Abstract
Purpose
Bi-specific antibody (BsAb) is an emerging novel format of antibody. We aimed to develop the natural killer (NK) cell receptor NK group 2, member D (NKG2D)-mediated, immune surveillance system. In this system, the NKG2D ligand MHC class I-related chain A (MICA) was fused with BsAb, which targeted a cluster of differentiation 24 (CD24), a tumor-initiating cell marker that is over-expressed on hepatocellular carcinoma (HCC).
Methods
The Homo MICA extracellular domains (hMICA) were fused to the end of the heavy chain of cG7 with the flexible pentapeptide (Gly-Gly-Gly-Gly-Ser; G4S), which formed the cG7-MICA that was further identified using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS–PAGE) and western blotting (WB). The targeting specificity was characterized using the Surface Plasmon Resonance (SPR) technology and a flow cytometry assay. Furthermore, the design of BsAb cG7-MICA that targeted CD24 and NKG2D was proven to enhance antibody-dependent, cell-mediated cytotoxicity (ADCC) in vitro by the CytoTox 96 Nonradioactive Cytotoxicity assay. Degranulation and a cytokine production assay of NK cells demonstrated that NK cells were activated effectively by cG7-MICA. Further, in HCC-bearing nude mice, the anti-tumor effects of cG7-MICA combined with sorafenib were verified again.
Results
We purified cG7-MICA successfully, and it has a high affinity. In vivo, cG7-MICA recruited NK cells to the tumor site and improved the anti-tumor efficacy of sorafenib. cG7-MICA also activated NK cells to release interferon γ (IFN-γ) and tumor necrosis factor α (TNF-α), and it increased the CD107a expression on the surface of the NK cells in vitro.
Conclusion
NK cells play a major role in the natural, innate immune system, and they have the function of identifying and killing target cells. cG7-MICA remodels the function of MICA molecules to activate NK cells, which provides a possible strategy for HCC-targeting immunotherapy.
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Acknowledgements
Thank Thomas A. Gavin, Professor Emeritus, Cornell University, for help with editing this paper.
Funding
This study was supported by the National Natural Science Foundation of China (NSFC81473125) and the Natural Science Foundation of Jiangsu Province (BK20161459), Jiangsu Province Qinglan Project (2014).
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All experimental procedures were conducted in conformity with institutional guidelines for the care and use of laboratory animals in Yangzhou University, Yangzhou, China, and they conformed to the National Institutes of Regulations for the Administration of Affairs Concerning Experimental Animals.
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Han, Y., Sun, F., Zhang, X. et al. CD24 targeting bi-specific antibody that simultaneously stimulates NKG2D enhances the efficacy of cancer immunotherapy. J Cancer Res Clin Oncol 145, 1179–1190 (2019). https://doi.org/10.1007/s00432-019-02865-8
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DOI: https://doi.org/10.1007/s00432-019-02865-8