Influence of genetic polymorphisms on frequency of micronucleated buccal epithelial cells in leukoplakia patients Manoj B. Mahimkar a,⇑ , Tanuja A. Samant a , Sadhana Kannan b , Tejas Patil a a Cancer Research Institute, Advanced Centre for Treatment, Research and Education in Cancer (ACTREC), Tata Memorial Centre, Navi Mumbai-410210, India b ECTU, Advanced Centre for Treatment, Research and Education in Cancer (ACTREC), Tata Memorial Centre (TMC), Navi Mumbai-410210, India article info Article history: Received 26 July 2010 Received in revised form 24 August 2010 Accepted 24 August 2010 Available online 17 September 2010 Keywords: Tobacco Micronucleus assay Genetic polymorphism Buccal epithelial cells Xenobiotic metabolizing genes DNA repair genes Oral leukoplakia Micronucleated cell frequency summary Micronuclei (MN) are extensively used as an indicator of chromosomal damage and early biomarker of cancer risk. The genetic host factors are known to influence the level of chromosomal alterations conse- quently affecting MN frequencies. Hence, in the present study, we investigated the extent of chromo- somal damage by analyzing micronucleated cell (MNC) frequency in exfoliated buccal epithelial cells (BEC) and its possible relationship with genetic polymorphisms in patients with oral leukoplakia (OL). The study group comprised of habit-free (NHC, n = 39), habit controls (HC, n = 62) and OL patients (n = 66). Micronucleus assay was carried out to determine the MNC frequency and the genotyping was performed by PCR–RFLP for metabolizing (CYP1A1, GSTM1, GSTT1, GSTP1) and DNA repair (hOGG1, XRCC1, XPD) genes. The correlation between MNC frequency and genetic polymorphisms was analyzed. We found significant increase in overall MNC frequency in OL patients as compared to habit-matched controls (p = 0.01). The higher proportion of multiple micronucleated cells (>5 MN per cell) indicate increased DNA damage in the buccal mucosa of OL patients than the controls (p = 0.004). The polymor- phic alleles of XPD751 and hOGG1 showed significant association with total MNC frequency in OLs (p = 0.034 and p = 0.03 respectively). In conclusion, the extent of chromosomal damage in target tissues is higher in patients with OL. MNC frequency in combination with genetic polymorphisms in DNA repair genes may serve as better predicator of risk. Ó 2010 Elsevier Ltd. All rights reserved. Introduction Higher incidence of oral squamous cell carcinoma (OSCC) has been reported in South-East Asia and is attributed to the popularity of tobacco habits. 1 OSCC is preceded by a number of precursor stages that induce some morphological changes in cells of oral mucosa, which result in clinically detectable oral pre-malignant lesions (OPLs). The biological behavior of these lesions is rather unpredictable, as some of them may regress, whereas about one-third may progress to invasive malignancy. 2 Our ability to predict the malignant potential of OPLs based on the pathologic findings is limited. Hence, it would be of practical importance to develop biomarkers which will help in identifying individuals at high risk of malignant transformation. Accumulation of genomic damage leads to genetic instability which may be manifested as chromosomal alterations. The forma- tion of micronuclei in dividing cells is the result of chromosome breakage due to unrepaired or misrepaired DNA lesions or chromo- some mal-segregation due to spindle dysfunction. The MN assay is rapid, sensitive, cost-effective and hence extensively used as reli- able early biomarker of elevated cancer risk in healthy human sub- jects. 3,4 The study of DNA damage in exfoliated buccal epithelial cells (BEC) collected from the oral cavity is a minimally invasive method for monitoring populations exposed to genotoxic agents. BEC are the first barrier for the inhalation or ingestion route and are capable of metabolizing proximate carcinogens to reactive products. About 90% of human cancers originate from epithelial cells. Therefore, oral epithelial cells represent a preferred target site for early genotoxic events induced by carcinogenic agents entering the body. 5,6 We investigated the extent of cytogenetic damage manifested as micronuclei in buccal cells of OLs compared to the matched healthy controls. The individual response to environmental exposure to genotoxic agents is linked with lifestyle and genetic host factors. Tobacco-specific pro-carcinogens require metabolic activation to cause oxidative DNA damage which is normally corrected by effi- cient DNA repair machinery. 1 The bioactivation of carcinogens is mediated by phase-I [cytochrome p450s (CYPs)] enzymes and the reactive metabolites generated during the process are conjugated by phase-II [glutathione-S-transferases, (GSTs)] enzymes. Hence, 1368-8375/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.oraloncology.2010.08.009 ⇑ Corresponding author. Tel.: +91 22 27405049; fax: +91 22 27405085. E-mail addresses: mmahimkar@actrec.gov.in (M.B. Mahimkar), tanujasmnt@ yahoo.com (T.A. Samant), skannan@actrec.gov.in (S. Kannan), tejas_patil1986@ yahoo.co.in (T. Patil). Oral Oncology 46 (2010) 761–766 Contents lists available at ScienceDirect Oral Oncology journal homepage: www.elsevier.com/locate/oraloncology