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An orally administered DNA vaccine targeting vascular endothelial growth factor receptor-3 inhibits lung carcinoma growth

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Tumor Biology

Abstract

Lung cancer is the leading cause of mortality and 5-year survival rate is very low worldwide. Recent studies show that vascular endothelial growth factor receptor-3 (VEGFR-3) signaling pathway contributes to lung cancer progression. So we hypothesize that an oral DNA vaccine that targets VEGFR-3 carried by attenuated Salmonella enterica serovar typhimurium strain SL3261 has impacts on lung cancer progression. In this study, the oral VEGFR-3-based vaccine-immunized mice showed appreciable inhibition of tumor growth and tumor lymphatic microvessels in lung cancer mice model. Moreover, the oral VEGFR-3-based vaccine-immunized mice showed remarkable increases in both VEGFR-3-specific antibody levels and cytotoxic activity. Furthermore, the oral VEGFR-3-based vaccine-immunized mice showed a significant increase in the levels of T helper type 1 (Th1) cell intracellular cytokine expression (IL-2, IFN-γ, and TNF-α). After inoculation with murine Lewis lung carcinoma (LLC) cells, CD4+ or CD8+ T cell numbers obviously declined in control groups whereas high levels were maintained in the oral VEGFR-3-based vaccine group. These results demonstrated that the oral VEGFR-3-based vaccine could induce specific humoral and cellular immune responses and then significantly inhibit lung carcinoma growth via suppressing lymphangiogenesis.

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Funding

This study was funded by grants from the National Natural Science Foundation of China (No. 81460358, No. 81060177, No. 81460441), the Joint Special Funds for the Department of Science and Technology of Yunnan Province-Kunming Medical University (No. 2012FB067), and the Yunnan Provincial Technology Project of Health (No. 2012WS0041, No. 2014NS024).

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All procedures performed in studies involving animals were in accordance with the ethical standards of the institution or practice at which the studies were conducted.

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    Authors and Affiliations

    1. Yunnan Tumor Institute, The Third Affiliated Hospital of Kunming Medical University (Tumor Hospital of Yunnan Province), 519 Kunzhou Road, Kunming, Yunnan, China

      Yan Chen, Xin Liu, Cong Guo Jin, Yong Chun Zhou, Xiao Qun Chen, Jia Li, Ting Ting Li, Lu Luo & Xi Cai Wang

    2. Yunnan Tumor Molecular Biomarker Research Center, Kunming, Yunnan, China

      Yan Chen, Xin Liu, Cong Guo Jin, Yong Chun Zhou, Xiao Qun Chen, Jia Li, Ting Ting Li, Lu Luo & Xi Cai Wang

    3. Yunnan Zhan Qimin Academician Workstation, Kunming, Yunnan, China

      Yan Chen, Xin Liu, Cong Guo Jin, Yong Chun Zhou, Xiao Qun Chen, Jia Li, Ting Ting Li, Lu Luo & Xi Cai Wang

    4. Princess Margaret Cancer Center and Campbell Family Institute for Cancer Research, University Health Network, Toronto, Ontario, Canada

      Roya Navab

    5. Department of Biomedicine, Aarhus University, Aarhus, Denmark

      Kristine Raaby Jakobsen

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    1. Yan Chen
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    Correspondence to Xi Cai Wang.

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    Yan Chen and Xin Liu contributed equally to this work.

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    Fig. S1

    The recombinant vector pcDNA3.1-VR-3 was successfully constructed, and verified by DNA sequencing and Western blotting. (a) DNA encoding VEGFR-3 extracellular domain was inserted into pcDNA3.1 vector between the restriction sites HindIII and XbaI and confirmed by DNA sequencing. DNA sequencing data showed that the recombinant gene matched the sequence in GenBank. (b) COS-7 cells were transiently transfected with the recombinant pcDNA3.1-VR-3 and the pcDNA3.1 empty vector respectively. Recombinant VEGFR-3 protein expression was confirmed by Western blotting in cell lysates at 24 h and 72 h after transfection. The protein appeared at approximately 180 kD. (GIF 61 kb)

    High resolution image (TIFF 7068 kb)

    Fig. S2

    Attenuated S. typhimurium SL3261 can infect macrophage cell line Ana-1 in vitro, and disseminate from the gut to other parts of body. (a) Fluorescence and optical microscopic images of Ana-1 cells in vitro co-incubated with EGFP-expressing SL3261 or negative control cells (NTC) without any treatment. 24 h after co-incubation, cells were detected by inverted fluorescence microscopy. Both panels, ×200 magnification. (b) C57BL/6 J mice were immunized 3 times at 2-weeks intervals by oral administration of 108 CFU of SL3261 harboring pEGFP-C2 or saline. Mice were sacrificed 4 h, 8 h or 96 h after the last immunization. The percentage of EGFP-expressing cells derived from small intestine, liver and spleen were detected by flow cytometry (n = 3 per time point). Error bars indicate S.D. (GIF 107 kb)

    High resolution image (TIFF 16038 kb)

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    Chen, Y., Liu, X., Jin, C.G. et al. An orally administered DNA vaccine targeting vascular endothelial growth factor receptor-3 inhibits lung carcinoma growth. Tumor Biol. 37, 2395–2404 (2016). https://doi.org/10.1007/s13277-015-4061-3

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