Basic Study
Copyright ©The Author(s) 2017. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Feb 7, 2017; 23(5): 817-829
Published online Feb 7, 2017. doi: 10.3748/wjg.v23.i5.817
Dendritic cells engineered to secrete anti-DcR3 antibody augment cytotoxic T lymphocyte response against pancreatic cancer in vitro
Jiang Chen, Xiao-Zhong Guo, Hong-Yu Li, Jia-Jun Zhao, Wen-Da Xu
Jiang Chen, Xiao-Zhong Guo, Hong-Yu Li, Jia-Jun Zhao, Wen-Da Xu, Department of Gastroenterology, Shenyang General Hospital of PLA, Shenyang 110016, Liaoning Province, China
Author contributions: Chen J performed the majority of experiments and analyzed the data; Li HY and Zhao JJ performed the molecular investigations; Guo XZ designed and coordinated the research; Chen J and Xu WD wrote the paper.
Supported by National Natural Science Foundation of China, No. 81071982.
Institutional review board statement: This study was approved by the Ethics Committee of Shenyang General Hospital of PLA.
Institutional animal care and use committee statement: There was no animal experimentation in this study.
Conflict-of-interest statement: The authors have no conflict of interests to declare.
Data sharing statement: 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: Xiao-Zhong Guo, MD, Department of Gastroenterology, Shenyang General Hospital of PLA, No. 83 Wenhua Road, Shenyang 110016, Liaoning Province, China. guoxiaozhong1962@aliyun.com
Telephone: +86-24-28897603 Fax: +86-24-28851113
Received: September 13, 2016
Peer-review started: September 14, 2016
First decision: October 20, 2016
Revised: November 4, 2016
Accepted: December 21, 2016
Article in press: December 21, 2016
Published online: February 7, 2017
Abstract
AIM

To investigate the enhanced cytotoxic T lymphocyte responses against pancreatic cancer (PC) in vitro induced by dendritic cells (DCs) engineered to secrete anti-DcR3 monoclonal antibody (mAb).

METHODS

DCs, T lymphocytes and primary PC cells were obtained from PC patients. DCs were transfected with a designed humanized anti-DcR3 monoclonal antibody heavy and light chain mRNA and/or total tumor RNA (DC-tumor-anti-DcR3 RNA or DC-total tumor RNA) by using electroporation technology. The identification, concentration and function of anti-DcR3 mAb secreted by DC-tumor-anti-DcR3 RNA were determined by western blotting and enzyme-linked immunosorbent assay. After co-culturing of autologous isolated PC cells with target DCs, the effects of secreting anti-DcR3 mAb on RNA-DCs’ viability and apoptosis were assessed by MTT assay and flow cytometry. Analysis of enhanced antigen-specific immune response against PC induced by anti-DcR3 mAb secreting DCs was performed using a 51Cr releasing test. T cell responses induced by RNA-loaded DCs were analyzed by measuring cytokine levels, including IFN-γ, IL-10, IL4, TNF-α and IL-12.

RESULTS

The anti-DcR3 mAb secreted by DCs reacted with recombinant human DcR3 protein and generated a band with 35 kDa molecular weight. The secreting mAb was transient, peaking at 24 h and becoming undetectable after 72 h. After co-incubation with DC-tumor-anti-DcR3 RNA for designated times, the DcR3 level in the supernatant of autologous PC cells was significantly down-regulated (P < 0.05). DCs secreting anti-DcR3 mAb could improve cell viability and slow down the apoptosis of RNA-loaded DCs, compared with DC-total tumor RNA (P < 0.01). The anti-DcR3 mAb secreted by DC-tumor-anti-DcR3 RNA could enhance the induction of cytotoxic T lymphocytes (CTLs) activity toward RNA-transfected DCs, primary tumor cells, and PC cell lines, compared with CTLs stimulated by DC-total tumor RNA or control group (P < 0.05). Meanwhile, the antigen-specific CTL responses were MHC class I-restricted. The CD4+ T cells and CD8+ T cells incubated with anti-DcR3 mAb secreting DCs could produce extremely higher level IFN-γ and lower level IL4 than those incubated with DC-total tumor RNA or controls (P < 0.01).

CONCLUSION

DCs engineered to secrete anti-DcR3 antibody can augment CTL responses against PC in vitro, and the immune-enhancing effects may be partly due to their capability of down-regulating DC apoptosis and adjusting the Th1/Th2 cytokine network.

Keywords: Dendritic cell, Antibody-encoding RNA, DcR3, Cytotoxic T lymphocyte response, Pancreatic cancer

Core tip: Dendritic cells co-transfection with tumor-associated antigens RNA and humanized anti-DcR3 monoclonal antibody mRNA may augment cytotoxic T lymphocyte responses against pancreatic cancer in vitro. This finding lays a good foundation for further investigation of tumor dendritic cells’ vaccine targeting DcR3 protein against pancreatic cancer.