title: Mechanism and role of growth arrest in programmed (apoptotic) death of prostatic cancer cells induced by thapsigargin.
abstract: BACKGROUND
  More than 95% of metastatic androgen independent prostatic cancer cells per day are in a proliferatively quiescent G0 state [Berges et al.: Clin Cancer Res 1:473-480, 1995] limiting their responsiveness to anti-proliferative chemotherapeutic agents. Novel therapeutics capable of activating the programmed (apoptotic) death pathway in these cells without requiring entrance into the proliferative cell cycle are urgently needed. Thapsigargin (TG) treatment of rapidly growing androgen independent prostatic cancer cells arrests such cells in G0 and induces their programmed death. This raises not only the issue of the mechanism for such growth arrest, but also whether this programmed death is simply a response of rapidly growing cells to growth arrest making cytotoxicity still dependent upon the initial rate of cell proliferation.
  
  
  METHODS
  To resolve the mechanism of TG induced growth arrest, rat AT3.1 prostatic cancer cells were analyzed for RNA and protein expression of the growth arrest gene, gadd153, intracellular free Ca2+ levels (Cai), and cell cycle distribution on exposure to TG alone and in combination with Ca2+ chelation induced by BAPTA-AM or BAPTA-AM/EGTA. To resolve whether growth arrest is required for TG cytotoxicity, primary cultures of proliferatively quiescent, human prostatic cancer cells were exposed to TG.
  
  
  RESULTS
  Co-treatment of androgen independent AT-3 rat prostatic cancer cells with the Cai chelator BAPTA plus TG prevented growth arrest, as monitored by DNA flow cytometry, and failure to induce mRNA and protein for gadd153, demonstrating that growth arrest is due to Cai elevation, not depletion of intracellular Ca2+ pools. In addition, proliferatively quiescent G0 primary cultures of human prostatic cancer cells were resistant to anti-proliferative agents, but could be induced to undergo programmed death by TG as documented by morphological criteria and 14C-labeled DNA fragmentation assays.
  
  
  CONCLUSIONS
  These results demonstrate that TG with its ability to elevate Cai induces proliferating prostate cancer cells to growth arrest. Such Cai dependent growth arrest is not required, however, since TG can induce the programmed death of proliferatively quiescent G0 prostatic cancer cells without requiring either growth arrest or progression through the proliferative cell cycle.
authors.affiliation: Johns Hopkins Oncology Center, Johns Hopkins School of Medicine, Baltimore, Maryland, USA.
authors.forename: X S
authors.initials: XS
authors.lastname: Lin
authors.forename: S R
authors.initials: SR
authors.lastname: Denmeade
authors.forename: L
authors.initials: L
authors.lastname: Cisek
authors.forename: J T
authors.initials: JT
authors.lastname: Isaacs
mesh_terms: D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000494:pharmacology*; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D015152:Blotting, Northern; D015153:Blotting, Western; D002118:Calcium / Q000032:analysis; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D002614:Chelating Agents / Q000494:pharmacology*; D004249:DNA Damage / Q000187:drug effects* / Q000235:genetics; D004273:DNA, Neoplasm / Q000187:drug effects* / Q000235:genetics; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D005434:Flow Cytometry; D005467:Floxuridine / Q000494:pharmacology; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D006801:Humans; D008297:Male; D011471:Prostatic Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000503:physiopathology*; D012333:RNA, Messenger / Q000032:analysis; D012334:RNA, Neoplasm / Q000032:analysis; D051381:Rats; D019284:Thapsigargin / Q000494:pharmacology*; D013997:Time Factors; D014407:Tumor Cells, Cultured
chemical_list: D000964:Antimetabolites, Antineoplastic; D000970:Antineoplastic Agents; D002614:Chelating Agents; D004273:DNA, Neoplasm; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D005467:Floxuridine; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D019284:Thapsigargin; D002118:Calcium