HEIM

Hematology and Immunology

6-Dec-11

HEIM

Faculty(building D level 0)

Bieghs LiesbethDe Bruyne ElkeDe Smedt AndreaDehui Xu De Zanger RonaldJinsong HuJoost de ter Beerst Marie Lemaire MiguelLub Susanne Maes Ken Menu ElineNur Haneen Seynaeve CarineVan Valckenborgh ElsVanderkerken KarinWillems Angelo

Clinical Hematology

Caron CharlotteDe Becker AnnFostier KarelSchots RikTrullemans Fabienne

Stem cell laboratory(BTC building level 1)

Arras NicoleDe Becker AnnDe Veirman KimXu SongVan Riet Ivan

Laboratory of Hematology

Bakkus MarleenDe Smet DieterDe Waele MarcDemanet ChristianJochmans KristinOrlando ChristelleVan der Meeren SamVanderstraeten JonaVercammen Martine Verheyden SonjaVerloes An

6-Dec-11

Overview of research lines

• Multiple myeloma: basic, translational, clinic

• Thrombophilia: basic, translational, clinic

• Clonal hemopoietic stem cell disorders: translational

• Natural killer cell receptors: translational

• Molecular targets for risk-group stratification in childhood acute lymphoblastic leukaemia: translational and clinic

6-Dec-11

MULTIPLE MYELOMA

� Plasma cell cancer

� Accumulation in the bone marrow

� Paraprotein

� Angiogenesis

� Osteolytic bone disease

� 500 new patients/year

� Older patients

6-Dec-11

INTERACTIONS BM-MM CELLS

CAM-DR

Lemaire et al. Advances in Cancer Research,

2011, 110:19-42.

angiogenesis

Drug resistance

Bone disease

Survival & proliferation of MM cells

Immune system

Within clinical trial context

Panobinostat

Carfilzomib

Pomalidomide

Elotuzumab

Routinely used

Melphalan

Prednisone

Dexamethasone

Adriamycine

Cyclophosphamide

Velcade

Revlimid

Thalidomide

Bendamustine

Caelyx (liposomal adriamycine)

MEDICATION USED FOR THE TREATMENT OF MYELOMA

6-12-2011

6

ChemotherapyCorticosteroids

Proteasome inhibitor

Immunomodulators

HDAC inhibitor

CS-1 antibody

6-Dec-11

*Methylation

*Post-T histon Modifications*mi RNA (in coll. with

T.VandenDriessche)*In vitro and in vivo* Epigenetic regulating agents(Dr. Elke De Bruyne)

*IGF-1*Notch*Hypoxia*MesenchymalStem cells(Prof. I. Van Riet)

*Different sub

Populations(Dr. Els Van Valckenborgh)*CAM-DR-integrines(Dr. Eline Menu)*Nanobodies(Miguel Lemaire-in coll. with T. Lahoutte)

*MDSc(Dr. Els Van Valckenborghin coll. with

J. Van Ginderachter)*iNKT cells(Dr. Eline Menu)*Immune therapy(Dr. R. Schots in

collaboration with K.

Thielemans)

*Participation toPhase II-III studies(Prof. R. Schots)

Myeloma Center Brussels www.myeloma.be

6-Dec-11

Division of Clinical Hematology

Clinical HematologyR SchotsF TrullemansA De BeckerC CaronK Fostier

Clinical Research

= participation in phase II-III clinical trialsMyeloma >> lymphoma/leukemia/transplantation

Translational researchDendritic cell therapy in myeloma (Collaboration with LMCT)Targeting residual cancer cells in myelomaTargeting BM microenvironment in myeloma

Clinical activity

Hospital Day Clinic Stem cell laboratoryI Van Riet

Hematopoietic stem cell transplantation

Research activity

6-Dec-11

Material

Human MM cell lines

Primary patient material

Syngeneic murine model: 5TMM

MATERIAL

�Similar characteristics as human disease

5T2moderate growth (12w)

osteolytic lesions

5T33

rapid growth (3w)

no osteolytic lesions

In vitro cell lines

6-Dec-11

Techniques-in vitro

• Cell culture and isolation (MACS, FACS)

• Proliferation, apoptosis, migration, invasion, clonogenic assays

• Cultures in hypoxic conditions (in collaboration with TROP)

• Flow cytometry: analysis + sorting (in collaboration with LMCT)

• Elisa

• Western blot analysis

• RT-PCR, Real Time PCR, Chip analysis

6-Dec-11

5T2 or 33MM model

0 weeks

Inject 5TMM cells SacrificeMyeloma Disease

Paraprotein

FACS

Angiogenesis

MVD

Bone disease

Techniques: in vivo

- 5T2 or 5T33MM model

- prophylactic or therapeutic treatment

6-Dec-11

BMEC

MM

BMSC

Osteoclast

BO

NE

CCR2

MCP-1

CD

44

v1

O

67kD LR

La

min

in-1

IGF-1R

IGF-I

CD44v6

67kD LR

Osteoblast

DKK1 RANK

RANKL

RANKL

MIP-1αααα

IL-1ββββ

MIP-1αααα

CCR5

VEGFPDGFbFGFAng-1

HGF

CXCR4

SDF-1ααααMMP9adipocyt

lept

in

6-Dec-11

Funding: FWO-Vlaanderen, OZR-VUB (GOA, HOA and projects), Stichting tegen Kanker, Vlaamse Kankerliga, MMRF, IMF, EUFP6, EUFP7

International Collaborations: •P. Croucher (Sheffield, UK)•H. Jernberg-Wiklund (Uppsala, USA)•S. Lentsch (Pittsburgh, USA)•W. Matsui (Baltimore, USA)•D. Hose (Heidelberg, Germany)•B. Klein (Montpellier, France)•S. Sahota (Southampton, UK)•T. Anderson (Danmark)•H. Johnson (Aarhus, Danmark)•X. Leleu (Lille, France)•I. Slaper (Utrecht, The Netherlands)

6-Dec-11

ThrombophiliaThrombophilia

• Acquired or inherited tendency for the development of

venous thromboembolism

• Our interest: inherited thrombophilia

– Clinical presentation: thrombosis at young age, recurrent events,

familial history, concomitant risk factors

– Treatment, prognosis and counselling in function of classification

6-Dec-11

ThrombophiliaCongenital AT and PC deficiency

• Objectives

– Full laboratory assessment of antithrombin (AT) and protein C

(PC) deficiency

– Molecular characterization of mutations in the AT and PC gene

– Investigation of correlation genotype/phenotype

– National and international collaborations as expert centre

for hereditary AT deficiency

• Researchers

– Kristin Jochmans, MD, PhD

– Christelle Orlando, Msc

6-Dec-11

Thrombophilia in assisted reproduction

• Objectives

– Prospective study: influence of thrombophilia on the outcome of

IVF/ICSI in couples with recurrent failure of assisted

reproduction

(implantation failure and miscarriage)

– Protein Z and Protein Z-dependent protease inhibitor as new

parameters in thrombophilia

• Researchers– Kristin Jochmans, MD, PhD

– Christelle Orlando, Msc

6-Dec-11

Thrombophilia Techniques

• Chromogenic substrate assays

• Clotting-based assays

• ELISA

• Immuno-electrophoresis

• Rocket electrophoresis

• PCR

• Sequencing

• MLPA (multiplex ligation-dependent probe amplification)

6-Dec-11

Clonal hemopoietic stem cell

disorders

• Myelodysplastic syndromes and chronic myeloproliferative neoplasms

• Team: M. De Waele, F. Trullemans, K. Jochmans, D. De Smet, S. Van der Meeren,

M. Bakkus

• Analysis of bone marrow aspirates from patients seen in the Department of Clinical Hematology (R. Schots) in the period 2002 -2009

• Immunophenotyping CD34+ cells:

– Question: Diagnostic potential?

– Myelodysplastic syndromes: D. De Smet

– Chronic myeloproliferative neoplasms: S. Van der Meeren

6-Dec-11

Clonal hemopoietic stem cell

disorders

• miRNA profiling:

– Myelodysplastic syndromes:

• miRNA profiling done in UZ Gent (Department of Medical Genetics, F. Speleman, P. Mestdagh)

• Validation and analysis of the data now performed

• Question: Is there a diagnostic miRNA signature for MDS compared to normal bone marrow and secondary cytopenia?

– Chronic myeloproliferative neoplasms:

• miRNA profiling still has to be performed.

• Funding: FWO-Vlaanderen

6-Dec-11

Natural killer cell receptors

• Natural killer cell receptor polymorphism in leukemia and hematopoietic

stem cell transplantation.

• Team: C. Demanet, S. Verheyden

• Analysis of samples of patients seen in the Department of Clinical

Hematology (R. Schots)

• Results:

– The identification of NK cell receptor phenotypes associated with

leukemia

– A defined donor activating NK cell receptor genotype protects against

leukemic relapse after related HLA-identical hematopoietic stem cell

transplantation

– The susceptibility to myeloid and lymphoid leukemia is mediated by

distinct inhibitory KIR-HLA ligand interactions

– The role of the inhibitory KIR ligand HLA-Bw4 and HLA-C expression

levels in the recognition of leukemic cells by natural killer cells.

• Funding: FWO-Vlaanderen

6-Dec-11

Molecular targets for therapy

stratification in ALL

• MRD analyses by specific Ig/TCR real-time PCR on predefined timepoints during therapy

• Prognostic impact of new molecular targets (IKZF, CRFL2)

• Team: – Marleen Bakkus, PhD

– Jona Van der Straeten, Msc

– Emmanuella Kabongo Kanjinga, Msc

– in collaboration with the BSPHO and the EORTC-CLG

• Analysis of bone marrow samples of all Belgian pediatric ALL patients

• Techniques: – Ig/TCR target identification by genescan analysis

– Sequencing of clonal Ig/TCR gene rearrangements

– patient specific design of TaqMan PCR assays

– MLPA (multiplex ligation-dependent probe amplification)

6-Dec-11

Molecular targets for therapy

stratification in ALL

• Funding: WFWG, ‘Stichting tegen Kanker’, ‘Nationaal Kankerplan’

(years)0 1 2 3 4 5

0

10

20

30

40

50

60

70

80

90

100

O N Number of patients at risk :23120 117 112 105 1009 12 7 5 4 4

Overall Logrank test: p<0.0001

EFS %

MRD > 10-2MRD < 10-2

TP1: MRD < 10-2

TP1: MRD > 10-2

VHR

SR

SR if MRD < 10-3 on TP2

VHR if MRD > 10-3 on TP2

No SCT if MRD < 10-3 on TP2

SCT if MRD > 10-3 on TP2

New EORTC-CLG guidelines

Results

EORTC 58881

EORTC 58951

Thank you !!

6-Dec-11