Factors affecting topotecan-induced programmed cell death: adhesion protects cells from apoptosis and impairs cleavage of poly(ADP-ribose)polymerase.
We have evaluated the influence of anchorage status together with endogenous levels of bcl-2 family members on the ability of the topoisomerase I inhibitor, topotecan (TPT), to induce programmed cell death (PCD) in human colon, breast, lymphoid, and cervical cancer cell lines. As part of this study, we assessed the use of measuring poly(ADP-ribose) polymerase (PARP) cleavage by Western blot, as an index of apoptosis, relative to measuring chromatin condensation by acridine orange analysis. Our results show a strong correlation between both assays, indicating that PARP cleavage is an accurate method to examine PCD. We have encountered a strong association between cell attachment and sensitivity to TPT-induced PCD. Cells growing attached to flasks appear to be relatively more resistant than suspension-growing cells in spite of endogenous bcl-2, bax, or bcl-x levels. Furthermore, we demonstrate that interference with attachment status alters the sensitivity of cells to TPT-induced PCD. Although cell attachment to ProNectin F confers protection against TPT-induced chromatin condensation and cleavage of PARP, cell detachment by poly(2-hydroxyethyl methacrylate) stimulates TPT-induced PCD and PARP cleavage.
Whitacre
C M
CM
Department of Medicine and Cancer Center, Case Western Reserve University, School of Medicine, Cleveland, Ohio 44106-4937, USA.
Berger
N A
NA
D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis; D015153:Blotting, Western; D002166:Camptothecin / Q000031:analogs & derivatives* / Q000494:pharmacology; D002448:Cell Adhesion; D002843:Chromatin / Q000378:metabolism; D005353:Fibronectins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008689:Methacrylates / Q000494:pharmacology; D009369:Neoplasms / Q000473:pathology*; D011065:Poly(ADP-ribose) Polymerases / Q000276:immunology / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000276:immunology / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000276:immunology / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D019772:Topotecan; D014407:Tumor Cells, Cultured; D051028:bcl-2-Associated X Protein
D000970:Antineoplastic Agents; C494826:BAX protein, human; D002843:Chromatin; D005353:Fibronectins; D008689:Methacrylates; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D011994:Recombinant Proteins; D051028:bcl-2-Associated X Protein; C070367:ProNectin F; C005044:hydroxyethyl methacrylate; D019772:Topotecan; D011065:Poly(ADP-ribose) Polymerases; D002166:Camptothecin