title: Immunobead multiplex RT-PCR detection of carcinoembryonic genes expressing cells in the blood of colorectal cancer patients.
abstract: Circulating cell detection using reverse transcriptase-polymerase chain reaction (RT-PCR) techniques has been studied as a new prognostic factor in colorectal cancer patients. With the view of enhancing detection sensitivity, we developed a new multiplex RT-PCR assay for circulating cell detection based on the expression of carcinoembryonic antigen-related cell adhesion molecule 5 (CEACAM5; formerly CEA) and CEACAM7 (formerly CGM2). Between November 2002 and December 2003, 45 stage III-IV, 39 stage I-II colorectal cancer patients, 32 non-colorectal cancer patients and 41 healthy individuals were included. Positive selection using HEA-125 immunobeads was applied to blood samples before mRNA extraction, cDNA synthesis and a multiplex CEACAM5/CEACAM7 RT-PCR assay. For both CEACAM5 and CEACAM7, the limit of detection was found to be as low as 1 expressing cell in 10(6) nucleated blood cells. The multiplex RT-PCR assay was negative for the 41 healthy individuals and the 32 non-colorectal cancer patients. The test was positive in 53/84 (63%) of the colorectal cancer patients for CEACAM5 and/or CEACAM7, whereas 32/84 (38%) were positive for both markers. Colorectal cancer patients were positive for one of the two markers in 80% of cases (36/45) for stage III-IV patients (CEACAM5 73%, CEACAM7 51%) and in 44% of cases (17/39) for stage I-II patients. This multiplex RT-PCR assay with two markers proved to be more sensitive than use of a single marker in detecting circulating tumour cells. The discrepant expression of CEACAM5 and CEACAM7 may label circulating tumour cells that have different levels of differentiation and subsequent aggressive behaviour.
authors.affiliation: General Surgery Department, George Pompidou AP-HP University Hospital, Paris, France.
authors.forename: Richard
authors.initials: R
authors.lastname: Douard
authors.forename: Stéphane
authors.initials: S
authors.lastname: Moutereau
authors.forename: Valérie
authors.initials: V
authors.lastname: Serru
authors.forename: Jean Patrick
authors.initials: JP
authors.lastname: Sales
authors.forename: Philippe
authors.initials: P
authors.lastname: Wind
authors.forename: Paul-Henri
authors.initials: PH
authors.lastname: Cugnenc
authors.forename: Michel
authors.initials: M
authors.lastname: Vaubourdolle
authors.forename: Sylvain
authors.initials: S
authors.lastname: Loric
mesh_terms: D000368:Aged; D001483:Base Sequence; D014408:Biomarkers, Tumor / Q000097:blood / Q000235:genetics; D002272:Carcinoembryonic Antigen / Q000097:blood* / Q000235:genetics; D002454:Cell Differentiation / Q000502:physiology; D015179:Colorectal Neoplasms / Q000097:blood*; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008875:Middle Aged; D009360:Neoplastic Cells, Circulating / Q000378:metabolism; D012333:RNA, Messenger / Q000097:blood / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D014407:Tumor Cells, Cultured
chemical_list: D014408:Biomarkers, Tumor; D002272:Carcinoembryonic Antigen; D012333:RNA, Messenger