Selection Of Highly Stable Genes For The Transcriptomic Evaluation Of Lung Cancer.

e24318 Background: Evaluation of healthy lung tissues sections from patients with lung cancer as comparator for genomic studies could produce an unwanted bias because the healthy tissue is also exposed to carcinogens producing alterations in gene expression. Our aim in this work was to identify highly stable genes in healthy lung tissues as reference genes with normalization purposes to evaluate lung-tumor transcriptomes. Methods: We evaluated all transcripts evaluated with RNA-Seq (RPKM) of non-tumoral lung tissues (n = 320) from the Genotype-Tissue Expression (GTEx) project. To select the candidate reference genes, we first choose genes with the most stable expression defined for this study as genes with coefficient of variation < 15%, with patterns of normal distribution, low frequencies of alterations ( < 10%) in the TCGA dataset for squamous cell lung cancer and whose expression is not associated with the clinical outcome (assessed in a meta-analysis in KM-plotter for lung cancer). Results: Eleven genes (GNB1, THRAP3, PEX19, TGOLN2, ARIH2, UBE2D2, TMED4, SART3, SNX1, VPS4A and RANBP3) had lowest coefficient of variation, of which PAX19 was excluded because it had a high frequency of alterations (11%). In the meta-analysis in KM-plotter, four 4 genes (GNB1, VPS4A, SART3 and RANBP3) were related with the outcome. Finally, SART3, RANBP3, GNB1 and VPS4A were selected as reference genes. The geometric mean (GM) of the four genes had a low coefficient of variation (9.2%) and was suitable for normalization when compared with the TCGA dataset of squamous cell lung cancer (Table 1). Conclusions: We suggest four genes with normalization purposes to improve the direct comparison between healthy lung tissues with lung tumors. Table 1. Candidate references genes in lung tissue Name Mean SD CV Standard error of the mean (SEM) Normality test SART3 9.85 1.30 13.17 0.073 0.914 RANBP3 14.26 2.08 14.56 0.116 0.741 GNB1 92.38 12.86 13.92 0.719 0.836 VPS4A 37.61 5.53 14.70 0.309 0.666 GM 26.3 2.4 9.16 0.134