Abstract 329: Establishment of three-dimensional primary tumor cell culture method and novel drug sensitivity test
Authors: Goji Hiroshi , Shimomura Manami , Uemura Yasushi , Nakatsura Tetsuya , Rahman M.Mamunur , Itoh Manabu 
Published: August 2015

Abstract
Background
Predicting sensitivity to anticancer drug and selecting the most effective drug may help to enhance the therapeutic value, leading to improve patients’ QOL. For these purposes, primary cancer cell from patient specimen is being studied on anticancer drug sensitivity test. In general cell culture method, primary cancer cells are difficult to culture because of fibroblast cells are increasing predominantly than cancer cells. For resolving the problem, collagen-gel-embedding culture method has been developed. However this culture method remain having some problems, for example, complexity of the cell culture preparation from small cancer tissue like a biopsy sample, low efficiency of primary cancer cells and difficulty in multi-drug sensitivity test.
Therefore we developed a three dimensional (3D) cancer cell culture kit that could conduct a primary tumor cell culture and an evaluation of the anticancer drug sensitivity in one-step process after dissociated small cancer tissue. For 3D cell culture we have used NanoCulture Plate (NCP) which is scaffold based 3D culture system and easily form spheroids than other culture systems. Spheroids microenvironment are closely resemble to actual tumor biology, and are therefore more relevant for drug sensitivity. In this report, we carried out a sensitivity test in a lung adenocarcinoma specimen harboring EGFR gene exon19 deletion mutation indicating susceptibility to gefitinib to confirm the efficacy of the system.
Method
Dissociation of cancer tissue and primary cancer cell culture were performed by using 3D cancer cell culture kit and NCP. Partially digested cancer fragments from surgical specimens were seeded and cultured for 3 days. The spheroids were composed of both CK-18 expressing cancer cells and α-SMA expressing fibroblast cells, then spheroids were treated with gefitinib. After 4 days of drug treatment, the susceptibility of the cells to gefitinib was investigated an index of viability by ATP assay. Detection of EGFR mutations was performed by SRL Inc., using real-time PCR that combines scorpion technology and Amplification Refractory Mutation System (ARMS).
Result and conclusion
We found that primary cells with EGFR exon19 deletion were highly susceptible to gefitinib, whereas the cells with intact EGFR were not. These finding indicated the feasibility of 3D cancer cell culture kit and NanoCulture Plate for primary cell culture and screening of effective anticancer drugs. The cell lines generated through this procedure may help to advance our knowledge of certain forms of lung cancer and may also be useful for developing patient-specific anti-cancer drug screening procedures. To clarify the causal relationship between the mutation type and susceptibility, additional experiment and optimization of this kit are currently underway.
Citation Format: Hiroshi Goji, Manami Shimomura, Yasushi Uemura, Tetsuya Nakatsura, M.Mamunur Rahman, Manabu Itoh. Establishment of three-dimensional primary tumor cell culture method and novel drug sensitivity test. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 329. doi:10.1158/1538-7445.AM2015-329