Colorectal cancer (CRC) is one of the most common cancers worldwide. It harbors several genetic and epigenetic alterations, which makes finding suitable biomarkers a challenge. CRC Cell lines are invaluable biomedical research tools for drug screening and biomarker discovery. Two emblematic cell lines for CRC were extensively used, HT29 and HCT116. We performed the whole transcriptome sequencing of these two cell lines and two normal epithelial cell lines, CR1780 and CRL18, derived from the colon as control. A methylation quantification of downregulated genes enables to screen potential biomarkers, which could be used as potential biomarkers for CRC diagnosis and prognosis.
Method: Total RNA was extracted from HT29, HCT116, CR1780 and CRL18 using RNeasy Mini Kit (Qiagen). mRNA libraries were generated using TruSeq Stranded mRNA Library Prep Kit (Illumina), and high-throughput sequencing were performed using Illumina systems (Hiseq and Miseq). All reads were filtered for high-quality reads (≥Q30, adapters trimming and broken pair-reads removing). Reads alignment and genes profiling analysis were done with CLCbio software. Selected genes downregulated with p-value <0.001 in cancer cell lines versus normal cell lines, were evaluated for methylation in DNA extracted from tumors and autologous normal tissues from CRC patients.
Results: The transcriptome analysis reveals 741 different splice variants are downregulated in both cancer cell lines HT29 and HCT116, when compared to the two normal cell lines, CR1780 and CRL18 (p-value <0.001). 10 relevant downregulated genes were selected, ACTG2, THY1, IGFBP3, WNT2, RAB32, SPARC, COL1A, MMP2, IGFBP3 and VIM. Interestingly, all the splice variants (10) of Vim, 7 of ATG2, 4 of THY1 and 2 of IGFBP3 were dramatically downregulated. Methylation quantification demonstrates the hypermethylation of all these 10 downregulated genes, in the tumors compared to matched normal tissues.
Conclusion. At the level of mRNA expression and methylation, this study enables to screen potential biomarkers for CRC diagnosis and prognosis.
Citation Format: Hicham Mansour, Noureddine Bouarourou, Jean Pierre Roperch, Fouazi Habib, Mourad Said. Potential biomarkers for colorectal cancer diagnosis and prognosis. [abstract]. In: Proceedings of the AACR Special Conference on Colorectal Cancer: From Initiation to Outcomes; 2016 Sep 17-20; Tampa, FL. Philadelphia (PA): AACR; Cancer Res 2017;77(3 Suppl):Abstract nr B08.
