Identification of protein binding partners of ALK-5 kinase inhibitors.
We have investigated the binding characteristics of a potent member of the bis-ortho-substituted five-membered nitrogen heterocycle class of ALK-5 kinase inhibitors with lysates of cultured HEK-293 cells to identify protein binding partners of potential biological significance. An affinity chromatographic resin containing an immobilized ALK-5 kinase inhibitor, 2-phenyl-4-[3-(pyridin-2-yl)-1H-pyrazol-4-yl]pyridine, was used to capture specific proteins from the cell lysate. The soluble inhibitor was then used to specifically elute the proteins which selectively bound to the pharmacophore ligand structure. Application of 2-D SDS-PAGE analysis with positive and negative controls demonstrated the inhibitor bound several different proteins via selective molecular recognition processes. The structural features of the specifically eluted proteins were identified by peptide mass fingerprinting (PMF) methods and included proteins with structural, metabolic and chaperone functions. Furthermore, these PMF results identified the therapeutic target in various cancer treatment studies, HSP-70, as a potential high-affinity binding partner. These observations warrant examination of bis-ortho-substituted five-membered nitrogen heterocycles as dual ALK-5/HSP-70 inhibitors for anti-cancer drug development.
Centre for Green Chemistry, Monash University, Clayton, Victoria 3800, Australia.
Rudy
R
Ciayadi
Geoffrey F
GF
Kelso
Mahesh K
MK
Potdar
Simon J
SJ
Harris
Kelly L
KL
Walton
Craig A
CA
Harrison
Milton T W
MT
Hearn
D000970:Antineoplastic Agents / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D057809:HEK293 Cells; D018840:HSP70 Heat-Shock Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D006801:Humans; D011485:Protein Binding; D047428:Protein Kinase Inhibitors / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D011725:Pyridines / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism; D000077293:Receptor, Transforming Growth Factor-beta Type I; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism
D000970:Antineoplastic Agents; D018840:HSP70 Heat-Shock Proteins; D047428:Protein Kinase Inhibitors; D011725:Pyridines; D018125:Receptors, Transforming Growth Factor beta; D017346:Protein Serine-Threonine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I; C000625841:TGFBR1 protein, human; C023666:pyridine
Activin-like kinase-5; Affinity chromatography; Kinase inhibitor; Kinase selectivity; Transforming growth factor β-1