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Molecular pathology & classification

Session: Poster Session

PATH-27 - Droplet digital PCR enhances detection of hotspot mutations in circulating tumor DNA from cerebrospinal fluid of patients with high-grade glioma

Friday, November 17, 2023
7:30 PM – 9:30 PM PT
Location: Exhibit Hall A/B
Non-CME
Molecular profiling of circulating tumor DNA (ctDNA) from cerebrospinal fluid (CSF) has the potential to enhance diagnosis, prognosis, and longitudinal monitoring for glioma patients through a minimally invasive approach. We previously detected somatic mutations from gliomas in CSF samples using MSK-IMPACT™, a 505-gene FDA-authorized next-generation DNA sequencing panel, and detected tumor-derived mutations in 50% of samples of patients with high-grade gliomas (HGG). To investigate whether we could enhance sensitivity of CSF ctDNA profiling and biomarker detection, we tested MSK-IMPACT™ pre-capture NGS residual DNA libraries from 43 CSF ctDNA samples by droplet digital PCR (ddPCR) for detection of key recurrent alterations in IDH1, BRAF, TERT, H3F3A, and TP53 using previously validated probes. Residual libraries contained a median of 18.0ng of DNA library (range, 0.0153-18.0ng). Of the 7 samples positive for IDH1 R132H, TERT C228T, or TERT C250T mutations in MSK-IMPACT™, we found that ddPCR accurately detected all mutations (7/7, 100%), confirming sensitivity of ddPCR in CSF ctDNA. We also tested 36 mutation-negative samples by MSK-IMPACT™ for the corresponding undetected mutations, of which 7 were tested for two mutations. Overall, ddPCR detected 41.9% (18/43) of previously undetected mutations. Specifically, we found that ddPCR rescued hotspot mutations such as BRAF V600E (2/3, 66.7%), IDH1 R132G/H (3/8, 37.5%), TERT C228T (11/26, 42.3%), and TERT C250T (2/4, 50.0%). Collectively, our results suggest that a combined-modality approach involving MSK-IMPACT™ and ddPCR enables comprehensive assessment of limited CSF ctDNA samples with high sensitivity and provides a minimally invasive methodology to molecularly profiling glioma.