Development and characterization of a WEHI-3B D+ monomyelocytic leukemia cell line resistant to novobiocin and cross-resistant to other topoisomerase II-targeted drugs.
A novobiocin-resistant subline of WEHI-3B D+ murine monomyelocytic leukemia cells was developed by the continuous exposure of cells to this agent in vitro. Sensitive (WEHI-3B/S) and novobiocin-resistant (WEHI-3B/NOVO) sublines were cloned in vitro. WEHI-3B/NOVO cells were stable in the absence of novobiocin for more than 3 months, and the sensitive and resistant clones displayed the same growth rate, cell cycle distribution, cell size, DNA and protein content, and cloning efficiency. Novobiocin has been shown to compete with ATP for the ATP-binding site of topoisomerase II; therefore, intracellular ATP levels can influence the cellular sensitivity to novobiocin. High-performance liquid chromatographic analysis of total cell extracts demonstrated that no difference exists between WEHI-3B/S and WEHI-3B/NOVO cells in the content of ATP. Furthermore, exposure of both cell lines to novobiocin did not affect intracellular ATP levels. In addition to an approximately 2-fold level of resistance to novobiocin, the WEHI-3B/NOVO subline was also 7- and 11-fold cross-resistant to the topoisomerase II-targeted drugs, teniposide and etoposide (VP-16), respectively. A lower level of cross-resistance, comparable to that of novobiocin, was observed in WEHI-3B/NOVO cells for the intercalating topoisomerase II-reactive drugs, doxorubicin, 4'-(9-acridinylamino)methanesulfon-m-anisidide and aclacinomycin A, while the sensitivity to the cytotoxic action of the non-topoisomerase II-acting agents, camptothecin and vincristine, was not altered. After 3-6 h of exposure to 1 microM VP-16, WEHI-3B/S cells accumulated in the S and G2 + M phases of the cell cycle. Similar changes were detected in WEHI-3B/NOVO cells only after exposure to a 10-fold higher concentration of VP-16. Exposure to 150 microM novobiocin caused an accumulation of WEHI-3B/S cells in the G0-G1 phase of the cell cycle but did not affect the cell cycle distribution of WEHI-3B/NOVO cells, while camptothecin induced the same type and extent of changes in the cell cycle distribution of both cell lines. Although the WEHI-3B/NOVO subline appeared to be less responsive to the differentiation-inducing activity of novobiocin and teniposide, the capacity of WEHI-3B/NOVO cells to respond to the differentiation-inducing agent 13-cis-retinoic acid was not significantly different from that of WEHI-3B/S cells. A slight decrease in the accumulation of VP-16 occurred in the resistant cell line, which did not appear to be of sufficient magnitude to account for the 11-fold increase in the degree of resistance to this agent.(ABSTRACT TRUNCATED AT 400 WORDS)
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Department of Pharmacology, Yale University School of Medicine, New Haven, Connecticut 06510.
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Sartorelli
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D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D002453:Cell Cycle / Q000187:drug effects; D004250:DNA Topoisomerases, Type II / Q000187:drug effects* / Q000378:metabolism; D004273:DNA, Neoplasm / Q000378:metabolism; D004351:Drug Resistance; D005047:Etoposide / Q000493:pharmacokinetics / Q000494:pharmacology; D007942:Leukemia, Experimental / Q000188:drug therapy / Q000201:enzymology / Q000473:pathology*; D015479:Leukemia, Myelomonocytic, Acute / Q000188:drug therapy / Q000201:enzymology / Q000473:pathology*; D051379:Mice; D009675:Novobiocin / Q000494:pharmacology*; D059005:Topoisomerase II Inhibitors; D014316:Tritium; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D004273:DNA, Neoplasm; D059005:Topoisomerase II Inhibitors; D014316:Tritium; D009675:Novobiocin; D005047:Etoposide; D000255:Adenosine Triphosphate; D004250:DNA Topoisomerases, Type II