Abstract 5577: Detection of RAS mutations in colorectal cancer patients using DNA from extracellular vesicles
Authors: Kuriyama Sho , Yamada Takeshi , Sonoda Hiromichi , Shinji Seiichi , Matsuda Akihisa , Yonaga Kazuhide , Iwai Takuma , Takeda Kohki , Ueda Koji , Miyasaka Toshimitsu , Kanaka Shintaro , Yoshida Hiroshi 
Published: April 2023

Abstract
Introduction: Precision medicine is becoming increasingly important in cancer treatment, and liquid biopsy (LB) is a core technique. Circulating tumor DNA (ctDNA) is the main target for LB; however, in some patients, its sensitivity is inadequate. Detection of post-operative ctDNA, called Minimal Residual Disease (MRD), indicates cancer recurrence; however, some patients experience recurrent disease without being MRD-positive. To improve sensitivity of ctDNA detection, we focused on exosomes, small vesicles of endosomal origin that contain DNA (exoDNA) derived from cells, including cancer cells. In this study, we assayed KRAS mutations in patients with colorectal cancer using exoDNA, and analyzed the utility of biomarkers in prognosis, using a machine learning algorithm.
Materials and Methods: Patients who underwent curative surgery for colorectal cancer from November 2018 to December 2020 were recruited prospectively. Blood samples were obtained before surgery, and ctDNA was extracted from 1 mL of plasma. Exosomes were isolated from this plasma, followed by exoDNA extraction. KRAS mutations in ctDNA and exoDNA were analyzed using droplet digital PCR. The Random Survival Forest (RSF) algorithm was used for variable importance analysis.
Results: 300 patients (123 with KRAS mutation and 177 without) were included. The median patient age was 71 years (range: 35 - 91). Concentrations of exoDNA were significantly higher in Stage IV patients compared to patients at other stages (p &lt; 0.01). Numbers of exosomes did not differ by stage. KRAS mutations were detected in exoDNA of 56 patients (45.5%) and in ctDNA of 28 patients with KRAS mutation in their tumors (22.7%, p&lt;0.001). Sensitivity of exoDNA was higher than that of ctDNA in Stage II/III patients with KRAS mutations (Stage II; 33.3% vs 14.2%, p=0.02, Stage III 46.7% vs 13.3%, p=0.001). However, sensitivity of cfDNA was almost equal to that of exoDNA in stage IV patients (53.8% vs 50.0%, p = 0.78). Surprisingly, in patients without KRAS mutations in their tumors, KRAS mutations were detected in exoDNA of 24 patients (13.5%) and in ctDNA of one patient (0.5%). Of 103 stage III patients, 31 (30.1%) experienced recurrence. The RSF algorithm showed that the exoDNA concentration is the most important risk factor for recurrence, followed by a lack of adjuvant chemotherapy.
Conclusion: Sensitivity of KRAS mutation detection using exoDNA is excellent. Additionally, the concentration of exoDNA appears to be a superior predictor for recurrence of Stage II/III colorectal cancer after curative surgery.
Citation Format: Sho Kuriyama, Takeshi Yamada, Hiromichi Sonoda, Seiichi Shinji, Akihisa Matsuda, Kazuhide Yonaga, Takuma Iwai, Kohki Takeda, Koji Ueda, Toshimitsu Miyasaka, Shintaro Kanaka, Hiroshi Yoshida. Detection of RAS mutations in colorectal cancer patients using DNA from extracellular vesicles. [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 5577.