The qRT-PCR assay and the genomic/epigenomic properties of the 10-gene Yin Yang expression ratio signature in non-small cell lung cancer
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Date
2019-08-23
Authors
Narrandes, Shavira
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Abstract
Lung cancer is the leading cause of cancer-related death in North America with a 5-year survival rate less than 20%. The ability to understand which lung cancer patients will progress and predict treatment responses will improve its management. Previously, our lab has shown a 10-gene YMR signature (GRM1, RECQL4, NRAS, and IGFBP5 are over-expressed and HOXA5, TNNC1, SOSTDC1, CRIP2, CD83, and GATA2 are under-expressed in tumor cells) correlates with prognosis and treatment prediction for non-small cell lung cancer (NSCLC). To further develop our signature for clinical use, other factors that regulate gene expression, such as changes in genomic sequences (mutations or copy number) or epigenomic factors (methylation of DNA), need to be investigated. Multiple linear regression models demonstrated that the combination of DNA methylation and copy number variation (CNV) correlate with gene expression for RECQL4, NRAS, IGFBP5, HOXA5, TNNC1, SOSTDC1, and CRIP2. Next, we evaluated six gene expression assay systems (qRT-PCR, DNA microarray, NanoString nCounter, RNA-seq, FISH, and tissue microarray) in a literature review to obtain our signature; qRT-PCR was determined to be the most feasible in a clinical setting. To validate our signature using qRT-PCR, we used an A549 cell line and lung tumor FFPE test samples obtained from the Manitoba Tumor Bank. IGFBP5 had the lowest mRNA expression level compared to TNNC1, CRIP2, and GATA2 and CRIP2 had the highest mRNA expression level in the A549 cell line, contrary to our expected signature. However, the expression levels of these genes correlated with the signature in lung tumor FFPE samples. To further confirm these results, I assessed 37 and 29 NSCLC cell lines from the Cancer Cell Line Encyclopedia (CCLE) and Genomics of Drug Sensitivity in Cancer (GDSC) databases, respectively, for the YMR signature. Only four cell lines (NCIH2066, NCIH810, NCIH292, and IA-LM) were similar to the expected signature expression pattern. These gene expression results indicate that cell lines may not be ideal for assessing the YMR signature. In the future, we will need to determine the 10-gene signature in NSCLC tumor samples in a prospective study of lung cancer patients using qRT-PCR to predict prognosis and treatment response.
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Keywords
Non-small cell lung cancer, Linear regression, Gene signature, DNA methylation, Copy number variation, qRT-PCR
Citation
Narrandes, S. & Xu, W. (2018). Gene Expression Detection Assay for Cancer Clinical Use. Journal of Cancer, 9(13), 2249–2265.