The MET oncogene:

From molecular biology to Clinical TrialsPaolo M. Comoglio, MD

Institute for Cancer Research @ Candiolo,University of Turin, Medical School, Italy

MET IS A ‘MASTER GENE’ IN THE CONTROL OF INVASIVE GROWTH

S-SKringles Protease-

like

Sema domain

G-P richIg like domains

500 AA

400 AA

Tyrosine Kinase

Met(HGF Receptor)

PP

PP

MRS (PSI)

HGF (Scatter Factor)

P

GAB1

P

P

GRB2RAS

P

P

CD44v6

MET

Plexin B1

Integrin 6 4

SHP2

SRC

Sustained MAP and PI3 kinase activation (invasion / apoptosis protection)

GRB2

Transient MAP kinaseactivation (proliferation)

Link to cytoskeleton

PSTAT

PLC

PI3Kp85

PI3Kp85

CRKL

C.Boccaccio and PM.Comoglio. Nature Rev. Cancer.2006, 6: 637-645.

Cell-Polarity and Morphogenesis

Xenopatient

Xenosphere(cancer stem cells)

Human metastaticcolorectal cancer

Spheropatient

MET IS A FUNCTIONAL MARKER OF

CANCER STEM CELLS

Xenopatients, Xenospheres and Spheropatients

Xenopatient Xenosphere(cancer stem cells)

Spheropatient(Secondary xenopatient)

C. Boccaccio et al., in preparation

Mutations retained and passed onResponse to targeted drugs retained and passed on

MET inhibitionpromotes differentiation of CRC colospheres

Control

JNJ inhib.

Cetuximab

Fibroblast conditioned medium0% serum

ACTIVATION OF THE MET-DRIVEN ‘INVASIVE GROWTH’ PROGRAMME

GENERATES AN AGGRESSIVE PHENOTYPE

• ONCOGENE ‘EXPEDIENCE’• ONCOGENE ‘ADDICTION’

THE CONCEPT OF ‘ONCOGENE EXPEDIENCE ‘

• Met can promote invasion and metastasis independent of the genetic lesions that caused tumour onset

• Met activation is a ‘stress’ response to unfavourable microenvironment (hypoxia, radiations , inflammatory cytokines)

• Met activation occurs mainly through transcriptional upregulation

• Met activation is an expedience that provides a pro-survival and invasive advantage

Trusolino and Comoglio., Nature Rev. Drug Discov, 2008Nature Rev. Mol. Cell. Biol. 2010

The MET promoter

Proteasomedegradation

Cellular [O2]

HIF-1(constitutive)

Nucleus

HIF-1(regulated) -OH

pVHL

Cytoplasm

HIF Proline Hydroxylase

HIF-1

Hypoxia promotes invasive growthby transcriptional activation of the MET Oncogene

P (-2619) S (-345) S (+89)

HRE-1 HRE-2 asHRE-1

AP-1

HRE-4

STARTA (-253)

asHRE-2/HRE-3

S (-411) A (-32)

S (-295)

A (+353)

HRE-5

met mRNA

21%

O2

3% O

2

CoC

l 2

Con

trol

28S

MET

HIF-1a

Pennacchietti, Michieli et al. , Cancer Cell 3: 347 (2003)

Hypoxia promotes invasive growthby transcriptional activation of the MET Oncogene

• In a limited number of cancer types, genetic lesions of Met are selected along the tumour natural history to become the driving force that maintains the transformed phenotype

• Met activation is the consequence of a fixed and transmissible genetic alteration; therefore, it is characterized by chronic firing and high steady-state signalling

THE CONCEPT OF ‘ONCOGENE ADDICTION ‘

• Amplification (e.g. gastric ca, NSCLCs and CRC mts resistant to targeted therapy)

• Point mutations - hereditary and sporadic carcinomas (e.g. kidney, hepatocellular, head

& neck ca.- early metastatic cancers of unknown primary origin (CUPs)

‘Oncogenic addiction’ to MET in tumourscan be achieved by:

Trusolino and Comoglio, Nature Rev. Drug Discov. 2008

TARGET THERAPIES AGAINST MET

• An adjuvant approach for a vast number of tumours:Overexpression: targeting ‘expedience’(almost all solid tumors expressing MET)- Endpoints: PFS, OS

• A front-line intervention for a limited fraction ofgenotype-specific tumours:Amplification, mutations : targeting ‘addiction’ (e.g. gastric ca., NSCLC, gefitinib-resistant NSCLCs)- Endpoints: Remission

THERAPEUTIC INACTIVATION OF THE MET ONCOGENECAN BE ACHIEVED BY:

1. Small molecules inhibiting the Tyrosine Kinase

2. Monoclonal Antibodies

RESPONSE TO MET INHIBITORS CORRELATES WITH GENE AMPLIFICATION

PHA-665752PHA-665752

97 %EXPEDIENCE

5.2NCI-H1993

5.6SNU5

6.3HS746T

6.1GTL-16

6MKN-45

5.8EBC-1

MET copy N°Cell Line

Hypoxia

(or HGF)

< 3 %ADDICTION

200 cancer l. tested

Clinical trials for MET inhibitors

• Inhibition of growth (reduction of tumor mass) is expected only in cancers “addicted” to MET

• Genetic lesions are expected in ~ 3-4 % of epithelial cancers

• It is mandatory to identify patiens with cancers “addicted” to MET before recruitment

111In DTPA-DN30 tumor uptake at 2h post injectionSPEC-CT scan

EBC1A2780

Bardelli. et al. Cancer Discovery 2013

Detection of MET-amplificationby Liquid Biopsy

Xenopatient-derived cancer cells display MET amplification and are sensitive to Met targeted drugs

00,20,40,60,8

11,21,4

NT 10 50 500 10 50 500

cell

viab

ility

%

JNJ CRIZOTINIB

Therapy with MET antibody

MV-DN30 Monoclonal AntibodyMonovalent, Chimeric, Stabilized

• Recombinant, properly assembled and PEGylated• Bind Met (IP4) with high affinity (Kd= 0,116 nM)• Down-regulate the Met receptor from the cell surface• Induce shedding of the extracellular domain (“decoy”)

O

OHn

Adam 10

p55

p125

p50Metp175

Inactive receptor

heterodimerS -S

Ligand neutralization

p55

p125

p50Metp175

Inactive receptor

heterodimer

Ligand neutralization

Shedding

Γ- secretase

DecoyInhibitoryeffects

Proteasome degradation

Exogenous administration of anti-MET DN-30 inhibits tumor growth and prevent metastasis

Mab DN30

Human Breast Carcinoma, transplanted in Athymic nu/nu mice

Petrelli, A. et al., Proc. Natl.Acad Sci. US: 2006, 28, 5090-5095

Irrelevant Mab

0

200

400

600

800

1000

1200

1400

1600

1800

2000

2200

0 2 4 6 8 10 12 14 16 18 20 22 24 26 28

Days after injection

Tum

or v

olum

e (m

m3)

.

NTCDDP5FUMV DN30CDDP + MVDN305FU + MVDN30

CHEMOTHERAPY (CDDP, 5FU) vs MET ANTIBODY TREATMENT OF EBC1

- Xenotransplants-

Days after injection

Tum

or V

olum

e

MECHANISMS OF ACQUIRED RESISTANCETO MET KINASE INHIBITORS

• MET amplification

• Activating point mutations

Amplification of MET contributes to acquired resistance to MET kinase inhibitors

GTL16 wt

Chromosome 7 centromereMET amplicon marker

Wt PHA resistant (150 nM)

WT

1 2 3 4 5 6 7 8 9

a-pMet

a-Met

PHA 250 nM

Y1349

Y1356

PP

PP

PP S985

Y1003

D1228H (Kit)D1228N (Kit)Y1230CY1230HM250T (Ret)

M1131TV1188LL1196VV1220I

Some MET mutations confer resistance to MET inhibitors

EBC1 wt EBC1 PHA Resistant

FAb DN30 treatment overcomes resistanceto the MET kinase Inhibitor PHA-665752

Viab

ility

in 5

0 nM

PHA

EBC1Wild Type

EBC1PHA Resistant

DN30 Fab 0.4 mM

MET and Invasive Growth: State of the art:

1. Scatter factors and the receptors of the MET oncogene family (RON, ROR) are key regulators of the Invasive Growth program in cancer stem cells.

2. The program is activated by MET over expression, induced by unfavourable micro-environmental conditions, such as hypoxia or ionizing radiations: (“Oncogene Expedience”).

3. The invasive growth program is constitutively activated in some cancers, by MET amplification, mutations or autocrine loops (“Oncogene Addiction”).

4. The invasive growth program driven by the oncogene MET is a good target for therapy.

5. Resistance may occur, that may be overcome by combined treatment with s.m. inhibitors and antibodies

Plexins L.TamagnoneE. GiraudoG. Serini

Met AddictionL.TrusolinoA. BertottiF. Galimi

Met GeneticsA. BardelliM.F. Di Renzo

Met & HypoxiaP. MichieliS. Pennacchietti C. Basilico

Met SignatureE. MedicoMet BiologyL. Lanzetti

Met & Stem cellsC. BoccaccioF. De BaccoP. LuraghiG. Reato

Met and resistanceS. GiordanoS. CorsoA. PetrelliC. Migliore

IRCCExperimental Clinical Molecular Oncology

Met Gene TherapyE. VignaS. CignettoR. Albano

Ron & RORS. BenvenutiA. GentileL. LazzariA. Arnesano

IRCC: Institute for Cancer Research @ Candiolo

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