A recommended testing region in MLH1 promoter for methylation detection in colorectal cancer patients

Abstract

AbstractBackground:Lynch syndrome screening should includeMLH1promoter methylation detection in patients with deficiency of MLH1 (dMLH1) in colorectal cancer, but there is still no consensus on the CpG detection sites. The aim of this study is to analyze the correlation between the specific CpG sites ofMLH1promoter and dMLH1 in a large sample, and to find the best detection region.Methods:Immunohistochemistry (IHC) was used to detect DNA mismatch repair (MMR) protein in patients with colorectal cancer (CRC), and bisulfite sequencing PCR (BSP) was used to detectMLH1promoter methylation.MLH1promoter was divided into 5 regions: region A (-755 to -574, relative to the start codon), region B (-597 to -393), region C (-420 to -188), region D (-286 to -53) and region E (-73 to +86), which were tested respectively. The correlation between methylation in each region and MLH1 expression was analyzed. The sample size was gradually expanded to 626 cases for testing the highest correlation region. Subsequently, the correlation betweenMLH1promoter methylation and clinical parameters was analyzed, and a nomogram model for the prediction ofMLH1promoter methylation was established. Survival analysis was performed to analyze the influence ofMLH1promoter methylation on the prognosis of CRC.Results:Thepvalues of correlation between methylation in five regions( regions A, B, C, D, and E) and MLH1 protein expression were 0.070, 0.070, 0.005, 0.002 and 0.002, respectively. Regions D and E were consistent. In the cohort of 626 cases, methylation in region E was significantly correlated with the female, family history, mutantBRAFV600E, dMLH1 and deficiency of PMS2 (dPMS2). The consistency of the nomogram model we established to predictMLH1methylation and BSP was 78.6%. And the sensitivity and specificity of this nomogram model were 97.5% and 72.8%, respectively. In this article, we found thatMLH1promoter methylation had no significant effect on the prognosis of CRC.Conclusions:In colorectal cancer, the CpG sites in the region E of theMLH1promoter are recommended to determine the status ofMLH1methylation. The nomogram model ofMLH1promoter methylation is valuable and reliable for predictingMLH1methylation status.