OBJECTIVE To investigate the mechanism of hMLH1 deregulation in non-small cell lung cancer (NSCLC).METHODS A genetic and epigenetic study of the hMLH1 gene was performed using surgical primary tumors from 40 NSCLC patients and their corresponding noncancerous tissues. The molecular alterations examined included promoter methylation by Hpa Ⅱ/Msp Ⅰ- based PCR analysis, loss of heterozygosity (LOH) by D3S1621 locus PCR-electrophoresis-silver staining, as well as the loss of protein expression by immunohistochemical analysis.RESULTS The frequencies of hypermethylation, LOH and loss of protein expression of hMLH1 were 67.5% (27/40), 65% (26/40) and 72.5% (29/ 40), respectively. Among 26 hMLH1 gene LOH (+) cases, 21 (80.8%)showed hypermethylation, which was significantly higher than the groupof LOH (-). The frequency of the double inactivation of the hMLH1 gene by hypermethylation and LOH related to a loss of protein expression of 72.4% (21/29).CONCLUSION Biallelic inactivation of the hMLH1 gene by hypermethylation and LOH most likely will cause loss of hMLH1 protein expression and play an important role in the development of NSCLC. Therefore, controlling and monitoring for hypermethylation of the hMLH1 gene may be partially useful for treatment and early diagnosis of NSCLC.