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In vitro effects of bryostatin 1 on the metabolism and cytotoxicity of 1-beta-D-arabinofuranosylcytosine in human leukemia cells.

Abstract
Bryostatin 1 is a macrocyclic lactone protein kinase C (PK-C) activator which has demonstrated promising antileukemic activity in preclinical studies. We have examined the effect of this agent on the metabolism and cytotoxicity of 1-beta-D-arabinofuranosylcytosine (ara-C) in both log phase and high-density human promyelocytic leukemia cells (HL-60). Exposure of low-density cells to 12.5 nM bryostatin 1 for 24 hr prior to a 4-hr incubation with 1 or 10 microM ara-C resulted in nearly a 2-fold increase in ara-CTP formation. When cells were maintained under high-cell density conditions (e.g. 5 x 10(6) cells/mL) for 24 hr prior to ara-C exposure, a 90% reduction in ara-CTP formation and ara-C DNA incorporation was observed. However, coincubation of high-density cells with bryostatin 1 for 24 hr increased ara-CTP formation 6- to 8-fold, yielding levels essentially equivalent to those achieved in low-density cells. Smaller (but still significant) increases in ara-C DNA incorporation were also noted. Enhancement of ara-CTP formation by bryostatin 1 occurred over a broad ara-C concentration range (0.1 to 100 microM), involved a temperature-dependent process, could not be mimicked by addition of hematopoietic growth factors, and was not related to neutralization of toxic or inhibitory substances in high-density medium. Exposure of cells to bryostatin 1 did not lead to morphologic or functional evidence of HL-60 cell maturation or an increase in cell viability, but did produce a decline in cellular proliferative activity as determined by thymidine and bromodeoxyuridine incorporation and cytofluorometric analysis. Bryostatin 1 did not exert its effects in high-density cells by inhibiting ara-C deamination or by interfering with ara-CTP dephosphorylation, but instead appeared to act by enhancing ara-C phosphorylation. Although cell-free extracts obtained from high-density cells exposed to bryostatin 1 exhibited levels of deoxycytidine kinase activity compared to controls, treated cells did display a significant decline in intracellular dCTP levels (e.g. 0.7 vs 1.3 pmol/10(6)), and nearly a 2-fold increase in ATP and UTP concentrations. Ara-CTP formation was also increased substantially by other PK-C activators including phorbol dibutyrate and mezerein (10-100 nM); this process was inhibited more than 70% by the PK-C inhibitor H-7 (50 microM), but not by the PK-C inhibitors staurosporine, tamoxifen, and HA1004. Finally, coadministration of ara-C and bryostatin 1 resulted in greater than expected inhibitory effects toward HL-60 cell clonogenic growth.(ABSTRACT TRUNCATED AT 400 WORDS)
AuthorsS Grant, L Boise, E Westin, C Howe, G R Pettit, A Turner, C McCrady
JournalBiochemical pharmacology (Biochem Pharmacol) Vol. 42 Issue 4 Pg. 853-67 (Jul 25 1991) ISSN: 0006-2952 [Print] England
PMID1867641 (Publication Type: Journal Article, Research Support, U.S. Gov't, P.H.S.)
Chemical References
  • Antineoplastic Agents
  • Bryostatins
  • DNA, Neoplasm
  • Diterpenes
  • Lactones
  • Macrolides
  • Terpenes
  • Cytarabine
  • Arabinofuranosylcytosine Triphosphate
  • mezerein
  • Phorbol 12,13-Dibutyrate
  • bryostatin 1
Topics
  • Antineoplastic Agents (pharmacology)
  • Arabinofuranosylcytosine Triphosphate (metabolism)
  • Bryostatins
  • Cell Count
  • Cytarabine (metabolism, toxicity)
  • DNA, Neoplasm (drug effects, metabolism)
  • Diterpenes
  • Humans
  • Kinetics
  • Lactones (pharmacology)
  • Leukemia, Experimental (drug therapy, metabolism, pathology)
  • Leukemia, Myeloid (drug therapy, metabolism, pathology)
  • Macrolides
  • Phorbol 12,13-Dibutyrate (pharmacology)
  • Phosphorylation
  • Terpenes (pharmacology)
  • Time Factors
  • Tumor Cells, Cultured

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