Apatinib prevents natural killer cell dysfunction to enhance the efficacy of anti-PD-1 immunotherapy in hepatocellular carcinoma


Apatinib, a selective vascular endothelial growth factor receptor 2-tyrosine kinase inhibitor, has demonstrated activity against a wide range of solid tumors, including advanced hepatocellular carcinoma (HCC). Preclinical and preliminary clinical results have confirmed the synergistic antitumor effects of apatinib in combination with anti-programmed death-1 (PD-1) blockade. However, the immunologic mechanism of this combination therapy remains unclear. Here, using a syngeneic HCC mouse model, we demonstrated that treatment with apatinib resulted in attenuation of tumor growth and increased tumor vessel normalization. Moreover, our results indicated that natural killer cells, but not CD4+ or CD8+ T cells mediated the therapeutic efficacy of apatinib in HCC mouse models. As expected, the combined administration of apatinib and anti-PD-1 antibody into tumor-bearing mice generated potent immune responses resulting in a remarkable reduction of tumor growth. Furthermore, increased interferon-γ and decreased tumor necrosis factor-α and interleukin-6 levels were observed, suggesting the potential benefits of combination therapy with PD-1 blockade and apatinib in HCC.

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Fig. 1: The therapeutic efficacy of apatinib in Hep1-6 C57BL/6 mouse model.
Fig. 2: The effects of apatinib on angiogenesis and vascular normalization in vivo.
Fig. 3: The changes in tumor-infiltrating immune cells after apatinib therapy in vivo.
Fig. 4: Functional changes of NK cells in TILs after apatinib therapy in vivo.
Fig. 5: The therapeutic efficacy of apatinib combined with PD-1 blockade.
Fig. 6: The changes in cytokine and PD-L1 expression after combination therapy with apatinib and PD-1 blockade.


  1. 1.

    Villanueva A. Hepatocellular carcinoma. N Engl J Med. 2019;380:1450–62.

    CAS  Article  Google Scholar 

  2. 2.

    Sartorius K, Sartorius B, Aldous C, Govender PS, Madiba TE. Global and country underestimation of hepatocellular carcinoma (HCC) in 2012 and its implications. Cancer Epidemiol. 2015;39:284–90.

    CAS  Article  Google Scholar 

  3. 3.

    El-Serag HB. Hepatocellular carcinoma. N Engl J Med. 2011;365:1118–27.

    CAS  Article  Google Scholar 

  4. 4.

    Park JW, Chen M, Colombo M, Roberts LR, Schwartz M, Chen PJ, et al. Global patterns of hepatocellular carcinoma management from diagnosis to death: the BRIDGE Study. Liver Int. 2015;35:2155–66.

    Article  Google Scholar 

  5. 5.

    Llovet JM, Ricci S, Mazzaferro V, Hilgard P, Gane E, Blanc JF, et al. Sorafenib in advanced hepatocellular carcinoma. N Engl J Med. 2008;359:378–90.

    CAS  Article  Google Scholar 

  6. 6.

    Kudo M, Finn RS, Qin S, Han KH, Ikeda K, Piscaglia F, et al. Lenvatinib versus sorafenib in first-line treatment of patients with unresectable hepatocellular carcinoma: a randomised phase 3 non-inferiority trial. Lancet. 2018;391:1163–73.

    CAS  Article  Google Scholar 

  7. 7.

    Cheng AL, Kang YK, Lin DY, Park JW, Kudo M, Qin S, et al. Sunitinib versus sorafenib in advanced hepatocellular cancer: results of a randomized phase III trial. J Clin Oncol. 2013;31:4067–75.

    CAS  Article  Google Scholar 

  8. 8.

    Llovet JM, Decaens T, Raoul JL, Boucher E, Kudo M, Chang C, et al. Brivanib in patients with advanced hepatocellular carcinoma who were intolerant to sorafenib or for whom sorafenib failed: results from the randomized phase III BRISK-PS study. J Clin Oncol. 2013;31:3509–16.

    CAS  Article  Google Scholar 

  9. 9.

    Zhu AX, Rosmorduc O, Evans TR, Ross PJ, Santoro A, Carrilho FJ, et al. SEARCH: a phase III, randomized, double-blind, placebo-controlled trial of sorafenib plus erlotinib in patients with advanced hepatocellular carcinoma. J Clin Oncol. 2015;33:559–66.

    CAS  Article  Google Scholar 

  10. 10.

    Prieto J, Melero I, Sangro B. Immunological landscape and immunotherapy of hepatocellular carcinoma. Nat Rev Gastroenterol Hepatol. 2015;12:681–700.

    CAS  Article  Google Scholar 

  11. 11.

    Nishida N, Kudo M. Role of immune checkpoint blockade in the treatment for human hepatocellular carcinoma. Dig Dis. 2017;35:618–22.

    Article  Google Scholar 

  12. 12.

    El-Khoueiry AB, Sangro B, Yau T, Crocenzi TS, Kudo M, Hsu C, et al. Nivolumab in patients with advanced hepatocellular carcinoma (CheckMate 040): an open-label, non-comparative, phase 1/2 dose escalation and expansion trial. Lancet. 2017;389:2492–502.

    CAS  Article  Google Scholar 

  13. 13.

    Nishida N, Kudo M. Immune checkpoint blockade for the treatment of human hepatocellular carcinoma. Hepatol Res. 2018;48:622–34.

    CAS  Article  Google Scholar 

  14. 14.

    Roviello G, Ravelli A, Polom K, Petrioli R, Marano L, Marrelli D, et al. Apatinib: a novel receptor tyrosine kinase inhibitor for the treatment of gastric cancer. Cancer Lett. 2016;372:187–91.

    CAS  Article  Google Scholar 

  15. 15.

    Scott AJ, Messersmith WA, Jimeno A. Apatinib: a promising oral antiangiogenic agent in the treatment of multiple solid tumors. Drugs Today (Barc). 2015;51:223–9.

    CAS  Article  Google Scholar 

  16. 16.

    Xue JM, Astere M, Zhong MX, Lin H, Shen J, Zhu YX. Efficacy and safety of apatinib treatment for gastric cancer, hepatocellular carcinoma and non-small cell lung cancer: a meta-analysis. Onco Targets Ther. 2018;11:6119–28.

    CAS  Article  Google Scholar 

  17. 17.

    Liang Q, Kong L, Du Y, Zhu X, Tian J. Antitumorigenic and antiangiogenic efficacy of apatinib in liver cancer evaluated by multimodality molecular imaging. Exp Mol Med. 2019;51:76.

    Article  Google Scholar 

  18. 18.

    Xu J, Zhang Y, Jia R, Yue C, Chang L, Liu R, et al. Anti-PD-1 antibody SHR-1210 combined with apatinib for advanced hepatocellular carcinoma, gastric, or esophagogastric junction cancer: an open-label, dose escalation and expansion study. Clin Cancer Res. 2019;25:515–23.

    Article  Google Scholar 

  19. 19.

    Motz GT, Coukos G. The parallel lives of angiogenesis and immunosuppression: cancer and other tales. Nat Rev Immunol. 2011;11:702–11.

    CAS  Article  Google Scholar 

  20. 20.

    Hodi FS, Lawrence D, Lezcano C, Wu X, Zhou J, Sasada T, et al. Bevacizumab plus ipilimumab in patients with metastatic melanoma. Cancer Immunol Res. 2014;2:632–42.

    CAS  Article  Google Scholar 

  21. 21.

    Wallin JJ, Bendell JC, Funke R, Sznol M, Korski K, Jones S, et al. Atezolizumab in combination with bevacizumab enhances antigen-specific T-cell migration in metastatic renal cell carcinoma. Nat Commun. 2016;7:12624.

    CAS  Article  Google Scholar 

  22. 22.

    Liu K, Zhang X, Xu W, Chen J, Yu J, Gamble JR, et al. Targeting the vasculature in hepatocellular carcinoma treatment: starving versus normalizing blood supply. Clin Transl Gastroenterol. 2017;8:e98.

    CAS  Article  Google Scholar 

  23. 23.

    Apte RS, Chen DS, Ferrara N. VEGF in signaling and disease: beyond discovery and development. Cell. 2019;176:1248–64.

    CAS  Article  Google Scholar 

  24. 24.

    De Palma M, Biziato D, Petrova TV. Microenvironmental regulation of tumour angiogenesis. Nat Rev Cancer. 2017;17:457–74.

    Article  Google Scholar 

  25. 25.

    Landskron G, De la Fuente M, Thuwajit P, Thuwajit C, Hermoso MA. Chronic inflammation and cytokines in the tumor microenvironment. J Immunol Res. 2014;2014:149185.

    Article  Google Scholar 

  26. 26.

    Jain RK. Antiangiogenesis strategies revisited: from starving tumors to alleviating hypoxia. Cancer Cell. 2014;26:605–22.

    CAS  Article  Google Scholar 

  27. 27.

    Huang Y, Yuan J, Righi E, Kamoun WS, Ancukiewicz M, Nezivar J, et al. Vascular normalizing doses of antiangiogenic treatment reprogram the immunosuppressive tumor microenvironment and enhance immunotherapy. Proc Natl Acad Sci USA. 2012;109:17561–6.

    CAS  Article  Google Scholar 

  28. 28.

    Zhao S, Ren S, Jiang T, Zhu B, Li X, Zhao C, et al. Low-dose apatinib optimizes tumor microenvironment and potentiates antitumor effect of PD-1/PD-L1 blockade in lung cancer. Cancer Immunol Res. 2019;7:630–43.

    CAS  PubMed  Google Scholar 

  29. 29.

    Sun C, Sun H, Zhang C, Tian Z. NK cell receptor imbalance and NK cell dysfunction in HBV infection and hepatocellular carcinoma. Cell Mol Immunol. 2015;12:292–302.

    CAS  Article  Google Scholar 

  30. 30.

    Krasnova Y, Putz EM, Smyth MJ, Souza-Fonseca-Guimaraes F. Bench to bedside: NK cells and control of metastasis. Clin Immunol. 2017;177:50–9.

    CAS  Article  Google Scholar 

  31. 31.

    Schmittnaegel M, Rigamonti N, Kadioglu E, Cassara A, Wyser Rmili C, Kiialainen A, et al. Dual angiopoietin-2 and VEGFA inhibition elicits antitumor immunity that is enhanced by PD-1 checkpoint blockade. Sci Transl Med. 2017;9:eaak9670.

    Article  Google Scholar 

  32. 32.

    Liu XD, Hoang A, Zhou L, Kalra S, Yetil A, Sun M, et al. Resistance to antiangiogenic therapy is associated with an immunosuppressive tumor microenvironment in metastatic renal cell carcinoma. Cancer Immunol Res. 2015;3:1017–29.

    CAS  Article  Google Scholar 

  33. 33.

    Liang L, Wen Y, Hu R, Wang L, Xia Y, Hu C, et al. Safety and efficacy of PD-1 blockade-activated multiple antigen-specific cellular therapy alone or in combination with apatinib in patients with advanced solid tumors: a pooled analysis of two prospective trials. Cancer Immunol Immunother. 2019;68:1467–77.

    CAS  Article  Google Scholar 

  34. 34.

    Zhang QF, Yin WW, Xia Y, Yi YY, He QF, Wang X, et al. Liver-infiltrating CD11b(-)CD27(-) NK subsets account for NK-cell dysfunction in patients with hepatocellular carcinoma and are associated with tumor progression. Cell Mol Immunol. 2017;14:819–29.

    CAS  Article  Google Scholar 

  35. 35.

    Allen E, Jabouille A, Rivera LB, Lodewijckx I, Missiaen R, Steri V, et al. Combined antiangiogenic and anti-PD-L1 therapy stimulates tumor immunity through HEV formation. Sci Transl Med. 2017;9:eaak9679.

    Article  Google Scholar 

  36. 36.

    Hsu J, Hodgins JJ, Marathe M, Nicolai CJ, Bourgeois-Daigneault MC, Trevino TN, et al. Contribution of NK cells to immunotherapy mediated by PD-1/PD-L1 blockade. J Clin Investig. 2018;128:4654–68.

    Article  Google Scholar 

  37. 37.

    Gupta KK, Khan MA, Singh SK. Constitutive inflammatory cytokine storm: a major threat to human health. J Interferon Cytokine Res. 2020;40:19–23.

    CAS  Article  Google Scholar 

  38. 38.

    Mace TA, Shakya R, Pitarresi JR, Swanson B, McQuinn CW, Loftus S, et al. IL-6 and PD-L1 antibody blockade combination therapy reduces tumour progression in murine models of pancreatic cancer. Gut. 2018;67:320–32.

    CAS  Article  Google Scholar 

  39. 39.

    Bertrand F, Montfort A, Marcheteau E, Imbert C, Gilhodes J, Filleron T, et al. TNFalpha blockade overcomes resistance to anti-PD-1 in experimental melanoma. Nat Commun. 2017;8:2256.

    Article  Google Scholar 

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This work was supported in part by grants from the National Natural Science Foundation of China (No. 81703786; to YY) and the Tianjin Science and Technology Committee (No. 18JCZDJC36700; to ZP).

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Correspondence to Zhanyu Pan.

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Yang, Y., Wang, C., Sun, H. et al. Apatinib prevents natural killer cell dysfunction to enhance the efficacy of anti-PD-1 immunotherapy in hepatocellular carcinoma. Cancer Gene Ther (2020). https://doi.org/10.1038/s41417-020-0186-7

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