Abstract 212: True2 duplex sequencing reveals a diverse landscape of unique somatic mutations beyond the surgical margins of high- and low-grade diffuse gliomas
Authors: Underhill Hunter , Karsy Michael , Davidson Christian , Hellwig Sabine , Stevenson Samuel , Vincenti Sydney , Dean Charlie , Harrison Brion , Goold Eric , Bronner Mary , Colman Howard , Jensen Randy 
Published: April 2023

Introduction: High- and low-grade diffuse gliomas commonly recur regardless of therapy. Here, we sought to detect and profile molecular evidence of disease in nearby brain tissue absent of tumor by standard-of-care clinical imaging.

Methods: Twenty-two patients presenting to the University of Utah with radiographic evidence of a new high- to low-grade intrinsic brain tumor were enrolled after providing written informed consent. Using intraoperative stereotactic guidance, tissue was acquired from bulky tumor (36 samples) and beyond the surgical margins in peritumoral edema (29 samples). All samples underwent somatic mutation discovery using a custom-designed 114 kb panel (115 genes) and True2 duplex sequencing. True2 is a duplex adapter-based technology developed and empirically validated to achieve &gt;95% sensitivity for an allele frequency ≥0.1% at a false positive rate of &lt;1 per 100 kb panel positions.

Results: Somatic mutations were detected in all samples regardless of radiographic location. The mean number of mutations was similar between bulky tumor and peritumoral edema (5.6±2.4 vs. 5.1±2.9 mutations/sample, respectively; P=0.48). Although the mean allele frequency of variants was higher in tumor compared to edema (16.0±21.2% vs. 11.2±17.4%, respectively; P=0.017), there was a strong overlap in the allele frequency ranges (tumor: 0.07% to 90.3% vs. edema: 0.10% to 75.6%). Notably, one-third of all the somatic mutations detected had an allele frequency &lt;0.5%. Common diagnostic mutations specific to each tumor type were present in both tumor and edema - TERT promoter mutations in glioblastoma and oligodendroglioma; IDH1/2 mutations in astrocytoma and oligodendroglioma. In edema, mutations associated with glioblastoma but unique to a single sample occurred in TP53, EGFR, PTEN, NF1, ATRX, SCN9A, and MTOR at an allele frequency ranging from 0.10% to 20.22%. The edema of astrocytomas similarly harbored unique mutations in TP53, PTEN, and NF1, but mutations in EGFR, ATRX, SCN9A, and MTOR were absent. In contrast to both glioblastoma and astrocytoma, oligodendrogliomas exhibited a diverse collection of unique CIC and PAFAH1B3 mutations in edema at an allele frequency between 0.13% and 3.63%.

Conclusions: The application of True2 technology to conduct a discovery search for very-low frequency somatic mutations found the molecular burden between tumor and adjacent edema to be remarkably similar. Our findings indicate an extensive tumor presence beyond the surgical margins in diffuse gliomas with unique and abundant somatic mutations that may form the foundation of recurrent lesions. Molecular analysis of adjacent tissue identified as radiographically unaffected by bulky tumor to better characterize the mutational landscape of migratory subclones may directly impact personalized medicine for high- and low-grade diffuse glioma patients.

Citation Format: Hunter Underhill, Michael Karsy, Christian Davidson, Sabine Hellwig, Samuel Stevenson, Sydney Vincenti, Charlie Dean, Brion Harrison, Eric Goold, Mary Bronner, Howard Colman, Randy Jensen. True2 duplex sequencing reveals a diverse landscape of unique somatic mutations beyond the surgical margins of high- and low-grade diffuse gliomas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 212.