title: Search for epithelial-specific mRNAs in peripheral blood of patients with colon cancer by RT-PCR.
abstract: Research has widely supported the efficacy of screening for colorectal cancer in reducing mortality. A blood-based assay potentially represents a more accessible early detection tool for the identification of solid tumor cells originating from a primary tumor site in the body. We demonstrate a relatively easy and highly reproducible technique for the detection of mRNA expression of genes as markers of malignancy in blood samples of patients with colon cancer. The present study aims to identify a set of specific mRNAs expressed in epithelial cells but not in blood cells, which may be useful as markers for early detection of circulating colon cancer cells by a simple, qualitative RT-PCR assay following semi-automated RNA extraction from peripheral blood samples. Our approach includes a systematic search for candidate markers using digital differential display, search on UniGene colon EST libraries and analysis of published data on colon cancer gene expression. A final list included the following genes: bone morphogenetic protein 4 (BMP4), cyclin D (CycD), family with sequence similarity 3, member D (FAM3D), gastrin (GAS), glycoprotein A33 transmembrane (GPA33), glutathione peroxidase 2 gastrointestinal (GPX2), galactoside-binding, soluble, 4 (galectin 4) (LGALS4), non-SMC, structural maintenance of chromosomes, element 1 protein (NSE1), tumor-associated calcium signal transducer 1 (TACSTD1), telomerase reverse transcriptase (hTERT), trefoil factor 3 intestinal (TFF3), transmembrane 4 superfamily member 3 (TM4SF3), UDP glycosyltransferase 1 family, polypeptide A9 (UGT1A9), villin 1 (VIL1), and the novel gene FLJ20127. The mRNA expression of these genes was evaluated in a pool of 16 samples from subjects diagnosed with colon cancer and from 16 normal-controls. We observed expression in 13 of the 15 investigated genes from the blood samples of the vast majority of patients considered, but also in a certain percentage of the controls (from 14.3 to 100%). This finding confirms that the extreme sensitivity of RT-PCR is able to detect minimal amounts of mRNA expressed in a non tissue-specific manner ('illegitimate transcription'). On the contrary, NSE1 and GAS mRNAs were not detected either in patient or in control blood samples; however, they were abundantly expressed in normal and cancerous colon mucosa, encouraging further search for useful markers able to detect epithelial cells in peripheral blood.
authors.affiliation: Institute of Histology and General Embriology, University of Bologna, Fondazione CARISBO Centre for Research into Molecular Genetics, I-40126 Bologna, Italy.
authors.forename: Rossella
authors.initials: R
authors.lastname: Solmi
authors.forename: Paola
authors.initials: P
authors.lastname: De Sanctis
authors.forename: Cinzia
authors.initials: C
authors.lastname: Zucchini
authors.forename: Giampaolo
authors.initials: G
authors.lastname: Ugolini
authors.forename: Giancarlo
authors.initials: G
authors.lastname: Rosati
authors.forename: Marco
authors.initials: M
authors.lastname: Del Governatore
authors.forename: Domenico
authors.initials: D
authors.lastname: Coppola
authors.forename: Thymothy J
authors.initials: TJ
authors.lastname: Yeatman
authors.forename: Luca
authors.initials: L
authors.lastname: Lenzi
authors.forename: Antonello
authors.initials: A
authors.lastname: Caira
authors.forename: Simone
authors.initials: S
authors.lastname: Zanotti
authors.forename: Mario
authors.initials: M
authors.lastname: Taffurelli
authors.forename: Paolo
authors.initials: P
authors.lastname: Carinci
authors.forename: Luisa
authors.initials: L
authors.lastname: Valvassori
authors.forename: Pierluigi
authors.initials: P
authors.lastname: Strippoli
mesh_terms: D000368:Aged; D000369:Aged, 80 and over; D014408:Biomarkers, Tumor / Q000097:blood*; D003110:Colonic Neoplasms / Q000097:blood* / Q000175:diagnosis; D018076:DNA, Complementary / Q000737:chemistry; D004848:Epithelium / Q000378:metabolism; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D012333:RNA, Messenger / Q000097:blood*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*
chemical_list: D014408:Biomarkers, Tumor; D018076:DNA, Complementary; D012333:RNA, Messenger