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Background: Intratumorally delivered immunotherapies have the potential to favorably alter the local tumor microenvironment and may stimulate systemic host immunity, offering an alternative or adjunct to other local and systemic treatments. Despite their potential, these therapies have had limited success in late-phase trials for advanced cancer resulting in few formal approvals. The Society for Immunotherapy of Cancer (SITC) convened a panel of experts to determine how to design clinical trials with the greatest chance of demonstrating the benefits of intratumoral immunotherapy for patients with cancers across all stages of pathogenesis., Methods: An Intratumoral Immunotherapy Clinical Trials Expert Panel composed of international key stakeholders from academia and industry was assembled. A multiple choice/free response survey was distributed to the panel, and the results of this survey were discussed during a half-day consensus meeting. Key discussion points are summarized in the following manuscript., Results: The panel determined unique clinical trial designs tailored to different stages of cancer development-from premalignant to unresectable/metastatic-that can maximize the chance of capturing the effect of intratumoral immunotherapies. Design elements discussed included study type, patient stratification and exclusion criteria, indications of randomization, study arm determination, endpoints, biological sample collection, and response assessment with biomarkers and imaging. Populations to prioritize for the study of intratumoral immunotherapy, including stage, type of cancer and line of treatment, were also discussed along with common barriers to the development of these local treatments., Conclusions: The SITC Intratumoral Immunotherapy Clinical Trials Expert Panel has identified key considerations for the design and implementation of studies that have the greatest potential to capture the effect of intratumorally delivered immunotherapies. With more effective and standardized trial designs, the potential of intratumoral immunotherapy can be realized and lead to regulatory approvals that will extend the benefit of these local treatments to the patients who need them the most., Competing Interests: Competing interests: JJL: Researcher: AbbVie, Astellas, Astrazeneca, Bristol-Myers Squibb, Corvus, Day 1, EMD Serono, Fstar, Genmab, Ikena, Immatics, Incyte, Kadmon, KAHR, Macrogenics, Merck, Moderna, Nektar, Next Cure, Numab, Palleon, Pfizer, Replimmune, Rubius, Servier, Scholar Rock, Synlogic, Takeda, Trishula, Tizona, XencorConsultant/Advisor/Speaker: Abbvie, Agenus, Alnylam, Atomwise, Bayer, Bristol-Myers Squibb, Castle, Checkmate, Codiak, Crown, Cugene, Curadev, Day One, Eisai, EMD Serono, Endeavor, Flame, G1 Therapeutics, Genentech, Gilead, Glenmark, HotSpot, Kadmon, KSQ, Janssen, Ikena, Inzen, Immatics, Immunocore, Incyte, Instil, IO Biotech, Macrogenics, Merck, Mersana, Nektar, Novartis, Partner, Pfizer, Pioneering Medicines, PsiOxus, Regeneron, Replimmune, Ribon, Roivant, Servier, STINGthera, Synlogic, Synthekine, 7 Hills, Affivant, Bright Peak, Exo, Fstar, Inzen, RefleXion, Xilio (stock) Actym, Alphamab Oncology, Arch Oncology, Duke Street Bio, Kanaph, Mavu, NeoTx, Onc.AI, OncoNano, physIQ, Pyxis, Saros, STipe, Tempest, Abbvie, Agenus, Amgen, Immutep, Evaxion. DD: Grants/Research Support (institutional): Arcus, Immunocore, Merck, Regeneron Pharmaceuticals, Tesaro/GSK. Consultant: ACM Bio, Ascendis, Castle, Clinical Care Options (CCO), Gerson Lehrman Group (GLG), Immunitas, Medical Learning Group (MLG), Replimmune, Trisalus, Xilio Therapeutics. CE Speakers’ Bureau: Castle Biosciences. Stockholder: None. Intellectual Property: US Patent 63/124,231, "Compositions and Methods for Treating Cancer", December 11, 2020 US Patent 63/208,719, "Compositions and Methods For Responsiveness to Immune Checkpoint Inhibitors (ICI), Increasing Effectiveness of ICI and Treating Cancer", June 9, 2021. AM: Owner: HiFiBio, Deka Bio, Hotspot Therapeutics, Shattuck Labs, Researcher: Astra Zeneca, BMS, Sanofi, Consultant/Advisor/Speaker: Gritstone, Innate Pharma, Neogene, Deka Bio, Hotspot therapeutics, J&J, Medicxi, Depth Charge, BiolineRx, Clover Pharma, Grey Wolf, Lytix, RedX, HiFiBio, ImCheck, Applied Materials, Takeda, Shattuck Labs, Marengo therapeutics, Pierre Fabre, Third Rock Ventures, SotioPublicly Traded Stocks: Centessa. RHA–Employee: Seven & Eight Biopharmaceuticals. Ownership Interest Less Than 5 Percent: Seven & Eight Biopharmaceuticals. NB: Consulting Fees: Novartis, Apricity, Rome Therapeutics, BreakBio, Carisma Therapeutics, CureVac, BioNTech, Gilead, Tempest Therapeutics, Boehringer Ingelheim, Contracted Research: Regeneron Pharmaceutics, Dragonfly Therapeutics, Harbour Biomed Sciences, Ownership Interest Less Than 5 Percent: BreakBio, Apricity. JDB: Researcher: Merck, Genentech, Astrazeneca, Kite/Gilead, BMS, Celldex, Oncovir, Seattle Genetics, ADC Therapeutics, Epizyme, Consultant/Advisor/Speaker: Merck, Genentech, Astrazeneca, Kite/Gilead, BMS, Seattle Genetics, ADC Therapeutics, Epizyme, Asgaard Therapeutics, Global BioAccess, SIRPant Immunotherapeutics. JC: Executive Role: UofL Health, Researcher: Amgen, Iovance, Fate, Replimune, Consultant/Advisor/Speaker: Replimune advisory board 2020-21, Royalties or Patent Beneficiary: US Patents: University of Louisville. RC: Employee: Replimune. TdB: Consultant/Advisor/Speaker: Terumo, Boston Scientific, Astra Zeneca, Nanobiotix, Quantum Surgical, Guerbet, Cook medical, Johnson & Johnson. TDdG: Patents: (1) The use of cytostatics for the accelerated differentiation of DC; WO2009019320-A2; WO2009019320-A3; AU2008285598-A1; EP2281030-A2; CA2724018-A1; US2011117051-A1. US8,470,789B2; DCprime BV. (2) Immunoglobulins binding human Vγ9VÎ’2 T cell receptors; P31885NL00. Consulting Fees: Mendus (formerly Immunicum, formerly DCPrime BV), Partner Therapeutics, GE Health, LAVA Therapeutics, Contracted Research: Idera Pharmaceuticals, Macrophage Parma, Ownership Interest Less Than 5 Percent: LAVA Therapeutics. MGF: Employee: Regeneron. GVG: Employee: Merck. Publicly Traded Stocks: Immunogen, Aveo, Beta Bionics. KJH: Researcher: AstraZeneca, Boehringer-Ingelheim, Replimune, Consultant/Advisor/Speaker: Arch Oncology, AstraZeneca, BMS, Boehringer-Ingelheim, Codiak, Eisai, Inzen, Merck-Serono, MSD, Oncolys, Pfizer, Replimune. HK: Employee: Ankyra Therapeutics. Consultant/Advisor/Speaker: Castle Biosciences, Marengo Therapeutics. CMK: Researcher: Amgen, Merck & Co., Kartos Pharmaceuticals, EMD Serono, Deciphera Pharmaceuticals, Eily lilly, Blueprint Medicines, Incyte Corporation, Clovis Oncology, BMS, Aadi Bioscience, Pfizer, Iterion Therapeutics, Springworks Therapeutics, Nektar Therapeutics, GSK, Adaptimmune, Medimmune, Bioatla, Oncternal Therapeutics, Daiichi- Sankyo Co. Traycon Pharm, Eisai. Ascentage Pharma, Hutchison Medipharma, Salarius Pharmaceuticals, Loxo Oncology, -BTG Specialty Pharmaceuticals, AStex Pharmaceuticals, Xencor, bayer healthcare, Athenex pharmaceuticals, servier, K- group Beta, Trilliium Pharmaceuticals, Ayala Pharmaceuticals, Ningbo Newbay Technology Development, Rain Therapeutics, Inhibrx, C4 Therapeutics, Foghorn Therapeutics, Theseus Pharmaceuticals, Cogent Therapeutics, Curadev PharmaRegeneron, Consultant/Advisor/Speaker: Chemocentryx, Kartos, Servier, Immunicom, Other: Spouse employed by Daichii Sankyo with stock options. ADK: Employee: Merck. KL: Employee: Marengo Therapeutics. SL: Executive Role: Big Against Cancer (Belgium), Breast Cancer Trials (Australia), International Breast Cancer Study Group (Switzerland) Researcher: Research funding to institution from Novartis, Bristol Meyers Squibb, Merck, Puma Biotechnology, Eli Lilly, Nektar Therapeutics, Astra Zeneca, Roche-Genentech and Seattle Genetics, Consultant/Advisor/Speaker: Consultant (not compensated) to Seattle Genetics, Novartis, Bristol Meyers Squibb, Merck, AstraZeneca, Eli Lilly, Pfizer, Gilead Therapeutics and Roche-Genentech. Consultant (paid to institution) to Aduro Biotech, Novartis, GlaxoSmithKline, Roche-Genentech, Astra Zeneca, Silverback Therapeutics, G1 Therapeutics, PUMA Biotechnologies, Pfizer, Gilead Therapeutics, Seattle Genetics, Daiichi Sankyo, Merck, Amunix, Tallac Therapeutics, Eli Lilly and Bristol Meyers Squibb, Role: Presenter (Speaker). GVL: Consultant/Advisor/Speaker: Consultant Advisor for Agenus, Amgen, Array Biopharma, AstraZeneca UK, Boehringer Ingelheim International, Bristol Myers Squibb, Evaxion Biotech A/S, Hexal AG (Sandoz Company), Highlight Therapeutics S.L., Innovent Biologics USA, Merck Sharpe & Dohme (Australia), Merck Sharpe & Dohme, Novartis Pharma AG, PHMR, Pierre Fabre, Provectus Australia, Qbiotics Group Limited, Regeneron Pharmaceuticals. IM: Consulting Fees: Bristol-Myers Squibb, F-STAR, Alligator, Pharma Mar, AstraZeneca, Numab Therapeutics, Roche, Amunix, Gossamer, Molecular Partners, Merck-Serono, Genmab, PharmaMar, Contracted Research: Roche, Bristol-Myers Squibb, Highlight Therapeutics, Alligator, Genmab, Astrazeneca. MRM: Researcher : Roche, Astrazeneca, Novartis, Immunocore, BMS, Pfizer, Merck/MSD, Regeneron, BiolineRx, Replimune, GRAIL, Alkermes, iOx, Vaccitech, Consultant/Advisor/Speaker: Infinitopes. BN: Researcher: As a principal investigator for clinical trials sponsored by UZ Brussel, my institution received support from Novartis, Pfizer, Bayer, BMS, MSD, Pierre-Fabre, Amgen, Consultant/Advisor/Speaker: Novartis, BMS, MSD, Pfizer, Pierre-Fabre, Amgen. DJP: Consulting Fees: ViiV Healthcare, Bayer, Hoffman La Roche, EISAI, H3B, MiNa Alpha Therapeutics, DaVolterra, Fees for Non CE Services: Hoffmann La Roche, EISAI, Contracted Research: Merck Sharpe and Dohme, Bristol Myers Squibb (to institution). RAS: Researcher: Boston ScientificConsultant/Advisor/Speaker: Cook Medical, TriSalus, Replimune, Medtronic. SBS: Owner: Aperture Medical Technology, Executive Role: Aperture Medical Technology, Researcher: GE Healthcare, Johnson & Johnson, Elesta, Consultant/Advisor/Speaker: GE Healthcare, XACT Robotics, Microbot, Candel Therapeutics, Varian, Merck & Co., Royalty and Patent Beneficiary: Aperture Medical Technology, Publicly Traded Stocks: Johnson & Johnson, Poseidx Therapeutics, Motus GI, Sientra, Avadel, Lantheus. PS: Employee: Johnson & Johnson, Publicly Traded Stocks: Johnson & Johnson. SITC staff members SMW, CG, AK, NL, EG and KJ have no disclosures., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Vγ9Vδ2 T cells are effector cells with proven antitumor efficacy against a broad range of cancers. This study aimed to assess the antitumor activity and safety of a bispecific antibody directing Vγ9Vδ2 T cells to EGFR-expressing tumors. An EGFR-Vδ2 bispecific T-cell engager (bsTCE) was generated, and its capacity to activate Vγ9Vδ2 T cells and trigger antitumor activity was tested in multiple in vitro, in vivo, and ex vivo models. Studies to explore safety were conducted using cross-reactive surrogate engagers in nonhuman primates (NHP). We found that Vγ9Vδ2 T cells from peripheral blood and tumor specimens of patients with EGFR+ cancers had a distinct immune checkpoint expression profile characterized by low levels of PD-1, LAG-3, and TIM-3. Vγ9Vδ2 T cells could be activated by EGFR-Vδ2 bsTCEs to mediate lysis of various EGFR+ patient-derived tumor samples, and substantial tumor growth inhibition and improved survival were observed in in vivo xenograft mouse models using peripheral blood mononuclear cells (PBMC) as effector cells. EGFR-Vδ2 bsTCEs exerted preferential activity toward EGFR+ tumor cells and induced downstream activation of CD4+ and CD8+ T cells and natural killer (NK) cells without concomitant activation of suppressive regulatory T cells observed with EGFR-CD3 bsTCEs. Administration of fully cross-reactive and half-life extended surrogate engagers to NHPs did not trigger signals in the safety parameters that were assessed. Considering the effector and immune-activating properties of Vγ9Vδ2 T cells, the preclinical efficacy data and acceptable safety profile reported here provide a solid basis for testing EGFR-Vδ2 bsTCEs in patients with EGFR+ malignancies., (©2023 American Association for Cancer Research.)

γδ T-cells directly recognize and kill transformed cells independently of HLA-antigen presentation, which makes them a highly promising effector cell compartment for cancer immunotherapy. Novel γδ T-cell-based immunotherapies, primarily focusing on the two major γδ T-cell subtypes that infiltrate tumors ( i.e. Vδ1 and Vδ2), are being developed. The Vδ1 T-cell subset is enriched in tissues and contains both effector T-cells as well as regulatory T-cells with tumor-promoting potential. Vδ2 T-cells, in contrast, are enriched in circulation and consist of a large, relatively homogeneous, pro-inflammatory effector T-cell subset. Healthy individuals typically harbor in the order of 50-500 million Vγ9Vδ2 T-cells in the peripheral blood alone (1-10% of the total CD3 + T-cell population), which can rapidly expand upon stimulation. The Vγ9Vδ2 T-cell receptor senses intracellular phosphorylated metabolites, which accumulate in cancer cells as a result of mevalonate pathway dysregulation or upon pharmaceutical intervention. Early clinical studies investigating the therapeutic potential of Vγ9Vδ2 T-cells were based on either ex vivo expansion and adoptive transfer or their systemic activation with aminobisphosphonates or synthetic phosphoantigens, either alone or combined with low dose IL-2. Immune-related adverse events (irAE) were generally \mild, but the clinical efficacy of these approaches provided overall limited benefit. In recent years, critical advances have renewed the excitement for the potential of Vγ9Vδ2 T-cells in cancer immunotherapy. Here, we review γδ T-cell-based therapeutic strategies and discuss the prospects of those currently evaluated in clinical studies in cancer patients as well as future therapies that might arise from current promising pre-clinical results., Competing Interests: JS-E, MJ and LK are funded by Lava therapeutics. HV, PP, EE, BW are employed by and hold stock of LAVA Therapeutics. TG holds stock of LAVA Therapeutics., (Copyright © 2022 Saura-Esteller, de Jong, King, Ensing, Winograd, de Gruijl, Parren and van der Vliet.)

Immune checkpoint blockade (ICB) has changed the therapeutic landscape of oncology but its impact is limited by primary or secondary resistance. ICB resistance has been related to a lack of T cells infiltrating into the tumor. Strategies to overcome this hurdle have so far focused on the tumor microenvironment, but have mostly overlooked the role of tumor-draining lymph nodes (TDLN). Whereas for CTLA-4 blockade TDLN have long since been implicated due to its perceived mechanism-of-action involving T cell priming, only recently has evidence been emerging showing TDLN to be vital for the efficacy of PD-1 blockade as well. TDLN are targeted by developing tumors to create an immune suppressed pre-metastatic niche which can lead to priming of dysfunctional antitumor T cells. In this review, we will discuss the evidence that therapeutic targeting of TDLN may ensure sufficient antitumor T cell activation and subsequent tumor infiltration to facilitate effective ICB. Indeed, waves of tumor-specific, proliferating stem cell-like, or progenitor exhausted T cells, either newly primed or reinvigorated in TDLN, are vital for PD-1 blockade efficacy. Both tumor-derived migratory dendritic cell (DC) subsets and DC subsets residing in TDLN, and an interplay between them, have been implicated in the induction of these T cells, their imprinting for homing and subsequent tumor control. We propose that therapeutic approaches, involving local delivery of immune modulatory agents for optimal access to TDLN, aimed at overcoming hampered DC activation, will enable ICB by promoting T cell recruitment to the tumor, both in early and in advanced stages of cancer., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 van Pul, Fransen, van de Ven and de Gruijl.)

Lessons Learned: The novel therapeutic vaccine hVEGF 26-104 /RFASE was found to be safe and well tolerated in patients with cancer. hVEGF 26-104 /RFASE failed to induce seroconversion against native hVEGF 165 and, accordingly, neither a decrease in circulating vascular endothelial growth factor (VEGF) levels nor clinical benefit was observed. Remarkably, hVEGF 26-104 /RFASE induced VEGF 165 -neutralizing antibodies in a nonhuman primate model. The absence of seroconversion in human calls for caution in the interpretation of efficacy of human vaccines in nonhuman primates., Background: Targeting vascular endothelial growth factor-A (VEGF) is a well-established anticancer therapy. We designed a first-in-human clinical trial to investigate the safety and immunogenicity of the novel vaccine hVEGF 26-104 /RFASE., Methods: Patients with advanced solid malignancies with no standard treatment options available were eligible for this phase I study with a 3+3 dose-escalation design. On days 0, 14, and 28, patients received intramuscular hVEGF 26-104 , a truncated synthetic three-dimensional (3D)-structured peptide mimic covering the amino acids 26-104 of the human VEGF 165 isoform, emulsified in the novel adjuvant Raffinose Fatty Acid Sulphate Ester (RFASE), a sulpholipopolysaccharide. Objectives were to determine safety, induction of VEGF-neutralizing antibodies, and the maximum tolerated dose. Blood was sampled to measure VEGF levels and antibody titers., Results: Eighteen of 27 enrolled patients received three immunizations in six different dose-levels up to 1,000 μg hVEGF 26-104 and 40 mg RFASE. No dose-limiting toxicity was observed. Although in four patients an antibody titer against hVEGF 26-104 was induced (highest titer: 2.77 10 log), neither a reduction in VEGF levels nor neutralizing antibodies against native VEGF 165 were detected., Conclusion: Despite having an attractive safety profile, hVEGF 26-104 /RFASE was not able to elicit seroconversions against native VEGF 165 and, consequently, did not decrease circulating VEGF levels. Deficient RFASE adjuvant activity, as well as dominant immunoreactivity toward neoepitopes, may have impeded hVEGF 26-104 /RFASE's efficacy in humans., (© 2020 The Authors. The Oncologist published by Wiley Periodicals LLC on behalf of AlphaMed Press.)

Priming of CD8 + T cells by dendritic cells (DCs) is crucial for the generation of effective antitumor immune responses. Here, we describe a liposomal vaccine carrier that delivers tumor antigens to human CD169/Siglec-1 + antigen-presenting cells using gangliosides as targeting ligands. Ganglioside-liposomes specifically bound to CD169 and were internalized by in vitro-generated monocyte-derived DCs (moDCs) and macrophages and by ex vivo-isolated splenic macrophages in a CD169-dependent manner. In blood, high-dimensional reduction analysis revealed that ganglioside-liposomes specifically targeted CD14 + CD169 + monocytes and Axl + CD169 + DCs. Liposomal codelivery of tumor antigen and Toll-like receptor ligand to CD169 + moDCs and Axl + CD169 + DCs led to cytokine production and robust cross-presentation and activation of tumor antigen-specific CD8 + T cells. Finally, Axl + CD169 + DCs were present in cancer patients and efficiently captured ganglioside-liposomes. Our findings demonstrate a nanovaccine platform targeting CD169 + DCs to drive antitumor T cell responses., Competing Interests: The authors declare no competing interest., (Copyright © 2020 the Author(s). Published by PNAS.)

Background: Chemotherapy (CT), radiotherapy (RT), and chemoradiotherapy (CRT) are able to alter the composition of the tumor immune microenvironment (TIME). Understanding the effect of these modalities on the TIME could aid in the development of improved treatment strategies. Our aim was to systematically review studies investigating the influence of CT, RT or CRT on different TIME markers., Methods: The EMBASE (Ovid) and PubMed databases were searched until January 2019 for prospective or retrospective studies investigating the dynamics of the local TIME in cancer patients (pts) treated with CT, RT or CRT, with or without targeted agents. Studies could either compare baseline and follow-up specimens - before and after treatment - or a treated versus an untreated cohort. Studies were included if they used immunohistochemistry and/or flow cytometry to assess the TIME., Results: In total we included 110 studies (n = 8850 pts), of which n = 89 (n = 6295 pts) compared pre-treatment to post-treatment specimens and n = 25 (n = 2555 pts) a treated versus an untreated cohort (4 studies conducted both comparisons). For several tumor types (among others; breast, cervical, esophageal, ovarian, rectal, lung mesothelioma and pancreatic cancer) remodeling of the TIME was observed, leading to a potentially more immunologically active microenvironment, including one or more of the following: an increase in CD3 or CD8 lymphocytes, a decrease in FOXP3 Tregs and increased PD-L1 expression. Both CT and CRT were able to immunologically alter the TIME., Conclusion: The TIME of several tumor types is significantly altered after conventional therapy creating opportunities for concurrent or sequential immunotherapy., Competing Interests: Declaration of Competing Interest Drs. Tom van den Ende, Héctor van den Boorn, Nadine Hoonhout, Faridi van Etten-Jamaludin, Dr. Sybren Meijer and Dr. Sarah Derks have nothing to disclose. Prof. Tanja de Gruijl reports stock ownership of LAVA therapeutics, consulting work for Macrophage Pharma, Partner Therapeutics DCPrime and research funding from Idera outside the submitted work. Dr. Bijlsma reports an advisory role for Servier outside the submitted work. Dr. Martijn van Oijen reports grants from Roche, grants from Servier, grants from Nordic, grants from Merck, grants from Amgen, outside the submitted work. Prof. Hanneke van Laarhoven reports grants from Bristol-Myers Squibb, grants from Bayer Schering Pharma, grants from Celgene, grants from Janssen-Cilag, grants from Lilly, grants from Nordic Group, grants from Philips Healthcare, grants from Roche, grants from Merck Sharp and Dohme, personal fees from Lilly, personal fees from AstraZeneca, personal fees from Lilly, personal fees from Nordic, personal fees from Bristol-Myers Squibb, outside the submitted work., (Copyright © 2020 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background: A recent randomized phase II trial evaluated stereotactic ablative radiotherapy (SABR) in a group of patients with a small burden of oligometastatic disease (mostly with 1-3 metastatic lesions), and found that SABR was associated with a significant improvement in progression-free survival and a trend to an overall survival benefit, supporting progression to phase III randomized trials., Methods: Two hundred and ninety-seven patients will be randomized in a 1:2 ratio between the control arm (consisting of standard of care [SOC] palliative-intent treatments), and the SABR arm (consisting of SOC treatment + SABR to all sites of known disease). Randomization will be stratified by two factors: histology (prostate, breast, or renal vs. all others), and disease-free interval (defined as time from diagnosis of primary tumor until first detection of the metastases being treated on this trial; divided as ≤2 vs. > 2 years). The primary endpoint is overall survival, and secondary endpoints include progression-free survival, cost effectiveness, time to development of new metastatic lesions, quality of life (QoL), and toxicity. Translational endpoints include assessment of circulating tumor cells, cell-free DNA, and tumor tissue as prognostic and predictive markers, including assessment of immunological predictors of response and long-term survival., Discussion: This study will provide an assessment of the impact of SABR on survival, QoL, and cost effectiveness to determine if long-term survival can be achieved for selected patients with 1-3 oligometastatic lesions., Trial Registration: Clinicaltrials.gov identifier: NCT03862911. Date of registration: March 5, 2019.

This review summarizes the use of high-voltage electrical pulses (HVEPs) in clinical oncology to treat solid tumors with irreversible electroporation (IRE) and electrochemotherapy (ECT). HVEPs increase the membrane permeability of cells, a phenomenon known as electroporation. Unlike alternative ablative therapies, electroporation does not affect the structural integrity of surrounding tissue, thereby enabling tumors in the vicinity of vital structures to be treated. IRE uses HVEPs to cause cell death by inducing membrane disruption, and it is primarily used as a radical ablative therapy in the treatment of soft-tissue tumors in the liver, kidney, prostate, and pancreas. ECT uses HVEPs to transiently increase membrane permeability, enhancing cellular cytotoxic drug uptake in tumors. IRE and ECT show immunogenic effects that could be augmented when combined with immunomodulatory drugs, a combination therapy the authors term electroimmunotherapy . Additional electroporation-based technologies that may reach clinical importance, such as gene electrotransfer, electrofusion, and electroimmunotherapy, are concisely reviewed. HVEPs represent a substantial advancement in cancer research, and continued improvement and implementation of these presented technologies will require close collaboration between engineers, interventional radiologists, medical oncologists, and immuno-oncologists., (© RSNA, 2020.)

Lymph nodes draining the primary tumor are essential for the initiation of an effective anti-tumor T-cell immune response. However, cancer-derived immune suppressive factors render the tumor-draining lymph nodes (TDLN) immune compromised, enabling tumors to invade and metastasize. Unraveling the different mechanisms underlying this immune escape will inform therapeutic intervention strategies to halt tumor spread in early clinical stages. Here, we review our findings from translational studies in melanoma, breast, and cervical cancer and discuss clinical opportunities for local immune modulation of TDLN in each of these indications.

Background: Stereotactic ablative radiotherapy (SABR) has emerged as a new treatment option for patients with oligometastatic disease. SABR delivers precise, high-dose, hypofractionated radiotherapy, and achieves excellent rates of local control for primary tumors or metastases. A recent randomized phase II trial evaluated SABR in a group of patients with a small burden of oligometastatic disease (mostly with 1-3 metastatic lesions), and found that SABR was associated with benefits in progression-free survival and overall survival. The goal of this phase III trial is to assess the impact of SABR in patients with 4-10 metastatic cancer lesions., Methods: One hundred and fifty-nine patients will be randomized in a 1:2 ratio between the control arm (consisting of standard of care palliative-intent treatments), and the SABR arm (consisting of standard of care treatment + SABR to all sites of known disease). Randomization will be stratified by two factors: histology (Group 1: prostate, breast, or renal; Group 2: all others), and type of pre-specified systemic therapy (Group 1: immunotherapy/targeted; Group 2: cytotoxic; Group 3: observation). SABR is to be completed within 2 weeks, allowing for rapid initiation of systemic therapy. Recommended SABR doses are 20 Gy in 1 fraction, 30 Gy in 3 fractions, or 35 Gy in 5 fractions, chosen to minimize risks of toxicity. The primary endpoint is overall survival, and secondary endpoints include progression-free survival, time to development of new metastatic lesions, quality of life, and toxicity. Translational endpoints include assessment of circulating tumor cells, cell-free DNA, and tumor tissue as prognostic and predictive markers, including assessment of immunological predictors of response and long-term survival., Discussion: This study will provide an assessment of the impact of SABR on clinical outcomes and quality of life, to determine if long-term survival can be achieved for selected patients with 4-10 oligometastatic lesions., Trial Registration: Clinicaltrials.gov identifier: NCT03721341 . Date of registration: October 26, 2018.

Innate immune cells are active at the front line of host defense against pathogens and now appear to play a range of roles under non-infectious conditions as well, most notably in cancer. Establishing the balance of innate immune responses is critical for the "flavor" of these responses and subsequent adaptive immunity and can be either "good or bad" in controlling cancer progression. The importance of innate NK cells in tumor immune responses has already been extensively studied over the last few decades, but more recently several relatively mono- or oligo-clonal [i.e., (semi-) invariant] innate T cell subsets received substantial interest in tumor immunology including invariant natural killer T (iNKT), γδ-T and mucosal associated invariant T (MAIT) cells. These subsets produce high levels of various pro- and/or anti-inflammatory cytokines/chemokines reflecting their capacity to suppress or stimulate immune responses. Survival of patients with cancer has been linked to the frequencies and activation status of NK, iNKT, and γδ-T cells. It has become clear that NK, iNKT, γδ-T as well as MAIT cells all have physiological roles in anti-tumor responses, which emphasize their possible relevance for tumor immunotherapy. A variety of clinical trials has focused on manipulating NK, iNKT, and γδ-T cell functions as a cancer immunotherapeutic approach demonstrating their safety and potential for achieving beneficial therapeutic effects, while the exploration of MAIT cell related therapies is still in its infancy. Current issues limiting the full therapeutic potential of these innate cell subsets appear to be related to defects and suppressive properties of these subsets that, with the right stimulus, might be reversed. In general, how innate lymphocytes are activated appears to control their subsequent abilities and consequent impact on adaptive immunity. Controlling these potent regulators and mediators of the immune system should enable their protective roles to dominate and their deleterious potential (in the specific context of cancer) to be mitigated.

There is a growing understanding of why certain patients do or do not respond to checkpoint inhibition therapy. This opens new opportunities to reconsider and redevelop vaccine strategies to prime an anticancer immune response. Combination of such vaccines with checkpoint inhibitors will both provide the fuel and release the brake for an efficient anticancer response. Here, we discuss vaccine strategies that use C-type lectin receptor (CLR) targeting of APCs, such as dendritic cells and macrophages. APCs are a necessity for the priming of antigen-specific cytotoxic and helper T cells. Because CLRs are natural carbohydrate-recognition receptors highly expressed by multiple subsets of APCs and involved in uptake and processing of Ags for presentation, these receptors seem particularly interesting for targeting purposes., (© The Author(s).)

Background: Only a small proportion of patients respond to anti-VEGF therapy, pressing the need for a reliable biomarker that can identify patients who will benefit. We studied the biological activity of anti-VEGF antibodies in patients' blood during anti-VEGF therapy by using the Ba/F3-VEGFR2 cell line, which is dependent on VEGF for its growth., Methods: Serum samples from 22 patients with cancer before and during treatment with bevacizumab were tested for their effect on proliferation of Ba/F3-VEGFR2 cells. Vascular endothelial growth factor as well as bevacizumab concentrations in serum samples from these patients were determined by enzyme linked immunosorbent assay (ELISA)., Results: The hVEGF-driven cell proliferation was effectively blocked by bevacizumab (IC 50 3.7 μg ml -1 ; 95% CI 1.7-8.3 μg ml -1 ). Cell proliferation was significantly reduced when patients' serum during treatment with bevacizumab was added (22-103% inhibition compared with pre-treatment). Although bevacizumab levels were not related, on-treatment serum VEGF levels were correlated with Ba/F3-VEGFR2 cell proliferation., Conclusions: We found that the neutralising effect of anti-VEGF antibody therapy on the biological activity of circulating VEGF can be accurately determined with a Ba/F3-VEGFR2 bioassay. The value of this bioassay to predict clinical benefit of anti-VEGF antibody therapy needs further clinical evaluation in a larger randomised cohort.

Purpose of Review: In this review, we focus on the recent findings and future challenges in cancer treatment with immune checkpoint inhibitors., Recent Findings: Major progress has been made in recent years as the first immune checkpoint inhibitors are approved by the US Food and Drug Administration for the treatment of cancer patients. Anticytotoxic T-lymphocyte-associated protein 4 and antiprogrammed death protein 1/programmed death-ligand 1 (PD-L1) monoclonal antibodies are being extensively studied in many different tumor types, often showing impressive response rates, but also a typical serious toxicity profile in the form of auto-immunity. Unfortunately, it is not yet possible to prevent or predict these immune-related adverse events. Studies on mutational load, neo-epitopes, lactate dehydrogenase, PD-L1 expression, and T-cell infiltration suggest that these markers are correlating with efficacy, but have not yet reached the status of a validated biomarker for checkpoint inhibitors. Other immune checkpoints are being investigated and new checkpoint inhibitors are on the brink of being evaluated in clinical trials., Summary: The main challenge for the near future will be to predict efficacy of immune checkpoint blockade and to predict and prevent immune-related adverse events. More research should be done in order to find potential biomarkers that predict treatment response and/or toxicity; the optimal administration route, dosage, and frequency; and possible combinations of therapies that have an added or synergetic effect.

Introduction: Over the past decade, the cytolytic capabilities of oncolytic viruses (OVs), exploited to selectively eliminate neoplastic cells, have become secondary to their use to elicit a tumor-directed immune response., Areas Covered: Here, based on an NCBI-PubMed literature survey, we review the efforts undertaken to arm OVs in order to improve therapeutic antitumor responses upon administration of these agents. Specifically, we explore the different options to modulate immune suppression in the tumor microenvironment (TME) and to facilitate the generation of effective antitumor responses that have been investigated in conjunction with OVs in recent years., Expert Opinion: Their induction of immunogenic tumor cell death and association with pro-inflammatory signals make OVs attractive immunotherapeutic modalities. The first promising clinical results with immunologically armed OVs warrant their further optimization and development. OVs should be modified to avoid detrimental effects of pre-existent anti-OV immunity as well as for increased tumor targeting and selectivity, so as to ultimately allow for systemic administration while achieving local immune potentiation and tumor elimination in the TME. In particular, a combination of trans-genes encoding bispecific T-cell engagers, immune checkpoint blockers and antigen-presenting cell enhancers will remove suppressive hurdles in the TME and allow for optimal antitumor efficacy of armed OVs.

Improvement of patient survival by anti-angiogenic therapy has proven limited. A vaccination approach inducing an immune response against the tumor vasculature combines the benefits of immunotherapy and anti-angiogenesis, and may overcome the limitations of current anti-angiogenic drugs. Strategies to use whole endothelial cell vaccines and DNA- or protein vaccines against key players in the VEGF signaling axis, as well as specific markers of tumor endothelial cells, have been tested in preclinical studies. Current clinical trials are now testing the promise of this specific anti-cancer vaccination approach. This review will highlight the state-of-the-art in this exciting field of cancer research., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Over the past decades advances in bioengineering and expanded insight in tumor immunology have resulted in the emergence of novel bispecific antibody (bsAb) constructs that are capable of redirecting immune effector cells to the tumor microenvironment. (Pre-) clinical studies of various bsAb constructs have shown impressive results in terms of immune effector cell retargeting, target dependent activation and the induction of anti-tumor responses. This review summarizes recent advances in the field of bsAb-therapy and limitations that were encountered. Furthermore, we will discuss potential future developments that can be expected to take the bsAb approach successfully forward., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

The ability of dendritic cells (DCs) to orchestrate innate and adaptive immune responses has been exploited to develop potent anti-cancer immunotherapies. Recent clinical trials exploring the efficacy of ex vivo modified autologous DC-based vaccines have reported some promising results. However, in vitro generation of autologous DCs for clinical administration, their loading with tumor associated antigens (TAAs) and their activation, is laborious and expensive, and, as a result of inter-individual variability in the personalized vaccines, remains poorly standardized. An attractive alternative approach is to load resident DCs in vivo by targeted delivery of TAAs, using viral vectors and activating them simultaneously. To this end, we have constructed genetically-modified adenoviral (Ad) vectors and bispecific adaptor molecules to retarget Ad vectors encoding TAAs to the CD40 receptor on DCs. Pre-clinical human and murine studies conducted so far have clearly demonstrated the suitability of a 'two-component' (i.e. Ad and adaptor molecule) configuration for targeted modification of DCs in vivo for cancer immunotherapy. This review summarizes recent progress in the development of CD40-targeted Ad-based cancer vaccines and highlights pre-clinical issues in the clinical translation of this approach., (Copyright © 2012 John Wiley & Sons, Ltd.)

Vγ9Vδ2-T cells constitute a proinflammatory lymphocyte subpopulation with established antitumor activity. Phosphoantigens activate Vγ9Vδ2-T cells in vivo and in vitro. We studied whether the antitumor activity of Vγ9Vδ2-T cells can be potentiated by invariant NKT cells (iNKT), an important immunoregulatory T cell subset. When activated by the glycolipid α-galactosylceramide (α-GalCer), iNKT produce large amounts of cytokines involved in antitumor immune responses. Monocyte-derived dendritic cells were loaded with both phosphoantigens (using aminobisphosphonates) and α-GalCer during maturation and subsequently co-cultured with Vγ9Vδ2-T and iNKT cells. Aminobisphosphonates dose-dependently enhanced Vγ9Vδ2-T cell activation, and this was potentiated by α-GalCer-induced iNKT co-activation. iNKT co-activation also enhanced the IFN-γ production and cytolytic potential of Vγ9Vδ2-T cells against tumor cells. Using transwell experiments and neutralizing antibodies cross-talk between iNKT and Vγ9Vδ2-T cells was found to be mediated by TNF-α. Our data provide a rationale for combining both activating ligands to improve Vγ9Vδ2-T cell based approaches in cancer-immunotherapy., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Appropriate activation of dendritic cells (DC) is essential for successful active vaccination and induction of cell-mediated immunity. The scarcity of precursor cells, as well as long culture methods, have hampered wide-scale application of DC vaccines derived from CD34(+) precursors, despite their suggested superior efficacy over the more commonly applied monocyte-derived DC (MoDC). Here, employing the CD34(+)/CD14(+) AML-derived human DC progenitor cell line MUTZ3, we show that cytostatic anthraquinone-derivatives (i.e., the anthracenedione mitoxantrone and the related anthracyclin doxorubicin) induce rapid differentiation of CD34(+) DC precursors into functional antigen-presenting cells (APC) in a three-day protocol. The drugs were found to act specifically on CD34(+), and not on CD14(+) DC precursors. Importantly, these observations were confirmed for primary CD34(+) and CD14(+) DC precursors from peripheral blood. Mitoxantrone-generated DC were fully differentiated within three days and after an additional 24 h of maturation, were as capable as standard 9-day differentiated and matured DC to migrate toward the lymph node-homing chemokines CCL19 and CCL21, to induce primary allogeneic T cell proliferation, and to prime functional MART1-specific CD8(+) T lymphocytes. Our finding that anthraquinone-derivatives like mitoxantrone support rapid high-efficiency differentiation of DC precursors may have consequences for in vitro production of DC vaccines as well as for novel immunochemotherapy strategies.

Progress in vector design and an increased knowledge of mechanisms underlying tumor-induced immune suppression have led to a new and promising generation of Adenovirus (Ad)-based immunotherapies, which are discussed in this review. As vaccine vehicles Ad vectors (AdVs) have been clinically evaluated and proven safe, but a major limitation of the commonly used Ad5 serotype is neutralization by preexistent or rapidly induced immune responses. Genetic modifications in the Ad capsid can reduce intrinsic immunogenicity and facilitate escape from antibody-mediated neutralization. Further modification of the Ad hexon and fiber allows for liver and scavenger detargeting and selective targeting of, for example, dendritic cells. These next-generation Ad vaccines with enhanced efficacy are now becoming available for testing as tumor vaccines. In addition, AdVs encoding immune-modulating products may be used to convert the tumor microenvironment from immune-suppressive and proinvasive to proinflammatory, thus facilitating cell-mediated effector functions that can keep tumor growth and invasion in check. Oncolytic AdVs, that selectively replicate in tumor cells and induce an immunogenic form of cell death, can also be armed with immune-activating transgenes to amplify primed antitumor immune responses. These novel immunotherapy strategies, employing highly efficacious AdVs in optimized configurations, show great promise and warrant clinical exploration., (Copyright © 2012 Elsevier Inc. All rights reserved.)

For over a century, research has sought ways to boost the immune system in order to eradicate tumors and viruses that exist after escaping immunosurveillance. For the treatment of cancer and hepatitis immunotherapeutic strategies have overall had limited clinical success. An urgent need exists therefore to introduce more effective therapeutic approaches. Invariant (i)NKT cells constitute a conserved T lymphocyte lineage with dominant immunoregulatory, antitumor and antiviral effector cell properties. iNKT specifically recognize the glycolipid α-galactosylceramide in the context of CD1d resulting in their activation. Activated iNKT can promote the development of a long-lasting Th1 biased proinflammatory immune response as demonstrated in multiple tumor-metastasis and viral infection models. Here, we will provide a brief overview of the preclinical data of α-galactosylceramide that formed the basis for subsequent clinical trials in patients with advanced cancer and chronic hepatitis B/C, and elaborate on our own clinical experience with α-galactosylceramide in these patient groups., (Copyright © 2010 Elsevier Inc. All rights reserved.)

Myeloid differentiation is often disturbed in cancer, leading to reduced frequencies of immunostimulatory dendritic cells and an over-representation of immunosuppressive immature myeloid cells, granulocytes and macrophages. As a result of this skewed myeloid differentiation, a highly immunosuppressive myeloid subset becomes prevalent during cancer development; these myeloid-derived suppressor cells are also recruited as a collateral to certain protumorigenic inflammatory processes, resulting in an effective downregulation of T-cell-mediated immune surveillance and antitumor immunity. In this article, some of the important myeloid cell subsets and mediators involved in cancer-related immune suppression are reviewed. Furthermore, cross-talk between tumors and the myeloid compartment, and ways in which it can suppress effective cell-mediated immunity, are discussed, as well as possible therapeutic approaches to tip the balance in favor of antitumor immunity.

ABC transporters were identified originally for their contribution to clinical MDR as a result of their capacity to extrude various unrelated cytotoxic drugs. More recent reports have shown that ABC transporters can play important roles in the development, differentiation, and maturation of immune cells and are involved in migration of immune effector cells to sites of inflammation. Many of the currently identified, endogenous ABC transporter substrates have immunostimulating effects. Increasing the expression of ABC transporters on immune cells and thereby enhancing immune cell development or functionality may be beneficial to immunotherapy in the field of oncology. On the contrary, in the treatment of autoimmune diseases, blockade of these transporters may prove beneficial, as it could dampen disease activity by compromising immune effector cell functions. This review will focus on the expression, regulation, and substrate specificity of ABC transporters in relation to functional activities of immune effector cells and discusses implications for the treatment of cancer on the one hand and autoimmune diseases on the other.

ATP-binding cassette (ABC) transporters are known for their involvement in clinical multidrug resistance (MDR) and their physiological defensive functions in barrier organs. More recently, attention has been focused on their possible involvement in the regulation of immune responses following the identification of their substrates as known immunomodulating agents (e.g. prostaglandins, leukotrienes and cyclic nucleotides) and their functional expression in various immune effector cells, most notably in dendritic cells (DCs). This review addresses the possible roles of ABC transporters in DC development and function, as well as the putative immunostimulatory potential of their cytostatic substrates and how this knowledge might benefit DC-based chemo-immunotherapies.

The field of tumor vaccination is currently undergoing a shift in focus, from individualized tailor-made vaccines to more generally applicable vaccine formulations. Although primarily predicated by financial and logistic considerations, stemming from a growing awareness that clinical development for wide-scale application can only be achieved through backing from major pharmaceutical companies, these new approaches are also supported by a growing knowledge of the intricacies and minutiae of antigen presentation and effector T-cell activation. Here, the development of whole-cell tumor and dendritic cell (DC)-based vaccines from an individualized autologous set-up to a more widely applicable allogeneic approach will be discussed as reflected by translational studies carried out over the past two decades at our laboratories and clinics in the vrije universiteit medical center (VUmc) in Amsterdam, The Netherlands.

Dendritic cells (DC) are increasingly applied as a cellular adjuvant in immunotherapy of cancer. Two major myeloid DC subsets are recognized: interstitial DC (IDC) that infiltrate connective tissues and Langerhans cells (LC) that line epithelial surfaces. Yet, functional differences between IDC and LC remain to be defined. We recently showed that the CD34(+) acute myeloid leukemia cell line MUTZ-3 supports differentiation of both DC-SIGN(+) IDC and Langerin-positive Birbeck granule-expressing LC. By comparative functional characterization of MUTZ-3 IDC and MUTZ-3 LC, we aimed to elucidate the relative abilities of these two DC subsets to induce a specific T cell response and reveal the more suitable candidate for use as a clinical vehicle of tumor vaccines. Although mature LC and IDC displayed comparable lymph node-homing potential, mature LC showed higher allogeneic T cell stimulatory capacity. Nevertheless, IDC supported the induction of tumor Ag-specific CD8(+) T cells at an overall higher efficiency. This might be related to the observed inability of LC to release T cell stimulatory cytokines such as IL-12p70, IL-23, and IL-15. Although this inability did not result in a detectable deviation in the cytokine expression profile of primed T cells, transduction with IL-12p70 significantly improved priming efficiency of LC, and ensured a functional equivalence with IDC in this regard. In conclusion, except for the inability of LC to release distinct type 1 T cell stimulatory cytokines, in vitro function of LC and IDC suggests comparable abilities of both subsets for the in vivo induction of antitumor T cells.

The adoptive transfer of in vitro-induced and expanded tumor-specific cytotoxic T lymphocytes (CTL) presents a promising immunotherapeutic approach for the treatment of cancer. The in vitro induction of tumor-reactive CTL requires repeated stimulation of CTL precursors with dendritic cells (DC). To circumvent problems like scarcity of blood DC precursors and donor variability, it would be attractive to use DC from a non-autologous, unlimited source. DCs derived from the human acute myeloid leukemia (AML) cell line MUTZ-3 are attractive candidates since these DCs closely resemble monocyte-derived DC (MoDC) in terms of phenotype and T cell stimulatory capacity. Here we demonstrate that functional CTL clones could be generated against multiple tumor-associated antigens, i.e., human telomerase reverse transcriptase (hTERT), ErbB3-binding protein-1 (Ebp1), carcinoembryonic antigen (CEA) and Her-2/neu, by stimulating CD8beta(+) CTL precursors with peptide-loaded allogeneic, HLA-A2-matched MUTZ-3-derived DC. A consistent induction capacity, as determined by MHC tetramer-binding, was found in multiple donors and comparable to autologous peptide-loaded MoDC. Functional characterization at the clonal level revealed the priming of CTL that recognized endogenously processed epitopes on tumor cell lines in an HLA-A2-restricted fashion. Our data indicate that MUTZ-3-derived DC can be used as stimulator cells for in vitro priming and expansion of functional TAA-specific effector CTL. MUTZ-3-derived DCs thus represent a ready and standardized source of allogeneic DC to generate CTL for therapeutic adoptive transfer strategies.

Immune responses against tumor antigens will initially occur in the first tumor-draining lymph node, the sentinel node (SN). Because of extensive diagnostic procedures, obtaining a piece of SN to isolate viable immune cells for functional analyses is often impossible. For this reason an alternative method to obtain viable cells from a lymph node (LN) was investigated, ie, scraping LNs with a surgical blade, and compared with dissociation of total LNs. Tumor-draining lymph nodes were retrieved from five oncological patients. The collected dendritic cells and T cells were phenotypically and functionally characterized by flow cytometry and antigen-specific interferon (IFN)-gamma release in an ELISPOT assay. Results were compared between the two isolation methods. Viabilities and phenotypic characteristics of the collected cells were entirely comparable for both methods. T-cell functionality was also comparable between both methods, with equal T-cell expansion factors and similar frequencies of cytotoxic T cells specifically recognizing the M1 matrix protein of Influenza haemophilus or the tumor antigen Her-2/neu. In conclusion, scraping LNs to obtain cells for analysis of immune functions in LNs is feasible and presents a good alternative to dissociation of LNs. Scraping may even be applied to small LNs that a pathologist will submit entirely for histological examination and may thus prove useful in the monitoring of immune responses in SNs.

Production of immunosuppressive factors is one of the mechanisms by which tumors evade immunosurveillance. Soluble factors hampering dendritic cell (DC) development have recently been identified in culture supernatants derived from tumor cell lines. In this study, we investigated the presence of such factors in 24-h culture supernatants from freshly excised solid human tumors (colon, breast, renal cell carcinoma, and melanoma). While primary tumor-derived supernatant (TDSN) profoundly hampered the in vitro DC differentiation from CD14(+) plastic-adherent monocytes or CD34(+) precursors (based on morphology and CD1a/CD14 phenotype), the effects of tested tumor cell line-derived supernatants were minor. Cyclooxygenase (COX)-1- and COX-2-regulated prostanoids present in the primary TDSN were found to be solely responsible for the observed hampered differentiation of monocyte-derived DC (MoDC). In contrast, both prostanoids and IL-6 were found to contribute to the TDSN-induced inhibition of DC differentiation from CD34(+) precursor cells. While the addition of TDSN during differentiation interfered with the ability of CD34-derived DC to stimulate a primary allogeneic T cell response, it actually increased this ability of MoDC. These opposite effects were correlated to different effects of the TDSN on the expression levels of CD86 and HLA-DR on the DC from the different precursor origins. Although TDSN increased the T cell-stimulatory capacity of MoDC, TDSN inhibited the IL-12 production and increased the IL-10 production of MoDC, thus skewing them to a type-2 T cell-inducing phenotype. In conclusion, this study demonstrates that primary tumors negatively impact DC development and function through COX-1 and -2 regulated factors, whereas tumor-derived cell lines may lose this ability upon in vitro propagation.