Poly I: C-activated dendritic cells that were generated in CellGro for use in cancer immunotherapy trials

BackgroundFor clinical applications, dendritic cells (DCs) need to be generated using GMP-approved reagents. In this study, we tested the characteristics of DCs generated in two clinical grade culture media and activated by three maturation stimuli, Poly I: C, LPS and the mixture of proinflammatory cytokines in order to identify the optimal combination of culture media and activation stimulus for the clinical use.MethodWe tested DCs generation using two GMP-certified culture media, CellGro and RPMI+5% human AB serum and evaluated DCs morphology, viability and capapability to mature. We tested three maturation stimuli, PolyI:C, LPS and the mixture of proinflammatory cytokines consisting of IL-1, IL-6, TNF and prostaglandin E2. We evaluated the capacity of activated DCs to induce antigen-specific T cells and regulatory T lymphocytes.ResultsCell culture in CellGro resulted in a higher yield of immature DCs resulting from increased number of adherent monocytes. DCs that were generated in CellGro and activated using Poly I:C were the most efficient in expanding antigen-specific T cells compared to the DCs that were generated in other media and activated using LPS or the cocktail of proinflammatory cytokines. A comparison of all tested combinations revealed that DCs that were generated in CellGro and activated using Poly I:C induced low numbers of regulatory T cells.ConclusionIn this study, we identified monocyte-derived DCs that were generated in CellGro and activated using Poly I:C as the most potent clinical-grade DCs for the induction of antigen-specific T cells.

[1]  Yong‐jun Liu,et al.  Mouse and human dendritic cell subtypes , 2002, Nature Reviews Immunology.

[2]  D. Neuberg,et al.  Engagement of CD83 ligand induces prolonged expansion of CD8+ T cells and preferential enrichment for antigen specificity. , 2006, Blood.

[3]  G. Schuler,et al.  Rapid Induction of Tumor-specific Type 1 T Helper Cells in Metastatic Melanoma Patients by Vaccination with Mature, Cryopreserved, Peptide-loaded Monocyte-derived Dendritic Cells , 2002, The Journal of experimental medicine.

[4]  E. Maraskovsky,et al.  Dramatic increase in the numbers of functionally mature dendritic cells in Flt3 ligand-treated mice: multiple dendritic cell subpopulations identified , 1996, The Journal of experimental medicine.

[5]  R. Steinman,et al.  Differentiation of monocytes into dendritic cells in a model of transendothelial trafficking. , 1998, Science.

[6]  H. Ueno,et al.  Human Dendritic Cell Subsets for Vaccination , 2005, Journal of Clinical Immunology.

[7]  J. Banchereau,et al.  Flt3-Ligand and Granulocyte Colony-Stimulating Factor Mobilize Distinct Human Dendritic Cell Subsets In Vivo1 , 2000, The Journal of Immunology.

[8]  G. Schuler,et al.  A method for the production of cryopreserved aliquots of antigen-preloaded, mature dendritic cells ready for clinical use. , 2000, Journal of immunological methods.

[9]  Z. Berneman,et al.  Clinical-grade manufacturing of autologous mature mRNA-electroporated dendritic cells and safety testing in acute myeloid leukemia patients in a phase I dose-escalation clinical trial. , 2009, Cytotherapy.

[10]  M. Grégoire,et al.  Transient exposure of dendritic cells to maturation stimuli is sufficient to induce complete phenotypic maturation while preserving their capacity to respond to subsequent restimulation , 2003, Cancer Immunology, Immunotherapy.

[11]  C. Sousa,et al.  Dendritic cells in a mature age , 2006, Nature Reviews Immunology.

[12]  W. Germeraad,et al.  Increased Tumor-Specific CD8+ T Cell Induction by Dendritic Cells Matured with a Clinical Grade TLR-Agonist in Combination with IFN-γ , 2010, International journal of immunopathology and pharmacology.

[13]  F. Sallusto,et al.  Efficient presentation of soluble antigen by cultured human dendritic cells is maintained by granulocyte/macrophage colony-stimulating factor plus interleukin 4 and downregulated by tumor necrosis factor alpha , 1994, The Journal of experimental medicine.

[14]  D. Pospíšilová,et al.  In vitro assessment of dendritic cells pulsed with apoptotic tumor cells as a vaccine for ovarian cancer patients. , 2007, Clinical immunology.

[15]  O. Hrusak,et al.  Generation of functional dendritic cells for potential use in the treatment of acute lymphoblastic leukemia , 2002, Cancer Immunology, Immunotherapy.

[16]  J. Kirkwood,et al.  alpha-type-1 polarized dendritic cells: a novel immunization tool with optimized CTL-inducing activity. , 2004, Cancer research.

[17]  M. Lotze,et al.  Dendritic cells prolong tumor-specific T-cell survival and effector function after interaction with tumor targets. , 2001, Clinical cancer research : an official journal of the American Association for Cancer Research.

[18]  A. Šedivá,et al.  FOCUS on FOCIS: combined chemo-immunotherapy for the treatment of hormone-refractory metastatic prostate cancer. , 2009, Clinical immunology.

[19]  Edgar Schmitt,et al.  Synergistic activation of dendritic cells by combined Toll-like receptor ligation induces superior CTL responses in vivo. , 2006, Blood.

[20]  G. Schuler,et al.  Large-scale generation of mature monocyte-derived dendritic cells for clinical application in cell factories. , 2002, Journal of immunological methods.

[21]  E. Baek,et al.  Two-step maturation of immature DCs with proinflammatory cytokine cocktail and poly(I:C) enhances migratory and T cell stimulatory capacity. , 2010, Vaccine.

[22]  N. Bhardwaj,et al.  Directing dendritic cell immunotherapy towards successful cancer treatment. , 2010, Immunotherapy.

[23]  R. Steinman,et al.  Expansion of FOXP3high regulatory T cells by human dendritic cells (DCs) in vitro and after injection of cytokine-matured DCs in myeloma patients. , 2006, Blood.

[24]  E. Kremmer,et al.  Generation of clinical grade dendritic cells with capacity to produce biologically active IL-12p70 , 2007, Journal of Translational Medicine.

[25]  A. Šedivá,et al.  Maturation of dendritic cells by bacterial immunomodulators. , 2004, Vaccine.

[26]  J. Fay,et al.  Dendritic cell based tumor vaccines. , 2000, Immunology letters.

[27]  C. Figdor,et al.  Polyinosinic polycytidylic acid prevents efficient antigen expression after mRNA electroporation of clinical grade dendritic cells , 2009, Cancer Immunology, Immunotherapy.

[28]  H. Kim,et al.  Generation of functionally mature dendritic cells from elutriated monocytes using polyinosinic : polycytidylic acid and soluble CD40 ligand for clinical application , 2008, Clinical and experimental immunology.

[29]  S. Adams,et al.  Manipulating dendritic cell biology for the active immunotherapy of cancer. , 2004, Blood.

[30]  T. Hartung,et al.  Anti‐inflammatory effects of granulocyte colony‐stimulating factor , 1998, Current opinion in hematology.

[31]  P. Chevallier,et al.  Induction of leukemia-specific cytotoxic response by cross-presentation of late-apoptotic leukemic blasts by autologous dendritic cells of nonleukemic origin. , 2002, Cancer research.

[32]  J. Crowley,et al.  Frequent and specific immunity to the embryonal stem cell–associated antigen SOX2 in patients with monoclonal gammopathy , 2007, The Journal of experimental medicine.

[33]  E. Bouřa,et al.  Polyomavirus EGFP‐pseudocapsids: Analysis of model particles for introduction of proteins and peptides into mammalian cells , 2005, FEBS letters.

[34]  C. Caux,et al.  Selective Recruitment of Immature and Mature Dendritic Cells by Distinct Chemokines Expressed in Different Anatomic Sites , 1998, The Journal of experimental medicine.

[35]  H. Ueno,et al.  Recent Developments in Cancer Vaccines , 2011, The Journal of Immunology.

[36]  M. Grégoire,et al.  Standardized generation of fully mature p70 IL-12 secreting monocyte-derived dendritic cells for clinical use , 2001, Cancer Immunology, Immunotherapy.

[37]  A. Enk,et al.  Pro‐inflammatory cytokines and prostaglandins induce maturation of potent immunostimulatory dendritic cells under fetal calf serum‐free conditions , 1997, European journal of immunology.

[38]  G. Schuler,et al.  Generation of mature dendritic cells from human blood. An improved method with special regard to clinical applicability. , 1997, Advances in experimental medicine and biology.

[39]  C Caux,et al.  Immunobiology of dendritic cells. , 2000, Annual review of immunology.

[40]  Liangji Zhou,et al.  CD14+ blood monocytes can differentiate into functionally mature CD83+ dendritic cells. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[41]  Gerold Schuler,et al.  Dendritic Cells as Vectors for Therapy , 2001, Cell.

[42]  E. Kaempgen,et al.  Efficient elutriation of monocytes within a closed system (Elutra) for clinical-scale generation of dendritic cells. , 2005, Journal of immunological methods.

[43]  P. Martiat,et al.  Poly(I:C) used for human dendritic cell maturation preserves their ability to secondarily secrete bioactive IL-12. , 2004, International immunology.

[44]  A. Enk,et al.  Mage-3 and Influenza-Matrix Peptide-Specific Cytotoxic T Cells Are Inducible in Terminal Stage HLA-A2.1+ Melanoma Patients by Mature Monocyte-Derived Dendritic Cells1 , 2000, The Journal of Immunology.

[45]  M. Dhodapkar,et al.  Immunoprevention of cancer. , 2006, Hematology/oncology clinics of North America.

[46]  P. Coulie,et al.  T-cell responses of vaccinated cancer patients. , 2003, Current opinion in immunology.

[47]  E. Gilboa.  DC-based cancer vaccines. , 2007, The Journal of clinical investigation.

[48]  I. Svane,et al.  Increase of Circulating CD4+CD25highFoxp3+ Regulatory T Cells in Patients With Metastatic Renal Cell Carcinoma During Treatment With Dendritic Cell Vaccination and Low-Dose Interleukin-2 , 2010, Journal of immunotherapy.

[49]  P. Martiat,et al.  Dendritic Cells Generated in Clinical Grade Bags Strongly Differ in Immune Functionality When Compared With Classical DCs Generated in Plates , 2010, Journal of immunotherapy.

[50]  Franco Patrone,et al.  The use of dendritic cells in cancer immunotherapy. , 2008, Critical reviews in oncology/hematology.

[51]  H. Ueno,et al.  Building on dendritic cell subsets to improve cancer vaccines. , 2010, Current opinion in immunology.

[52]  G. Kvalheim,et al.  A full scale comparative study of methods for generation of functional Dendritic cells for use as cancer vaccines , 2007, BMC Cancer.

[53]  R. Steinman,et al.  Rapid generation of broad T-cell immunity in humans after a single injection of mature dendritic cells. , 1999, The Journal of clinical investigation.