کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2020372 1542332 2015 6 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Construction, expression, purification, and characterization of a dual-targeting PD-1/VEGF-A fusion protein (P-V)
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی زیست شیمی
پیش نمایش صفحه اول مقاله
Construction, expression, purification, and characterization of a dual-targeting PD-1/VEGF-A fusion protein (P-V)
چکیده انگلیسی
Targeting programmed death-1 (PD-1) is regarded as a novel and promising means for the treatment of many types of solid tumor. In the tumor microenvironment (TME), VEGF expression is dramatically up-regulated, and compounds that neutralize VEGF or block the interaction of VEGF with its receptors exhibit potent antitumor activity, and blocking PD-1 might promote T cell infiltration into TME and significantly enhance local immune activation. Thus, we fused domain II and domain III of kinase-insert domain receptor (KDR), the receptor of VEGF-A, to the Fc side of an anti-PD-1 monoclonal antibody with a (Gly4Ser)3 linker to generate a dual targeting fusion protein. The recombinant plasmid was successfully constructed and the fusion protein was expressed in 293E cells. Protein purification was performed in a single step by using protein A affinity chromatography. The molecular weight of the fusion protein was approximately 220 kDa, and the yield was approximately 2.97 g/L. Specific binding of recombinant protein to PD-1 and VEGF was detected by enzyme-linked immunosorbent assay (ELISA) analysis. Half maximal effective concentration (EC50) values were 0.561 nM for PD-1 and 0.682 nM for VEGF-A; accordingly, half maximal inhibitory concentration (IC50) values were 0.914 nM and 0.583 nM, respectively. Proliferation inhibition assays indicated that the fusion protein could inhibit the growth of human umbilical vein endothelial cells effectively. Taken together, the results indicate that this novel fusion protein can simultaneously target PD-1 and VEGF and may be beneficial for combining anti-angiogenesis with immunotherapeutic approaches for the treatment of patients with cancer.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Protein Expression and Purification - Volume 109, May 2015, Pages 1-6
نویسندگان
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