Abstract
Aberrant cytosine 5-methylation underlies many deregulated elements of cancer. Among paired non-small cell lung cancers (NSCLC), we sought to profile DNA 5-methyl-cytosine features which may underlie genome-wide deregulation. In one of the more dense interrogations of the methylome, we sampled 1.2 million CpG sites from twenty-four NSCLC tumor (T)–non-tumor (NT) pairs using a methylation-sensitive restriction enzyme- based HELP-microarray assay. We found 225,350 differentially methylated (DM) sites in adenocarcinomas versus adjacent non-tumor tissue that vary in frequency across genomic compartment, particularly notable in gene bodies (GB; p<2.2E-16). Further, when DM was coupled to differential transcriptome (DE) in the same samples, 37,056 differential loci in adenocarcinoma emerged. Approximately 90% of the DM-DE relationships were non-canonical; for example, promoter DM associated with DE in the same direction. Of the canonical changes noted, promoter (PR) DM loci with reciprocal changes in expression in adenocarcinomas included HBEGF, AGER, PTPRM, DPT, CST1, MELK; DM GB loci with concordant changes in expression included FOXM1, FERMT1, SLC7A5, and FAP genes. IPA analyses showed adenocarcinoma-specific promoter DMxDE overlay identified familiar lung cancer nodes [tP53, Akt] as well as less familiar nodes [HBEGF, NQO1, GRK5, VWF, HPGD, CDH5, CTNNAL1, PTPN13, DACH1, SMAD6, LAMA3, AR]. The unique findings from this study include the discovery of numerous candidate The unique findings from this study include the discovery of numerous candidate methylation sites in both PR and GB regions not previously identified in NSCLC, and many non-canonical relationships to gene expression. These DNA methylation features could potentially be developed as risk or diagnostic biomarkers, or as candidate targets for newer methylation locus-targeted preventive or therapeutic agents.
Highlights
Lung cancer is responsible for the highest number of cancer-related deaths in the United States [1]
Among 24 non-small cell lung cancers (NSCLC) lung resection donors (S1 Table), using the HELP-microarray assay we identified 452,754 HpaII fragments significantly differentially methylated (DM; p
Of these DM sites, 57% were found in coding regions. (Fig 1) Another 39% of these were found in intergenic regions (48% of those sites represented on the array) and were mostly hypomethylated in tumors
Summary
Lung cancer is responsible for the highest number of cancer-related deaths in the United States [1]. Investigating the same specimens for differential gene expression would allow identification of higher impact DM loci, by virtue of potential impact on expression To test these hypotheses, we used the HELP assay [26] to assay the CpG methylation of 24 pairs of tumor (T) and adjacent non-tumor (NT) human samples. We used the HELP assay [26] to assay the CpG methylation of 24 pairs of tumor (T) and adjacent non-tumor (NT) human samples This assay queries 1.2 million CCGG motif-defined fragments across the genome by restriction enzymes HpaII (methylation sensitive) and MspI (methylation resistant) to isolate differentially methylated fragments of the genome. Reasoning that methylation-deregulated genes might be more apparent if cognate/proximate gene expression is altered, we further examined the association of differentially methylated (DM) regions with differentially expressed (DE) genes, using mRNA expression data from the same paired T and NT surgical resection samples
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
