Major contribution of the multidrug transporter P-glycoprotein to reduced susceptibility of poly(ADP-ribose) polymerase-1 knock-out cells to doxorubicin action.
Inactivation of poly(ADP-ribose) polymerase-1 (PARP-1) has been shown to potentiate the cytotoxicity of distinct DNA targeting agents including topoisomerase I inhibitors. On the other hand, the PARP-1 deficient cells exhibited resistance to conventional inhibitors of topoisomerase II such as etoposide or doxorubicin (DOX). Recently, we observed the extreme sensitivity of PARP-1 knock-out (KO) cells to C-1305, a new biologically active triazoloacridone compound. C-1305 permanently arrested the cells in G2-phase of the cell-cycle. These observations prompted us to investigate more thoroughly the susceptibility of PARP-1 KO cells to DOX and to examine the effect of DOX on the progression of cell-cycle. We determined the uptake of DOX and P-glycoprotein (P-gp) expression in mouse cells and compared it with that in human myeloma 8226/Dox40 cells overexpressing P-gp. Exposure of mouse cells to DOX revealed a reduced drug uptake in cells lacking PARP-1. However, combined treatment with verapamil, a potent MDR modulator increased the DOX accumulation. Detailed immunoblotting experiments revealed an approximately threefold higher P-gp level in PARP-1 KO cells as compared with normal counterparts. Interestingly, DOX induced in normal fibroblasts very rapidly G2 arrest whereas in PARP-1 KO cells it blocked primarily the transition between S and G2 resulting in the increase of cells remaining in S-phase. This coincided with the lack of the site-specific phosphorylation of CDK2. Simultaneous inhibition of P-gp in cells lacking PARP-1 resulted in an accumulation of cells in G2. Exposure of mouse cells to high DOX dose activated significantly caspase-3/7 in PARP-1 KO cells.
Wesierska-Gadek
Józefa
J
Department of Medicine I, Cell Cycle Regulation Group, Institute of Cancer Research, Medical University of Vienna, Vienna, Austria. Jozefa.Gadek-Wesierski@meduniwien.ac.at
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism*; D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000378:metabolism* / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D002121:Calcium Channel Blockers / Q000494:pharmacology; D053148:Caspase 3; D053179:Caspase 7; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured / Q000187:drug effects; D004317:Doxorubicin / Q000378:metabolism* / Q000494:pharmacology; D018432:Drug Resistance, Multiple; D019008:Drug Resistance, Neoplasm; D004359:Drug Therapy, Combination; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D016195:G2 Phase / Q000187:drug effects; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D009101:Multiple Myeloma / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D010766:Phosphorylation / Q000187:drug effects; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases / Q000235:genetics / Q000502:physiology*; D016196:S Phase / Q000187:drug effects; D014700:Verapamil / Q000494:pharmacology
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000903:Antibiotics, Antineoplastic; D000970:Antineoplastic Agents; D002121:Calcium Channel Blockers; D004317:Doxorubicin; D014700:Verapamil; C499321:Parp1 protein, mouse; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; C505407:CASP3 protein, human; C505520:CASP7 protein, human; C505409:Casp3 protein, mouse; C505524:Casp7 protein, mouse; D053148:Caspase 3; D053179:Caspase 7; D020169:Caspases