Methods Patient’s accrual From January 2007 to December 2012, patients with a history of colorectal cancer (CRC) and age at diagnosis ≤ 50 years, who were referred to Hereditary CRC Clinic of Regina Elena National Cancer Institute, were prospectively https://www.selleckchem.com/products/px-478-2hcl.html recruited in the present study. Patients with Familial GSK3326595 Adenomatous Polyposis (FAP), Hyperplastic Polyposis, Hamartomatous Polyposis syndromes, MUTYH associated polyposis and inflammatory bowel disease were excluded from the study. For each patient an informed consent form was signed and

approved by the IFO Institutional Ethics Committee and personal medical history, detailed oncological family history were recorded and evaluated according to the Amsterdam II Criteria [35]. Immunohistochemistry for MMR proteins and microsatellite instability (MSI) analysis on tumour sampling were performed in all the patients. Tumors were considered MMR deficient if they were MSI-H and/or showed lack of MMR protein expression. Germline mutation analysis of MLH1 or MSH2 was carried out in all cases with a total lack of expression for MLH1 and no promoter hypermethylation VX-809 nmr or loss of MSH2 at immunohistochemistry, respectively. MSH6 genetic testing was done in patients whose tumor showed loss of MSH6 expression or a combined lack for MSH2

and MSH6 expression but did not have MSH2 mutations. Patients with a loss of MSH2 expression with no MSH2 or MSH6 mutations detected were analysed for EpCAM rearrangements. PMS2 genetic testing was performed in patients showing isolated loss of PMS2 expression or a combined lack of MLH1 and PMS2 expression but did not have MLH1 mutations. In patients with MSI-H tumor and normal or not available MMR protein expression, the four MMR genes were investigated in order of decreasing prevalence. Immunohistochemistry and microsatellite instability analysis Tissues (surgical sample) from colorectal adenocarcinoma patients were collected and stored in the Institute’s Tissue Bank. Patients who did not undergo surgery at our Institution www.selleck.co.jp/products/Adrucil(Fluorouracil).html were asked to apply for pathological specimens/slides at the Pathology Unit of the Hospital

in which they had surgery. The expression of MLH1, MSH2, MSH6, PMS2 genes was assessed by IHC on 2 micron thick sections of routinely formaline-fixed and paraffin-embedded blocks of selected colon adenocarcinoma tissues. Monoclonal antibodies BioCARE MEDICAL, MLH1 (clone G168-18), MSH2 (clone FE11), MSH6 (BC/44) and PMS2 (tipo clone A16-4) were used in an automated Bond immunostainer (Vision-Biosystem. Menarini, Florence, Italy). A pathologist with vast gastrointestinal experience scored the gene as expressed (positive) when nuclear staining in tumour tissue was present or, as not expressed (negative), when nuclear staining was absent. Microsatellite instability was assessed on DNA extracted from microsections of paraffin-embedded blocks of selected colon adenocarcinoma tissues.