Nitrogen-bisphosphonates block retinoblastoma phosphorylation and cell growth by inhibiting the cholesterol biosynthetic pathway in a keratinocyte model for esophageal irritation.
The surprising discovery that nitrogen-containing bisphosphonates (N-BPs) act via inhibition of the mevalonate-to-cholesterol pathway raised the possibility that esophageal irritation by N-BPs is mechanism-based. We used normal human epidermal keratinocytes (NHEKs) to model N-BP effects on stratified squamous epithelium of the esophagus. The N-BPs alendronate and risedronate inhibited NHEK growth in a dose-dependent manner without inducing apoptosis. N-BPs (30 microM) caused accumulation of cells in S phase and increased binucleation (inhibited cytokinesis). Consistent with N-BP inhibition of isoprenylation, geranylgeraniol or farnesol prevented accumulation in S phase. Binucleation was also induced by the 3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitor lovastatin and by the squalene synthase inhibitor zaragozic acid A and was prevented by adding low-density lipoprotein. At 300 microM, N-BPs reduced expression of cyclin-dependent kinase (cdk) 2 and cdk4 and enhanced expression of p21(waf1) and p27(kip1) and their binding to cdks with corollary hypophosphorylation of retinoblastoma. Lovastatin and zaragozic acid A produced similar effects, except that p21(waf1) expression and binding to cdks was not induced. Growth inhibition, but not binucleation, was also caused by the geranylgeranyl transferase I inhibitor, GGTI-298, which also enhanced cdk2 and cdk4 association with p27(kip1). These findings are consistent with suppression of epithelial cell growth by N-BPs via inhibition of the mevalonate pathway and the consequent reduction in cholesterol synthesis, which blocks cytokinesis, and in geranylgeranylation, which interferes with progression through the cell cycle.
Reszka
A A
AA
Department of Bone Biology and Osteoporosis Research, Merck Research Laboratories, West Point, Pennsylvania 19486, USA. alfred_reszka@merck.com
Halasy-Nagy
J
J
Rodan
G A
GA
D002453:Cell Cycle / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D002784:Cholesterol / Q000096:biosynthesis* / Q000378:metabolism; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D004164:Diphosphonates / Q000737:chemistry / Q000494:pharmacology*; D004935:Esophageal Diseases / Q000378:metabolism* / Q000473:pathology; D013186:Farnesyl-Diphosphate Farnesyltransferase / Q000037:antagonists & inhibitors; D006801:Humans; D015603:Keratinocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008148:Lovastatin / Q000494:pharmacology; D008798:Mevalonic Acid / Q000378:metabolism; D008954:Models, Biological; D010766:Phosphorylation / Q000187:drug effects; D017368:Protein Prenylation; D016160:Retinoblastoma Protein / Q000378:metabolism*
D004164:Diphosphonates; D016160:Retinoblastoma Protein; D002784:Cholesterol; D008148:Lovastatin; D013186:Farnesyl-Diphosphate Farnesyltransferase; D018844:Cyclin-Dependent Kinases; D008798:Mevalonic Acid