the 30th Annual Conference of the

21-23 September 2016 Amsterdam, The Netherlands www.emds2016-amsterdam.nl

Dendritic cells and macrophages: Distant relatives or just close neighbours?

Photo Source: Sriram Subramaniam National Cancer Institute (NCI).

Welcome in Amsterdam

“To be or not to be a macrophage or a dendritic cell…is that the question?”

We wish everybody fruitful discussions!

30th Annual Conference of the EMDS, September 21st-23rd, Amsterdam, The Netherlands

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Contents Welcoming address

4

Local Organizing Scientific Committee and Conference Agency

5

The European Macrophage and Dendritic Cell Society (EMDS)

6

Sponsors

7

General Information

8

Conference Venue

9

Party information

10

Invited Speakers

12

Scientific Programme

13

Abstracts of oral presentations

17

Abstracts of poster presentations

33

Participants Contact List

82

Schedule of Poster Walks

87

30th Annual Conference of the EMDS, September 21st-23rd, Amsterdam, The Netherlands

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A word of welcome from the conference chairs Dear colleagues and friends, On behalf of the Organizing and Scientific Committee it is our great pleasure to welcome you to the 30 th Annual Meeting of the European Macrophage and Dendritic Cell Society (EMDS). We are delighted that you decided to join us in Amsterdam. Amsterdam is a great small metropole. The city has a rich historic culture, many museums, wide gastronomic diversity, good transportation and the nightlife of a big city. It is however small enough to see most things by foot, or by bike, which is the most popular form of transportation in the Netherlands. Going around by boat is a great way to see the city from the perspective of its extensive canals. Do not miss any of Amsterdam’s famous museums, such as the Rijksmuseum, - the home of the Nightwatch by Rembrandt -, the Van Gogh Museum or the Anne Frank House, which is located close to the Rode Hoed, where the EMDS is organized. Additionally, Amsterdam is a hotspot of science in the Netherlands. Three top institutes – the VU University Medical Center/Cancer Center Amsterdam, Academic Medical Center, University of Amsterdam and Sanquin Research – have joined forces to organise this exciting 30th edition of the annual EMDS meeting. Basic aspects, such as control of APC function, metabolism, signalling, heterogeneity, antigen processing & presentation, and microbe-host crosstalk will be discussed. Additionally, many clinical aspects of APC dysfunction in chronic inflammatory disorders and cancer will be addressed as well as the possibility to target dendritic cells and/or macrophages in therapeutic strategies. We are honoured by the 13 eminent scientists who have accepted the invitation to present their latest scientific insights during this meeting, and feel privileged that 115 abstracts have been submitted. Over 200 participants from 13 countries, including Australia and China, will participate in the meeting, many of which will share their data in either an oral presentation or during the poster walks. There is ample time to interact and exchange views during the coffee breaks, lunches and the congress party. After all, ‘To be a macrophage or a dendritic cell…’ Does it really matter? We wish you an enjoyable, satisfying, interesting scientific and socially interactive meeting. On behalf of the local Organizing and Scientific Committee,

Marjolein van Egmond

Yvett e van Kooyk

30th Annual Conference of the EMDS, September 21st-23rd, Amsterdam, The Netherlands

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Local Organizing and Scientific Committee Anja ten Brinke

Sanquin

Marjolein van Egmond

VUmc-CCA

Tanja de Gruijl

VUmc-CCA

Joke den Haan

VUmc-CCA

Marieke van Ham

UvA, Sanquin

Esther de Jong

AMC, UvA

Yvette van Kooyk

VUmc-CCA

Arjan van de Loosdrecht

VUmc-CCA

Sandra van Vliet

VUmc-CCA

Conference Agency Congres Office De Vlinder, Nieuw Bergen, The Netherlands [email protected]

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The European Macrophage and Dendritic Cell Society (EMDS) The European Macrophage and Dendritic Cell Society (EMDS) has emerged from the activities of the former European Macrophage Study Group (EMSG), a loose association of scientists interested in basic and clinical aspects of monocytes, macrophages, dendritic cells and other myleoid cells in man and experimental animal models. The group was constituted in 1992 as a result of a successful series of annual conferences called THE MACROPHAGE. The annual macrophage conference originated from meetings in the Upper Rhine area organized by scientists from the universities and research centers of Strasbourg, Freiburg and Basel, but rapidly grew to a European format. On April 28, 1999, the European Macrophage Society (EMS) was founded in Regensburg. At the end of year 2000, the members of the EMS decided to rename the society as European Macrophage and Dendritic Cell Society (EMDS) in order to better emphasize the two main streams of research within the Society. The EMDS currently has 485 members from 34 countries. For more information and registration as a member, please see our website: www.macrophage.de

Council of the EMDS

Advisory Board of the EMDS

President Guenter Weiss, Innsbruck, Austria

Reinhard Andreesen, Regensburg, Germany Robert HJ Beelen, Amsterdam, The Netherlands Christian Bogdan, Erlangen, Germany Patrick De Baetselier, Brussels, Belgium Antonio Celada, Barcelona, Spain Thomas Decker, Vienna, Austria Jean-Claude Drapier, Gif-sur-Yvette, France Hans D. Flad, Freiburg, Germany Regine Landmann, Basel, Switzerland Michael Rehli, Regensburg, Germany Silvano Sozzani, Brescia, Italy Andreas Spittler, Wien, Austria Alexander Steinkasserer, Erlangen, Germany Jo Van Ginderachter, Brussels, Belgium Loems Ziegler-Heitbrock, Gauting, Germany

Officers Amaya Puig Kröger, Madrid, Geert Raes, Brussels, Belgium Maciek Siedlar, Krakow, Poland Treasurer Ulrike Schleicher, Erlangen, Germany Secretary Luigi Varesio, Genova, Italy

30th Annual Conference of the EMDS, September 21st-23rd, Amsterdam, The Netherlands

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Thanks to our sponsors Platinum

Gold

Bronze

Sponsors

30th Annual Conference of the EMDS, September 21st-23rd, Amsterdam, The Netherlands

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General Information

Registration desk The registration desk is located on the ground level of the ‘Rode Hoed’. Opening hours: Wednesday, September 21st: 08:00 am-17:30 pm Thursday, September 22nd: 8:00 am- 17:30 pm Friday, September 23rd: 8:00 am- 13:30 pm

Oral presentations Exactly 15 minutes, including discussion, are allocated for each oral presentation. Please stay within the allocation time. Oral presenters are requested to upload their presentation 20 minutes before the start of their session on the computer of the Rode Hoed, which is present on the first balcony. There is no possibility of using your own computer during the presentation.

Poster presentations The posters session will take place at the 2 balconies of the Rode Hoed. Please set up your poster before the start of the first session of the day of your poster presentation, and remove the poster at the end of the day. Poster boards are 2 meters high and 1 meter wide, and will be numbered. Material to attach the poster to the poster boards will be available. Posters can be viewed during the coffee breaks and lunch. Additionally, please be on time for the scheduled poster walk in which you have the opportunity to present your poster in 3 minutes, after which it can be discussed. The poster walk are scheduled: Wednesday, September 21st: 11:30-12.30 pm Thursday, September 22nd: 11:00-12:00 am

Coffee breaks and lunch Coffee/ tea and lunch will be served on the ground floor and the first balcony.

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Conference Venue De Rode Hoed Keizersgracht 102, 1015 CV Amsterdam

Central Station Congress venue Rode Hoed

WIFI network: derodehoed Password: rodehoed1987

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Party information Conference Party with EMDS and Society of Leukocyte Biology Travel Awards and Walking Dinner at the “IJ-kantine” As local organizing committee we have selected a program for you by which you can experience the different aspects of Amsterdam, from the historical canals to the trendy and industrial northern shore of the IJ. At 17.45 hrs. at the dock in front of the Rode Hoed (the conference venue) a canal boat will pick us up. After a cruise through the canals of Amsterdam, we will cross the IJ were we will depart the boat. We will arrive at the IJ-kantine around 1900. IJ-kantine is located at northern shore of the IJ. From the industrial and spacious place of the IJkantine you can enjoy the beautiful view looking out over the harbor of Amsterdam and the water of the IJ. You can feel yourself far away while being close to the city center. At the IJ-kantine a walking dinner will be served, giving you plenty of time for socializing and dancing with the other conference participants. Dinner and the first 3 drinks are included. Together with DJ John Valk we will try to make it a conference party you will not easily forget. The party ends around 2400. Return to the city center of Amsterdam is your own responsibility and will not be offered by the conference organization. From the NDSM dock, situated immediately next to the IJ-kantine, a free ferry departs to the Center of Amsterdam and will take you in about 10 minutes to the dock at the back of the Central station. The ferry is leaving 2 times an hour at the half and whole hour, with the last one leaving exactly at 2400.

Address: Restaurant-Café IJ-kantine MT. Ondinaweg 15-17 1033 RE Amsterdam (NDSM-WERF)

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Invited speakers Joachim Schultze, Limes Institute, University of Bonn, Bonn, Germany Menno de Winther, Amsterdam Medical Center, University of Amsterdam, Netherlands Marc Raaijmakers, Erasmus Medical Center, Rotterdam, Netherlands Edward Pearce Max Planck Institute of Immunobiology and Epgenetics, Freiburg, Germany Mihai Netea, Radboud University Medical Center, Nijmegen, Netherlands Paul Kubes, Snyder Institute for Chronic Diseases, University of Calgary, Canada Marjolein van Egmond, VU University Medical Center, Amsterdam, Netherlands Peter Cresswell, Howard Hughes Medical Institute, Yale University School of Medicine, New Haven, USA Esther de Jong, Amsterdam Medical Center, University of Amsterdam, Netherlands Muzlifah Haniffa, Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, UK Sjoerd van der Burg, Leiden University Medical Center, Leiden, Netherlands Kris Thielemans, Free University, Brussels, Belgium Benjamin Marsland, University Hospital of Lausanne, Lausanne, France

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EMDS 2016 Program Wednesday 21 September 8.30

Registration

9.15

Opening by Marjolein van Egmond

9.30- 11.00

Session

I:

Control

of

APC

function

(chairs: Joachim Schultze and Joke den Haan)

9.30- 10.00

Joachim Schultze next?

The multi-dimensional model of macrophage activation: What’s

10.00-10.15

Carla Ribeiro Distinct signaling by C-type lectin receptors dictates HIV-1 restriction by E3 ligase TRIM5α in human dendritic cell subsets

10.15-10.30

Gera Goverse Interaction between enteric glia and myeloid cells as critical players in intestinal immune homeostasis

10.30-11.00

Menno de Winther

11.00-11.30

Coffee break and poster viewing

11.30- 12.30

Guided posterwalks

12.00-13.00

EMDS council meeting

12.30-13.30

Lunch and poster viewing

13.30-15.00

Session

Epigenetic pathways regulating macrophage function

II:

Chronic

inflammatory

disorders

(chairs: Marc Raaijmakers and Arjan van de Loosdrecht)

13.30-14.00

Marc Raaijmakers

The inflammatory niche in myelodysplastic syndromes

14.00-14.15

Roham Parsa BAFF-secreting neutrophils drive plasma cell responses during emergency granulopoiesis

14.15-14.30

Arjan Boltjes CD141+ conventional dendritic cells (cDC) are enriched at the site of autoimmune inflammation and display a regulatory phenotype.

14.30-14.45

Monika Linke Active mTORC1 signaling induces macrophage granuloma formation and sarcoidosis progression

14.45-15.00

Manon E. Wildenberg Benzimidazole anti-helminthics promote anti-TNF induced regulatory macrophage formation and potentiate therapeutic effect in a mouse model of inflammatory bowel disease

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15.00

Coffee break and poster viewing

15.30-17.15

Session

III:

Metabolism

and

Signaling

(chairs: Edward Pearce and Sandra van Vliet)

15.30-16.00

Mihai Netea

Immunometabolic circuits in innate immune responses

16.00-16.15

Leonard R Pelgrom by dendritic cells

Distinct metabolic requirements for T helper 1 and 2 polarization

16.15-16.30

Hweixian Leong Penny Warburg metabolism in tumor-conditioned macrophages promotes metastasis in human pancreatic ductal adenocarcinoma

16.30-16.45

Veronika Grau Alpha-1 antitrypsin inhibits ATP-dependent release of monocytic IL-1 via CD36, phospholipase A2 and nicotinic acetylcholine receptors

16.45-17.15

Edward Pearce activation

Metabolic reprogramming during plasmacytoid dendritic cell

17.15 End

Thursday 22 September 9.00- 10.30

Session

IV:

Macrophage

function

in

health

and

disease

(chairs: Paul Kubes and Anja ten Brinke)

9.00- 9.30

Paul Kubes

A novel macrophage recruitment paradigm

9.30-9.45

Geert Raes

Imaging of myeloid cell behaviour

9.45-10.00

Aoife Kelly Human peripheral blood monocytes and intestinal macrophage populations activate TGF-β via expression of the integrin αvβ8

10.00-10.30

Marjolein van Egmond Enlisting myeloid cells in the fight against cancer

10.30-11.00

Coffee break and poster viewing

11.00- 12.00

Guided posterwalks

12.00-13.00

Lunch and poster viewing

13.00- 14.30

Session

V:

Antigen

processing

and

presentation

(chairs: Peter Cresswell and Marieke van Ham)

13.00- 13.30

Peter Cresswell Mechanistic elements of conventional MHC class I antigen processing and cross-presentation

13.30-13.45

Katrien van der Borght Tissue necrosis is a driver of organ directed autoimmunity through activation of dendritic cells

13.45-14.00

Andrew S MacDonald A central role for Type I IFN in Th2 induction by dendritic cells

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14.00-14.15

Dieke van Dinther melanoma

14.15-14.30

Martijn Verdoes In vivo imaging of tumor-associated macrophages and subcellular compartment characterization in dendritic cells using novel quenched fluorescent cysteine cathepsin probes

14.30-15.00

Coffee break and poster viewing

15.00 – 16.45 Session

VI:

Development of a macrophage-based vaccination strategy for

Functional

heterogeneity

of

APCs

(chairs: Muzlifah Haniffa and Yvette van Kooyk)

15.00 – 15.30 Esther de Jong development

DCs, neutrophils and T cells: trinity for human Th17 cell

15.30-15.45

Rieneke van de Ven Correlating the ILT4/HLA-G inhibitory pathway to an immune suppressive microenvironment in sentinel lymph nodes draining head and neck squamous cell carcinoma

15.45-16.00

Anouk Zaal The anaphylatoxin C5a regulates the pro-inflammatory potential of human 6-sulfo LacNAc dendritic cells (slanDC) through crosstalk with TLR-induced ERK/p38/CREB signalling

16.00-16.15

Dorine Sichien IRF8 controls survival and function in terminally differentiated conventional and plasmacytoid dendritic cells respectively

16.15 – 16.45 Muzlifah Haniffa health and disease

Functional heterogeneity of human mononuclear phagocytes in

16.45-17.45

General Assembly (EMDS members only)

17.45-19.00

Canal cruise

19.00-24.00

Conference Party with EMDS and Society of Leukocyte Biology Travel Awards and Walking Dinner at the “IJkantine”

Friday 23 September 9.00- 10.45

Session

VII:

Clinical

applications

and

tumor

immunology

(chairs: Kris Thielemans and Tanja de Gruijl)

9.00- 9.30

Sjoerd van der Burg Chemotherapy driven modulation of intratumoral myeloid cells to improve immunotherapy

9.30-9.45

Mario Kuttke Myeloid PTEN deficiency impairs tumor immune surveillance via immune checkpoint inhibition

9.45-10.00

Camilla Salvagno Therapeutic synergy between macrophage blockade and chemotherapy in breast cancer elicits neutrophil-dependent therapy resistance

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10.00-10.15

Rebekka Wehner Accumulation of tolerogenic human 6-sulfo LacNAc+ dendritic cells in renal cell carcinoma is associated with poor prognosis

10.15- 10.45

Kris Thielemans

10.45-11.15

Coffee break

11.15- 12.45

Session

Dendritic cell and mRNA based immunotherapy

VIII:

Host-microbe

crosstalk

(chairs: Benjamin Marsland and Esther de Jong)

11.15-11.30

Ulrike Schleicher Suppression of arginase 1 by TNF facilitates the production of NO by type 2 NO synthase and is a novel mechanism by which TNF confers protection against Leishmania major parasites in vivo

11.30-11.45

Melissa Newling IgG opsonization of viruses functions as an endogenous suppressor of type I and III IFN-related anti-viral immunity by human myeloid cells through FcγRIIa

11.45-12.00

Joris K. Sprokholt Follicular T helper cell differentiation induced by Dengue virus through RIG-I like receptor and Type I Interferon receptor crosstalk

12.00-12.15

Late breaking abstract Diana Dudziak Human Lympoid tissue dendritic cells are less influenced by their microenvironment their counterparts in non-lymphoid tissues

12.15- 12.45

Benjamin Marsland

12.45

Closing remarks and awarding prizes by Marjolein van Egmond

13.00

End

Implications of the microbiota in airway inflammatory responses

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Abstracts Oral presentations Wednesday September 21st Poster number 2101

Carla Ribeiro Distinct signaling by C-type lectin receptors dictates HIV-1 restriction by E3 ligase TRIM5α in human dendritic cell subsets. 1

1

1

1

Carla M. S. Ribeiro , Ramin Sarrami-Forooshani , Laurentia C. Setiawan , Esther M. Zijlstra-Willems , John 1 2 2 1 1 L. van Hamme , Wikky Tigchelaar , Nicole van der Wel , Neeltje A. Kootstra , Sonja I. Gringhuis and 1 Teunis B. H. Geijtenbeek 1

Department of Experimental Immunology, Academic Medical Center, University of Amsterdam, 2 Amsterdam, the Netherlands Department of Cell Biology & Histology, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands Mucosal Langerhans cells (LCs) act as barriers against HIV-1 infection. HIV-1 capture by the C-type lectin receptor (CLR) langerin prevents infection of LCs. However, the underlying molecular mechanism of langerin-mediated HIV-1 restriction remains unknown. Here we show that E3 ligase TRIM5α potently + restricts HIV-1 infection of LCs but not of submucosal DC-SIGN dendritic cells (DCs). TRIM5α associates at steady-state with langerin via adaptor protein LSP-1 and mediates the recruitment of autophagic molecule Atg5 to TRIM5α-Atg16L1-HIV-1 capsid complexes upon HIV-1 exposure in LCs. The TRIM5αmediated autophagy-activating scaffolds targets langerin-HIV-1 capsid complexes into autophagic vesicles for degradation in LCs. Notably, HIV-1 infection of DCs leads to disassociation of TRIM5α from the DCSIGN signalosome and abrogates TRIM5α-mediated autophagic degradation. Remarkably, HIV-1 restriction by TRIM5α was thus far considered to be reserved for rhesus TRIM5α in monkeys but our data strongly indicate that human TRIM5α is a cell-specific restriction factor dependent on CLR signalling. Hence, our data strongly suggest that HIV-1 binding to either langerin or DC-SIGN dictates TRIM5α restriction or viral escape, respectively. Funding: Dutch Scientific Organization NWO (VENI 863.13.025)

2102

Gera Goverse Interaction between enteric glia and myeloid cells as critical players in intestinal immune homeostasis. G. Goverse, M. van Winge, M. Stakenborg, E. Labeeuw, M. Hao, P.J. Gomez-Pinilla, P. VandenBerghe, G.E. Boeckxstaens and G. Matteoli KU Leuven, Translational Research Center for Gastrointestinal Disorders (TARGID), Department of Clinical and eperimental medicine, Leuven, Belgium In the gastrointestinal tract a balance between immune activation and tolerance is essential to preserve intestinal homoeostasis. Recently we have demonstrated that stimulation of the enteric nervous system (ENS) has potent anti-inflammatory effects on macrophages (MFs). In the current study, we investigate if also enteric glial cells (EGCs), the major cellular constituent of the ENS, may have immunomodulatory properties. Immunohistochemical analysis revealed that EGCs are in close contact with MFs within all layers of the intestine. Moreover, we found that glial-secreted molecules were able to decrease expression of pro-inflammatory cytokines in MFs in response to LPS stimulation. In addition, typical antiinflammatory MF markers, such as IL-10, MRC-1 and MSR-1, were increased in MFs stimulated with EGC supernatant. Furthermore, analysis of the EGCs phenotype in vivo during inflammation revealed expression of several monocyte-chemoattractant proteins, as well as genes involved in the synthesis of

30th Annual Conference of the EMDS, September 21st-23rd, Amsterdam, The Netherlands

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the tolerogenic molecule retinoic acid. In conclusion, we showed that EGCs exert immunomodulatory effects on MFs, suggesting that these interactions might be crucial to maintain intestinal immune homeostasis and prevent immune-mediated diseases such as inflammatory bowel disease. Funding: FWO postdoctoral fellow

2108

Roham Parsa BAFF-secreting neutrophils drive plasma cell responses during emergency granulopoiesis. Dr. Roham Parsa , Mr. Harald Lund , Dr. Anna-Maria Georgoudaki , Dr. Xing-mei Zhang, Dr. André Ortlieb Guerreiro-Cacais , Mr. David Grommisch , Mr. Andreas Warnecke , Dr. Andrew L Croxford , Dr. Maja Jagodic , Prof. Burkhard Becher , Dr. Mikael C. I. Karlsson, Prof. Robert A. Harris Prolonged infections or adjuvant usage can trigger emergency granulopoiesis, leading to dysregulation in neutrophil blood counts. However, the impact of emergency granulopoiesis on T and B cell function remains largely unknown. To address this question we used a mouse model of neutropenia and studied immune activation following adjuvant administration. The initial neutropenic state fostered an environment of increased DC activation and T cell-derived IL-17 production. Interestingly, neutropenic LysM-DTA mice exhibited striking emergency granulopoiesis and amplified neutrophil recruitment to the lymph nodes that was dependent on IL-17-induced prostaglandin activity. The recruited neutrophils secreted BAFF that highly accelerated plasma cell generation and antigen-specific antibody production. Reduction of neutrophil functions via G-CSF neutralization significantly diminished plasma cell formation, directly linking emergency granulopoiesis with the humoral immune response. We conclude that neutrophils are capable of directly regulating T cell-dependent B cell responses in the lymph nodes.

2109

Arjan Boltjes CD141+ conventional dendritic cells (cDC) are enriched at the site of autoimmune inflammation and display a regulatory phenotype. 1

1

2

2

1

Arjan Boltjes , Michal Mokry , Bas Castelijns , Menno Creyghton and Femke van Wijk y 1

Laboratory for Translational Immunology, Wilhelmina Children’s Hospital, University Medical Center 2 Utrecht (UMC Utrecht), Utrecht, The Netherlands Hubrecht Institute, Utrecht, The Netherlands Dendritic cells (DC) are crucial in initiating and shaping immune responses. However, so far, little is known about the heterogeneity and functional specialization of human DC subsets, especially in (local) inflammatory conditions. In synovial fluid (SF), derived from inflamed joints of juvenile idiopathic arthritis (JIA) patients, we found a remarkable enrichment of CD141+ cDC. We therefore compared SF CD1c+ cDC, CD141+ cDC, and inflammatory monocytes based on phenotype, transcriptome, epigenome (enhancer activity determined with H3K27ac Chip-sequencing), and function. Phenotypically, SF CD1c+ cDC resembled monocytes, based on monocyte-associated markers, while CD141+ cDC selectively expressed CLEC9A and CD26. RNA-seq confirmed related transcriptional profiles between SF CD1c+ DC and SF monocytes. CD1c+ cDC were clearly superior in inducing T-cell proliferation, while all APC subsets were similarly capable of cross-presentation. Monocytes constitutively produced pro-inflammatory cytokines like IL-6 and TNF, but upon TLR induction also CD1c+ cDC showed a pro-inflammatory profile, which was confirmed by transcriptome and epigenome data. In contrast, CD141+ cDC revealed a more quiescent signature, and functional data with DC-subset co-cultures suggest a regulatory function. In conclusion, while CD1c+ cDC and monocytes at the site of inflammation show a related proinflammatory profile, enriched CD141+ cDC possess regulatory features, which emphasizes the differential programming of these subsets, especially in inflammation. Funding: VIDI grant form the Netherlands Organization for Scientific Research (NWO, ZonMw)

30th Annual Conference of the EMDS, September 21st-23rd, Amsterdam, The Netherlands

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2110

Monika Linke A Active mTORC1 signaling induces macrophage granuloma formation and sarcoidosis progression. 1

1

1

1

2

1

Monika Linke , Ha Thi Thanh Pham , Karl Katholnig , Thomas Schnöller , Anne Miller , Florian Demel , 1 1 3 3 4 5 Andrea Preitschopf , Boris Kovacic , Birgit Niederreiter , Stephan Blüml , Peter Kuess , Veronika Sexl , 6 1 7 2 8 Mathias Müller , Mario Mikula , Wolfram Weckwerth , Arvand Haschemi , Martin Susani , Markus 1 9 1 Hengstschläger , Michael J Gambello , Thomas Weichhart

1I

2

nstitute of Medical Genetics, Medical University of Vienna, 1090 Vienna, Austria Department of 3 Laboratory Medicine, Medical University of Vienna, 1090 Vienna, Austria Division of Rheumatology, 4 Internal Medicine III, Medical University of Vienna, 1090 Vienna, Austria Department of Radiation 5 Oncology, Medical University of Vienna, 1090 Vienna, Austria Institute of Pharmacology and Toxicology, 6 University of Veterinary Medicine, 1210 Vienna, Austria Biomodels Austria and Institute of Animal 7 Breeding and Genetics, University of Veterinary Medicine, 1210 Vienna, Austria Department of Ecogenomics and Systems Biology, Vienna Metabolomics Center (VIME), University of Vienna, 1090 8 Vienna, Austria Clinical Institute of Pathology, Medical University of Vienna, 1090 Vienna, Austria 9 Department of Human Genetics, Emory University, Atlanta, Georgia 30322, United States Aggregation of hypertrophic macrophages constitutes the basis of all granulomatous diseases such as tuberculosis or sarcoidosis and is decisive for disease pathogenesis. However, intrinsic pathways driving granuloma initiation and maintenance still remain elusive. Here we show that activation of mTORC1 in macrophages by deletion of Tsc2 was sufficient to induce hypertrophy and proliferation resulting in excessive granuloma formation in vivo. Remarkably, Tsc2-deficient macrophages formed mTORC1dependent hypertrophic granulomatous structures in vitro and showed constitutive proliferation mediated by the neo-expression of cyclin-dependent kinase 4 (CDK4). Moreover, mTORC1 promoted metabolic reprogramming via CDK4 towards increased glycolysis, while simultaneously inhibiting NF-κB signaling and apoptosis. Inhibition of mTORC1 rapidly induced apoptosis and completely resolved granulomas in myeloid Tsc2-deficient mice. Notably, in human sarcoidosis, mTORC1 activation, macrophage proliferation, and glycolysis were identified as hallmarks that correlated with clinical disease progression. Collectively, TSC2 maintains macrophage quiescence and prevents mTORC1-dependent granulomatous disease with clinical implications for sarcoidosis. Funding: T.W. is supported by grants from the Austrian Science Fund (FWF) grant FWF-P27701-B20, the Else-Kröner-Fresenius-Stiftung (P2013_A149), and the Herzfelder’sche Familienstiftung. M.L. is supported by the [DOC] Doctoral Fellowship Programme of the Austrian Academy of Sciences. V.S. is funded by FWF SFB F28 and SBF F47; M.Mü. is funded by FWF SFB F28. M.Mi. is supported by the FWF grant P25336-B13.

2111

Manon E. Wildenberg Benzimidazole anti-helminthics promote anti-TNF induced regulatory macrophage formation and potentiate therapeutic effect in a mouse model of inflammatory bowel disease. 1,2

2

3

2

2

Manon E. Wildenberg , Alon D. Levin , Alessandro Ceroni , Zhen Huo , Pim J. Koelink , Theodorus B.M. 2 2 1 3 1 Hakvoort , Felicia M. Bloemendaal , Johannan F. Brandse , Alison Simmons , Geert R.A.M. D’Haens , 3 1,2 Daniel Ebner and Gijs R. van den Brink 1

Tytgat Institute for Intestinal and Liver Research, Academic Medical Center, Amsterdam, The Netherlands Department of Gastroenterology and Hepatology, Academic Medical Center, Amsterdam, The Netherlands 3 Nuffield Department of Medicine, University of Oxford, Oxford, UK Introduction 2

The clinical benefit of anti-TNF in IBD correlates with their capacity to induce immunosuppressive macrophages in vitro and in vivo. The aim of this study was to screen a library of 1600 existing drugs for their capacity to potentiate this induction and thus enhance clinical efficacy of anti-TNF.

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Methods: Induction of CD14+CD206+ macrophages was analyzed by flow cytometry. Positive hits were validated in vitro and in the T cell transfer model of colitis. Results: Ninety-eight compounds potentiated anti-TNF mediated macrophage induction, including the commonly used anti-helminthic drug albendazole. Anti-TNF + albendazole combination treated macrophages were phenotypically comparable to macrophages induced by anti-TNF alone (CD14+, CD206+, CD163+), but immunosuppressive capacity was enhanced. In vivo, albendazole + anti-TNF combination therapy increased the presence of regulatory macrophages in the colon. Consequently, combination therapy was clinically superior in the treatment of experimental colitis compared to either compound alone. Conclusions: The anti-helminthic drug albendazole enhances the induction of regulatory macrophages by anti-TNF In vitro and the clinical efficacy in a colitis model. Given the good safety profile of this drug, antiTNF plus albendazole combination therapy may enhance clinical response in IBD. Funding: European Crohn’s and Colitis Organization, Dutch Society for Gastoenterology (NVGE)

2128

Leonard R Pelgrom Distinct metabolic requirements for T helper 1 and 2 polarization by dendritic cells. Leonard R. Pelgrom, Alwin J. van der Ham, Leonie Hussaarts, Bruno Guigas, Maria Yazdanbakhsh & Bart 1 Everts * *Corresponding author 1

Department of Parasitology, Leiden University Medical Center, Leiden, The Netherlands

Dendritic cells (DCs) play central role in the activation and polarization of T cell responses. We recently found that toll-like receptor signalling promotes a shift to glycolysis to support the anabolic demands of murine DC activation and effective priming of T cell responses. However, the metabolic requirements for polarization of distinct T helper cell (Th) responses by DCs remain poorly defined. Through genome wide expression analysis of human monocyte-derived DCs stimulated with helminth-derived antigens (Schistosoma mansoni derived egg antigen [SEA] and omega-1) or pharmacological compounds (rapamycin) that are known to prime Th2 responses, we identify suppression of genes involved in glycolysis as a key characteristic of Th2-polarizing DCs. Conversely, stimuli that promote Th1 responses enhance glycolysis in DCs. Furthermore, consistent with these observations, when glycolysis is blocked, DCs fail to induce Th1 responses and instead favour Th2 polarization. On the other hand, DCs that have been conditioned with SEA rely on mitochondrial fatty acid oxidation to promote effective differentiation of Th2 cells. These findings suggest that DC-driven polarization of different T cell responses is dependent on the activation of distinct metabolic programs in DCs and highlights that metabolic manipulation of DCs could hold promise as a novel therapeutic approach to control immune-polarization in disease settings.

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Hweixian Leong Penny Warburg metabolism in tumor-conditioned macrophages promotes metastasis in human pancreatic ductal adenocarcinoma. 1

1

2

1

1

1

Hweixian Leong Penny , Je Lin Sieow , Giulia Adriani , Wei Hseun Yeap , Peter See Chi Ee , Boris San Luis , 1 3 4 4 4 5 Bernett Lee , Terence Lee , Shi Ya Mak , Ying Swan Ho , Kong Peng Lam , Choon Kiat Ong , Ruby YJ 6 1 1 2,7 1 Huang , Florent Ginhoux , Olaf Rotzschke , Roger D. Kamm , Siew Cheng Wong 1

Singapore Immunology Network (SIgN), Biomedical Sciences Institute, A*STAR, 8A Biomedical Grove, 2 Immunos, Singapore 138648 BioSystems and Micromechanics IRG, Singapore-MIT Alliance for Research 3 and Technology (SMART), 1 CREATE Way, Singapore 138602 Raffles Institution, 1 Raffles Institution Lane, 4 Singapore 575954 Bioprocessing Technology Institute, A*STAR, 20 Biopolis Way, Centros, Singapore 5 138668 NCCS-VARI Translational Research Laboratory, National Cancer Centre, 11 Hospital Drive, 6 Singapore 169610 Centre for Translational Medicine NUS Yong Loo Lin School of Medicine, CSI Singapore,

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7

14 Medical Drive, Level 11, MD6, Singapore 117599 Department of Biological Engineering, Massachusetts Institute of Technology, 77 Massachusetts Ave, Cambridge, MA 02139, USA Patients with pancreatic ductal adenocarcinoma (PDAC) face a clinically intractable disease with exceptionally high rates of metastasis. Although Epithelial-to-mesenchymal transition (EMT) is pronounced at inflammatory foci within the tumor, the immunological mechanisms promoting tumor dissemination remain unclear. It is well established that tumors exhibit the Warburg effect, a preferential use of glycolysis for energy production to support rapid growth. We hypothesized that the metabolic pathways utilized by tumor-infiltrating macrophages are altered in PDAC, conferring a pro-metastatic phenotype. Human peripheral blood monocytes were cultured with conditioned media generated from normal pancreatic or PDAC cell lines to obtain steady-state and tumor-associated macrophages (TAMs), respectively. Compared with steady-state macrophages, TAMs augmented vascular network formation, promoted extravasation of tumor cells out of blood vessels, and induced higher levels of EMT. This prometastatic phenotype correlated with a pronounced glycolytic signature. Inhibiting glycolysis in TAMs with a single competitive inhibitor to Hexokinase II (HK2), 2-deoxyglucose (2DG), was sufficient to disrupt this pro-metastatic phenotype, reversing the observed increases in TAM-supported angiogenesis, extravasation, and EMT. Our results indicate a key role for metabolic reprogramming of tumor-infiltrating macrophages in PDAC metastasis, and highlight the therapeutic potential of using pharmacologics to modulate these metabolic pathways.

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Veronika Grau Alpha-1 antitrypsin inhibits ATP-dependent release of monocytic IL-1 via CD36, phospholipase A2 and nicotinic acetylcholine receptors. 1

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1

1

1

Robin Siebers , Katrin Richter , Anna Zakrzewicz , Bijan Fink , Sigrid Wilker , Mira Küllmar , Andreas 1 2 1 3 4 Hecker , Nupur Aggarwal , Winfried Padberg , J. Michael McIntosh , Sentot Santoso , Sabina 2,5 1,5 Janciauskiene , Veronika Grau 1

Laboratory of Experimental Surgery, Department of General and Thoracic Surgery, Justus-Liebig2 University, Giessen, Germany Department of Pulmonology, Hannover Medical School, Hannover, 3 4 Germany Department of Biology, University of Utah, Salt Lake City, Utah, USA Institute for Clinical 5 Immunology and Transfusion Medicine , Justus-Liebig-University, Giessen , Germany. Member of the German Centre for Lung Research High systemic levels of IL-1beta, a potent cytokine involved in host defense, cause life-threatening inflammatory diseases. Monocytes/macrophages synthesize a cytoplasmic pro-form of IL-1beta in response to triggers such as lipopolysaccharide. Extracellular ATP originating from the cytoplasm of damaged cells is a typical signal inducing inflammasome-dependent maturation and release of IL-1beta. Ample evidence suggests that the anti-protease alpha-1 antitrypsin (AAT) and IL-1beta regulate each other via largely unknown mechanisms. Here we demonstrate that AAT dose-dependently inhibits ATPinduced IL-1beta release by monocytic U937 cells and primary human monocytes, whereas ATPindependent release is unimpaired. Evidence is provided that AAT signals via long-chain fatty acid receptor CD36 that activates calcium-independent phospholipase A2 and leads to the secretion of a small soluble factor X. Factor X acts as an agonist of nicotinic acetylcholine receptors containing subunit alpha9 that, in turn, efficiently inhibit ion channel functions at ATP receptor P2X7. In conclusion, we suggest a novel triple-membrane-passing signaling pathway, by which AAT controls ATP-induced IL-1beta release from human monocytes. AAT might be a potent medicament for the prevention of trauma-associated systemic inflammatory response syndrome. Funding: LOEWE Program of the State of Hesse (Non-neuronal cholinergic systems), by the German Centre for Lung Research, by the Excellence Cluster Cardio-Pulmonary Systems, and by the German Research Foundation to V.G. (GR 1094/7-1).

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Oral presentations September Thursday 22nd

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Geert Raes Imaging of myeloid cell behaviour. 1,2,3

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Fang Zheng , PhD, Harris Perlman , PhD, Patrick Matthys , PhD, Tony Lahoutte , MD/PhD, Serge 2 3 2,3 2,3 Muyldermans , PhD, Shemin Lu , PhD, Patrick De Baetselier , PhD, Steve Schoonooghe *, PhD, Nick 2,6 2,3 Devoogdt * PhD and Geert Raes *, PhD Department of Biochemistry and Molecular Biology, Health Science Center, Xi'an Jiaotong University, 2 Xi’an, China; Research group of Cellular and Molecular Immunology, Vrije Universiteit Brussel, Brussels, 3 Belgium; Laboratory of Myeloid Cell Immunology, VIB Inflammation Research Center, Ghent, Belgium; 4 Division of Rheumatology, Feinberg School of Medicine, Northwestern University, Chicago, IL60611,USA; 5 6 Laboratory of Immunobiology, Rega Institute, Katholieke Universiteit Leuven, Leuven, Belgium; In Vivo 7 Cellular and Molecular Imaging Center, Vrije Universiteit Brussel, Brussels, Belgium; Department of Nuclear Medicine, UZ Brussel, Vrije Universtiteit Brussel, Brussels, Belgium. * S.S, N.D. and G.R. share senior authorship Single-photon emission computed tomography combined with micro-CT (SPECT/µCT) imaging using Nanobodies against complement receptor of the Ig superfamily (CRIg), found on tissue macrophages such as synovial macrophages, has promising potential to visualize joint inflammation in experimental arthritis. Here, we further addressed the specificity and assessed the potential for arthritis monitoring. Methods: 99m Signals obtained with Tc-labelled NbV4m119 Nanobody were compared in joints of wild type (WT) -/versus CRIg mice with collagen-induced arthritis (CIA) or K/BxN serum transfer-induced arthritis (STIA). In addition, SPECT/µCT imaging was used to investigate arthritis development in STIA and in CIA under 99m dexamethasone treatment. Results: Tc-NbV4m119 accumulated in inflamed joints of WT, but not CRIg /mice with CIA and STIA. Development and spontaneous recovery of symptoms in STIA was reflected in 99m initially increased and subsequently reduced joint accumulation of Tc-NbV4m119. Dexamethasone 99m treatment of CIA mice reduced Tc-NbV4m119 accumulation as compared to saline control in most 99m joints except knees. Conclusions: SPECT/µCT imaging with Tc-NbV4m119 allows specific assessment of inflammation in different arthritis models and provides complementary information to clinical scoring for quantitatively and non-invasively monitoring the pathological process and the efficacy of arthritis treatment.

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Aoife Kelly Human peripheral blood monocytes and intestinal macrophage populations activate TGF-β via expression of the integrin αvβ8. 1,2,3

1,2,3

1,2,3

Aoife Kelly *, Elinor Shuttleworth *, Tom Fenton , Catherine Smedley 1,2,3 Travis * Both authors contributed equally to this work 1

1,2,3

, Scott

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Levison , Mark

2

Manchester Collaborative Centre for Inflammation Research, Wellcome Trust Centre for Cell-Matrix 3 4 Research, Manchester Immunology Group, University of Manchester, UK. Central Manchester University Hospital NHS Foundation Trust, Manchester, UK. Transforming growth factor beta (TGF-β) plays a key role in maintaining intestinal homeostasis, acting to control inflammation and promote tissue repair. The cytokine is secreted as a latent complex, requiring activation to be functional. Although recent work has identified activators of TGF-β in mice, how the cytokine is activated to regulate human immune responses is poorly understood. Here, we show that human CD14+ classical monocytes activate high levels of TGF-β, which is not apparent in murine monocytes. Mechanistically, we show that activation of TGF-β by human monocytes requires expression of the integrin, αvβ8. When monocytes are differentiated to macrophages, integrin αvβ8 expression levels and TGF-β-activating function is elevated on tolerogenic versus inflammatory macrophages.

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Interestingly, we find that integrin αvβ8 is expressed by a proposed anti-inflammatory human intestinal macrophage subset expressing CD163 and CD206, with this macrophage population decreased in patients with active Crohn’s disease. Thus, our data suggest that expression of integrin αvβ8 by human monocytes may play an important role in the induction and function of anti-inflammatory macrophages in the intestine via TGF-β activation.

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Katrien van der Borght Tissue necrosis is a driver of organ directed autoimmunity through activation of dendritic cells. Katrien A Van der Borght(1,2), Charlotte L Scott(1,4), Veronica Nindl(5), Ann Bouché(3), Liesbet Martens(1,2,4), Dorine Sichien(1,4), Justine Van Moorleghem(1,2), Manon Vanheerswynghels(1,2), Sofie De Prijck(1,4), Gert Van Isterdael(1,2), Burkhard Ludewig(5), Thierry Gillebert(2), Martin Guilliams(1,4), Peter Carmeliet(3,7) and Bart N Lambrecht (1,2,6,7_ Katrien A Van der Borght(1,2), Charlotte L Scott(1,4), Veronica Nindl(5), Ann Bouché(3), Liesbet Martens(1,2,4), Dorine Sichien(1,4), Justine Van Moorleghem(1,2), Manon Vanheerswynghels(1,2), Sofie De Prijck(1,4), Gert Van Isterdael(1,2), Burkhard Ludewig(5), Thierry Gillebert(2), Martin Guilliams(1,4), Peter Carmeliet(3,7) and Bart N Lambrecht (1,2,6,7) (1)VIB Inflammation Research Center, VIB and Ghent University, Ghent, Belgium(2)Department of Internal Medicine, Ghent University, Ghent, Belgium(3)Vesalius Research Center, VIB and KULeuven, Leuven, Belgium(4)Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium(5)Institute of Immunobiology, Kantonal hospital St. Gallen, St. Gallen, Switzerland(6)Department of Pulmonary Medicine, ErasmusMC, Rotterdam, The Netherlands Peripheral tolerance is crucial for avoiding activation of self-reactive T cells to tissue restricted antigens. Autoimmunity can be triggered by sterile tissue injury, but it is unclear how immune cells get activated to respond to self-antigen. A good example of sterile injury is acute myocardial infarction (MI). We hypothesized that tissue necrosis is an activator of dendritic cells (DC) that control tolerance to self+ antigens. DC subsets of the murine healthy heart consisted of IRF8-dependent XCR-1 conventional + + (c)DC1, IRF4-dependent CD172α cDC2 and CD64 monocyte-derived DCs, that shared a common gene signature with DC subsets elsewhere in the body. In steady state, cardiac self-antigen α-myosin was presented in the heart-draining mediastinal lymph node (mLN) by cDC1s, driving the proliferation of + antigen-specific CD4 TCR-M T cells, and expansion of Treg cells. Following MI, all DC subsets infiltrated the infarcted heart, while only cDCs migrated to the mLN. Here, cDC2s induced TCR-M proliferation and differentiation into IL-17/IFNγ producing effector cells. RNA sequence analysis revealed that MI changed the heart DC gene expression profile, inducing genes involved in migration and T cell activating capacity. Thus, tissue necrosis is a major driver of autoimmunity to tissue restricted self-antigen via activation of DCs.

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Andrew S MacDonald A central role for Type I IFN in Th2 induction by dendritic cells. 1,7

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Lauren M. Webb , Rachel J. Lundie , Jessica G. Borger , Sheila L. Brown , Lisa M. Connor , Adam N. R. 1 4 5 1 2 Cartwright , Annette M. Dougall , Ruud H. P. Wilbers , Peter C. Cook , Lucy H. Jones , Alexander T. 1 6 1 4 3 Phythian-Adams , Cecilia Johansson , Daniel M. Davis , Benjamin G. Dewals , Franca Ronchese and 1 Andrew S. MacDonald . 1

Manchester Collaborative Centre for Inflammation Research, University of Manchester, Manchester, U.K. 3 University of Edinburgh, Edinburgh, U.K. Malaghan Institute of Medical Research, Wellington, New 4 5 Zealand. University of Liege, Belgium. Wageningen University and Research Centre, Wageningen, The 6 7 Netherlands. Imperial College London, London, U.K. Current address: Cornell University, Ithaca, U.S.A. 8 Current address: Monash University, Victoria, Australia. 2

Although dendritic cells (DCs) are vital for Th2 induction against helminths or allergens, relatively little is known about how they are activated and function in response to Th2-polarizing antigens. We have

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discovered a previously unappreciated role for Type I IFN (IFN-I) in the activation and function of DCs following exposure to strongly Th2-polarizing antigens. So far, IFN-I has chiefly been associated with antiviral immunity, while its role in Th2 settings is much less clear. DCs cultured with total egg antigens from the parasitic helminth Schistosoma mansoni, or the dominant immunostimulatory component of S. mansoni eggs omega-1, produced IFN-I. IFN-I was also detected in response to the common allergen house dust mite (HDM). DCs lacking the IFN-I receptor displayed dramatically impaired Th2 induction in vivo, but unimpaired ability to support CD4+ T cell polarization in vitro. Further, Th2-promoting DCs depended on IFN-I signaling for optimal activation, efficient migration to the draining LN, and effective localization within the T cell zone. Additionally, challenge of mice with S. mansoni eggs or HDM in the absence of the IFN-I receptor resulted in significantly reduced Th2 cytokine induction in vivo. Together, our data suggest a key and unexpected role for IFN-I responsiveness in enabling Th2 induction by DCs in vivo. Funding: Medical Research Council UK

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Dieke van Dinther Development of a macrophage-based vaccination strategy for melanoma. Dieke van Dinther (1), Henrike Veninga (1), Leoni Hoogterp (1), Mirjam Revet (1), Ellen G.F. Borg (1), Hakan Kalay (1), Jan Wouter Drijfhout (2), Yvette van Kooyk (1), Joke M.M. den Haan (1) (1) Department of Melcular Cell Biology and Immunology, Vumc, Amsterdam, The Netherlands (2) Departement of Immunohematology and Blood Transfusion, Leiden University Medical Centre Leiden, the Netherlands. Melanoma causes the majority of skin cancer related deaths, but because it is highly immunogenic, treatment with immunotherapies such as immune blockade inhibitors might work synergistically with vaccination strategies. As an in vivo vaccination strategy we propose to target melanoma antigens (Ag) to a specific subset of Ag-capturing macrophages in the spleen that are characterized by the expression of CD169. Previously, we have shown that these CD169+ macrophages transfer Ag to B cells and DCs for the induction of strong humoral responses and T cell responses, respectively. In the current study we compare the cytotoxic T cell response after targeting Ag to CD169 in the presence or absence of CD4 T cell help and a humoral response. We conjugated protein or peptide to anti-CD169 antibodies and then monitored (i) the primary, (ii) the recall and (iii) the anti-tumor immune responses after immunization with these conjugates. Both protein and peptide targeting to CD169 resulted in strong primary and recall immune responses and preliminary data on the protective immunity against melanoma show promising results. In conclusion, this macrophage-based vaccination strategy stimulates strong immune responses and can be applied to raise protective immunity against melanoma. Funding: KWF

2204

Martijn Verdoes In vivo imaging of tumor-associated macrophages and subcellular compartment characterization in dendritic cells using novel quenched fluorescent cysteine cathepsin probes 1,2

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Martijn Verdoes, * Kristina Oresic Bender, Carl Figdor, Matthew Bogyo.

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Department of Tumor Immunology, Radboud Institute for Molecular Life Sciences, Radboudumc and 3 Radboud University, Institute for Molecules and Materials, Nijmegen, The Netherlands. Departments of Pathology, Stanford School of Medicine, United States. *Correspondence: [email protected] 2

Cysteine cathepsins (CCTS) are highly expressed by myeloid immune cells. They are involved in antigen presentation, migration as well as tumor progression. Several studies suggest that CCTS regulate the activity of their family members and have seemingly redundant as well as very specific functions. This suggests a high degree of control of localization of their individual proteolytic activities. However, no tools

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existed for live cell activity imaging of specific CCTS with respect to their family members. We developed a quenched activity-based probe (qABP) specific for cathepsin S. In combination with a complementary panreactive qABP we generated a set of tools which enable us to perform dual color live cell activity localization studies. Using this set of qABPs we identified sub-cellular compartments containing exclusively cathepsin S activity in live dendritic cells. Furthermore, we use these qABPs to perform noninvasive and multiphoton intravital imaging of mouse models of cancer and we found that within the tumor microenvironment a subpopulation of macrophages with a pro-tumor, M2 type phenotype are the major source of CCTS activity. We are currently investigating the functional characterization of this subpopulation of tumor resident myeloid cells, as well as designing molecules to manipulate these cells in vivo. Funding: Institute for Chemical Immunology, ERC Starting Grant

2210

Rieneke van de Ven Correlating the ILT4/HLA-G inhibitory pathway to an immune suppressive microenvironment in sentinel lymph nodes draining head and neck squamous cell carcinoma. 1

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A.M Heeren , A.G.M. Stam , E.S. Jordanova , S. van Weert , K.H. Karagozoglu , T.D. de Gruijl , C.R. 3 5 2 Leemans , E. Bloemena and R. van de Ven * 1

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Departments of Gynecology, Medical Oncology, Otolaryngology-Head and Neck surgery, Oral and 5 Maxillofacial Surgery and Oral Pathology, Pathology, VU University medical center – Cancer Center Amsterdam, De Boelelaan 1117, 1081 HV Amsterdam, The Netherlands. The sentinel lymph node (SLN) is the first tumor-draining LN and consequently the first site after the primary tumor where tumor-mediated immune suppression can hamper the development of an efficient anti-tumor immune response. Presence of the inhibitory molecule immunoglobulin-like transcript 4 (ILT4) on tolerogenic dendritic cells (DC) has been described to result in induction of regulatory T cells (Treg). In SLN (n=12) draining head and neck squamous cell carcinomas (HCSCC) we studied by flow cytometry whether the presence of ILT4 and its ligand the non-classical MHC class I molecule HLA-G, correlated with immune suppression. Migratory DC, lymph node resident conventional DC (LNR-cDCs) and CD14+ macrophages (MFs) in the SLN all expressed HLA-G. ILT4 expression was highest on CD14+ MFs. However, increased expression of ILT4 (mean fluorescence index) on LNR-cDC was found to correlate with increased frequencies of activated Treg (aTreg) (Pearson r= 0.84, p