Loss expression of O 6 -methylguanine DNA methyltransferase by promoter hypermethylation and its relationship to betel quid chewing in oral squamous cell carcinoma Sung-Hsien Huang, MS, a Herng-Sheng Lee, MD, PhD, b Kwei Mar, DDS, c Dar-Der Ji, PhD, d Mao-Suan Huang, DDS, MS, e and Kan-Tai Hsia, DDS, PhD, f Taipei, Taiwan NATIONAL YANG-MING UNIVERSITY, NATIONAL DEFENSE MEDICAL CENTER, TAIPEI CITY HOSPITAL, CENTERS FOR DISEASE CONTROL, AND SHIN KONG WU HO-SU MEMORIAL HOSPITAL Objective. O 6 -methylguanine-DNA methyltransferase (MGMT) ameliorates mutagenic, carcinogenic, and cytotoxic adducts from O 6 -methylguanine in DNA through a direct reversal mechanism. Decreased expression of MGMT has been reported in a variety of human malignant tumors. The purpose of this study was to clarify the correlation of MGMT expression levels in oral squamous cell carcinoma (OSCC) with promoter hypermethylation and with betel quid chewing and cigarette smoking. Study design. MGMT protein expression in 63 cases of oral squamous cell carcinoma by immunohistochemistry was investigated. Methylation status of the MGMT was analyzed by methylation-specific PCR. Correlation with clinicopathologic parameters was then tested by statistical analysis. Results. MGMT immunohistochemistry revealed nuclear staining in normal epithelium, whereas 47 (75%) of 63 OSCC tumors were devoid of MGMT expression and this was related to tumor cell differentiation. Furthermore, the association between loss of MGMT expression and promoter hypermethylation was significant. Lacking protein expression for MGMT in OSCC was also associated with the use of betel quid. Conclusions. The results suggest that the absence of MGMT expression, which would seem to be associated with promoter hypermethylation, is related to betel quid chewing and, thus, in turn, might be a significant event in oral carcinogenesis. (Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2010;109:883-889) O 6 -methylguanine-DNA methyltransferase (MGMT) is a specific DNA direct reversal repair protein that re- moves mutagenic, carcinogenic, and cytotoxic adducts from O 6 -methylguanine lesions in DNA. 1,2 MGMT transfers alkyl groups at the O 6 position of guanine to an internal cysteine residue through a stoichiometric and auto-inactivating reaction. 3,4 In the absence of MGMT activity, the ability of these adducts to pair with thymine frequently gives rise to a GC¡AT transition in the K-ras oncogene and the p53 tumor suppressor gene, thereby contributing to carcinogenesis and tumor pro- gression. 5-8 MGMT knockout mice have also been found to be more sensitive than wild-type mice to tumor induction and lethality induced by alkylating agents. 9-11 Inactivation of MGMT occurs rarely owing to dele- tion, mutation, or rearrangement of the MGMT gene. 12 Previous studies have found that loss of MGMT protein expression is predominantly associated with hyper- methylation of the promoter region in a variety of primary human cancers, 13-17 suggesting that inactiva- tion of this DNA repair mechanism may be an impor- tant step in human tumorigenesis. However, loss ex- pression of MGMT by promoter hypermethylation in oral squamous cell carcinoma (OSCC) has received relatively little attention. OSCC is one of the leading cancers worldwide. 18 In Taiwan, it ranked as the sixth most prevalent cancer and the fourth leading cause of cancer deaths for men. 19 Epidemiological studies have shown that betel quid The first two authors contributed equally to this work. This study was supported by grants from the National Science Coun- cil NSC 94-2314-B-010-030, Aim for Top University plan from Department of Education, Taiwan and Department of Health, Taipei City Government 95003-62-130. a PhD student, Institute of Oral Biology, School of Dentistry, National Yang-Ming University. b Associate Professor, Department of Pathology, Tri-Service General Hospital, and National Defense Medical Center. c Doctor, Department of Dentistry, Zhongxiao Branch, Taipei City Hospital. d Assistant Professor, Research and Diagnostic Center, Centers for Disease Control, and Department of Microbiology and Immunology, National Defense Medical Center. e Doctor, Department of Dentistry, Shin Kong Wu Ho-Su Memorial Hospital. f Associate Professor, Institute of Oral Biology, School of Dentistry, National Yang-Ming University, and Department of Research and Education, Taipei City Hospital. Received for publication May 3, 2009; returned for revision Dec 9, 2009; accepted for publication Dec 9, 2009. 1079-2104/$ - see front matter © 2010 Mosby, Inc. All rights reserved. doi:10.1016/j.tripleo.2009.12.019 883