Dna methylation markers associated with the cpg island methylator phenotype (cimp) in human colorectal cancer

Abstract

Particular aspects confirm the existence of a CpG island methylator phenotype (CIMP) in colorectal cancer, and provide novel validated DNA methylation markers associated with CIMP. Additional aspects provide novel methods and compositions for: determining CIMP status in colorectal cancers, determining the relationship between CIMP status and other molecular features of the cancers (e.g., BRAF mutation, KRAS mutation and MSI status); determining the relationship between CIMP status and other variables (e.g., age, sex, tumor location, family history, race, country of origin, tumor characteristics (including, tumor type, tumor grade, invasive margin characteristics, lymphocyte infiltration characteristics, direct spread, lymph node spread, venous spread and type of residual adjacent polyp, if present)); and determining, between subgroups defined by CIMP status and BRAF mutations, effects of selected risk factors (e.g., body mass index, smoking history, alcohol intake, dietary folate intake, folate metabolic enzyme polymorphisms and history of hormonal use).

1 .- 21 . (canceled) 
     
     
         22 . A treated nucleic acid derived from genomic SEQ ID NOS:128-141, 114-127 and 100-113, wherein the treatment is suitable to convert at least one unmethylated cytosine base of the genomic DNA sequence to uracil or another base that is detectably dissimilar to cytosine in terms of hybridization. 
     
     
         23 . A nucleic acid, comprising at least 16 contiguous nucleotides of a treated genomic DNA sequence selected from the group consisting of SEQ ID NOS:170-197, 226-253, 142-169 and 198-225, and sequences complementary thereto, wherein the treatment is suitable to convert at least one unmethylated cytosine base of the genomic DNA sequence to uracil or another base that is detectably dissimilar to cytosine in terms of hybridization. 
     
     
         24 . A nucleic acid, comprising at least 50 contiguous nucleotides of a DNA sequence selected from the group consisting of SEQ ID NOS:170-197, 226-253, 142-169 and 198-225, and sequences complementary thereto. 
     
     
         25 . The nucleic acid of any one of  claims 22  to  24 , wherein the contiguous base sequence comprises at least one CpG, TpG or CpA dinucleotide sequence. 
     
     
         26 . A nucleic acid, comprising at least 16 contiguous nucleotides of nucleic acid sequence selected from the group consisting of SEQ ID NOS:128-141, 114-127 and 100-113, SEQ ID NOS:170-197, 226-253, 142-169 and 198-225 and sequences complementary thereto as a diagnostic means. 
     
     
         27 . A kit suitable for performing the method according to claim  3  comprising a) a plurality of oligonucleotides or polynucleotides able to hybridise under stringent or moderately stringent conditions to the transcription products of at least one gene or genomic sequence selected from the group consisting of BCL2, BDNF, CACNA1G, CALCA, CRABP1, DLEC1, GATA3, HOXA1, IGF2, KL, NEUROG1, NR3C1, RUNX3, SOCS1 (including all transcript variants thereof), and SEQ ID NOS:128-141, 114-127 and 100-113; (b) a container suitable for containing the oligonucleotides or polynucleotides and a biological sample of the patient comprising the transcription products wherein the oligonucleotides or polynucleotides can hybridise under stringent or moderately stringent conditions to the transcription products, (c) means to detect the hybridisation of (b); and optionally, (d) instructions for use and interpretation of the kit results. 
     
     
         28 . A kit suitable for performing the method according to claim  6 , comprising (a) a means for detecting polypeptides of at least one gene or genomic sequence selected from the group consisting of BCL2, BDNF, CACNA1G, CALCA, CRABP1, DLEC1, GATA3, HOXA1, IGF2, KL, NEUROG1, NR3C1, RUNX3, SOCS1 (including all transcript variants thereof); (b) a container suitable for containing the said means and the biological sample of the patient comprising the polypeptides wherein the means can form complexes with the polypeptides; (c) a means to detect the complexes of (b). 
     
     
         29 . A kit suitable for performing the method according to claim  9 , comprising (a) a bisulfite reagent; (b) a container suitable for containing the said bisulfite reagent and the biological sample of the patient; (c) at least one set of oligonucleotides containing two oligonucleotides whose sequences in each case are identical, are complementary, or hybridize under stringent or highly stringent conditions to a 9 or more preferably 18 base long segment of a sequence selected from SEQ ID NOS:170-197, 226-253, 142-169 and 198-225. 
     
     
         30 . A kit suitable for performing the method according to claim  9 , comprising (a) a methylation sensitive restriction enzyme reagent; (b) a container suitable for containing the said reagent and the biological sample of the patient; (c) at least one set of oligonucleotides one or a plurality of nucleic acids or peptide nucleic acids which are identical, are complementary, or hybridize under stringent or highly stringent conditions to an at least 9 base long segment of a sequence selected from SEQ ID NOS:128-141, 114-127 and 100-113; and optionally (d) instructions for use and interpretation of the kit results. 
     
     
         31 .- 33 . (canceled)