georg speyer haus institut fÜr tumorbiologie und...

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GEORG SPEYER HAUSINSTITUT FÜR TUMORBIOLOGIE UND EXPERIMENTELLE THERAPIE

2017Annual ReportGeorg-Speyer-Haus

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Die Grundfinanzierung des Georg-Speyer-Hauses wird vom Bundesministerium für Gesundheit und dem Hessischen Ministerium für Wissenschaft und Kunst getragen.

The basic funding of the Georg-Speyer-Haus is provided by the Federal Ministry of Health and the Ministry of Higher Education, Research and the Arts of the State of Hessen.

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Forschen für das Leben Research for Life

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Inhalt

VorwortOrganisationsstruktur Das Georg-Speyer-HausHighlights

Zelluläre Kommunikation in der StammzellnischeCellular Communication in the Stem Cell NicheProf. Dr. D. Krause Dr. H. MedyoufProf. Dr. M. Zörnig

Zell-Zell Interaktionen im TumorstromaCell-Cell Interaction in the Tumor StromaPD Dr. M. C. ArkanDr. H. Farin Prof. Dr. F. R. GretenDr. L. Sevenich

Experimentelle TherapieExperimental TherapyDr. U. DietrichProf. Dr. W. Wels

PublikationenFinanzen und AdministrationServiceVeranstaltungen, Lehre und NachwuchsförderungDer Verein »Freunde und Förderer des Georg-Speyer-Hauses«Zuwendungsgeber

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High speed cell sorting

in the cytometry unit

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Introduction

unsere in den letzten Jahren durchgeführte Umstruk-turierung und Fokussierung auf die onkologische Forschung unter besonderer Berücksichtigung des Tumormikromilieus ist inzwischen gut etabliert. Insbesondere unsere neuen Nachwuchsgruppen sind sehr erfolgreich und weisen erste sehenswerte Erfolge auf. Die Bemühungen, gemeinsam mit unseren klinischen Partnern unsere Ergebnisse und therapeutischen Konzepte in die klinische Anwendung zu überführen, machen große Fortschritte. Dieses Jahr hat das Paul-Ehrlich-Institut die Genehmigung für die Durchführung der CAR2BRAIN Studie erteilt, um den Verträglichkeit von sogenannten chimären Antigen-Rezeptor (CAR)-exprimierenden NK Zellen (NK-92/5.28.z) für die Behandlung von Glioblastom-Patienten zu testen. Diese Studie, die auf der Entwicklung einer CAR-NK Zelllinie durch die Gruppe von Prof. Wels in Kollaboration mit den Kollegen vom Blutspendedienst beruht, ist weltweit die erste ihrer Art. Wir gehen davon aus, dass die ersten Patienten, die derzeit in der Neuro-Onkologie in die Studie eingeschleust werden, demnächst behandelt werden.

Neben diesen exzellenten Entwicklun-gen in Bezug auf die klinische Trans-lation sind auch unsere Bemühungen, das Frankfurt Cancer Institut (FCI) zu gründen, sehr erfolgreich verlaufen. Zusammen mit unseren Kollegen

der Goethe-Universität haben wir eine Antragsskizze zur Finanzierung von Hochschulbauten nach Artikel 91b Grundgesetz eingereicht. Diese wurde durch den Wissenschaftsrat sehr positiv evaluiert und zur Voll-antragstellung aufgefordert, so dass wir jetzt mit der

Our focus on translational oncology with a special emphasis on the tumor microenvironment has been firmly established, and together with our clinical partners our efforts to efficiently translate our findings and therapeutic concepts into clinical medicine significantly advance. This year the Paul-Ehrlich Institute has approved a clinical trial (CAR2BRAIN) using chimeric antigen receptor (CAR)-engineered NK cells (NK-92/5.28.z) to treat glioblastoma patients. Enrollment of patients for this trial – which is worldwide the first of its kind – is commencing, and we expect treatment of the first patient by our neuro-oncological colleagues very soon. Apart from this excellent development in translation our efforts to create the Frankfurt Cancer Institute (FCI) are nicely progressing as well. Together with our colleagues from the Goethe University we submitted a proposal to obtain funding according to Article 91b of the German “Grundgesetz”. This first proposal was positively evaluated by the German Science Council, thus enabling us now to initiate the precise construc-tion planning for this building. We are optimistically looking forward to the final decision due in April 2018. Moreover, upon positive evaluation of our

Dear Readers,dear friends of the Georg-Speyer-Haus,

Liebe Leserinnen und Leser, liebe Freunde des Georg-Speyer-Hauses,

Florian R. Greten | DirektorGeorg-Speyer-HausInstitut für Tumorbiologie und experimentelle Therapie

Paul-Ehrlich-Str. 42 – 44D-60596 Frankfurt / M.Tel. (069) 63395-232Fax (069) [email protected]

FRANKFURT CANCER INSTITUTE

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konkreten Planung beginnen können. Eine endgültige Entscheidung in diesem Verfahren erwarten wir für April 2018. Darüber hinaus haben wir einen Vollan-trag zur Gründung eines neuen LOEWE-Zentrums (LOEWE-FCI) eingereicht und sehen mit Spannung der Vor-Ort-Begutachtung im März 2018 entgegen. Die

erfolgreiche Einrichtung dieses LOEWE-Zentrums wird es dann ermöglichen, die innovativen wissenschaftlichen Konzepte, die wir im FCI verfolgen wollen, auch tatsächlich umzusetzen.

Neben den positiven wissenschaftlichen und strukturellen Entwicklungen gab es auch einige andere Höhepunkte während des Jahres. Die über die Jahre fest etablierte und auch in diesem Jahr wieder außerordentlich gut besuchte Schülervorle-sungsreihe, sowie das Schülerpraktikum, organisiert von Dr. Ursula Dietrich, ermöglichten interessierten Oberstufenschülern, mehr über die experimentelle Grundlagenforschung zu erfahren. Im Juni führten wir unser im Abstand von zwei Jahren durchgeführtes wissenschaftliches Retreat auf der Burg Rothenfels im Spessart durch. Neben einer Vielzahl von spannenden wissenschaftlichen Vorträgen und sehr lebhaften Poster-Diskussionen kam auch der gesellschaftliche Teil nicht zu kurz. Im September fand wie in jedem Jahr das Auswahlsymposium für den Paul Ehrlich- und Ludwig Darmstaedter-Nachwuchspreis bei uns am GSH statt, und auch in diesem Jahr waren die Kandidaten exzellent und die Vorträge ungemein spannend. Im Oktober diskutierten internationale Experten im Rahmen des von Dr. Henner Farin organisierten Symposiums „Dynamics of adult stem cells and cancer” für zwei Tage über ihre Ergebnisse und Visionen zur Rolle von Tumor-Stammzellen.

Wir sind sehr froh über die positive Entwicklung, die das GSH in den letzten Jahren durchlaufen hat, und sind sehr optimistisch über die vielzähligen Möglichkeiten, die die Gründung des FCI in der Zukunft bieten wird. Eine spannende und sicherlich erfolgreiche Zeit liegt vor uns.

pre-proposal we have been asked to submit a full application for a new LOEWE Center (LOEWE – FCI), which will be reviewed in depth in Frankfurt in March 2018, and which would provide the required funding of our proposed innovative scientific concepts within the FCI. Apart from this year’s scientific and structural progress we experienced several other highlights. The over the years well-established and in this year again extremely well attended student lecture series as well as the student internships, organized by Dr. Ursula Dietrich, allowed interested and talented high school students to gain a detailed insight into experimental basic research. In June all GSH groups gathered for our bi-annual Scientific Retreat on Burg Rothenfels in the Spessart, where everyone enjoyed the bal-anced combination of science and outdoor activities. In September the six candidates for the Paul Ehrlich- and Ludwig Darmstaedter-Junior Award presented their excellent work, and in October Dr. Henner Farin organized the very well attended symposium „Dynamics of adult stem cells and cancer”. During two days international experts discussed and exchanged ideas on various aspects of tumor cell stemness.

We are very happy about the positive development at the GSH over the last years and are enthusiastic about the possibilities that the FCI will be able to provide. Exciting and most likely even more successful years are lying ahead of us.

Introduction

Florian R. Greten, Direktor

GEORG SPEYER HAUSINSTITUTE FOR TUMOR BIOLOGY

AND EXPERIMENTAL THERAPY

CONFIRMED SPEAKERS

Renée van Amerongen Swammerdam Institute for Life Sciences,

University of Amsterdam, NL

Thomas Brabletz Nikolaus-Fiebiger-Center for Molecular

Medicine, Erlangen

Dominik GrünMax Planck Institute of Immunobiology

and Epigenetics, Freiburg

Borhane GuezguezMedizinische Klinik und Poliklinik,

Johannes Gutenberg-Universität Mainz

Meritxell Huch Gurdon Institute,

Cambridge, UK

Christoph A. KleinExperimental Medicine and Therapy

Research, Universität Regensburg

Daniel Peeper Netherlands Cancer Institute,

Amsterdam, NL

Freddy Radtke ISREC/EPFL,

Lausanne, Switzerland

Martin SprickHI-STEM Deutschen Krebsforschungs-

zentrum DKFZ, Heidelberg

Sabine TejparMolecular Digestive Oncology,

KU Leuven, Belgium

Zlatko TrajanoskiBiocenter, Division for Bioinformatics,

Innsbruck Medical University, Austria

Lars ZenderClinical Tumor Biology, University Hospital

and Faculty of Medicine, Tübingen

KEY NOTE SPEAKER

Fiona Watt Centre for Stem Cells & Regenerative

Medicine, King‘s College London

ORGANIZER

Henner FarinGeorg-Speyer-Haus, Frankfurt

REGISTRATION

Stefanie Schütt

[email protected]

Phone: +49 (0)69 63395-183

Dynamics of adult stem cells and cancer

OCT 25–26, 2017

FRANKFURT/MAIN

GERMANY

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Organizational Structure

* Kooperationsgruppe mit Institut für Biochemie II, Goethe-Universität Frankfurt In cooperation with Institute of Biochemistry II Goethe University Frankfurt

FORSCHUNGSBEREICH 3

Experimentelle Therapie

Dr. U. DietrichProf. Dr. W. Wels

STIFTUNGSRAT

Vorsitzender G. Wiesheu

Dr. U. BollertMinDirig. Dr. V. GrigutschProf. Dr. W. Müller-Esterl Prof. Dr. S. Offermanns Prof. Dr. J. Pfeilschifter

MinR‘in A. Steinhofer-Adam

WISSENSCHAFTLICHER BEIRAT

VorsitzenderProf. Dr. A. Radbruch

Prof. Dr. T. BrunnerProf. Dr. A. Eggert

Prof. Dr. L. HennighausenProf. Dr. K. L. RudolphProf. Dr. D. TuvesonProf. Dr. E. Wiertz

DIREKTORIUM

Wissenschaftlicher DirektorProf. Dr. F. R. Greten

StellvertreterProf. Dr. W. Wels

Kaufmännischer LeiterR. Dornberger

SERVICE-EINRICHTUNGEN

DurchflusszytometrieHistologie / Präklinische Einheit

Transgenic Core Facility

Dr. B. BrillDr. M. Karimova

Dr. S. Stein

VERWALTUNG

Personal, Finanzen, IT, Innendienst, Einkauf,

Arbeitssicherheit

R. Dornberger

FORSCHUNGSBEREICH 1

Zelluläre Kommunikation in der Stammzellnische

Prof. Dr. D. Krause Dr. H. Medyouf

Prof. Dr. M. Zörnig

FORSCHUNGSBEREICH 2

Zell-Zell Interaktionen im Tumorstroma

PD Dr. M.C. Arkan* Dr. H. Farin

Prof. Dr. F. R. GretenDr. L. Sevenich

Hygiene- und Labor-

management

Hana Kunkel

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Die Stiftung privaten Rechts „Chemo-therapeutisches Forschungsinstitut Georg-Speyer-Haus“ wurde 1904 in Frankfurt am Main gegründet, um eine Forschungsstätte für Paul Ehrlich, den ersten Direktor des Hauses, zu schaffen. Die Stiftungsverfassung bestimmt als Zweck der Stiftung die wissenschaftliche Forschung auf den Gebieten der Chemo-therapie und verwandter Wissenschaften, die dem Fortschritt der Biomedizin dienen. Es werden ausschließlich und unmittelbar gemeinnützige Zwecke verfolgt.

Die laufenden Geschäfte des heutigen Instituts für Tumorbiologie und experimentelle Therapie nimmt der Direktor wahr. Er ist in dieser Tätigkeit dem Stiftungsvorstand verantwortlich. Das Georg-Speyer-Haus ist durch einen Kooperationsvertrag mit der Goethe-Universität Frankfurt verbunden.

Das Gebäude des Georg-Speyer-Hauses in der Paul-Ehrlich-Straße 42 – 44, 1906 eröffnet, wurde von der Stadt Frankfurt am Main zur Nutzung

für Institutszwecke zur Verfügung gestellt. Der gesamte Gebäudekomplex wurde in den Jahren 1995 – 1997 aus Mitteln des Bundesministeriums für Gesundheit und des Hessischen Ministeriums für Wissen-schaft und Kunst saniert und modernisiert. Er umfasst eine Gesamtfläche von 4710 qm. Die Laboratorien sind für Arbeiten unter verschiedenen biologischen und gentechnischen Sicherheitsstufen (L2, L3, S1, S2, S3) zugelassen.

The private foundation “Chemothera-peutisches Forschungsinstitut Georg-Speyer-Haus” (Chemotherapeutic Research Institute Georg-Speyer-House) was established in 1904 in order to provide a research institute for Paul Ehrlich, its first director. The constitution of the institute, originating from its foundation, defines its purpose as an establishment for scientific research in the field of chemotherapy and related sciences. It is an independent institution under public law which is exclusively engaged in non-profit work.

Today’s Institute for Tumor Biology and Experimental Therapy is headed by the Scientific Director who reports to the Board of the Foundation. The Georg-Speyer-Haus has a cooperative agreement with the Goethe University Frankfurt.

The Georg-Speyer-Haus is located in a building on Paul-Ehrlich-Str. 42- 44, which has been provided by the City of Frankfurt. The building which was opened in 1906 was renovated in the years from 1995 – 1997 with support

Research for LifeForschen für das Leben

The Georg-Speyer-Haus

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The Georg-Speyer-Haus

Das Georg-Speyer-Haus wird finanziell vom Bundesministerium für Gesundheit (BMG) sowie vom Hessischen Ministerium für Wissenschaft und Kunst (HMWK) unterstützt. Zusätzlich stehen Mittel aus der Drittmittelförderung öffentlicher und privater Forschungsförderungsorganisatio-nen, aus Kooperationsvereinbarungen mit Unternehmen, aus Erträgen des Stiftungs-kapitals und aus Spenden zur Verfügung.

Als Partner im Universitären Centrum für Tumorerkran-kungen (UCT), dem LOEWE Zentrum für Zell-und Gen-therapie (LOEWE-CGT) sowie dem Deutschen Konsortium

für translationale Krebsforschung (DKTK) führt das Georg-Speyer-Haus international kompetitive Grundlagenforschung auf dem Gebiet der Tumorbiologie unter besonderer Berücksichtigung des Tumormikromilieus durch. Durch die enge Kollaboration mit den klinischen Partnern der Goethe-Universität im Rahmen der oben genannten Verbünde werden die Ergebnisse aus der Grundlagenforschung in frühe klinische Studien überführt. Darüberhinaus engagiert sich das Georg-Speyer-Haus in der Wissensvermittlung sowie in der Umsetzung neuer Einsichten in therapeutische Applikationen, Dienst-leistungen und Produkte und kann so als ein Zentrum der translationalen onkolo-gischen Forschung angesehen werden.

from the Federal Ministry of Health and the Ministry of Higher Education, Research and the Arts of the State of Hessen. It comprises an area of 4710 m2. The laboratories are certified for work under different biological and gene technology safety regulations (L2, L3, S1, S2, S3).

The Georg-Speyer-Haus is supported by the Federal Ministry of Health and the Ministry of Higher Education, Research and the Arts of the State of Hessen. Additional funding is provided by competitive grants, by cooperation agreements with companies, by returns from the investment of the foundation and by private donations.

As a strong partner within the University Cancer Center, the LOEWE Center für Cell and Gene Therapy as well as the German Cancer Consortium the Georg-Speyer-Haus is performing internationally competitive basic research in the field of tumor biology with a particular focus on the tumor microenvironment. In close collaboration with clinical partners at the Goethe-University, results are translated into early clinical trials and the Georg-Speyer-Haus can therefore be considered a center of translational oncology.

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Highlights 2017

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10. SeptemberLauf für mehr Zeit

25.-26. OktoberSymposium „Dynamics of adult stem cells and cancer“Organisiert von Henner Farin.Organized by Henner Farin.

1. Dezember 2016Welt-AIDS-Tag A. Haberl/H. Rabenau, Uniklinik Frankfurt.Das Audimax der Uniklinik Frankfurt gefüllt mit 440 Schülern.The Audimax of the Uniklinik Frankfurt filled with 440 students.

GEORG SPEYER HAUSINSTITUTE FOR TUMOR BIOLOGY AND EXPERIMENTAL THERAPY

CONFIRMED SPEAKERS

Renée van Amerongen Swammerdam Institute for Life Sciences, University of Amsterdam, NL

Thomas Brabletz Nikolaus-Fiebiger-Center for Molecular Medicine, Erlangen

Dominik GrünMax Planck Institute of Immunobiology and Epigenetics, Freiburg

Borhane GuezguezMedizinische Klinik und Poliklinik, Johannes Gutenberg-Universität Mainz

Meritxell Huch Gurdon Institute, Cambridge, UK

Christoph A. KleinExperimental Medicine and Therapy Research, Universität Regensburg

Daniel Peeper Netherlands Cancer Institute, Amsterdam, NL

Freddy Radtke ISREC/EPFL, Lausanne, Switzerland

Martin SprickHI-STEM Deutschen Krebsforschungs-zentrum DKFZ, Heidelberg

Sabine TejparMolecular Digestive Oncology, KU Leuven, Belgium

Zlatko TrajanoskiBiocenter, Division for Bioinformatics, Innsbruck Medical University, Austria

Lars ZenderClinical Tumor Biology, University Hospital and Faculty of Medicine, Tübingen

KEY NOTE SPEAKER

Fiona Watt Centre for Stem Cells & Regenerative Medicine, King‘s College London

ORGANIZER

Henner FarinGeorg-Speyer-Haus, Frankfurt

REGISTRATION

Stefanie Schü[email protected]: +49 (0)69 63395-183

Dynamics of adult stem cells and cancer

OCT 25–26, 2017FRANKFURT/MAINGERMANY

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20.-21. Juni GSH-RetreatDas diesjährige Retreat des Instituts fand am 20. und 21. Juni auf der Burg Rothenfels im bayrischen Landkreis Main-Spessart statt.This year´s GSH retreat took place from the 20th until 21st of June at the Burg Rothenfels which is located at the River Main in the Bavarian county Main-Spessart.

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Zwei Tage lang haben wir zusammen mit den eingeladenen Gastsprechern angeregt über wissenschaftliche Projekte diskutiert und gemeinsame Freizeit verbracht.

For two days we discussed scientific projects with our invited guest speakers and had fun during our leisure time.

Neben spannender Wissenschaft ...Apart from fascinating science ...

... galt es, interessante Geschichte spielerisch zu erkunden ...

... we followed the traces of interesting history ...

... und alte Sportarten auszuprobieren.

... and tried medieval sporting activities.

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Zelluläre Kommunikation in der Stammzellnische Cellular Communication in the Stem Cell Niche

Laboratories I

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Bone marrow microenvironmentDaniela Krause

Die Rolle des Knochenmarksmikromilieus bei den Leukämien

The bone marrow microenvironment (BMM) is increasingly being considered as a novel target to augment existing thera-pies for haematological malignancies. This is important, as the overall survival rate for all leukaemias in adults is only 44% and leukaemic stem cells are rarely eradicated. Eradication of cancer stem cells or leukaemia stem cells in leukaemia, however, is important for cure of a cancer.

Based on our previous work our labora-tory focuses on various pathways of interaction of leukaemia cells with their surrounding bone marrow microenviron-ment in an effort to eventually target these interactions and eradicate leukae-mic stem cells (LSC). The vascular niche, the extracellular matrix, the coagulation system and novel pathways of adhesion to the BMM, hereby, studied by various in vitro and in vivo modelling systems, as well as in vivo 2-photon based imag-ing (in collaboration with Prof. S. Dim-meler) form the basis of our studies.

The role of the bone marrow microenvironment in leukaemia

MitarbeiterRahul Kumar Thanh Van HoangValentina MinciacchiDivij VermaSonika GodavarthyChristina Karantanou Raquel PereiraCostanza ZanettiJennifer Butenschön Nina Hayduk

GruppenleiterinDaniela KrauseTel.: +49 69 63395-500Fax: +49 69 [email protected]

leukaemia

bone marrow microenvironment

pharmacological modulation

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Trotz verbesserter Therapien, z.B. in Form von Tyrosinkinaseinhibi-toren, liegt die 5-Jahres-Überlebensrate bei Erwachsenen für alle Leukämien bei nur 40%. Deshalb hat es sich unsere Arbeitsgruppe zur Aufgabe gemacht, neue Therapien, vor allem solche mit neuem Therapieansatz, zu entwickeln.Wie bereits von uns und anderen Gruppen publiziert, kann eine gezielte Modulation des Knochenmarksmikromilieus (KMMM), dem Ort, wo eine Leukämie entsteht und voranschreitet, eine Ver-ringerung von leukämischen Stammzellen nach sich ziehen. Dies ist notwendig, denn leukämische Stammzellen können zu Thera-pieresistenz und Krankheitsrückfall führen. Das KMMM, welches leukämische Stammzellen vor der Chemotherapie „beschützen“ kann, besteht aus verschiedenen Zelltypen wie Osteoblasten, Osteoklasten, mesenchymalen Stammzellen, Endothelzellen, und der extrazellulären Matrix.

Wir testen experimentell, durch welchen Mechanismus eine Blo-ckade eines auf Endothelzellen exprimierten Proteins, E-Selektin, das Überleben von leukämischen Stammzellen beeinträchtigt und wie die Lokalisation von leukämischen Stammzellen innerhalb des KMMMs und ihre spezifische Interaktion mit der extrazellulären Matrix des KMMMs den Krankheitsverlauf einer Leukämie beein-flussen kann. Dazu wird in Kürze eine klinische Studie zur Modu-lation der extrazellulären Matrix im KMMM bei chronisch myeloi-scher Leukämie in enger Kollaboration mit der Medizinischen Klinik für Hämatologie/Onkologie des Klinikums der Goethe-Universität gestartet. Ferner sind die Rolle des Blutgerinnungssystems im KMMM und die Adhäsion von Leukämie-induzierenden Zellen im KMMM Fokus unserer Arbeitsgruppe.

As we have previously shown that CD44 and selectins and their ligands play a role for the engraftment of LSC in chronic myeloid leukaemia (CML) we are now testing, if the E-selectin-specific inhibitor GMI-1271 is beneficial for the reduction of LSC in CML. Indeed, our data indicate that inhibition of E-selectin in murine CML may lead to ‘non-adherence’ to the vascular niche, reduced engraftment of leukaemia-initiating cells and pos-sibly an altered cell cycle of LSC, likely via non-adhesion to the vascular niche, a novel finding in leukaemia. Simultane-ously, we have implicated that BCR-ABL1, the causative oncogene giving rise to CML, regulates the expression of CD44, which is essential for the engraftment of CML-initiating cells, via Akt and Scl/Tal-1.

In other work we have shown by confo-cal 2-photon in-vivo microscopy of the calvarium in live mice that leukaemic stem cells in CML have a distinct location in the bone marrow niche, which is different from the location of normal haemato-poietic stem cells. Prior in-vitro treatment of leukaemic stem cells with imatinib, considered standard of care in CML, reversed this phenotype. Furthermore, LSC in CML, harbouring the T315I point mutation in BCR-ABL1 (BCR-ABL1T315I), which conveys resistance to imatinib, were found closest to the endosteum. As patients and mice with CML due to BCR-ABL1T315I have an accelerated clinical course and a worse outcome, we are currently investigating whether an altered interaction with the bone marrow micro-environment via altered signaling from BCR-ABL1 or its respective mutants may be a cause of this. Indeed, in compari-son to BCR-ABL1WT+ cells we have been able to implicate an altered expression of adhesion molecules, increased engraft-ment, increased adhesion, alterations of the actin cytoskeleton and differences in signaling pathways. We hold a patent for these discoveries, on the basis of which a

clinical trial is being initiated in collabora-tion with a pharmaceutical company.

Thirdly, the coagulation system is a com-plex system comprised of various prote-ases and other factors which result in the formation of a blood clot. The bone mar-row microenvironment has been implicat-ed in the maintenance of haematopoietic stem and progenitor cells (HSPC), but pro-teases such as metalloproteinase 9 have only been implicated in the mobilization of HSPC, but not for HSPC homeostasis.Therefore, we hypothesized that the proteases of the coagulation system may play a role in the normal physiology of the BMM and that modulation of coagulation factors may have an effect on HSPC, for instance via degradation of the extracel-lular matrix and increased mobilization of HPSC. Indeed, by this approach we discovered a prominent role for certain proteins in the BMM which influence the homeostasis and maintenance of HSPC.

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In a fourth project we are investigat-ing the role of lipid raft associated molecules for adhesion of leukaemia cells to the BMM. We have found that lipid raft-associated molecules play a prominent role for the engraftment of leukaemia cells, possibly via associa-tion with certain adhesion molecules.

We continue to work on the role of the transcriptional regulator Far upstream element-binding protein 1 (FUBP1) and its role in the development of leukaemia in a productive collabora-tion with the Zörnig Group in a project funded by the Sander Foundation.

Figure 1.Immunofluorescence staining of an extracellular matrix protein.

In summary, the laboratory focuses on the role of the different constituents of the BMM on the initiation, maintenance and progression of leukaemias in an at-tempt to develop novel therapies which can augment our existing armamen-tarium against this intractable disease.

Figure 2.Transplanted, fluorescently labelled leukaeamia-initiating cells in the bone marrow cavity of a Tie2 GFP reporter mouse. Endothelial cells fluoresce in green, bone in blue. The image was taken by 2-photon in-vivo microscopy.

Figure 3.Brightfield image of endothelial cells.

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Ausgewählte Publikationen

Krause DS, Lazarides K, Lewis JB, von Andrian UH, Van Etten RA. Selectins and their ligands are required for homing and engraftment of BCR-ABL1+ leukemic stem cells in the bone marrow nicheBlood 2014; 123(9): 1361-1371 Krause DS, Fulzele K, Catic A, Sun CC, Dombkow-ski D, Hurley MP, Lezeau S, Attar E, Wu JY, Lin HY, Divieti-Pajevic P, Hasserjian RP, Schipani E, Van Etten RA, Scadden DT. Differential regulation of my-eloid leukemias by the bone marrow microenviron-ment. Nature Medicine 2013; 19(11):1513-1517*

Fulzele K*, Krause DS*, Panaroni C, Saini V, Barry KJ, Lotinun S, Baron R, Bonewald L, Feng JQ, Chen M, Weinstein LS, Wu JY, Kronenberg HM, Scadden DT, Divieti-Pajevic P. Myelopoiesis is regu-lated by osteocytes through Gsα-dependent signaling Blood 2013 Feb 7;121(6):930-9.

Krause DS, Lazarides K, von Andrian UH, Van Etten RA. Requirement for CD44 in homing and engraftment of BCR-ABL-expressing leukemic stem cells. Nat Med 2006; 12 (10):1175-80

*co-first authorship

... weitere Publikationen finden Sie auf Seite 57

Figure 4.Brightfield image of differentiated stroma cells.

Figure 5.Brightfield image of differentiated osteoblasts.

Other activitiesWe coorganized the second interna-tional scientific workshop on the “Tumor environment in haematological malig-nancies and its therapeutic targeting” under the umbrella of the European School of Haematology in Berlin. The third workshop will take place in 2019.

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Bone Marrow MicroenvironmentHind Medyouf

Die Knochenmarksnische bei malignen Erkrankungen des menschlichen Blut-systems

Hematological malignancies are driven by self-renewing malignant cells that, similar to their healthy counterpart, reside in a highly complex and dynamic cellular microenvironment in the bone marrow. This so called “niche” is composed of a complex mixture of cell types, includ-ing mesenchymal, nervous, immune and endothelial cells all of which pro-vide signals, that tightly control cardinal features of hematopoietic stem cells (HSCs) that are essential to maintain-ing cellular homeostasis and blood regeneration throughout life. Although hallmarks of cancer are thought to be acquired via somatic mutational events that progressively provide neoplastic cells with a fitness advantage (Mossner et al, Blood, 2016), compelling evi-dence points at the critical importance of the surrounding microenvironment (reviewed in Medyouf, Blood, 2017).

Our goal is to further our understand-ing of the complex biology of human hematologic malignancies by focusing on the complex cellular cross-talk that takes place in the bone marrow niche and dissecting the extrinsic mechanisms

The Bone Marrow Microenvironment in Human Hematological Malignancies

Bone marrow niche

Cellular crosstalk

Patient-derived models

MitarbeiterEwelina CzlonkaArnaud DescotIrene Tirado GonzalezAleksandra NevmerzhitskayaDevona SoetopoMaresa Weitmann

GruppenleiterinHind MedyoufTel.: +49 69 63395-540Fax: +49 69 [email protected]

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Stammzellen des Blutsystems sind in einer höchst komplexen und dynamischen Mikroumgebung im Knochenmark zu finden. Vermutlich tragen Störungen dieser Knochenmarksnische ganz wesentlich zur Entstehung bösartiger Blutkrebsarten bei, indem sie das Wachstum der entarteten auf Kosten der normalen Zellen fördern (Medyouf, CSC, 2014; Medyouf, Blood, 2017). Mit Hilfe von OMICs-Analysen möchte unser Labor diese spezifischen, pathogenen Veränderungen der Nische identifizieren und näher charakterisieren. Außerdem sollen neuartige Modelle entwickelt werden, um die biologische Bedeutung der Nischenveränderun-gen herauszuarbeiten.

Dazu gehören die Editierung von Xenotransplantationsmodellen (PDX) aus Patientenmaterial mit Hilfe der CRISPR-Cas Technologie wie auch ex vivo konstruierte und vollständig humanisierte dreidi-mensionale Nischengerüste. Solcherart „individualisierte“ Modelle können experimentell manipuliert (Auswahlprüfverfahren für neue Medikamente, gentechnische Veränderungen) und mittels bildgebender Systeme „live“ untersucht werden, um so qualitative und quantitative Fakten zu den spezifischen Wechselwirkungen zu sammeln. Als oberstes Ziel möchte unser Labor aber diese Platt-formen nutzen, um einen eventuellen therapeutischen Nutzen bei der gezielten Intervention mit spezifischen Nischeneigenschaften herauszufinden, den wir dann letztendlich auf neue Behandlungs-ansätze für die betroffenen Patienten übertragen können.

that modulate the biology of leukemic cells. The overarching goal is to eventu-ally provide novel therapies to patients with blood cancer (Figure 1). To achieve this, we carry out a translational research program that is based on key observa-tions that emerge from the molecular analysis of stromal and leukemic cells derived from primary patient biopsies. We then apply state-of-the-art genetic tools (CRISPR/Cas9) and inhibitor studies to evaluate the functional relevance of our findings using in vitro co-culture systems (2D and 3D), as well as in vivo modeling of human diseases using both syngeneic models and patient-derived xenografts.

New tools to interrogate the relevance of niche-produced factors in leukemiaCRISPR/Cas9 based workflow for the rapid generation of genetically modified (GM) NSG mice Studies addressing the role of the microenvironment in human leukemia are scarce due to the general lack of genetically modified NSG mice, the most commonly used strain to study human hematologic malignancies, in vivo. We took advantage of CRISPR/Cas9

technology and cytoplasmic injection in NSG zygotes, to establish a workflow that allows for the fast and cost-effective generation of GM NSG mice (Figure 2A). We took advantage of operating HDR mechanisms to carry out precise genome editing in NSG zygotes using CRISPR/Cas technology. Knock out lines can then be generated by using a template DNA that incorporates an in frame stop codon in an early translated exon, which drives non-sense mediated mRNA decay and halts translation in a premature fashion (Figure 2B). We used this strategy to generate an NSG line in which the matricellular protein, Secreted Protein Acidic and Rich in Cysteine (SPARC), is knocked out to study the specific function of niche produced SPARC in human leukemia (see below, Tirado-Gonzalez & Medyouf, Leukemia, accepted). Similar to their C57BL/6 counterpart, NSG SPARC-deficient mice appear normal until around 4-6 months of age, when they develop severe cataract formation (Figure 2C). Of note, this workflow is highly efficient and significantly faster than the time required to backcross a genetically modified C57BL/6 mouse strain onto

an NSG background. Notably, our work will greatly facilitate the study of cellular cross-talk in the tumor microenvironment in the context of patient-derived xenograft models.

Fully humanized 3D niche modelsAlthough 2D systems in which tumor cells are co-cultured on stromal feeder layers are widely used, they often poorly recapitulate the complexity observed in vivo. To further improve our studies on the role of the bone marrow microenviron-ment in the pathogenesis of hematologi-cal malignancies, we recently developed a fully humanized bioengineered 3D bone marrow niche model. As opposed to current models, which are based on hydrogel scaffolds seeded with mesenchy-mal niche cells (MSCs) only, our models are engineered using the two essential components of the bone marrow niche, namely MSCs and endothelial cells. Importantly both cell types are directly isolated from patient-derived bone marrow and shown to actively contribute to the pathogenesis of leukemia (Figure 3A; Członka & Medyouf, in preparation). When cultured under optimized condi-

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tions, MSCs can instruct the formation of vessel-like structures and the scaffold adopts a meshwork structure that sup-ports the maintenance of slowly cycling human CD34+ hematopoietic stem/pro-genitor cells (Figure 3B). These 3D models are now being used as a valuable and easily accessible platform to functionally explore and validate the cellular interac-tions between tumor cells and bone marrow niche in a setting that is highly amenable to experimental manipulation.

The matricellular protein SPARC is a potential novel therapeutic target in relapsed pediatric BCP-ALL. SPARC is a multi-faceted matricellular protein that regulates key physiological processes by modulating cell-cell and cell-matrix interactions. SPARC is shown to be differentially expressed in various tumors and proposed to modulate cancer cell activity. In leukemic contexts, cell intrinsic expression of SPARC has been attributed opposing effects, depending on the leukemia type or even the specific

subtype under study. Importantly its leukemia specific upregulation appears to be a recurrent event in relapsed pediatric B-ALL (Chow YP, BMC cancer, 2017). Despite its high stromal expres-sion, so far, no study has investigated the specific role of niche-produced SPARC in human leukemia, in an in vivo setting. Because mouse and human SPARC have a 92% sequence identity at the protein level, we used our newly generated NSG SPARC deficient line to explore the functional importance of niche-produced


Figure 1.Targeting the niche as a new avenue for leu-kemia therapy is an active area of research in our group. Our work, currently aims to (1) explore the molecular impact of disease associated niche changes on normal and malignant hematopoiesis, (2) identify the nature of the instructive signal responsible for the niche re-shaping and (3) evaluate the therapeutic benefit of targeting the specific cross-talk between niche and hematopoietic cells.

Figure 2.Generation of genetically modi-fied NSG mice using CRISPR/cas9 General workflow and details of template DNA used for targeting the Sparc locus are depicted. Homozygous SPARC edited mice exhibit complete loss of Sparc mRNA and SPARC protein and develop severe cataract pheno-type at 4-6 onths of age, similar to their C57BL/6 counterpart.

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SPARC on the propagation of relapsed pediatric B-cell precursor-ALL (BCP-ALL) in vivo, using PDX modeling. We focused on relapsed pediatric BCP-ALL, as these children often have a dismal prognosis. When compared to wild type mice, SPARC deficient animals exhibit significantly reduced leukemic burden, pointing to a potentially critical role of SPARC in BCP-ALL biology. Ongoing studies are exploring the specific programs that medi-ate SPARC downstream effects, in vivo. Identifying such programs might pave

the way to the design of new therapeutic strategies, in particular for patients in urgent need for alternative therapies.

Conclusion and future directionsDeciphering the complex interplay between niche and hematopoietic cells in normal and disease contexts is increasing our understanding of dis-ease pathogenesis and providing us the knowledge necessary to devise new strategies to specifically dampen the niche support towards malignant cells.



Tirado-Gonzalez I, Czlonka E, Nevmerzhitskaya A, Soetopo D, Bergonzani E, Mahmoud A, Contreras A, Jeremias I, Platzbecker U, Bourquin JP, Kloz U, Van der Hoeven F, and Medyouf H (2017). CRISPR/Cas9 edited NSG mice as PDX models of human leukemia to address the role of niche-derived SPARC. Leukemia. In press.

Medyouf H. The microenvironment in human myeloid malignancies: emerging concepts and therapeutic implications. Blood. 2017. Mar 23;129(12):1617-1626. doi: 10.1182/blood-2016-11-696070.

Mossner M, Jann JC, Wittig J, Nolte F, Fey S, Nowak V, Obländer J, Pressler J, Palme I, Xanthopoulos C, Boch T, Metzgeroth G, Röhl H, Witt SH, Dukal H, Klein C, Schmitt S, Gelß P, Platzbecker U, Balaian E, Fabarius A, Blum H, Schulze TJ, Meggendorfer M, Haferlach C, Trumpp A, Hofmann WK, Medyouf H*, Nowak D*. Mutational hierarchies in myelodysplastic syndromes dynamically adapt and evolve upon therapy response and failure.Blood. 2016 Sep 1;128(9):1246-59.

Medyouf H, Mossner M, Jann JC, Nolte F, Raffel S, Herrmann C, Lier A, Eisen C, Nowak V, Zens B, Müdder K, Klein C, Obländer J, Fey S, Vogler J, Fabarius A, Riedl E, Roehl H, Kohlmann A, Staller M, Haferlach C, Müller N, John T, Platzbecker U, Metzgeroth G, Hofmann WK, Trumpp A, Nowak D.Myelodysplastic Cells in Patients Re-program Mesenchymal Stromal Cells to Establish a Transplantable Stem Cell-Niche Disease Unit. Cell Stem Cell. 2014. Jun 5;14(6):824-37.

* Co-senior authors... weitere Publikationen

finden Sie auf Seite 57

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Figure 3.Fully humanized 3D niche models Schematic view of the 3D co-culture setting combining human derived MSCs and ECs from bone marrow biopsies. Human CD31 and vWF mark endothelial cells and the vessel like structures that can be observed, NG2 marks MSCs and DAPI nuclei. Bright CD34 staining identifies human HSPCs that were subsequently seeded on the scaffold. CFSE labeled CD34+ cells were cultured under standard culture conditions or seeded on human 3D scaffold. As anticipated, CFSE analysis 72h post seeding shows a reduced cycling in the scaffold, likely indicative of maintenance of more quiescent stem/progenitor cells.

Figure 4.Niche-produced SPARC modulates the biology of relapsed pediatric BCP-ALL. Non-conditioned NSG WT or SPARC deficient mice were engrafted with two independent relapsed pediatric BCP-ALL cases (ALL-199 & ALL-265). Leukemic burden is compared using human specific antibodies by flow cytometry. Data show a significantly lower burden in SPARC deficient recipients. Student’s t-test. *p<0.05, ** p<0.01.

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Regulation and deregulationMartin Zörnig

Regulationsmechanismen des programmierten Zelltodes (Apoptose)

Alteration of the control mechanisms of cell death contributes to the pathogenesis of many human diseases, including cancer and neurodegenerative diseases. Our group is interested in identifying novel anti-apoptotic oncoproteins that are responsible for tumor initiation, progres-sion and/or therapy resistance in particular cancer entities. We are analyzing the precise molecular mechanisms of how these proteins inhibit cell death and support tumor growth, and we are validating their potential as targets for future molecular therapies. At the same time, we are investigating their physiologi-cal role in vivo to predict potential side effects of molecular targeting strategies.

Identification of novel mam-malian proteins that regulate apoptosome activitySeveral apoptotic stimuli, including cancer treatment regimens of radiation and chemotherapy, ultimately result in the activation of the mitochondrial apoptosis pathway. Inhibition of this pathway allows malignant cells to evade cell death successfully during tumorigenesis

Regulation and deregulation of apoptosis

Regulation of the mitochondrial apoptosis pathway

Targeting of anti-apoptotic oncoproteins

Analysis of the transcriptional FUBP1 network

MitarbeiterKatharina GerlachViktoria von MansteinVan Thanh HoangMarlene SteinerAlisa MaringSusanne Bösser

GruppenleiterMartin ZörnigTel.: +49 69 63395-115Fax: +49 69 [email protected]

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Unsere Arbeitsgruppe beschäftigt sich mit der Identifizierung und Analyse neuer anti-apoptotischer Onkogene, die für die Tumor-entstehung sowie für Therapieresistenzen verantwortlich sind. In der Vergangenheit konnten wir in einem selbst entwickelten „Hefe-Survival-Screen“ mehrere interessante anti-apoptotische Kandidatengene identifizieren, die in bestimmten Tumorentitä-ten überexprimiert werden. Wir untersuchen diese Moleküle in Zellkulturexperimenten und in geeigneten Mausmodellen in vivo daraufhin, welche Rolle sie während der Tumorentstehung und -progression spielen, und ob sie sich als Zielstrukturen für zukünf-tige molekulare Krebstherapien eignen. Parallel sind wir auch

daran interessiert herauszufinden, welche Funktionen diese Gene im gesunden Organismus ausüben. Letzteres erlaubt auch die Abschätzung möglicher Nebenwirkungen bei molekularen Thera-pien, in deren Verlauf die tumorrelevante Funktion solcher Gene gestört werden soll. Für ein „Targeting“ geeigneter Kandidaten-gene bzw. der entsprechenden Onkoproteine versuchen wir, kleine inhibitorische Moleküle und shRNA-basierte Strategien für thera-peutische Zwecke zu entwickeln, um eine Resistenzentwicklung der Tumorzellen zu umgehen und diese für weitere Behandlungen zu sensibilisieren.

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and to acquire therapy resistance which represents the biggest challenge in cancer treatment. Therefore, it is important to identify and characterize novel anti-apoptotic proteins that inhibit Caspase-9 activation within the apoptosome complex during execution of the intrinsic mitochon-drial apoptosis pathway. In several screens, we isolated human genes that inhibit cell death at the level of the apoptosome, and we identified promising candidates as potential targets for cancer therapy.

FUSE Binding Protein 1 (FUBP1) binds to single-stranded DNA and regulates the expression of various target genes. Transcription of the proto-oncogene c-myc for example is activated by FUBP1, while the cell cycle inhibitor gene p21 is repressed by the same protein. We isolated FUBP1 from a breast carcinoma-derived cDNA library and studied its involvement in tumorigenesis. FUBP1 is overexpressed in more than 80% of hepatocellular carcinomas (HCCs) and supports HCC tumor growth by inhibiting anti-proliferative and pro-apoptotic genes. We are currently developing small molecule inhibitors of FUBP1 for

HCC therapy, and we are testing these inhibitors in several different systems, including liver organoid cultures (Fig. 1).FUBP1 also plays an oncogenic role in leukemia. Our results obtained in cell culture and with mouse transplantation leukemia models demonstrate that FUBP1 is required for chronic myelogenous and acute myeloid leukemia (CML and AML) development (Fig. 2). As in HCC tumor cells, interference with FUBP1 activity leads to decreased cell proliferation and elevated apoptosis rates in the leukemia cells. In collaboration with clinicians from the Frankfurt University Clinic, we could show that increased FUBP1 levels correlate with shorter patient survival. Our aim is to identify the FUBP1 target genes that are relevant for leukemogenesis and to explore possible leukemia therapy options using FUBP1 inhibitors.

In parallel studies, we are analyzing the physiological function of FUBP1 in suitable mouse models. Our experiments unraveled a crucial role of FUBP1 for the mainte-nance and self-renewal of both, adult and fetal, hematopoietic stem cells (HSCs) by regulating relevant target genes. The tran-

scriptional FUBP1 network is also involved in the differentiation of embryonic stem cells into cells of the mesoderm germ layer. We also discovered a prominent role for FUBP1 for murine and human erythroid differentiation of hematopoietic cells (Fig. 3). The upregulation of FUBP1 during erythropoiesis is mediated by the transcriptional master regulator TAL1 (T-cell acute lymphocytic leukemia protein 1) which is co-expressed with FUBP1.

The anti-apoptotic protein AVEN was also isolated in one of our functional screenings, and it interacts with various regulators of apoptosis, such as the apoptosome adaptor protein APAF-1. We could demonstrate that AVEN requires proteolytic processing by the lysosomal protease Cathepsin D to unleash its full anti-apoptotic potential, thereby implying that AVEN may be involved in the lysosomal apoptotic pathway.

Published data indicate a strong association between poor prognosis in acute childhood lymphoblastic leukemia (ALL) and AVEN expression, suggesting that AVEN has oncogenic activity. We

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established a transgenic mouse line with T cell-specific overexpression of the full-length AVEN protein, which acceler-ates leukemogenesis in heterozygous p53+/- knockout mice. Moreover, the downregulation of AVEN in T-ALL cell lines reduces tumor growth in xenograft experiments. Both findings demonstrate the significant oncogenic potential of AVEN. AVEN has also been implicated in both DNA repair and the activation of ataxia telangiectasia mutated (ATM) protein kinase, a major regulator of the cell cycle and the DNA damage response. We developed and analyzed a constitutive Aven-/- knockout mouse model, and the results suggest that AVEN plays a vital role in embryonic development. Lack of AVEN expression leads to accumulation of DNA damage and growth arrest, thereby resulting in embryonic lethality at approximately day E9.5. To further study the physiological role of AVEN in adult mouse organs and tissues, and to inves-tigate its oncogenic function in leukemia and breast carcinoma, we established conditional Aven-/- knockout mice. The detailed analyses of FUBP1 and AVEN activity will help to further clarify the

mechanism by which apoptosome assem-bly and Caspase-9 activation are regulated in response to mitochondrial Cytochrome c release. Based on this knowledge, we aim to develop inhibitors for therapeutic intervention targeting the anti-apoptotic activities of these molecules.


Figure 1.Generation of a HCC biobank Schematic overview of liver organoid culture. Adult bile duct-derived bi-potent progenitor cells are isolated from human liver biopsies. Histological and immunohisto-chemical staining of a human HCC patient sample. Upper left: nearby tissue, stained with hematoxylin/eosin (H/E); lower left: nearby tissue, anti-FUBP1; upper right: tumor tissue, H/E; lower right, anti-FUBP1. The tumor cells express significant amounts of FUBP1 protein (brownish color). Cultured organoids of the cor-responding patient sample. Left: organoids derived from nearby tissue; right: tumor tissue-derived organoids. FUBP1 expression in the organoids correlates with FUBP1 expression in normal tissue (low/absent) and HCC biopsies (high).

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Wesely J, Steiner M, Schnütgen F, Kaulich M, Rie-ger MA, Zörnig M. Delayed mesoderm and erythroid differentiation of murine embryonic stem cells in the absence of the transcriptional regulator FUBP1.Stem Cells Int. 2017;2017:5762301.doi: 10.1155/2017/5762301. Epub 2017 May 15.

Rabenhorst U¹, Thalheimer FB¹, Gerlach K¹, Kijonka M, Böhm S, Krause DS, Vauti F, Arnold HH, Schroeder T, Schnütgen F, von Melchner H, Rieger MA¹, Zörnig M1. Single-stranded DNA-bin-ding transcriptional regulator FUBP1 is essential for fetal and adult hematopoietic stem cell self-renewal. Cell Rep. 2015, 11:1847-55.

Eißmann M1, Melzer IM1, Mateus Fernández SB1, Michel G, Hrabě de Angelis M, Hoefler G, Finkenwirth P, Jauch A, Schoell B, Grez M, Schmidt M, Bartholomae CC, Newrzela S, Haetscher N, Rieger MA, Zachskorn C, Mittelbronn M, Zörnig M. Overexpression of the anti-apoptotic protein AVEN contributes to increased malignancy in hematopoietic neoplasms. Oncogene 2013, 32: 2586-91.

¹ these authors contributed equally to the work


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Figure 2.FUBP1 plays an important role in promoting survival and proliferation of CML cells in a BCR-ABL1-induced CML mouse transplantation model. BCR-ABL1+ leukemic cells expressing Fubp1 shRNA showed a growth disadvantage upon transplantation into recipient mice compared to control leukemic cells transduced with scrambled shRNA in all analyzed cell popula-tions. Cell cycle analysis using Ki-67 and DAPI revealed that a higher proportion of Fubp1 shRNA+-transduced progenitor cells was quiescent than in cells transduced with scrambled shRNA. FUBP1-deficient cells also showed an increased apoptosis rate. In concordance, the CML recipient mice transplanted with Fubp1 knockdown cells survived longer than the animals that received transformed cells with FUBP1 wildtype levels. Working hypothesis: Our results indicate that the oncoprotein Fubp1 promotes proliferation and survival in CML leukemia cells, and that inhibition of FUBP1 in combination with the standard chemotherapeutic drug imatinib might represent a promising CML therapy option.

Figure 3.The knockdown of FUBP1 in primary human CD34+ donor cells leads to a reduced number of erythroid colonies during hematopoietic differentiation. Primary human CD34+ cells were transduced to generate FUBP1 knockdown and control cells. Successfully transduced cells were sorted and plated in methyl cellulose under cytokine conditions that allowed growth of colonies derived from different hematopoietic lineages. The FUBP1 knockdown in the colony cells was verified by western blotting. , . No difference was observed in the total number of colonies formed by FUBP1 knockdown and control cells , but the number of erythroid colonies BFU-E (burst forming unit-erythroid) and CFU-E (colony forming unit-erythroid) was significantly reduced in FUBP1-deficient cells .

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Zell-Zell Interaktionen im Tumorstroma Cell-Cell Interaction in the Tumor Stroma

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Diet and Cancer LinkMelek Canan Arkan

Ernährung und Krebs

Precision Medicine calls for Precision NutritionPrecision medicine is a developing concept in cancer therapy that aims at tailoring the right drug for the right type of patient. So far, much of the effort to achieve this goal has been based on genomics. However, the majority of the cancer patients who receive genomic testing do not apparently benefit from a genomic precision medicine strategy. This suggests targeting not only the initial or existing genetic profiles of tumors, but also the functional perturbations of the primary tumor cells, might be a more complete approach especially for patients who are less responsive to conventional therapy. The variation in cancer risk and the inter-individual differences in tumor therapy response in patients is in most part due to tissue- or cell- specific differences and tumor heterogeneity. Derangement in bio-energetic pathways, which is a hallmark of all cancers, may indeed play a dominant role in this heterogeneity. Indeed, alterations in metabolic phenotype and metagenome profiles of different tumors and individuals vary in between along with changes in microbial community and

Diet and Cancer Link

Diet as a major risk factor for cancer

Metabolic vulnerabilities in host and tumor cells

Microbial community changes and disease susceptibility

GruppenleiterinMelek Canan ArkanTel.: +49 69 63395-600Fax: +49 69 [email protected]

MitarbeiterBegüm AküzümSanam SaeifarVeronika Liptakova

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Die Präzisionsmedizin ist ein im Wachsen befindliches Konzept der Krebstherapie, welches darauf abzielt, maßgeschneiderte Medika-mente für den richtigen Patiententyp zu finden. Bisher beruhten die meisten Bemühungen zu diesem Ziel auf der Genomforschung. Allerdings konnte die Mehrheit der Krebspatienten, die genetisch getestet wurden, scheinbar nicht von der genomischen Präzisi-onstherapie profitieren. Eine bessere Herangehensweise wäre daher, nicht nur das initiale oder existierende genetische Profil der Tumore, sondern auch die funktionellen Störungen der primären Tumorzellen zu analysieren, um Therapieansätze zu vervollständi-gen, insbesondere bei Patienten die auf konventionelle Therapien wenig ansprechen.Die Schwankungen des Krebsrisikos und die individuellen Unter-schiede in der Tumortherapieantwort einzelner Patienten, ist größtenteils auf spezifische Unterschiede von Gewebe und Zellen und auf die Tumorheterogenität zurückzuführen. Eine Störung der bioenergetischen Signalwege, die für alle Krebsarten typisch ist,

kann in dieser Heterogenität eine führende Rolle spielen. Tatsäch-lich variieren der metabolische Phänotyp und das Metagenom-Profil von verschiedenen Tumoren und Individuen untereinander ebenso, wie das Mikrobiom und die metabolischen Funktionen. Ob dies jedoch eine Ursache oder Folge der Krankheitsprogres-sion ist, ist bisher weitgehend unbekannt. Angesichts der hohen Plastizität von Tumoren, sich an verfügbare Energieressourcen anzupassen, um ihr Wachstum und ihre Verbreitung zu sichern, sind unsere Studien, die darauf abzielen die wichtigsten Verände-rungen im Energiestoffwechsel während der Karzinogenese und der Tumortherapie aufzudecken, von kritischer Bedeutung. Die gewonnenen Erkenntnisse sollen erörtern ob Kontertherapien wie eine Präzisionsernährung, welche metabolische Schwachstellen angreifen oder verändern kann, ein neuartiges Therapiekonzept in Krebspatienten darstellen könnte.Unsere Gruppe wird vom Institut für Biochemie II, Goethe Univer-sität Frankfurt finanziert.

metabolic functions, although whether this is a cause or consequence of disease progression remains largely unknown. Thus, given the tumors’ high plasticity in shifting and adapting to available energy sources for rapid growth and dissemina-tion, our studies that aim to define the key changes in energy metabolism during carcinogenesis as well as during therapy will be of critical importance. This will help us to define whether counter approaches such as precision nutrition that target and/or change metabolic vulnerability of a tumor may offer therapeutic options in cancer patients.

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Figure 1.Measuring derangements in tumor energy metabolism: Different tumor cells were injected with Oligomycin, FCCP, Rotenon/antimycin A and their oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) were measured using Seahorse Metabolic Analyzer.

Metabolic Derangements in Diet-induced CancerPancreatic ductal adenocarcinoma (PDAC) is one of the most lethal types of cancer, with high death rates shortly after diagnosis. This is majorly due to the hidden localization of the organ, the rather non-specific symptoms of the disease and the low efficacy of imaging tools in early diagnosis. There are several genes and signaling pathways involved in pancreatic cancer. However, the mutation of K-RAS is the major driving force of this pathology and is detected in early premalignant lesions, termed as pan-creatic intraepithelial neoplasia (PanIN). Although, there has been considerable

amount of research that focuses on therapeutic approaches such as specific inhibition of these signaling pathways, unfortunately cancer-associated mortality rates continue to increase questioning the validity of these current approaches in the past decades. Epidemiological studies suggest diet-induced obesity is closely associated to pancreatic cancer and increased BMI correlates positively with the mortality rates. These studies suggest a strong correlation between altered tumor metabolism and disease development. Thus, targeted derange-ments in energy metabolism may provide potential approaches for therapeutic interventions in PDAC (Figure 1).

OCR ECAR Energy Map

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The crosstalk between diet, microbiota, metabolites and cancer The microbial community in the gut has a decisive role in determining our health. Indeed, dysbiotic microbiota is closely associated with tumor development in colorectal cancer patients. Metabolic activity of bacteria affects food diges-tion, absorption, immunity, and energy production. However, how microbiota composition and functional profiles shape cancer susceptibility under deranged metabolism needs further insight. Recent evidence suggests that bacterial metabolites may indeed serve as the main hub for communication between the gut and extra-intestinal organs. Even though far from their site of production, bacterial metabolites can employ systemic changes providing an important means of microbiome-host-tumor interactions. Microbial community is highly personal-ized and exhibits a high degree of inter-individual variability due to hosts’ genetic background, physiology, and lifestyle. Although, the gut microbiota is shaped early on during birth, there are several factors that affect the composition of microbiota during childhood and

adulthood. Among them diet appears to be a consistent and prominent one. The dynamics of the bacterial community is critical because inter-bacterial interactions may correlate with and/or determine the inter-individual variation in susceptibility to cancer as well as varying therapy responses. Thus, determining the role of community changes and their functional profiles under a cancer setup with altered energy metabolism can pave the way for individual-based interventions (Figure 2).The current complexity of cancer as a disease and the lack of effective treatment options is in most part due to tumor heterogeneity and tissue- or cell- specific differences. This, combined with the complex nature of food, the nutritional values as well as the inter-individual differ-ences in dietary intake, makes it a difficult task to define and use food as medicine. Indeed, there are studies that suggest personalized diets over general dietary recommendations to be more useful especially in the treatment of metabolic diseases. Nevertheless, there is strong evidence that diet is a major risk factor for the development of certain types of cancer and modifications in nutrition

already exist in the guidelines for cancer survivors. Interventional approaches currently hold great promise for the use of dietary factors in malnourished cancer patients to increase quality of life and in reducing cancer risk/cancer prevention. Approaches for deliberately reshaping the highly adaptive and durable relationship between the gut microbiome and its host in a way to promote health is going to be the ultimate aim in cancer therapies. In order to increase the effectivity of nutritional therapies, strategies on how to avoid the confounding effect of prior dietary practices and to ensure that gut microbiota respond consistently to prescribed dietary interventions will be of great importance in cancer patients.

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Arkan MC. The intricate connection between diet, microbiota, and cancer: A jigsaw puzzle. Semin. Immunol. 2017. Sep 1. pii: S1044-5323(17)30023-4.

Arkan MC. Cancer: Fat and the fate of pancreatic tumours. Nature 2016. 536 (7615): 157-8.

Schulz MD, Atay C, Heringer J, Romrig FK, Schwitalla S, Aydin B, Ziegler PK, Varga J, Reindl W, Pommerenke C, Salinas-Riester G, Böck A, Alpert C, Blaut M, Polson SC, Brandl L, Kirchner T, Greten FR, Polson SW, Arkan MC. High-fat-diet-mediated dysbiosis promotes intestinal carcinogenesis independently of obesity. Nature 2014. 514 (7523): 508-12.

Khasawneh J, Schulz MD, Walch A, Rozman J, Hrabe de Angelis M, Klingenspor M, Buck A, Schwaiger M, Saur D, Schmid RM, Klöppel G, Sipos B, Greten FR, Arkan MC. Inflammation and mitochondrial fatty acid beta-oxidation link obesity to early tumor promotion. Proc. Natl. Acad. Sci. U.S.A. 2009. 106 (9): 3354-9.


Figure 2. Determining bacterial metabolism in cancer models: Different diets can lead to varying alterations in microbiota. Bacterial community changes are detected by 16S rRNA gene sequencing and bacterial metabolic func-tions are expressed as KEGG annotations.


Organismal Systems; Digestive System;

Metabolism; Lipid Metabolism;

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Microenvironmental crosstalkHenner Farin

Gewebsinteraktionen und Signalmechanismen im Darmkrebs

Colorectal cancer (CRC) is the third leading cause of death from cancer among adults in Germany. Cancer genome sequencing programs have identified a heterogeneous spectrum of oncogenic mutations in patients. However, we currently cannot predict therapeutic responses based on the genetic composition of a tumor. This is due to the complexity of cell signaling processes that involve an intrinsic crosstalk between all tissue compartments. Our lab explores the 3-D ‘organoid’ culture system as a solid tumor model. This system allows expansion of primary intestinal stem cells under full control of the exogenous ‘microenvironment’. Exposure to stromal signals such as inflammatory cytokines, growth factors, microbial and metabolic cues are analyzed to dissect tumor-specific vulnerabilities. Our goal is to understand how tumor cells respond to and influ-ence their microenvironment to be able to specifically target this crosstalk.

The organoid culture system as a human colorectal cancer modelTumor cells successively acquire muta-tions that confer unrestricted local growth before progression to metastatic disease.

Signaling crosstalk in the colon cancer microenvironment

3-D epithelial cultures from endoscopic biopsies

Paracrine signaling mechanisms of the intestinal stem cell niche

Targeting of the colon cancer microenvironment

MitarbeiterMohammed MosaBirgitta MichelsMoyo GrebbinConstantin MencheTahmineh DarvishiTheresa Schnalzger

GruppenleiterHenner FarinTel.: +49 69 63395-520Fax.: +49 69 [email protected]

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Unsere Arbeitsgruppe am Georg-Speyer-Haus erforscht die zellulären und molekularen Vorgänge bei der Entstehung von Darmkrebs. Insbesondere interessiert uns die Kommunikation ver-schiedener Zelltypen in der unmittelbaren Umgebung des Tumors, dem so genannten “Tumor-microenvironment”. Dabei nutzen wir „Organoide“, ein neuartiges dreidimensionales Gewebekultur-System. Organoide können unter definierten Kulturbedingungen aus humanen Darm-Stammzellen etabliert werden und bilden Darmepithel-spezifische Strukturen wie Krypten (Furchen) oder Villi (Zotten) aus (so genannte "Mini-Därme"). Dieses System ermög-licht die Expansion von Stammzellen in einem Gewebe-ähnlichen Zustand, was die Untersuchung von molekularen Signalen in einer definierbaren Mikroumgebung ermöglicht. So kann z.B. durch Zugabe von nicht-epithelialen Zellen wie Fibroblasten, Gefäß- oder Immunzellen der Organkontext nachgebildet werden.

Im Mittelpunkt unserer Forschung steht die genetische Analyse der Entstehung und Progression des Darm-Karzinoms sowie der Einfluss körpereigener Abwehrmechanismen wie Entzündungs-reaktionen. Dazu werden in klinischer Kollaboration Tumorbiop-sien expandiert um Patienten-spezifische Signalmechanismen zu identifizieren. Mit Hilfe von genetischen Techniken (CRISPR/Cas9) und Hochdurchsatzanalysen (z.B. RNA-Sequenzierung und Prote-omanalyse) versuchen wir zu verstehen, wie einzelne onkogene Mutationen den zellulären Phänotyp beeinflussen, als Ansatzpunkt für zukünftige Therapien.

Unsere Gruppe wird vom Deutschen Krebsforschungszentrum (DKFZ) im Rahmen des Deutschen Konsortiums für Translationale Krebsforschung (DKTK) am Georg-Speyer-Haus finanziert.

Although great advances have been achieved in prevention and therapy, the clinical options for progressed CRC patients are still very limited. Metastatic coloniza-tion to distant sites, which are mainly the liver and the lungs, often precludes surgical and radiological intervention. The response rates to classical chemotherapy and targeted therapies are modest with a high degree of relapse. To better predict drug efficiency and prevent develop-ment of resistance, new tumor models are required that recapitulate the signal-ing processes in CRC. Currently used transformed cell lines have lost important physiologic traits of tumor cells and cannot fully reflect the heterogenic nature of the disease. Species barriers limit the use of animal models to test genetic hypotheses.

The ‘organoid’ culture system (first described by Sato et al., 2009) allows expansion of gastrointestinal stem cells over long periods ex vivo. In a 3-D extra-cellular matrix epithelial structures are formed that undergo continuous self-renewal and differentiation, recapitulating a normal crypt-villus architecture (Figure 1). Organoids can be established from

Figure 1.Organoid cultures recapitulate the intestinal stem cell nicheSchematic illustration of the intestinal organoid culture system. The medium composition mimics the stem cell niche environment in the intestine, which is characterized by high WNT/EGF and low BMP signals. In 3-D Matrigel ‘mini guts’ are formed that consist of a single-layered epithelium. Proliferation occurs in the crypt–like structures, each composed of a stem cell niche compartment (red fluorescence: Paneth cells).

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endoscopic patient biopsies of normal and tumor tissue. Importantly, the culture conditions preserve cells in a native state and stem cell niche signals have to be supplemented with the culture medium. This allows functional selection for pres-ence of oncogenic mutations e.g. affecting the WNT, BMP or the EGF pathways. In our lab we exploit and further improve this technological platform as a CRC model.

Characterization of the normal and tumorigenic stem cell nicheIIn the normal intestine, the Wnt path-way controls epithelial self-renewal and differentiation. Constitutive Wnt path-way activation is the main driver of CRC affecting >90% of all cases. We have shown previously that mouse intestinal organoids produce Wnt3 by a specific cell type, the Paneth cells, and moreover critically depend on Wnt3 as an epithelial niche signal (Farin et al., Gastroenterology 2012). Thus organoids provide a unique

and accessible experimental system to investigate the mechanisms that confine Wnt signaling. By combination of mouse transgenesis, in vitro manipulation of organoids and confocal microscopy we could recently demonstrate that Wnt3 represents a short-range signal that is directly transferred from Paneth cells to the adjacent stem cells (Farin et al., Nature 2016). For the first time we could visualize that endogenous Wnt3 protein covers the basolateral membranes of crypt cells (Figure 2). Once stem cells loose their

Figure 2.Localized Wnt signals compartmentalize stem cells and differentiation in intestinal organoids Endogenous Wnt3 protein is produced by Paneth cells and acts as a niche factor. Confocal microscopy shows membranous distribution of Wnt3 that covers the crypts. Model for the short-range action of the Wnt3. Niche contact-dependent transfer and limited diffusion result in a Wnt3 gradient that orchestrates self-renewal and

differentiation (Farin et al., Nature 2016).

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Farin HF*, Jordens I, Mosa MH, Basak O, Korving J, Tauriello DVF, de Punder K, Angers S, Peters PJ, Maurice MM, Clevers H.* (2016). Visualization of a short-range Wnt gradient in the intestinal stem-cell niche. Nature 530, 340–343.

Tetteh PW, Farin HF, and Clevers H. (2015). Plasticity within stem cell hierarchies in mammalian epithelia. Trends in Cell Biology 25, 100–108.

Yin X, Farin HF, Es JH, Clevers H, Langer R, Karp JM. (2014) Niche-independent high-purity cultures of Lgr5+ intestinal stem cells and their progeny. Nature Methods, 11: 106 – 112.

Farin HF, Van Es JH, Clevers H. Redundant sources of Wnt regulate intestinal stem cells and promote formation of Paneth cells. Gastroenterology 2012, 143: 1518 – 1529.

* co-corresponding authors


physical contact to the Paneth cell niche, the surface-bound Wnt3 can only support limited divisions as transit amplifying (TA) cells. Thus self-renewal, cell cycle exit and differentiation critically depend on the cellular context within the crypt.

The oncogenic microenvironment as a potential therapeutic targetIn vivo the gastro-intestinal epithelium is in close contact to surrounding tissues such as mesenchymal, immune and endothe-lial cells and is furthermore continuously

exposed to the gut lumen that contains potentially toxic microbes. In this dynamic environment, the stem cell niche secures homeostasis. Non-controlled epithelial expansion in colon cancer also results in a modified microenvironment (Figure 3): Cancer cells actively recruit stromal cells such as immune cells, vasculature and fibroblasts to obtain trophic signals and repress immune surveillance. In order to survive at secondary sites, disseminated tumor cells need to re-establish a sup-portive stroma. Given this complexity,

tumor organoids can provide a ‘reduc-tionist approach’ to dissect and to model the epithelial-stromal crosstalk: To study cell-intrinsic features we have engineered defined oncogenic mutations in normal human colon organoids using the CRISPR/Cas9 system. To reflect the clinical hetero-geneity we generate organoid biobanks from primary tumors and metastases. The resulting microenvironmental changes are then addressed in co-cultures with stromal cells and by xenotransplantation. Diverse cellular and molecular read-outs (RNA sequencing, Proteome) allow us to link genotypes and phenotypes with the goal to identify new strategies to interfere with the stromal crosstalk.

The research group is funded by the German Consortium for Trans-lational Cancer Research (DKTK) which is part of the German Cancer Research Centre (DKFZ).

Figure 3.Paracrine signaling in colon-cancer microenvironment Histology (HE staining) of grafted tumor organoids (subcutaneous). The tumor epithelium shows a glandular architecture. Recruitment of stromal cells is observed such as blood vessels and fibroblasts. Schematic representation of tissue interactions in colon cancer. Boxes highlight cellular processes that offer potential anti-cancer targets.

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Cell PlasticityFlorian Greten

Zellplastizität im Mikromilieu des Kolonkarzinoms

Carcinogenesis is not a cell autonomous process. Instead, it requires an intricate reciprocal interplay between mutagenized cells and their surrounding stroma. Initiated tumor cells secrete chemokines and growth factors that cause remodeling of recruited inflammatory immune cells, stromal fibroblasts and vascular cells. These stroma cells communicate to each other and induce their respective polarization to ultimately form a niche that supports tumor cell growth. Thus, tumorigenesis relies greatly on the plasticity of both tumor and surrounding cells within the tumor microenvironment: cells of the adaptive immune system as well as the innate immune system create an immune tolerant network in the tumor stroma that affects basically every aspect of tumor cell biology by secretion of cytokines and growth factors that act in paracrine, autocrine, and juxtacrine manners to activate signaling pathways in both cancer and stromal cells. However, tumor growth does not simply depend on the presence of these cells. Instead, particularly infiltrating immune cells are characterized by a high degree of plasticity and therefore, it is their respective polar-

Cell Plasticity in the Intestinal Tumor Microenvironment

Tumorigenesis relies on the plasticity of both tumor and stroma cells

Inflammation controls cell plasticity

Stemness and EMT are controlled by the microenvironment

MitarbeiterVerawan Boonsanay-Michel Özge CanliFatih CeticiClaire Conche Christin DanneilNatalia DelisTiago de OliveiraFabian FinkelmeierJalaj GuptaPreeti GuptaHana KunkelBirgit LehmannKathleen MohsTobias NeumannAdele NicolasMarina PesicMallika RamakrishnanEva RudolfJulia VargaPaul Ziegler

GruppenleiterFlorian R. Greten, DirektorTel.: +49-69-63395-232Fax: [email protected]

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Der Fokus unserer Forschung liegt auf der funktionellen Analyse des Mikromilieus im Kolonkarzinom. Unter Verwendung von kon-ditionalen Knockout-Mäusen, mit denen wir eine Zelltyp-spezifische Aktivierung oder Inaktivierung bestimmter Gene erreichen, führen wir funktionelle Untersuchungen durch. Hierbei kommen relevante Mausmodelle für das Kolonkarzinom zum Einsatz, welche die ver-schiedenen Stadien der Tumorentstehung valide abbilden. Zell-Zell Interaktionen spielen eine besonders große Rolle in der Entwicklung einer Zellplastizität sowohl in Tumorzellen als auch in Stromazellen. Der Tumor benötigt ein hohes Maß an Plastizität dieser verschiede-

nen Zellen und schafft sich die geeigneten Voraussetzungen dafür während aller Stadien seiner Entwicklung: Initiation, Promotion und Progression. Seit vielen Jahren beschäftigen wir uns mit der systema-tischen Analyse eines entzündlichen Mikromilieus im sporadischen und Entzündungs-assoziierten Kolonkarzinom. Darüber hinaus ent-wickeln wir die einzusetzenden Mausmodelle kontinuierlich weiter, um die Situation in Patienten so gut wie möglich in Mäusen abzubil-den. Diese Modelle verwenden wir einerseits um Pathogenese des Kolonkarzinoms zu untersuchen, insbesondere aber auch um neue therapeutische Ansätze zu validieren.

ization profile rather than just their number that decides whether these immune cells confer pro- or anti-tumorigenic properties. A second important and extremely plastic cell type comprises cancer-associated fibroblasts (CAFs). CAFs enhance stiffness of the extracellular matrix, stimulate stemness, cancer survival, growth and invasion, contribute to angiogenesis by releasing pro-angiogenic factors, contrib-ute to a pro-inflammatory milieu, and have impact on the polarization of immune cells thereby affecting every aspect of multi-step tumorigenesis. Different stromal cell types control essential effector mecha-nisms involved in their polarization and activation state. Yet, upon remodeling they also have great impact on the actual tumor cells. Importantly, this is not only limited to cancer cell proliferation and survival but stromal cells are capable of controlling cancer cell fate decisions as well.

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Figure 1.Dedifferentiation promotes both tumor initiation and tumor progression Normal ISCs reside in a specialized niche which maintains their stemness and is crucial to proper stem cell function. Inflammatory conditions mimic or expand the normal stem cell niche and allow and promote dedifferentiation of post-mitotic cells, thereby increasing the number of tumor initiating cells. The mutual interaction between the tumor initiating cell and the stroma is key to tumor initiation. Cells that have acquired an oncogenic mutation will change the surrounding stroma in a way that favors tumor initiation. On the other hand, primary changes in the stroma will re-wire key signaling pathways and induce a tumor-initiating phenotype in the epithelial compartment. Similarly to their normal counterparts CSCs also reside in specialized niches. Inflammation induced expansion of the CSC niche pushes more differentiated cancer cells towards the CSCs state, which increases the number of CSCs and enhances tumor progression. The mutual interaction between the tumor stroma and tumor cells constantly remodels the tumor micro-environment and contributes to tumor progression. Green cells represent normal- and cancer stem cells or cells that underwent dedifferentiation. Blue circles correspond to the normal stem cell niche; red circles indicate an abnormal niche.

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Most tumors, albeit to different extent, resemble the tissue they originate from. Thus, it was suggested that they also preserve some degree of the original hier-archical organization implying that tumors contain more differentiated cells that depend on the presence of stem cell-like cells. These stem cell-like cells are termed cancer stem cells (CSCs) and are believed to maintain tumor growth, be responsible for metastatic dissemination and mediate therapy resistance. One possible concept is that CSCs are basically transformed counterparts of the normal adult SCs and they are the only cells that can be placed to the top of the tumor cell hierarchy. In line with this concept it has been shown that cells that express the intestinal stem cell marker Lgr5 continuously fuel tumor growth, indicating that they are not only important for tumor initiation, but also for tumor maintenance. An alternative and more likely concept suggests that a one directional hierarchy in the tumor or true CSC do not exist suggesting that the term “CSC” describes more a functional state rather than a distinct cell type. As a consequence, any tumor cell could acquire CSC properties when exposed

to the respective environmental signals (Fig. 1). The fact that tumor cells have been proven to be even more plastic than normal non-transformed cells supports this concept. However, if any cell in the tumor can become a CSC, then after their depletion, CSCs would be regenerated from more differentiated cells and their elimination alone would have only little effect on tumor homeostasis. Indeed, it has been recently reported that loss of Lgr5+ cells from established tumors does not lead to tumor shrinkage indicating that non stem cells can compensate for the loss of Lgr5+ cells. CSCs, similarly to their normal counterparts, also reside in specialized niches that may induce and maintain the CSC state. Since tumors are characterized by intense and constant remodelling of the tumor microenviron-ment, changes in the stem cell fates might be highly dynamic. These findings do not necessarily suggest that tumors are not hierarchically organized, but this hierarchy is just as plastic and transient as the differ-ent tumor cells states are. The analysis of such putative CSC and their dependence on signals by the surrounding stroma is one of our main research interests.

Also during later stages of tumorigenesis cell plasticity is essential for tumors to progress. A process called epithelial-to-mesenchymal transition (EMT) which plays fundamental roles in many physiological processes, including embryonic develop-ment and regeneration can often also be observed in pathological conditions, such as organ fibrosis and cancer. It has been suggested that EMT, alongside with the gain of a mesenchymal phenotype, is closely associated with the acquisition of stem cell-like characteristics. While the mesenchymal phenotype is required for the early steps of the metastatic cascade, reversion to the epithelial state via mesenchymal-to-epithelial transition (MET) is necessary for metastatic out-growth, which would support a link between the epithelial state and stemness. Based on mathematical modeling the existence of a third EMT category with both epithelial and mesenchymal properties was suggested. Cells in this hybrid EMT state still maintain cell-cell contacts but acquire migratory phenotype, which allows them to travel in small clusters. It is possible that this hybrid state endows cells with stem cell properties, while

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Figure 2. Proposed model of the relationship between EMT and stemness Neither the epithelial nor the mesenchymal state can be clearly linked to the stem cell state. Instead there is a broader stemness window positioned along the EMT scale. Cells that will fall into this stemness window will exhibit a broad range of EMT phenotypes with the majority being in a plastic state. The fixed epithelial or mesenchymal states are located outside of the stemness window. The position of the stemness window is not fixed and can be moved along the EMT scale depending on the microenvironment (Varga, J & Greten F.R., Nat Cell Biol. 2017;19(10):1133-1141.)

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Diamanti MA, Gupta J, Bennecke M, DeOliveira T, Ramakrishnan M, Braczynski AK, Richter B, Beli P, Hu Y, Saleh M, Mittelbronn M, Dikic I and Greten FR. IKKα controls ATG16L1 degradation to prevent ER stress during inflammation. J Exp Med. 2017;214(2):423-437.

Greten FR.Cancer: Tumor stem-cell surprisesNature. 2017;543(7647):626-627

Varga J and Greten FR.Cell plasticity in epithelial homeostasis and tumori-genesisNat Cell Biol. 2017;19(10):1133-1141

Canli Ö, Nicolas AM, Gupta J, Finkelmeier F, Gon-charova O, Pesic M, Neumann T, Horst D, Löwer M, Sahin U and Greten FR. Myeloid cell-derived reactive oxygen species induce epithelial mutagenesis. Cancer Cell, 2017, in press


being fixed in an extreme epithelial or mesenchymal state does not (Fig. 2).

Just like primary tumors also metastatic out-growth is profoundly influenced by the stroma and metastasizing cells which leave the primary tumor are exposed to a completely new microenvironment. This new microenvironment is often hostile, where disseminated cells are exposed to several harmful stimuli, such as lack of resources and attacks by the immune system. To bypass the adverse environment tumor cells hijack the normal stem cell niche at the initial stages of the metastatic seeding. However later, by re-educating already preexisting niches and by recruiting new stromal components tumor cells can build up a more favorable niche which will help their survival and proliferation.

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Microenvironmental regulationLisa Sevenich

Die Rolle der Tumormikroumgebung in der Hirnmetastasierung

Brain metastasis remains one of the most lethal aspects of cancer with limited effective treatment options that still mostly depend on surgical resection and/or radio- or chemotherapy. To provide better care for brain metastasis patients novel therapies have to be developed. The identification of therapeutic targets however requires a detailed understand-ing of the underlying mechanisms that drive brain metastasis. It is increasingly recognized that in addition to tumor cell intrinsic properties, the crosstalk between tumor cells and the surrounding tissue is an important regulator of disease progression and therapeutic response.

Cancer-associated inflamma-tion in brain metastasisThe brain represents a unique tissue envi-ronment with various highly specialized cell types such as neurons and neuroglia. Neuroglia are a heterogeneous cell popu-lation that comprises astrocytes, oligoden-drocytes, and microglia, the brain resident macrophages. Unlike other tissue resident macrophages, microglia derive exclusively from the yolk sac and populate the devel-oping brain before the blood-brain barrier

Microenvironmental regulation of brain metastasis

MitarbeiterKatja NieselAnna Salamero BoixMichael SchulzWoon Hyung ChaeSebastian FriemelMaria NolteAlexander SchäfferTijna Alekseeva

GruppenleiterinLisa SevenichTel.: +49 69 63395-560Fax: +49 69 [email protected]

Cellular cross-talk in brain metastasis

Therapy-induced inflammatory response

Tumor microenvironment targeted therapy

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Viele Krebserkrankungen können dank intensiver Forschung und den daraus resultierenden Therapiefortschritten erfolgreich behan-delt werden. Metastasen stellen jedoch weiterhin die Haupttodes-ursache bei Tumorpatienten dar, da die verfügbaren Behandlungs-möglichkeiten, insbesondere bei Hirnmetastasen, nur begrenzt wirksam sind. Die Entwicklung neuartiger Therapieansätze zur Bekämpfung von Hirnmetastasen ist daher von großer Bedeutung. Neuere Studien verdeutlichen den Einfluss der Gewebsumgebung auf die Tumorprogression und die organspezifische Metastasie-rung. Die Mikroumgebung von Hirnmetastasen weist aufgrund des Vorhandenseins hirn-spezifischer Zelltypen, wie z.B. Mikroglia oder Astrozyten, im Vergleich zu anderen Organen einige Besonderhei-

ten auf. Der Einfluss dieser hirn-residenten Zelltypen sowie weiterer Entzündungszellen, die in Hirnmetastasen einwandern, ist derzeit weitgehend unbekannt. Das Forschungsziel unserer Nachwuchs-gruppe besteht darin, die komplexen Interaktionen zwischen Tumorzellen unterschiedlicher Entitäten (Melanom, Bronchial- oder Mammakarzinom) und hirnresidenten- sowie rekrutierten Entzün-dungszellen während der Hirnmetastasierung zu entschlüsseln. Ein besonderer Fokus liegt hierbei auf der funktionellen Analyse der Mechanismen, durch die Krebszellen tumor-fördernde Entzün-dungsreaktionen im Gehirn hervorrufen sowie der Fragestellung wie Strahlen- bzw. Chemotherapie diese Vorgänge auf zellulärer und molekularer Ebene beeinflusst.

is fully established. Thus, the brain has long been regarded as an immunologi-cally sanctuary site with restricted entry of immune and inflammatory cells from the periphery. However, brain tumors diminish the integrity of the blood-brain barrier and thus lead to massive invasion of inflam-matory cells from the periphery. The main cellular players of neuro-inflammation in brain metastasis are astrocytes and microglia, as well as blood-borne myeloid and lymphoid cells. In addition, neuro-genic neuro-inflammation is a unique feature of the central nervous system. There is accumulating evidence that inflammatory responses in brain metas-tases support distinct steps within the metastatic cascade. However, the underly-ing mechanisms remain largely unknown. Our goal is to investigate the complex cellular crosstalk in brain metastases from different tumor entities that frequently metastasize to brain, i.e. melanoma, breast- and lung cancer to identify (1) oncogene-driven signals that are involved in stromal education to generate a cancer-permissive environment and (2) to identify stromal-derived stimuli that support metastatic seeding and outgrowth. We

use a comprehensive set of experimental systems to investigate molecular and cel-lular players of CNS inflammation in brain metastasis. These models include in vitro co-culture assays, organo-typic brain slice assays for time lapse imaging of tumor-stroma interactions and experimental brain metastasis models representing different tumor entities that frequently metastasize to the brain (melanoma, breast- and lung cancer) (Figure 1). Close collaboration with the neuropathologists Prof. Plate and PD Dr. Harter from the Edinger Institute (KGU Frankfurt) allows us to analyze specimen from brain metastasis patients which provides important insight into the clinical relevance of our experimental data. Histological assessment on 1mm thick cleared sections of brain metastasis-bearing mice is used for 3D reconstruction of changes in cellular morphology and spatial organization of tumor-associated stromal cells in brain metastasis (Figure 2). Of particular interest is the population of tumor-associated macrophages/microglia in brain metastasis. Under physiological conditions, almost 100% of the macro-phages are yolk sac-derived microglia. Tumor cell invasion and metastatic colo-

nization leads to recruitment of blood-borne macrophages (BMDM) leading to a mixed macrophage population in brain metastases (Figure 2D+E). While both cell types share many physiological features, it remains largely unknown if they exert redundant or cell type-specific roles in brain metastasis and if therapeutic inter-vention differentially affects their effector functions. However, detailed knowledge on brain metastasis-associated and therapy induced inflammation is necessary to develop therapeutic approaches that specifically target aberrant functions of brain resident and recruited inflammatory cells while preserving important physiolog-ical functions of the respective cell types.

Radiotherapy as an amplifier of brain metastasis-associated inflammationThere is accumulating evidence of a close link between radiotherapy and an induc-tion of tumor immunogenicity in various cancer types. Consequently, radiotherapy is regarded as a potential sensitizer of tumors towards a range of immunothera-pies. Our goal is to investigate effects of ionizing radiation on brain metastasis-associated inflammatory cells to character-

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ize changes of immune responses in the CNS (e.g. pro-inflammatory / anti-tumor vs immune suppressive / tumor-promoting responses). We use the Small Animal Radiation Research Platform (SARRP) in collaboration with Prof. Rödel (KGU Frank-furt) to apply radiotherapy to preclinical models as single high dose or fractionated whole brain radiotherapy (WBRT) or as stereotactic radiosurgery (SRS). Magnetic resonance imaging (MRI) using a 7T Small Animal MRI is used to analyze the effects of therapeutic intervention (Figure 3). Moreover, in collaboration with Dr. Pilatus (Brain Imaging Center, KGU Frankfurt) we optimized the use of NMR spectroscopy to follow the radio-response in brain metastasis. NMR spectroscopy allows for non-invasive detection of metabolites (e.g. NAA, Cho, Cr) that can be used as a mea-sure for tumor progression and therapeu-tic response as shown for the ratio of Cho and NAA in Figure 3C that is indicative for cellular density. Analysis of the cellular

composition of the brain metastasis-associated microenvironment by histol-ogy as well as flow cytometry revealed that ionizing radiation leads to enhanced recruitment of blood-borne inflammatory cells in brain metastasis. RNA sequenc-ing showed profound transcriptomic changes at distinct time points after irradiation in different brain metastasis-associated inflammatory cell populations.

Target tumorstroma interactions in brain metastases in monotherapy or adjuvant settings in combina-tion with standard of careCombination of standard therapy with targeted- or immune therapies shows synergistic efficacy in different tumor entities. A central goal of our work is the preclinical testing of different combina-tions of standard of care and immune modulatory therapies in brain metas-tasis models. These strategies include (i) depletion of individual stromal cell

Figure 1. Schematic of the generation of experimental brain metastasis models representing tumor entities that frequently metastasize to the brain (melanoma, breast- and lung cancer). Parental tumor cell lines that show low brain tropism are injected into the left ventricle of mice. Brain metastatic cells are isolated and reinjected to select for variants with high brain tropism. Representative MRI images of a breast-to-brain metastasis model that develops several contrast-enhancing metastatic lesions in the brain parenchyma. Representative images of brain metastasis from the breast-to-brain model. H&E staining and EpCAM staining show the lobular growth pattern of this model that closely mimics the histopathology of patient brain metastasis.

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types, (ii) blockade of their activation or recruitment, (iii) disruption of cellular communication between tumor cells and stromal cells or (iv) targeting of aberrant functions of tumor-associated stromal cells. TME-targeted therapies are tested as monotherapies or in combination with radio- or chemotherapy. While strate-

gies that aim at depleting or blocking the recruitment of individual cell types shows limited efficacy, there is growing evidence that strategies that target aber-rant functions or alter the mode of the immune response are more promising for efficient and long lasting disease control. The long-term goal of our research is

to translate our pre-clinical findings into clinical applications by contribut-ing to the understanding of the complex mechanisms that drive brain metastases to identify novel targets for immune therapies against brain metastases.

Figure 2. Schematic of the procedure of tissue clearance based on X-Clarity. Tissue clearing leads to removal of lipids in organs resulting in transparent tissues that can be used for immunofluorescent staining and 3D reconstruction of tissue architecture and spatial organization of different cell types. Representative images of GFAP+ astrocytes (magenta) and NeuN+ neurons (red) in breast-to-brain and lung-to-brain metastasis. Detection of CX3CR1+ macrophages/microglia in normal brain parenchyma and breast-to-brain metas-tasis. Microglia constitute almost 100% of the brain-resident macrophages in normal brain parenchyma. In contrast, blood-borne macrophages are recruited to brain metastatic lesions and form together with microglia the complex and dynamic population of brain metastasis-associated macrophages/microglia. Flow cytometry allows to discriminate both populations based on CD49d expression.


Bowman RL, Klemm F, Akkari L, Pyonteck SM, Sevenich L, Quail DF, Dhara S, Simpson K, Gardner E.E., Iacobuzio-Donahou C, Brennan CW, Tabar V, Gutin P.H., Joyce J.A. (2016) Macrophage ontogeny underlines differences in tumor-specific education in brain malignancies. Cell Reports 22;17(9):2445-2459

Sevenich L, Bowman RL, Mason SD, Quail DF, Ra-paport F. Elie BT, Brogi E, Brastianos PK, Hahn WC, Holsinger LJ, Massague J, Leslie CS, Joyce JA. Analysis of tumour- and stroma-supplied proteolytic networks reveals a brain-metastasis-promoting role for cathepsin S. Nat Cell Biol. 2014 Sep;16(9):876-88

Pyonteck SM, Akkari L, Schuhmacher AJ, Bowman RL, Sevenich L, Quail DF, Olson OC, Quick ML, Huse JT, Teijeiro V, Setty M, Leslie CS, Oei Y, Pedra-za A, Zhang J, Brennan CW, Sutton JC, Holland EC, Daniel D, Joyce JA. CSF-1R inhibition alters mac-rophage polarization and blocks glioma progression. Nat Med. 2013 Oct;19(10):1264-72



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Figure 3. Dose distribution in whole brain radiation (WBRT) using the SARRP. Nearly 100% of the relative dose is focused on the head region with minimal involvement of the body. Volumetric analysis of tumor progression in control animals compared to irradiated animals using the Small Animal MRI. NMR spectroscopy was used to measure specific metabolites in tumor-free animals, non-treated tumor-bearing animals and tumor-bearing animals after radiotherapy. Shown here is the ratio between Cho and NAA which can be used as a measure cellular density which is changed during tumor progression and therapeutic response.

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Experimentelle Therapie Experimental Therapy

Laboratories III

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Molecular therapyUrsula Dietrich

Molekulare Therapie viraler Infektionen

Antiretroviral therapy (ART) has turned infection with HIV-1 into a chronic disease and, furthermore, the broader availability of antiretroviral drugs has stabilized the number of new HIV-1 infections in many countries. Nevertheless, 1.8 million new infections were still acquired in 2016 worldwide reflecting the urgent need for a preventive vaccine. In the absence of HIV-1 Env immunogens able to induce broadly neutralizing antibodies (bnAbs) against the virus, vaccine research has focused on bnAbs generated during chronic HIV-1 infection in a subset of infected persons. These bnAbs have shown efficacy in animal models and have entered first clinical trials. However, the high production costs and multiple parenteral applications represent a major obstacle in view of broad preventive applications. We therefore concentrated our efforts on broadly neutralizing nanobodies, which are antibody frag-ments corresponding to the variable domain of heavy chain only antibodies from camelids. Due to their small size and favourable physicochemical properties bn nanobodies are promising candidates for clinical development in view of prophy-

Molecular therapy/prevention of viral infections

Identification of nanobodies broadly neutralizing HIV-1

Expression of nanobody constructs in Lactobacillus vectors

Characterization of primary CXCR4-using HIV-1 strains

MitarbeiterSarah KaluscheFelix LehmannBianca PetriSebastian ReneltOliver RingelPatricia Schult-Dietrich

GruppenleiterinUrsula DietrichTel.: +49 69 63395-216Fax: +49 69 [email protected]

Wissenschaftliche GästeBenjamin Koch (Medizinische Klinik III, Nephrologie, Goethe Universität)

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Im Mittelpunkt unserer Arbeit steht die Identifizierung und Charakterisierung HIV-neutralisierender Antikörper sowie ihrer Epitope im viralen Hüllprotein (Env). Hierbei gelang es uns, Anti-körperfragmente („Nanobodies“) zu identifizieren, die ein breites Spektrum primärer HIV-1 Isolate verschiedener Subtypen neutrali-sieren können. Wegen ihrer breit neutralisierenden Eigenschaften, aber auch aufgrund ihrer geringen Größe und ihrer günstigen physikochemischen Eigenschaften, eignen sich diese nanobodies besonders gut für klinische Anwendungen. Im Hinblick auf einen präventiven Einsatz in Hochrisikokohorten exprimieren wir die Nanobodies in Lactobazillen, natürlichen Mikroorganismen der Vaginalflora, um die sexuelle Übertragung des Virus direkt bei der Infektion und somit vor der Etablierung der lebenslang persistieren-den Virusreservoire im Körper verhindern zu können.

Weiterhin entwickeln und charakterisieren wir verschiedene Env-Immunogene klinisch relevanter HIV-Stämme, um die immunologischen, funktionellen und strukturellen Eigenschaften zu untersuchen. Dazu gehören primäre X4 Viren, die über den CXCR4-Rezeptor infizieren und mit einer schnelleren Krankheits-progression assoziiert sind. Wir konnten zeigen, dass diese Viren sich in ihrem Neutralisationsverhalten von den „klassischen“ CCR5-nutzenden Viren unterscheiden. Eine genaue Analyse der immunologischen und strukturellen Eigenschaften der X4-Viren soll Aufschluss darüber geben, ob ggf. primäre X4-Immunogene in Vakzinestudien eingeschlossen werden müssen.

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lactic and therapeutic applications. In a second group of projects we anal-yse Env characteristics of HIV-1 strains infecting via the CXCR4 coreceptor (X4 viruses). These viruses are generally more aggressive than CCR5-using HIV-1 (R5 viruses) and are associated with a faster progression to AIDS. Furthermore, they expand the spectrum of HIV-1 target cells by including early stem and progenitor cells. We therefore gener-ated a panel of primary X4 viruses to characterize their biological, immuno-logical and structural properties and the implications for vaccine development.

Identification of broadly neutral-izing nanobodies against HIV-1We could select very effective bn nano-bodies from phage immune libraries generated from dromedaries immu-nized with trimeric HIV-1 Env proteins from the most prevalent HIV-1 subtype C (Fig. 1). Two nanobodies showing a complementary neutralization pattern using a standard pseudovirus panel could neutralize 19 of 21 HIV-1 strains of dif-ferent subtypes. Structural analysis of the nanobodies in complex with trimeric Env constructs is ongoing by our collaboration partners at SCRIPPS to map the epitopes at the CD4 binding site in more detail. We are currently expressing the most effective nanobodies in Lactobacillus vectors derived from strains populating the human vagina (collaboration with H. Marcotte, Karolinska Institute, Stockholm) with the aim to derive effective and safe vaccines for HIV prevention in high risk cohorts. The respective recombinant Lactobacillus strains will be analysed in a humanized mouse model for the prevention of HIV-1 infection (collabora-tion with F. Klein, Institute of Virology and Molecular Medicine, Cologne).

Characterization of CXCR4 using (X4) Env immunogens and virusesX4 viruses have generally been described so far as late population arising in persons initially infected by CCR5 using viruses (R5) (Fig. 2A). However, in recent years multiple publications reported an increase in X4 viruses in several countries and their appearance at a much earlier timepoint in infection. To study these new primary X4 viruses we generated the first panel of primary X4 viruses (Fig. 2B) comprising all major HIV-1 subtypes in collaboration with C. Kücherer/N. Bannert (Robert Koch Institute, Berlin) and R. Dürr (NYU, New York) and compared it to lab-adapted strains as well as to a standard panel of R5 viruses. Neutralization assays not only revealed differences in sensitivity to a set of bnAbs between primary and lab-adapted X4 viruses, but interestingly also less sensitivity to CD4 binding site anti-bodies of primary X4 viruses compared to R5 viruses. This argues for a more closed and compact structure of primary X4 Envs, which may escape neutralization and facilitate the increased emergence of X4 viruses in primary infections. To verify this hypothesis we generated two replication

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Figure 1.Broadly neutralizing nanobodies against HIV-1. Schematic comparison of IgG, heavy chain only antibodies from camelids and nanobodies. Nanobodies VHH28 and A6 show a complementary neutralization pattern of HIV-1 strains (boxed in blue) neutralizing 19 of 21 strains of a standard panel comprising different clades. Right: VHH28 binds to the CD4 binding site in the Env trimer (orange), as shown in the high resolution model of the PGV04 Fab (blue/green)/BG505 SOSIP complex docked into 3D reconstructions of ZM197M SOSIP/VHH28 (A. Ward, I. Wilson, SCRIPPS). Nanobodies were expressed in Lactobacillus vectors habitating the human vagina in a soluble (1) and surface-attached (2) form (H. Marcotte, Karolisnka Institute) to achieve high-level expression to prevent sexual transmission of HIV-1. Lactobacillus vectors expressing the nanobodies will be analyzed in a humanized mouse model of HIV-1 infection, which is suited for vaginal transmission studies.

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competent X4 viruses, which are currently being analysed by cryo-electron micros-copy by our collaboration partners at NIH.

We further analyze the infectability of primary human haematopoietic stem and progenitor cells (HSPC) derived from cord blood and bone marrow samples by X4 HIV-1 strains. After showing the presence of both, CD4 and CXCR4 receptors on HSPC of either origin, we proved the in vitro infectability of these cells, whereby infectability correlated with the degree of maturation. We currently establish pro-tocols based on in situ hybridization and immunohistochemistry to detect proviral genomes in HSPC derived from bone marrow samples of HIV-positive patients.

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Koch K, Kalusche S, Torres JL, Stanfield RL, Dan-quah W, Khazanehdari K, von Briesen H, Geertsma ER, Wilson IA, Wernery U, Koch-Nolte F, Ward AB, Dietrich U. Selection of nanobodies with broad neutralizing potential against primary HIV-1 strains using soluble subtype C gp140 envelope trimers. Scientific Reports 2017, 7:8390 doi: 10.1038/s41598-017-08273-7.

Arnold P*, Himmels P*, Weiß S*, Decker TM, Markl J, Gatterdam V, Tampé R, Bartholomäus P, Dietrich U+, Dürr R+. Antigenic and 3D structural charac-terization of soluble X4 and hybrid X4-R5 HIV-1 Env trimers. Retrovirology 2014, 11(1):42 (*equal contribution, +joint last authors).

Zhou M, Meyer T, Koch S, Koch J, Brill B, von Briesen H, Benito JM, Soriano V, Haberl A, Bickel M, Dübel S, Hust M, Dietrich U. Identification of a new epitope for HIV neutralizing antibodies in the gp41membrane proximal external region by an Env-tailored phage display library. Eur J Immunol 2013, 43: 499 – 509.


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Figure 2.Characterization of CXCR4 using (X4) Env immunogens and viruses. HIV-1 uses the CD4 receptor and either CCR5 (R5), CXCR4 (X4) or both coreceptors (R5/X4). A panel of recombinant primary X4 viruses and Env immu-nogens was generated from patients harbouring X4 and R5 viruses or from very rare patients with a homozygous Δ32bp ccr5 deletion infected by X4 viruses. Haematopoietic stem cells (HSC) and multipotent progenitor cells (MPP) are infectable in vitro by X4 HIV-1 strains, as shown here by green GFP-positive cells.

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Immunotherapy of malignanciesWinfried Wels

Immuntherapie maligner Erkrankungen

Expression of chimeric antigen recep-tors (CARs) in cytotoxic lymphocytes constitutes a promising strategy for adoptive cancer immunotherapy with effector cells of defined specificity. CARs consist of a tumor-specific single chain Fv (scFv) antibody fragment connected via a flexible spacer and a transmembrane domain to intracellular signaling domains such as CD3ζ chain or CD3ζ together with one or more costimulatory protein domains. CAR-engineered T cells targeting CD19 have demonstrated remarkable clinical efficacy in patients with malignancies of B-cell origin. Natural killer (NK) cells represent another valuable effector cell population for adoptive cancer immunotherapy, but experience with CAR-engineered NK cells is still limited. NK cells are part of the innate immune system and play an important role in cancer immunosurveillance, with their cytotoxicity being triggered rapidly upon appropriate stimulation through germline-encoded cell surface receptors. In addition, NK cells modulate T-cell mediated antitumor immune responses by maintaining the quality of dendritic cells and enhancing the

Immunotherapy of malignancies

chimeric antigen receptors

natural killer cells

adoptive cancer immunotherapy

MitarbeiterPranav OberoiCongcong ZhangMalena BoddenAline LindnerJasmin RöderAnja WaldmannJanina HenzeChristina KoppThorsten GeyerBarbara Uherek

GruppenleiterWinfried Wels, stellvertretender Direktor Tel.: +49 69 63395-188Fax: +49 69 [email protected]

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Ziel unserer Arbeiten ist die Erforschung und Entwicklung effektiver Immuntherapien zur Behandlung von Krebserkrankungen. Einen Schwerpunkt bilden dabei natürliche Killerzellen (NK-Zellen), die Teil des angeborenen Immunsystems sind und eine wichtige Rolle bei der Abwehr maligner Zellen spielen. Durch Expression sogenannter chimärer Antigenrezeptoren (CARs) generieren wir mittels lentivira-lem Gentransfer genmodifizierte NK-Zellen, die Tumorzellen selek-tiv abtöten. CARs tragen ein extrazelluläres Antikörperfragment mit Tumorzellspezifität, das über eine flexible Verbindungsregion und eine Transmembrandomäne mit intrazellulären Signaldomänen verbunden ist. Damit lösen die Rezeptoren nach Zielzellerkennung

gerichtete zytotoxische Aktivität der Effektorzellen aus. Daneben modulieren CAR-NK-Zellen indirekt auch die endogene adaptive Anti-Tumor-Immunantwort. Als Zielantigene nutzen wir tumoras-soziierte Oberflächenantigene wie das zelluläre Proto-Onkogen ErbB2 (HER2), den epidermalen Wachstumsfaktor-Rezeptor EGFR, das Gangliosid GD2 und Differenzierungsantigene wie CD19 und CD20. Gegenwärtig entwickeln wir in enger Kooperation mit aka-demischen Partnern am Standort Frankfurt eine ErbB2-spezifische Variante der klinisch einsetzbaren humanen NK-Zelllinie NK-92 für den Einsatz in einer Phase-I-Studie bei Patienten mit rezidiviertem, ErbB2-positivem Glioblastom.

presentation of tumor antigens (Figure 1).Nevertheless, in cancer patients NK cells are often functionally compromised due to the immunosuppressive activity of the tumor. Hence, for adoptive cancer immunotherapy donor-derived allogeneic NK cells are being preferred since they do not recognize tumor cells as 'self', thereby bypassing inhibitory signals.

Tumor-specific natural killer cellsSimilar to donor-derived primary NK cells, the continuously expanding human NK cell line NK-92 has been safely applied in clinical trials as an allogeneic cell therapeutic, with durable responses observed in some of the cancer patients treated. In continuous work over the past 15 years, we demonstrated that the therapeutic utility of NK-92 can be further enhanced by expression of CARs targeting antigens such as CD19 and CD20 present on B-cell malignancies, or ErbB2 (HER2), epidermal growth factor receptor (EGFR), the tumor-specific EGFR mutant EGFRvIII, epithelial cell adhesion molecule (EpCAM), and the disialo-ganglioside GD2 expressed by cancer cells from solid tumors. While earlier

approaches used CARs harboring a single intracellular signaling domain derived from CD3ζ, our current work is based on optimized CAR sequences employing in addition to CD3ζ different costimulatory

protein domains for signaling. In a recent example, we engineered NK-92 cells by lentiviral gene transfer to express CARs that target CD19 and contain CD3ζ, or composite CD28-CD3ζ or CD137-CD3ζ

Figure 1.Modulation of adaptive antitumor immunity by natural killer cells. Activation of NK cells by target tumor cells induces production of IFN-γ and TNF-α that promote maturation of dendritic cells (DCs). Mature DCs (mDC) in turn produce IL-12, IL-15 and IL-18 which enhance cytotoxicity and IFN-γ secretion of NK cells. NK cells also eliminate immature DCs (iDC) thereby maintaining the quality of the mDC population (DC editing). NK-induced tumor cell lysis provides antigens which can be taken up by DCs for antigen presenta-tion. Once maturated, antigen-loaded mDCs migrate into tumor-draining lymph nodes and cross-present tumor antigens to T cells.

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signaling domains (Figure 2). Exposure of CD19-positive targets to these CAR NK-92 cells resulted in secretion of IFN-γ, NK-cell degranulation and selective cytotoxicity towards established B-cell lymphoma and leukemia cells as well as primary pre-B-ALL blasts. While all three CAR NK-92 cell variants were functionally active, cells harboring the CD137-CD3ζ receptor were less effective in cell killing and cytokine production than cells with the CD28-CD3ζ CAR, pointing towards differential effects of these costimula-tory protein domains in NK-92 cells.

Genetic modification of primary lymphocytesIn addition to our work with clinically usable NK-92 cells, we are developing improved CAR vectors and optimized transduction protocols suitable for primary NK cells that are either expanded from peripheral blood or differentiated ex vivo from mobilized hematopoietic stem or progenitor cells. In a collaborative effort together with the Department of Stem Cell Transplantation and Immunol-ogy, Clinic for Pediatric and Adolescent Medicine at the University of Frankfurt,

we also investigated tumor-specific cytokine-induced killer (CIK) cells target-ing CD19 or ErbB2 as potential immuno-therapies for acute leukemia and sarcoma. CIK cells consist mainly of T cells and NK-T cells, and are expanded ex vivo from peripheral blood mononuclear cells. In preclinical models we could demonstrate potent and selective cytotoxicity of CD19-specific CAR CIK cells against cancer cell lines and primary pre-B-ALL blasts, which was retained in vivo, resulting in cures in a murine leukemia xenograft model in the majority of animals.

CAR-NK cells for clinical applicationsIn close collaboration with the German Red Cross Blood Donation Service in Frankfurt we generated a clonal ErbB2-specific CAR NK-92 cell line that may become useful as an off-the-shelf cell therapeutic for cancer immunotherapy. These NK-92/5.28.z cells selectively recognized ErbB2-expressing cells of different tumor origins and displayed high and selective antitumor activity in in vitro and in vivo models. Ongoing work focuses on the development of these cells for adoptive immunotherapy of ErbB2-

positive glioblastoma. Together with colleagues from the Institute of Neuroon-cology at the University of Frankfurt, we demonstrated potent activity of NK-92/5.28.z against established and primary glioblastoma cells in vitro and orthotopic glioblastoma xenografts in immuno-deficient mice. In immunocompetent animals, local therapy with NK-92/5.28.z cells resulted in cures of transplanted syngeneic glioblastoma tumors, induction of endogenous antitumor immunity and long-term protection against tumor rechallenge at distant sites. Based on these promising data, the protocol for a monocentric phase I clinical trial was developed to evaluate NK-92/5.28.z cells as a treatment for patients with recurrent ErbB2-positive glioblastoma (phase I study CAR2BRAIN; sponsor: Goethe University, Frankfurt). Very recently, authorization to conduct this trial has been obtained from the respective regula-tory authorities, and patient recruitment will commence in the near future.


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Zhang C, Burger MC, Jennewein L, Genßler S, Schönfeld K, Zeiner P, Hattingen E, Harter PN, Mittelbronn M, Tonn T, Steinbach JP, Wels WS. ErbB2/HER2-specific NK cells for targeted the-rapy of glioblastoma. J Natl Cancer Inst. 2016 May;108:djv375.

Genßler S, Burger MC, Zhang C, Oelsner S, Mil-denberger I, Wagner M, Steinbach JP, Wels WS. Dual targeting of glioblastoma with chimeric antigen receptor-engineered natural killer cells overcomes he-terogeneity of target antigen expression and enhances antitumor activity and survival. OncoImmunology. 2016 Apr;5(4):e1119354.

Zhang C, Oberoi P, Oelsner S, Waldmann A, Lind-ner A, Tonn T, Wels WS. Chimeric antigen receptor-engineered NK-92 cells: An off-the-shelf cellular therapeutic for targeted elimination of cancer cells and induction of protective anti-tumor immunity. Front Immunol. 2017 May 18;8:533.


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Figure 2. Schematic representation of chimeric antigen receptors for expression in NK-92 cells that consist of an extracellular scFv antibody domain for target recognition (scFv) fused via a hinge region derived from CD8α (hinge) to transmembrane and intracellular domains of CD3ζ (left), transmembrane and intracel-lular domains of CD28 and the intracellular domain of CD3ζ (middle), or transmembrane and intracellular domains of CD137 (4-1BB) and the intracellular domain of CD3ζ (right). For analysis of CAR expression, lysates of NK-92 cells harboring CD19-specific CARs containing CD3ζ, composite CD28-CD3ζ or CD137-CD3ζ signaling domains as represented in were subjected to SDS-PAGE under reducing conditions and subsequent immunoblotting with CD8α-specific antibody which detects the hinge domain. As a measure for activation, secretion of IFN-γ by NK-92 cells carrying CD19-specific CARs with CD3ζ, CD28-CD3ζ or CD137-CD3ζ domains was measured after co-incubation with CD19-positive Raji lymphoma cells for 6 hours. Parental NK-92 cells without CAR and NK cells kept in the absence of target cells were included as controls.

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Publikationen

AG MedyoufAG Krause

Medyouf H and De Fontenay M. Book Chapter: Pathophysiologie des syndromes myélodysplasiques: biologie cellulaire. Syndromes myélodysplasiques. John Libbey, 4th Edition. 2016 (French).

Medyouf H. Mouse models of MDS. The European MDS Studies Coordination Organisation (EMSCO) Newsletter. March 2015.

Ekaterina B, Rauner M, Medyouf H, Theurl I, Bornhauser M, Hofbauer L, Platzbecker U. Myelodysplasia is in the niche - novel con-cepts and emerging therapies. Leukemia. Review. 2015. Feb;29(2):259-68.

Tirado-Gonzalez I, Czlonka E, Nevmer-zhitskaya A, Soetopo D, Bergonzani E, Mahmoud A, Contreras A, Jeremias I, Platzbecker U, Bourquin JP, Kloz U, Van der Hoeven F, and Medyouf H (2017). CRISPR/Cas9 edited NSG mice as PDX models of human leukemia to address the role of niche-derived SPARC. Leukemia. In press

Schanda J, Lee CW, Wohlan K, Müller-Kuller U, Kunkel H, Coco IQ, Stein S, Metz A, Koch J, Lausen J, Platzbecker U, Medyouf H, Gohlke H, Heuser M, Eder M, Grez M, Scherr M, Wichmann C. Suppression of RUNX1/ETO oncogenic activity by a small molecule inhibitor of tetramerization. Haematologica. 2017 May;102(5):e170-e174. doi: 10.3324/haematol.2016.161570.

Medyouf H. The microenvironment in human myeloid malignancies: emerging concepts and therapeutic implications. Blood. 2017. Mar 23;129(12):1617-1626. doi: 10.1182/blood-2016-11-696070.

Mossner M, Jann JC, Wittig J, Nolte F, Fey S, Nowak V, Obländer J, Pressler J, Palme I, Xanthopoulos C, Boch T, Metzgeroth G, Röhl H, Witt SH, Dukal H, Klein C, Schmitt S, Gelß P, Platzbecker U, Balaian E, Fabarius A, Blum H, Schulze TJ, Meggendorfer M, Haferlach C, Trumpp A, Hofmann WK, Medyouf H*, Nowak D*. Mutational hierarchies in myelodysplastic syndromes dynamically adapt and evolve upon therapy response and failure.Blood. 2016 Sep 1;128(9):1246-59.

Botezatu L, Michel LC, Helness A, Vad-nais C, Makishima H, Hönes JM, Robert F, Vassen L, Thivakaran A, Al-Matary Y, Lams RF, Schütte J, Giebel B, Görgens A, Heuser M, Medyouf H, Maciejewski J, Dührsen U, Möröy T, Khandanpour C.Epigenetic therapy as a novel approach for GFI136N-associated murine/human AML. Exp Hematol. 2016 Aug;44(8):713-726.e14.

Kohrs N, Kolodziej S, Kuvardina ON, Herglotz J, Yillah J, Herkt S, Pie-chatzek A, Salinas Riester G, Lingner T, Wichmann C, Bonig H, Seifried E, Platzbecker U, Medyouf H, Grez M, Lausen J.MiR144/451 Expression Is Repressed by RUNX1 During Megakaryopoiesis and Disturbed by RUNX1/ETO. PLoS Genet. 2016 Mar 18;12(3):e1005946.

Publications 2015 – 2017

Kumar R, Godavarthy PS, Krause DS.The bone marrow microenvironment at a glanceJ Cell Sci; in press.

Karantanou C, Krause DS.Targeting the bone marrow microenvironment in acute leukaemiaLeuk Lymphoma; in press.

Verma D, Krause DS.Targeting the bone marrow niche in haematological malignanciesAdvances in Stem Cells and Their Niches, In Dominique Bonnet, editor (2017): Hematopoietic Stem Cell Niche, Vol 1, ASN, UK: Academic Cell, 155-175

Weissenberger ES, Krause DS.Histological and In Vivo Microscopic Analysis of the Bone Marrow Microenvironment in a Murine Model of Chronic Myelogenous LeukemiaMethods Mol Biol. 2016;1465:59-72.

Krause DS, Scadden DT.A hostel for the hostile: The stem cell niche in haematological neoplasmsHaematologica 2015;100(11):1376-87.

Krause DS.Illness and artistic creativity (on the 70th anniversary of the death of Béla Bartók, composer, ethnomusicologist and leukemia patient).Leukemia 2015;29(12):2417-8.

Naka K, Jin CH, Ishihara K, Jomen Y, Kim D-H, Gu Y-K, Jeong E-S, Li S, Krause DS, Bae E, Ooshima A, Sheen YY, Kim S-J, Kim D-K.The novel oral TGF-β signaling inhibitor EW-7197 targets chronic myeloid leukemia stem cellsCancer Sci. 2015 Nov 19. doi: 10.1111/cas.12849.

Rabenhorst U, Thalheimer F, Gerlach K, Kijonka M, Böhm S, Krause DS, Vauti F, Arnold HH, Schroeder T, Schnütgen F, von Melchner H, Rieger M, Zörnig M.Single-stranded DNA-binding transcriptional regulator FUBP1 is essential for fetal and adult hematopoietic stem cell self-renewalCell Rep. 2015 Jun 30;11(12):1847-55.

Masia R, Krause DS, Yellen G.The inward rectifier potassium channel Kir2.1 is expressed in mouse neutrophils from bone marrow and liverAm J Physiol Cell Physiol. 2015 Feb 1;308(3):C264-76.

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Khageh Hosseini S, Kolterer S, Steiner M, von Manstein V, Gerlach K, Trojan J, Waidmann O, Zeuzem S, Schulze JO, Hahn S, Steinhilber D, Gatterdam V, Tampé R, Biondi RM, Proschak E, Zörnig M.Camptothecin and its analog SN-38, the active metabolite of irinotecan, inhibit binding of the transcriptional regulator and oncoprotein FUBP1 to its DNA target sequence FUSE.Biochem Pharmacol. 2017 Oct 13. pii: S0006-2952(17)30633-0. doi: 10.1016/j.bcp.2017.10.003. [Epub ahead of print]

Wesely J, Steiner M, Schnütgen F, Kaulich M, Rieger MA, Zörnig M. Delayed mesoderm and erythroid differentiation of murine embryonic stem cells in the absence of the transcriptional regulator FUBP1. Stem Cells Int. 2017;2017:5762301.doi: 10.1155/2017/5762301. Epub 2017 May 15.

Hauck S, Hiesinger K, Khageh Hosseini S, Achenbach J, Biondi RM, Proschak E, Zörnig M1, Odadzic D1 (2016).Pyrazolo[1,5a]pyrimidines as a new class of FUSE binding protein 1 (FUBP1) inhibitors. Bioorg Med Chem. 2016 Nov 15; 24(22):5717-29.

Tan D, Tao S, Chen Z, Koliesnik IO, Calmes PG, Hoerr V, Han B, Gebert N, Zörnig M, Löffler B, Morita Y, Rudolph KL.Dietary restriction improves repopulation but impairs lymphoid differentiation capacity of hematopoietic stem cells in early aging.J Exp Med. 2016 Apr 4; 213(4):535-53.

Rabenhorst U, Thalheimer FB, Gerlach K, Kijonka M, Böhm S, Krause DS, Vauti F, Arnold HH, Schroeder T, Schnütgen F, von Melchner H, Rieger MA, Zörnig M.Single-stranded DNA-binding transcriptional regulator FUBP1 is essential for fetal and adult hematopoietic stem cell self-renewal. Cell Rep. 2015 Jun 30; 11(12):1847-55.

Harter PN, Jennewein L, Baumgarten P, Ilina E, Burger MC, Thiepold AL, Tichy J, Zörnig M, Senft C, Steinbach JP, Mittelbronn M, Ronellenfitsch MW.Immunohistochemical assessment of phosphorylated mTORC1-pathway proteins in human brain tumors. PLoS One. 2015 May 19; 10(5):e0127123.

Ordog T, Zörnig, M., Hayashi Y.Targeting disease persistence in gastrointestinal stromal tumors. Stem Cells Transl Med. 2015 Jul; 4(7):701-7.

Schwamb B, Pick R, Mateus Fernández SB, Völp K, Heering J, Dötsch V, Bösser S, Jung J, Beinoraviciute-Kellner R, Wesely J, Zörnig I, Hammerschmidt M, Nowak M, Penzel R, Zatloukal K, Joos S, Rieker R, Agaimy A, Söder S, Reid-Lombardo K, Kendrick ML, Bardsley MR, Hayashi Y, Asuzu DT, Syed SA, Ordog T, Zörnig M.FAM96A is a novel pro-apoptotic tumor suppressor in gastrointestinal stromal tumors. Int J Cancer. 2015 Sep; 137(6):1318-29

Kramer D, Schön M, Bayerlová M, Bleckmann A, Schön MP, Zörnig M, Dobbelstein M.A pro-apoptotic function of iASPP by stabilizing p300 and CBP through inhibition of BRMS1 E3 ubiquitin ligase activity. Cell Death Dis. 2015 Feb 12; 6:e1634.

1 both senior authors contributed equally to the work

Akademische Ausbildung Josephine Wesely: “The role of FUBP1 during differentiation of murine ESCs”. Dissertation am Fachbereich 15: Biowissenschaften der Goethe-Universität Frankfurt am Main, 2017.

Stefanie Hauck: “Mechanistic studies on FUSE binding protein 1 (FUBP1) inhibition in cancer cells”. Dissertation am Fachbereich 14: Biochemie, Chemie und Pharmazie der Goethe-Universität Frankfurt am Main, 2017.


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Arkan MC. The intricate connection between diet, microbiota, and cancer: A jigsaw puzzle. Semin. Immunol. 2017. Sep 1. pii: S1044-5323(17)30023-4.

Arkan MC. Cancer: Fat and the fate of pancreatic tumours. Nature 2016. 536 (7615): 157-8

Taniguchi K, Moroishi,T, de Jong PR, Krawczyk M, Grebbin BM (AG Farin), Luo H, Xu R-H, Golob-Schwarzl N, Schweiger C, Wang K, Di Caro G, Feng Y, Fearon ER, Raz E, Kenner L, Farin HF, Guan KL, Haybaeck J, Datz C, Zhang K, and Karin M. (2017).YAP-IL-6ST autoregulatory loop activated on APC loss controls colonic tumorigenesis. PNAS doi: 10.1073/pnas.1620290114

Tetteh PW, Kretzschmar K, Begthel H, Born M, van den Korving J, Morsink F, Farin H, Es JH van, Offerhaus GJA, and Clevers H. (2016). Generation of an inducible colon-specific Cre enzyme mouse line for colon cancer research. PNAS 201614057.

Farin HF*, Jordens I, Mosa MH (AG Farin), Basak O, Korving J, Tauriello DVF, de Punder K, Angers S, Peters PJ, Maurice MM, Clevers H.* (2016). Visualization of a short-range Wnt gradient in the intestinal stem-cell niche. Nature 530, 340–343.* co-corresponding authors

Tetteh PW, Basak O, Farin HF, Wiebrands K, Kretzschmar K, Begthel H, van den Born M, Korving J, de Sauvage F, van Es JH, et al. (2016). Replacement of Lost Lgr5-Positive Stem Cells through Plasticity of Their Enterocyte-Lineage Daughters. Cell Stem Cell 18, 203–213.

Lüdtke TH, Rudat C, Wojahn I, Weiss A-C, Kleppa M-J, Kurz J, Farin HF, Moon A, Christoffels VM, and Kispert A. (2016). Tbx2 and Tbx3 Act Downstream of Shh to Maintain Canonical Wnt Signaling during Branching Morphogenesis of the Murine Lung. Developmental Cell 39, 239–253.

Greulich F, Rudat C, Farin HF, Christoffels VM, and Kispert A. (2016). Lack of Genetic Interaction between Tbx18 and Tbx2/Tbx20 in Mouse Epicardial Development. PLoS ONE 11, e0156787.

Greulich F, Trowe M-O, Leffler A, Stoetzer C, Farin HF, and Kispert A. (2016). Misexpression of Tbx18 in cardiac chambers of fetal mice interferes with chamber-specific developmental programs but does not induce a pacemaker-like gene signature. J. Mol. Cell. Cardiol. 97, 140–149.

Weren RD, Venkatachalam R, Cazier JB, Farin HF, Kets CM, de Voer RM, Vreede L, Verwiel ET, van Asseldonk M, Kamping EJ, Kiemeney LA, Neveling K, Aben KK, Carvajal-Carmona L, Nagtegaal ID, Schackert HK, Clevers H, van de Wetering M, Tomlinson IP, Ligtenberg MJ, Hoogerbrugge N, Geurts van Kessel A, Kuiper RP. Germline deletions in the tumour suppressor gene FOCAD are associated with polyposis and colorectal cancer development. J Pathol. 2015 Jun;236(2):155-64.

Tetteh PW, Farin HF, and Clevers H. (2015). Plasticity within stem cell hierarchies in mammalian epithelia. Trends in Cell Biology 25, 100–108.

Canli Ö, Nicolas AM, Gupta J, Finkelmeier F, Goncharova O, Pesic M, Neumann T, Horst D, Löwer M, Sahin U, Greten FR.Myeloid Cell-Derived Reactive Oxygen Species Induce Epithelial Mutagenesis (2017). Cancer Cell [in press]

Varga J, Greten FR.Cell plasticity in epithelial homeostasis and tumorigenesis. Nat Cell Biol. 2017 Oct;19(10):1133-1141.

Srivatsa S, Paul MC, Cardone C, Holcmann M, Amberg N, Pathria P, Diamanti MA, Linder M, Timelthaler G, Dienes HP, Kenner L, Wrba F, Prager GW, Rose-John S, Eferl R, Liguori G, Botti G, Martinelli E, Greten FR, Ciardiello F, Sibilia M.EGFR in Tumor-Associated Myeloid Cells Promotes Development of Colorectal Cancer in Mice and Associates With Outcomes of Patients. Gastroenterology. 2017 Jul;153(1):178-190.

Bergmann H, Roth S, Pechloff K, Kiss EA, Kuhn S, Heikenwälder M, Diefenbach A, Greten FR, Ruland J.Card9-dependent IL-1β regulates IL-22 production from group 3 innate lymphoid cells and promotes colitis-associated cancer. Eur J Immunol. 2017 Aug;47(8):1342-1353

Cammareri P, Vincent DF, Hodder MC, Ridgway RA, Murgia C, Nobis M, Campbell AD, Varga J, Huels DJ, Subramani C, Prescott KLH, Nixon C, Hedley A, Barry ST, Greten FR, Inman GJ, Sansom OJ.TGFβ pathway limits dedifferentiation following WNT and MAPK pathway activation to suppress intestinal tumourigenesis. Cell Death Differ. 2017 Oct;24(10):1681-1693.

Van Wijk SJL, Fricke F, Herhaus L, Gupta J, Hötte K, Pampaloni F, Grumati P, Kaulich M, Sou YS, Komatsu M, Greten FR, Fulda S, Heilemann M, Dikic I.Linear ubiquitination of cytosolic Salmonella Typhimurium activates NF-κB and restricts bacterial proliferation. Nat Microbiol. 2017 May 8;2:17066.

Greten FR.Cancer: Tumour stem-cell surprises. Nature 2017 Mar 29; 543(7647):626-627.

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Koliaraki V, Pallangyo CK, Greten FR, Kollias G.Mesenchymal Cells in Colon Cancer. Gastroenterology 2017 Apr;152(5):964-979.

Diamanti MA, Gupta J, Bennecke M, De Oliveira T, Ramakrishnan M, Braczynski AK, Richter B, Beli P, Hu Y, Saleh M, Mittelbronn M, Dikic I, Greten FR.IKKα controls ATG16L1 degradation to prevent ER stress during inflammation. J Exp Med. 2017 Feb; 214(2):423-437

Greten FR.The Irony of Tumor-Induced Inflammation. Cell Metab. 2016 Sep 13;24(3):368-9.

Pesic M, Greten FR.Inflammation and cancer: tissue regeneration gone awry. Curr Opin Cell Biol. 2016 Aug 10;43:55-61.

Bocklitz TW, Salah FS, Vogler N, Heuke S, Chernavskaia O, Schmidt C, Waldner MJ, Greten FR, Bräuer R, Schmitt M, Stallmach A, Petersen I, Popp J.Pseudo-HE images derived from CARS/TPEF/SHG multimodal imaging in combination with Raman-spectroscopy as a pathological screening tool. BMC Cancer. 2016 Jul 26;16:534.

Vogler N, Bocklitz T, Subhi Salah F, Schmidt C, Bräuer R, Cui T, Mireskandari M, Greten FR, Schmitt M, Stallmach A, Petersen I, Popp J.Systematic evaluation of the biological variance within the Raman based colorectal tissue diagnostics. J Biophotonics. 2016 May;9(5):533-41

Varga J, Greten FR.Lifting the Mist on Gastric Stem Cells. Cell Stem Cell. 2016 Jan 7;18(1):7-9.

Van der Heijden M, Zimberlin CD, Nicholson AM, Colak S, Kemp R, Meijer SL, Medema JP, Greten FR, Jansen M, Winton DJ, Vermeulen L. Bcl-2 is a critical mediator of intestinal transformation. Nat Commun. 2016 Mar 9;7:10916.

Göktuna SI, Shostak K, Chau TL, Heukamp LC, Hennuy B, Duong HQ, Ladang A, Close P, Klevernic I, Olivier F, Florin A, Ehx G, Baron F, Vandereyken M, Rahmouni S, Vereecke L, van Loo G, Büttner R, Greten FR, Chariot A.The Prosurvival IKK-Related Kinase IKKε Integrates LPS and IL17A Signaling Cascades to Promote Wnt-Dependent Tumor Development in the Intestine. Cancer Res. 2016 May 1;76(9):2587-99.

Finkelmeier F, Canli Ö, Tal A, Pleli T, Trojan J, Schmidt M, Kronenberger B, Zeuzem S, Piiper A, Greten FR, Waidmann O. High levels of the soluble programmed death-ligand (sPD-L1) identify hepatocellular carcinoma patients with a poor prognosis. Eur J Cancer. 2016 Mar 31;59:152-159.

Canli Ö, Alankuş YB, Grootjans S, Vegi N, Hültner L, Hoppe PS, Schroeder T, Vandenabeele P, Bornkamm GW, Greten FR (2015).Glutathione peroxidase 4 prevents necroptosis in mouse erythroid precursors. Blood. 2016 Jan 7;127(1):139-48.

Davis H, Irshad S, Bansal M, Rafferty H, Boitsova T, Bardella C, Jaeger E, Lewis A, Freeman-Mills L, Giner FC, Rodenas-Cuadrado P, Mallappa S, Clark S, Thomas H, Jeffery R, Poulsom R, Rodriguez-Justo M, Novelli M, Chetty R, Silver A, Sansom OJ, Greten FR, Wang LM, East JE, Tomlinson I, Leedham S. (2015)Abberant epithelial GREM1 expression initiates colorectal tumour formation from cells outside the crypt based stem cell niche. Nature Medicine, 21:62-70.

Drube S, Weber F, Loschinski R, Beyer M, Rothe M, Rabenhorst A, Göpfert C, Meininger I, Diamanti MA, Stegner D, Häfner N, Böttcher M, Reinecke K, Herdegen T, Greten FR, Nieswandt B, Hartmann K, Krämer OH, Kamradt T. (2015)Subthreshold IKK activation modulates the effector functions of primary mast cells and allows specific targeting of transformed mast cells. Oncotarget, 6(7):5354-68

Drube S, Weber F, Göpfert C, Loschinski R, Rothe M, Boelke F, Diamanti MA, Löhn T, Ruth J, Schütz D, Häfner N, Greten FR, Stumm R, Hartmann K, Krämer OH, Dudeck A, Kamradt T. (2015)TAK1 and IKK2, novel mediators of SCF-induced signaling and potential targets for c-Kit-driven diseases. Oncotarget 6(30):28833-50

Pallangyo CK, Ziegler PK, Greten FR.IKKß acts as a tumor suppressor in cancer-associated fibroblasts during intestinal tumorigenesis. J Exp Med. 2015 Dec 14;212(13):2253-66.

Akademische Ausbildung Tiago De Oliveira: Divergent Functions of IL-4Ra and STAT6 in a Model of Colitis-Associated Carcinogenesis". Dissertation am Fachbereich Biologie, Ludwig-Maximilians-Universität München, 2017

Bowman RL, Klemm F, Akkari L, Pyonteck SM, Sevenich L, Quail DF, Dhara S, Simpson K, Gardner EE, Iacobuzio-Donahou C, Brennan CW, Tabar V, Gutin PH, Joyce JA. (2016) Macrophage ontogeny underlines differences in tumor-specific education in brain malignancies. Cell Reports 22;17(9):2445-2459.

Akademische Ausbildung Sebastian Friemel: „CX3CL1-CX3CR1 signaling in neuronal-glial crosstalk in brain metastasis-associated neuro-inflammation”. Masterarbeit im Fachbereich 16 (Molekulare Medizin) der Goethe Universität Frankfurt 2017

Koch K, Kalusche S, Torres JL, Stanfield RL, Danquah W, Khazanehdari K, von Briesen H, Geertsma ER, Wilson IA, Wernery U, Koch-Nolte F, Ward AB, Dietrich U. Selection of nanobodies with broad neutralizing potential against primary HIV-1 strains using soluble subtype C gp140 envelope trimers. Scientific Reports 2017, 7:8390 doi: 10.1038/s41598-017-08273-7

Alam MI, Mostafa A, Kanrai P, Mueller C, Dzieciolowski J, Lenz E, Kuznetsova I, Schult-Dietrich P, Ziebuhr J, Dietrich U, Pleschka S. Verapamil has antiviral activities that target different steps of the influenza virus replication cycle. Antivir & Antiretrovir 2016, 8 (4):121-130.

Schmier S+, Mostafa A+, Haarmann T, Bannert N, Ziebuhr J, Veljkovic V*, Dietrich U*, Pleschka S*. In silico prediction and experimental confirmation of amino acids in the HA conferring enhanced receptor specificity for H5N1 Influenza A viruses. Scientific Reports 2015, 5:11434. doi: 10.1038/srep11434. (+equal contribution; *joint last authors)

Geiß Y, Dietrich U. Catch me if you can – the race between HIV and neutralizing antibodies. AIDS Rev 2015, 17(2): 107-113.

Dietrich U, Landersz M, Stahl-Hennig C, Geiger C, Foley BT.Genetic characterization of near full length SIVdrl genomes from four captive drills (Mandrillus leucophaeus). AIDS Res Hum Retrovir 2015, 31(3): 353-357.

Akademische AusbildungKathrin Koch: „Generation, selection and characterization of single domain antibody fragments (VHHs) derived from HIV-1 Env immunized dromedaries”. Dissertation am Fachbereich 14: Biochemie, Chemie und Pharmazie der Goethe Universität, 2017

Svenja Weiß: “Characterization of emerging HIV-1 X4 viruses with respect to biochemical, immunological and structural properties of their envelopes”. Dissertation am Fachbereich 14: Biochemie, Chemie und Pharmazie der Goethe Universität, 2017

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Maria Trott, geb. Hertje: „Analyse und Selektion von HIV-neutralisierenden scFv Antikörpern aus HIV-Positiven mit nicht progredierendem Krankheitsverlauf“. Dissertation am Fachbereich 15: Biowissenschaften der Goethe Universität, 2017

Felix Lehmann: „Selection and characterization of HIV-1 Env-specific nanobodies from phage immune libraries”. Masterarbeit am Fachbereich 14: Biochemie, Chemie und Pharmazie der Goethe Universität, 2017

Bianca Petri: „Charakterisierung von HIV-1 Hüllproteinen rezent infizierter Patienten“. Masterarbeit im Fachbereich Chemie und Biotechnologie der Hochschule Darmstadt, 2017

Nowakowska P, Romanski A, Miller N, Odendahl M, Bönig H, Zhang C, Seifried E, Wels WS, Tonn T.Clinical grade manufacturing of genetically modified, CAR-expressing NK-92 cells for the treatment of ErbB2-positive malignancies. Cancer Immunol Immunother. (Epub 2017 Sep 06).

Merker M, Pfirrmann V, Oelsner S, Fulda S, Klingebiel T, Wels WS, Bader P, Rettinger E.Generation and characterization of ErbB2-CAR-engineered cytokine-induced killer cells for the treatment of high-risk soft tissue sarcoma in children. Oncotarget. 2017 Sep 12;8(39):66137-53, 2017.

Ahmed N, Brawley V, Hegde M, Bielamowicz K, Kalra M, Landi D, Robertson C, Gray TL, Diouf O, Wakefield A, Ghazi A, Gerken C, Yi Z, Ashoori A, Wu MF, Liu H, Rooney C, Dotti G, Gee A, Su J, Kew Y, Baskin D, Zhang YJ, New P, Grilley B, Stojakovic M, Hicks J, Powell SZ, Brenner MK, Heslop HE, Grossman R, Wels WS, Gottschalk S.HER2 chimeric antigen receptor-modified virus-specific T cells for progressive glioblastoma: a phase I dose-escalation trial. JAMA Oncol. 2017 Aug 1;3(8):1094-101.

Wagner J, Pfannenstiel V, Waldmann A, Bergs JWJ, Brill B, Hünecke S, Klingebiel T, Rödel F, Buchholz CJ, Wels WS, Bader P, Ullrich E. A two-phase expansion protocol combining IL-15 and IL-21 improves NK cell proliferation and cytotoxicity against rhabdomyosarcoma. Front Immunol. 2017 Jun 12;8:676.

Zhang C, Oberoi P, Oelsner S, Waldmann A, Lindner A, Tonn T, Wels WS. Chimeric antigen receptor-engineered NK-92 cells: An off-the-shelf cellular therapeutic for targeted elimination of cancer cells and induction of protective anti-tumor immunity. Front Immunol. 2017 May 18;8:533.

Oelsner S, Friede ME, Zhang C, Wagner J, Badura S, Bader P, Ullrich E, Ottmann OG, Klingemann H, Tonn T, Wels WS. Continuously expanding CAR NK-92 cells display selective cytotoxicity against B-cell leukemia and lymphoma. Cytotherapy. 2017 Feb;19(2):235-49.

Oelsner S, Wagner J, Friede ME, Pfirrmann V, Genßler S, Rettinger E, Buchholz CJ, Pfeiffer H, Schubert R, Ottmann OG, Ullrich E, Bader P, Wels WS. Chimeric antigen receptor-engineered cytokine-induced killer cells overcome treatment resistance of pre-B-cell acute lymphoblastic leukemia and enhance survival. Int J Cancer. 2016 Oct 15;139(8):1799-809.

Hegde M, Mukherjee M, Grada Z, Pignata A, Landi D, Navai SA, Wakefield A, Fousek K, Bielawowicz K, Chow KK, Brawley VS, Byrd TT, Krebs S, Gottschalk S, Wels WS, Baker ML, Dotti G, Mamonkin M, Brenner MK, Orange JS, Ahmed N. Tandem CAR T cells targeting HER2 and IL13Rα2 mitigate tumor antigen escape. J Clin Invest. 2016 Aug 1;126(8):3036-52.

Romanski A, Uherek C, Bug G, Seifried E, Klingemann H, Wels WS, Ottmann OG, Tonn T. CD19-CAR engineered NK-92 cells are sufficient to overcome NK cell resistance in B-cell malignancies. J Cell Mol Med. 2016 Jul;20(7):1287-94.

Alkins RD, Burgess A, Kerbel R, Wels WS, Hynynen K.Early Treatment of HER2-amplified brain tumors with targeted NK-92 cells and focused ultrasound improves survival. Neuro Oncol. 2016 Jul;18(7):974-81.

Zhang C, Burger MC, Jennewein L, Genßler S, Schönfeld K, Zeiner P, Hattingen E, Harter PN, Mittelbronn M, Tonn T, Steinbach JP, Wels WS.ErbB2/HER2-specific NK cells for targeted therapy of glioblastoma. J Natl Cancer Inst. 2016 May;108:djv375.

Genßler S, Burger MC, Zhang C, Oelsner S, Mildenberger I, Wagner M, Steinbach JP, Wels WS. Dual targeting of glioblastoma with chimeric antigen receptor-engineered natural killer cells overcomes heterogeneity of target antigen expression and enhances antitumor activity and survival. OncoImmunology. 2016 Apr;5(4):e1119354.

Suck G, Odendahl M, Nowakowska P, Seidl C, Wels WS, Klingemann HG, Tonn T. NK-92: an 'off-the-shelf therapeutic' for adoptive natural killer cell-based cancer immunotherapy. Cancer Immunol Immunother. 2016 Apr;65(4):485-92.

Zhou Q, Uhlig KM, Muth A, Kimpel J, Lévy C, Münch RC, Seifried J, Pfeiffer A,Trkola A, Coulibaly C, von Laer D, Wels WS, Hartwig UF, Verhoeyen E, Buchholz CJ.Exclusive transduction of human CD4+ T cells upon systemic delivery of CD4-targeted lentiviral vectors. J Immunol. 2015 Sep 1;195(5): 2493-501.

Pfirrmann V, Oelsner S, Rettinger E, Huenecke S, Boenig H, Merker M, Wels WS, Cinatl J, Schubert R, Klingebiel T, Bader P. Cytomegalovirus-specific cytokine-induced killer cells: concurrent targeting of leukemia and infections. Cytotherapy. 2015 Aug;17(8): 1139-51.

Ahmed N, Brawley VS, Hegde M, Robertson C, Ghazi A, Gerken C, Liu E, Dakhova O, Ashoori A, Corder A, Gray T, Wu MF, Liu H, Hicks J, Rainusso N, Dotti G, Mei Z, Grilley B, Gee A, Rooney CM, Brenner MK, Heslop HE, Wels WS, Wang LL, Anderson P, Gottschalk S. Human epidermal growth factor receptor 2 (HER2)-specific chimeric antigen receptor-modified T cells for the immunotherapy of HER2-positive sarcoma. J Clin Oncol. 2015 May;33(15): 1688-96.

Seidel D, Shibina A, Siebert N, Wels WS, Reynolds CP, Huebener N, Lode HN. Disialoganglioside-specific human natural killer cells are effective against drug-resistant neuroblastoma. Cancer Immunol Immunother. 2015 May;64(5): 621-34.

Schönfeld K, Sahm C, Zhang C, Naundorf S, Brendel C, Odendahl M, Nowakowska P, Bönig H, Köhl U, Kloess S, Köhler S, Holtgreve-Grez H, Jauch A, Schmidt M, Schubert R, Kühlcke K, Seifried E, Klingemann HG, Rieger MA, Tonn T, Grez M, Wels WS. Selective inhibition of tumor growth by clonal NK cells expressing an ErbB2/HER2-specific chimeric antigen receptor. Mol Ther. 2015 Feb;23(2): 330-8.

Glienke W, Esser R, Priesner C, Suerth JD, Schambach A, Wels WS, Grez M, Kloess S, Arseniev L, Koehl U. Advantages and applications of CAR-expressing natural killer cells. Front Pharmacol. 2015 Feb 12;6:21.

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Financial affairs

Finanzen und Administration

Die Abteilung Finanzen/Administration setzt jährlich ein durchschnittliches Finanzvolumen von 9 Millionen Euro um und betreut dabei rund 100 Mitarbeiterin-nen und Mitarbeiter. Sie wird geführt von Robert Dornberger, der dabei von Christof Kaiser unterstützt wird.

Christiane Strack im Personalbüro erledigt sämtliche Personalfragen. Verstärkt wird sie dabei von Ilka Grau, die auch in der Finanzbuchhaltung/Drittmittelverwaltung insbesondere die Projektfördermittel des Bundes betreut. Gabriele Heckl erstellt die Bilanz, bearbeitet alle Ausgangsrech-nungen, die Reisekostenabrechnungen und ist für die Abrechnung internationaler Drittmittel zuständig. Giuseppina Virgillito betreut die Kreditorenrechnungen sowie die Drittmittel nationaler Stiftungen.

Ansprechpartner in der Telefonzentrale und am Empfang ist Bernd Würdemann. Adrian Gresik ist verantwortlich für die vielfältigen Aufgaben des Innendienstes. Er, Michael Paul und Heinrich Krompietz kümmern sich um die haustechnischen Ausstattungen und Installationen und arrangieren alle Arten von wissenschaft-lichen Tagungen und Veranstaltungen. Dabei werden sie von Volker Hopf unter-stützt.

Yoseph Alazar, Maria Fernandes, Yasemin Piskin und Neriman Sarac reinigen die Laboratorien, entsorgen die anfallenden Abfälle und kümmern sich um die Bereitstellung von Laborbedarf. Sie werden dabei von Keziban Ata unterstützt.

Our administration and services de-partment is led by Robert Dornberger who oversees a yearly budget of ap-proximately 9 million Euros. He also takes care of the administrative needs of about 100 staff members and is supported by Christof Kaiser.

He is assisted by Christiane Strack who heads the personnel department with the support of Ilka Grau. Ilka Grau is also engaged in financial accounting and the administration of external fundings. Gabriele Heckl prepares the balance sheets, is responsible for all outgoing invoices, all claims of travel expenses and is in charge of accounting of interna-tional grants. Giuseppina Virgillito handles grants from nationwide foundations and took care of all incoming invoices.

Bernd Würdemann is our receptionist and is the first contact with the Institute.Adrian Gresik is responsible for the internal service tasks. Together with Michael Paul and Heinrich Krompietz they take care of the technical equip-ment and the installations and arrange all kinds of scientific meetings and events. They are supported by Volker Hopf.

Yoseph Alazar, Maria Fernandes, Yasemin Piskin and Neriman Sarac clean the laboratories, dispose of the waste and restock the supplies. They are supported by Keziban Ata.

Robert DornbergerLeiter der Abteilung Finanzen/Administration Tel.: +49 69 63395-333Fax: +49 69 63395-353 [email protected]

Christoph Kaiser Stellv. Leiter der Abteilung Finanzen/Administration

Tel.: +49 69 63395-106Fax: +49 69 63395-353 [email protected]

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Scientific Services

Zentrale Einheit HistologieZur Anfertigung von histologischen Präparaten betreibt das Georg-Speyer-Haus eine Histologie-Serviceeinheit unter der Leitung von Dr. Boris Brill. Hier werden von Frau Petra Dinse, meist automatisiert, die Gewebeaufarbeitung sowie immuno-histochemische Färbungen und Stan-dardfärbungen durchgeführt. Weiterhin verfügt das Labor über ein automatisiertes Präparate-Scanner- und Bildanalyse-system, Aperio ScanScope CS2, einen Färbeautomat Leica Autostainer XL sowie einen Leica BOND max zur Anfertigung von automatisierten Immunfärbungen.

Das Labor stellt seine Leistungen den Arbeitsgruppen des Georg-Speyer-Hauses sowie externen Forschergruppen zur Verfügung.

TierhaltungDas Georg-Speyer-Haus betreibt eine Tierhaltung, geleitet von Dr. Boris Brill, um den Forschungsgruppen die Zucht von Mäusen und Experimente zu ermöglichen. Die Tierhaltung stellt in vivo Imaging Systeme, wie Endoskopie (Storz Coloview Set), in vivo konfokale Mikroskopie (Cellvi-zio), ein Bruker 7T MRT und ein Detekti-onssystem für Fluoreszenz und Luciferase in vivo (IVIS Lumina II) zur Verfügung.

Zentrale Einheit DurchflusszytometrieDie zentrale Zytometrie-Einrichtung besteht aus drei Durchflusszytometern zur Zellanalyse (BD LSRFortessa, FACSCantoII, FACSCalibur) und zwei Zellsortern (BD FACSAriaI, FACSAria Fusion). Geleitet wird die Serviceeinheit von Dr. Stefan Stein, der auch Ansprechpartner für allgemeine Fragen zur Durchflusszytometrie und bei der Entwicklung und Anpassung neuer Mess- und Sortieransätze ist. Tefik Merovci führt die anfallenden Hochgeschwindigkeits-Zellsortierungen durch und ist für den einwandfreien Zustand aller FACS-Geräte am Institut verantwortlich. In einigen Fällen fungiert Thorsten Geyer als zusätzlicher Operator an den Zellsortern. Die Sortiereinheit steht primär den Arbeitsgruppen des

Core Facility HistologyThe Georg-Speyer-Haus operates a histology core facility. It is supervised by Dr. Boris Brill. Petra Dinse is responsible for the mostly automated procedures of tissue processing and immunohisto-chemistry as well as hematoxylin / eosin staining. The laboratory is equipped with a slide scanner and image analysis system, Aperio ScanScope CS2, a Leica AutostainerXL and a Leica BOND max for automated immunostaining.

The services of the histology core facility are available to all scientists of the Georg-Speyer-Haus as well as to external research partners in collaboration.

Animal HusbandryThe Georg-Speyer-Haus has an animalfacility, led by Dr. Boris Brill, which provides capacity for our research groups for mouse breeding and experiments. Included in the animal facility are imaging techniques like endoscopy (Storz Coloview Set), in vivo confocal microscopy (Cell-vizio), a Bruker 7T MRI and a detection system for fluorescence and luciferase in vivo (IVIS Lumina II).

Core Facility Flow CytometryThe Flow Core Unit of the Georg Speyer Haus operates three flow cytometer instruments (BD LSRFortessa, FACS-CantoII, FACSCalibur) and two cell sorters (BD FACSAriaI and BD FACSAria Fusion). Dr. Stefan Stein oversees the perfor-mance of the core facility and is available for scientific questions regarding flow cytometry in general and the establish-ment of new FACS based assays. Tefik Merovci is responsible for high-speed cell sorting as operator in this central service unit for all research groups of the GSH as well as for external researchers. Tefik also takes care of the maintenance and functionality of the flow cytometers in the institute. Occasionally, Thorsten Geyer serves as an additional sorting

Wissenschaftlicher Service

Dr. Boris BrillTel.: +49 69 63395-205Fax: +49 69 [email protected]

Dr. Stefan SteinTel.: +49 69 63395-260Fax: +49 69 [email protected] [email protected]

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Scientific Services

Georg-Speyer-Hauses, aber auch externen Forschergruppen zur Verfügung. Seit 2017 wird in der Einrichtung zusätzlich ein CQ1 Confocal Quantitative Image Cytometer (Yogokawa) zur 3D High-throughput Zellanalyse betrieben.

Zentrale Einheit Transgene Techniken(Transgenic Core Facility, TCF) Die „Transgenic Core Facility“ (TCF) des Georg-Speyer-Hauses startete im Oktober 2016. Ziel dieser Einheit ist es in Zusam-menarbeit mit den Forschungsgruppen des GSH neuartige Mausmodelle zu entwickeln und zur Verfügung zu stellen. Dazu werden neueste Methoden (wie z.B. die CRISPR/Cas Technologie) mit etablierten Techniken (wie z.B. Zellkultur, Mikroinjektionstechniken) kombiniert. Im vergangenen, ersten Jahr wurden erfolgreich die strukturellen, metho-dischen und personellen Grundlagen geschaffen und erste vielversprechende Ergebnisse bei der Etablierung dieser anspruchsvollen Anwendung erzielt.

Geleitet wird die Einrichtung von Dr. Madina Karimova. Sie wird dabei von ihren Mitarbeitern Kübra Erdel und Manuel Klein unterstützt.

operator. A CQ1 Confocal Quantitative Image Cytometer (Yogokawa) for 3D high-throughput cell measurements was installed in 2017.

Transgenic Core Facility, TCF The Transgenic Core Facility at GSH started off in October 2016. The goal of the facil-ity is to generate novel mouse models in collaboration with the research groups at the GSH. To this end, innovative methods, such as CRISPR/Cas9, are combined with established techniques such as embryonic cell culture and microinjections. In its first year, TCF has successfully built up the structural, methodological and personnel basis for its functioning. On its way to establish this challenging application at the GSH first promising results were achieved.

The facility is led by Dr. Madina Karimova who is supported by her team members Kübra Erdel and Manuel Klein.

Ex vivo embryo culture stages, dpc – days post copulation

Pronuclear microinjection into FVB/N 1 cell stage fertilized oocyte

Optimizing pronuclear capillar parameters and injection pressure with fluorescently labelled dex-tran (teramethylrhodamine)

Blastocyst microinjection setup. On the left: early blastocyst with inner cell mass on the top, on the right: mES cells in microinjection capillar

Viable hatching blastocysts post microinjection

Hatching blastocyst 24h post microinjection with eGFP expressing mES cells

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Dr. Madina KarimovaGruppenleiterin Transgenic Core FacilityTel.: +49 69 63395-620Fax: +49 69 [email protected]

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Scientific Services

Dr. Herbert KühnelTel.: +49 69 63395-120Fax: +49 69 [email protected]

Hana KunkelTel.: +49 69 [email protected]

Steffen LuftTel.: +49 69 [email protected]

ITSteffen Luft leitet als Chief Information Officer (CIO) die IT. Er nimmt die zentralen Tätigkeiten der Unterstützung der Mitar-beiterinnen und Mitarbeiter des Hauses in allen Fragen der IT, der Serverbetreuung, des IT-Projekt-managements, der Netzwerkadministration und des Einkaufs wahr.

Arbeitssicherheit und Genehmigungen Dr. Herbert Kühnel beschafft die nötige Laborausstattung / Arbeitsgeräte, holt behördliche Genehmigungen ein und kümmert sich um die Arbeitssicherheit.

Gefahrstoff- und Abfall-management Dr. Klaus Lehmen hat die Verantwortung für den Bereich Gefahrstoff- und Abfall-management sowie den Aufbau eines digitalen Betriebsmittelkatasters.

Hygiene- und Labormanagement Hana Kunkel achtet auf die Einhaltung der geltenden Laborstandards und Arbeitssi-cherheitsbedingungen. Sie verantwortet den Spülküchenbereich und koordiniert die Reinigungsdienstleistungen.

ITSteffen Luft is Chief Information Officer (CIO) of the Institute. His main tasks are the maintenance of the servers, IT project management, administration of the networks and the support of the colleagues in the institute.

Occupational safety and permissionsDr. Herbert Kühnel attends for equipment and supply, permissions, and occupational safety.

Dangerous material and waste management Dr. Klaus Lehmen is in charge with manag-ing of hazardous goods and disposals and implementing a digital register of all technical equipment and documents.

Facility- / Lab ManagementHana Kunkel ensures the compliance with the current laboratory standards and safety regulations. She is responsible for managing all Cleaning services.

Dr. Klaus LehmenTel.: +49 69 63395-118Fax: +49 69 [email protected]

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Veranstaltungen

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Meetings and Lectures 2017

20.12.2017 “Signaling integration by the autophagy adaptor p62 in the tumor microenvi-ronment”Prof. Dr. Jorge Moscat, Sanford Burnham Prebys Medical Discovery Institute, La Jolla, CA

30.11.2016Welt-AIDS-Tag in Zusammenarbeit mit der Goethe Universität und der Arbeitsgruppe von Dr. Ursula Dietrich vom Georg-Speyer-Haus.

23.11.2017“Acute myeloid leukemia associated alteration of the bone marrow microen-vironment”Dr. Diana Passaro, Haematopoietic Stem Cell Laboratory, The Francis Crick Institute, London

09.11.2017“Expanding the CRISPR Toolbox with 3Cs-gRNAs”Dr. Manuel Kaulich, Institut Biochemie II, Universitätsklinikum Frankfurt

25.-26.10.2017 Symposium „Dynamic of adult stem cells and cancer”

12.10.2017“Models and Insights in Translational Research” 2017

05.10.2017 “A Highly Specific Inhibitor Targeting the Effector Binding Site of Ras”Dr. Andreas Ernst, Institut Biochemie II, Universitätsklinikum Frankfurt

28.09.2017“Metabolism, metabolites and endothe-lial plasticity”PD Dr. Michael Potente, Max Planck Institute for Heart and Lung Research, Bad Nauheim

18.09.2017“The onco-metabolic role and liabilities of the TCA cycle in cancer”Prof. Eyal Gottlieb, Technion-Israel Institute of Technology Haifa, Israel

12.09.2017Auswahlsymposium zum Paul-Ehrlich- und-Ludwig-Darmstaedter-Nachwuch-spreis

“Harnessing the power of genome engineering to deconvolute genetic traits of cancer”Darjus Tschaharganeh, German Cancer Research Center (DKFZ),Heidelberg

“Using computational methods to unlock the neural code”Tatjana Tchumatchenko, Max Planck Institute for Brain Research, Frankfurt/M.

31.05.2017“Beyond Targeting Oncogenes: Emerging Anti-Cancer Strategies”Professor Dr. Tak W. Mak, University Health Networks, Princess Margaret Cancer Center, Toronto, Canada

17.05.2017“The role of the transcription factor and epigenetic modulator NFE2 in myelop-roliferative neoplasms”Prof. Dr. Heike L. Pahl, Center for Tumor Biology, University Hospital Freiburg18.04.2017 PerkinElmer: Research solutions for cancer immunology & immunotherapy

06.04.2017“Dual role of chromosome instability in cancer”Prof. Dr. Rocio Sotillo, Molecular Thoracic Oncology, German Cancer Research Center (DKFZ)

03.04.-06.04.2017Schülerpraktikum in den Laborgrup-pen des Georg-Speyer-Hauses

30.03.2017“The Power of Single Cells: BD Genomics Integrated Workflow to Gene Expression Studies”Dr. Wieland Keilholz, BD Life Sciences Genomics, Heidelberg

23.03.2017“Regulation and Heterogeneity of Dor-mant Hematopoietic Stem Cells”Dr. Nina Cabezas-Wallscheid, Division of Stem Cells and Cancer, German Cancer Research Center (DKFZ), Heidelberg

02.03.2017„Novel Targeted Therapies in T-Cell Acute Lymphoblastic Leukemia“Prof. Dr. Jacques Ghysdael, Institut Curie, Paris

01.03.2017ACD Seminar

02.02.2017 “Targeting treatment-resistant leukemia in preclinical mouse models“Prof. Dr. med. Irmela Jeremias, Helm-holtz Zentrum München

31.01.-06.03.2016 Schülervorlesungsreihe 2017

25.01.2017“in vivo dynamics of healthy and mali-gnant haematopoiesis - from population studies to single cell resolution“Dr. Cristina Lo Celso, Imperial College London

“Deciphering the adipocytic lineage: Stem cell biology and development of metabolic disorders”Tim J. Schulz, German Institute of Human Nutrition (DifE), Potsdam-Rehbrücke

“Breaking barriers: TREK1 channel mediated barrier function in brain and muscle”Tobias Ruck, University of Münster Department of Neurology with Insti-tute of Translational Neurology

“How bacteria talk: Lessons from Vibrio cholera”Kai Papenfort, Ludwig Maximilians University, Munich, Department of Biology I, Microbiology

“Synapses. Memories. On the fly.”David Owald, The Charité University Medical Center, Institute of Neuro-physiology

07.09.2017“Tumour Microenvironment Triggers Neuroblastoma Cell Migration and Invasion”Dr. Olga Piskareva, Molecular & Cellu-lar Therapeutics (MCT), Royal College of Surgeons in Ireland RCSI

25.08.2017”CRISPR screening with single-cell transcriptome readout establishes a high-throughput method for dissecting gene-regulatory mechanisms”Prof. Dr. Christoph Bock, CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna

13.07.2017”Malignant alterations of the human bone marrow microenvironment”PD Dr. Manja Wobus, Universitätsklini-kum Carl Gustav Carus, Dresden, Stem Cell Lab

06.07.2017“Mechanisms of immune evasion and metastasis in colorectal cancer”Dr. Eduard Batlle, ICREA Research Professor, Institute for Research in Biomedicine (IRB Barcelona), Spanien

20.–21.06.2017 GSH-Retreat auf der Burg Rothenfels

08.06.2017“G-quadruplex DNA revolution in cancer research: A novel biomarker for tracking cancer gene activity and genome instability”Dr. Robert Hänsel-Hertsch, Cancer Research UK, Cambridge Institute, Li Ka Shing Centre

19.01.2017“Use of a fungal toxin to unravel the intracellular signaling pathway of anoi-kis (detachment-induced apoptosis)“Prof. Dr. Dr. h.c. Christoph Borner, Ins-titute of Molecular Medicine and Cell Research, University of Freiburg

10.01.2017“FUBP1: one more evidence as a player in hematological lineages“Dr. Marie-Bérengère Troadec, Institute of Genetics and Development of Rennes, Université de Rennes

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Regular Research MeetingRecent results, advances and problems of individual research projects are presented and discussed in English

30.01.-06.03.2017Schülervorlesung und –praktikum Lecture series and practical course for high school studentsOrganisation: Dietrich U

„Krebsentstehung: genetische Grundlagen, Diagnostik und neue Therapieansätze“Greten F

„Stammzellen und Krebsstammzellen“Krause D

„Immuntherapie“Sevenich L

„Gentechnologie: Grundlagen und Konsequenzen“Dingermann T

„HIV/AIDS - ein Immundefizienzvirus erobert die Welt“Dietrich U

03.04.–06.04.2017Schülerpraktikum in den Laborgruppen des Georg-Speyer-Hauses

WS 2016 | 17, WS 2017 | 18 SemesterbegleitendSeminarreihe Masterstudiengang Molekulare Medizin: “Frankfurter Forschung“Medyouf H, Sevenich L, Greten F

WS 2016 | 17, SS 2017, WS 2017 | 18Anleitung zum wissenschaftlichen Arbei-ten für Bachelor- und MasterstudentenGreten F, Wels W, Zörnig M, Dietrich U, Medyouf H, Sevenich L, Farin H, Krause D

WS 2016 | 17, SS 2017, WS 2017 | 18Individuelle 6-wöchige Laborpraktika in Zusammenarbeit mit der Goethe-UniversitätGreten F, Wels W, Zörnig M, Dietrich U, Medyouf H, Sevenich L, Farin H, Krause D und Mitarbeiter

WS 2017 | 18Vorlesung Molekulare Onkologie und Tumorimmunologie (MOT) Tumormik-roenvironment/Tumor und Entzündung/TumormakrophagenGreten F, Zörnig M, Krause D, Medyouf H, Farin H, Sevenich L

SS 2017Anwendungen der Biologie in der Häma-tologie für MedizinstudentenKrause D

WS 2017 | 18, SS 2018Vorlesung GraduiertenkollegGreten F

Education

Lehrveranstaltungen

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Der Verein „Freunde und Förderer des Georg-Speyer-Hauses“

The Association„Friends and Sponsors of the Georg-Speyer-Haus“

Innovative Forschung und wissenschaft-licher Fortschritt in unserer Gesellschaft sind nur möglich durch das Engagement der Wissenschaftler/innen und die aktive Unterstützung von Forschungsförderern aus Öffentlichkeit, Wissenschaft und Wirtschaft. Diesem Engagement hat sich der Verein „Freunde und Förderer des Georg-Speyer-Hauses“ verpflich-tet: Sein Ziel ist es, über die Grundfi-nanzierung durch Bund und Länder hinaus für weitere erforderliche Mittel zu sorgen und so das hohe Niveau der Grundlagenforschung zu sichern. Mitglied im Verein kann werden, wer den wissenschaftlichen Fortschritt im Bereich der Krebsforschung und der experimentellen Therapie zum Wohle der Allgemeinheit fördern möchte und Interesse hat am Forschungsprozess und am Diskurs über Ergebnisse und deren Nutzen für die Allgemeinheit.Neben der einfachen Mitgliedschaft (Freund/innen) und der Forschermitglied-schaft (Wissenschaftler/innen, Student/in-nen) besteht die Möglichkeit der fördern-den Mitgliedschaft für Einzelpersonen oder Firmen. Förderer können im Jahrbuch und auf der Spendentafel aufgeführt werden.Da der Verein eine gemeinnützige Einrichtung ist, sind Mitgliedsbeiträge und Spenden im Rahmen der zulässigen Höchstbeträge von der Steuer absetzbar.

Innovative research and scientific ad-vances are only possible through generous financial support from public and private sponsors. The association „Friends and Sponsors of the Georg-Speyer-Haus“ has committed itself to this task. The major goal of the association is to raise the necessary funds to supplement the basic financing provided by the federal and state governments. This should ensure a continuing high quality of basic research.Everybody who would like to support research in the fields of cancer and experimental therapy is welcome to join the association. Private persons can become supporting members („friend“) or research members (scientists and students). Moreover, private individu-als and companies may obtain cor-porate membership. Sponsors will be listed in both the annual report and the table of benefactors in the Institute.

Since the association is a non-profit organisation, all membership fees and donations are tax deductable.

Jährliche MitgliedsbeiträgeAnnual membership fees

ForschermitgliedScientist 100,– € StudentenStudents 12,– €

FreundFriend 150,– €

FördererSponsor 1000,– €

Firmenmitgliedschaft Company membership 5000,– € Kontakt

Gabriele HecklMitgliederbetreuung und Schatzmeister / members care and treasurerTel.: +49 (0) 69 63395-212Fax: +49 (0) 69 63395-280E-Mail: [email protected]

Prof. Dr. Bernd Groner1. Vorsitzender / ChairmanTel.: +49 (0) 69 63395-0E-Mail: [email protected]

www.georg-speyer-haus.de/friends/index.htm

The Association

FREUNDE UND FÖRDERER DES GEORG SPEYER HAUSES E.V.

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Finanzierung des Georg-Speyer-Hauses: Die Grundfinanzierung des Georg-Speyer-Hauses wird vom Bundesministerium für Gesundheit und dem Hessischen Ministerium für Wissenschaft und Kunst getragen.

Funding of the Georg-Speyer-Haus:The basic funding of the Georg-Speyer-Haus is provided by the Federal Ministry of Health and the Ministry of Higher Education, Research and the Arts of the State of Hessen.

Einzelne Projekte werden unterstützt durch:Individual projects are supported by:

BerGenBio ASBoehringer Ingelheim GmbHBundesministerium für Bildung und Forschung (BMBF) Deutsche Forschungsgemeinschaft (DFG) Deutsche José Carreras Leukämie-Stiftung Deutsche Kinderkrebsstiftung Deutsches Konsortium für Translationale Krebsforschung (DKTK) Dr. Bodo Sponholz-StiftungDr. Hans-Joachim und Ilse Brede-StiftungDr. Rolf M. Schwiete Stiftung Else Kröner-Fresenius-StiftungEuropean Commission European Research Council (ERC) Hans und Wolfgang Schleussner-StiftungH.W. & J. Hector Stiftung Janssen Research & Development LLCKlinikum der Johann-Wolfgang-Goethe UniversitätLOEWE Zentrum für Zell- und Gentherapie FrankfurtLOEWE Forschungsschwerpunkt: Ubiquitin-Netzwerk Stiftung Deutsche Krebshilfe Werner Schaefer-Stiftung Wilhelm Sander-StiftungWilly Robert Pitzer Stiftung

Für Zuwendungen von Privatpersonen und Organisationen sind wir dankbar. Gerne stellen wir eine Spenden-bescheinigung aus.

Unsere Bankverbindung lautet: Deutsche Bank Frankfurt IBAN: DE93 5007 0010 0255 1604 00BIC: DEUTDEFF

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Impressum

Herausgegeben vonGeorg-Speyer-HausInstitut für Tumorbiologie und experimentelle TherapiePaul-Ehrlich-Straße 42 – 44D-60596 Frankfurt am Main

RedaktionProf. Dr. Florian R. GretenStefanie Schütt

GestaltungStählingdesign, Darmstadt

BildnachweisPortraits: Stefan Streit, KönigsteinPortraits/Veranstaltungen: Andreas Reeg, DarmstadtAlle übrigen Fotos: Georg-Speyer-Haus

DruckWerbedruck Petzold GmbH, Gernsheim

72www.georg-speyer-haus.de

„… Großtat des Geistes ...? Mein lieber Kollege, es ist nichts anderes, als daß ich sieben Jahre Pech und einen Moment Glück gehabt habe.“Paul Ehrlich

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