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. 1992 Nov;66(5):840–843. doi: 10.1038/bjc.1992.370

O6-alkylguanine-DNA-alkyltransferase activity and nitrosourea sensitivity in human cancer cell lines.

M C Walker 1, J R Masters 1, G P Margison 1
PMCID: PMC1978000  PMID: 1419626

Abstract

The DNA repair enzyme, O6-alkylguanine-DNA-alkyltransferase (ATase), is thought to be the principal mechanism controlling resistance to nitrosoureas and related alkylating agents. We compared the sensitivities of five human testis and five bladder tumour cell lines to two nitrosoureas (N-nitroso-N-methylurea (MNU) and mitozolomide) with cellular levels of ATase. Enzyme levels ranged from 3 to 206 fmol mg-1 protein (0.1 x 10(4) to 5.1 x 10(4) molecules/cell) in the testis lines and from 11 to 603 fmol mg-1 (0.4 x 10(4) to 9.1 x 10(4) molecules/cell) in the bladder lines. Based on IC50s in an MTT assay, the testis tumour cell lines were, on average, four times more sensitive to MNU and six times more sensitive to mitozolomide than the bladder cell lines. The cytotoxicities of MNU and mitozolomide were closely related (R = 0.9). In the testis cell lines ATase activity (molecules/cell) was related to IC50s for mitozolomide (R = 0.97) but not MNU (R = 0.78). In the bladder cell lines and overall, ATase activity correlated with cellular sensitivity to neither agent. Relatively high levels of resistance occurred in cells expressing low levels of ATase, and amongst cell lines expressing high levels of ATase, large differences in IC50s were observed. These results support the suggestion that resistance to nitrosoureas can be mediated by mechanisms other than ATase and that at relatively high levels of expression, ATase does not confer resistance in proportion to its activity.

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Selected References

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  1. Bedford P., Fichtinger-Schepman A. M., Shellard S. A., Walker M. C., Masters J. R., Hill B. T. Differential repair of platinum-DNA adducts in human bladder and testicular tumor continuous cell lines. Cancer Res. 1988 Jun 1;48(11):3019–3024. [PubMed] [Google Scholar]
  2. Bodell W. J., Aida T., Berger M. S., Rosenblum M. L. Increased repair of O6-alkylguanine DNA adducts in glioma-derived human cells resistant to the cytotoxic and cytogenetic effects of 1,3-bis(2-chloroethyl)-1-nitrosourea. Carcinogenesis. 1986 Jun;7(6):879–883. doi: 10.1093/carcin/7.6.879. [DOI] [PubMed] [Google Scholar]
  3. Boyle J. M., Durrant L. G., Wild C. P., Saffhill R., Margison G. P. Genetic evidence for nucleotide excision repair of O6-alkylguanine in mammalian cells. J Cell Sci Suppl. 1987;6:147–160. doi: 10.1242/jcs.1984.supplement_6.10. [DOI] [PubMed] [Google Scholar]
  4. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  5. Day R. S., 3rd, Ziolkowski C. H., Scudiero D. A., Meyer S. A., Lubiniecki A. S., Girardi A. J., Galloway S. M., Bynum G. D. Defective repair of alkylated DNA by human tumour and SV40-transformed human cell strains. Nature. 1980 Dec 25;288(5792):724–727. doi: 10.1038/288724a0. [DOI] [PubMed] [Google Scholar]
  6. Erickson L. C., Bradley M. O., Ducore J. M., Ewig R. A., Kohn K. W. DNA crosslinking and cytotoxicity in normal and transformed human cells treated with antitumor nitrosoureas. Proc Natl Acad Sci U S A. 1980 Jan;77(1):467–471. doi: 10.1073/pnas.77.1.467. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Erickson L. C., Laurent G., Sharkey N. A., Kohn K. W. DNA cross-linking and monoadduct repair in nitrosourea-treated human tumour cells. Nature. 1980 Dec 25;288(5792):727–729. doi: 10.1038/288727a0. [DOI] [PubMed] [Google Scholar]
  8. Fox M., Roberts J. J. Drug resistance and DNA repair. Cancer Metastasis Rev. 1987;6(3):261–281. doi: 10.1007/BF00144267. [DOI] [PubMed] [Google Scholar]
  9. Fry A. M., Chresta C. M., Davies S. M., Walker M. C., Harris A. L., Hartley J. A., Masters J. R., Hickson I. D. Relationship between topoisomerase II level and chemosensitivity in human tumor cell lines. Cancer Res. 1991 Dec 15;51(24):6592–6595. [PubMed] [Google Scholar]
  10. Gerson S. L., Trey J. E., Miller K. Potentiation of nitrosourea cytotoxicity in human leukemic cells by inactivation of O6-alkylguanine-DNA alkyltransferase. Cancer Res. 1988 Mar 15;48(6):1521–1527. [PubMed] [Google Scholar]
  11. Gibson N. W., Hartley J. A., Barnes D., Erickson L. C. Combined effects of streptozotocin and mitozolomide against four human cell lines of the Mer+ phenotype. Cancer Res. 1986 Oct;46(10):4995–4998. [PubMed] [Google Scholar]
  12. Gibson N. W., Hickman J. A., Erickson L. C. DNA cross-linking and cytotoxicity in normal and transformed human cells treated in vitro with 8-carbamoyl-3-(2-chloroethyl)imidazo[5,1-d] -1,2,3,5-tetrazin-4(3H)-one. Cancer Res. 1984 May;44(5):1772–1775. [PubMed] [Google Scholar]
  13. Gibson N. W., Zlotogorski C., Erickson L. C. Specific DNA repair mechanisms may protect some human tumor cells from DNA interstrand crosslinking by chloroethylnitrosoureas but not from crosslinking by other anti-tumor alkylating agents. Carcinogenesis. 1985 Mar;6(3):445–450. doi: 10.1093/carcin/6.3.445. [DOI] [PubMed] [Google Scholar]
  14. Jelinek J., Kleibl K., Dexter T. M., Margison G. P. Transfection of murine multi-potent haemopoietic stem cells with an E. coli DNA alkyltransferase gene confers resistance to the toxic effects of alkylating agents. Carcinogenesis. 1988 Jan;9(1):81–87. doi: 10.1093/carcin/9.1.81. [DOI] [PubMed] [Google Scholar]
  15. Kohn K. W. Interstrand cross-linking of DNA by 1,3-bis(2-chloroethyl)-1-nitrosourea and other 1-(2-haloethyl)-1-nitrosoureas. Cancer Res. 1977 May;37(5):1450–1454. [PubMed] [Google Scholar]
  16. Laurent G., Erickson L. C., Sharkey N. A., Kohn K. W. DNA cross-linking and cytotoxicity induced by cis-diamminedichloroplatinum(II) in human normal and tumor cell lines. Cancer Res. 1981 Sep;41(9 Pt 1):3347–3351. [PubMed] [Google Scholar]
  17. Lefebvre P., Laval F. Enhancement of O6-methylguanine-DNA-methyltransferase activity induced by various treatments in mammalian cells. Cancer Res. 1986 Nov;46(11):5701–5705. [PubMed] [Google Scholar]
  18. Margison G. P., Cooper D. P., Brennand J. Cloning of the E. coli O6-methylguanine and methylphosphotriester methyltransferase gene using a functional DNA repair assay. Nucleic Acids Res. 1985 Mar 25;13(6):1939–1952. doi: 10.1093/nar/13.6.1939. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Maynard K., Parsons P. G., Cerny T., Margison G. P. Relationships among cell survival, O6-alkylguanine-DNA alkyltransferase activity, and reactivation of methylated adenovirus 5 and herpes simplex virus type 1 in human melanoma cell lines. Cancer Res. 1989 Sep 1;49(17):4813–4817. [PubMed] [Google Scholar]
  20. Morten J. E., Margison G. P. Increased O6-alkylguanine alkyltransferase activity in Chinese hamster V79 cells following selection with chloroethylating agents. Carcinogenesis. 1988 Jan;9(1):45–49. doi: 10.1093/carcin/9.1.45. [DOI] [PubMed] [Google Scholar]
  21. Parris C. N., Arlett C. F., Lehmann A. R., Green M. H., Masters J. R. Differential sensitivities to gamma radiation of human bladder and testicular tumour cell lines. Int J Radiat Biol Relat Stud Phys Chem Med. 1988 Apr;53(4):599–608. doi: 10.1080/09553008814550921. [DOI] [PubMed] [Google Scholar]
  22. Parris C. N., Walker M. C., Masters J. R., Arlett C. F. Inherent sensitivity and induced resistance to chemotherapeutic drugs and irradiation in human cancer cell lines: relationship to mutation frequencies. Cancer Res. 1990 Dec 1;50(23):7513–7518. [PubMed] [Google Scholar]
  23. Peckham M. Testicular cancer. Acta Oncol. 1988;27(4):439–453. doi: 10.3109/02841868809093570. [DOI] [PubMed] [Google Scholar]
  24. Pegg A. E. Mammalian O6-alkylguanine-DNA alkyltransferase: regulation and importance in response to alkylating carcinogenic and therapeutic agents. Cancer Res. 1990 Oct 1;50(19):6119–6129. [PubMed] [Google Scholar]
  25. Samson L., Linn S. DNA alkylation repair and the induction of cell death and sister chromatid exchange in human cells. Carcinogenesis. 1987 Feb;8(2):227–230. doi: 10.1093/carcin/8.2.227. [DOI] [PubMed] [Google Scholar]
  26. Smith D. G., Brent T. P. Response of cultured human cell lines from rhabdomyosarcoma xenografts to treatment with chloroethylnitrosoureas. Cancer Res. 1989 Feb 15;49(4):883–886. [PubMed] [Google Scholar]
  27. Stevens M. F., Hickman J. A., Stone R., Gibson N. W., Baig G. U., Lunt E., Newton C. G. Antitumor imidazotetrazines. 1. Synthesis and chemistry of 8-carbamoyl-3-(2-chloroethyl)imidazo[5,1-d]-1,2,3,5-tetrazin-4(3 H)-one , a novel broad-spectrum antitumor agent. J Med Chem. 1984 Feb;27(2):196–201. doi: 10.1021/jm00368a016. [DOI] [PubMed] [Google Scholar]
  28. Walker M. C., Parris C. N., Masters J. R. Differential sensitivities of human testicular and bladder tumor cell lines to chemotherapeutic drugs. J Natl Cancer Inst. 1987 Aug;79(2):213–216. [PubMed] [Google Scholar]
  29. Walker M. C., Povey S., Parrington J. M., Riddle P. N., Knuechel R., Masters J. R. Development and characterization of cisplatin-resistant human testicular and bladder tumour cell lines. Eur J Cancer. 1990;26(6):742–747. doi: 10.1016/0277-5379(90)90133-e. [DOI] [PubMed] [Google Scholar]

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