hairy cell leukemia prof. arcangelo liso università di foggia rieti 27 ottobre 2006

Hairy cell leukemia

Prof. Arcangelo Liso

Università di Foggia

Rieti 27 ottobre 2006

HCL: PERIPHERAL BLOOD

CD22HAIRY CELL LEUKEMIA

- Pancytopenia- Dry-tap (marrow fibrosis)- Marked splenomegaly- No lymphadenopathy- Circulating hairy cells

HAIRY CELL LEUKEMIA: UNSOLVED ISSUES

• ORIGIN OF HCL CELLS

• MORPHOLOGICAL APPEARANCE (HAIRY PROJECTIONS)

• SELECTIVE HOMING OF TUMOR CELLS TO BM, PB, LIVER,

AND SPLEEN (WITH SPARING OF LYMPH NODES)

• PHAGOCYTIC ACTIVITY (LATEX PARTICLES)

• CONSISTENT BONE MARROW FIBROSIS

• DIFFERENTIAL DIAGNOSIS WITH RELATED CONDITIONS (HCL VARIANT, SPLENIC MARGINAL ZONE LYMPHOMA WITH VILLOUS LYMPHOCYTES)

• GOOD RESPONSE TO INTERFERONS AND PURINE ANALOGUES

<-2 >2 B

HCL BM biopsies

MCLB-CLLDLBCLFLBL

HCLHCL CD19+ enriched

A

Bone marrowsignature

J. Ex. Med 199:59,

2004

HCL SHOWS HOMOGENEOUS SIGNATURE (UNSUPERVISED ANALYSIS)

THE ORIGIN OF HCL CELLS

POST-GERMINAL CENTER CELLOF UNKNOWN STAGE (GIVEN THE UNIQUE PHENOTYPE OF HCL ) (REAL/WHO)

Somatic IgV mutations (no ongoing)

No GC-related markers

N&M

GC

0 0.1

-0.1

0.3

0.5

0.7

0.9

-0.3

-0.5

-0.7

-0.9

N&M GC HCL N HCLM M

N

HCL are more related to memory B cells

Pathogenetic implications: Lack of chromosomal translocations (typical of HCL) may be to the fact that the mechanisms generating these aberrations (i.e. VDJ recombination, somatic hypermutation, etc. ) are switched off in memory B cells.

(Basso et al, J Exp Med, 199: 59, 2004)

- In spite of similarities, a peculiar expression pattern

of cytokines, chemokine receptors and adhesion

molecules, distinguishes HCL from tonsil normal

memory B cells

- this finding might be caused by unknown oncogenic

event(s) leading to HCL or reflect derivation from

a particular subset of memory B cells (splenic mz

B-cells ?)

DIFFERENCES BETWEEN HCL AND MEMORY B CELLS

N CC HCLCB M

Activation

DNA metabolism

Transcription

Proliferation

- CHC1L

- CCND1

Signaling

- IkB epsilon

- CAMK1

- DGKA

- SIPA1

- IRF4- EGR3

- MADH3- LYL1- ZNF135

Chemokine-receptorsChemokines

Cytokines - IL-6- TNSF11

- CCR7

antiApoptosis

- BIKpro

Cytokine-Receptors- TNFRSF1B

- WSX1

N CC HCLCB M

Adhesion - CD1c

- PCDH9

- GARP- CD9- CD103

- ICAM3

Cellular programs affected by malignant transformation

HCL: ADHERENCE TO GLASS SURFACE

ACTINANNEXIN I

PHAGOCYTOSIS MO/MØ FEATURES

• TRAP lysosomal activity

• Expression of Mo/Mø associated molecules: - CD11c - CD68 Mo/Mø-restricted - MIR10 (CD85d)

- Endoglin (CD105)

- CD63 - CPVL

- c-MAF

HCL-specific genes (n=89)

CB CC N MHCL FL BL B-CLLDLBCL MCL

Normal B cells B cell lymphomas

- ANXA1

- FLT3

- CD63

- SCNB1- SDC3

- FGF2

- CCND1

- IL3Ra

- TIMP1

- MYF6

- PLXNC1

- FGFR1

- PTPRM

- SYT1

- TIMP4- arrestin B

Previously knownIHC performed

- EPB41L2

- CXCR5

- GAS7

GAS-7

F-ACTIN

OVERLAY

GAS7

• BELONGS TO THE FAMILY OF THE GROWTH-ARREST-SPECIFIC (GAS) PROTEINS

• IS EXPRESSED IN ADULT MOUSE BRAIN AND TESTICLE

• ESSENTIAL FOR NEURITE OUTGROWTH IN CULTURED CEREBELLAR NEURONS

• GAS7 CAN INDUCE THE FORMATION OF EXTENDED CELLULAR PROCESSES IN NIHT3 BY INTERACTING WITH ACTIN AND MEDIATING RE-ORGANIZATION OF MICROFILAMENTS

(She B-R et al., Exp. Cell Research 273:34-44, 2002)

THE ECTOPIC EXPRESSION OF GAS7 IN NIH3T3 CELLS INDUCES

THE FORMATION OF EXTENSIVE CELLULAR PROCESSES WITH

BRANCHES AND KNOBS, AND SHIRINKAGE OF THE CELL BODY

GAS-7 F-ACTIN OVERLAY

GAS-7 F-ACTIN OVERLAY

HCL: SPLEEN (pre-IFN era)

HOMING OF HCL CELLS TO SELECTED BOBY SITES

HCL: LIVER INVOLVEMENT

SPLEENBASEMENTMEMBRANE

(RING FIBER)

LATERAL FIBER

A NORMAL SINUS

LONGITUDINALRETICULUMFIBERS(CORE FIBERS)

ENDOTHELIALCELL

DEGENERATIVEENDOTHELIALCELL

HAIRY CELL

B PARTIALLY INVOLVED

REDBLOODCELL

CPSEUDOSINUS

D BLOOD LAKE

HAIRY CELLS

Interaction VLA-4 (HCL) / VCAM-1 (sinusoids)

HCL-specific genes

CB CC N MHCL FL BL B-CLLDLBCL MCL

Normal B cells B cell lymphomas

- ANXA1

- FLT3

- CD63

- SCNB1- SDC3

- FGF2

- CCND1

- IL3Ra

- TIMP1

- MYF6

- PLXNC1

- FGFR1

- PTPRM

- SYT1

- TIMP4- arrestin B

Previously knownIHC performed

- EPB41L2

- CXCR5

- GAS7

BM

SPLEENRED PULP

PE

RIP

HE

RA

L B

LO

OD

ANNEXIN I

TRANS-ENDOTHELIALMIGRATION

CXCR5 CCR7

HOMING T0LYMPH NODES

- Degradation of the pericellular matrix- Crossing of tissue barriers- Extravasation- Neo-angiogenesis

EN

DO

TH

EL

IUM

HCL REMAINS CONFINED TO BLOOD COMPARTMENTS

EXTRANODALINVOLVEMENT

MMPs ACTIVITY (INVASION AND METASTASIS)

NODAL INVOLVEMENT

TIMP-1TIMP-4RECKTHROMBOSPONDIN-1

(inhibitors of metalloproteases)

HCL: BONE MARROW FIBROSIS

- BM fibrosis: Fibronectin + argyrophilic reticulin fibers

- HCLs bind to fibronectin

- HCLs synthetize/assemble fibronectin matrix (Burthem)

- Production rate fibronectin under control of autocrine b-FGF secreted by HCLs (Aziz)

- Microarray study confirms up-regulation of b-FGF and FGFR (autocrine loop)

b-FGF

PRODUCTION OF

FLT3-L

VLA4

VLA5

FIBRONECTINRECEPTORS

ADHESION TO

FIBRONECTIN MATRIX

BM MICRO-ENVIRONMENT

HAIRY CELL

RETICULINFIBROSIS

IL-3

b-FGF

FGFR

HYPOTHETICAL MODEL OF BONE MARROW FIBROSIS IN HCL

IL3R

FLT3

TGF-beta

Collagen III

OTHER GENES

6.1 (12.0) Gene 1 6.0 (41.5) Gene 2 5.3 (10.1) Gene 3

5.2 (-26.5) Gene 35.1 (-6.4) Unknown 2.7 (-8.5) Gene 4

IMMUNIZATION

PRODUCTION OF mAb

IMMUNOHISTOCHEMISTRY

AVAILABLE ANTIBODY

DISEASE (A) DISEASE (B)

HCL-specific genes

CB CC N MHCL FL BL B-CLLDLBCL MCL

Normal B cells B cell lymphomas

- ANXA1

- FLT3

- CD63

- SCNB1- SDC3

- FGF2

- CCND1

- IL3Ra

- TIMP1

- MYF6

- PLXNC1

- FGFR1

- PTPRM

- SYT1

- TIMP4- arrestin B

Previously knownIHC performed

- EPB41L2

- CXCR5

- GAS7

Annexin 1 expression

0

4000

8000

12000

16000

20000

HC

L

CB

CC N M FL

BL

DL

BC

L

MC

L

B-C

LL

Ave

rage

Diff

eren

ce (

MA

S5.

0)

Annexin 1 expression

0

4000

8000

12000

16000

20000

HC

L

CB

CC N M FL

BL

DL

BC

L

MC

L

B-C

LL

Ave

rage

Diff

eren

ce (

MA

S5.

0)

- ANXA1: REGULATION OF EARLY INFLAMMATORY RESPONSES (INHIBITION OF ADHESION,

MIGRATION, ETC.)

- ANXA1 BINDS TO ACTIN FIBERS (CYTOSKELETON) - EXPRESSED IN MYELOID CELLS, STROMAL CELLS, MACROPHAGES, SUBSET T CELLS

ANXA1 PROTEIN EXPRESSION IN B-CELL LYMPHOMAS

LYMPHOMA TYPE N=500 ANXA1

HCL 64 62/64*

HCL variant (HCLv) 8 0/8

B-CLL 80 0/80

Prolymphocytic leukemia 3 0/3

Splenic MZL** 50 0/50

Nodal MZL 15 0/15

Lymphoplasmocytic lymphoma 30 0/30

Follicular lymphoma 65 0/65

Mantle cell lymphoma 14 0/14

DLCL-B 100 0/100

Burkitt lymphoma 10 0/10

Myeloma 50 0/50

* Two cases turn out to be HCLv; ** With and without circulating villous lymphocytes

HAIRY CELL LEUKEMIA

HCL cell Annexin A1

CLASSIC HODGKIN’S DISEASE

B-CELL LYMPHOMAS SIMULATING HCL

• HCL-VARIANT (HCLv)

• LYMPHOCYTES (SLVL) SPLENIC LYMPHOMA

WITH VILLOUS

• OTHER LYMPHOMAS WITH CIRCULATING

VILLOUS LYMPHOCYTES

Biphasic morphology

Villous lymphocytes

Splenic MZL 1% of all lymphomas

Indolent tumor

Leukemic phase: Villous lymphocytes

Phenotype: Similar to HCL

Genetic: Allelic loss 7q31-32 (SLVL)

Clinic: Splenomegaly

HCV+ SLVL respond to IFN

Falini B., Lancet 363:1869, 2004

SLVL: INTRASINUSOIDAL PATTERN (CD79a)

SLVL (Annexin A1)

*

CONCLUSIONS

• HCL DISPLAYS A DISTINCTIVE AND HOMOGENOUS GEP SUGGESTING THAT IT IS A SINGLE DISEASE, CONSISTENT WITH ITS HOMOGENOUS MORPHOLOGY AND CLINICAL BEHAVIOUR

• HCL SIGNATURE IS MORE RELATED TO MEMORY B CELLS THAN TO NAIVE AND GERMINAL CENTER B CELLS, OR LYMPHOBLASTOID B CELLS AND PLASMA CELLS

• HCL DIFFERS FROM MEMORY B CELLS MAINLY IN THE EXPRESSION OF CYTOKINES, CHEMOKINE RECEPTORS AND ADHESION MOLECULES

• GEP ANALYSIS IDENTIFIES 89 GENES WHICH ARE SPECIFICALLY UP- OR DOWN-REGULATED IN HCL. THEY ARE POSSIBLY RESPONSIBLE FOR THE DISTINCTIVE FEATURES OF HCL CELLS ( EXPERIMENTAL VALIDATION REQUIRED)

• INFORMATION OF GEP HAS BEEN ALREADY TRANSLATED INTO A SPECIFIC, CLINICALLY USEFUL ANNEXIN A1-BASED IMMUNOASSAY

BRUNANGELO FALINI (PERUGIA UNIVERSITY)

ENRICO TIACCI (PERUGIA UNIVERSITY)

ROBERTA BENEDETTI (PERUGIA UNIVERSITY)

ELENA SABBATINI (BOLOGNA UNIVERSITY)

STEFANO PILERI (BOLOGNA UNIVERSITY)

ALESSANDRO PULSONI ( ROME UNIVERSITY)

ROBIN FOA (ROME UNIVERSITY)

FRANCESCO DI RAIMONDO (CATANIA UNIVERSITY)

ACHILLE AMBROSETTI (VERONA UNIVERSITY)

KATIA BASSO (COLUMBIA UNIVERSITY)

RICCARDO DALLA FAVERA(COLUMBIA UNIVERSITY)

IMMUNOPHENOTYPE OF CHRONIC B-CELL LEUKEMIAS

Antigen CLL HCL SLVL

CD5 + + - - CD23 ++ - -

FMC7 ± + + + +

CD10 - - -

CD25 - + + -/+

CD68 - + -/+

CD11c -/+ + +

DBA44 - + +

CD103 - -/+ -

ANNEXIN A1 - +++ -