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Mechanism of action differences in the antitumor effects of transmembrane and secretory tumor necrosis factor-alpha in vitro and in vivo

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Abstract

The proinflammatory cytokine tumor necrosis factor-alpha (TNFα) exists naturally in two forms, a 26 kDa transmembrane form (TM-TNFα), and a 17 kDa secretory form (S-TNFα). The biological roles for each of these forms of TNFα in tumor killing have not been completely elucidated. Therefore, in this study, three different recombinant retroviral vectors, wild-type TNFα, solely secretable TNFα mutant, and uncleavable transmembrane TNFα mutant, were constructed by molecular techniques and stably transfected into a murine hepatic carcinoma cell line (H22). TNFα, either secreted in cell culture supernatants by secretable TNFα mutant- or wild-type TNFα-producing tumor cells, or as a treansmembrane form expressed on the cell surface of uncleavable TNFα mutant- or wild-type TNFα-synthesizing tumor cells, was demonstrated to be cytotoxic against the TNF sensitive L929 cell line. The H22 cells transfected with the three different forms of TNFα were shown to kill parental H22 cells in an in vitro cytotoxicity assay [effect/target (E/T) ratio-dependent manner], and their maximal killing rates were ~38–43% at E/T ratio of 5:1. The injection of total 2.5×105 mixed cells containing transfected and parental H22 tumor cells at different ratios into syngeneic mice resulted in the inhibition of tumor growth with a maximal inhibition rates of ~57~72% at E/T ratio of 5:1. A transient weight loss was found in mice bearing solely secretable TNFα mutant producing tumors, whereas no obvious side effects were seen in mice bearing uncleavable TNFα mutant or wild-type TNFα expressing tumors. Finally, we demonstrate that tumors secreting S-TNFα promoted the subsequent infiltration of CD4+ T cells, and to a lesser extent CD8+ T cells, to the tumor site. The TM-TNFα expressing tumors up-regulated Fas (CD95) expression and inhibited the expression of tumor metastasis associated molecule CD44v3. These results suggest that S-TNFα and TM-TNFα kill cancer cells in vivo through different mechanisms of action. We conclude that the non-secreted form of TNFα may be an ideal candidate for cancer gene therapy due to its therapeutic potential and lowered side effect profile.

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Acknowledgments

The authors wish to thank Mary Reiche for careful review of the manuscript and Professor Xuetao Cao for his generous gift of the PA317 and H22 cell lines. This project was supported by the National Natural Science Foundation of China (30200257), Key Project of Science and Technology from the Education Ministry of China (02136), the Hi-tech Research and Development Program of China (2001AA215431) and National Key Basic Research Program of China from the Ministry of Science and Technology of the People’s Republic of China (2001CB510008), and The Leukemia and Lymphoma Society of the United States of America (6249-05, to M.Z.).

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Authors and Affiliations

  1. Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, 430030, Wuhan, Hubei, China

    Qingfen Li, Li Li, Wenfang Shi, Xiaodan Jiang, Yong Xu, Feili Gong & Zhuoya Li

  2. Division of Pediatric Hematology/Oncology, Emory University, Atlanta, GA, 30322, USA

    Muxiang Zhou

  3. Department of Dermatology, University of Colorado Health Sciences Center at Fitzsimons, Aurora, CO, 80045, USA

    Carl K. Edwards III

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  1. Qingfen Li
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  9. Zhuoya Li
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Correspondence to Zhuoya Li.

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Li, Q., Li, L., Shi, W. et al. Mechanism of action differences in the antitumor effects of transmembrane and secretory tumor necrosis factor-alpha in vitro and in vivo. Cancer Immunol Immunother 55, 1470–1479 (2006). https://doi.org/10.1007/s00262-006-0150-x

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  • DOI: https://doi.org/10.1007/s00262-006-0150-x

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