Method of determining dihydropyrimidine dehydrogenase gene expression
The present invention relates to prognostic methods which are useful in medicine, particularly cancer chemotherapy. The object of the invention to provide a method for assessing Dihydropyrimidine dehydrogenase (DPD) expression levels in tissues and prognosticate the probable resistance of a patient's tumor to treatment with 5-FU based therapies by examination of the amount of DPD mRNA in a patient's tumor cells and comparing it to a predetermined threshold expression level. More specifically, the invention provides to oligonucleotide primer pairs DPD3A and DPD3B and methods comprising their use for detecting levels of Dihydropyrimidine dehydrogenase (DPD) mRNA.
1. An isolated and purified oligonucleotide consisting of the sequence of SEQ ID NO: 1; wherein said isolated and purified oligonucleotide is capable of amplifying a portion of the 5′ untranslated region and Exon 1 of a Dihydropyrimidine Dehydrogenase (DPD) mRNA isolated from fixed and paraffin embedded (FPE) tumor tissue when used with SEQ ID NO: 2.
2. An isolated and purified oligonucleotide consisting of the sequence of SEQ ID NO: 2; wherein said isolated and purified oligonucleotide is capable of amplifying a portion of the 5′ untranslated region and Exon 1 of a Dihydropyrimidine Dehydrogenase (DPD) mRNA isolated from fixed and paraffin embedded (FPE) tumor tissue when used with SEQ ID NO: 1.
3. An isolated and purified oligonucleotide consisting of the sequence of SEQ ID NO: 7; wherein said isolated and purified oligonucleotide is capable of amplifying a portion of Exon 6 of a Dihydropyrimidine Dehydrogenase (DPD) mRNA isolated from fixed and paraffin embedded (FPE) tumor tissue when used with SEQ ID NO: 8.
4. An isolated and purified oligonucleotide consisting of the sequence of SEQ ID NO: 8; wherein said isolated and purified oligonucleotide is capable of amplifying a portion of Exon 6 of a Dihydropyrimidine Dehydrogenase (DPD) mRNA isolated from fixed and paraffin embedded (FPE) tumor tissue when used with SEQ ID NO: 7.
5. A kit for detecting expression of a Dihydropyrimidine Dehydrogenase (DPD) gene in a tissue comprising at least two oligonucleotide primers, said two oligonucleotide primers consisting of SEQ ID NO: 1; capable of amplifying a portion of a Dihydropyrimidine Dehydrogenase (DPD) mRNA isolated from fixed and paraffin embedded (FPE) tumor tissue when used with SEQ ID NO:2; and SEQ ID NO: 2; capable of amplifying a portion of a Dihydropyrimidine Dehydrogenase (DPD) mRNA isolated from fixed and paraffin embedded (FPE) tumor tissue when used with SEQ ID NO: 1.
6. A kit for detecting expression of a Dihydropyrimidine Dehydrogenase (DPD) gene in a tissue comprising at least two oligonucleotide primers, said two oligonucleotide primers consisting of SEQ ID NO:7 capable of amplifying portion of a Dihydropyrimidine Dehydrogenase (DPD) mRNA isolated from fixed and paraffin embedded (FPE) tumor tissue when used with SEQ ID NO: 8; and SEQ ID NO: 8; capable of amplifying a portion of a Dihydropyrimidine Dehydrogenase (DPD) mRNA isolated from fixed and paraffin embedded (FPE) tumor tissue when used with SEQ ID NO: 7.