PATOGENESI MOLECOLARE E PATOGENESI MOLECOLARE E POSSIBILI TARGET TERAPEUTICI POSSIBILI TARGET TERAPEUTICI

NEL MIELOMA MULTIPLONEL MIELOMA MULTIPLO

Alberto RocciOspedale San Giovanni Battista di Torino

Università degli Studi di Torino

Mediterranean School of OncologyOrvieto, 20 novembre 2009

MULTIPLE MYELOMAAccumulation of abnormal

plasma cells in the bone marrowProduction of monoclonal protein

detectable in serum or urine

Organ damage: kidney, bone, blood

Hematological Neoplasia Pathogenesis

Mantle Cell Lymphoma Bcl1

Primary Effusion Lymphoma HHV-8

Burkitt Lymphoma c-MYC

Follicular Lymphoma Bcl2

Chronic Myeloid Leukemia BCR/ABL

Multiple Myeloma multifactorialpathogenesis ?

MM PATHOGENESIS

GENOMIC INSTABILITYMICROENVIRONMENT ALTERATIONS

IMMUNE SYSTEM IMPAIRMENT

GENOMIC INSTABILITYIN MULTIPLE MYELOMA

MemoryB Lymphocyteplasmacells

B-cell precursor

Memory B lymphocyte

Mantle-zone Germinal

Centre

Marginal zone

Naive B lymphocyte

Ext

ra G

C-m

atu

rati

on

GC

-mat

ura

tio

n

BM and PB Lymph Node

VDJ RECOMBINATION

IgH switch recombination

somatic hypermutation

Alterations regarding these three mechanisms are commonly involved in the pathogenesis of hematological malignancies.

In particular it is involved the 14q32 chromosomal region

t(4p16;14q32) 15% FGFR3 GF rec TK MMSET Transcription regulator

TACC3 ND

t(6p21;14q32) 5% CICLINA D3 Cell cycle regulator

t(6p25;14q32) 5% MUM/IRF4 Regulation of IFN transc.

t(14q32;16q23) 5%-10% c-maf Transcriptional factor

t(14q32;20q12) 5% b-MAF Transcriptional factor

t(11q13,14q32) 15%-20% CICLINA D1 Cell cycle regulator MYEOV ND

TRASLOCATION % ONCOGENE ROLE

PRIMARY Translocations

PRIMARY Translocations

Almost all the genetic aberrations identified in MM are present in MGUS

MemoryB Lymphocyteplasmacells

Mantle-zone Germinal

Centre

Marginal zone

MM

GC

-mat

ura

tio

n

THE MM STORY PROBABLY STARTS HERE

Two pathway hypotesisMultiple Myeloma

and MGUS

Hyperdiploid pathway(multiple chromosome

trisomies)

Trisomies of chromosome 3,5,7,9,11,15,19,21

50%

Nonhyperdiploid pathway(high prevalence of IgH translocations)

11q13 (CCND1)4p16 (FGFR3 and MMSET)6p21 (CCND3)16q23 (MAF)20q11 (MAFB)

50%

Chromosome 13 monosomy/deletion is an early event that can occur in both groups

Cyclin D disregulation

Nonhyperdiploid pathway

Virtually all MM and MGUS plasmacells have increased levels of Cyclin D1, D2 or D3, despite the low proliferative rate

Cyclin overespression does not lead to a growth improvement, however it seems that this situation can make plasma cells

more prone to growth stimuli

RB 211540_s_at

208712_at cyclin D1

200951_s_at cyclin D2

206363_at MAF

204379_s_at FGFGR3

209053_s_at MMSET

218559_s_at MAFB

201700_at cyclin D3

FGFR3

MMSET

C-MAF

MAFB

CCND3

CCND1

CCND2

Bergsagel PL, Blood 2005

4p1614%

Maf8%

11q1316%

D132%

D1+

D27%

D219%

6p212%

None

RB1

2%

TC classification

Alteration of Rb pathway

Bergsagel PL and Kuehl WM, JCO 2005

DEL13 e/o1q gain

NON-HYPERDIPLOID

HYPER DIPLOID55%-60%

TRISOMY3, 5, 7, 9,

11,15,19,21

11q136p21

16q2320q11

4p16

IgH tx

ONCOGENIC EVENTS

GENOMIC INSTABILITY

Bergsagel PL, Blood 2005

Supervised analysis 263 genes are differentially expressed between healthy and MGUS

380 genes are differentially expressed between healthy and MM

74 genes are differentially expressed between MGUS and MM

There are fewer differences at the gene expression level

between MGUS and MM than between healthy and MM or

healthy and MGUS

Davies FE et al., Blood 2003

FROM MGUS TO MULTIPLE MYELOMA

SECONDARY Translocations

Occurres as progression event

Rare or absent in MGUS patients

Can be mediated by mechanisms that do not involve the three B-cell-specific DNA-modification mechanisms

Bergsagel PL and Kuehl WM, JCO 2005

DEL13 e/o1q gain

NON-HYPERDIPLOID

HYPER DIPLOID55%-60%

TRISOMY3, 5, 7, 9,

11,15,19,21

11q136p21

16q2320q11

4p16

IgH tx

ONCOGENIC EVENTS

MYC dysregulation (45%)

Secondary (Ig) TLC

Karyotypic abnormalities

DEL 17p DEL 1p

MAPK pathway activating mutations

N-ras (codon 12,13,61)K-ras (40%-55%)FGFR3

NFkB pathway activating mutations

TRAF3cIAP1/2CYLDNIK

RB1 pathway p18INK4c

inactivating muts

TP53

GENOMIC INSTABILITY

Bergsagel PL, Blood 2005

Disease stages and timing of oncogenic events

Bergsagel PL and Kuehl WM, JCO 2005

DEL13 e/o1q gain

NON-HYPERDIPLOID

HYPER DIPLOID55%-60%

TRISOMY3, 5, 7, 9,

11,15,19,21

11q136p21

20q11 16q23

4p16

IgH

MYC dysregulation (45%)

Secondary (Ig) TLC

Karyotypic abnormalities

DEL 17p DEL 1p

MAPK pathway activating mutations

N-ras (codon 12,13,61)K-ras (40%-55%)FGFR3

NFkB pathway activating mutations

TRAF3cIAP1/2CYLDNIK

RB1 pathway p18INK4c

inactivating muts

TP53

ONCOGENIC EVENTS

GENOMIC INSTABILITY and Prognosis

GENOMIC ALTERATION AND EFS – OS

Avet-Loiseau H et al., Blood 2007

Sagaster V et al, Leukemia 2007

GENOMIC ALTERATIONSAND NEW DRUGS

BORTEZOMIB LENALIDOMIDE

Del(13q)

T(4;14)

GENOMIC ALTERATION AND TARGET THERAPY

Clin Cancer Res 2009

Leukemia 2009

GENOMIC INSTABILITY: CONCLUSIONS

MM seems to include several diseases that have differences in early events, global gene expression patterns,

clinical features, prognosis and response to therapy

Common translocations have a major prognostic role in patients treated with standard or high dose chemotherapy,

their role in patients treated with novel drugs is still under investigation

IgH translocation are common in half of MM and MGUSpatients whereas other patients display trisomies

MICROENVIRONMENT ALTERATION IN

MULTIPLE MYELOMA

A TYPICAL CASE OF MM DOES NOT HAVE ALL THESE FEATURESA TYPICAL CASE OF MM DOES NOT HAVE ALL THESE FEATURES

HOW MICROENVIROMENT SUSTAINS MALIGNANT GROWTH

MICROEVIRONMENT AND MM(main actors)

Plasma cellPlasma cellOsteoclastOsteoclast

Endothelial cellEndothelial cellHematopoietic cellsHematopoietic cellsMesenchymal cellsMesenchymal cells

OsteoblastOsteoblastECMECM

Dendritic cellsDendritic cellsNatural immunity Natural immunity

cellscellsT-cellsT-cellsB-cellsB-cells

MM cells and bone marrow interaction

Hideshima T & Anderson KC Nat Rev Cancer 2007:585-598

CYTOCHINE-MEDIATED SIGNALING

ADHESION-MEDIATED SIGNALING

Hideshima T et al., Nat Rev Canc 2007

Molecular modification due to cell-to-cell interaction

Hideshima T & Anderson KC Nat Rev Cancer 2007:585-598Hideshima T et al., Nat Rev Canc 2007

HGF/MET in Multiple Myeloma

Börset M et al, Blood 1996

HGF

Met

β-actin

Primary myeloma cells

Normal BM Cells

Normal PB Cells

Seidel C et al, Blood 1998

p= 0.007

p= 0.047

RESULTS: MET and PFS - OS

The two groups displayno differences about

beta2microglobulin andalbumin levels or

FISH feature

Rocci A. et al, ASH 2009

MICROEVIRONMENT AND MM(How does it plays)

IL-6IL-6

TNF-TNF-

RANK-LRANK-L

VEGFVEGF

TGF-TGF-

ProstaglandinsProstaglandins

Multiple role and indipendence to Multiple role and indipendence to growth stimuligrowth stimuli

resistance to apoptosisresistance to apoptosis

homing and migrationhoming and migration

angiogenesisangiogenesis

replication and proliferation replication and proliferation

resistance to inhibition stimuliresistance to inhibition stimuli

Hideshima T and Anderson KC, Nat Rev Cancer 2002

MULTIPLE ROLE OF IL-6 IN MALIGNANT PC

IL-6 induces expression and secretion of VEGF

Blood vessel

angiogenesis

IL-6VEGF

VEGFIL-6

MMcell

VEGF, IL-6

VEGF

Laminin,

fibronectinBMSCs

Inhibition ofmaturation

Dendritic Cell

Osteoclast

Increase of bone-resorbing

activity

Proliferation,migration

Lin et al. Cancer Res 2002

ROLE OF VEGF IN Myeloma

MM PC

BONE DISEASE

Very debilitating aspect of multiple myeloma due to painDangerous due to pathological fractures

Increased in bone resorption

(osteoclast activity)

Reduction in new bone formation(osteoblast activity)

The development of lytic lesion is related to an uncoupled bone remodelling:

Scheletal involvment is observed in approximately 80% of newly diagnosed multiple myeloma patients

Bone Physiology

TNFTNF, IL-1,, IL-1,IL-6, PGEIL-6, PGE22,,

IFNIFN, RANKL, RANKL

Bone FormationPTH, Vit. DPTH, Vit. D33,,Vit. K, IL-11,Vit. K, IL-11,estrogeni estrogeni

Bone Resorption

OSTEOBLASTOSTEOBLAST OSTEOCLASTOSTEOCLAST

Bone FormationPTH, Vit. DPTH, Vit. D33,,Vit. K, IL-11,Vit. K, IL-11,estrogeni estrogeni

OSTEOBLASTOSTEOBLAST

TNFTNF, IL-1,, IL-1,IL-6, PGEIL-6, PGE22,,

IFNIFN, RANKL, RANKL

Bone Resorption

OSTEOCLASTOSTEOCLAST

Bone in Multiple Myeloma

Osteoclast

RANKL

RANK

HBMSC

Myeloma Cells

+ve+ve

B. Myeloma

OPG

MICROEVIRONMENT AND MM(role of RANK/RANKL)

RANK is found on the surface of osteoclast precursors

OPG is produced by osteoblasts and can antagonize RANKL activity

RANKL is expressed on marrow stromal cells and osteoblasts

RANKL binds to RANK Inducing osteoclast formation

Myeloma cells- downregulate expression of OPG - upregulate expression of RANKL

Myeloma cells produce DKK-1 that antagonize the Wnt signaling, vital for osteoblast differentiation

Terpos E. et al, Leukemia 2007

DRUGS ACTIVITY ON BONE

CONCLUSIONS

Treatment paradigm in MM: concurrent targeting of both tumor cells and bone marrow

microenvironment to overcome drug resistance

Despite the absence of a peculiar molecular signature, malignant plasma cells display many genomic alterations usefull to divide myeloma patients in several groups

Genomic alterations are similar in MGUS and MM, highlighting the essential role of microenvironment in disease maintenance and progression

Several genetic alteration, cytokine overespression or microenvironment alteration can be used as a target for anti myeloma therapy, HOWEVER…

IMiDs in myeloma treatment

Richardson P. et al, JCO 2004

modified from Armand et al., Oncologist 2007

Triggers MM cell apoptosis by the

accumulation of improperly folded

proteins

Antiangiogenic effects:reduce cell adesion and IL-6 production

Increase Immunity:disrupts tumor-DC interaction

and enhances DC-mediated immunity

Reduce growth factors:Decrease cytokine levels

Bortezomib in myeloma treatmentPROTEASOME

INHIBITOR

Bone remodelling:Reduce resorptionIncrease formation