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Archive > Volume 23 Issue 4 > 2016,23(4):468-475. DOI:10.3872/j.issn.1007-385X.2016.04.002 Prev Next
Prompt Report
Inhibitory effect of DC vaccine sensitized by GnRH/M2 fusion protein on melanoma B16F10 cell xenografts
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Abstract:
Objective:To prepare fusion protein of gonadotropin releasing hormone (GnRH) and M2 (GnRH/M2), and investigate inhibitory effect of DC vaccine sensitized with the fusion protein on melanoma B16F10 cell xenografts. Methods: An expression vector of pET28a-ansB-C-GnRH3-hinge-MVP-M2 plasmid was constructed and transferred into engineering bacteria. Under the induction of lactose, fusion protein of ansB-C-GnRH3-hinge-MVP-M2 was expressed as inclusion body in the transferred engineering bacteria. Then the fusion protein was purified by means of ultrasonic broken, washings and ethanol fractionation precipitation. After purification, protein polypeptide GnRH3-hinge-MVP-M2 was released by acid hydrolysis and isolated with DEAE-52 anion exchange chromatography. DCs were sensitized with the fusion polypeptide to obtain DC vaccine. Mouse models with melanoma B16F10 xenografts were established and divided into cyclophosphamide group (CTX), DC sensitized with GnRH/M2 fusion protein group (GDC), DC sensitized with tumor cell lysate group (BDC), DC sensitized with GnRH/M2 fusion protein plus cyclophosphamide group (GDCTX), DC sensitized with tumor cell lysate plus cyclophosphamide group (BDCTX) and normal saline group (NS), according to different vaccination. Effects of GnRH/M2 vaccine on growth of the xenografts, killing ability of CTL and proliferation of T cells in the mouse models were observed. Results: The pET28a-ansB-C-GnRH3-hinge-MVP-M2 plasmid was successfully constructed and the fusion protein efficiently expressed. Growth of the xenografts in GDC group was more slower than that in NS group obviously (P<0.05), and similar to that in BDC group (P>0.05). Although inhibitory effect on tumor in GDCTX group was further increased, but there not was any significant differences comparing with CTX group (P>0.05). Killing effect on B16F10 cells and proliferation effect on T cells in various treatment groups were significantly better than those in negative control groups (P<0.05 or P<0.01), but differences of them between GDC and BDC groups were not obvious (P>0.05). Conclusion: It was preliminarily proved that the DC vaccine sensitized with the fusion polypeptide GnRH/M2 could effectively inhibit growth of the melanoma B16F10 xenografts in the mouse model.
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Project Supported:
Project supported by the National Natural Science Foundation of China (No．81172973, 81373232), the National Programs of Training Innovative Durg Development Abilities for Undergraduates(No. J1030830), the Priority Academic Program Development of Jiangsu Higher Education Institutions(PAPD), and the Excellent Talent Leading Foundation of Jiangsu Province (No. BK20140029)
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