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Test Code: LABIDH
Description: The IDH1 and IDH2 genes encode isocitrate dehydrogenases 1 and 2, respectively. These enzymes catalyze the conversion of isocitrate to alpha-ketoglutarate as part of the citric acid cycle during normal metabolism. Highly recurrent hotspot mutations occur in these genes in the majority of gliomas as well as in a significant percentage of myeloid neoplasms, and less commonly in other tumors. The 2016 WHO Classification of Tumors of the Central Nervous System stratifies gliomas into subtypes based on their IDH mutation status 1. Hotspot mutations affecting R132 in IDH1 and R140 and R172 in IDH2 shift the enzyme's activity, losing the ability to convert isocitrate to alpha-ketoglutarate and gaining the capacity to convert alpha-ketoglutarate (the final product of the wild-type enzyme) into 2-hydroxyglutarate2-3. This metabolite appears to favor oncogenesis by effecting broad changes in promoter methylation, which impairs cell differentiation4 and may dysregulate other genes relevant to tumor formation, such as PDGFRA5. IDH1/2 mutations are associated with improved overall survival in gliomas6.

This assay employs the SNaPshot™ methodology, which uses a single base extension step with a labeled ddNTP to detect mutations in IDH1 and IDH2. Analysis of mutation sites in exon 4 of both IDH1 and IDH2 in DNA extracted from FFPE tissue containing a minimum of 10% tumor is accomplished by first amplifying these exons with multiplexed PCR primers. The amplicons are then treated to remove primers and single nucleotides, and reamplified to identify specific nucleotides in a "SNaPshot" reaction. The chemistry of the SNaPshot Multiplex kit is based on the dideoxy single-base extension of an unlabeled oligonucleotide extension primer. Each primer binds to a complementary template in the presence of fluorescently labeled ddNTPs and DNA polymerase. The polymerase extends the primer by one nucleotide, adding a single labeled ddNTP to its 3' end.

1. 22.1 International Academy for Research on Cancer, Louis, D.N., et al. WHO Classification of Tumours of the Central Nervous System. Lyon: IARC Press, 2016.

2. Dang, L. et al. Cancer-associated IDH1 mutations produce 2-hydroxyglutarate. Nature. 2009; 462(7274):739-44.

3. Ward, P.S. et al. The common feature of leukemia-associated IDH1 and IDH2 mutations in a neomorphic enzyme activity converting alpha-ketoglutarate to 2-hydroxyglutarate. Cancer Cell. 2010; 17(3):225-34.

4. Figueroa, M.E. et al. Leukemic IDH1 and IDH2 mutations result in a hypermethylation phenotype, disrupt TET2 function, and impair hematopoietic differentiation. Cancer Cell. 2010; 18(6):553-67.

5. Flavahan, W.A. et al. Insulator dysfunction and oncogene activation in IDH mutant glioma. Nature. 2016; 529(7584):110-4.

6. Yan, H. et al. IDH1 and IDH2 mutations in gliomas. N Engl J Med. 2009; 360(8):765-73.


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