Abstract 3422: Label-free circulating tumor cells isolation platform for lung cancer EGFR mutation identification and colorectal cancer with KRAS mutation detection

Abstract

Abstract Circulating tumor cell becomes an urgent issue in personalized cancer treatment studies. AS the liquid biopsy, many previous reports showed the number and the characteristic of CTC are very important for patient prognosis. In cancer therapy, specific gene mutation in primary tumor decides the therapy strategy and drug response. However, the patient who cannot perform an operation or take biopsy would become a risk factor for treatment and affect therapy outcome. Hence, drug treatable related gene mutation detection by CTC is necessary. Platforms for mutation detection in tissue were well established but less useful for CTC. Most of them were lower efficacy for rare cell or DNA. In order to solve this dilemma, scientists use next-generation sequence (NGS) or droplet digital PCR (ddPCR) for gene mutation detection. But, in most clinical situations, we only have to follow-up some specific gene mutation, instead of many mutant types. Thus, we eager to develop a fast and low-cost methodology for drug treatable related gene mutation detection by CTC. In this study, 4mL blood was collected by EDTA vacuum blood collection tube from 37 patients. Whole blood was processed with RBC lyse steps, then added CD45 and glycophorin A antibodies conjugated with the nanoparticle to remove leukocyte and remained RBC. Further, Genomic DNA was extracted from negative selection cell mixture. An ARMS-PCR (Amplification-refractory mutation system PCR) combined with a touchdown PCR program was used for EGFR mutations identification. We enrolled 31 lung cancer patients. Nineteen patients’ CTC were matched with tissue mutation results (14 mutant and 5 wild-types), and 12 unmatched. In those 12 patients, 2 patients with mutant tissue but wild-type CTC due to their blood were drawn at disease-free status after long-term therapy, 4 patients with wild-type tissue but mutant CTC were still had tumor burden after therapy and need to more long-term follow-up. Overall, we have a mutation correlation rate of (19 + 2) / 31 = 68% between CTC and primary tumor. Furthermore, an LNA-PCR (Locked nucleic acid PCR) system was designed for colorectal cancer KRAS codon 12 and codon 13 mutations detection. In this platform, modified RNA was designed for blocked wild-type DNA amplification in PCR. We enrolled 6 patients with primary tumor mutant positive, 5 of 6 (83%) patients were also mutant positive in their CTC sample. In conclusion, an easy CTC isolation platform and PCR design could hugely help cancer patient mutation detection. Only less than 4 hours and 100 USD were needed for drug treatable gene mutation detection by CTC. Citation Format: Hung-Chih Lin, Yu-Lin Yang, Chia-Hsun Hsieh. Label-free circulating tumor cells isolation platform for lung cancer EGFR mutation identification and colorectal cancer with KRAS mutation detection [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3422.