Explore the vast range of titles available.

Tumor ablation technologies, such as radiofrequency-, cryo- or high-intensity focused ultrasound (HIFU) ablation will destroy tumor tissue in a minimally invasive manner. Ablation generates large volumes of tumor debris in situ, releasing multiple bio-molecules like tumor antigens and damage-associated molecular patterns. To initiate an adaptive antitumor immune response, antigen-presenting cells need to take up tumor antigens and, following activation, present them to immune effector cells. The impact of the type of tumor ablation on the precise nature, availability and suitability of the tumor debris for immune response induction, however, is poorly understood. In this review, we focus on immune effects after HIFU-mediated ablation and compare these to findings using other ablation technologies. HIFU can be used both for thermal and mechanical destruction of tissue, inducing coagulative necrosis or subcellular fragmentation, respectively. Preclinical and clinical results of HIFU tumor ablation show increased infiltration and activation of CD4 + and CD8 + T cells. As previously observed for other types of tumor ablation technologies, however, this ablation-induced enhanced infiltration alone appears insufficient to generate consistent protective antitumor immunity. Therapies combining ablation with immune stimulation are therefore expected to be key to boost HIFU-induced immune effects and to achieve systemic, long-lasting, antitumor immunity.

Objectives: Immunotherapy utilizing immune-stimulating cytokines such as IL12 holds great promise for the treatment of cancer. However, clinical use of IL12 is hampered due to severe toxicity following systemic administration. We here present a novel treatment strategy in which IL12 is chemically silenced by conjugation to PEG masks that sterically hinder the receptor binding. Subsequently, the masks can be released on demand using a bioorthogonal click reaction, cleaving the linker connecting the masks, thereby restoring the native cytokine. This “click-to-release” approach is based on the highly selective Inverse electron-demand Diels–Alder (IEDDA) pyridazine elimination reaction between a tetrazine (Tz) and a trans-cyclooctene (TCO), optimized for fast reaction kinetics and in vivo compatibility. Selective activation in the tumor microenvironment is achieved by pretargeting one component of this reaction to the tumor, triggering local activation of the masked IL12 once it is given in a secondary i.v. injection. Methods: IL12 masking and unmasking were evaluated in vitro with PAGE and HEK-Blue reporter cells and ex vivo with ELISA. Biodistribution in mice was evaluated with I-125 radiolabeling and biotin-click histochemistry. Results: Several designs were evaluated and optimized in vitro, resulting in an IL12-TCO-PEG construct that exhibited superior masking and subsequent reactivation upon reaction with a tetrazine bound to a TAG-72-targeted diabody. In tumor-bearing mice, we demonstrated that this diabody-tetrazine could efficiently pre-localize tetrazine in the tumor. Administration of IL12-TCO-PEG 24 h later afforded efficient and selective unmasking in tumors, but not in the blood. Conclusions: These results demonstrate proof of principle of the click-cleavable IL12 prodrug approach and showcase the versatility of the click-to-release reaction.

Sialic acid sugars cover the surface of dendritic cells (DCs) and have been suggested to impact several aspects of DC biology. Research into the role of sialic acids in DCs, however, is complicated by the limited number of tools available to modulate sialic acid expression. Here we report on a synthetic, fluorinated sialic acid mimetic, Ac53FaxNeu5Ac, which potently blocks sialic acid expression in human monocyte‐derived DCs (moDCs). Sialic acid blockade enhanced the responsiveness of moDCs to Toll‐like receptor (TLR) stimulation as measured by increased maturation marker expression and cytokine production. Consequently, the T‐cell activation capacity of Ac53FaxNeu5Ac‐treated moDCs was strongly increased. In addition to sialic acids, moDCs also expressed the sialic acid‐binding immunoglobulin‐like lectins (Siglecs) ‐3, ‐5, ‐7, ‐9 and ‐10, immune inhibitory receptors recognizing these sialic acids. Treatment with Ac53FaxNeu5Ac abrogated putative cis and trans interactions between sialic acids and Siglec‐7/‐9. Together, these data indicate that sialic acids limit the activation of moDCs via the TLR pathway, potentially by interacting with Siglec‐7 or Siglec‐9. Metabolic sialic acid blockade with Ac53FaxNeu5Ac could therefore potentially be used to generate more potent DC‐based vaccines for induction of robust anti‐viral or anti‐tumor immune responses.

The TLR9 agonist CpG is increasingly applied in preclinical and clinical studies as a therapeutic modality to enhance tumor immunity. The clinical application of CpG appears, however, less successful than would be predicted from animal studies. One reason might be the different administration routes applied in most mouse studies and clinical trials. We studied whether the efficacy of CpG as an adjuvant in cancer immunotherapy is dependent on the route of CpG administration, in particular when the tumor is destructed in situ. In situ tumor destruction techniques are minimally invasive therapeutic alternatives for the treatment of (nonresectable) solid tumors. In contrast to surgical resection, tumor destruction leads to the induction of weak but tumor-specific immunity that can be enhanced by coapplication of CpG. As in situ tumor destruction by cryosurgery creates an instant local release of antigens, we applied this model to study the efficacy of CpG to enhance antitumor immunity when administrated via different routes: peritumoral, intravenous, and subcutaneous but distant from the tumor. We show that peritumoral administration is superior in the activation of dendritic cells, induction of tumor-specific CTL, and long-lasting tumor protection. Although the intravenous and subcutaneous (at distant site) exposures are commonly used in clinical trials, they only provided partial protection or even failed to enhance antitumor responses as induced by cryosurgery alone. CpG administration greatly enhances the efficacy of in situ tumor destruction techniques, provided that CpG is administered in close proximity of the released antigens. Hence, this study helps to provide directions to fully benefit from CpG as immune stimulant in a clinical setting.

Saponin-based adjuvants (SBAs) are being used in animal and human (cancer) vaccines, as they induce protective cellular immunity. Their adjuvant potency is a factor of inflammasome activation and enhanced antigen cross-presentation by dendritic cells (DCs), but how antigen cross-presentation is induced is not clear. Here we show that SBAs uniquely induce intracellular lipid bodies (LBs) in the CD11b+ DC subset in vitro and in vivo . Using genetic and pharmacological interference in models for vaccination and in situ tumour ablation, we demonstrate that LB induction is causally related to the saponin-dependent increase in cross-presentation and T-cell activation. These findings link adjuvant activity to LB formation, aid the application of SBAs as a cancer vaccine component, and will stimulate development of new adjuvants enhancing T-cell-mediated immunity. Saponin-based adjuvants are being explored as vaccine components as they induce high levels of antigen cross-presentation, but it is unknown how. Here the authors show that these adjuvants enhance cross-presentation by driving production of lipid bodies inside CD11b dendritic cells.

In around half of the patients with neuroblastoma (NBL), the primary tumor is located in one of the adrenal glands. We have previously reported on a transplantable TH-MYCN model of subcutaneous (SC) growing NBL in C57Bl/6 mice for immunological studies. In this report, we describe an orthotopic TH-MYCN transplantable model where the tumor cells were injected intra-adrenally (IA) by microsurgery. Strikingly, 9464D cells grew out much faster in IA tumors compared to the subcutis. Tumors were infiltrated by equal numbers of lymphocytes and myeloid cells. Within the myeloid cell population, however, tumor-infiltrating macrophages were more abundant in IA tumors compared to SC tumors and expressed lower levels of MHC class II, indicative of a more immunosuppressive phenotype. Using 9464D cells stably expressing firefly luciferase, enhanced IA tumor growth could be confirmed using bioluminescence. Collectively, these data show that the orthotopic IA localization of TH-MYCN cells impacts the NBL tumor microenvironment, resulting in a more stringent NBL model to study novel immunotherapeutic approaches for NBL.

Background. Infection with Coxiella burnetii can lead to acute and chronic Q fever. Toll-like receptor 1 (TLR1), TLR2, TLR4, TLR6, nucleotide-binding oligomerization domain receptor 1 (NODI), NOD2, and the mitogenactivated protein kinases are central in the innate immune response against microorganisms, but little is known about their role in the recognition of C. burnetii in humans. Methods. Human peripheral blood mononudear cells (PBMCs) were stimulated with C. burnetii Nine Mile and the Dutch outbreak isolate C. burnetii 3262. TLRs were inhibited using specific antibodies or antagonists. Additionally, the influence of human polymorphisms in TLRs and Nod-like receptors (NLRs) on C. burnetii-induced cytokine production was assessed. Results. Inhibition of TLR2, p38, JNK, and ERK led to decreased cytokine responses in G frurnetf/-stimulated human PBMCs. Humans with polymorphisms in TLR1 and N0D2 had reduced cytokine production, compared with humans with wild-type genotypes, after stimulation. Interestingly, polymorphisms in TLR6 led to decreased cytokine production after C. burnetii 3262 stimulation but not after C. burnetii Nine Mile stimulation. Conclusions. The TLR1/TLR2 heterodimer and NOD2 are important recognition receptors for the induction of cytokine responses against C. burnetii in humans. Furthermore, an interesting finding was the divergent recognition of C. burnetii Nine Mile and C. burnetii 3262.

BackgroundTumor ablation techniques, like cryoablation, are successfully used in the clinic to treat tumors. The tumor debris remaining in situ after ablation is a major antigen depot, including neoantigens, which are presented by dendritic cells (DCs) in the draining lymph nodes to induce tumor-specific CD8+ T cells. We have previously shown that co-administration of adjuvants is essential to evoke strong in vivo antitumor immunity and the induction of long-term memory. However, which adjuvants most effectively combine with in situ tumor ablation remains unclear.Methods and resultsHere, we show that simultaneous administration of cytidyl guanosyl (CpG) with saponin-based adjuvants following cryoablation affects multifunctional T-cell numbers and interleukin (IL)-1 induced polymorphonuclear neutrophil recruitment in the tumor draining lymph nodes, relative to either adjuvant alone. The combination of CpG and saponin-based adjuvants induces potent DC maturation (mainly CpG-mediated), antigen cross-presentation (mainly saponin-based adjuvant mediated), while excretion of IL-1β by DCs in vitro depends on the presence of both adjuvants. Most strikingly, CpG/saponin-based adjuvant exposed DCs potentiate antigen-specific T-cell proliferation resulting in multipotent T cells with increased capacity to produce interferon (IFN)γ, IL-2 and tumor necrosis factor-α in vitro. Also in vivo the CpG/saponin-based adjuvant combination plus cryoablation increased the numbers of tumor-specific CD8+ T cells showing enhanced IFNγ production as compared with single adjuvant treatments.ConclusionsCollectively, these data indicate that co-injection of CpG with saponin-based adjuvants after cryoablation induces an increased amount of tumor-specific multifunctional T cells. The combination of saponin-based adjuvants with toll-like receptor 9 adjuvant CpG in a cryoablative setting therefore represents a promising in situ vaccination strategy.

Saponin-based adjuvants (SBAs) are promising new adjuvants that stand out as they not only enforce CD4 + T cell-mediated immunity and antibody responses, but also induce an unprecedented level of antigen cross-presentation by dendritic cells (DC) and subsequent CD8 + T cell activation. We discovered that SBA’s ability to boost cross-presentation depends on the induction of lipid bodies (LBs). Moreover, the MHCII lo CD11b hi DC subset was identified to be most responsive to SBA-induced cross-presentation. The aim is to further unravel the mechanisms behind the induction of DC cross-presentation by SBAs. Here we show that SBAs specifically induce the PKR-like Endoplasmic Reticulum kinase (PERK) pathway and that SBA-induced DC cross-presentation is dependent on activation of the PERK pathway. PERK activation and LB formation are both crucial for SBA-induced cross-presentation and PERK inhibition has little or no effect on SBA-induced LB formation. SBA’s responsiveness, LB formation and PERK activation are specific for the MHCII lo CD11b hi DCs. These findings contribute to understanding the pathways involved in SBA-induced cross-presentation and immune activation which will ultimately lead to the development of vaccines with improved efficiency and safety.

Background Dendritic cells (DCs) are the highly specialized antigen presenting cells of the immune system that play a key role in regulating immune responses. DCs can efficiently initiate immune responses or induce tolerance. Due to this dual function, DCs are studied in the context of immunotherapy for both cancer and autoimmune diseases. Characterization of DC-specific genes, leading to better understanding of DC immunobiology, will help to guide their use in clinical settings. We previously identified DC-STAMP, a multi-membrane spanning protein preferentially expressed by DCs. DC-STAMP resides in the endoplasmic reticulum (ER) of immature DCs and translocates towards the Golgi compartment upon maturation. In this study we knocked down DC-STAMP in mouse bone marrow-derived DCs (mBMDCs) to determine its function. Results We demonstrate that DC-STAMP knock-down mBMDCs secrete less IL-6, IL-12, TNF-α and IL-10 while IL-1 production is enhanced. Moreover, LPS-matured DC-STAMP knock-down mBMDCs show impaired T cell activation potential and induction of Th1 responses in an alloreaction. Conclusions We show that DC-STAMP plays an important role in cytokine production by mBMDCs following LPS exposure. Our results reveal a novel function of DC-STAMP in regulating DC-initiated immune responses.