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Digital Droplet PCR-based Detection of KRAS, NRAS, and BRAF Mutation in Plasma Samples from Patients with Colorectal Cancer.

Journal of Clinical Oncology(2024)

Seoul National University Boramae Medical Center | Gencurix Inc

Cited 0|Views18
Abstract
e15626 Background: Digital droplet PCR (ddPCR) is known for high sensitivity to detect mutated DNA in plasma. The aim of this study was to investigate the concordance of KRAS, NRAS and BRAF mutation detection in patients with colorectal cancers (CRCs) between plasma samples (PSs) and tissue samples (TSs) in pair. Methods: We prospectively collected paired PS and TS from patients with CRC (N = 200) who underwent surgery or biopsy for metastasis at Seoul National University Boramae Medical Center from Mar 2017 to January 2020. The ddPCR was performed using QX200™ Droplet Digital PCR System (Bio-Rad, USA) according to the manufacturer's recommended protocol with PCR kits developed for each gene (Gencurix, South Korea). Results: A total of 187 TSs and PSs were valid for ddPCR. Among these, 111 had results for KRAS, 51 had results for NRAS and 21 had results for BRAF, which were tested through routine clinical practice (RCP) using the pyrosequencing or Sanger sequencing method. The experiment's outcomes were analyzed for KRAS, NRAS, and BRAF, respectively. The analysis initially compared the results of RCP TS and ddPCR TS, and subsequently, intended to compare the mutation detection rates between TS and PS by comparing the results of ddPCR TS and ddPCR PS for FFPE/PS paired specimens. For KRAS, compared to RCP TS, the sensitivity of ddPCR TS was 100%, specificity was 66.7%, and the overall concordance rate was 79.3%. In TS/PS paired specimens, the sensitivity of ddPCR PS for ddPCR TS results was 38.5%, specificity was 97.6%, and the overall concordance rate was 64.7%. For NRAS, compared to RCP TS, the sensitivity of ddPCR TS was 100%, specificity was 98.0%, and the overall concordance rate was 98.0%. In TS/PS paired specimens, the sensitivity of ddPCR PS for ddPCR TS results was 33.3%, specificity was 100%, and the overall concordance rate was 97.4%. For BRAF, sensitivity of ddPCR TS was not available (no RCP positive sample), specificity was 85.7% compared to RCP, corresponded to a concordance rate of 97.0%. The sensitivity of ddPCR PS for ddPCR TS results was 71.4%, specificity was 100%, and the overall concordance rate was 97.4%. The sensitivity of ddPCR PS for ddPCR TS results was 71.4%, specificity was 100%, and the overall concordance rate was 97.4%. Conclusions: The present study showed that ddPCR has high accuracy in detecting KRAS, NRAS and BRAF mutation in plasma samples from patients with CRC. Analysis of PS could be useful in early response assessment and longitudinal disease monitoring for patient with metastatic CRC with specific mutation.
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