title: Cyclin-dependent kinase inhibitor, P276-00, inhibits HIF-1α and induces G2/M arrest under hypoxia in prostate cancer cells.
abstract: BACKGROUND
  Hypoxia-inducible factor-1 (HIF-1) is a master regulator of the transcriptional response to oxygen deprivation and controls genes involved in glycolysis, angiogenesis, migration and invasion. Overexpression of HIF-1α has been demonstrated in many common human cancers.
  
  
  METHODS
  Luciferase reporter gene assay under hypoxia and normoxia was used to demonstrate transcriptional inhibition of HIF-1 by P276-00. Detailed studies such as western blotting, reverse-transcriptase-PCR and immunofluorescence were carried out to elucidate its mechanism of action. Cytotoxic potential of P276-00 under normoxia and hypoxia was determined on prostate cancer cells using CCK-8 assay, and cell-cycle analysis was carried out using flow cytometry. Antiangiogenic activity of P276-00 was demonstrated by migration assay and tube-formation assay. Efficacy study of P276-00 was performed in a PC-3 xenograft model.
  
  
  RESULTS
  P276-00 inhibits transcriptional activation of HIF-1 under hypoxia. It suppressed hypoxia-mediated nuclear HIF-1α expression, as well as phosphorylation of Akt and 4E-BP1 and abrogated expression of HIF-1-inducible gene viz. vascular endothelial growth factor. Under hypoxia, P276-00 did not exhibit enhanced cytotoxic activity in prostate cancer cells but arrested them in the G2/M phase of the cell cycle. The tubular formation of human umbilical vein endothelial cells and migration of prostate cancer cells were also inhibited by P276-00 in vitro. In addition, it demonstrated significant in vivo efficacy in the PC-3 xenograft model.
  
  
  CONCLUSIONS
  Given its low toxicity profile, its demonstrated antitumor activity and its potential to inhibit the HIF-1 pathway, P276-00 should be considered as antiangiogenic chemotherapy for prostate cancer.
authors.affiliation: Oncology Franchisee, Cancer Pharmacology, Piramal Life Sciences, Mumbai, Maharashtra, India.
authors.forename: S M
authors.initials: SM
authors.lastname: Manohar
authors.forename: A A
authors.initials: AA
authors.lastname: Padgaonkar
authors.forename: A
authors.initials: A
authors.lastname: Jalota-Badhwar
authors.forename: S V
authors.initials: SV
authors.lastname: Rao
authors.forename: K S
authors.initials: KS
authors.lastname: Joshi
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology* / Q000627:therapeutic use; D015687:Cell Hypoxia; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D019938:Cyclin D1 / Q000235:genetics / Q000378:metabolism; D004357:Drug Synergism; D047309:Flavones / Q000494:pharmacology* / Q000627:therapeutic use; D059565:G2 Phase Cell Cycle Checkpoints / Q000187:drug effects*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D055785:Gene Knockdown Techniques; D017930:Genes, Reporter; D061307:Human Umbilical Vein Endothelial Cells / Q000187:drug effects / Q000502:physiology; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D008156:Luciferases / Q000096:biosynthesis / Q000235:genetics; D008297:Male; D051379:Mice; D016513:Mice, SCID; D011471:Prostatic Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology* / Q000627:therapeutic use; D059748:Proteolysis; D034741:RNA, Small Interfering / Q000235:genetics / Q000494:pharmacology; D011897:Random Allocation; D015398:Signal Transduction / Q000187:drug effects; D015533:Transcriptional Activation / Q000187:drug effects; D023041:Xenograft Model Antitumor Assays
chemical_list: D000970:Antineoplastic Agents; D047309:Flavones; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; C520467:P276-00; D047428:Protein Kinase Inhibitors; D034741:RNA, Small Interfering; D019938:Cyclin D1; D008156:Luciferases; D046988:Proteasome Endopeptidase Complex