Induction of thymidylate synthase associated with multidrug resistance in human breast and colon cancer cell lines

E. Chu, J. C. Drake, D. M. Koeller, S. Zinn, Carlos Alberto Jamis-Dow, G. C. Yeh, C. J. Allegra

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Abstract

A series of Adriamycin-resistant human breast MCF-7 and human colon DLD-1 cancer cell lines were established by stepwise selection. The concentration of Adriamycin required to inhibit cell proliferation by 50% (IC50) in the parent breast line (MCF-7), Adriamycin-resistant lines (MCF-Ad5 and MCF-Adl 0), and a 5-fluorouracil (5-FU)-revertant line (MCF-R) was 0.005, 3.3, 6, and 4.9 μM, respectively. The Adriamycin IC50 value for the resistant colon line (DLD-Ad) was 8.2 μM, 68-fold higher than that for its parent line (DLD-1) (IC50 = 0.12 μM). The MCF-Ad5 and MCF-Ad10 cells were cross-resistant to 5-FU, with respective 5-FU IC50 values of 11.7 and 22.5 μM, or 7.3- and 14-fold less sensitive than their parent MCF-7 (IC50 = 1.6 μM) line. The MCF-R line completely reverted in sensitivity to 5-FU, with an IC50 of 1.7 μM. The resistant DLD-Ad line was 3.5-fold more resistant to 5-FU than was the parent DLD-1 line. Using both the 5-fluoro-2'-deoxyuridine-5'-monophosphate binding and catalytic assays for measurement of thymidylate synthase (TS) activity, there was significantly increased TS activity in the resistant MCF-Ad5 (2.4 and 2.5-fold), MCF-Ad10 (11.5- and 6.8-fold), and DLD-Ad (4.8- and 10.7-fold) lines, for binding and catalytic assays, respectively, compared with their parent MCF-7 and DLD-1 lines. The level of TS in cytosolic extracts, as determined by Western immunoblot analysis, was markedly increased for the resistant MCF-Ad5 (31-fold), MCF-Ad10 (46-fold), and DLD-Ad (52-fold) cells. Measurement of TS mRNA levels by Northern analysis revealed elevation of TS mRNA in the resistant MCF-AD5 (16.7-fold), MCF-Ad10 (31-fold), and DLD-Ad (55-fold) cells. Southern analysis showed that this increase in TS mRNA was not accompanied by any major rearrangements or amplification of the TS gene. Incorporation of 5-FU into the RNA and DNA of the resistant MCF-Ad10 cells was not significantly different, compared with that for parent MCF-7 cells. These studies suggest that exposure of human breast and human colon cancer cells to Adriamycin leads to overexpression of TS, with concomitant development of resistance to 5-FU.

Original languageEnglish (US)
Pages (from-to)136-143
Number of pages8
JournalMolecular Pharmacology
Volume39
Issue number2
Publication statusPublished - Jan 1 1991

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All Science Journal Classification (ASJC) codes

  • Molecular Medicine
  • Pharmacology

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Chu, E., Drake, J. C., Koeller, D. M., Zinn, S., Jamis-Dow, C. A., Yeh, G. C., & Allegra, C. J. (1991). Induction of thymidylate synthase associated with multidrug resistance in human breast and colon cancer cell lines. Molecular Pharmacology, 39(2), 136-143.