Study On The Potential Of Epigenetic Biomarkers For Early Colorectal Cancer Detection

In recent years,with the improvement of living standards,colorectal cancer(CRC)has become one of the leading causes of cancer-associated death worldwide.Detection of CRC at an early curable stage and removal of precancerous lesions can reduce the mortality.At present,the conventional methods,fecal occult blood test(FOBT)and colonoscopy are used most frequently to screen for CRC.However,the sensitivity and specificity of FOBT are still not optimal.Colonoscopy is invasive,costly and low adherence rates,although it has highly positive predictive value.Therefore,it is imperative to develop molecular biomarkers as a noninvasive screening method for the diagnosis of early CRC.Development of colorectal cancer is a complex process with accompanying genetic and epigenetic changes and accumulation,and epigenetic changes of the related gene is an early event in the process of colorectal cancer.Epigenetics refers to the study of mechanisms that alter gene expression without permanently altering the DNA sequence.Epigenetic alterations are reversible and heritable,and include changes in DNA methylation,and non-coding RNA-mediated gene silencing.At present,DNA methylation and non-coding RNA including microRNAs(miRNAs)represent the largest body of available literature on epigenetic biomarkers with the highest potential for cancer diagnosis.The recent breakthrough research findings is that stool intestinal mucosal cells and the tumor tissues have the similar genetic change such as DNA methylation,the dysregulation of microRNAs has been linked to tumor progression.Epigenetic for early detection and prognosis judgment of colorectal cancer is of great significance.In view of this,this study focus on the potential of stool DNA methylation and serum microRNA as candidate biomarkers for early colorectal cancer detection.Part one:Identification the potential of stool-based DNA methylation as a candidate biomarker for colorectal neoplasia early detectionBackground and ObjectiveColorectal cancer has become one of the leading causes of cancer-associated death worldwide.Recent advances in stool-based DNA methylation as noninvasive biomarkers have provided promising prospect in detecting colorectal cancer.This study aimed to investigate the potential of stool-based DNA methylation for colorectal cancer detection,especially in the curable stage.Materials and MethodsMethylation status of eight candidate biomarkers(SNCA,SPG20,CNRIP1,FBN1,MAL,INA,OSMR,TFPI2)selected from the literature was identified in adenoma(n=10),colorectal tumor(n=10)and their adjacent normal tissues(n=10)using bisulfite sequencing PCR(BSP).The most hypermethylated genes were then validated in 144 stool samples including adenoma groups(n=49),CRC groups(n=31)and normal controls(n=64).The stool samples were processed by the stool intestinal mucosal cells collector and analyzed by quantitative methylation-specific polymerase chain reaction(qMSP).Promoter methylation status of genes was denoted as percentage of methylated reference(PMR)values.Receiver operator characteristic(ROC)curve analysis and multivariable logistic analysis were used to determine the detecting performance of the biomarker.ResultsSNCA,SPG20 and FBN1 were detected frequently hypermethylated in the CRC and adenoma tissue samples compared to their adjacent normal tissues.In stool samples,The SNCA PMR values were significantly higher in CRC and adenoma groups than in the normal controls.The SPG20 and FBN1 methylation status was not significantly different among groups.Stool-based SNCA methylation yield an AUC of 0.836 with 83.9%sensitivity and 75%specificity in discriminating CRC from normal controls,and yield an AUC of 0.772 with 75.5%sensitivity and 75%specificity in discriminating adenoma from normal controls.Multivariable logistic regression analyses showed that the odds ratio for CRC and adenoma patients with high stool-based SNCA methylation was 11.291 and 9.234,respectively,after adjustment for patients' age and sex.SNCA promoter methylation increased with disease progression.No significant difference in stool-based SNCA methylation was found between proximal and distal lesions.SNCA methyaltion significantly reduced after tumor resection compared to the initial status.ConclusionThe results validated the capability of SNCA methylation in stool samples for the identification of patients with colorectal cancer and adenoma.Part two:Study on serum miR-20a and miR-486 as potential biomarkers for discriminating colorectal neoplasiaObjectiveThis study aimed to identify the potential of serum miRNAs(miR-20a,miR-486,miR-92a,miR-135b)selected from the literature for the identification of patients with colorectal neoplasia.Materials and MethodsThe serum samples from 46 colorectal neoplasia patients and 33 normal controls were analyzed the potential of four serum miRNAs by quantitative RT-PCR.Receiver operator characteristic(ROC)curve analysis was used to determine the discriminating performance of the biomarkers.ResultsSerum levels of miR-20a and miR-486 were significantly decreased in colorectal neoplasia patients than in healthy controls.Serum miR-92a expression was examined no differential expression between two groups,while miR-135b level in serum was too low to be accurately quantified.Serum miR-486 level was lower in tubulovillous adenoma and HGINs patients than in healthy controls.The AUC values for miR-20a and miR-486,and their combined value in discriminating colorectal neoplasia patients were 0.676,0.629 and 0.698,respectively.There were no significant correlations between serum miR-20a and miR-486 levels and age,gender,location or lesion size.ConclusionThe results validated the capability of serum miR-20a and miR-486 to detect colorectal neoplasia which provide rationale for further development of the epigenetic biomarker in noninvasive early detection of CRC.