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:124 and 138, wherein the treatment converts 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 to provide the treated nucleic acid comprising the at least one converted base, and wherein the treated nucleic acid is not derived from nucleotides 942-1,719 of SEQ ID NO:124, nucleotides 41-63, 278-321, 325-347, 373-1150, and 1209-1734 of SEQ ID NO:162, nucleotides 358-883 of SEQ ID NO:163, nucleotides 281-1058 and 1117-1642 of SEQ ID NO:190, and nucleotides 879-929, 1273-1798, and 1857-2634 of SEQ ID NO:191. 
     
     
         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:162, 163, 190, 191, 218, 219, 246, and 247, and sequences complementary thereto, wherein the treatment converts 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, to provide the nucleic acid comprising the at least 16 contiguous nucleotides having the at least one converted base, wherein the treated genomic DNA sequence does not include nucleotides 942-1,719 of SEQ ID NO:163, nucleotides 41-63, 278-321, 325-347, 373-1150, and 1209-1734 of SEQ ID NO:162, nucleotides 358-883 of SEQ ID NO:163, nucleotides 281-1058 and 1117-1642 of SEQ ID NO:190, nucleotides 879-929, 1273-1798, and 1857-2634 of SEQ ID NO:191, and nucleotides 176-210, 887-929, and 1872-1909 of SEQ ID NO:247. 
     
     
         24 . A nucleic acid, comprising at least 50 contiguous nucleotides of a treated genomic DNA sequence selected from the group consisting of SEQ ID Seattle NOS:162, 163, 190, 191, 218, 219, 246, and 247, and sequences complementary thereto, wherein the treatment converts at least one unmethylated cytosine base of a genomic DNA sequence to uracil or another base that is detectably dissimilar to cytosine in terms of hybridization to provide the nucleic acid comprising the at least 50 contiguous nucleotides having the at least one converted base, and wherein the treated genomic DNA sequence does not include nucleotides 358-883 and 942-1,719 of SEQ ID NO:163, nucleotides 373-1150 and 1209-1734 of SEQ ID NO:162, nucleotides 281-1058 and 1117-1642 of SEQ ID NO:190, and nucleotides 879-929, 1273-1798, and 1857-2634 of SEQ ID NO:191. 
     
     
         25 . The nucleic acid of any one of  claims 22  to  24 , wherein the treated contiguous base sequence comprises at least one CpG, TpG, or CpA dinucleotide sequence. 
     
     
         26 . A nucleic acid, comprising at least 16 contiguous nucleotides of a nucleic acid sequence selected from the group consisting of SEQ ID NOS:162, 163, 190, 191, 218, 219, 246, and 247 and sequences complementary thereto as a diagnostic means, wherein the treated genomic DNA sequence does not include nucleotides 358-883 and 942-1,719 of SEQ ID NO:163, nucleotides 41-63, 278-321, 325-347, 373-1150, and 1209-1734 of SEQ ID NO:162, nucleotides 281-1058 and 1117-1642 of SEQ ID NO:190, nucleotides 879-929, 1273-1798, and 1857-2634 of SEQ ID NO:191, and nucleotides 176-210, 887-929, and 1872-1909 of SEQ ID NO:247. 
     
     
         27 - 28 . (canceled) 
     
     
         29 . A kit comprising:
 a bisulfite reagent;   a container suitable for containing the bisulfite reagent and a biological sample of a patient; and   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 an at least 9 or an at least 18 base-long contiguous segment of a treated genomic Seattle sequence selected from the group consisting of SEQ ID NOS:162, 163, 190, 191, 218, 219, 246, and 247, wherein the treatment is suitable to convert at least one unmethylated cytosine base of a genomic DNA sequence to uracil or another base that is detectably dissimilar to cytosine in terms of hybridization to provide the at least 9 or the at least 18 contiguous segment of the treated genomic sequence having the at least one converted base, and wherein the treated genomic DNA sequence does not include nucleotides 358-883 and 942-1,719 of SEQ ID NO:163, nucleotides 41-63, 278-321, 325-347, 373-1150, and 1209-1734 of SEQ ID NO:162, nucleotides 281-1058 and 1117-1642 of SEQ ID NO:190, nucleotides 879-929, 1273-1798, and 1857-2634 of SEQ ID NO:191, and nucleotides 176-210, 887-929, and 1872-1909 of SEQ ID NO:247.   
     
     
         30 - 33 . (canceled)