Datasets:

Brizape

/

Variome_split_0404_dev

like 0

[]

[]

[ "Age,", "age", "at", "diagnosis;", "DL,", "duodenal", "mutation", " ", "in", "APC", "or", "MUTYH" ]

[ 0, 0, 0, 0, 0, 3, 5, 0, 0, 0, 0, 0 ]

[ "MSI", "was", "detected", "in", "three", "of", "the", "eight", "cell", "lines", "i.e.", "A2780", "(no", "MLH1", "mRNA", "expression", "due", "to", "promoter", "methylation),", "SKOV3", "(no", "MLH1", " ", "mRNA", "expression)", "and", "2774", "(no", "altered", "expression", "of", "MMR", "genes).", "Overall,", "there", "was", "no", "association", "between", "cisplatin", "response", "and", "MMR", "status", "in", "these", "eight", "cell", "lines." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 21, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "The", "mutation", "between", "the", "MSI-L", "and", "MSI-H" ]

[ 0, 0, 0, 0, 0, 0, 0 ]

[ "Genetic", "instability", "is", "seen", "in", "most", "types", "of", "cancer", "[10].", "Two", "distinct", "types", "of", "genetic", "instability", "appear", "to", "occur", "in", "colorectal", "cancer", "[11]:", "chromosomal", "and", "microsatellite", "instability.", "Chromosomal", "instability", "results", "in", "gains", "or", "losses", "of", "entire", "chromosomes", "or", "parts", "of", "them,", "and", "gives", "rise", "to", "aneuploid", "tumours", "and", "occurs", "in", "the", "majority", "of", "cancers.", "A", "smaller", "proportion", "of", "colorectal", "cancers", "displays", "microsatellite", "instability,", "represented", "by", "diploid", "cells", "acquiring", "high", "mutation", "rates,", "and", "was", "found", "to", "be", "associated", "with", "defective", "mismatch", "repair", "[12].", "These", "tumours", "are", "less", "likely", "to", "harbour", "mutations", "in", "genes", "associated", "with", "chromosomally", "instable", "and", "generally", "aneuploid", "tumours,", "such", "as", "APC,", "K-ras", "and", "TP53", "[13-21],", "suggesting", "that", "these", "tumours", "form", "a", "distinct", "group.", "Moreover,", "microsatellite", "instable", "tumours", "are", "found", "predominantly", "in", "the", "proximal", "colon", "[22,23],", "are", "more", "likely", "to", "occur", "in", "patients", "with", "a", "positive", "family", "history", "of", "colorectal", "cancer", "[22,23],", "are", "often", "less", "differentiated", "than", "microsatellite", "stable", "tumours", "colorectal", "colorectal", "cancer", " ", "has", "been", "proposed,", "which", "describes", "the", "sequential", "accumulation", "of", "specific", "genetic", "alterations", "in", "various", "pathways,", "involving", "tumour", "suppressor", "genes", "(e.g.", "APC,", "SMAD4,", "TP53)", "and", "oncogenes", "(e.g.", "CTNNB1,", "K-ras)", "[3,4].", "Important", "molecular", "pathways", "that", "upon", "activation", "affect", "the", "early", "and", "intermediate", "stages", "of", "colorectal", "carcinogenesis", "are", "the", "Wnt", "and", "Ras", "signalling", "pathways,", "whereas", "TP53", "inactivation", "is", "considered", "a", "late", "event." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 3, 1, 2, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Amplification", "of", "exon", "3", "of", "the", "CTNNB1", "gene", "entailed", "a", "semi-nested", "PCR", "strategy,", "which", "covered", "codons", "33,", "37,", "41", "and", "45.", "Flank", "PCR", "was", "performed", "to", "generate", "a", "308", "bp", "fragment", "(primers,", "forward:", "5'-CCAATCTACTAATGCTAATACTG-3',", "reverse:", "5'-GCATTCTGACTTTCAGTAAGGC-3')", "that", "was", "used", "in", "a", "1:100", "dilution", "for", "amplification", "of", "the", "final", "PCR", "product", "(primers,", "forward:", "5'-CCAATCTACTAATGCTAATACTG-3',", "reverse:", "5'-CTTCCTCAGGATTGCCTTTACC-3').", "In", "each", "PCR,", "one", "round", "of", "35", "cycles", "was", "performed." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "In", "tumors", "different", "genetic", "mechanisms", "are", "known", "to", "affect", "gene", "expression", "in", "wider", "chromosomal", "regions.", "Chromosomal", "aberrations,", "like", "homozygous", "and", "heterozygous", "deletions", "or", "amplifications,", "alter", "the", "DNA", "copy", "number", "of", "large", "genomic", "regions", "or", "even", "whole", "chromosome", "arms,", "leading", "to", "inactivation", "of", "tumor", "suppressor", "genes", "[7,8]", "or", "to", "activation", "of", "oncogenes.", "Another", "genetic", "phenomenon", "that", "is", "assumed", "to", "have", "drastic", "effects", "on", "gene", "expression", "in", "cancer", "cells", "is", "the", "aberrant", "alteration", "of", "chromatin", "structure.", "Methylation", "of", "genomic", "DNA,", "histone", "acetylation,", "and", "histone", "methylation", "are", "assumed", "to", "have", "a", "large", "impact", "on", "the", "accessibility", "of", "DNA", "for", "transcription", "initiation", "[9].", "Such", "epigenetic", "mechanisms", "can", "affect", "large", "genomic", "regions", "by", "possibly", "either", "silencing", "or", "activating", "large", "arrays", "of", "genes.", "However,", "the", "regulatory", "mechanisms", "governing", "chromatin", "assembly", "and", "disassembly", "are", "only", "beginning", "to", "emerge.", "So", "far,", "due", "to", "methodological", "limitations", "it", "has", "not", "been", "possible", "to", "study", "the", "role", "of", "such", "phenomena", "for", "gene", "expression", "in", "cancer", "cells", "on", "a", "genome-wide", "scale.", "Nevertheless,", "evidence", "from", "single-gene", "focused", "studies", "suggests", "that", "chromatin", "regulation", "does", "play", "an", "important", "role", "in", "tumorigenesis", "[10,11]." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Families", "carrying", "the", "p.Lys618Ala", "variant" ]

[ 0, 0, 0, 0, 0 ]

[ "Of", "656", "tumours", "for", "which", "the", "other", "molecular", "alterations,", "i.e.", "mutations", "in", "the", "APC", "and", "K-ras", "genes", "and", "hMLH1", "expression,", "were", "all", "successfully", "and", "completely", "analysed,", "103", "colorectal", "tumours", "tumours", "Tumours", " ", "from", "464", "of", "656", "patients,", "which", "did", "not", "harbour", "a", "truncating", "APC", "mutation", "or", "lacked", "hMLH1", "expression,", "were", "analysed", "for", "mutations", "in", "exon", "3", "of", "the", "CTNNB1", "gene.", "Table", "1", "describes", "the", "tumour", "and", "patient", "characteristics", "of", "seven", "colorectal", "tumours", "tumour", " ", "and", "patient", "characteristics", "of", "seven", "colorectal", "tumours", "that", "harboured", "a", "mutation", "in", "CTNNB1", "exon", "3.", "In", "five", "colorectal", "cancers,", "a", "CTNNB1", "mutation", "that", "would", "lead", "to", "loss", "of", "one", "of", "the", "Ser/Thr", "phosphorylation", "sites", "and", "subsequent", "stabilisation", "of", "the", "protein,", "occurred", "at", "codons", "37", "and", "45,", "all", "were", "C→T", "transitions,", "leading", "to", "Ser→Phe", "amino", "acid", "changes", "and", "occurred", "in", "the", "proximal", "colon.", "All", "bar", "one", "also", "had", "an", "activating", "mutation", "in", "the", "K-ras", "gene.", "Three", "of", "these", "five", "tumours", "showed", "hMLH1", "deficiency.", "Two", "colorectal", "cancer", "patients", "harboured", "a", "mutation", "in", "the", "CTNNB1", "gene,", "that", "did", "not", "occur", "at", "the", "Ser/Thr", "phosphorylation", "sites,", "but", "would", "result", "in", "an", "amino", "acid", "alteration", "at", "codons", "22", "and", "29,", "the", "effects", "of", "which", "are", "unknown.", "Because", "of", "the", "very", "low", "frequency", "of", "tumours", "harbouring", "a", "CTNNB1", "mutation,", "these", "mutations", "were", "not", "included", "in", "further", "analyses.", "In", "addition,", "mutation", "analysis", "of", "remaining", "samples", "was", "abandoned,", "since", "this", "was", "deemed", "irrelevant", "as", "these", "harboured", "truncating", "APC", "mutations", "and", "are", "considered", "to", "be", "unlikely", "to", "also", "have", "CTNNB1", "mutations", "[7]." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 1, 1, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 1, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "a", "Data", "was", "unavailable", "for", "MSI", "status", "in", "40", "cases,", "methylation", "status", "in", "83", "cases,", "KRAS", "mutation", "in", "26", "cases", "and", "TP53", "colorectal", "cancer", "." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 21, 1, 2, 0 ]

[ "Not", "unexpected,", "we", "found", "loss", "of", "expression", "in", "region", "5q22.2-5q23.1", "(see", "Figures", "27,", "28,", "29).", "This", "interval", "harbors", "two", "known", "TSGs", "in", "colon", "cancer,", "the", "adenomatous", "polyposis", "coli", "gene", "(APC)", "gene", "and", "the", "mutated", "in", "colorectal", "cancer", "(MCC).", "We", "were", "not", "able", "to", "obtain", "expression", "values", "for", "APC.", "APC", "is", "located", "at", "the", "border", "of", "a", "region", "at", "5q22.2-5q22.3", "that", "harbors", "several", "drastically", "down-regulated", "genes.", "Central", "in", "this", "region", "is", "the", "MCC", "gene.", "The", "distal", "border", "is", "the", "CDO1", "gene.", "We", "assume", "that", "deletion", "or", "epigenetic", "silencing", "of", "this", "region", "is", "a", "frequent", "mechanism", "contributing", "to", "colorectal", "tumorigenesis.", "It", "is", "possible", "that", "also", "APC", "or", "MCC", "show", "reduced", "expression,", "that", "genes", "in", "this", "region", "other", "than", "APC", "and", "MCC", "are", "piggy-back", "genes,", "and", "that", "their", "misregulation", "is", "not", "of", "functional", "significance", "for", "tumorigenesis." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "The", "region", "20q11.22-q11.23", "was", "among", "the", "most", "frequently", "up-regulated", "regions", "(see", "Figures", "12,", "13,", "14).", "Amplifications", "of", "regions", "on", "chromosome", "20q", "have", "been", "identified", "independently", "by", "several", "groups", "in", "CRCs", "[19,21,23,24].", "The", "interval", "comprises", "the", "known", "tumor", "gene", "SRC", "(located", "between", "MANBAL", "and", "BLCAP", "in", "Figures", "12,", "13,", "14)", "for", "which", "no", "informative", "expression", "measures", "were", "obtained.", "We", "note", "that", "it", "is", "possible", "that", "the", "SRC", "gene", "is", "the", "primary", "target", "of", "up-regulation", "in", "our", "CRC", "patients", "tumor", "tumor", "-versus-normal", "expression.", "Genes", "are", "given", "in", "chromosomal", "order", "on", "the", "horizontal", "axis.", "Patient", "codes", "are", "given", "on", "the", "vertical", "axis.", "The", "legend", "depicts", "which", "colors", "code", "for", "which", "expression", "changes", "on", "a", "loge", "scale", "(green:", "down", "in", "tumor;", "red:", "up", "in", "tumor).", "View", "in", "conjunction", "with", "Figures", "7", "and", "8." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 9, 1, 1, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "The", "fraction", "of", "large", "deletions", "high", "age", "at", "diagnosis", ",", "and", "a", "low", "frequency", "of", "extracolonic", "manifestations.", "Whether", "these", "patients", "really", "are", "affected", "by", "APC-associated", "FAP", "can", "(of", "course)", "be", "called", "into", "question.", "However,", "among", "attenuated", "cases", "of", "FAP", "in", "these", "study", "we", "have", "found", "very", "subtle", "mutations", "such", "as", "the", "mosaic", "case", "as", "well", "as", "the", "c.70C", ">", "T", "mutation", "and", "splice-site", "mutations.", "Considering", "these", "facts,", "at", "least", "some", "of", "the", "cases", "could", "be", "caused", "by", "mutations", "in", "APC", "resulting", "in", "only", "partially", "inactivation", "of", "the", "gene", "function.", "Since", "the", "main", "purpose", "of", "this", "study", "was", "to", "achieve", "as", "high", "mutation-detection", "rate", "as", "possible", "in", "families", "with", "colorectal", "polyposis", "syndromes,", "using", "a", "range", "of", "different", "molecular", "genetic", "techniques,", "we", "have", "not", "yet", "performed", "any", "further", "analyses", "of", "the", "relatively", "few", "mutation", "negative", "cases", "to", "determine", "if", "they", "belong", "to", "non-polyposis", "CRC", "syndromes." ]

[ 0, 0, 0, 5, 6, 15, 16, 16, 16, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Down-regulation", "of", "mRNA", "expression", "in", "human", "chromosomal", "region", "18q21.2-18q23", "–", "the", "BCL2", "region", "(patient", "counts", "with", "coordinate", "down-regulation).", "Grayscale", "cross-comparison", "plot", "of", "down-regulation", "patterns", "across", "patients", "(analogous", "to", "Figures", "8,", "11,", "14).", "View", "this", "plot", "in", "conjunction", "with", "Figures", "24", "and", "25.", "Note,", "that", "many", "more", "patients", "show", "down-regulation", "as", "indicated", "by", "dark", "spots", "in", "this", "plot", "than", "up-regulation", "as", "indicated", "by", "dark", "spots", "in", "Figure", "25.", "This", "region", "has", "been", "reported", "in", "other", "studies", "to", "be", "frequently", "deleted", "in", "colon", "cancer", "(see", "Table", "4).", "Note", "the", "expression", "down-regulation", "of", "BCL2.", "SMAD4", "(Hs.298320)", "and", "TCF4", "are", "only", "weakly", "down-regulated.", "The", "DCC", "gene", "is", "also", "located", "in", "this", "region", "between", "LOC51320", "and", "MBD2", "but", "no", "informative", "expression", "measures", "were", "obtained." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "4p15.31-4p15.2" ]

[ 0 ]

[ "Methods" ]

[ 0 ]

[ "Amplification", "of", "exon", "3", "of", "the", "CTNNB1", "gene", "entailed", "a", "semi-nested", "PCR", "strategy,", "which", "covered", "codons", "33,", "37,", "41", "and", "45.", "Flank", "PCR", "was", "performed", "to", "generate", "a", "308", "bp", "fragment", "(primers,", "forward:", "5'-CCAATCTACTAATGCTAATACTG-3',", "reverse:", "5'-GCATTCTGACTTTCAGTAAGGC-3')", "that", "was", "used", "in", "a", "1:100", "dilution", "for", "amplification", "of", "the", "final", "PCR", "product", "(primers,", "forward:", "5'-CCAATCTACTAATGCTAATACTG-3',", "reverse:", "5'-CTTCCTCAGGATTGCCTTTACC-3').", "In", "each", "PCR,", "one", "round", "of", "35", "cycles", "was", "performed." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Conclusions" ]

[ 0 ]

[ "An", "epigenetic", "mechanism", "may", "help", "explain", "why", "sporadic", "MSI+", "cancers", "colorectal", "germline", "mutations", " ", "(APC", "and", "MMR", "loci)", "in", "both", "common", "colorectal", "familial", "cancer", "familial", " ", "cancer", "syndromes", "(", "FAP", " ", "and", "HNPCC)", "appear", "to", "advance", "progression", "by", "more", "than", "a", "single", "mutation", "relative", "to", "their", "sporadic", "counterparts." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 3, 5, 6, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 17, 0, 0, 0, 0, 1, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "As", "screening", "method", "for", "HNPCC", "we", "searched", "for", "MSI", "in", "the", "proband's", "primary", "tumor", "and", "metastasis", "tissue", "blocks.", "The", "absence", "of", "proband's", "non-tumor", "DNA", "for", "MSI", "testing", "was", "overcome", "studying", "the", "alleles", "carried", "by", "his", "progenitors.", "Once", "the", "MSI", "was", "established,", "a", "second", "blood", "sample", "was", "obtained", "from", "the", "proband's", "mother", "(obligate", "carrier", "but", "at", "the", "moment", "of", "the", "study", "still", "unaffected", "poorly", "differentiated", " ", "colorectal", "adenocarcinoma", "at", "age", "23", "contacted", "us", "for", "genetic", "counseling.", "A", "detailed", "family", "and", "medical", "history", "was", "obtained", "through", "interview", "with", "the", "proband", "relatives", "and", "their", "consent", "for", "release", "of", "medical", "records", "and", "use", "of", "the", "pathological", "tissue", "blocks", "still", "available." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 7, 8, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Details", "of", "the", "8", "patients", "in", "MSS", "group", "identified", "to", "have", "MMR", "gene", "germline", "mutation" ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Ovarian", "cancer", "is", "the", "leading", "cause", "of", "death", "from", "gynecological", "cancer", "in", "the", "Western", "world", "[1].", "The", "treatment", "of", "ovarian", "adenocarcinoma", "has", "improved", "over", "the", "last", "20", "years", "owing", "to", "the", "combined", "treatment", "of", "cytoreductive", "surgery", "and", "chemotherapy", "[2].", "Although", "the", "response", "of", "the", "primary", "tumor", "to", "taxane", "and", "platinum-based", "chemotherapy", "is", "high,", "about", "20%", "of", "patients", "never", "achieve", "a", "clinical", "response", "and", "the", "majority", "of", "the", "patients", "will", "relapse", "and", "eventually", "die", "of", "drug-resistant", " ", "disease", "[3]." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 17, 0, 0, 0 ]

[ "20q11.22-q11.23" ]

[ 0 ]

[ "*Amsterdam", "criteria", "II" ]

[ 0, 0, 0 ]

[ "MSI", "is", "an", "important", "phenotype", "of", "MMR", "gene", "mutation.", "In", "1997,", "the", "National", "Cancer", "Institute", "(NCI)", "recommended", "screening", "MSI", "CRCs", "using", "the", "Bethesda", "guidelines", "[5].", "After", "compiling", "evidence", "from", "years", "of", "global", "studies", "and", "follow", "up,", "NCI", "revised", "the", "Bethesda", "guidelines", "in", "2004,", "which", "is", "called", "the", "revised", "Bethesda", "standard", "[6].", "NCI", "recommended", "screening", "of", "HNPCC", "based", "on", "detection", "of", "MSI", "in", "the", "tumor", "and", "loss", "of", "expression", "of", "a", "MMR", "gene", "using", "immunohistochemistry", "(IHC)", "staining", "[6].", "However,", "until", "now,", "there", "has", "been", "no", "research", "about", "the", "applicability", "of", "the", "NCI", "recommendations", "to", "the", "Chinese", "population", "with", "colorectal", "cancers.", "The", "aim", "of", "this", "study", "is", "to", "detect", "and", "study", "MSI", "carrier", "and", "mismatch", "repair", "(MMR)", "gene", "germline", "mutation", "carriers", "HNPCC", " ", "is", "an", "autosomal", "dominant", "syndrome,", "which", "accounts", "for", "about", "1–5%", "of", "colorectal", "cancer", "[1].", "HNPCC", " ", "patients", "are", "characterized", "by", "earlier", "symptoms,", "more", "mucinous", "carcinoma,", "more", "synchronous", "and", "metachronous", "colorectal", "tumors", "and", "more", "extra-colonic", "tumors,", "but", "have", "better", "survival", "[1,2].", "It", "is", "believed", "that", "HNPCC", "is", "secondary", "to", "a", "germline", "mutation", "resulting", "in", "a", "defective", "MMR", "gene.", "A", "defective", "MMR", "gene", "results", "in", "increased", "DNA", "replication", "errors", "and", "MSI,", "which", "causes", "the", "occurrence", "of", "tumors", "in", "different", "organs,", "especially", "in", "the", "colorectum.", "Identification", "of", "HPNCC", "families", "is", "important", "because", "the", "diagnosis,", "treatment", "and", "follow", "up", "of", "these", "individuals", "should", "be", "different", "from", "those", "with", "sporadic", "colorectal", "cancer", "[2].", "However,", "the", "clinical", "diagnosis", "of", "HNPCC", "patients", "is", "very", "difficult", "for", "lack", "of", "specific", "clinical", "phenotype.", "Though", "Amsterdam", "criteria", "I", "and", "II", "were", "established", "for", "HNPCC", "diagnosis[3,4],", "many", "HNPCC", "families", "still", "do", "not", "meet", "the", "criteria." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 9, 1, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "ACF,", "Aberrant", "crypt", "foci", "(hyperplastic", "or", "dysplastic);", "HP,", "hyperplastic", "polyp;", "SSA,", "sessile", "serrated", " ", "adenoma;", "SA", "SSA", ",", "sessile", " ", "serrated", "adenoma;", "SA,", "serrated", "adenoma;", "TA,", "tubular", "serrated", " ", "adenoma;", "TA", "lesions", "serrated", "mutation", "BRAF", "polyps", "dysplasia", "polyps", "serrated", " ", "polyps", "with", "dysplasia", "evolve", "through", "at", "least", "two", "independent", "histogenetic", "pathways.", "Group", "A", "polyps", "are", "likely", "to", "be", "initiated", "by", "BRAF", "mutation", "and", "are", "serrated", "lesions", "at", "the", "outset.", "Group", "B", "polyps", "may", "begin", "as", "conventional", "adenomas", "and", "then", "become", "serrated", "and", "villous", "following", "mutation", "of", "KRAS.", "There", "may", "be", "instances", "in", "which", "KRAS", "mutation", "can", "initiate", "lesions", "which", "are", "serrated", "at", "the", "outset", "and", "become", "dysplastic.", "While", "this", "suggestion", "remains", "speculative,", "it", "may", "apply", "to", "the", "Group", "A", "serrated", "polyps", "that", "show", "a", "distinct", "likeness", "to", "the", "goblet", "cell", "variant", "of", "HP", "in", "which", "columnar", "cells", "are", "eosinophilic", "and", "lack", "mucin-filled", "microvesicles", "and", "KRAS", "mutation", "is", "frequent", "(Figure", "1E,F).16,25", "The", "literature", "refers", "to", "the", "concept", "of", "‘traditional’", "SA.43", "Based", "on", "the", "current", "findings,", "it", "is", "likely", "that", "several", "mechanisms", "can", "account", "for", "adenomatous", "lesions", "with", "glandular", "serration", "and", "that", "‘traditional’", "SA", "is", "not", "a", "single", "entity." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 17, 0, 0, 1, 1, 0, 17, 0, 0, 0, 0, 0, 0, 0, 17, 17, 0, 0, 1, 1, 17, 5, 21, 1, 19, 1, 17, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Families", "carrying", "the", "p.Lys618Ala", "variant" ]

[ 0, 0, 0, 0, 0 ]

[ "Genotyping", "of", "the", "MLH1", "p.Lys618Ala", "variant" ]

[ 0, 0, 0, 0, 0, 0 ]

[ "Mutation", "frequencies", "for", "both", "KRAS", "(P", "<", "0.0001)", "and", "BRAF", "(P", "<", "0.0001)", "are", "distributed", "differently", "across", "the", "seven", "classes", "of", "polyp", "(see", "Results", "for", "individual", "comparisons).", "Distribution", "of", "MGMT", "loss", "differs", "across", "the", "seven", "classes", "of", "polyp", "(P", "<", "0.001)." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "The", "method", "for", "mutation", "analysis", "of", "the", "APC", "mutation", "cluster", "region", "and", "exon", "1", "of", "K-ras", "is", "based", "on", "nested", "amplification", "and", "direct", "sequencing", "of", "purified", "PCR", "fragments,", "a", "highly", "sensitive", "method.", "Since", "no", "screening", "step", "was", "performed", "prior", "to", "the", "sequencing", "of", "the", "gene", "fragments,", "it", "is", "unlikely", "that", "mutations", "would", "have", "escaped", "detection.", "The", "reproducibility", "of", "the", "applied", "assays", "was", "good,", "with", "a", "reproducibility", "of", "85%", "and", "88%", "for", "APC", "and", "K-ras,", "respectively.", "Arguably,", "this", "indicates", "the", "extent", "of", "heterogeneity", "present", "in", "the", "tumour", "samples." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "**", "IGNORE", "LINE", "**" ]

[ 0, 0, 0, 0 ]

[ "Amplification", "of", "exon", "3", "of", "the", "CTNNB1", "gene", "entailed", "a", "semi-nested", "PCR", "strategy,", "which", "covered", "codons", "33,", "37,", "41", "and", "45.", "Flank", "PCR", "was", "performed", "to", "generate", "a", "308", "bp", "fragment", "(primers,", "forward:", "5'-CCAATCTACTAATGCTAATACTG-3',", "reverse:", "5'-GCATTCTGACTTTCAGTAAGGC-3')", "that", "was", "used", "in", "a", "1:100", "dilution", "for", "amplification", "of", "the", "final", "PCR", "product", "(primers,", "forward:", "5'-CCAATCTACTAATGCTAATACTG-3',", "reverse:", "5'-CTTCCTCAGGATTGCCTTTACC-3').", "In", "each", "PCR,", "one", "round", "of", "35", "cycles", "was", "performed." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Activation", "of", "the", "Wnt", "pathway", "plays", "a", "central", "role", "in", "the", "aetiology", "of", "most", "colorectal", "cancers", "and", "is", "often", "the", "result", "of", "mutations", "in", "the", "N-terminal", "domain", "of", "the", "APC", "gene,", "that", "lead", "to", "partial", "or", "complete", "loss", "of", "this", "region", "and", "thereby", "to", "loss", "of", "the", "β-catenin", "regulating", "function", "[5,6].", "Conversely,", "in", "tumours", "lacking", "these", "APC", "mutations", "[7],", "activating", "missense", "mutations", "at", "one", "of", "the", "phosphorylation", "sites", "at", "codons", "31,", "33,", "37", "and", "45", "of", "exon", "3", "of", "the", "CTNNB1", "gene", "(encoding", "the", "β-catenin", "protein)", "can", "render", "it", "stable", "as", "it", "can", "no", "longer", "be", "tagged", "for", "cellular", "degradation.", "Activation", "of", "the", "Ras", "pathway", "in", "cancer", "is", "marked", "by", "the", "loss", "of", "the", "intrinsic", "GTPase", "activity", "of", "the", "Ras", "protein,", "which", "can", "be", "ascribed", "to", "missense", "mutations", "in", "codons", "12", "and", "13", "of", "exon", "1,", "which", "are", "responsible", "for", "90%", "activating", "mutations", "in", "the", "of", "the", "K-ras", "gene", "[8]." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Concomitant", "deleterious", "variants", "were", "detected", "in", "two", "of", "the", "families:", "one", "in", "the", "MLH1", "gene", "(c.676C>T;", "p.Arg226X)", "and", "the", "other", "in", "the", "MSH6", "gene", "(c.3013C>T;", "p.Arg1005X).", "Seventeen", "affected", "and", "unaffected", "46.7%", "373", " ", "sporadic", "CRC", "patients,", "250", "index", "subjects", "from", "families", "suspected", "of", "having", "LS", "[revised", "Bethesda", "Guidelines]", "and", "411", "controls).", "The", "controls", "were", "selected", "from", "the", "same", "hospitals,", "had", "no", "personal", "histories", "of", "cancer", "and", "had", "diagnoses", "unrelated", "to", "the", "variables", "of", "interest.", "They", "were", "matched", "for", "age,", "gender", "and", "race/ethnicity", "with", "the", "sporadic", "CRC", "patients." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 11, 11, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "To", "establish", "and", "explain", "the", "pattern", "of", "molecular", "signatures", "across", "colorectal", "polyps." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Results" ]

[ 0 ]

[ "p53", "immunohistochemistry", "and", "scoring" ]

[ 0, 0, 0, 0 ]

[ "In", "recent", "years,", "the", "fundamental", "division", "of", "colorectal", "polyps", "into", "precancerous", "adenomas", " ", "and", "innocent", "HPs", "has", "begun", "to", "erode", "and", "the", "concept", "of", "an", "alternative", "serrated", "pathway", "has", "gained", "support.", "This", "revision", "began", "with", "the", "description", "of", "an", "intermediate", "lesion", "described", "as", "serrated", "adenoma", " ", "(SA).10", "Initially,", "however,", "SA", "was", "not", "conceived", "as", "an", "intermediate", "category", "of", "polyp", "but", "essentially", "as", "an", "adenoma", "with", "a", "superimposed", "serrated", "architecture", "that", "conferred", "only", "a", "superficial", "likeness", "to", "a", "HP.10", "Additionally,", "the", "mixed", "polyp", "(MP)", "was", "perceived", "as", "a", "chance", "collision", "between", "a", "HP", "and", "an", "adenoma,", "giving", "a", "combined", "polyp.", "These", "preliminary", "interpretations", "did", "not", "represent", "a", "major", "departure", "from", "the", "traditional", "classification", "of", "colorectal", "polyps", "but", "preserved", "the", "fundamental", "distinction", "of", "neoplastic", "adenomas", "versus", "non-neoplastic", "HPs." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Details", "of", "the", "8", "patients", "in", "MSS", "group", "identified", "to", "have", "MMR", "MSS", "CRCs", " ", "are", "summarized", "in", "Table", "6.", "Six", "CRCs", "with", "mutations", "were", "MSI-H", "and", "two", "patients", "with", "mutations", "were", "MSI-L." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 21, 19, 1, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "APC", "mutation", "analysis" ]

[ 0, 0, 0 ]

[ "The", "semi-nested", "PCR", "products", "from", "samples", "without", "a", "truncating", "APC", "mutation", "were", "screened", "for", "mutations", "using", "denaturing", "high-pressure", "liquid", "chromatography", "(dHPLC)", "on", "a", "WAVE", "3500", "HT", "system", "(Transgenomic", "Inc.,", "UK).", "WAVE", "analysis", "was", "optimised", "and", "validated", "using", "specific", "mutations", "in", "cell", "line", "DNA,", "i.e.", "HCT116", "(codon", "45:", "3", "bp", "deletion)", "and", "SW48", "(codon", "33:", "C→A)", "as", "well", "as", "DNA", "derived", "from", "desmoid", "tumours", "from", "patients", "(codon", "41:", "A→G", "and", "codon", "45:", "C→T)", "as", "positive", "controls", "APC", "mutation", "K-ras", " ", "mutation", "analysis" ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 9, 10, 21, 5, 21, 0, 0, 0 ]

[ "In", "a", "group", "of", "656", "unselected", "sporadic", "colorectal", "cancer", "patients,", "aberrations", "in", "the", "APC,", "K-ras", ",", "CTNNB1", "genes,", "and", "expression", "of", "hMLH1", "were", "investigated.", "Additionally,", "tumours", "were", "divided", "in", "groups", "based", "on", "molecular", "features", "and", "compared", "with", "respect", "to", "patient's", "age", "at", "diagnosis,", "sex,", "family", "history", "of", "colorectal", "cancer,", "tumour", "sub-localisation,", "Dukes'", "stage", "and", "differentiation." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 21, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Mutation", "spectrum", "of", "the", "APC", "gene.", "(A)", "The", "spectrum", "of", "APC", "mutations", "identified", "among", "families", "from", "the", "Swedish", "Polyposis", "Registry", "showing", "the", "distribution", "between", "previously", "reported", "and", "novel", "mutations", "in", "our", "patients.", "(B)", "A", "schematic", "representation", "of", "the", "APC", "coding", "region,", "shown", "in", "the", "same", "scale", "as", "in", "(A).", "The", "arrow", "with", "an", "asterisk", "indicates", "codon", "24", "and", "the", "second", "arrow", "points", "at", "codon", "184.", "(C)", "Distribution", "of", "six", "large", "deletions", "found", "in", "seven", "unrelated", "patients", "of", "the", "Swedish", "Polyposis", "Registry.", "Novel", "deletions", "are", "marked", "with", "an", "asterisk.", "Patient", "numbers", "are", "shown", "to", "the", "left.", "Scale", "as", "in", "(A)." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Large", "deletions", "of", "the", "APC", "CRC", "mutations", "in", "the", "region", "from", "codon", "1250", "to", "1464", "Probands", " ", "with", "mutations", "in", "the", "region", "from", "codon", "1250", "to", "1464", "of", "the", "APC", "gene", "which", "predicts", "a", "severe", "phenotype", "[36]", "had", "a", "median", "age", "at", "diagnosis", "of", "21.8", "(range,", "11–49)", "years", "compared", "with", "34.4", "(range,", "14–57)", "years", "among", "those", "with", "mutations", "outside", "this", "region", "range,", "14–57", "34.4", "range,", "11–49", ")", "years", "compared", "with", "34.4", "(range,", "14–57)", "years", "among", "those", "with", "mutations", "outside", "this", "region", "(P", "<", "0.017).", "Available", "data", "on", "colorectal", "polyp", "number", "shows", "that,", "in", "spite", "of", "higher", "age", "at", "diagnosis,", "dense", "polyposis", "(>", "1000)", "only", "occurred", "in", "30%", "of", "the", "probands", "compared", "with", "75%", "in", "those", "with", "mutations", "between", "codon", "1250", "and", "1464.", "In", "the", "former", "group", "29%", "(7", "out", "of", "24)", "had", "CRC", "at", "diagnosis", "compared", "with", "25%", "(2", "out", "of", "8)", "in", "the", "latter", "group.", "The", "mean", "age", "at", "CRC", "was", "46.6", "(range", "28–57)", "and", "37.5", "(range", "26–49)", "years,", "respectively.", "The", "total", "morbidity", "in", "CRC", "among", "probands", "was", "34%", "(11", "out", "of", "32).", "Of", "all", "probands", "diagnosed", "after", "1996,", "four", "out", "of", "nine", "(44%)", "had", "cancer", "at", "diagnosis.", "The", "median", "age", "in", "this", "group", "was", "47.5", "(range", "45–51)", "years", "and", "none", "had,", "despite", "high", "age", "at", "diagnosis", "of", "CRC,", "dense", "colorectal", "polyposis", "at", "diagnosis", "indicating", "a", "less-severe", "phenotype.", "A", "compilation", "of", "clinical", "status", "of", "all", "patients", "analyzed", "in", "this", "study", "is", "shown", "in", "Additional", "file", "1." ]

[ 0, 0, 0, 0, 21, 1, 5, 6, 6, 6, 6, 6, 6, 6, 6, 9, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 9, 10, 10, 10, 10, 10, 15, 16, 15, 15, 16, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Classification", "of", "the", "MLH1", "p.Lys618Ala", "variant", "according", "to", "the", "InSiGHT", "database", "(accessed", "on", "07/2010)." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Associations", "between", "BRAF", "mutation", "and", "molecular", "features", "of", "colorectal", "cancer." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Whole-chromosome", "plots", "of", "running", "average", "of", "fractions", "of", "samples", "showing", "up-/down-regulation", "in", "tumor", "versus", "normal", "normal", " ", "samples", "(Chromosomes", "1,", "2,", "3,", "4).", "For", "each", "chromosome", "you", "see", "a", "separate", "figure.", "Gray", "dots", "denote", "the", "number", "of", "patients", "with", "up-", "or", "down-regulation", "for", "a", "single", "gene.", "Orange/green", "lines", "represent", "a", "running", "average", "of", "these", "values.", "The", "plots", "are", "made", "to", "be", "easily", "comparable", "with", "whole-genome", "CGH", "plots", "(like", "e.g.", "those", "in", "Knösel", "et", "al.", "[21])", "Further", "details", "of", "plot", "construction", "are", "described", "in", "the", "methods", "section." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 1, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "This", "paper", "explores", "the", "possibility", "that", "the", "early", "evolution", "of", "colorectal", "cancer", " ", "is", "not", "limited", "to", "two", "essentially", "independent", "pathways,", "but", "often", "combines", "components", "of", "these", "pathways.", "Indeed,", "the", "successful", "‘fusion’", "of", "the", "hyperproliferation", "and", "crypt", "fission", "that", "characterize", "adenomas21", "with", "the", "inhibition", "of", "apoptosis", "that", "has", "been", "linked", "with", "serrated", "polyps22,23", "may", "generate", "lesions", "with", "enhanced", "aggressiveness.", "Specifically,", "it", "is", "suggested", "that", "methylation", "of", "the", "DNA", "repair", "gene", "O-6-methylguanine", "DNA", "methyltransferase", "(MGMT),", "mutation", "of", "KRAS", "and", "inactivation", "BRAF", " ", "and", "extensive", "DNA", "methylation.12", "This", "viewpoint", "was", "consolidated", "through", "the", "formal", "recognition", "of", "two", "largely", "independent", "pathways", "of", "colorectal", "tumorigenesis:", "(i)", "the", "traditional", "adenoma–", "carcinoma", "DNA", "methylation", ".12", "This", "viewpoint", "was", "consolidated", "through", "the", "formal", "recognition", "of", "two", "largely", "independent", "pathways", "of", "colorectal", "tumorigenesis:", "(i)", "the", "traditional", "adenoma", "colorectal", " ", "tumorigenesis", ":", "(i)", "the", "traditional", "adenoma–carcinoma", "sequence", "associated", "with", "chomosomally", "unstable", "CRCs,13", "and", "(ii)", "the", "‘serrated", "pathway’", "culminating", "in", "CRCs", "with", "DNA", "microsatellite", "instability", "(MSI),", "mutation", "of", "BRAF", "and", "extensive", "DNA", "methylation.12,14–20" ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 2, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 5, 21, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 7, 8, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 3, 0, 1, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Inactivation", "of", "MMR", "leads", "to", "the", "occurrence", "of", "unrepaired", "deletions", "in", "mono-", "and", "dinucleotide", "repeats", "resulting", "in", "variable", "lengths", "of", "these", "repeats.", "This", "is", "called", "microsatellite", "instability", "(MSI)", "and", "MSI", "is", "therefore", "used", "as", "a", "marker", "for", "MMR", "deficiency.", "MSI", "can", "be", "caused", "by", "genetic", "or", "epigenetic", "inactivation", "of", "several", "genes", "involved", "in", "MMR.", "Mouse", "knockout", "models", "have", "demonstrated", "that", "MSH2-/-,", "MSH3-/-,", "MLH1-/-", "and", "PMS2-/-", "leads", "to", "a", "high", "frequency", "of", "MSI", "while", "MSH6-/-", "and", "PMS1-/-", "cause", "a", "low", "frequency", "(reviewed", "by", "Wei", "et", "al.", "[10]).", "However,", "in", "hereditary", "nonpolyposis", "colon", "cancer", "(HNPCC)", "families", "ovarian", "ovarian", " ", "adenocarcinoma", "has", "improved", "over", "the", "last", "20", "years", "owing", "to", "the", "combined", "treatment", "of", "cytoreductive", "surgery", "and", "chemotherapy", "[2].", "Although", "the", "response", "of", "the", "primary", "tumor", "to", "taxane", "and", "platinum-based", "chemotherapy", "is", "high,", "about", "20%", "of", "patients", "never", "achieve", "a", "clinical", "response", "and", "the", "majority", "of", "the", "patients", " ", "will", "relapse", "and", "eventually", "die", "of", "drug-resistant", "disease", "[3]." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 9, 3, 3, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 9, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Conclusion" ]

[ 0 ]

[ "When", "comparing", "tumours", "with", "a", "missense", "(but", "not", "a", "truncating)", "mutation", "in", "APC", "to", "tumours", "with", "a", "truncating", "mutation", "in", "APC,", "missense", "mutations", "occurred", "relatively", "more", "frequently", "in", "the", "colon", "(P", "=", "0.002),", "less", "often", "also", "harboured", "an", "activating", "K-ras", "mutation", "(P", "=", "0.004),", "and", "more", "often", "also", "lacked", "hMLH1", "hMLH1", " ", "expression,", "and", "these", "results", "are", "presented", "in", "table", "4.", "Patients", "harbouring", "hMLH1", " ", "deficient", "tumours", "were", "slightly", "older", "when", "diagnosed", "with", "colorectal", "cancer", "(69.3", "yr", "(68.0–70.5)", "versus", "67.8", "(67.4–68.3),", "P", "=", "0.03),", "were", "relatively", "less", "frequently", "men", "(40%", "versus", "58%,", "P", "=", "0.02).", "Tumours", "without", "hMLH1", "expression", "occurred", "relatively", "more", "frequently", "in", "the", "proximal", "colon", "(P", "<", "0.001)", "and", "relatively", "more", "frequently", "showed", "poor", "differentiation", "or", "are", "undifferentiated", "(P", "<", "0.001)." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 21, 21, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 21, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "The", "preceding", "findings", "indicate", "that", "serrated", "polyps", "with", "dysplasia", "evolve", "through", "at", "least", "two", "independent", "histogenetic", "pathways.", "Group", "A", "polyps", "are", "likely", "to", "be", "initiated", "by", "BRAF", "mutation", "and", "are", "serrated", "lesions", "at", "the", "outset.", "Group", "B", "polyps", "may", "begin", "as", "conventional", "adenomas", "and", "then", "become", "serrated", "and", "villous", "following", "mutation", "of", "KRAS.", "There", "may", "be", "instances", "in", "which", "KRAS", "mutation", "can", "initiate", "lesions", "which", "are", "serrated", "at", "the", "outset", "and", "become", "dysplastic.", "While", "this", "suggestion", "remains", "speculative,", "it", "may", "apply", "to", "the", "Group", "A", "serrated", "polyps", "that", "show", "a", "distinct", "likeness", "to", "the", "goblet", "cell", "variant", "of", "HP", "in", "which", "columnar", "cells", "are", "eosinophilic", "and", "lack", "mucin-filled", "microvesicles", "and", "KRAS", "mutation", "is", "frequent", "(Figure", "1E,F).16,25", "The", "literature", "refers", "to", "the", "concept", "of", "‘traditional’", "SA.43", "Based", "on", "the", "current", "findings,", "it", "is", "likely", "that", "several", "mechanisms", "can", "account", "for", "adenomatous", "lesions", "with", "glandular", "serration", "and", "that", "‘traditional’", "SA", "is", "not", "a", "single", "entity." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Methods" ]

[ 0 ]

[ "*Amsterdam", "criteria", "II" ]

[ 0, 0, 0 ]

[ "Associations", "between", "BRAF", "mutation", "and", "clinicopathological", "features", "of", "colorectal", "cancer." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "There", "were", "no", "significant", "associations", "in", "the", "case-control", "and", "case-case", "studies", "(80%", "detection", "power,", "OR", "=", "3.0;", "two-sided", "test,", "alpha", "level", "=", "5%)", "(Table", "2)", "and", "no", "statistically", "significant", "associations", "when", "the", "OR", "was", "adjusted", "for", "age", "and", "sex." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Three", "characterized", "LS", "families", "that", "fulfilled", "the", "Amsterdam", "II", "Criteria", "and", "that", "consisted", "of", "members", "with", "the", "p.Lys618Ala", "variant", "were", "included", "to", "assess", "co-occurrence", "and", "co-segregation.", "Two", "families", "attended", "the", "Genetic", "Counselling", "in", "Cancer", "Units", "of", "the", "Elche", "and", "La", "Fe", "Hospitals", "and", "one", "family", "was", "a", "member", "of", "the", "EPICOLON", "cohort", "[7]." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Results" ]

[ 0 ]

[ "In", "the", "present", "study", "patients", "included", "in", "the", "Swedish", "Polyposis", "Registry", "were", "subjected", "to", "a", "thorough", "clinical", "characterization", "and", "mutational", "screening", "of", "the", "APC", "gene", "including", "screening", "for", "large", "deletions", "and", "detection", "of", "low-frequency", "alleles", "caused", "by", "germline", "mosaicism.", "Sixty-one", "mutations", "Sixty-one", "APC", " ", "gene", "including", "screening", "for", "large", "deletions", "and", "detection", "of", "low-frequency", "alleles", "caused", "by", "germline", "mosaicism.", "Sixty-one", "mutations,", "including", "27", "not", "described", "previously,", "are", "reported.", "Among", "the", "characterized", "mutations", "are", "elusive", "changes", "such", "as", "a", "case", "of", "mosaicism,", "splicing", "defects,", "and", "a", "mutation", "in", "APC", "exon", "1", "which", "is", "the", "most", "5'", "APC", "APC", " ", "exon", "1", "which", "is", "the", "most", "5'", "APC", "mutation", "hitherto", "reported.", "The", "detection", "of", "reduced", "APC", "expression", "in", "one", "family", "splicing", "defects", ",", "and", "a", "mutation", "in", "APC", "exon", "1", "which", "is", "the", "most", "5'", "APC", "mutation", "hitherto", "reported.", "The", "detection", "of", "reduced", "APC", "patients", " ", "included", "in", "the", "Swedish", "Polyposis", "Registry", "were", "subjected", "to", "a", "thorough", "clinical", "characterization", "and", "mutational", "screening", "of", "the", "APC", "gene", "including", "screening", "for", "large", "deletions", "and", "detection", "of", "low-frequency", "alleles", "caused", "by", "germline", "mosaicism.", "Sixty-one", "mutations,", "including", "27", "not", "described", "previously,", "are", "reported.", "Among", "the", "characterized", "mutations", "are", "elusive", "changes", "such", "as", "a", "case", "of", "mosaicism,", "splicing", "defects,", "and", "a", "mutation", "in", "APC", "exon", "1", " ", "which", "is", "the", "most", "5'", "APC", "mutation", " ", "hitherto", "reported.", "The", "detection", "of", "reduced", "APC", "expression", " ", "in", "one", "Swedish", " ", "Polyposis", " ", "Registry", "were", "subjected", "to", "a", "thorough", "clinical", "characterization", "and", "mutational", "screening", "of", "the", "APC", "gene", "including", "screening", "for", "large", "deletions", "and", "detection", "of", "low-frequency", "alleles", "caused", "by", "germline", "mosaicism.", "Sixty-one", "mutations,", "including", "27", "not", "described", "previously,", "are", "reported.", "Among", "the", "characterized", "mutations", "are", "elusive", "changes", "such", "as", "a", "case", "of", "mosaicism", ",", "splicing", "defects,", "and", "a", "mutation", "in", "APC", "exon", "1", "which", "is", "the", "most", "5'", "APC", "mutation", "hitherto", "reported.", "The", "detection", "of", "reduced", "APC", "expression", "in", "one", "family", "is", "also", "described.", "A", "combination", "of", "mutation", "screening", "techniques", "was", "used", "to", "achieve", "as", "high", "a", "mutation-detection", "frequency", "as", "possible." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 5, 11, 21, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 21, 21, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 9, 5, 6, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 21, 9, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 5, 6, 6, 6, 6, 0, 0, 0, 0, 0, 5, 6, 6, 0, 0, 0, 0, 0, 0, 7, 8, 8, 0, 0, 11, 25, 0, 1, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 5, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Multistage", "models", "are", "mechanistically", "different", "from", "tumor", "progression", "models", "and", "more", "consistent", "with", "a", "hypothesis", "that", "mutations", "acquired", "early", "during", "progression", "help", "determine", "extent", "of", "invasion", "(Figure", "3).", "Mutations", "sequentially", "accumulate", "before", "transformation", "in", "both", "models,", "but", "the", "adenoma-cancer", "sequence", "suggests", "most", "cancer", "mutations", "start", "to", "accumulate", "after", "the", "age", "of", "50", "years", "in", "adenomas", "[7].", "Such", "tumor", "progression", "imposes", "purpose", "to", "early", "mutations", "because", "each", "additional", "mutation", "confers", "incremental", "changes", "to", "a", "non-invasive", "adenoma", "phenotype.", "Therefore,", "tumor", "progression", "models", "would", "likely", "differ", "between", "MSI+", " ", "and", "MSI-", "cancers", "because", "their", "biology", "and", "types", "of", "mutations", "are", "quite", "different", "[7,8]." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 19, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Sixty-one", "different", "APC", "mutations", "in", "81", "of", "the", "96", "families", "were", "identified", "and", "27", "of", "those", "are", "novel.", "We", "have", "previously", "shown", "that", "6", "of", "the", "96", "patients", "27", " ", "of", "those", "are", "novel.", "We", "have", "previously", "shown", "that", "6", "of", "the", "96", "patients", "carried", "biallelic", "MUTYH", "mutations.", "The", "9", "mutation-negative", "cases", "families" ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 9, 5, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 9, 10, 9 ]

[ "12q14.2-12q22" ]

[ 0 ]

[ "The", "mutation", "between", "the", "MSI-L", "and", "MSI-H" ]

[ 0, 0, 0, 0, 0, 0, 0 ]

[ "**", "IGNORE", "LINE", "**" ]

[ 0, 0, 0, 0 ]

[ "**", "IGNORE", "LINE", "**" ]

[ 0, 0, 0, 0 ]

[ "Results", "of", "genotyping", "for", "the", "p.Lys618Ala", "variant", "using", "the", "iPLEX", "Sequenom", "(A)", "and", "sequencing", "(B)", "methods." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Incident", "cancer", "cases", "are", "identified", "by", "monitoring", "of", "the", "entire", "cohort", "for", "cancer", "occurrence", "through", "annual", "record", "linkage", "to", "the", "Netherlands", "Cancer", "Registry,", "i.e.", "nine", "regional", "cancer", "registries", "throughout", "the", "Netherlands,", "and", "to", "PALGA,", "a", "nationwide", "network", "and", "registry", "of", "histo-", "and", "cytopathology", "[30].", "Together,", "the", "NCR", "and", "PALGA", "provide", "a", "near", "100%", "coverage", "of", "the", "municipalities", "included", "in", "the", "NLCS.", "The", "first", "2.3", "years", "of", "follow", "up", "were", "excluded", "because", "of", "possible", "pre-clinical", "disease", "affecting", "exposure", "status", "and", "because", "of", "incomplete", "nationwide", "coverage", "of", "PALGA", "in", "some", "of", "the", "municipalities", "included", "in", "the", "NLCS", "in", "that", "period.", "From", "1989", "until", "1994,", "929", "incident", "cases", "with", "histologically", "confirmed", "colorectal", "cancer", "were", "identified", "within", "the", "cohort,", "of", "whom", "819", "could", "also", "be", "linked", "to", "a", "PALGA", "report", "of", "the", "lesion." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Genetic", "instability", "is", "seen", "in", "most", "types", "of", "cancer", "[10].", "Two", "distinct", "types", "of", "genetic", "instability", "appear", "to", "occur", "in", "colorectal", "cancer", "[11]:", "chromosomal", "and", "microsatellite", "instability.", "Chromosomal", "instability", "results", "in", "gains", "or", "losses", "of", "entire", "chromosomes", "or", "parts", "of", "them,", "and", "gives", "rise", "to", "aneuploid", "tumours", "and", "occurs", "in", "the", "majority", "of", "cancers.", "A", "smaller", "proportion", "of", "colorectal", "cancers", "displays", "microsatellite", "instability,", "represented", "by", "diploid", "cells", "acquiring", "high", "mutation", "rates,", "and", "was", "found", "to", "be", "associated", "with", "defective", "mismatch", "repair", "[12].", "These", "tumours", "are", "less", "likely", "to", "harbour", "mutations", "in", "genes", "associated", "with", "chromosomally", "instable", "and", "generally", "aneuploid", "tumours,", "such", "as", "APC,", "K-ras", "and", "TP53", "[13-21],", "suggesting", "that", "these", "tumours", "form", "a", "distinct", "group.", "Moreover,", "microsatellite", "instable", "tumours", "are", "found", "predominantly", "in", "the", "proximal", "colon", "[22,23],", "are", "more", "likely", "to", "occur", "in", "patients", "with", "a", "positive", "family", "history", "of", "colorectal", "cancer", "[22,23],", "are", "often", "less", "differentiated", "than", "microsatellite", "stable", "tumours", "[22],", "and", "occur", "more", "frequently", "in", "women", "[24]", "and", "at", "older", "age", "[25].", "Moreover,", "in", "tumours", "colorectal", " ", "cancer", "has", "been", "proposed,", "which", "describes", "the", "sequential", "accumulation", "of", "specific", "genetic", "alterations", "in", "various", "pathways,", "involving", "tumour", "suppressor", "genes", "(e.g.", "APC,", "SMAD4,", "TP53)", "and", "oncogenes", "(e.g.", "CTNNB1,", "K-ras)", "[3,4].", "Important", "molecular", "pathways", "that", "upon", "activation", "affect", "the", "early", "and", "intermediate", "stages", "of", "colorectal", "carcinogenesis", "are", "the", "Wnt", "and", "Ras", "signalling", "pathways,", "whereas", "TP53", "inactivation", "is", "considered", "a", "late", "event." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 3, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "**", "IGNORE", "LINE", "**" ]

[ 0, 0, 0, 0 ]

[ "The", "semi-nested", "PCR", "products", "from", "samples", "without", "a", "truncating", "APC", "mutation", "were", "screened", "for", "mutations", "using", "denaturing", "high-pressure", "liquid", "chromatography", "(dHPLC)", "on", "a", "WAVE", "3500", "HT", "system", "(Transgenomic", "Inc.,", "UK).", "WAVE", "analysis", "was", "optimised", "and", "validated", "using", "specific", "mutations", "in", "cell", "line", "DNA,", "i.e.", "HCT116", "(codon", "45:", "3", "bp", "deletion)", "and", "SW48", "(codon", "33:", "C→A)", "as", "well", "as", "DNA", "derived", "from", "desmoid", "tumours", "from", "patients", "(codon", "41:", "A→G", "and", "codon", "45:", "C→T)", "as", "positive", "controls.", "All", "of", "these", "mutations", "were", "repeatedly", "confirmed", "by", "sequencing.", "WAVE", "analysis", "was", "carried", "out", "at", "two", "different", "temperatures", "(57.7", "en", "60°C).", "Samples", "showing", "an", "aberrant", "elution", "profile", "were", "re-amplified", "and", "re-screened.", "When", "an", "aberrant", "elution", "profile", "was", "confirmed,", "direct", "sequencing", "was", "performed.", "All", "samples", "without", "hMLH1", "expression", "codon", "45:", "C→T", ")", "as", "positive", "controls.", "All", "of", "these", "mutations", "were", "repeatedly", "confirmed", "by", "sequencing.", "WAVE", "analysis", "was", "carried", "out", "at", "two", "different", "temperatures", "(57.7", "en", "60°C).", "Samples", "showing", "an", "aberrant", "elution", "profile", "were", "re-amplified", "and", "re-screened.", "When", "an", "aberrant", "elution", "profile", "was", "confirmed,", "direct", "sequencing", "was", "performed.", "All", "samples", "hMLH1", "APC", " ", "mutation", "analysis" ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 7, 8, 8, 5, 6, 6, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 9, 21, 21, 0, 0, 0 ]

[ "P", "=", "0.112" ]

[ 0, 0, 0 ]

[ "Next,", "we", "compared", "the", "patient", "and", "tumour", "characteristics", "of", "tumours", "harbouring", "a", "truncating", "APC", "and/or", "an", "activating", "K-ras", "mutation", "to", "those", "of", "tumours", "without", "hMLH1", "expression,", "and", "these", "results", "are", "presented", "in", "table", "4.", "Patients", "harbouring", "hMLH1", "deficient", "tumours", "were", "slightly", "older", "when", "diagnosed", "with", "colorectal", "cancer", "(69.3", "yr", "(68.0–70.5)", "versus", "67.8", " ", "(67.4–68.3),", "P", "=", "0.03),", "were", "relatively", "less", "frequently", "men", "(40%", "versus", "58%,", "P", "=", "0.02).", "Tumours", "without", "hMLH1", "expression", "occurred", "relatively", "more", "frequently", "in", "the", "proximal", "colon", "(P", "<", "0.001)", "and", "relatively", "more", "frequently", "showed", "poor", "differentiation", "or", "are", "undifferentiated", "(P", "<", "0.001)." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 15, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "No", "familial", "history", "of", "cancer", "was", "available", "from", "the", "control", "group.", "Patients", "diagnosed", "at", "an", "age", "over", "50", "years", "and", "not", "referred", "to", "Genetic", "Counselling", "Units", "were", "considered", "as", "sporadic", "CRC.", "Samples", "from", "sporadic", "CRC", "patients", "were", "obtained", "from", "the", "Elche", "University", "Hospital", "BioBank", "and", "the", "Castellon", "Provincial", "Hospital", "BioBank.", "Written", "consent", "to", "be", "included", "in", "the", "respective", "biobanks", "was", "obtained", "from", "each", "patient.", "CRC", "patients,", "as", "index", "subjects", "from", "families", "with", "suspicion", "of", "LS", "that", "attended", "Genetic", "Counselling", "at", "the", "Cancer", "Units", "of", "the", "Elche", "and", "La", "Fe", "Hospitals,", "were", "recruited.", "The", "study", "was", "approved", "by", "the", "Ethics", "Committee", "of", "the", "Elche", "University", "Hospital." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "**", "IGNORE", "LINE", "**" ]

[ 0, 0, 0, 0 ]

[ "After", "establishing", "the", "familial", "mutation", "in", "the", "proband's", "mother,", "located", "in", "the", "exon", "12", "of", "hMSH2", "hMSH2", " ", "and", "hMLH1", "genes" ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 21, 21, 0, 0, 0, 0 ]

[ "14q24.3" ]

[ 0 ]

[ "The", "MSI", "state,", "mutation", "sites,", "and", "consequential", "changes", "in", "amino-acid", "sequences", "of", "MMR", "genes", "are", "summarized", "in", "Table", "4.", "In", "34", "MSI", "CRCs", "and", "one", "MSS", "CRC", "with", "MLH1", "negative", "staining,", "8", "had", "MMR", "gene", "germline", "mutations,", "accounting", "for", "22.9%", "of", "MSI", "colorectal", "cancers", "and", "5.5%", "of", "all", "colorectal", "cancers.", "Three", "patients", "had", "MSH6", "germline", "mutations", "and", "5", "had", "MSH2", "germline", "mutations;", "the", "clinical", "features", "of", "these", "patients", "are", "summarized", "in", "Table", "5.", "None", "of", "the", "patients", "had", "MLH1", "gene", "mutations.", "Seven", "patients", "in", "the", "MSI", "group", "had", "A/T", "heterozygosis", "in", "MSH6", "codon", "380", "of", "exon", "5,", "but", "it", "did", "not", "cause", "changes", "in", "the", "amino", "acid", "sequence.", "The", "germline", "mutations", "of", "the", "MSI-L", "and", "MSI-H", "CRCs", "are", "summarized", "in", "Table", "6.", "Six", "CRCs", "with", "mutations", "were", "MSI-H", "and", "two", "patients", "with", "mutations", "were", "MSI-L." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Results" ]

[ 0 ]

[ "In", "a", "group", "of", "656", "unselected", "sporadic", "colorectal", "cancer", "patients,", "aberrations", "in", "the", "APC,", "K-ras,", "CTNNB1", "genes,", "and", "expression", "of", "hMLH1", "were", "investigated.", "Additionally,", "tumours", "were", "divided", "in", "groups", "based", "on", "molecular", "features", "and", "compared", "with", "respect", "to", "patient's", "age", "at", "diagnosis,", "sex,", "family", "history", "of", "colorectal", "cancer,", "tumour", "sub-localisation,", "Dukes'", "stage", "and", "differentiation." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "CTNNB1", "mutation", "analysis" ]

[ 0, 0, 0 ]

[ "Patient", "C896,", "with", "only", "five", "adenomas,", "was", "included", "because", "of", "a", "family", "history", "of", "FAP", "FAP", "FAP", ").", "Sixty-one", "of", "these", "families", "are", "now", "extinct", "but", "135", "families", "with", "at", "least", "one", "living", "disease-affected", "member", "remain.", "Presently,", "315", "disease-affected", "living", "patients", "are", "included", "in", "the", "registry.", "The", "geographical", "catchment", "area", "comprises", "the", "whole", "of", "Sweden.", "Details", "of", "how", "patients", "have", "been", "eligible", "for", "accession", "into", "the", "registry", "are", "given", "in", "[30].", "In", "this", "study", "we", "have", "analyzed", "96", "families", "included", "in", "the", "registry", "for", "mutations", "in", "the", "APC", "gene.", "Twenty-four", "of", "the", "remaining", "39", "families", "have", "been", "analyzed", "for", "APC", "gene", "mutations", "at", "other", "genetic", "laboratories", "and", "15", "families", "remain", "to", "be", "tested.", "We", "have", "previously", "reported", "six", "of", "these", "patients", "who", "carried", "bi-allelic", "MUTYH", "mutations", "[31].", "Probands", "were", "defined", "as", "those", "diagnosed", "on", "the", "basis", "of", "the", "occurrence", "of", "symptoms", "and", "irrespective", "of", "other", "cases", "in", "the", "family", "and", "call-up", "patients,", "as", "those", "identified", "as", "subjects", "at", "risk", "on", "the", "basis", "of", "studies", "of", "pedigrees", "and", "found", "to", "have", "FAP.", "De", "novo", "mutations", "were", "defined", "in", "those", "individuals", "where", "none", "of", "the", "parents", "carried", "the", "mutation", "or", "where", "the", "parents", "had", "a", "negative", "colonoscopy", "after", "the", "age", "of", "50", "or", "died", "of", "a", "non-FAP", "related", "cause", "after", "the", "age", "of", "75.", "All", "patients", "have", "given", "their", "consent", "and", "the", "local", "ethics", "committees", "have", "approved", "the", "study.", "The", "clinical", "features", "of", "index", "patients", "of", "each", "of", "the", "families", "families", "families", "families", "families", "families", " ", "are", "now", "extinct", "but", "135", "families", "with", "at", "least", "one", "living", "disease-affected", "member", "remain.", "Presently,", "315", "disease-affected", "living", "patients", "are", "included", "in", "the", "registry.", "The", "geographical", "catchment", "area", "comprises", "the", "whole", "of", "Sweden.", "Details", "of", "how", "patients", "have", "been", "eligible", "for", "accession", "into", "the", "registry", "are", "given", "in", "[30].", "In", "this", "study", "we", "have", "analyzed", "96", "families", "included", "in", "the", "registry", "for", "mutations", "in", "the", "APC", "gene.", "Twenty-four", "of", "the", "remaining", "39", "families", "have", "been", "analyzed", "for", "APC", "gene", "mutations", "at", "other", "genetic", "laboratories", "and", "15", "families", "remain", "to", "be", "tested.", "We", "have", "previously", "reported", "six", "of", "these", "patients", "who", "carried", "bi-allelic", "MUTYH", "mutations", "[31].", "Probands", "were", "defined", "as", "those", "diagnosed", "on", "the", "basis", "of", "the", "occurrence", "of", "symptoms", "and", "irrespective", "of", "other", "cases", "in", "the", "family", "and", "call-up", "patients,", "as", "those", "identified", "as", "subjects", "at", "risk", "on", "the", "basis", "of", "studies", "of", "pedigrees", "and", "found", "to", "have", "FAP.", "De", "novo", "mutations", "were", "defined", "in", "those", "individuals", "where", "none", "of", "the", "parents", "carried", "the", "mutation", "or", "where", "the", "parents", "had", "a", "negative", "colonoscopy", "after", "the", "age", "of", "50", "or", "died", "of", "a", "non-FAP", "related", "cause", "after", "the", "age", "of", "75.", "All", "patients", "have", "given", "their", "consent", "and", "the", "local", "ethics", "committees", "have", "approved", "the", "study.", "The", "clinical", "features", "of", "index", "patients", "of", "each", "of", "the", "families", "analyzed", "are", "listed", "in", "Additional", "file", "1", "and", "Table", "1." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 1, 1, 1, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 9, 9, 9, 9, 9, 9, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "This", "autosomal", "dominantly", "inherited", "disorder", "is", "caused", "by", "germline", "mutations", "in", "genes", "coding", "proteins", "responsible", "for", "the", "repair", "of", "DNA", "replication", "errors,", "which", "are", "referred", "to", "as", "DNA", "mismatch", "repair", "(MMR)", "genes", "[5].", "DNA", "mismatch", "repair", "machinery", "plays", "a", "critical", "role", "in", "genomic", "stability,", "including", "correction", "of", "mispaired", "bases", "associated", "with", "DNA", "replication", "and", "recombination.", "Germline", "mutations", "in", "one", "allele", "of", "any", "of", "these", "genes", "followed", "by", "the", "somatic", "loss", "or", "inactivation", "of", "the", "wild-type", "allele", "leads", "to", "a", "defective", "mismatch", "repair", "mechanism.", "The", "current", "\"gold", "standard\"", "for", "assessing", "tumor", "DNA", "MMR", " ", "activity", "is", "molecular", "microsatellite", "instability", "(MSI)", "testing.", "In", "most", "cases,", "it", "involves", "extracting", "DNA", "from", "both", "tumor", "and", "normal", "tissue.", "The", "DNA", "is", "subjected", "to", "polymerase", "chain", "reaction", "(PCR)", "amplification", "of", "five", "or", "more", "different", "chromosomal", "loci", "that", "compare", "\"microsatellites\",", "running", "the", "PCR", "products", "through", "a", "gel", "to", "separate", "DNA", "fragments", "by", "size,", "comparing", "the", "tumor-normal", "pairs,", "and", "scoring", "for", "differences", "between", "the", "two.", "Instability", "at", "two", "or", "more", "out", "of", "five", "markers", "defines", "a", "tumor", "as", "MSI-H", "and", "prompts", "further", "analysis,", "as", "sequencing", "of", "DNA", "MMR", "genes.", "A", "number", "of", "them", "have", "been", "associated", "with", "HNPCC,", "including", "hMSH2,", "hMLH1,", "hPMS1,", "hPMS2,", "hMSH3", ",", "and", "hMSH6", ".", "Most", "of", "the", "HNPCC", "families", "in", "which", "mutations", "have", "been", "identified", "involved", "hMSH2", "and", "hMLH1", "genes", "[6]." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 21, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 21, 0, 0, 21, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "There", "were", "no", "significant", "associations", "in", "the", "case-control", "and", "case-case", "studies", "(80%", "detection", "power,", "OR", "=", "3.0;", "two-sided", "test,", "alpha", "level", "=", "5%)", "(Table", "2)", "and", "no", "statistically", "significant", "associations", "when", "the", "OR", "was", "adjusted", "for", "age", "and", "sex." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Tissues" ]

[ 0 ]

[ "In", "previous", "studies", "of", "KRAS", "in", "colorectal", "adenomas,", "mutation", "has", "been", "associated", "negatively", "with", "flat", "and", "depressed", "TAs", "and", "positively", "with", "polypoid", "appearance,", "increasing", "size,", "dysplasia,", "villous", "change", "and", "synchronous", "colorectal", "cancer", ".6,33–35", "In", "by", "far", "the", "largest", "study,", "which", "included", "738", "adenomas", "obtained", "from", "639", "participants", "in", "a", "dietary", "intervention", "trial,", "multivariate", "analysis", "showed", "that", "the", "independent", "predictors", "of", "KRAS", "mutation", "were", "age", "of", "subject,", "presence", "of", "villous", "architecture", "and", "high-grade", "dysplasia,", "but", "not", "size", "of", "adenoma.6", "It", "is", "well", "known", "that", "adenoma", "size,", "dysplasia", "and", "villous", "architecture", "are", "interrelated", "and", "account", "has", "to", "be", "taken", "of", "this", "in", "assessing", "results.", "In", "this", "study", "KRAS", "mutation", "occurred", "with", "the", "same", "frequency", "in", "small", "(18%)", "and", "large", "(17%)", "adenomas", "but", "was", "significantly", "more", "frequent", "in", "adenomas", "that", "included", "a", "villous", "architecture", "(50%).", "A", "previous", "study", "showed", "a", "very", "high", "frequency", "of", "KRAS", "mutation", "(93%)", "in", "flat", "synchronous", "adenomas", ",", "second", "on", "non-dysplastic", "serrated", "polyps", "and", "will", "then", "conclude", "with", "the", "concept", "that", "features", "of", "these", "two", "types", "of", "polyp", "can", "co-occur", "or", "become", "‘fused’", "in", "subtypes", "of", "advanced", "polyps", "with", "mixed", "cytomorphology", "(hyperplastic", "and", "dysplastic)", "and", "a", "serrated", "and/or", "villous", "architecture." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Aberrant", "expression", "of", "p53", "and", "correlation", "with", "MGMT", "loss" ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "p53", "immunohistochemistry", "and", "scoring" ]

[ 0, 0, 0, 0 ]

[ "Duke's", "stage", "and", "age", "at", "clinical", "presentation", "(Figure", "1B)", "were", "documented", "for", "884", "of", "the", "895", "MSI-", "sporadic", "cancers", "(Table", "1).", "Average", "ages", "were", "68.6", "years", "for", "stage", "A,", "69.0", "years", "for", "stage", "B,", "65.2", "years", "for", "stage", "C,", "and", "65.4", "years", "for", "stage", "D.", "The", "most", "likely", "numbers", "of", "oncogenic", "mutations", "were", "seven", "for", "stage", "A", "cancers,", "eight", "for", "stage", "B", "cancers,", "and", "six", "for", "stage", "C", "or", "D", "cancers", "(Table", "1)." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "LS", "is", "the", "most", "common", "hereditary", "CRC-predisposing", "syndrome", "and", "accounts", "for", "3%", "of", "unselected", "CRC", "cases.", "A", "significant", "proportion", "of", "DNA", "variations", "found", "in", "patients", "suspected", "of", "having", "LS", "are", "UVs", "(32%,", "18%", "and", "38%", "for", "MLH1,", "MSH2", "and", "MSH6,", "respectively)", "[6].", "The", "pathogenicity", "of", "the", "MLH1", "p.Lys618Ala", "variant", "remains", "controversial", "because", "of", "conflicting", "data", "[InSiGHT,", "http://www.insight-group.org]", "(Figure", "5)." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "MSI", "was", "detected", "in", "three", "of", "the", "eight", "cell", "lines", "i.e.", "A2780", "(no", "MLH1", "mRNA", "expression", "due", "to", "promoter", "methylation),", "SKOV3", "(no", "MLH1", "mRNA", "expression)", "and", "2774", "(no", "altered", "expression", "of", "MMR", "genes).", "Overall,", "there", "was", "no", "association", "between", "cisplatin", "response", "and", "MMR", "status", "in", "these", "eight", " ", "cell", "lines." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 11, 0, 0, 0 ]

[ "Numbers", "of", "oncogenic", "alterations", "(genetic", "mutations", "or", "epigenetic", "alterations)", "required", "for", "transformation", "were", "estimated", "from", "ages", "at", "cancer", "using", "a", "Bayesian", "approach", "as", "previously", "described", "[11].", "This", "method", "requires", "the", "use", "of", "a", "life", "table", "from", "census", "data:", "for", "the", "Finnish", "data", "set", "we", "used", "a", "Finnish", "adenocarcinoma", ",", "ICD-0-2", "codes", "8000–8500),", "and", "stage", "(localized,", "regional,", "or", "distant).", "These", "cancers", "were", "not", "characterized", "with", "respect", "to", "HNPCC", "or", "MSI." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 25, 1, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Results", "of", "case-control", "and", "case-case", "analyses.", "Odds", "ratios", "(ORs)", "and", "95%", "confidence", "intervals", "(95%", "CIs)", "for", "the", "p.Lys618Ala", "variant" ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Our", "data", "may", "imply", "that", "the", "characteristics", "of", "HNPCC", "in", "the", "Chinese", "population", "are", "probably", "different", "from", "those", "of", "Western", "countries.", "Application", "of", "NCI", "recommended", "criteria", "may", "not", "be", "effective", "enough", "to", "identify", "Chinese", "HNPCC", "families.", "Further", "studies", "are", "necessary", "to", "echo", "or", "refute", "our", "results", "so", "as", "to", "make", "the", "NCI", "recommendation", "more", "universally", "applicable." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Splice-site", "affecting", "mutations" ]

[ 0, 0, 0 ]

[ "Methods" ]

[ 0 ]

[ "Results" ]

[ 0 ]

[ "Frequency", "of", "KRAS", "and", "BRAF", "mutation", "and", "loss", "of", "expression", "of", "O-6-methylguanine", "DNA", "methyltransferase", "(MGMT)", "by", "polyp", "type" ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[]

[]

[ "Mutations", "in", "exon", "3", "of", "the", "CTNNB1", "gene", "leading", "to", "loss", "of", "one", "of", "the", "phosphorylation", "sites", "were", "rare.", "Strikingly,", "all", "five", "of", "these", "mutations", "occurred", "in", "the", "proximal", "colon", "and", "three", "of", "these", "also", "had", "absent", "hMLH1", "expression.", "This", "may", "indicate", "that", "these", "proximal", "colon", "tumours,", "which", "often", "also", "show", "mismatch", "repair", "deficiency,", "are", "more", "likely", "to", "harbour", "CTNNB1", "mutations.", "This", "was", "also", "found", "in", "a", "study", "of", "microsatellite", "instable", "colorectal", "tumours", "[26].", "The", "WAVE", "screening", "technique", "has", "not", "been", "used", "previously", "for", "formalin-fixed,", "paraffin-embedded", "tissue,", "and", "therefore", "it", "seems", "plausible", "that", "samples", "harbouring", "a", "CTNNB1", "mutation", "have", "escaped", "detection.", "However,", "all", "58", "hMLH1", "APC", ",", "CTNNB1", "and", "K-ras", "genes", "as", "well", "as", "expression", "of", "the", "hMLH1", "protein", "in", "tumour", "tissue", "of", "656", "sporadic", "colorectal", "cancer", "cases", "were", "investigated.", "The", "occurrence", "of", "mutations", "in", "the", "CTNNB1", "gene,", "which", "codes", "for", "β-catenin,", "was", "rare:", "only", "five", "of", "464", "tumours", "analysed", "were", "found", "to", "have", "a", "mutation", "at", "one", "of", "the", "phosphorylation", "sites", "in", "exon", "3.", "Truncating", "mutations", "in", "APC", "and", "activating", "mutations", "in", "K-ras", "appeared", "to", "occur", "at", "similar", "frequencies.", "Although", "tumours", "harbouring", "both", "mutations", "were", "relatively", "rare,", "mutations", "in", "APC", "and", "K-ras", "seemed", "to", "occur", "co-dependently.", "Nine", "percent", "of", "all", "tumours", "(58/656)", "lacked", "hMLH1", "expression,", "and", "in", "these", "tumours", "almost", "no", "APC", "or", "K-ras", "mutations", "was", "detected.", "Patients", "harbouring", "a", "tumour", "with", "absent", "hMLH1", "expression", "were", "older,", "more", "often", "women,", "more", "often", "had", "proximal", "colon", "tumours", "that", "showed", "poorer", "differentiation", "when", "compared", "to", "patients", "who", "harboured", "a", "tumour", "with", "an", "APC", "and/or", "K-ras", "mutation." ]

[ 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 21, 21, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0, 0 ]

[ "Conclusions" ]

[ 0 ]