TY - JOUR
T1 - Biochemical characterization of a mitomycin C-resistant human bladder cancer cell line
AU - Singh, Shivendra V.
AU - Scalamogna, Domenic
AU - Xia, Hong
AU - O'Toole, Stacy
AU - Roy, Deodutta
AU - Emerson, Erling O.
AU - Gupta, Vicram
AU - Zaren, Howard A.
PY - 1996/3/15
Y1 - 1996/3/15
N2 - This study describes characteristics of a mitomycin C (MMC)resistant human bladder cancer cell line, J82/MMC-2, which was established by repeated in vitro exposures of a 6-fold MMC-resistant variant (J82/MMC) to 18 nM MMC. A 9.6-fold higher concentration of MMC was required to kill 50% of the J82/MMC-2 sub-line compared with parental cells (J82/WT). NADPH cytochrome P450 reductase and DT-diaphorase activities were significantly lower in J82/MMC-2 cells compared with J82/WT, suggesting that reduced sensitivity of J82/MMC-2 cells to MMC resulted from impaired drug activation. Consistent with this hypothesis, the formation of MMC-alkylating metabolites was significantly lower in J82/MMC-2 cells compared with J82/WT. Furthermore, DT-diaphorase activity in J82/MMC-2 cells was significantly lower compared with the 6-fold MMC-resistant variant. Glutathione (GSH) levels were comparable in all 3 cell lines. Although GSH transferase (GST) activity was significantly higher in the J82/MMC-2 cells compared with J82/WT, this enzyme activity did not differ between 6- and 9.6-fold MMC-resistant variants. Whereas DNA polymerase a mRNA expression was comparable in these cell lines, levels of DNA ligase I mRNA were slightly lower in both MMC-resistant variants relative to J82/WT. However, the DNA polymerase β mRNA level was markedly higher in the J82/MMC-2 cell line compared with either J82/WT or 182/MMC. Thus, emergence of a higher level of resistance to MMC in J82/MMC-2 cells compared with J82/MMC may be attributed to (i) impaired drug activation through further reduction in DT-diaphorase activity and (ii) enhanced DNA repair through over-expression of DNA polymerase 8.
AB - This study describes characteristics of a mitomycin C (MMC)resistant human bladder cancer cell line, J82/MMC-2, which was established by repeated in vitro exposures of a 6-fold MMC-resistant variant (J82/MMC) to 18 nM MMC. A 9.6-fold higher concentration of MMC was required to kill 50% of the J82/MMC-2 sub-line compared with parental cells (J82/WT). NADPH cytochrome P450 reductase and DT-diaphorase activities were significantly lower in J82/MMC-2 cells compared with J82/WT, suggesting that reduced sensitivity of J82/MMC-2 cells to MMC resulted from impaired drug activation. Consistent with this hypothesis, the formation of MMC-alkylating metabolites was significantly lower in J82/MMC-2 cells compared with J82/WT. Furthermore, DT-diaphorase activity in J82/MMC-2 cells was significantly lower compared with the 6-fold MMC-resistant variant. Glutathione (GSH) levels were comparable in all 3 cell lines. Although GSH transferase (GST) activity was significantly higher in the J82/MMC-2 cells compared with J82/WT, this enzyme activity did not differ between 6- and 9.6-fold MMC-resistant variants. Whereas DNA polymerase a mRNA expression was comparable in these cell lines, levels of DNA ligase I mRNA were slightly lower in both MMC-resistant variants relative to J82/WT. However, the DNA polymerase β mRNA level was markedly higher in the J82/MMC-2 cell line compared with either J82/WT or 182/MMC. Thus, emergence of a higher level of resistance to MMC in J82/MMC-2 cells compared with J82/MMC may be attributed to (i) impaired drug activation through further reduction in DT-diaphorase activity and (ii) enhanced DNA repair through over-expression of DNA polymerase 8.
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U2 - 10.1002/(SICI)1097-0215(19960315)65:6<852::AID-IJC24>3.0.CO;2-4
DO - 10.1002/(SICI)1097-0215(19960315)65:6<852::AID-IJC24>3.0.CO;2-4
M3 - Article
C2 - 8631603
AN - SCOPUS:0029960764
SN - 0020-7136
VL - 65
SP - 852
EP - 857
JO - International Journal of Cancer
JF - International Journal of Cancer
IS - 6
ER -