Establidhment of highly-sensitive TERT mutation assay in glioma

2020 Fiscal Year Final Research Report

• Project/Area Number: 18K09003
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Review Section: Basic Section 56010:Neurosurgery-related
• Research Institution: Saitama Medical University
• Principal Investigator: ADACHI JUNICHI 埼玉医科大学, 医学部, 准教授 (70291143)
• Co-Investigator(Kenkyū-buntansha): 西川 亮 埼玉医科大学, 医学部, 教授 (90237678)
• Project Period (FY): 2018-04-01 – 2021-03-31
• Keywords: Glioma / TERT / ddPCR

Outline of Final Research Achievements

It is important to accurately detect TERT promoter mutations in glioma. Sanger DNA sequencing is the currently standard method for analyzing TERT mutations. In this report, we described a novel droplet digital PCR (ddPCR) assay to evaluate TERT hot spot mutations in fresh frozen and formalin-fixed paraffin-embedded (FFPE) specimens of glioma and verified the difference in results from the Sanger DNA sequencing results. We obtained the mutant allele fraction for TERT mutations of in a single ddPCR run in all cases, including the microdissected FFPE sections. On the contrary, up to twice the DNA sequences were required from fresh frozen tissue to obtain the results, consistent with ddPCR assay. When FFPE specimens were used, more time was required to evaluate TERT mutations through DNA sequencing. DdPCR is an effective and sensitive assay compared to the conventional standard Sanger DNA sequencing.

Free Research Field

脳神経外科

Academic Significance and Societal Importance of the Research Achievements

ddPCR法はサンプル内にPCR増幅される遺伝子があれば必ず増幅検出されるため、0.01％の変異DNAでも検出が可能な高感度な遺伝子変異検出法であり再現性も高い。特に変異のホットスポットが判明しているTERT遺伝子変異の解析に対しては極めて有用な方法である。