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An Integrated Approach For Uncovering Novel DNA Methylation Biomarkers For Non-small Cell Lung Carcinoma

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Introduction - Lung cancer is one of primal and ubiquitous cause of cancer related fatalities in the world. Leading cause of these fatalities is non-small cell lung cancer (NSCLC) with a proportion of 85%. The major subtypes of NSCLC are Lung Adenocarcinoma (LUAD) and Lung Small Cell Carcinoma (LUS

C). Early-stage surgical detection and removal of tumor offers a favorable prognosis and better survival rates. However, a major portion of 75% subjects have stage III/IV at the time of diagnosis and despite advanced major developments in oncology survival rates remain poor. Carcinogens produce wide

spread DNA methylation changes within cells. These changes are characterized by globally hyper or hypo methylated regions around CpG islands, many of these changes occur early in tumorigenesis and are highly prevalent across a tumor type.Structure - This research work took advantage of publicly avai

lable methylation profiling resources and relevant comorbidities for lung cancer patients extracted from meta-analysis of scientific review and journal available at PubMed and CNKI search which were combined systematically to explore effective DNA methylation markers for NSCLC. We also tried to iden

tify common CpG loci between Caucasian, Black and Asian racial groups for identifying ubiquitous candidate genes thoroughly. Statistical analysis and GO ontology were also conducted to explore associated novel biomarkers. These novel findings could facilitate design of accurate diagnostic panel for

practical clinical relevance.Methodology - DNA methylation profiles were extracted from TCGA for 418 LUAD and 370 LUSC tissue samples from patients compared with 32 and 42 non-malignant ones respectively. Standard pipeline was conducted to discover significant differentially methylated sites as prim

ary biomarkers. Secondary biomarkers were extracted by incorporating genes associated with comorbidities from meta-analysis of research articles. Concordant candidates were utilized for NSCLC relevant biomarker candidates. Gene ontology annotations were used to calculate gene-pair distance matrix fo

r all candidate biomarkers. Clustering algorithms were utilized to categorize candidate genes into different functional groups using the gene distance matrix. There were 35 CpG loci identified by comparing TCGA training cohort with GEO testing cohort from these functional groups, and 4 gene-based pa

nel was devised after finding highly discriminatory diagnostic panel through combinatorial validation of each functional cluster.Results – To evaluate the gene panel for NSCLC, the methylation levels of SCT(Secritin), FOXD3(Forkhead Box D3), TRIM58(Tripartite Motif Containing 58) and TAC1(Tachikinin

1) were tested. Individually each gene showed significant methylation difference between LUAD and LUSC training cohort. Combined 4-gene panel AUC, sensitivity/specificity were evaluated with 0.9596, 90.43%/100% in LUAD; 0.949, 86.95%/98.21% in LUSC TCGA training cohort; 0.94, 85.92%/97.37 in GEO 66

836; 0.91,89.17%/100% in GEO 83842 smokers; 0.948, 91.67%/100% in GEO83842 non-smokers independent testing cohort. Our study validates SCT, FOXD3, TRIM58 and TAC1 based gene panel has great potential in early recognition of NSCLC undetermined lung nodules. The findings can yield universally accurate

and robust markers facilitating early diagnosis and rapid severity examination.