Expression of serine/threonine protein-kinases and related factors in normal monkey and human retinas: the mechanistic understanding of a CDK2 inhibitor induced retinal toxicity.
Protein-kinase inhibitors are among the most advanced compounds in development using the new drug discovery paradigm of developing small-molecule drugs against specific molecular targets in cancer. After treatment with a cyclin dependent kinase CDK2 inhibitor in monkey, histopathological analysis of the eye showed specific cellular damage in the photoreceptor layer. Since this CDK2 inhibitor showed activity also on other CDKs, in order to investigate the mechanism of toxicity of this compound, we isolated cones and rods from the retina of normal monkey and humans by Laser Capture Microdissection. Using Real-Time PCR we first measured the expression of cyclin dependent protein-kinases (CDK)1, 2, 4, 5, Glycogen synthase kinase 3beta (GSK3beta) and microtubule associated protein TAU. We additionally verified the presence of these proteins in monkey eye sections by immuno-histochemistry and immunofluorescence analysis and afterwards quantified GSK3beta, phospho-GSK3beta and TAU by Reverse Phase Protein Microarrays. With this work we demonstrate how complementary gene expression and protein-based technologies constitute a powerful tool for the understanding of the molecular mechanism of a CDK2 inhibitor induced toxicity. Moreover, this investigative approach is helpful to better understand and characterize the mechanism of species-specific toxicities and further support a rational, molecular mechanism-based safety assessment in humans.
D000818:Animals; D051357:Cyclin-Dependent Kinase 2 / Q000037:antagonists & inhibitors*; D005260:Female; D005455:Fluorescent Antibody Technique; D038362:Glycogen Synthase Kinase 3 / Q000032:analysis; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D007150:Immunohistochemistry; D008252:Macaca fascicularis; D008297:Male; D042282:Microdissection; D010766:Phosphorylation; D016133:Polymerase Chain Reaction; D047428:Protein Kinase Inhibitors / Q000633:toxicity*; D017346:Protein Serine-Threonine Kinases / Q000032:analysis*; D012160:Retina / Q000187:drug effects* / Q000201:enzymology* / Q000473:pathology; D016875:tau Proteins / Q000032:analysis
D047428:Protein Kinase Inhibitors; D016875:tau Proteins; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; D017346:Protein Serine-Threonine Kinases; D051357:Cyclin-Dependent Kinase 2; D038362:Glycogen Synthase Kinase 3