Basic Study
Copyright ©The Author(s) 2015. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Jul 14, 2015; 21(26): 8052-8060
Published online Jul 14, 2015. doi: 10.3748/wjg.v21.i26.8052
Conjugation of toll-like receptor-7 agonist to gastric cancer antigen MG7-Ag exerts antitumor effects
Xiao-Dong Wang, Ning-Ning Gao, Yu-Wen Diao, Yu Liu, Dong Gao, Wang Li, Yan-Yan Wan, Jing-Jing Zhong, Guang-Yi Jin
Xiao-Dong Wang, Ning-Ning Gao, Yu-Wen Diao, Yu Liu, Dong Gao, Wang Li, Yan-Yan Wan, Jing-Jing Zhong, Guang-Yi Jin, School of Medicine, Shenzhen University, Shenzhen 518060, Guangdong Province, China
Xiao-Dong Wang, Ning-Ning Gao, Yu-Wen Diao, Yu Liu, Dong Gao, Wang Li, Yan-Yan Wan, Jing-Jing Zhong, Guang-Yi Jin, Shenzhen Engineering Laboratory of Synthetic Biology, Shenzhen 518060, Guangdong Province, China
Yu-Wen Diao, National-Regional Key Technology Engineering Laboratory for Medical Ultrasound, School of Medicine, Shenzhen University, Shenzhen 518060, Guangdong Province, China
Yu Liu, Key Laboratory of Optoelectronic Devices and Systems of Ministry of Education and Guangdong Province, College of Optoelectronic Engineering, Shenzhen University, Shenzhen 518060, Guangdong Province, China
Author contributions: Wang XD, Gao NN, Diao YW, Liu Y and Jin GY designed the research; Wang XD, Gao NN, Gao D, Li W, Wan YY and Zhong JJ performed the research; Wang XD, Gao NN, Diao YW and Liu Y analyzed the data; and Wang XD wrote the paper.
Supported by National Natural Science Foundation of China, No. 81202396 and No. 81273374; and grants from the Science Foundation of Shenzhen, No. JCYJ20130326112757843.
Institutional animal care and use committee statement: All procedures involving animals were reviewed and approved by the Institutional Animal Care and Use Committee of Shenzhen University, China.
Conflict-of-interest statement: The authors declare that there are no conflicts of interest related to this study.
Data sharing statement: Technical appendix, statistical code, and dataset available from the corresponding author at gyjin@szu.edu.cn. No additional data are available.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Correspondence to: Guang-Yi Jin, PhD, Professor of Medicinal Chemistry, School of Medicine, Shenzhen University, No. 3688 Nanhai Avenue, Shenzhen 518060, Guangdong Province, China. gyjin@szu.edu.cn
Telephone: +86-755-86670303 Fax: +86-755-86671906
Received: December 26, 2014
Peer-review started: December 27, 2014
First decision: January 22, 2015
Revised: April 1, 2015
Accepted: May 20, 2015
Article in press: May 21, 2015
Published online: July 14, 2015
Processing time: 199 Days and 23 Hours
Abstract

AIM: To investigate the effects of our tumor vaccines on reversing immune tolerance and generating therapeutic response.

METHODS: Vaccines were synthesized by solid phase using an Fmoc strategy, where a small molecule toll-like receptor-7 agonist (T7) was conjugated to a monoclonal gastric cancer 7 antigen mono-epitope (T7-MG1) or tri-epitope (T7-MG3). Cytokines were measured in both mouse bone marrow dendritic cells and mouse spleen lymphocytes after exposed to the vaccines. BALB/c mice were intraperitoneally immunized with the vaccines every 2 wk for a total of three times, and then subcutaneously challenged with Ehrlich ascites carcinoma (EAC) cells. Three weeks later, the mice were killed, and the tumors were surgically removed and weighed. Serum samples were collected from the mice, and antibody titers were determined by ELISA using an alkaline phosphate-conjugated detection antibody for total IgG. Antibody-dependent cell-mediated cytotoxicity was detected by the lactate dehydrogenase method using natural killer cells as effectors and antibody-labeled EAC cells as targets. Cytotoxic T lymphocyte activities were also detected by the lactate dehydrogenase method using lymphocytes as effectors and EAC cells as targets.

RESULTS: Vaccines were successfully synthesized and validated by analytical high performance liquid chromatography and electrospray mass spectrometry, including T7, T7-MG1, and T7-MG3. Rapid inductions of tumor necrosis factor-α and interleukin-12 in bone marrow dendritic cells and interferon γ and interleukin-12 in lymphocytes occurred in vitro after T7, T7-MG1, and T7-MG3 treatment. Immunization with T7-MG3 reduced the EAC tumor burden in BALB/c mice to 62.64% ± 5.55% compared with PBS control (P < 0.01). Six or nine weeks after the first immunization, the monoclonal gastric cancer 7 antigen antibody increased significantly in the T7-MG3 group compared with the PBS control (P < 0.01). As for antibody-dependent cell-mediated cytotoxicity, antisera obtained by immunization with T7-MG3 were able to markedly enhance cell lysis compared to PBS control (31.58% ± 2.94% vs 18.02% ± 2.26%; P < 0.01). As for cytotoxic T lymphocytes, T7-MG3 exhibited obviously greater cytotoxicity compared with PBS control (40.92% ± 4.38% vs 16.29% ± 1.90%; P < 0.01).

CONCLUSION: A successful method is confirmed for the design of gastric cancer vaccines by chemical conjugation of T7 and multi-repeat-epitope of monoclonal gastric cancer 7 antigen.

Keywords: Gastric cancer; Immunotherapy; Monoclonal gastric cancer 7 antigen; Toll-like receptor-7; Vaccine

Core tip: Immunization with toll-like receptor-7 agonist conjugated with a monoclonal gastric cancer 7 antigen tri-epitope was efficacious in reversing tolerance and generating a therapeutic response in Ehrlich ascites carcinoma tumor-bearing mice. This occurred by enhancing the specific humoral and cellular immunity, which were displayed as higher antibody titers, antibody-dependent cell-mediated cytotoxicity, and cytotoxic T lymphocyte activity.