Effects of bryostatin 1 and other pharmacological activators of protein kinase C on 1-[β-d-arabinofuranosyl]cytosine-induced apoptosis in HL-60 human promyelocytic leukemia cells

W. David Jarvis, Lawrence F. Povirk, Amy J. Turner, Rebecca S. Traylor, David A. Gewirtz, George Pettit, Steven Grant

Research output: Contribution to journalArticle

95 Citations (Scopus)

Abstract

We have demonstrated previously that bryostatin 1, a macrocyclic lactone with putative protein kinase C (PKC)-activating properties, synergistically augments the antileukemic actions of the deoxycytidine analog 1-[β-d-arabinofuranosyl]cytosine (ara-C) in HL-60 human promyelocytic leukemia cells (Grant et al., Biochem Pharmacol 42: 853-867, 1991), and that this effect appears to be related to sensitization to ara-C-induced apoptosis (Grant et al., Cancer Res 52: 6270-6278, 1992). In the present studies, we have assessed the extent of this damage by quantitative spectrofluorophotometry of small molecular weight, double-stranded DNA fragments in order to provide: (a) a more complete characterization of the interaction between ara-C and bryostatin 1, and (b) a direct comparison of the relative effects of bryostatin 1 treatment with other pharmacological manipulations known to modulate protein kinase C activity. Exposure of cells to ara-C (10-9 to 10-4 M; 1-24 hr) induced time- and concentration-related increases in the extent of DNA fragmentation. Treatment with bryostatin 1 (10-11 to 10-7 M; 1-24 hr) alone failed to induce DNA damage, but promoted substantial time- and concentration-related increases in the extent of fragmentation induced by a subsequent 6-hr exposure to ara-C. Maximal potentiation of fragmentation (e.g. 2- to 3-fold greater than that obtained with ara-C alone) was observed following a 24-hr pretreatment with 10-8 M or 10-7 M bryostatin 1, and correlated closely with enhanced inhibition of HL-60 cell clonogenicity. The stage-1 tumor-promoter phorbol dibutyrate potentiated the effects of ara-C in a biphasic manner, maximally augmenting the response at 2.5 × 10-8 M, but exerting no effect at 10-7 M, whereas the stage-2 tumor-promoter mezerein failed to augment ara-C-related DNA fragmentation at low concentrations, and antagonized ara-C action at high concentrations. In contrast, ara-C-related DNA fragmentation was attenuated or abolished either by continual preexposure to synthetic diglyceride or by pretreatment with exogenous phospholipase C at all concentrations tested. Increased DNA fragmentation was not specifically related to recruitment of cells into S-phase or enhancement of ara-C-related cellular differentiation. Finally, concentrations of bryostatin 1 that maximally potentiated ara-C-related DNA fragmentation were associated with virtually complete down-regulation of total cellular PKC activity, whereas diglyceride and phospholipase C, which suppressed the response to ara-C, moderately increased total PKC activity. Taken together, these findings provide a quantitative basis for assessing the ability of bryostatin 1 and related compounds to potentiate ara-C-induced damage to DNA, and underscore the complex and pleiotropic effects that modulation of the PKC family of isoenzymes may exert on ara-C-related apoptosis in human leukemia cells.

Original languageEnglish (US)
Pages (from-to)839-852
Number of pages14
JournalBiochemical Pharmacology
Volume47
Issue number5
DOIs
StatePublished - Mar 2 1994

Fingerprint

Cytosine
Protein Kinase C
Leukemia
Pharmacology
Apoptosis
DNA Fragmentation
DNA
Diglycerides
Type C Phospholipases
bryostatin 1
Carcinogens
DNA Damage
Deoxycytidine
HL-60 Cells
Cytarabine
Lactones
S Phase
Isoenzymes

Keywords

  • apoptosis
  • ara-C
  • bryostatin 1
  • leukemia
  • PKC activators
  • protein kinase C

ASJC Scopus subject areas

  • Pharmacology

Cite this

Effects of bryostatin 1 and other pharmacological activators of protein kinase C on 1-[β-d-arabinofuranosyl]cytosine-induced apoptosis in HL-60 human promyelocytic leukemia cells. / David Jarvis, W.; Povirk, Lawrence F.; Turner, Amy J.; Traylor, Rebecca S.; Gewirtz, David A.; Pettit, George; Grant, Steven.

In: Biochemical Pharmacology, Vol. 47, No. 5, 02.03.1994, p. 839-852.

Research output: Contribution to journalArticle

David Jarvis, W. ; Povirk, Lawrence F. ; Turner, Amy J. ; Traylor, Rebecca S. ; Gewirtz, David A. ; Pettit, George ; Grant, Steven. / Effects of bryostatin 1 and other pharmacological activators of protein kinase C on 1-[β-d-arabinofuranosyl]cytosine-induced apoptosis in HL-60 human promyelocytic leukemia cells. In: Biochemical Pharmacology. 1994 ; Vol. 47, No. 5. pp. 839-852.
@article{4d1281733ef74af99caed795d26c56bb,
title = "Effects of bryostatin 1 and other pharmacological activators of protein kinase C on 1-[β-d-arabinofuranosyl]cytosine-induced apoptosis in HL-60 human promyelocytic leukemia cells",
abstract = "We have demonstrated previously that bryostatin 1, a macrocyclic lactone with putative protein kinase C (PKC)-activating properties, synergistically augments the antileukemic actions of the deoxycytidine analog 1-[β-d-arabinofuranosyl]cytosine (ara-C) in HL-60 human promyelocytic leukemia cells (Grant et al., Biochem Pharmacol 42: 853-867, 1991), and that this effect appears to be related to sensitization to ara-C-induced apoptosis (Grant et al., Cancer Res 52: 6270-6278, 1992). In the present studies, we have assessed the extent of this damage by quantitative spectrofluorophotometry of small molecular weight, double-stranded DNA fragments in order to provide: (a) a more complete characterization of the interaction between ara-C and bryostatin 1, and (b) a direct comparison of the relative effects of bryostatin 1 treatment with other pharmacological manipulations known to modulate protein kinase C activity. Exposure of cells to ara-C (10-9 to 10-4 M; 1-24 hr) induced time- and concentration-related increases in the extent of DNA fragmentation. Treatment with bryostatin 1 (10-11 to 10-7 M; 1-24 hr) alone failed to induce DNA damage, but promoted substantial time- and concentration-related increases in the extent of fragmentation induced by a subsequent 6-hr exposure to ara-C. Maximal potentiation of fragmentation (e.g. 2- to 3-fold greater than that obtained with ara-C alone) was observed following a 24-hr pretreatment with 10-8 M or 10-7 M bryostatin 1, and correlated closely with enhanced inhibition of HL-60 cell clonogenicity. The stage-1 tumor-promoter phorbol dibutyrate potentiated the effects of ara-C in a biphasic manner, maximally augmenting the response at 2.5 × 10-8 M, but exerting no effect at 10-7 M, whereas the stage-2 tumor-promoter mezerein failed to augment ara-C-related DNA fragmentation at low concentrations, and antagonized ara-C action at high concentrations. In contrast, ara-C-related DNA fragmentation was attenuated or abolished either by continual preexposure to synthetic diglyceride or by pretreatment with exogenous phospholipase C at all concentrations tested. Increased DNA fragmentation was not specifically related to recruitment of cells into S-phase or enhancement of ara-C-related cellular differentiation. Finally, concentrations of bryostatin 1 that maximally potentiated ara-C-related DNA fragmentation were associated with virtually complete down-regulation of total cellular PKC activity, whereas diglyceride and phospholipase C, which suppressed the response to ara-C, moderately increased total PKC activity. Taken together, these findings provide a quantitative basis for assessing the ability of bryostatin 1 and related compounds to potentiate ara-C-induced damage to DNA, and underscore the complex and pleiotropic effects that modulation of the PKC family of isoenzymes may exert on ara-C-related apoptosis in human leukemia cells.",
keywords = "apoptosis, ara-C, bryostatin 1, leukemia, PKC activators, protein kinase C",
author = "{David Jarvis}, W. and Povirk, {Lawrence F.} and Turner, {Amy J.} and Traylor, {Rebecca S.} and Gewirtz, {David A.} and George Pettit and Steven Grant",
year = "1994",
month = "3",
day = "2",
doi = "10.1016/0006-2952(94)90484-7",
language = "English (US)",
volume = "47",
pages = "839--852",
journal = "Biochemical Pharmacology",
issn = "0006-2952",
publisher = "Elsevier Inc.",
number = "5",

}

TY - JOUR

T1 - Effects of bryostatin 1 and other pharmacological activators of protein kinase C on 1-[β-d-arabinofuranosyl]cytosine-induced apoptosis in HL-60 human promyelocytic leukemia cells

AU - David Jarvis, W.

AU - Povirk, Lawrence F.

AU - Turner, Amy J.

AU - Traylor, Rebecca S.

AU - Gewirtz, David A.

AU - Pettit, George

AU - Grant, Steven

PY - 1994/3/2

Y1 - 1994/3/2

N2 - We have demonstrated previously that bryostatin 1, a macrocyclic lactone with putative protein kinase C (PKC)-activating properties, synergistically augments the antileukemic actions of the deoxycytidine analog 1-[β-d-arabinofuranosyl]cytosine (ara-C) in HL-60 human promyelocytic leukemia cells (Grant et al., Biochem Pharmacol 42: 853-867, 1991), and that this effect appears to be related to sensitization to ara-C-induced apoptosis (Grant et al., Cancer Res 52: 6270-6278, 1992). In the present studies, we have assessed the extent of this damage by quantitative spectrofluorophotometry of small molecular weight, double-stranded DNA fragments in order to provide: (a) a more complete characterization of the interaction between ara-C and bryostatin 1, and (b) a direct comparison of the relative effects of bryostatin 1 treatment with other pharmacological manipulations known to modulate protein kinase C activity. Exposure of cells to ara-C (10-9 to 10-4 M; 1-24 hr) induced time- and concentration-related increases in the extent of DNA fragmentation. Treatment with bryostatin 1 (10-11 to 10-7 M; 1-24 hr) alone failed to induce DNA damage, but promoted substantial time- and concentration-related increases in the extent of fragmentation induced by a subsequent 6-hr exposure to ara-C. Maximal potentiation of fragmentation (e.g. 2- to 3-fold greater than that obtained with ara-C alone) was observed following a 24-hr pretreatment with 10-8 M or 10-7 M bryostatin 1, and correlated closely with enhanced inhibition of HL-60 cell clonogenicity. The stage-1 tumor-promoter phorbol dibutyrate potentiated the effects of ara-C in a biphasic manner, maximally augmenting the response at 2.5 × 10-8 M, but exerting no effect at 10-7 M, whereas the stage-2 tumor-promoter mezerein failed to augment ara-C-related DNA fragmentation at low concentrations, and antagonized ara-C action at high concentrations. In contrast, ara-C-related DNA fragmentation was attenuated or abolished either by continual preexposure to synthetic diglyceride or by pretreatment with exogenous phospholipase C at all concentrations tested. Increased DNA fragmentation was not specifically related to recruitment of cells into S-phase or enhancement of ara-C-related cellular differentiation. Finally, concentrations of bryostatin 1 that maximally potentiated ara-C-related DNA fragmentation were associated with virtually complete down-regulation of total cellular PKC activity, whereas diglyceride and phospholipase C, which suppressed the response to ara-C, moderately increased total PKC activity. Taken together, these findings provide a quantitative basis for assessing the ability of bryostatin 1 and related compounds to potentiate ara-C-induced damage to DNA, and underscore the complex and pleiotropic effects that modulation of the PKC family of isoenzymes may exert on ara-C-related apoptosis in human leukemia cells.

AB - We have demonstrated previously that bryostatin 1, a macrocyclic lactone with putative protein kinase C (PKC)-activating properties, synergistically augments the antileukemic actions of the deoxycytidine analog 1-[β-d-arabinofuranosyl]cytosine (ara-C) in HL-60 human promyelocytic leukemia cells (Grant et al., Biochem Pharmacol 42: 853-867, 1991), and that this effect appears to be related to sensitization to ara-C-induced apoptosis (Grant et al., Cancer Res 52: 6270-6278, 1992). In the present studies, we have assessed the extent of this damage by quantitative spectrofluorophotometry of small molecular weight, double-stranded DNA fragments in order to provide: (a) a more complete characterization of the interaction between ara-C and bryostatin 1, and (b) a direct comparison of the relative effects of bryostatin 1 treatment with other pharmacological manipulations known to modulate protein kinase C activity. Exposure of cells to ara-C (10-9 to 10-4 M; 1-24 hr) induced time- and concentration-related increases in the extent of DNA fragmentation. Treatment with bryostatin 1 (10-11 to 10-7 M; 1-24 hr) alone failed to induce DNA damage, but promoted substantial time- and concentration-related increases in the extent of fragmentation induced by a subsequent 6-hr exposure to ara-C. Maximal potentiation of fragmentation (e.g. 2- to 3-fold greater than that obtained with ara-C alone) was observed following a 24-hr pretreatment with 10-8 M or 10-7 M bryostatin 1, and correlated closely with enhanced inhibition of HL-60 cell clonogenicity. The stage-1 tumor-promoter phorbol dibutyrate potentiated the effects of ara-C in a biphasic manner, maximally augmenting the response at 2.5 × 10-8 M, but exerting no effect at 10-7 M, whereas the stage-2 tumor-promoter mezerein failed to augment ara-C-related DNA fragmentation at low concentrations, and antagonized ara-C action at high concentrations. In contrast, ara-C-related DNA fragmentation was attenuated or abolished either by continual preexposure to synthetic diglyceride or by pretreatment with exogenous phospholipase C at all concentrations tested. Increased DNA fragmentation was not specifically related to recruitment of cells into S-phase or enhancement of ara-C-related cellular differentiation. Finally, concentrations of bryostatin 1 that maximally potentiated ara-C-related DNA fragmentation were associated with virtually complete down-regulation of total cellular PKC activity, whereas diglyceride and phospholipase C, which suppressed the response to ara-C, moderately increased total PKC activity. Taken together, these findings provide a quantitative basis for assessing the ability of bryostatin 1 and related compounds to potentiate ara-C-induced damage to DNA, and underscore the complex and pleiotropic effects that modulation of the PKC family of isoenzymes may exert on ara-C-related apoptosis in human leukemia cells.

KW - apoptosis

KW - ara-C

KW - bryostatin 1

KW - leukemia

KW - PKC activators

KW - protein kinase C

UR - http://www.scopus.com/inward/record.url?scp=0028181584&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0028181584&partnerID=8YFLogxK

U2 - 10.1016/0006-2952(94)90484-7

DO - 10.1016/0006-2952(94)90484-7

M3 - Article

C2 - 8135859

AN - SCOPUS:0028181584

VL - 47

SP - 839

EP - 852

JO - Biochemical Pharmacology

JF - Biochemical Pharmacology

SN - 0006-2952

IS - 5

ER -