title: Molecular and pathological characterization of the EZH2 rs3757441 single nucleotide polymorphism in colorectal cancer.
abstract: BACKGROUND
  The enhancer of zeste-homolog 2 (EZH2) is involved in cancer development through gene silencing by trimethylation of lysine 27 of histone 3 (H3K27me3). The C/C genotype for the EZH2 rs3757441 single-nucleotide polymorphism (SNP) is linked with poor prognosis in metastatic colorectal cancer (CRC), but molecular and pathological characterization of this SNP is lacking.
  
  
  METHODS
  119 primary CRCs were analyzed. SNP was evaluated by real-time PCR from colonic healthy tissue, while EZH2 and H3K27me3 expression were studied by immunohistochemistry. We primarily looked for correlation between EZH2 rs3757441 genotypes and EZH2/H3K27me3 expression. Potential associations between EZH2/H3K27me3 expression and clinico-pathological features or KRAS exon 2 and BRAF exon 15 mutations were secondary endpoints. Statistical analysis was performed by chi-square test, T-test or ANOVA.
  
  
  RESULTS
  The C/C genotype was significantly associated with higher EZH2 (100 vs. 44 %; P = 0.019) and H3K27me3 (100 vs. 38 %; P = 0.009) staining intensity compared with C/T and T/T. EZH2 3+ staining significantly correlated with stronger H3K27me3 expression (P = 0.039). KRAS and BRAF mutations were not associated with EZH2 or H3K27me3 expression.
  
  
  CONCLUSIONS
  EZH2 rs3757441 C/C genotype is associated with stronger EZH2 and H3K27me3 immunoreactivity in primary CRC: this SNP may serve as a promising biomarker for EZH2-targeting agents and may add independent information to KRAS and BRAF testing.
authors.affiliation: Unit of Medical Oncology 2, Azienda Ospedaliero-Universitaria Pisana, Istituto Toscano Tumori, Pisa, Italy. lorenzo.fornaro@gmail.com.
authors.forename: Lorenzo
authors.initials: L
authors.lastname: Fornaro
authors.affiliation: Unit of Pathology, Department of Surgical, Medical and Molecular Pathology and Critical Care, University of Pisa, Pisa, Italy. pinuccia.faviana@med.unipi.it.
authors.forename: Pinuccia
authors.initials: P
authors.lastname: Faviana
authors.affiliation: Unit of Medical Oncology 2, Azienda Ospedaliero-Universitaria Pisana, Istituto Toscano Tumori, Pisa, Italy. veronicadegregorio@libero.it.
authors.forename: Veronica
authors.initials: V
authors.lastname: De Gregorio
authors.affiliation: Unit of Medical Oncology 2, Azienda Ospedaliero-Universitaria Pisana, Istituto Toscano Tumori, Pisa, Italy. caterinavivaldi@gmail.com.
authors.forename: Caterina
authors.initials: C
authors.lastname: Vivaldi
authors.affiliation: Department of Biology, Unit of Genetics, University of Pisa, Pisa, Italy. eli.paolicchi@gmail.com.
authors.forename: Elisa
authors.initials: E
authors.lastname: Paolicchi
authors.affiliation: Unit of Medical Oncology 2, Azienda Ospedaliero-Universitaria Pisana, Istituto Toscano Tumori, Pisa, Italy. gl.masi@tin.it.
authors.forename: Gianluca
authors.initials: G
authors.lastname: Masi
authors.affiliation: Unit of Medical Oncology 2, Azienda Ospedaliero-Universitaria Pisana, Istituto Toscano Tumori, Pisa, Italy. fotiosloupakis@gmail.com.
authors.forename: Fotios
authors.initials: F
authors.lastname: Loupakis
authors.affiliation: Unit of Pathology, Department of Surgical, Medical and Molecular Pathology and Critical Care, University of Pisa, Pisa, Italy. e.sensi@med.unipi.it.
authors.forename: Elisa
authors.initials: E
authors.lastname: Sensi
authors.affiliation: Unit of Pathology, Department of Surgical, Medical and Molecular Pathology and Critical Care, University of Pisa, Pisa, Italy. cristianalupi@libero.it.
authors.forename: Cristiana
authors.initials: C
authors.lastname: Lupi
authors.affiliation: Unit of Pathology, Department of Surgical, Medical and Molecular Pathology and Critical Care, University of Pisa, Pisa, Italy. gabriella.fontanini@med.unipi.it.
authors.forename: Gabriella
authors.initials: G
authors.lastname: Fontanini
authors.affiliation: Experimental Therapeutics, BCCA Cancer Research Centre, Vancouver, BC, Canada. ywang@bccrc.ca.|Vancouver Prostate Centre, Vancouver, BC, Canada. ywang@bccrc.ca.
authors.forename: Yuzhuo
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Pharmacology Unit, Department of Clinical and Experimental Medicine, University of Pisa, Pisa, Italy. romano.danesi@med.unipi.it.
authors.forename: Romano
authors.initials: R
authors.lastname: Danesi
authors.affiliation: Unit of Medical Oncology 2, Azienda Ospedaliero-Universitaria Pisana, Istituto Toscano Tumori, Pisa, Italy. alfredo.falcone@med.unipi.it.
authors.forename: Alfredo
authors.initials: A
authors.lastname: Falcone
authors.affiliation: Experimental Therapeutics, BCCA Cancer Research Centre, Vancouver, BC, Canada. francesco.crea@open.ac.uk.|Department of Life Health and Chemical Sciences, The Open Universit, Milton Keynes, UK. francesco.crea@open.ac.uk.
authors.forename: Francesco
authors.initials: F
authors.lastname: Crea
mesh_terms: D014408:Biomarkers, Tumor / Q000096:biosynthesis / Q000235:genetics*; D015179:Colorectal Neoplasms / Q000235:genetics* / Q000473:pathology; D000071221:Enhancer of Zeste Homolog 2 Protein; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D005838:Genotype; D006657:Histones / Q000235:genetics; D006801:Humans; D008297:Male; D009154:Mutation; D009367:Neoplasm Staging; D063151:Polycomb Repressive Complex 2 / Q000096:biosynthesis / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide; D011379:Prognosis; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics; D016283:Proto-Oncogene Proteins p21(ras) / Q000235:genetics
chemical_list: D014408:Biomarkers, Tumor; D006657:Histones; C117307:KRAS protein, human; C498409:EZH2 protein, human; D000071221:Enhancer of Zeste Homolog 2 Protein; D063151:Polycomb Repressive Complex 2; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf; D016283:Proto-Oncogene Proteins p21(ras)