Clinical Cancer Research, Vol 2, Issue 3 577-584, Copyright © 1996 by American Association for Cancer Research

ARTICLES

Quantitative immunohistochemical estimates of O6-alkylguanine-DNA alkyltransferase expression in normal and malignant human colon

NH Zaidi, L Liu and SL Gerson
Division of Hematology, Department of Medicine, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106-4937, USA.

A major mechanism of resistance to nitrosoureas is O6-alkylguanine-DNA-alkyltransferase. The alkyltransferase biochemical assay measures mean tissue activity but requires availability of fresh tissue and cannot assess tumor heterogeneity, an important component of tumor resistance to alkylating agents. We assessed the levels of alkyltransferase in human colon carcinoma and normal colon by biochemical assay, Western blot, conventional immunohistochemistry, and quantitative immunohistochemistry (using 5H7 and mT3.1 monoclonal IgGs) to correlate whole tissue levels with cell-specific expression. Alkyltransferase activity was 18.0 +/- 4.6 fmol/microgram DNA in normal colon and 15.0 +/- 6.5 fmol/microgram DNA in tumors. By Western blot estimates, alkyltransferase in normal colon was 14.8 +/- 4.2 fmol/microgram DNA and in tumors was 16.2 +/- 7.8 fmol/microgram DNA. Alkyltransferase estimates by biochemical and Western blots were correlated strongly (P < 0.0001). Conventional immunohistochemistry demonstrated that alkyltransferase was predominantly nuclear and in normal colon was concentrated in glandular epithelial mucosal cells close to the lumen, whereas in tumors, expression was heterogenous but localized to malignant epithelial cells. Two parameters of quantitative immunohistochemistry, integrated gray and mean gray, were correlated strongly with each other (P < 0.002) and with biochemical and Western blot estimates (P = 0.004-0.04). Thus, quantitative immunohistochemical estimates of alkyltransferase in fixed tissues are a reasonable alternative to biochemical analysis and have an added advantage of identifying heterogeneity of alkyltransferase expression in tumors.

This article has been cited by other articles:

				M. E. Hegi, L. Liu, J. G. Herman, R. Stupp, W. Wick, M. Weller, M. P. Mehta, and M. R. Gilbert Correlation of O6-Methylguanine Methyltransferase (MGMT) Promoter Methylation With Clinical Outcomes in Glioblastoma and Clinical Strategies to Modulate MGMT Activity J. Clin. Oncol., September 1, 2008; 26(25): 4189 - 4199. [Abstract] [Full Text] [PDF]

				D. Fontijn, A. D. Adema, K. K. Bhakat, H. M. Pinedo, G. J. Peters, and E. Boven O6-Methylguanine-DNA-methyltransferase promoter demethylation is involved in basic fibroblast growth factor induced resistance against temozolomide in human melanoma cells Mol. Cancer Ther., October 1, 2007; 6(10): 2807 - 2815. [Abstract] [Full Text] [PDF]

				L. Zhang, W. Lu, X. Miao, D. Xing, W. Tan, and D. Lin Inactivation of DNA repair gene O6-methylguanine-DNA methyltransferase by promoter hypermethylation and its relation to p53 mutations in esophageal squamous cell carcinoma Carcinogenesis, June 1, 2003; 24(6): 1039 - 1044. [Abstract] [Full Text] [PDF]

				N. Kohya, K. Miyazaki, S. Matsukura, H. Yakushiji, Y. Kitajima, K. Kitahara, M. Fukuhara, Y. Nakabeppu, and M. Sekiguchi Deficient Expression of O6-Methylguanine-DNA Methyltransferase Combined With Mismatch-Repair Proteins hMLH1 and hMSH2 Is Related to Poor Prognosis in Human Biliary Tract Carcinoma Ann. Surg. Oncol., May 1, 2002; 9(4): 371 - 379. [Abstract] [Full Text] [PDF]

				S. L. Gerson Clinical Relevance of MGMT in the Treatment of Cancer J. Clin. Oncol., May 1, 2002; 20(9): 2388 - 2399. [Abstract] [Full Text] [PDF]

				S. Matsukura, K. Miyazaki, H. Yakushiji, A. Ogawa, K. Harimaya, Y. Nakabeppu, and M. Sekiguchi Expression and Prognostic Significance ofO6-Methylguanine-DNA Methyltransferase in Hepatocellular, Gastric, and Breast Cancers Ann. Surg. Oncol., December 1, 2001; 8(10): 807 - 816. [Abstract] [Full Text] [PDF]

				M. Y. Hong, J. R. Lupton, J. S. Morris, N. Wang, R. J. Carroll, L. A. Davidson, R. H. Elder, and R. S. Chapkin Dietary Fish Oil Reduces O6-Methylguanine DNA Adduct Levels in Rat Colon in Part by Increasing Apoptosis during Tumor Initiation Cancer Epidemiol. Biomarkers Prev., August 1, 2000; 9(8): 819 - 826. [Abstract] [Full Text]

				P. J. O'Connor, F. C.R. Manning, A. T. Gordon, M. A. Billett, D. P. Cooper, R. H. Elder, and G. P. Margison Thresholds for Genotoxic Carcinogens: DNA Repair: Kinetics and Thresholds Toxicol Pathol, May 1, 2000; 28(3): 375 - 381. [Abstract] [PDF]

				M. Y. Hong, R. S. Chapkin, C. P. Wild, J. S. Morris, N. Wang, R. J. Carroll, N. D. Turner, and J. R. Lupton Relationship between DNA Adduct Levels, Repair Enzyme, and Apoptosis as a Function of DNA Methylation by Azoxymethane Cell Growth Differ., November 1, 1999; 10(11): 749 - 758. [Abstract] [Full Text]

				M. Esteller, S. R. Hamilton, P. C. Burger, S. B. Baylin, and J. G. Herman Inactivation of the DNA Repair Gene O6-Methylguanine-DNA Methyltransferase by Promoter Hypermethylation is a Common Event in Primary Human Neoplasia Cancer Res., February 1, 1999; 59(4): 793 - 797. [Abstract] [Full Text] [PDF]

				L. Liu, X. Qin, and S. L. Gerson Reduced lung tumorigenesis in human methylguanine DNA—methyltransferase transgenic mice achieved by expression of transgene within the target cell Carcinogenesis, February 1, 1999; 20(2): 279 - 284. [Abstract] [Full Text] [PDF]