Abstract 283: Biomarker search using gene expression databases in a phase III, controlled clinical trial of postoperative adjuvant chemotherapy for stage III colon cancer (acts-cc): Correlation between gene expression and DNA copy number

Abstract

Abstract Background: The ACTS-CC trial is a randomized, controlled phase III study designed to validate the noninferiority of S-1 to UFT/LV as adjuvant chemotherapy for stage III colon cancer and rectosigmoid cancer. A prospective biomarker search was performed as an auxiliary study of the ACTS-CC trial, using formalin-fixed, paraffin-embedded (FFPE) specimens obtained from 892 patients. The gene expression levels of 5-FU metabolizing enzymes and folate metabolizing enzymes and alterations of genome-wide copy numbers in tumor have been reported (2012 and 2013 Annual Meetings of AACR). In the present study, we performed correlation analyses of gene expression and DNA copy number data to characterize colorectal cancer and to prepare for a pre-planned analysis of predictive or prognostic biomarkers. Methods: FFPE specimens for this biomarker study were obtained after receiving informed consent from 892 (gene expression analysis) or 795 (DNA copy number analysis) of 1535 patients enrolled between April 2009 and January 2010. Macro-dissections were performed to extract total RNA and genomic DNA from 10-μm-thick FFPE specimens of colon cancer. Gene expression levels of 11 enzymes (TS, DPD, TP, OPRT, FPGS, GGH, DHFR, MTHFR, MTHFD, FOLRA, GART) related to 5-FU and folic acid metabolism were studied according to the Danenberg Tumor ProfileTM method (Shirota, Y. JCO 2001). Somatic copy-number alterations in cancer were analyzed by high-density SNP array analysis (Human 250K StyI array, Affymetrix, Santa Clara, CA). To detect copy-number alterations, the scanned image CEL files were imported into Partek Genomics Suite v6.6 (Partek Inc., St. Louis, MO, USA). We compared mRNA level, gene copy number and genome-wide segment copy number across a total of 161 colorectal cancers. Results: Gene expression levels were successfully estimated in 521-808 samples (84.0-90.0%, median 89.4%). Genome-wide DNA copy numbers profiles of 161 samples were obtained by GeneChip. Extracted genomic DNA from 616 of 777 samples that were not subjected to microarray analysis served as validation samples. Frequencies of copy number gain (>2.3) ranged from 0% to 57% for 11 genes, whereas frequencies of copy number loss (<1.7) ranged from 24.8% to 87%. Among 11 genes, the DNA copy numbers of GART, GGH, MTHFD1, and FPGS most strongly correlated with mRNA expression: correlation coefficients of these genes were 0.48, 0.45, 0.42, and 0.41, respectively. Gene expressions of OPRT or TP were influenced not only by their own DNA copy numbers but also by other genomic segments of 20q or 18q on genome-wide analysis. Conclusions: Gene expression of folate metabolizing enzymes positively correlated with DNA copy numbers in patients with stage III colorectal cancer. Such correlations may merit further evaluation to identify potential biomarkers. Citation Format: Hiroyuki Uetake, Toshiaki Ishikawa, Megimi Ishiguro, Shigeyuki Matsui, Kenichi Sugihara, ACTS-CC Study Group. Biomarker search using gene expression databases in a phase III, controlled clinical trial of postoperative adjuvant chemotherapy for stage III colon cancer (acts-cc): Correlation between gene expression and DNA copy number. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 283. doi:10.1158/1538-7445.AM2014-283