Methods and compositions for detecting cancers associated with methylation of hMLH1 promoter DNA

A method for monitoring over time a cancer associated with methylation of hMLH1 promoter DNA in a subject;

• comprising
• a) assaying for the presence of methylated hMLH1 promoter DNA in a bodily fluid taken from the subject at a first time, said bodily sample selected from the group consisting of blood, serum, plasma, a blood-derived fraction, stool, and a colonic effluent, and
• b) assaying for the presence of methylated hMLH1 promoter DNA in a corresponding bodily fluid taken from the subject at a later time;
• wherein absence of methylated hMLH1 promoter DNA in the bodily fluid taken at a later time and the presence of methylated hMLH1 promoter DNA in the bodily fluid taken at the first time is indicative of cancer regression and wherein presence of methylated hMLH1 promter DNA in the bodily fluid taken at a later time and absence of methylated hMLH1 promoter DNA in the bodily fluid taken at the first time is indicative of cancer progression.

A primer set for detecting the presence of a cancer associated with methylation of hMHL1 promoter DNA in a subject;

• wherein the primer set is a first primer set and comprises a forward primer and the reverse primer for amplifying a bisulfite converted template in a CpG rich region of the Watson strand of hMLH1 promoter DNA,
• wherein said forward primer of the first primer set comprises a sequence which is substantially identical to a first target sequence in the bisulfite converted template in the Watson strand, said first target sequence being at least 8 nucleotides in length, said first target sequence being located at the 5′ end of the bisulfite-converted template;
• and
• wherein said reverse primer of set first primer set comprises a sequence which is identical to the reverse complement of a second target sequence in the bisulfite converted template in the Watson strand, said second target sequence being at least 8 nucleoitdes in length, said second target sequence being located at the 3′end of the bisulfite-converted template;
• or
• wherein the primer set is a second primer set and comprises a forward primer and the reverse primer for amplifying a bisulfite converted template in a CpG rich region of the Crick strand of hMLH1 promoter DNA,
• wherein said forward primer of the second primer set comprises a sequence which is substantially identical to a first target sequence in the bisulfite converted template of the Crick strand, said first target sequence being at least 8 nucleotides in length, said first target sequence being located at the 5′ end of the bisulfite-converted template;
• and
• wherein said reverse primer of set second primer set comprises a sequence which is identical to the reverse complement of a second target sequence in the bisulfite converted template in the Crick strand, said second target sequence being at least 8 nucleoitdes in length, said second target sequence being located at the 3′end of the bisulfite-converted template