Structural and functional characterization of β2 -glycoprotein I domain 1 in anti-melanoma cell migration.

Shr-Jeng Jim Leu, Tzong-Yi Lee, Shu-Wei Cheng, Meng-Ying Tsai, Yu-Shan Lin, Tzeon-Jye Chiou, Kai-Yao Huang, An-Na Chiang

Cancer science 2019 Jun

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We previously found that circulating β2 -glycoprotein I inhibits human endothelial cell migration, proliferation, and angiogenesis by diverse mechanisms. In the present study, we investigated the antitumor activities of β2 -glycoprotein I using structure-function analysis and mapped the critical region within the β2 -glycoprotein I peptide sequence that mediates anticancer effects. We constructed recombinant cDNA and purified different β2 -glycoprotein I polypeptide domains using a baculovirus expression system. We found that purified β2 -glycoprotein I, as well as recombinant β2 -glycoprotein I full-length (D12345), polypeptide domains I-IV (D1234), and polypeptide domain I (D1) significantly inhibited melanoma cell migration, proliferation and invasion. Western blot analyses were used to determine the dysregulated expression of proteins essential for intracellular signaling pathways in B16-F10 treated with β2 -glycoprotein I and variant recombinant polypeptides. Using a melanoma mouse model, we found that D1 polypeptide showed stronger potency in suppressing tumor growth. Structural analysis showed that fragments A and B within domain I would be the critical regions responsible for antitumor activity. Annexin A2 was identified as the counterpart molecule for β2 -glycoprotein I by immunofluorescence and coimmunoprecipitation assays. Interaction between specific amino acids of β2 -glycoprotein I D1 and annexin A2 was later evaluated by the molecular docking approach. Moreover, five amino acid residues were selected from fragments A and B for functional evaluation using site-directed mutagenesis, and P11A, M42A, and I55P mutations were shown to disrupt the anti-melanoma cell migration ability of β2 -glycoprotein I. This is the first study to show the therapeutic potential of β2 -glycoprotein I D1 in the treatment of melanoma progression. © 2019 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

Citation

Shr-Jeng Jim Leu, Tzong-Yi Lee, Shu-Wei Cheng, Meng-Ying Tsai, Yu-Shan Lin, Tzeon-Jye Chiou, Kai-Yao Huang, An-Na Chiang. Structural and functional characterization of β2 -glycoprotein I domain 1 in anti-melanoma cell migration. Cancer science. 2019 Jun;110(6):1974-1986