Biology of Driver OncogenesEGFR, EML4-ALK, ROS1…

Tetsuya MitsudomiKinki University Faculty of Medicine

Perspectives in Lung Cancer5th Asian Pacific Conference

24-25 August, 2012, Hong Kong

Interchromosomal translocation

Intrachromosomal translocation

LOH

Copy number gain

Copy number loss

一 single nucleotide variation

51 y/o male caucacian with smoking hxp/d adenocarcinomaKRAS mutation + 391 somatic mutation43 large-scale structural variants17.7 mutation/Mb

First adenocarcinoma genome

Driver and Passenger

Passenger gene mutation

Neutral genetic changes that are unrelated to cancer development caused by

exposure to mutagens, or genetic instability or many

mitoses

Driver gene mutation

Gene mutation essential for cancer cell development, growth

or survival

Tumor heterogeneity in Patient 1

Of all somatic mutations found on sequencing, 63 to 69% were heterogeneous

Intratumor heterogeneity Geringer et al., NEJM 2012

Phylogenetic relationships of tumor regions

Driver gene

Intratumor heterogeneity Geringer et al., NEJM 2012

The phenomenon by which some cancers that contain multiple genetic and epigenetic abnormalities remain dependent on (addicted to) one or a few genes for both maintenance of the malignant phenotype and cell survival

Weinstein and Joe, Nature Rev Clin Oncol, 2006

siRNA mediated specific knockdown of the mutant EGFR allele in NSCLC cells

Sordella et al., Science 2004

Models of oncogene addictionTorti and Trusolino, 2011

Genetic streamlining Oncogenic shock

A common signaling cascade may underlie ‘‘addiction’’ to the Src, BCR-ABL, and EGF receptor oncogenes

Sharma et al., Cancer Cell 2006

Pro-apoptotic

Pro-survival

How can Driver gene / oncogene addiction be identified?

Search for mutations that occur exclusively with known mutations of addicted oncogenes

Search for oncogenes whose inhibition result in induction of apoptosissiRNA screenInhibitor screen

RNAi screening of the tyrosine kinome identifies therapeutic targets in AML

Tyner et al., Blood, 2008

CMK cell: JAK3 A572V

HMC1.1 cell: c-KIT (V560G)

We established a high-throughput platform to profile500 cell lines derived from diverse epithelial cancers for sensitivity to14 kinase inhibitors.

Profiling of 500 cell lines with a variety of selective kinase inhibitors

Erlotinib(EGFR)

PHA665752(MET)

CL-387,785(EGFR irrev.)

HKI-272(EGFR, HER2)

Sunitinib(PDGFR, FLT-2, c-Kit, VEGFR2)

AZ628(BRAF)

AZD0530(Src family)

Compound 14(Bcr-Abl, Src family)

MK-0457 (Aurola, FLT-3)

Sorafenib (BRAF, c-KIT, FLT-3, PDGFRb)

Imatinib (Bcr-Abl, c-KIT, PDGFR)

NVP-TAE684 (ALK)

PD-173074(FGFR)

PF-2341066(c-MET, ALK)

HER

BRAF

MET

Kris et al., ASCO 2011

Kris et al., ASCO 2011

EGFR

Phase III trial for patients selected by EGFR mutation (as of ASCO 2012)

Study Race TKI NPFS OS

TKI CTx HR TKI CTx HR

NEJ002 Japanese G 228

10.8 5.4

0.32(0.24– 0.44)

27.7 26.60.88

(0.63-1.24)

WJTOG3405 Japanese G 172 9.6 6.6 052

(038-072) 35.5 38.81.18

(0.77-1.83)

OPTIMAL Chinese E 154

13.7 4.6 0.16

(0.11-0.26) 22.7 28.91.04

(0.69-1.58)

EURTAC Caucasian E 17

3 9.7 5.2 0.37(0.25-0.54) 19.3 19.5

1.04(0.65-1.68)

Approx. % of patients with PD harboring sensitive EGFR mutation treated with EGFR-

TKI

0mo 3mo 6mo 9mo 12mo0

10

20

30

40

50

60

70

80

90

100

IPASS

NEJ

WJTOG

OPTIMAL

% o

f pati

ents

wit

h P

D

About 20-30% of the patients progressed before 6 months

Why is response heterogenous?

Mutation classX19del ≥ L858R > G719X >>>> X20 ins = T790M

Amount of T790M allele

FAS, NFkB

BIM

CRKL

Heterogeneity of driver?

PFS in patients treated with gefitinib in patients with activating EGFR mutation according to presence or absence of T790M as detected Scorpion–Arms

Maheswaran et al., NEJM 2008

Bivona, et al. Nature, 2011

BIM Expression in Treatment-Naive Cancers Predicts Responsiveness to Kinase Inhibitors

Faber et al., Cancer Discovery, 2011

A common BIM deletion polymorphism mediates intrinsic resistance and inferior responses to

tyrosine kinase inhibitors in cancerNg et al., Nature Med., 2012

Amplification of CRKL Induces Transformationand Epidermal Growth Factor Receptor

Inhibitor Resistance in Human Non–SmallCell Lung Cancers

Cheung et al., Cancer Discovery 2011

KRAS

KRAS Dependency in KRAS Mutant Lung Adenocarcinoma Cell Lines

Ras dependency indices (RDI) =relative cell densities following K-Ras shRNAs treatment

1

A Gene Expression Signature Associated with ‘‘K-Ras Addiction’’ Reveals Regulators of EMT and Tumor Cell Survival. Singh et al., Cancer Cell 15: 489-500, 2009

KRAS dependent

KRAS independent

KRAS 変異のある肺癌はかならずしも KRAS 経路に addict していない

Synthetic lethality by siRNA library in KRAS mutated cell linesSTK33　　　Scholl et al., Cell

Serine threonine kinaseｍTOR downstream?Non-oncogene

PLK-1 Luo et al., Cell, 2009identified ~50 synthetic lethal genes from 74905 shRNAsAssociated with mitotic functionNon-oncogene

TBK1 Barbie et al., Nature 2009Suppresses IkB, activates NFkBNFkB is associated with cytokine production, proliferation,

survival , angiogenesis, and invasionCdk 4 Puyol et al., Cancer Cell, 2010

Ablation of Cdk4 results in immediate senescenceLung cancer specific

HER2

Incidence of HER 2 mutations in lung cancer (compilation of the 14 recent reports)

Tomizawa et al., Lung Cancer 2011

Overall Ethnicity Sex Smoking Histology0

1

2

3

4

2 2.1

3.63.9

2.7

1.5

1 1

0.3

N=3248

Asian

Cauca

sian

Fem

ale

Male

Never

Ever

Aden

o

Other

Minami et al., Oncogene 2007

Adenocarcinoma with HER2 mutation (ins 776 YVMA) treated by trastuzumab (anti HER2 Ab) plus vinorelbine

6/3/09 9/2/094/14/093/17/09

trastuzumab

vinorelbine

x5

Tomizawa et al., Lung Cancer 2011

MET

Met amplification in lung cancer

Investigator N Met amplification %

EGFR-TKI treated

Engelman 2007 Adeno 18 4 22.2

Bean 2007 Adeno 43 9 20.9

Suda 2010 Adeno(autopsy) 6 3 50.0

Sequist 2011 Adeno 37 2 5.4

EGFR-TKI untreated

Engelman 2007 Adeno 8 0 0.0

Bean 2007 Adeno 62 2 3.2 Beau-Faller

2008 Adeno 49 10 20.4

Cappuzzo 2008 NSCLC 166 12 7.2

Okuda 2008 Adeno 136 6 4.4

Kubo 2008 Adeno 75 2 2.7

Onozato 2009 Adeno 148 2 1.4

Cappuzzo 2009 NSCLC 435 18 4.1

Molecular predictors of sensitivity to the MET inhibitor (PHA665752) in lung carcinoma cells

Matsubara et al., JTO, 2010

Cell lines sensitivity IC50

Met amp

EGFR amp

HER2

amp

Met mut

EGFR mut

Her2 mut

KRAS mut

sens

itivi

ty

Activity of crizotinib in a NSCLC patients with de novo MET amplification

Ou et al., JTO, 2011

MET FISH

ALK

Waterfall Plot in Profile 1001**

** 早期死亡および未評価の患者を除く (n=106)*** 早期死亡および評価期間不足のため腫瘍評価が得られていなかった症例を含む (n=116)

Objective response details(all evaluable patients)

N=116**

ORR (95% CI) 61% (52, 70)

Median response duration 48 weeks

Median time to response 8 weeks

Disease control rate at 8, 16 weeks

79%, 67%

BOR CR PR SD PD

Bes

t %C

hang

e in

Tar

get L

esio

ns fr

om B

asel

ine

-100.0

-80.0

-60.0

-40.0

-20.0

0.0

20.0

40.0

60.0

80.0

100.0

* ** * * * * * * * * * * *

* : Japanese patient

N N=116***

RR (95% CI) 61% (52 ～ 70)

Median of respose 48w

Median until response 8 w

Progression-Free Survival (N=119)

Censored

95% Hall-Wellner Band

1.0

0.8

0.6

0.4

0.2

0Su

rviv

al d

istr

ibu

tio

n f

un

ctio

n

0 5 10 15 20

Months119 73 29 8 1n at risk

Median PFS = 10.0 months (95% CI: 8.2, 14.7)50 events (42%; 40 PD events) 69 patients (58%) censored, 59/69 (86%) in follow-up for PFS

EGFR and ALK lung cancer

EGFR ALK

discovery 2004 2007

Clinical backgroundNon-

smoker、 female,ad,Asian

Non-smoker, young

histologyReplacemt growth,

non-mucinousAcinar,cribriform,

signet

Incidence in Ad 40-50% 5-8%

Mechanism of acativation

Deletion~point mutation tramnslocation

TKI gefitinib/ erlotinib crizotinib

RR 60-80% 50-60%

MST 9~10~12 mo 10 month

m

ROS1

2

2

2

3

2

1

Variants of ROS fusionTakeuchi et al., Nature Med 2012

Key results: summary of tumour responses in patients with advanced ROS1+ NSCLC (n=14*)

† ‡

PD SD PR CR100

80

60

40

20

0

–20

–40

–60

–80

–100

16+

15+

18+4+ 12+

8+22+ 18 44+

20+35+ 48+

*Response-evaluable population†Tumour ROS1 FISH-positive, but negative for ROS1 fusion gene expression‡Crizotinib held for >6 wks prior to first scans which showed PD+, treatment ongoing

Dec

reas

e o

r in

crea

se f

rom

bas

elin

e (%

) Shaw et al. J Clin Oncol 30, 2012 (suppl; abstr 7508)

Response rate 8/14 = 57 ％

RET

Variants of RET fusion

KIF5B RET Ju Kohno Takeuchi

Lipsin Total

K15 R12 1 3 8 8 20

K16 R12 1 1 1 3 6

K22 R12 1 1 2

K23 R12 1 1 1 3

K24 R11 1 1

K15 R11 1 1

K24 R8 1 1

CCDC6 R12 2 2

Takeuchi et al., Nat Med., 2012

RET lung cancer is sensitive to RET TKI

Lipson et al., 2012

Kohno et al, 2012

Lung cancer and receptor tyrosine kinases

mutationtranslocationamplification

Acquired resistance

獲得耐性

مقاومة اكتسبت

획득 내성

获得性耐药

獲得性耐藥 kháng lại

רכשה ההתנגדות

oporność nabyta

erworbene Resistenz

hankitun resistenssin

erhvervet resistens

resistencia adquirida

ความต้�านทานทได้�มาεπίκτητης αντοχής

verworven resistentie

resistenza acquisita

приобретенной устойчивости

acquiritur resistentia

résistance acquise

Acquired resistance to EGFR-TKI therapy

A:　 secondary alteration of the target gene

YYT790M

D761YL747ST854A

EGFR-TKI

Kobayashi, NEJM 2005, Pao , PLoS Med 2005Balak, CCR 2006Costa, JCO 2008Bean, CCR 2008

Y

B:　 activation of other kinases

Y YYYMETamp

Engelman et al., Science 2007Bean et al., PNAS 2007

ATP

IGF1?, HGF?

Mechanisms of acquired resistance

Unknown Oncogene/ ALK-

9%EGFR mut/ALK-9%

KRAS mut/ALK-9%

KRAS mut/

ALK+*9%

ALK CNG9%

ALK Mutation/CNG9%

ALK Mutation27%

ALK+/Unknown Mechanism*

18%

ResistanceMutation

ALK Copy NumberGain (CNG)

Doebele et al., Clin Can Res 2012

to crizotinib in ALK+ NSCLC

AlternateOncogene

N=37

to EGFR-TKI in EGFR+ NSCLC

Sequist et al., Sci Translational Med., 2012

Summary…Driver gene mutations in lung cancer

All the driver genes so far identified that addict lung cancer cells are receptor tyrosine kinases activated by mutation (EGFR, HER2, DDR2), amplification (MET, FGFR1) or translocation (ALK, ROS1, RET)

Pharmacologic inhibition of driver gene product typically achieve ~60-70% response rate and ~10 month PFS

There is heterogeneity in response even within patients having driver gene mutation caused by additional alterations

Acquired resistance is inevitable, and its overcoming is of utmost importance.