Download The TLR8 agonist R848 primes human monocyte

The TLR8 agonist R848 primes human monocyte-derived dendritic cells for a secondary, CD40dependent, burst of IL-12p70 production
Author Block L. Bergqvist1, S. Holmgren1, S. Johnson1, K. Gustafsson2, J. Alder1, B. Andersson1, L.
Pellettieri2, V. d´Angelo3, P. Eriksson2, A. Karlsson-Parra1;
1Department of Clinical Immunology, Gothenburg, Sweden, 2Department of Neuroscience and
Physiology, Gothenburg, Sweden, 3Department of Neurosurgery, San Giovanni Rotondo, Italy.
Abstract:
Effective induction of antitumor CTL responses requires fully mature DCs that express high levels of
costimulatory molecules. In addition, high interleukin-12p70 (IL-12p70) secretion dramatically
enhances the ability of DC-based vaccines to induce tumor-specific Th1 cells and CTLs, and promotes
tumor rejection in therapeutic mouse models. A major concern, however, is whether mature DCs are
still capable of secreting IL-12p70 after in vivo administration at the time of interaction with CD40
ligand expressing T cells. Here, we report that maturation with the TLR8 agonist R848, but not
maturation with LPS or poly-I:C, primes monocyte-dervied human DCs (propagated in a clinically
relevant serum-free medium supplemented with GM-CSF and IL-4) for substantial production of IL12p70 upon CD40 cross-linking 48-72 hours apart from initial stimulation. Combining R848 with LPS
or poly-I:C induced a significant primary burst of IL-12p70 production followed by strong secondary
burst of IL-12p70 production after CD40-mediated restimulation. Addition of IFN-gamma to R848
during the maturation step syngergistically enhanced the secondary, CD40-mediated, burst of IL12p70 production. In contrast, re-stimulation of R848/IFN-gamma-primed DCs with TLR ligands
(R848, LPS or poly I:C) did not induce any substantial IL-12p70 production. Our data thus suggest that
R848 may be one of the most appropriate agents to generate mature DC by retaining the ability of
these DCs to secrete bioactive IL-12 upon CD40-stimulation.
Profuse and sustained production of MIG/CXCL9 by a clinical grade dendritic cell vaccine: A
feature to provide NK cell-dependent boosting for TH1 polarization
Author Block K. Gustafsson1, M. Persson2, L. Bergqvist3, J. Alder3, J. Nyström2, B. Andersson3, L.
Pellettieri1, V. d´Angelo4, P. Eriksson1, A. Karlsson-Parra3;
1Department of Neuroscience and Physiology, Gothenburg, Sweden, 2Department of Medicine,
Gothenburg, Sweden, 3Department of Clinical Immunology, Gothenburg, Sweden, 4Department of
Neurosurgery, San Giovanni Rotondo, Italy.
Abstract:
CXCR3-dependent recruitment of circulating NK-cells into inflamed nymph nodes is known to provide
a potent, interferon (IFN)-gamma-dependent, boost for TH1-polarized immune responses in mouse
models. Such NK cell recruitment into draining lymph nodes is induced by certain subcutaneously
injected adjuvants, including mature dendritic cells (DCs). We here demonstrate that monocytederived immature human DCs stimulated with poly I:C in combination with IFN-alpha, IFN-gamma, IL1 beta and TNF-alpha (alpha-DC1) exhibit a sustained and profuse production (> 150 ng/mL/106 DCs)
of the CXCR3-ligand monokine induced by IFNgamma (MIG)/CXCL9 after withdrawal of maturation
stimuli. In contrast, no measurable or low levels (< 1 ng/mL/106 DCs) of MIG/CXCL9 are produced by
DCs after maturation with the current “gold standard” maturation protocol for human DC-based cancer
vaccines consisting of TNF-alpha, IL-1 beta, IL-6 and prostaglandin E2 (PGE2-DC). Functional studies
in vitro further demonstrate that alpha-DC1-supernatants actively recruit NK cells, and that addition of
anti-MIG/CXCL9 antibodies to the supernatant blocks this recruitment. Finally, alpha-DC1, but not
PGE2-DC, significantly activate cocultured autologous NK cells as determined by CD69 expression
and intracellular IFN-gamma production. These novel findings indicate that fully matured and
subsequently injected human alpha-DC1-based clinical grade vaccines have the potential to recruit
and activate NK-cells during their arrival to draining lymph nodes and that this feature may be of
relevance for efficient priming of tumor-specific Th1 cells and CTLs.
Allogeneic monocyte-derived cells loaded with tumor antigens as a combined antigen-delivery
vehicle and adjuvant in cancer immunotherapy
Author Block J. Alder1, K. Gustafsson2, A. Wallgren1, B. Andersson1, E. Jennische3, L. Pellettieri2, V.
d´Angelo4, P. Eriksson2, S. Lange5, A. Karlsson-Parra1;
1Department of Clinical Immunology, Gothenburg, Sweden, 2Department of Neuroscience and
Physiology, Gothenburg, Sweden, 3Department of Anatomy and Cell Biology, Gothenburg, Sweden,
4Department of Neurosurgery, San Giovanni Rotondo, Italy, 5Department of Clinical Bacteriology,
Gothenburg, Sweden.
Abstract:
We have recently shown that addition of antigen-presenting cells (APCs) into an allogeneic immune
compartment in vitro elicits an inflammatory reaction that promotes maturation of bystander dendritic
cells with T helper 1 (TH1)-inducing capacity. We therefore proposed that vaccination with allogeneic,
tumor-antigen-loaded, APCs would lead to efficient cross-priming of tumor-specific T cells. Fisher344
female rats were challenged subcutaneously with the highly malignant breast cancer cell line
13762MAT B III. The rats were vaccinated in both prophylactic and therapeutic settings. In initial
experiments, the vaccine cells consisted of allogeneic peripheral blood monocytes cultivated for 2
days in GM-CSF. After 1 day the monocytes were loaded with apoptotic tumor cells with or without
addition of Vibrio cholerae neuraminidase (NAS), an enzyme known to induce inflammatory
chemokine production in monocytes and to enhance their allogenicity. Prophylactic vaccinations
reduced tumor take from 80% in non-vaccinated rats to 20% in vaccinated rats. This immunity was
long-lasting since re-challenge of tumor-rejecting rats with tumor cells 6 weeks later failed to induce
any tumor growth. In the therapeutic setting all rats developed tumors but tumor growth was
significantly reduced in rats given allogeneic and NAS-treated tumor cell loaded monocytes (p< 0.05)
or monocyte-derived dendritic cells (differentiated with GM-CSF+ IL-4 for 5 days, p<0.001). Our results
indicate that NAS-treated allogeneic monocyte-derived cells may act as antigen carrier as well as
adjuvant in cancer vaccination and immunotherapy.