Cancer Immunology, Immunotherapy

, Volume 67, Issue 7, pp 1091–1103 | Cite as

Role of MDA5 and interferon-I in dendritic cells for T cell expansion by anti-tumor peptide vaccines in mice

  • Hussein Sultan
  • Juan Wu
  • Takumi Kumai
  • Andres M. Salazar
  • Esteban Celis
Original Article


Cytotoxic T lymphocytes (CTLs) are effective components of the immune system capable of destroying tumor cells. Generation of CTLs using peptide vaccines is a practical approach to treat cancer. We have previously described a peptide vaccination strategy that generates vast numbers of endogenous tumor-reactive CTLs after two sequential immunizations (prime-boost) using poly-ICLC adjuvant, which stimulates endosomal toll-like receptor 3 (TLR3) and cytoplasmic melanoma differentiation antigen 5 (MDA5). Dendritic cells (DCs) play an important role not only in antigen presentation but are critical in generating costimulatory cytokines that promote CTL expansion. Poly-ICLC was shown to be more effective than poly-IC in generating type-I interferon (IFN-I) in various DC subsets, through its enhanced ability to escape the endosomal compartment and stimulate MDA5. In our system, IFN-I did not directly function as a T cell costimulatory cytokine, but enhanced CTL expansion through the induction of IL15. With palmitoylated peptide vaccines, CD8α+ DCs were essential for peptide crosspresentation. For vaccine boosts, non-professional antigen-presenting cells were able to present minimal epitope peptides, but DCs were still required for CTL expansions through the production of IFN-I mediated by poly-ICLC. Overall, these results clarify the roles of DCs, TLR3, MDA5, IFN-I and IL15 in the generation of vast and effective antitumor CTL responses using peptide and poly-IC vaccines.


Peptide vaccines Poly-IC Dendritic cells MDA5 Type-I interferon Interleukin-15 



Adoptive cell transfer


Antigen-presenting cell


Bone marrow


Cytotoxic T lymphocyte


Dendritic cell


Diphtheria toxin


Diphtheria toxin receptor


Type-I interferon


Interferon beta


Type-I interferon receptor


IL-2 immune complex




Monoclonal antibody


Melanoma differentiation-associated protein 5


MHC class I


MHC class II


Non-professional antigen-presenting cell






Professional APC


Plasmacytoid DC




Polyinosinic–polycytidylic acid


Poly-IC stabilized with poly-lysine and carboxymethyl cellulose


Poly-IC stabilized with PEI


Signal 3


T cell receptor for antigen


Toll-like receptor 3


Tyrosinase-related protein 1


Wild type


Authorship contributions

HS designed, performed experiments, analyzed the data and helped to write the manuscript. TK and JW performed experiments. Andres Salazar discussed the results and provided reagents. EC designed and analyzed the experiments, and wrote the manuscript.


This work was supported by National Cancer Institute grant R01CA157303, and by start-up funds from Augusta University, Georgia Cancer Center and the Georgia Research Alliance (GRA).

Compliance with ethical standards

Conflict of interest

A. Salazar is President and CEO of Oncovir, Inc. and is developing poly-ICLC (Hiltonol ™) for the clinic. Esteban Celis has filed patent applications based on the use of synthetic peptides and poly-IC combinatorial vaccines. The rights of the patent applications have been transferred to the Moffitt Cancer Center (Tampa, FL). Other authors declare no conflict of interest.

Statement on the welfare of animals

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. All procedures performed in the experiments involving animals were in accordance with the ethical standards of the Augusta University Institutional Animal Care and Use Committee where all the studies were conducted (Protocol No. 2013 − 0598, approved on 11/21/2016).

Human subjects

This article does not contain any studies with humans done by any of the authors.

Supplementary material

262_2018_2164_MOESM1_ESM.pdf (1.1 mb)
Supplementary material 1 (PDF 1117 KB)


  1. 1.
    Rosenberg SA, Dudley ME (2009) Adoptive cell therapy for the treatment of patients with metastatic melanoma. Curr Opin Immunol 21:233–240. CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Cho HI, Barrios K, Lee YR, Linowski AK, Celis E (2013) BiVax: a peptide/poly-IC subunit vaccine that mimics an acute infection elicits vast and effective anti-tumor CD8 T-cell responses. Cancer Immunol Immunother 62:787–799. CrossRefPubMedGoogle Scholar
  3. 3.
    Cho HI, Celis E (2009) Optimized peptide vaccines eliciting extensive CD8 T-cell responses with therapeutic antitumor effects. Cancer Res 69:9012–9019. CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Barrios K, Celis E (2012) TriVax-HPV: an improved peptide-based therapeutic vaccination strategy against human papillomavirus-induced cancers. Cancer Immunol Immunother 61:1307–1317. CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Wang Z, Celis E (2015) STING activator c-di-GMP enhances the anti-tumor effects of peptide vaccines in melanoma-bearing mice. Cancer Immunol Immunother 64:1057–1066. CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Kato H, Takeuchi O, Sato S et al (2006) Differential roles of MDA5 and RIG-I helicases in the recognition of RNA viruses. Nature 441:101–105. CrossRefPubMedGoogle Scholar
  7. 7.
    Dougan SK, Dougan M, Kim J, Turner JA, Ogata S, Cho HI, Jaenisch R, Celis E, Ploegh HL (2013) Transnuclear TRP1-specific CD8 T cells with high or low affinity TCRs show equivalent antitumor activity. Cancer Immunol Res 1:99–111. CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Duran-Struuck R, Dysko RC (2009) Principles of bone marrow transplantation (BMT): providing optimal veterinary and husbandry care to irradiated mice in BMT studies. J Am Assoc Lab Anim Sci 48:11–22PubMedPubMedCentralGoogle Scholar
  9. 9.
    Fuertes Marraco SA, Grosjean F, Duval A et al (2012) Novel murine dendritic cell lines: a powerful auxiliary tool for dendritic cell research. Front Immunol 3:331. CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Winzler C, Rovere P, Rescigno M, Granucci F, Penna G, Adorini L, Zimmermann VS, Davoust J, Ricciardi-Castagnoli P (1997) Maturation stages of mouse dendritic cells in growth factor-dependent long-term cultures. J Exp Med 185:317–328CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Shen Z, Reznikoff G, Dranoff G, Rock KL (1997) Cloned dendritic cells can present exogenous antigens on both MHC class I and class II molecules. J Immunol 158:2723–2730PubMedGoogle Scholar
  12. 12.
    Assudani D, Cho HI, DeVito N, Bradley N, Celis E (2008) In vivo expansion, persistence, and function of peptide vaccine-induced CD8 T cells occur independently of CD4 T cells. Cancer Research 68:9892–9899CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Sultan H, Fesenkova VI, Addis D, Fan AE, Kumai T, Wu J, Salazar AM, Celis E (2017) Designing therapeutic cancer vaccines by mimicking viral infections. Cancer Immunol Immunother. 66:203–213. CrossRefPubMedGoogle Scholar
  14. 14.
    Sultan H, Kumai T, Fesenkova VI, Fan AE, Wu J, Cho HI, Kobayashi H, Harabuchi Y, Celis E (2018) Sustained persistence of IL2 signaling enhances the antitumor effect of peptide vaccines through T-cell expansion and preventing PD-1 inhibition. Cancer Immunol Res doi. Google Scholar
  15. 15.
    Banchereau J, Steinman RM (1998) Dendritic cells and the control of immunity. Nature 392:245–252. CrossRefPubMedGoogle Scholar
  16. 16.
    den Haan JM, Lehar SM, Bevan MJ (2000) CD8(+) but not CD8(−) dendritic cells cross-prime cytotoxic T cells in vivo. J Exp Med 192:1685–1696CrossRefGoogle Scholar
  17. 17.
    Heath WR, Belz GT, Behrens GM et al (2004) Cross-presentation, dendritic cell subsets, and the generation of immunity to cellular antigens. Immunol Rev 199:9–26. CrossRefPubMedGoogle Scholar
  18. 18.
    Jung S, Unutmaz D, Wong P et al. (2002) In vivo depletion of CD11c + dendritic cells abrogates priming of CD8 + T cells by exogenous cell-associated antigens. Immunity 17:211–220CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Kunzmann V, Kretzschmar E, Herrmann T, Wilhelm M (2004) Polyinosinic-polycytidylic acid-mediated stimulation of human gammadelta T cells via CD11c dendritic cell-derived type I interferons. Immunology. 112:369–377. CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Baranek T, Manh TP, Alexandre Y et al (2012) Differential responses of immune cells to type I interferon contribute to host resistance to viral infection. Cell Host Microbe 12:571–584. CrossRefPubMedGoogle Scholar
  21. 21.
    Berard M, Brandt K, Bulfone-Paus S, Tough DF (2003) IL-15 promotes the survival of naive and memory phenotype CD8 + T cells. J Immunol 170:5018–5026CrossRefPubMedGoogle Scholar
  22. 22.
    Mattei F, Schiavoni G, Belardelli F, Tough DF (2001) IL-15 is expressed by dendritic cells in response to type I IFN, double-stranded RNA, or lipopolysaccharide and promotes dendritic cell activation. J Immunol 167:1179–1187CrossRefPubMedGoogle Scholar
  23. 23.
    Yao X, Wu J, Lin M et al (2016) Increased CD40 expression enhances early STING-mediated type I interferon response and host survival in a rodent malaria model. PLoS Pathog 12:e1005930. CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Buhtoiarov IN, Lum H, Berke G, Paulnock DM, Sondel PM, Rakhmilevich AL (2005) CD40 ligation activates murine macrophages via an IFN-gamma-dependent mechanism resulting in tumor cell destruction in vitro. J Immunol 174:6013–6022CrossRefPubMedGoogle Scholar
  25. 25.
    Ellermeier J, Wei J, Duewell P et al (2013) Therapeutic efficacy of bifunctional siRNA combining TGF-beta1 silencing with RIG-I activation in pancreatic cancer. Cancer Res 73:1709–1720. CrossRefPubMedGoogle Scholar
  26. 26.
    McKenna K, Beignon AS, Bhardwaj N (2005) Plasmacytoid dendritic cells: linking innate and adaptive immunity. J Virol 79:17–27. CrossRefPubMedPubMedCentralGoogle Scholar
  27. 27.
    Perussia B, Fanning V, Trinchieri G (1985) A leukocyte subset bearing HLA-DR antigens is responsible for in vitro alpha interferon production in response to viruses. Nat Immun Cell Growth Regul 4:120–137PubMedGoogle Scholar
  28. 28.
    Hildner K, Edelson BT, Purtha WE et al (2008) Batf3 deficiency reveals a critical role for CD8alpha + dendritic cells in cytotoxic T cell immunity. Science 322:1097–1100. CrossRefPubMedPubMedCentralGoogle Scholar
  29. 29.
    Sammons ML, Stephen EL, Levy HB, Baron S, Hilmas DE (1977) Interferon induction in cynomolgus and rhesus monkey after repeated doses of a modified polyriboinosinic-polyribocytidylic acid complex. Antimicrob Agents Chemother. 11:80–83CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Lampson GP, Field AK, Tytell AA, Hilleman MR (1981) Poly I:C/poly-l-lysine: potent inducer of interferons in primates. J Interferon Res 1:539–549CrossRefPubMedGoogle Scholar
  31. 31.
    Talmadge JE, Adams J, Phillips H et al (1985) Immunomodulatory effects in mice of polyinosinic-polycytidylic acid complexed with poly-l-lysine and carboxymethylcellulose. Cancer Res 45:1058–1065PubMedGoogle Scholar
  32. 32.
    Gitlin L, Barchet W, Gilfillan S, Cella M, Beutler B, Flavell RA, Diamond MS, Colonna M (2006) Essential role of mda-5 in type I IFN responses to polyriboinosinic:polyribocytidylic acid and encephalomyocarditis picornavirus. Proc Natl Acad Sci USA 103:8459–8464. CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    McCartney S, Vermi W, Gilfillan S, Cella M, Murphy TL, Schreiber RD, Murphy KM, Colonna M (2009) Distinct and complementary functions of MDA5 and TLR3 in poly(I:C)-mediated activation of mouse NK cells. J Exp Med 206:2967–2976. CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Yamamoto A, Tagawa Y, Yoshimori T, Moriyama Y, Masaki R, Tashiro Y (1998) Bafilomycin A1 prevents maturation of autophagic vacuoles by inhibiting fusion between autophagosomes and lysosomes in rat hepatoma cell line, H-4-II-E cells. Cell Struct Funct 23:33–42CrossRefPubMedGoogle Scholar
  35. 35.
    Zou J, Kawai T, Tsuchida T, Kozaki T, Tanaka H, Shin KS, Kumar H, Akira S (2013) Poly IC triggers a cathepsin D- and IPS-1-dependent pathway to enhance cytokine production and mediate dendritic cell necroptosis. Immunity 38:717–728. CrossRefPubMedGoogle Scholar
  36. 36.
    Aichele P, Unsoeld H, Koschella M, Schweier O, Kalinke U, Vucikuja S (2006) CD8 T cells specific for lymphocytic choriomeningitis virus require type I IFN receptor for clonal expansion. J Immunol 176:4525–4529CrossRefPubMedGoogle Scholar
  37. 37.
    Kolumam GA, Thomas S, Thompson LJ, Sprent J, Murali-Krishna K (2005) Type I interferons act directly on CD8 T cells to allow clonal expansion and memory formation in response to viral infection. J Exp Med 202:637–650. CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Xiao Z, Casey KA, Jameson SC, Curtsinger JM, Mescher MF (2009) Programming for CD8 T cell memory development requires IL-12 or type I IFN. J Immunol 182:2786–2794. CrossRefPubMedPubMedCentralGoogle Scholar
  39. 39.
    Boyman O, Kovar M, Rubinstein MP, Surh CD, Sprent J (2006) Selective stimulation of T cell subsets with antibody-cytokine immune complexes. Science 311:1924–1927. CrossRefPubMedGoogle Scholar
  40. 40.
    Zhang X, Sun S, Hwang I, Tough DF, Sprent J (1998) Potent and selective stimulation of memory-phenotype CD8 + T cells in vivo by IL-15. Immunity 8:591–599CrossRefPubMedGoogle Scholar
  41. 41.
    Zhang M, Yao Z, Dubois S, Ju W, Muller JR, Waldmann TA (2009) Interleukin-15 combined with an anti-CD40 antibody provides enhanced therapeutic efficacy for murine models of colon cancer. Proc Natl Acad Sci USA 106:7513–7518. CrossRefPubMedPubMedCentralGoogle Scholar
  42. 42.
    Sultan H, Trillo-Tinoco J, Rodriguez P, Celis E (2017) Effective antitumor peptide vaccines can induce severe autoimmune pathology. Oncotarget 8:70317–70331. CrossRefPubMedPubMedCentralGoogle Scholar
  43. 43.
    Kumai T, Lee S, Cho HI, Sultan H, Kobayashi H, Harabuchi Y, Celis E (2017) Optimization of peptide vaccines to induce robust antitumor CD4 T-cell responses. Cancer Immunol Res 5:72–83. CrossRefPubMedGoogle Scholar
  44. 44.
    Wang Y, Cella M, Gilfillan S, Colonna M (2010) Cutting edge: polyinosinic:polycytidylic acid boosts the generation of memory CD8 T cells through melanoma differentiation-associated protein 5 expressed in stromal cells. J Immunol 184:2751–2755. CrossRefPubMedGoogle Scholar
  45. 45.
    Pack DW, Hoffman AS, Pun S, Stayton PS (2005) Design and development of polymers for gene delivery. Nat Rev Drug Discov. 4:581–593. CrossRefPubMedGoogle Scholar
  46. 46.
    Sonawane ND, Szoka FC Jr, Verkman AS (2003) Chloride accumulation and swelling in endosomes enhances DNA transfer by polyamine-DNA polyplexes. J Biol Chem 278:44826–44831. CrossRefPubMedGoogle Scholar
  47. 47.
    Wagner E, Zatloukal K, Cotten M, Kirlappos H, Mechtler K, Curiel DT, Birnstiel ML (1992) Coupling of adenovirus to transferrin-polylysine/DNA complexes greatly enhances receptor-mediated gene delivery and expression of transfected genes. Proc Natl Acad Sci USA 89:6099–7103CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Cancer Immunology, Immunotherapy and Tolerance Program, Georgia Cancer CenterAugusta UniversityAugustaUSA
  2. 2.Department of Otolaryngology, Head and Neck SurgeryAsahikawa Medical UniversityAsahikawaJapan
  3. 3.Oncovir, Inc.Washington, DCUSA

Personalised recommendations