Abstract 3832: Cancer tissue-derived spheroid, CTOS, for evaluation of drug response from individual patient tumor samples.
Authors: Inoue Masahiro , Endo Hiroko , Ohue Masayuki , Nishimura Kazuo , Imamura Fumio , Okami Jiro , Okuyama Hiroaki 
Published: August 2014

Abstract
Primary culture of cancer cells can be an excellent platform for investigating the biology of cancer and predicting chemosensitivity for individual patients, yet has been hampered by technical difficulties. We established a method of primary culture for human colorectal cancer: cancer tissue-originated spheroid (CTOS) method. The principle of the method is to maintain cell-cell contact throughout preparation and culture; never dissociating the samples into single cells. The clusters of cancer cells turn out to spheroids, CTOSs, within a short period. CTOSs of colorectal cancer consisted of highly purified cancer cells, and they were prepared with high efficiency. We applied the CTOS method to non-small cell lung cancer and urothelial cancer.
For non-small cell lung cancer, we used 125 surgical specimens and 18 pleural effusions. The success rate of CTOS preparation from surgical specimens was 80.0%. CTOSs were subjected to sensitivity assay and pathway activation assay for the EGFR tyrosine kinase inhibitor (TKI) erlotinib. The sensitivity differed among individual patient CTOSs. The EGFR mutation status and expression levels corresponded with erlotinib sensitivity, confirming previous clinical findings. Next, we examined urothelial cancer. The success rate of CTOS preparation from the non muscle-invasive urothelial cancer was 90.7%, whereas that from the muscle-invasive urothelial cancer was 68.2%. Total development rate was 84.2%.
Using these CTOSs, we investigated the effects of growth factors on the growth of CTOS in vitro. Among the growth factors examined, including EGF, IGF-1, basic FGF, and activin A, we found that heregulin (HRG)/neuregulin1, a ligand of HER3, potently induced CTOS growth in both lung and urothelial cancers. HRG promoted the growth of primary CTOSs in 10 out of 13 (71.4%) cases of lung cancer, and in 2 out of 4 (50%) cases of urothelial cancer. Along with the growth, HRG most potently stimulated the phosphorylation of AKT. In lung cancer CTOSs, a neutralizing antibody against HER3 inhibited the growth and the phosphorylation of HER3 and AKT.
In all three tumor types examined, CTOSs formed xenograft tumors in immunodeficient mice, and the tumors retained the features of the parental tumors. As CTOS can be cryopreserved, we are accumulating stocks of CTOSs derived from patient samples as well as the CTOSs from xenograft tumors. CTOS experiments can be shuttled between in vitro and in vivo.
Thus, the CTOS method enables us to obtain primary colorectal, lung, and urothelial tumor cells of high viability and purity. CTOS could be a new platform for studying lung cancer biology, and be useful to select patients for molecular targeting drugs.
Citation Format: Masahiro Inoue, Hiroko Endo, Masayuki Ohue, Kazuo Nishimura, Fumio Imamura, Jiro Okami, Hiroaki Okuyama. Cancer tissue-derived spheroid, CTOS, for evaluation of drug response from individual patient tumor samples. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3832. doi:10.1158/1538-7445.AM2013-3832