Tumor-specific neoantigens can be highly immunogenic, but their identification for each patient and the production of personalized cancer vaccines can be time-consuming and prohibitively expensive. In contrast, tumor-associated antigens are widely expressed and suitable as an off the shelf immunotherapy. Here, we developed a PLGA-based nanoparticle vaccine that contains both the immunogenic cancer germline antigen NY-ESO-1 and an α-GalCer analog IMM60, as a novel iNKT cell agonist and dendritic cell transactivator. Three peptide sequences (85-111, 117-143, and 157-165) derived from immunodominant regions of NY-ESO-1 were selected. These peptides have a wide HLA coverage and were efficiently processed and presented by dendritic cells via various HLA subtypes. Co-delivery of IMM60 enhanced CD4 and CD8 T cell responses and antibody levels against NY-ESO-1 in vivo. Moreover, the nanoparticles have negligible systemic toxicity in high doses, and they could be produced according to GMP guidelines. Together, we demonstrated the feasibility of producing a PLGA-based nanovaccine containing immunogenic peptides and an iNKT cell agonist, that is activating DCs to induce antigen-specific T cell responses.

Aix Marseille Univ CNRS INSERM CIML Centre d'Immunologie de Marseille Luminy Marseille France

Department of Tumor Immunology Radboud University Medical Center Radboud Institute for Molecular Life Sciences Nijmegen Netherlands

Institute of Macromolecular Chemistry v v i Academy of Sciences of the Czech Republic Prague Czechia

Medical Research Council Human Immunology Unit Radcliffe Department of Medicine Weatherall Institute of Molecular Medicine University of Oxford Oxford United Kingdom

Oncode Institute Nijmegen Netherlands

TRON Translational Oncology at the University Medical Center of the Johannes Gutenberg University Mainz gGmbH Mainz Germany

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Janelle V, Rulleau C, Del Testa S, Carli C, Delisle JS. T-Cell Immunotherapies targeting histocompatibility and tumor antigens in hematological malignancies. Front Immunol. (2020) 11:276. 10.3389/fimmu.2020.00276 PubMed DOI PMC

Akers SN, Odunsi K, Karpf AR. Regulation of cancer germline antigen gene expression: implications for cancer immunotherapy. Future Oncol. (2010) 6:717–32. 10.2217/fon.10.36 PubMed DOI PMC

Thomas R, Al-Khadairi G, Roelands J, Hendrickx W, Dermime S, Bedognetti D, et al. . NY-ESO-1 based immunotherapy of cancer: current perspectives. Front Immunol. (2018) 9:947. 10.3389/fimmu.2018.00947 PubMed DOI PMC

Esfandiary A, Ghafouri-Fard S. New York esophageal squamous cell carcinoma-1 and cancer immunotherapy. Immunotherapy. (2015) 7:411–39. 10.2217/imt.15.3 PubMed DOI

Bendelac A, Savage PB, Teyton L. The biology of NKT cells. Annu Rev Immunol. (2007) 25:297–336. 10.1146/annurev.immunol.25.022106.141711 PubMed DOI

Brennan PJ, Brigl M, Brenner MB. Invariant natural killer T cells: an innate activation scheme linked to diverse effector functions. Nat Rev Immunol. (2013) 13:101–17. 10.1038/nri3369 PubMed DOI

Cerundolo V, Silk JD, Masri SH, Salio M. Harnessing invariant NKT cells in vaccination strategies. Nat Rev Immunol. (2009) 9:28–38. 10.1038/nri2451 PubMed DOI

Chen YT, Scanlan MJ, Sahin U, Tureci O, Gure AO, Tsang S, et al. . A testicular antigen aberrantly expressed in human cancers detected by autologous antibody screening. Proc Natl Acad Sci USA. (1997) 94:1914–8. 10.1073/pnas.94.5.1914 PubMed DOI PMC

Jungbluth AA, Chen YT, Stockert E, Busam KJ, Kolb D, Iversen K, et al. . Immunohistochemical analysis of NY-ESO-1 antigen expression in normal and malignant human tissues. Int J Cancer. (2001) 92:856–60. 10.1002/ijc.1282 PubMed DOI

Gnjatic S, Nishikawa H, Jungbluth AA, Gure AO, Ritter G, Jager E, et al. . NY-ESO-1: review of an immunogenic tumor antigen. Adv Cancer Res. (2006) 95:1–30. 10.1016/S0065-230X(06)95001-5 PubMed DOI

Kakimi K, Isobe M, Uenaka A, Wada H, Sato E, Doki Y, et al. . A phase I study of vaccination with NY-ESO-1f peptide mixed with Picibanil OK-432 and Montanide ISA-51 in patients with cancers expressing the NY-ESO-1 antigen. Int J Cancer. (2011) 129:2836–46. 10.1002/ijc.25955 PubMed DOI

Odunsi K, Matsuzaki J, James SR, Mhawech-Fauceglia P, Tsuji T, Miller A, et al. . Epigenetic potentiation of NY-ESO-1 vaccine therapy in human ovarian cancer. Cancer Immunol Res. (2014) 2:37–49. 10.1158/2326-6066.CIR-13-0126 PubMed DOI PMC

Sahin U, Oehm P, Derhovanessian E, Jabulowsky RA, Vormehr M, Gold M, et al. . An RNA vaccine drives immunity in checkpoint-inhibitor-treated melanoma. Nature. (2020) 585:107–12. 10.1038/s41586-020-2537-9 PubMed DOI

Dolen Y, Kreutz M, Gileadi U, Tel J, Vasaturo A, van Dinther EA, et al. . Co-delivery of PLGA encapsulated invariant NKT cell agonist with antigenic protein induce strong T cell-mediated antitumor immune responses. Oncoimmunology. (2016) 5:e1068493. 10.1080/2162402X.2015.1068493 PubMed DOI PMC

Dölen Y, Valente M, Tagit O, Jäger E, Van Dinther EAW, van Riessen NK, et al. . Nanovaccine administration route is critical to obtain pertinent iNKt cell help for robust anti-tumor T and B cell responses. OncoImmunology. (2020) 9:1738813. 10.1080/2162402X.2020.1738813 PubMed DOI PMC

Valente M, Dolen Y, van Dinther E, Vimeux L, Fallet M, Feuillet V, et al. . Cross-talk between iNKT cells and CD8 T cells in the spleen requires the IL-4/CCL17 axis for the generation of short-lived effector cells. Proc Natl Acad Sci USA. (2019) 116:25816–27. 10.1073/pnas.1913491116 PubMed DOI PMC

Bartkowiak T, Singh S, Yang G, Galvan G, Haria D, Ai M, et al. . Unique potential of 4-1BB agonist antibody to promote durable regression of HPV+ tumors when combined with an E6/E7 peptide vaccine. Proc Natl Acad Sci USA. (2015) 112:E5290–9. 10.1073/pnas.1514418112 PubMed DOI PMC

Jukes JP, Gileadi U, Ghadbane H, Yu TF, Shepherd D, Cox LR, et al. . Non-glycosidic compounds can stimulate both human and mouse iNKT cells. Eur J Immunol. (2016) 46:1224–34. 10.1002/eji.201546114 PubMed DOI PMC

Robson NC, McAlpine T, Knights AJ, Schnurr M, Shin A, Chen W, et al. . Processing and cross-presentation of individual HLA-A, -B, or -C epitopes from NY-ESO-1 or an HLA-A epitope for Melan-A differ according to the mode of antigen delivery. Blood. (2010) 116:218–25. 10.1182/blood-2009-10-249458 PubMed DOI

Chen JL, Dawoodji A, Tarlton A, Gnjatic S, Tajar A, Karydis I, et al. . NY-ESO-1 specific antibody and cellular responses in melanoma patients primed with NY-ESO-1 protein in ISCOMATRIX and boosted with recombinant NY-ESO-1 fowlpox virus. Int J Cancer. (2015) 136:E590–601. 10.1002/ijc.29118 PubMed DOI

Choi EM, Palmowski M, Chen J, Cerundolo V. The use of chimeric A2K(b) tetramers to monitor HLA A2 immune responses in HLA A2 transgenic mice. J Immunol Methods. (2002) 268:35–41. 10.1016/S0022-1759(02)00198-9 PubMed DOI

Jackson H, Dimopoulos N, Mifsud NA, Tai TY, Chen Q, Svobodova S, et al. . Striking immunodominance hierarchy of naturally occurring CD8+ and CD4+ T cell responses to tumor antigen NY-ESO-1. J Immunol. (2006) 176:5908–17. 10.4049/jimmunol.176.10.5908 PubMed DOI

Nguyen DT. Cancer Antigenic Peptide Database. (2019). Available online at: https://caped.icp.ucl.ac.be/Peptide/list (accessed January 21, 2019).

Gnjatic S, Jager E, Chen W, Altorki NK, Matsuo M, Lee SY, et al. . CD8(+) T cell responses against a dominant cryptic HLA-A2 epitope after NY-ESO-1 peptide immunization of cancer patients. Proc Natl Acad Sci USA. (2002) 99:11813–8. 10.1073/pnas.142417699 PubMed DOI PMC

IEDB Analysis Resource: National Institute of Allergy Infectious Diseases. (2019). Available online at: https://tools.iedb.org/population/ (accessed January 21, 2019).

Bidmon N, Attig S, Rae R, Schroder H, Omokoko TA, Simon P, et al. . Generation of TCR-engineered T cells and their use to control the performance of T cell assays. J Immunol. (2015) 194:6177–89. 10.4049/jimmunol.1400958 PubMed DOI

Simon P, Omokoko TA, Breitkreuz A, Hebich L, Kreiter S, Attig S, et al. . Functional TCR retrieval from single antigen-specific human T cells reveals multiple novel epitopes. Cancer Immunol Res. (2014) 2:1230–44. 10.1158/2326-6066.CIR-14-0108 PubMed DOI

Jia J, Zhang Y, Xin Y, Jiang C, Yan B, Zhai S. Interactions between nanoparticles and dendritic cells: from the perspective of cancer immunotherapy. Front Oncol. (2018) 8:404. 10.3389/fonc.2018.00404 PubMed DOI PMC

Chenthamara D, Subramaniam S, Ramakrishnan SG, Krishnaswamy S, Essa MM, Lin FH, et al. . Therapeutic efficacy of nanoparticles and routes of administration. Biomater Res. (2019) 23:20. 10.1186/s40824-019-0166-x PubMed DOI PMC

Benlalam H, Linard B, Guilloux Y, Moreau-Aubry A, Derre L, Diez E, et al. . Identification of five new HLA-B*3501-restricted epitopes derived from common melanoma-associated antigens, spontaneously recognized by tumor-infiltrating lymphocytes. J Immunol. (2003) 171:6283–9. 10.4049/jimmunol.171.11.6283 PubMed DOI

Lopes L, Dewannieux M, Gileadi U, Bailey R, Ikeda Y, Whittaker C, et al. . Immunization with a lentivector that targets tumor antigen expression to dendritic cells induces potent CD8+ and CD4+ T-cell responses. J Virol. (2008) 82:86–95. 10.1128/JVI.01289-07 PubMed DOI PMC

Firat H, Garcia-Pons F, Tourdot S, Pascolo S, Scardino A, Garcia Z, et al. . H-2 class I knockout, HLA-A2.1-transgenic mice: a versatile animal model for preclinical evaluation of antitumor immunotherapeutic strategies. Eur J Immunol. (1999) 29:3112–21. PubMed

Newberg MH, Smith DH, Haertel SB, Vining DR, Lacy E, Engelhard VH. Importance of MHC class 1 alpha2 and alpha3 domains in the recognition of self and non-self MHC molecules. J Immunol. (1996) 156:2473–80. PubMed

Kageyama S, Wada H, Muro K, Niwa Y, Ueda S, Miyata H, et al. . Dose-dependent effects of NY-ESO-1 protein vaccine complexed with cholesteryl pullulan (CHP-NY-ESO-1) on immune responses and survival benefits of esophageal cancer patients. J Transl Med. (2013) 11:246. 10.1186/1479-5876-11-246 PubMed DOI PMC

Qiu L, Valente M, Dolen Y, Jager E, Beest MT, Zheng L, et al. . Endolysosomal-escape nanovaccines through adjuvant-induced tumor antigen assembly for enhanced effector CD8(+) T cell activation. Small. (2018) 14:e1703539. 10.1002/smll.201703539 PubMed DOI

Wada H, Isobe M, Kakimi K, Mizote Y, Eikawa S, Sato E, et al. . Vaccination with NY-ESO-1 overlapping peptides mixed with Picibanil OK-432 and montanide ISA-51 in patients with cancers expressing the NY-ESO-1 antigen. J Immunother. (2014) 37:84–92. 10.1097/CJI.0000000000000017 PubMed DOI

Baumgaertner P, Costa Nunes C, Cachot A, Maby-El Hajjami H, Cagnon L, Braun M, et al. . Vaccination of stage III/IV melanoma patients with long NY-ESO-1 peptide and CpG-B elicits robust CD8(+) and CD4(+) T-cell responses with multiple specificities including a novel DR7-restricted epitope. Oncoimmunology. (2016) 5:e1216290. 10.1080/2162402X.2016.1216290 PubMed DOI PMC

Gasser O, Sharples KJ, Barrow C, Williams GM, Bauer E, Wood CE, et al. . A phase I vaccination study with dendritic cells loaded with NY-ESO-1 and alpha-galactosylceramide: induction of polyfunctional T cells in high-risk melanoma patients. Cancer Immunol Immunother. (2018) 67:285–98. 10.1007/s00262-017-2085-9 PubMed DOI PMC

Dutoit V, Taub RN, Papadopoulos KP, Talbot S, Keohan ML, Brehm M, et al. . Multiepitope CD8(+) T cell response to a NY-ESO-1 peptide vaccine results in imprecise tumor targeting. J Clin Invest. (2002) 110:1813–22. 10.1172/JCI16428 PubMed DOI PMC

Bijker MS, van den Eeden SJ, Franken KL, Melief CJ, van der Burg SH, Offringa R. Superior induction of anti-tumor CTL immunity by extended peptide vaccines involves prolonged, DC-focused antigen presentation. Eur J Immunol. (2008) 38:1033–42. 10.1002/eji.200737995 PubMed DOI

Globisch T, Steiner N, Fulle L, Lukacs-Kornek V, Degrandi D, Dresing P, et al. . Cytokine-dependent regulation of dendritic cell differentiation in the splenic microenvironment. Eur J Immunol. (2014) 44:500–10. 10.1002/eji.201343820 PubMed DOI

Fujii S, Shimizu K, Smith C, Bonifaz L, Steinman RM. Activation of natural killer T cells by alpha-galactosylceramide rapidly induces the full maturation of dendritic cells in vivo and thereby acts as an adjuvant for combined CD4 and CD8 T cell immunity to a coadministered protein. J Exp Med. (2003) 198:267–79. 10.1084/jem.20030324 PubMed DOI PMC

Hermans IF, Silk JD, Gileadi U, Salio M, Mathew B, Ritter G, et al. . NKT cells enhance CD4+ and CD8+ T cell responses to soluble antigen in vivo through direct interaction with dendritic cells. J Immunol. (2003) 171:5140–7. 10.4049/jimmunol.171.10.5140 PubMed DOI

Hunder NN, Wallen H, Cao J, Hendricks DW, Reilly JZ, Rodmyre R, et al. . Treatment of metastatic melanoma with autologous CD4+ T cells against NY-ESO-1. N Engl J Med. (2008) 358:2698–703. 10.1056/NEJMoa0800251 PubMed DOI PMC

Fonteneau JF, Brilot F, Munz C, Gannage M. The tumor antigen NY-ESO-1 mediates direct recognition of melanoma cells by CD4+ T cells after intercellular antigen transfer. J Immunol. (2016) 196:64–71. 10.4049/jimmunol.1402664 PubMed DOI PMC

Tonti E, Galli G, Malzone C, Abrignani S, Casorati G, Dellabona P. NKT-cell help to B lymphocytes can occur independently of cognate interaction. Blood. (2009) 113:370–6. 10.1182/blood-2008-06-166249 PubMed DOI

Dellabona P, Abrignani S, Casorati G. iNKT-cell help to B cells: a cooperative job between innate and adaptive immune responses. Eur J Immunol. (2014) 44:2230–7. 10.1002/eji.201344399 PubMed DOI

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