Inhibition of MG-63 cell cycle progression by synthetic vitamin D3 analogs mediated by p27, Cdk2, cyclin E, and the retinoblastoma protein.
Progression through eukaryotic cell division cycle is regulated by synergistic activities of both positive and negative regulatory factors. The active form of vitamin D(3) (1alpha,25(OH)(2)D(3), 1,25D) and a number of its synthetic analogs have been shown to arrest cells in the G(1) phase of the cell cycle. In the present study, 1alpha,25(OH)(2)D(3) and the analogs KH1060, EB1089, and CB1093 were used to study the mechanism of the cell cycle arrest and to compare the effectiveness of these compounds in human MG-63 osteosarcoma cells. The 20-epi analogs KH1060 and CB1093, as well as the 20-normal analog EB1089, were found to be more potent than 1alpha,25(OH)(2)D(3) in inhibiting cell proliferation and arresting the MG-63 cells in the G(1) phase. These analogs were more active than 1alpha,25(OH)(2)D(3) in increasing the cyclin dependent kinase inhibitor p27 protein levels (approximately 2.3-2.5-fold compared to 1alpha,25(OH)(2)D(3)) by both increasing its formation and decreasing its degradation rate. The increased p27 formation was accompanied by stabilization of binding of nuclear proteins to the Sp1+NF-Y responsive promoter region of the p27 gene. The increase in p27 protein levels and the simultaneous decrease in cyclin E protein levels was accompanied by decreased Cdk2 kinase activity, retinoblastoma (Rb) protein hypophosphorylation and, finally, cell cycle arrest in the G(1) phase. In summary, the analogs KH1060, EB1089, and CB1093 keep Rb protein in its growth-suppressing, hypophosphorylated form and prevent cell cycle progression through the restriction point. Therefore, these synthetic vitamin D(3) analogs may be potential candidates for treating diseases, where cell cycle regulation is needed.
Ryhänen
Sanna
S
Department of Biochemistry, University of Kuopio, Kuopio, Finland. sanna.ryhanen@uku.fi
Jääskeläinen
Tiina
T
Mahonen
Anitta
A
Mäenpää
Pekka H
PH
D042846:CDC2-CDC28 Kinases; D002453:Cell Cycle / Q000187:drug effects*; D002762:Cholecalciferol / Q000138:chemical synthesis / Q000494:pharmacology*; D019927:Cyclin E / Q000378:metabolism*; D051357:Cyclin-Dependent Kinase 2; D051358:Cyclin-Dependent Kinase 4; D051361:Cyclin-Dependent Kinase 6; D050759:Cyclin-Dependent Kinase Inhibitor p21; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D016213:Cyclins / Q000378:metabolism; D016193:G1 Phase / Q000187:drug effects; D006801:Humans; D008840:Microfilament Proteins / Q000378:metabolism*; D009124:Muscle Proteins; D011401:Promoter Regions, Genetic / Q000187:drug effects / Q000502:physiology; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins; D016160:Retinoblastoma Protein / Q000378:metabolism*; D014407:Tumor Cells, Cultured
C493956:CDKN1A protein, human; D019927:Cyclin E; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D008840:Microfilament Proteins; D009124:Muscle Proteins; D011518:Proto-Oncogene Proteins; D016160:Retinoblastoma Protein; C058992:Tagln protein, mouse; D002762:Cholecalciferol; D017346:Protein Serine-Threonine Kinases; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; C495900:CDK4 protein, human; C495911:CDK6 protein, human; D051357:Cyclin-Dependent Kinase 2; D051358:Cyclin-Dependent Kinase 4; D051361:Cyclin-Dependent Kinase 6; D018844:Cyclin-Dependent Kinases