A quantitative detection method for glioma IDH1 R132H mutation based on pyrosequencing

Abstract

Objective To develop a method based on pyrosequencing that can be used to accurately and quantitatively detect the IDH1 R132H mutation of gliomas and to evaluate the clinical application of that method. Methods Thirty cases of formalin-fixed and parrffin-embedded (FFPE) samples of gliomas and clinical database were retrospectively obtained from the Chinese Glioma Genome Atlas (CGGA). There are 15 IDH1 wild type and 15 IDH1 R132H mutant samples. The pyrosequencing results were compared with the findings of immunohistochemistry. Results The pyrosequencing results indicated that the percentage of CAT in glioma sample ranged from 10% to 52% in all 30 cases. The percentage of CAT in IDH1 R132H mutant glioma was less than 19%, while the percentage of CAT in IDH1 R132H wildtype glioma was more than 27%. Immunohistochemistry results indicated negative staining in 15 samples and positive staining in the other 15 samples including 8 positive staining and 7 strong positive staining. Further statistical analysis showed that the quantitative IDH1 R132H mutant ratios of pyrosequencing were significantly different between immunohistochemistry-negative and immunohistochemistry-positive glioma FFPE samples, and the former was below 20% (10%-18%), while the latter was above 20% (28%-52%). Conclusion The quantitative IDH1 R132H mutant detection method based on pyrosequencing could accurately detect the IDH1 R132H mutant level in FFPE glioma samples, which seems suitable for routine molecular pathological practice. Key words: Glioma; Pathology, molecular; Mutation; Isocitrate dehydrogenase 1; Pyrosequencing