PMID- 30122654 OWN - NLM STAT- MEDLINE DCOM- 20191008 LR - 20191008 IS - 1477-2566 (Electronic) IS - 1465-3249 (Linking) VI - 20 IP - 9 DP - 2018 Sep TI - An accelerated, clinical-grade protocol to generate high yields of type 1-polarizing messenger RNA-loaded dendritic cells for cancer vaccination. PG - 1164-1181 LID - S1465-3249(18)30548-6 [pii] LID - 10.1016/j.jcyt.2018.06.006 [doi] AB - BACKGROUND: Many efforts have been devoted to improve the performance of dendritic cell (DC)-based cancer vaccines. Ideally, a DC vaccine should induce robust type 1-polarized T-cell responses and efficiently expand antigen (Ag)-specific cytotoxic T-cells, while being applicable regardless of patient human leukocyte antigen (HLA) type. Production time should be short, while maximally being good manufacturing practice (GMP)-compliant. We developed a method that caters to all of these demands and demonstrated the superiority of the resulting product compared with DCs generated using a well-established "classical" protocol. METHODS: Immunomagnetically purified monocytes were cultured in a closed system for 3 days in GMP-compliant serum-free medium and cytokines, and matured for 24 h using monophosphoryl lipid A (MPLA)+ interferon-gamma (IFN-gamma). Mature DCs were electroporated with messenger RNA (mRNA) encoding full-length antigen and cryopreserved. "Classical" DCs were cultured for 8 days in flasks, with one round of medium and cytokine supplementation, and matured with tumor necrosis factor alpha (TNF-alpha) + prostaglandin E2 (PGE2) during the last 2 days. RESULTS: Four-day MPLA/IFN-gamma-matured DCs were superior to 8-day TNF-alpha/PGE2-matured DCs in terms of yield, co-stimulatory/co-inhibitory molecule expression, resilience to electroporation and cryopreservation and type 1-polarizing cytokine and chemokine release after cell thawing. Electroporated and cryopreserved DCs according to our protocol efficiently present epitopes from tumor antigen-encoding mRNA, inducing a strong expansion of antigen-specific CD8+ T-cells with full cytolytic capacity. CONCLUSION: We demonstrate using a GMP-compliant culture protocol the feasibility of generating high yields of mature DCs in a short time, with a superior immunogenic profile compared with 8-day TNF-alpha/PGE2-matured DCs, and capable of inducing vigorous cytotoxic T-cell responses to antigen from electroporated mRNA. This method is now being applied in our clinical trial program. CI - Copyright (c) 2018 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved. FAU - Brabants, E AU - Brabants E AD - Tumor Immunology Laboratory, Department of Respiratory Medicine, Ghent University Hospital, Ghent, Belgium. Electronic address: Elisabeth.Brabants@ugent.be. FAU - Heyns, K AU - Heyns K AD - Tumor Immunology Laboratory, Department of Respiratory Medicine, Ghent University Hospital, Ghent, Belgium. FAU - De Smet, S AU - De Smet S AD - Cell Therapy Unit, Department of Regenerative Medicine, Ghent University Hospital, Ghent, Belgium. FAU - Devreker, P AU - Devreker P AD - Cell Therapy Unit, Department of Regenerative Medicine, Ghent University Hospital, Ghent, Belgium. FAU - Ingels, J AU - Ingels J AD - Cell Therapy Unit, Department of Regenerative Medicine, Ghent University Hospital, Ghent, Belgium. FAU - De Cabooter, N AU - De Cabooter N AD - Tumor Immunology Laboratory, Department of Respiratory Medicine, Ghent University Hospital, Ghent, Belgium; Primary Immunodeficiencies Research Laboratory, Department of Pediatric Lung Diseases;-Immunodeficiencies; and-Infectious Diseases, Ghent University Hospital, Ghent, Belgium. FAU - Debacker, V AU - Debacker V AD - Tumor Immunology Laboratory, Department of Respiratory Medicine, Ghent University Hospital, Ghent, Belgium; Primary Immunodeficiencies Research Laboratory, Department of Pediatric Lung Diseases;-Immunodeficiencies; and-Infectious Diseases, Ghent University Hospital, Ghent, Belgium. FAU - Dullaers, M AU - Dullaers M AD - Tumor Immunology Laboratory, Department of Respiratory Medicine, Ghent University Hospital, Ghent, Belgium; Primary Immunodeficiencies Research Laboratory, Department of Pediatric Lung Diseases;-Immunodeficiencies; and-Infectious Diseases, Ghent University Hospital, Ghent, Belgium. FAU - VAN Meerbeeck, J P AU - VAN Meerbeeck JP AD - Center for Oncological Research, Department of Pulmonology, Antwerp University Hospital, Antwerp, Belgium. FAU - Vandekerckhove, B AU - Vandekerckhove B AD - Cell Therapy Unit, Department of Regenerative Medicine, Ghent University Hospital, Ghent, Belgium. FAU - Vermaelen, K Y AU - Vermaelen KY AD - Tumor Immunology Laboratory, Department of Respiratory Medicine, Ghent University Hospital, Ghent, Belgium. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20180816 PL - England TA - Cytotherapy JT - Cytotherapy JID - 100895309 RN - 0 (Antigens, Neoplasm) RN - 0 (Cancer Vaccines) RN - 0 (Epitopes) RN - 0 (Lipid A) RN - 0 (RNA, Messenger) RN - 0 (Tumor Necrosis Factor-alpha) RN - 82115-62-6 (Interferon-gamma) RN - K7Q1JQR04M (Dinoprostone) RN - MWC0ET1L2P (monophosphoryl lipid A) SB - IM MH - Antigens, Neoplasm/genetics/immunology MH - *Cancer Vaccines MH - Cell Culture Techniques/*methods MH - Cell Differentiation MH - Cryopreservation MH - Dendritic Cells/*cytology/immunology MH - Dinoprostone/pharmacology MH - Electroporation MH - Epitopes MH - Humans MH - Interferon-gamma/pharmacology MH - Lipid A/analogs & derivatives/pharmacology MH - Monocytes/cytology MH - *RNA, Messenger/genetics MH - T-Lymphocytes, Cytotoxic/immunology MH - Tumor Necrosis Factor-alpha/pharmacology OTO - NOTNLM OT - dendritic cell OT - immunotherapy OT - interferon-gamma OT - messenger RNA OT - monophosphoryl lipid A OT - prostaglandin E2 OT - tumor necrosis factor-alpha OT - vaccination EDAT- 2018/08/21 06:00 MHDA- 2019/10/09 06:00 CRDT- 2018/08/21 06:00 PHST- 2018/03/22 00:00 [received] PHST- 2018/05/24 00:00 [revised] PHST- 2018/06/26 00:00 [accepted] PHST- 2018/08/21 06:00 [pubmed] PHST- 2019/10/09 06:00 [medline] PHST- 2018/08/21 06:00 [entrez] AID - S1465-3249(18)30548-6 [pii] AID - 10.1016/j.jcyt.2018.06.006 [doi] PST - ppublish SO - Cytotherapy. 2018 Sep;20(9):1164-1181. doi: 10.1016/j.jcyt.2018.06.006. Epub 2018 Aug 16.