Liberator
Paul
P
Nare
Bakela
B
D000595:Amino Acid Sequence; D000818:Animals; D047389:Casein Kinase I / Q000037:antagonists & inhibitors* / Q000302:isolation & purification; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism*; D007093:Imidazoles / Q000378:metabolism; D007536:Isomerism; D018320:Leishmania major / Q000201:enzymology / Q000254:growth & development*; D008024:Ligands; D013058:Mass Spectrometry / Q000379:methods; D047428:Protein Kinase Inhibitors / Q000378:metabolism*; D015800:Protozoan Proteins / Q000032:analysis; D011725:Pyridines / Q000378:metabolism; D011758:Pyrroles / Q000378:metabolism; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification; D013379:Substrate Specificity; D053719:Tandem Mass Spectrometry / Q000379:methods; D014346:Trypanosoma brucei brucei / Q000378:metabolism
D007093:Imidazoles; D008024:Ligands; D047428:Protein Kinase Inhibitors; D015800:Protozoan Proteins; D011725:Pyridines; D011758:Pyrroles; D011994:Recombinant Proteins; D047389:Casein Kinase I; D017869:Cyclic GMP-Dependent Protein Kinases