P03.15 Detection of 1p19q co-deletion in oligodendrogliomas with droplet digital PCR

摘要

>Introduction: Complete 1p19q co-deletion is a molecular marker with favorable prognostic and predictive value in oligodendrogliomas (ODG) and is together with mutations in IDH1/2 the most abundant alteration. Moreover, the new WHO classification of tumors of the central nervous system emphasizes the importance of molecular testing for diagnostic purposes. The most commonly used methods for detection of 1p19q co-deletion are FISH, MLPA or microsatellite analysis. All are burdened with some flaws, for instance susceptibility to subjective evaluation, complicated analysis or necessity of parallel testing of blood samples. We propose a new method which uses droplet digital PCR (ddPCR) technology characterized by very high sensitivity and specificity. >Materials and Methods: Tumors from Linköping University Hospital, by the neuro-pathologist diagnosed as being of oligodendroglial differentiation, were analyzed. Using the ddPCR approach and copy number variation analysis in regard to a reference gene with stable 2 copies, we analyzed 8 selected genes distributed along the chromosomal arms 1p and 19q (TP73, CDKN2C, FUBP1, LRIG2 on 1p and NLRP9, ERCC1, CEBPA, CCNE1 on 19q). For complete loss of both arms, deletion of all gene markers had to be stated. Sanger sequencing and pyrosequencing were used for IDH1/2 mutation detection. Survival was calculated from date of first surgery. >Results: Eighty four ODGs collected between 2000 and 2013 and histologically reviewed as ODG grade II-IV (28 ODGs grade II, 50- grade III, 5- grade IV and 1 oligoastrocytoma) were analyzed for 1p19q co-deletion and IDH1/2 mutation. 55% of patients were male and the median age was 57 years. In 35 samples (42%) complete co-deletion of chromosomal arms 1p and 19q was found. In 20 cases (24%) no chromosomal arm was completely deleted, however local deletions of genes were present. Deletion of 1p only was observed in one sample and only 19q loss was found in 28 samples (33%). In 47 (56%) tumors mutations in IDH1 or IDH2 were detected and 24 of the 35 (69%) patients with co-deleted tumor were IDH mutated. Kaplan-Meier analysis showed significantly longer survival for patients with co-deletion (77 vs. 41 months, p=0.000) and those with mutated IDH (92 vs. 29 months). In the multivariate analysis co-deletion, IDH mutation and age were independent factors influencing survival (HR=0.502, CI=0.255–0.987, p=0.046, HR=0.184, CI=0.084–0.404, p=0.000 and HR=1.069, CI=1.043–1.096, p=0.000, respectively). >Conclusions: The ddPCR based method provides a good alternative for 1p19q co-deletion detection. It can be successfully used on FFPE samples, it is relatively fast, with short hands-on time requirement, simple analysis, high sensitivity and specificity. We could confirm that survival of ODG patients depends on co-deletion of 1p/19q, presence of IDH mutations and age at diagnosis, as has been shown previously.