Cancer Immunology, Immunotherapy

, Volume 56, Issue 11, pp 1831–1843 | Cite as

Dendritic cells modified with 6Ckine/IFNγ fusion gene induce specific cytotoxic T lymphocytes in vitro

  • Gang Xue
  • Ran-yi Liu
  • Yan Li
  • Ying Cheng
  • Zhi-hui Liang
  • Jiang-xue Wu
  • Mu-sheng Zeng
  • Fu-zhou Tian
  • Wenlin Huang
Original Article


Backgroud and objective

Dendritic cells play an important role in initiation and regulation of immune responses. Previous studies demonstrated that intratumoral administration of 6Ckine-modified DCs enhanced local and systemic antitumor effects. Herein we report the investigation of the specific CTL responses elicited by adenoviral 6Ckine/IFNγ fusion gene-modified DCs in vitro.


Human monocyte-derived DCs were modified with an adenoviral vector encoding 6Ckine/IFNγ fusion protein (Ad-6Ckine/IFNγ), and then investigated the effect of 6Ckine/IFNγ fusion protein on the maturation, cytokine and chemokine secretion of DCs, and their activities of recruiting and activating T cells in vitro were investigated.


6Ckine/IFNγ fusion protein induced DC maturation characterized with the upregulation of CD83 and CCR7. And it up-regulated the expression of RANTES and IL-12p70, down-regulated that of IL-10 in DCs. Additionally, 6Ckine/IFNγ markedly increased DC’s recruiting ability for naive T cells, benefiting from the enhanced expression of chemokines 6Ckine and RANTES in DCs. Fusion gene-modified DCs significantly promoted the proliferation of autologous T cells, induced Th1 differentiation by upregulating the expression of IL-2 and T-bet in T cells, and increased specific cytotoxicity of CTLs against specific tumor cells, HepG2 or LoVo cells, respectively.


Combining the effects of 6Ckine and IFNγ, Ad-6Ckine/IFNγ modified DCs induced enhanced CTL responses in vitro, which indicated that Ad-6Ckine/IFNγ modified DCs might be used as an adjuvant to trigger an effective antitumor immune response.


Dendritic cells Cytokines Chemokines Tumor immunology Gene therapy 



Recombinant adenovirus carrying 6Ckine/IFNγ fusion gene


Recombinant adenoviruses carrying 6Ckine gene


Recombinant adenoviruses carrying IFNγ gene


Recombinant adenoviruses carrying β-galactosidase gene


Recombinant adenoviruses carrying green fluorescent protein gene


Tumor-associated antigens


DC transfected with Ad-6Ckine


DC transfected with Ad-6Ckine/IFNγ


Non-transfected DC


DC transfected with Ad-IFNγ


DC transfected with Ad-LacZ


Lactate dehydrogenase


National Institute for the Control of Pharmaceutical and Biological Products



This study was supported by the grants from National Basic Research Program of China (No.2004CB518801), Research & Development Fund of Guangdong Provine (No.2003A10902), Natural Science Foundation of Guangdong Province (No.021810), Research & Development Fund of Guangzhou City (No.2004Z3-E4011), and Medical Science Grant from Guangdong Province (No.B2002050). We thank Miao-la Ke, Jie-min Chen, Ying-hui Zhu and Xia Xiao (Sun Yat-sen University) for their technical assistance. We thank Dr. Changyou Wu, Dr. Li-min Zeng, and Dr. Qiang Liu (Sun Yat-sen University) for their review of this article. We also thank Ying-jun Ji (Doublle Bioproduct Inc.) and Yin Duan (Simon Fraser University) for their linguistic help in the preparation of this manuscript.

Supplementary material


  1. 1.
    Lanzavecchia A, Sallusto F (2001) Regulation of T cell immunity by dendritic cells. Cell 106:263–266PubMedCrossRefGoogle Scholar
  2. 2.
    Mihich E (2003) Cellular immunity for cancer chemoimmunotherapy–an overview. Cancer Immunol Immunother 52:661–662PubMedCrossRefGoogle Scholar
  3. 3.
    Bell D, Young JW, Banchereau J (1999) Dendritic cells. Adv Immunol 72:255–324PubMedCrossRefGoogle Scholar
  4. 4.
    Evel-Kabler K, Chen SY (2006) Dendritic cell-based tumor vaccines and antigen presentation attenuators. Mol Ther 13:850–858PubMedCrossRefGoogle Scholar
  5. 5.
    Yang SC, Batra RK, Hillinger S et al (2006) Intrapulmonary administration of CCL21 gene-modified dendritic cells reduces tumor burden in spontaneous murine bronchoalveolar cell carcinoma. Cancer Res 66:3205–3213PubMedCrossRefGoogle Scholar
  6. 6.
    Lundqvist A, Choudhury A, Nagata T et al (2002) Recombinant adenovirus vector activates and protects human monocyte-derived dendritic cells from apoptosis. Hum Gene Ther 13:1541–1549PubMedCrossRefGoogle Scholar
  7. 7.
    Miller G, Lahrs S, Pillarisetty VG et al (2002) Adenovirus infection enhances dendritic cell immunostimulatory properties and induces natural killer and T-cell-mediated tumor protection. Cancer Res 62:5260–5266PubMedGoogle Scholar
  8. 8.
    Kirk CJ, Hartigan-O’Connor D, Mule JJ (2001) The dynamics of the T-cell antitumor response: chemokine-secreting dendritic cells can prime tumor-reactive T cells extranodally. Cancer Res 61:8794–8802PubMedGoogle Scholar
  9. 9.
    Kirk CJ, Hartigan-O’Connor D, Nickoloff BJ et al (2001) T cell-dependent antitumor immunity mediated by secondary lymphoid tissue chemokine: augmentation of dendritic cell-based immunotherapy. Cancer Res 61:2062–2070PubMedGoogle Scholar
  10. 10.
    Baggiolini M, Dewald B, Moser B (1997) Human chemokines: an update. Annu Rev Immunol 15:675–705PubMedCrossRefGoogle Scholar
  11. 11.
    Gunn MD, Tangemann K, Tam C et al (1998) A chemokine expressed in lymphoid high endothelial venules promotes the adhesion and chemotaxis of naive T lymphocytes. Proc Natl Acad Sci USA 95:258–263PubMedCrossRefGoogle Scholar
  12. 12.
    Cyster JG (1999) Chemokines and the homing of dendritic cells to the T cell areas of lymphoid organs. J Exp Med 189:447–450PubMedCrossRefGoogle Scholar
  13. 13.
    Hromas R, Kim CH, Klemsz M et al (1997) Isolation and characterization of Exodus-2, a novel C–C chemokine with a unique 37-amino acid carboxyl-terminal extension. J Immunol 159:2554–2558PubMedGoogle Scholar
  14. 14.
    Sharma S, Stolina M, Luo J et al (2000) Secondary lymphoid tissue chemokine mediates T cell-dependent antitumor responses in vivo. J Immunol 164:4558–4563PubMedGoogle Scholar
  15. 15.
    Sharma S, Stolina M, Zhu L et al (2001) Secondary lymphoid organ chemokine reduces pulmonary tumor burden in spontaneous murine bronchoalveolar cell carcinoma. Cancer Res 61:6406–6412PubMedGoogle Scholar
  16. 16.
    Perussia B (1991) Lymphokine-activated killer cells, natural killer cells and cytokines. Curr Opin Immunol 3:49–55PubMedCrossRefGoogle Scholar
  17. 17.
    Boehm U, Klamp T, Groot M et al (1997) Cellular responses to interferon-gamma. Annu Rev Immunol 15:749–795PubMedCrossRefGoogle Scholar
  18. 18.
    Shankaran V, Ikeda H, Bruce AT et al (2001) IFNgamma and lymphocytes prevent primary tumour development and shape tumour immunogenicity. Nature 410:1107–1111PubMedCrossRefGoogle Scholar
  19. 19.
    Yanagihara K, Seyama T, Watanabe Y (1994) Antitumor potential of interferon-gamma: retroviral expression of mouse interferon-gamma cDNA in two kinds of highly metastatic mouse tumor lines reduces their tumorigenicity. Nat Immun 13:102–112PubMedGoogle Scholar
  20. 20.
    Musiani P, Allione A, Modica A et al (1996) Role of neutrophils and lymphocytes in inhibition of a mouse mammary adenocarcinoma engineered to release IL-2, IL-4, IL-7, IL-10, IFN-alpha, IFN-gamma, and TNF-alpha. Lab Invest 74:146–157PubMedGoogle Scholar
  21. 21.
    Sadanaga N, Nagoshi M, Lederer JA et al (1999) Local secretion of IFN-gamma induces an antitumor response: comparison between T cells plus IL-2 and IFN-gamma transfected tumor cells. J Immunother 22:315–323PubMedCrossRefGoogle Scholar
  22. 22.
    Ito T, Amakawa R, Inaba M et al (2001) Differential regulation of human blood dendritic cell subsets by IFNs. J Immunol 166:2961–2969PubMedGoogle Scholar
  23. 23.
    Akbar SM, Kajino K, Tanimoto K et al (1999) Unique features of dendritic cells in IFN-gamma transgenic mice: relevance to cancer development and therapeutic implications. Biochem Biophys Res Commun 259:294–299PubMedCrossRefGoogle Scholar
  24. 24.
    Koski GK, Lyakh LA, Rice NR (2001) Rapid lipopolysaccharide-induced differentiation of CD14(+) monocytes into CD83(+) dendritic cells is modulated under serum-free conditions by exogenously added IFN-gamma and endogenously produced IL-10. Eur J Immunol 31:3773–3781PubMedCrossRefGoogle Scholar
  25. 25.
    Ohteki T, Fukao T, Suzue K et al (1999) Interleukin 12-dependent interferon gamma production by CD8alpha+ lymphoid dendritic cells. J Exp Med 189:1981–1986PubMedCrossRefGoogle Scholar
  26. 26.
    Mizuguchi H, Kay MA (1998) Efficient construction of a recombinant adenovirus vector by an improved in vitro ligation method. Hum Gene Ther 9:2577–2583PubMedCrossRefGoogle Scholar
  27. 27.
    Liu RY, Wu LZ, Huang BJ et al (2005) Adenoviral expression of a truncated S1 subunit of SARS-CoV spike protein results in specific humoral immune responses against SARS-CoV in rats. Virus Res 112:24–31PubMedCrossRefGoogle Scholar
  28. 28.
    Finney DJ (1964) Statistical method in biological assay, 2nd edn. Hafner Pub. Co., New YorkGoogle Scholar
  29. 29.
    Nishimura N, Nishioka Y, Shinohara T et al (2001) Novel centrifugal method for simple and highly efficient adenovirus-mediated green fluorescence protein gene transduction into human monocyte-derived dendritic cells. J Immunol Methods 253:113–124PubMedCrossRefGoogle Scholar
  30. 30.
    Nishimura N, Nishioka Y, Shinohara T et al (2001) Enhanced efficiency by centrifugal manipulation of adenovirus-mediated interleukin 12 gene transduction into human monocyte-derived dendritic cells. Hum Gene Ther 12:333–346PubMedCrossRefGoogle Scholar
  31. 31.
    Delemarre FG, Simons PJ, de Heer HJ et al (1999) Signs of immaturity of splenic dendritic cells from the autoimmune prone biobreeding rat: consequences for the in vitro expansion of regulator and effector T cells. J Immunol 162:1795–1801PubMedGoogle Scholar
  32. 32.
    Duperrier K, Eljaafari A, Dezutter-Dambuyant C et al (2000) Distinct subsets of dendritic cells resembling dermal DCs can be generated in vitro from monocytes, in the presence of different serum supplements. J Immunol Methods 238:119–131PubMedCrossRefGoogle Scholar
  33. 33.
    Terando A, Roessler B, Mule JJ (2004) Chemokine gene modification of human dendritic cell-based tumor vaccines using a recombinant adenoviral vector. Cancer Gene Ther 11:165–173PubMedCrossRefGoogle Scholar
  34. 34.
    Arthur JF, Butterfield LH, Roth MD et al (1997) A comparison of gene transfer methods in human dendritic cells. Cancer Gene Ther 4:17–25PubMedGoogle Scholar
  35. 35.
    Zhong L, Granelli-Piperno A, Choi Y et al (1999) Recombinant adenovirus is an efficient and non-perturbing genetic vector for human dendritic cells. Eur J Immunol 29:964–972PubMedCrossRefGoogle Scholar
  36. 36.
    Fasbender A, Zabner J, Chillon M et al (1997) Complexes of adenovirus with polycationic polymers and cationic lipids increase the efficiency of gene transfer in vitro and in vivo. J Biol Chem 272:6479–6489PubMedCrossRefGoogle Scholar
  37. 37.
    Dietz AB, Vuk-Pavlovic S (1998) High efficiency adenovirus-mediated gene transfer to human dendritic cells. Blood 91:392–398PubMedGoogle Scholar
  38. 38.
    Aiba S, Tagami H (1998) Dendritic cell activation induced by various stimuli, e.g. exposure to microorganisms, their products, cytokines, and simple chemicals as well as adhesion to extracellular matrix. J Dermatol Sci 20:1–13PubMedCrossRefGoogle Scholar
  39. 39.
    Bayry J, Lacroix-Desmazes S, Kazatchkine MD et al (2005) Modulation of dendritic cell maturation and function by B lymphocytes. J Immunol 175:15–20PubMedGoogle Scholar
  40. 40.
    Pan J, Zhang M, Wang J et al (2004) Interferon-gamma is an autocrine mediator for dendritic cell maturation. Immunol Lett 94:141–151PubMedCrossRefGoogle Scholar
  41. 41.
    Mosca PJ, Hobeika AC, Clay TM et al (2000) A subset of human monocyte-derived dendritic cells expresses high levels of interleukin-12 in response to combined CD40 ligand and interferon-gamma treatment. Blood 96:3499–3504PubMedGoogle Scholar
  42. 42.
    Riedl K, Baratelli F, Batra RK et al (2003) Overexpression of CCL-21/secondary lymphoid tissue chemokine in human dendritic cells augments chemotactic activities for lymphocytes and antigen presenting cells. Mol Cancer 2:35–47PubMedCrossRefGoogle Scholar
  43. 43.
    Morelli AE, Larregina AT, Ganster RW et al (2000) Recombinant adenovirus induces maturation of dendritic cells via an NF-kappaB-dependent pathway. J Virol 74:9617–9628PubMedCrossRefGoogle Scholar
  44. 44.
    Miller G, Lahrs S, Shah AB et al (2003) Optimization of dendritic cell maturation and gene transfer by recombinant adenovirus. Cancer Immunol Immunother 52:347–358PubMedGoogle Scholar
  45. 45.
    Hensley SE, Giles-Davis W, McCoy KC et al (2005) Dendritic cell maturation, but not CD8+ T cell induction, is dependent on type I IFN signaling during vaccination with adenovirus vectors. J Immunol 175:6032–6041PubMedGoogle Scholar
  46. 46.
    Bacon KB, Premack BA, Gardner P et al (1995) Activation of dual T cell signaling pathways by the chemokine RANTES. Science 269:1727–1730PubMedCrossRefGoogle Scholar
  47. 47.
    Dairaghi DJ, Soo KS, Oldham ER et al (1998) RANTES-induced T cell activation correlates with CD3 expression. J Immunol 160:426–433PubMedGoogle Scholar
  48. 48.
    Appay V, Dunbar PR, Cerundolo V et al (2000) RANTES activates antigen-specific cytotoxic T lymphocytes in a mitogen-like manner through cell surface aggregation. Int Immunol 12:1173–1182PubMedCrossRefGoogle Scholar
  49. 49.
    Guo Z, Zhang M, Tang H et al (2005) Fas signal links innate and adaptive immunity by promoting dendritic-cell secretion of CC and CXC chemokines. Blood 106:2033–2041PubMedCrossRefGoogle Scholar
  50. 50.
    Rodolfo M, Colombo MP (1999) Interleukin-12 as an adjuvant for cancer immunotherapy. Methods 19:114–120PubMedCrossRefGoogle Scholar
  51. 51.
    Glimcher LH, Townsend MJ, Sullivan BM et al (2004) Recent developments in the transcriptional regulation of cytolytic effector cells. Nat Rev Immunol 4:900–911PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2007

Authors and Affiliations

  • Gang Xue
    • 3
  • Ran-yi Liu
    • 1
  • Yan Li
    • 1
  • Ying Cheng
    • 4
  • Zhi-hui Liang
    • 1
  • Jiang-xue Wu
    • 1
  • Mu-sheng Zeng
    • 1
  • Fu-zhou Tian
    • 3
  • Wenlin Huang
    • 1
    • 2
  1. 1.State Key Laboratory of Oncology in South China, Room 619, Cancer CenterSun Yat-sen UniversityGuangzhouChina
  2. 2.Beijing Institute of MicrobiologyChinese Academy of SciencesBeijingChina
  3. 3.Department of General SurgeryChengdu Army General HospitalChengduChina
  4. 4.Department of EndocrinologyChengdu Army General HospitalChengduChina

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