Targeting KRAS and down streaming pathways Federico Cappuzzo Istituto Toscano Tumori Ospedale Civile Livorno-Italy

Targeting KRAS and down streaming pathways

Federico CappuzzoIstituto Toscano Tumori

Ospedale CivileLivorno-Italy

Istituto Toscano Tumori – Livorno, Italy

Molecular events in lung cancer

Adenocarcinoma

Unknown53.8%

KRAS27%

EGFR9.5%

EGFR resistance mutations0.8%

HER20.9%

BRAF1.7%

PI3K2.6%ALK

3.7%

Squamous-cell carcinoma

KRAS mutations in NSCLC

• KRAS is the most frequently mutated oncogene in NSCLC outside Asia. Mutations in ~20% of tumors

• KRAS mutations occur in lung adenocarcinoma and less frequently in the squamous cell carcinoma subtype

• KRAS mutations are usually associated with a history of tobacco use (transversion mutations), even if have been detected in 10% of never smoker (transition mutations)

• The vast majority of KRAS mutations involves codons 12 or 13


Questions on KRAS mutations in lung cancer

• Are KRAS mutations prognostic?• Are KRAS mutations predictive for chemotherapy

sensitivity?• Are KRAS mutations predictive for targeted therapy

sensitivity?• Has KRAS any influence on sensitivity to checkpoint

inhibitors?• Any drug really working?


KRAS mutations and NSCLC prognosis


KRAS mutations and prognosis• Over 50 studies published• Different methods for detection (IHC versus PCR)• Conflicting results

Reference N Mutated (%) p value for OS

Cappelletti 258 27 0.7

Grossi 249 19 0.07

Kern 44 36 0.16

Ma 718 14 0.31

Tsao 450 26 0.4

Mascaux* 3779 18 0.01

*2005 meta-analysis


No prognostic effect of KRAS mutations in the LACE-Bio pooled analysis


Shepherd et al, JCO 2013

All Patients Adenocarcinoma Patients

Pooled analysis of prognostic and predictive effect of KRAS mut in patients treated with

EGFR-TKIs


Zer et al, ESMO 2014

Prognostic effect of KRAS mutationin placebo arm only

9

All patients

Median OS 4.7 v 5.0 m HR 1.095, p=0.48

Resected Adenocarcinoma

Advanced adenocarcinoma

Median OS 4.7 v 4.4 m HR 0.91, p=0.63

Median OS 5.2 v 5.1 m HR 1.0, p=0.98

Time in months



Prognostic Effect of KRAS Codon 12 Mutation Subtype

10

mOS 6.3 mo(n=75)

mOS 3.9 mo(n=12)

mOS 1.8 mo(n=21)

p=0.01

Pe

rce

nta

ge

0

20

40

60

80

100

Time (Months) # At Risk(G12C or G12V) # At Risk(G12D or G12S) # At Risk(G12A or G12R)

0 752112

2 5788

4 4564

6 2130

8 1710

101310

SUMMARY STATISTICS:Stratified Log Rank test: p=0.0105Median for G12C or G12V: 6.28 -95% C.I. (4.08 ,9.80 )Median for G12D or G12S: 1.77 -95% C.I. (1.28 ,6.31 )Median for G12A or G12R: 3.88 -95% C.I. (1.05 ,4.54 )Stratified Hazard Ratio of G12D or G12S/G12C or G12V: 2.734 - 95 % C.I. (1.498, 4.993)Stratified Hazard Ratio of G12A or G12R/G12C or G12V: 1.818 - 95 % C.I. (0.846, 3.909)

KM Curve for Overall SurvivalKRAS mutation genotype - Placebo and stratified by trials

G12C or G12VG12D or G12SG12A or G12R



KRAS mutations as predictive factors for survival with adjuvant chemotherapy


KRAS mutations are not predictive for adjuvant CT in the LACE-Bio pooled analysis



Patients with KRAS mut Patients with KRAS wild-type

Interaction p value=0.37

Adjuvant CT potentially detrimental in codon 13 KRAS mutations: LACE-Bio pooled analysis



Patients withcodon 12 KRAS mut Patients with codon 13 KRAS mut

KRAS mutations as predictive factors for survival to EGFR-TKIs


Predictive Effect of KRAS Mutation in patients treated with EGFR-TKIs

15Istituto Toscano Tumori – Livorno, Italy


KRAS Mutant KRAS WT

Predictive effect of KRAS mutations All Patients

HR=1.13, p=0.395

HR=0.91, p=0.18

Interaction p=0.17



Predictive effect of KRAS mutation Adenocarcinoma EGFRWT

KRAS Mutant KRAS WT

Perc

enta

ge

0

20

40

60

80

100

Time (Months) # At Risk(Placebo)

# At Risk(EGFR_TKI)

0 106198

2 80

155

4 54

125

6 3992

8 2974

102157

SUMMARY STATISTICS:Stratified Log Rank test: p=0.1904Median for Placebo: 4.30 -95% C.I. (3.02 ,5.91 )Median for EGFR_TKI: 5.36 -95% C.I. (4.60 ,7.52 )Stratified Hazard Ratio of EGFR_TKI/Placebo: 0.838 - 95 % C.I. (0.643, 1.092)

KM Curve for Overall Survival (Adenocarcinoma, EGFR WT) - excluding BR19KRAS mutation status - Wild type

Placebo EGFR_TKI

Perc

enta

ge

0

20

40

60

80

100


# At Risk(EGFR_TKI)

0 3986

2 2869

4 2249

6 1437

8 1225

109

18


KM Curve for Overall Survival (Adenocarcinoma, EGFR WT) - excluding BR19KRAS mutation status - Mutated

Placebo EGFR_TKI

HR=1.04, p=0.84

HR=0.83, p=0.19

Interaction p=0.31



Predictive effect of KRAS mutation, Codon 12 subtype in advanced adenocarcinoma

G12D, G12S, G12A, G12R

Median OS 3.3 v 6.4m HR 0.43, p=0.08

Interaction p=0.003

Perc

entag

e

0

20

40

60

80

100


# At Risk(EGFR_TKI)

0 3058

2 2445

4 2230

6 1621

8 1513

101110

SUMMARY STATISTICS:Stratified Log Rank test: p=0.0438Median for Placebo: 8.28 -95% C.I. (4.08 ,13.14 )Median for EGFR_TKI: 4.32 -95% C.I. (3.32 ,6.08 )Stratified Hazard Ratio of EGFR_TKI/Placebo: 1.644 - 95 % C.I. ( 1.01, 2.677)

KM Curve for Overall Survival (Adenocarcinoma) - excluding BR19KRAS codon 12 - G12C/G12V Mutated

Placebo EGFR_TKI

G12C or G12V

Median OS 8.2 v 4.3 m HR 1.64, p=0.04

Perc

enta

ge

0

20

40

60

80

100


# At Risk(EGFR_TKI)

0 1222

2 7

20

4 3

17

6 2

12

8 19

1016


KM Curve for Overall Survival (Adenocarcinoma) - excluding BR19KRAS codon 12 - G12D/G12S/G12A/G12R Mutated

Placebo EGFR_TKI



(n=350)Placebo

(N=973) Randomizationstratified by:

histology, stage, prior adjuvant chemo, EGFR FISH status, smoking

status, country

(n=623)Erlotinib

150mg/day

Up to 4 cycles ofplatinum-based

doublet

Tumor samplesEGFR IHC+ and/or EGFR FISH+

90 d

180 d

RADIANT trial design

• Radiology assessment: every 3 months on treatment and yearly during long-term follow up

• Primary endpoint: DFS• Secondary endpoints: Overall survival (OS); DFS and OS in patients with del19/L858R (EGFR M+)

No adjuvant chemotherapy

2:1

StageIB–IIIA NSCLC

Complete surgical

resection

2-yr treatment period


Kelly K, ASCO 2014

RADIANT: DFS and OS in the whole study population


DFS OS

ErlotinibErlotinib

Placebo

Placebo

Kelly K, ASCO 2014

RADIANT Trial: Prognostic Effect of KRAS:Placebo Arm, All patients

DFS OS

0 6 12 18 24 30 36 42 48 54 60 660.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0

Disease-free Survival (Months)

Dis

ease

-fre

e S

urv

ival

Pro

bab

ility

KRAS M+ WT

0 6 12 18 24 30 36 42 48 54 60 660.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0


Dis

ease

-fre

e S

urv

ival

Pro

bab

ility

KRAS M+ WT

HR=1.14 (95% CI: 0.72, 1.81) HR=1.19 (95% CI: 0.67, 2.12)


Shepherd et al, ESMO 2014

Predictive Value of KRASon DFS: All Randomized

22

0 6 12 18 24 30 36 42 48 54 60 660.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0


Dis

ease-f

ree S

urv

ival P

rob

ab

ilit

y

Erlotinib Placebo

0 6 12 18 24 30 36 42 48 54 60 660.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0


Dis

ease-f

ree S

urv

ival P

robabili

ty

Erlotinib Placebo

Interaction HR: 0.87; P=0.64

KRAS M+ KRAS WT

HR: 0.81 (95% CI: 0.48, 1.38) HR: 0.93 (95% CI: 0.73, 1.19)



23

Predictive Value of KRASon OS: All Randomized

0 6 12 18 24 30 36 42 48 54 60 660.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0


Dis

ease

-fre

e S

urv

ival

Pro

bab

ility

PlaceboErlotinib

0 6 12 18 24 30 36 42 48 54 60 660.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0


Dis

ease

-fre

e S

urv

ival

Pro

bab

ility

Erlotinib Placebo

Interaction HR: 0.90; P=0.78

KRAS M+ KRAS WT

HR: 1.05 (95% CI: 0.56, 1.99) HR: 1.15 (95% CI: 0.85, 1.55)



Can we use KRAS testing for precluding an EGFR-TKI to pretreated NSCLC?


KRAS mutations are predictive for lack of response to EGFR-TKIs

Reference N % Mutated RR (%)

Massarelli 70 22.8 0

Miller 80 22.5 0

BR21 118 16.9 5.0

Eberhadt 264 21.0 8.0

Zer• Codon 12• Codon 13

79685

19.3 1.31.50


Gefitinib versus docetaxelOverall survival by biomarkers

0.5 1.0 1.5 2.0HR (gefitinib vs docetaxel) and 95% CI

Overall

EGFR FISH +

EGFR FISH -

EGFR expression +

EGFR expression -

EGFR mutation +

EGFR mutation -

K-RAS mutation +

K-RAS mutation -

0

Favors gefitinib Favors docetaxel


TAILOR Study Design

ERLOTINIB 150 mg po, daily

DOCETAXEL75 mg/m2 iv day 1,22 OR 35 mg/m2 iv day 1,8,15,29

R 1:1

CRO

SS O

VER

N

OT

ALLO

WED

Garassino MC, et al Lancet Oncol 2013

N 222

EGFR wild-type


Prognostic impact of KRASUnivariate analysis

PFS OS

Garassino MC, et al ESMO 2013


HR 1.02, p=0.93 HR 1.28, p=0.14

Predictive impact of KRAS on OS

KRAS mutants KRAS wild-type

Test for interaction Univariate p=0.82; Multivariate =0.97

HR 0.81 (95%CI 0.45-1.47) p=0.492 HR 0.79 (95%CI 0.57-1.10) p=0.167

Garassino MC, et al ESMO 2013


HR 0.81, p=0.492 HR 0.79, p=0.167

Can we use KRAS testing for selecting patients candidate for checkpoint inhibitors?


Clinical Development of Inhibitors of PD-1 Immune Checkpoint

Target Agent Molecule Company Development

PD-1 Nivolumab-BMS-936558

Fully human IgG4 mAb Bristol-Myers Squibb Phase II, III multiple tumors

PidilizumabCT-011

Humanized IgG1 mAb CureTech Phase II multiple tumors

LambrolizumabMK-3475

Humanized IgG4 mAb Merck Phase I-II

AMP-224 Recombinant PD-L2-Fc fusion protein

GlaxoSmithKline Phase I

PD-L1 BMS-936559 Fully human IgG4 mAb Bristol-Myers Squibb Phase I

MedI-4736 Engineered human IgG1 mAb

MedImmune Phase I

MPDL-3280A Engineered human IgG1 mAb

Genentech Phase I-II


a ORR includes investigator-assessed unconfirmed and confirmed PR. Patients first dosed at 1-20 mg/kg by Oct 1, 2012. Data cutoff: Apr 30, 2013.

Patie

nts

With

PR,

%

Response by Smoking Status (ORRa)

n = 43n = 10

MPDL3280A Phase Ia: Response by Smoking Status - NSCLC

Former/current smokers

Never smokers

Patient Smoking Status (n = 53)

Soria et al et al. ESMO 2013


Activity of pembrolizumab according to clinical characteristics

N ORR (%)

Total 236 21

Previous treatment 236

• Treatment naive 42 26

• Previuos treated 194 20

Histology 230

• Non-squamous 191 23

• Squamous 39 18

Smoking history 230

• Current/Former 165 27

• Never 65 9

Garon et al et al. ESMO 2014


Nivolumab more effective in smokers

Variable ORR, % (n/N) [95% CI] P-value

Smoking exposure 0.018

≤5 pack-yrs 0 (0/14) [0, 23]

>5 pack-yrs 30 (20/66) [20, 43]

Time since quitting 0.22

Current smoker 27 (6/22) [11, 50]

1–5 yrs prior 46 (6/13) [19, 75]

6–15 yrs prior 17 (2/12) [2, 48]

>15 yrs prior 18 (6/33) [7, 36]

0

20

80

60

40

Months Since Treatment Initiation

100

PFS

(%)

PFS by smoking exposure

Never/minimal smokers (mPFS 1.7 months)Former/current smokers (mPFS 2.2 months)HR (95% CI) = 0.41 (0.22, 0.74), P = 0.003

mPFS = median Progression-free survival


Hellmann et al et al. ASCO 2014


Altered protein contain new epitopes for immune recognition, providing a common denominator for immunotherapy

High mutational rates may contribute to increased immunogenicity

Response to anti-PD-1 is independent of EGFR or KRAS mutation status: nivolumab as an example

Subgroup ORR, % (n/N) [95% CI]

EGFR status

Mutant 17 (2/12) [2.1, 48.4]

Wild-type 20 (11/56) [10.2, 32.4]

Unknown 15 (9/61) [7.0, 26.2]

KRAS status

Mutant 14 (3/21) [3.0, 36.3]

Wild-type 25 (9/36) [12.1, 42.2]

Unknown 14 (10/72) [6.9, 24.1]

Ch

ang

e in

tu

mo

ur

size

, %

-100

-80-60

-40

120

-20

0

20

40

60

80

100

Patients

EGFR mutation statusMutantUnknownWild-type

-100

-80

-60

-40

120

-200

20

40

60

80

100

Patients

Ch

ang

e in

tu

mo

ur

size

, %

KRAS mutation statusMutantUnknownWild-type

CA209-003: phase 1 follow-up study, up to 5 prior lines of therapy, NSCLC cohort

Brahmer J, et al. ASCO 2014 (Abstract 8112).


Response to anti-PD-1 is independent of EGFR or KRAS mutations: pembrolizumab and MPDL 3280A

Subgroup N ORR (%)

Total 53 23

EGFR status 46

• Mutant 6 17

• Wild type 40 23

KRAS status 37

• Mutant 10 10

• Wild type 27 30

MPDL 3280A PembroSubgroup N ORR (%)

Total 236 21

EGFR status

• Mutant 36 14

• Wild type 214 -

KRAS status

• Mutant 39 28

• Wild type 117 -


Garon et al et al. ESMO 2014Soria et al et al. ESMO 2013

There is any drug really working in KRAS mutated NSCLC?



Selumetinib

• Selumetinib (AZD6244, ARRY-142886) is a potent and selective allosteric inhibitor of MEK 1/21

• Tendency for greater sensitivity to selumetinib in BRAF/RAS-mutant cell lines2

1. Yeh et al. Clin Cancer Res 2007;13:1576–83; 2. Davies et al. Mol Cancer Ther 2007;6:2209–19

Ras

Raf

MEK 1/2

ERK 1/2

SelumetinibCe

ll vi

abili

ty in

hibi

tion

IC50

(µM

)

Cell line

RAS/BRAF mutation

0

5

10

15

20

25

30

NoneKRASBRAFNRAS

Selumetinib: preclinical/early phase clinical studies Preclinically, the combination of selumetinib and docetaxel demonstrated tumor regression in a KRAS-mutant cancer1

Phase I study showed that selumetinib plus docetaxel had a manageable tolerability profile2

1. Holt et al. Br J Cancer 2012; 2 Kim et al. Mol Can Ther 2011;

HCT-116 human tumor xenografts (KRAS-mutant)

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

0 2 4 6 8 10 12 14

Mea

n tu

mou

r vol

ume

(cm

3) +

/-s.

e.m

ControlSelumetinib-25mg kg-1 per qdDocetaxel-15mg kg-1Combo



Phase II, double-blind, randomized, placebo-controlled, multi-center trial; NCT00890825

• Docetaxel was administered every 21 days; selumetinib/placebo administered daily• Following completion of patient enrollment, the primary endpoint was changed from PFS to OS, without

changing the sample size‡ – OS analysis was planned for after approximately 58 events; HR 0.57, 80% power assuming a 1-sided

10% significance level

Selumetinib 75 mg BID+ docetaxel 75 mg/m2

Placebo BID + docetaxel 75 mg/m2

EndpointsPrimary• OS

Secondary• PFS• ORR• Duration of response• Change in tumor size• Alive and progression-

free at 6 months• Safety and tolerability

Randomization(1:1 ratio)

Patients

• Locally advanced or metastatic NSCLC (stage IIIB-IV)

• Failed first-line therapy

• Confirmed KRAS mutant tumor*

• WHO PS 0-1

• Excluding symptomatic brain metastases

Janne PA et al, Lancet Oncol 2014

Overall survival

• There was a numerical increase in OS (median follow-up 7.2 mo); hazards non-proportional– 56/83 deaths (67% maturity): selumetinib + docetaxel 29/43, placebo + docetaxel 27/40

Median OSSelumetinib + docetaxel, n=43 9.4 moPlacebo + docetaxel, n=40 5.2 moHR 0.80; 80% CI 0.56, 1.14; 1-sided p=0.2069*

Prop

ortio

n of

pati

ents

aliv

e

Days

0 50 100 150 200 250 300 350 400 450 500 550 600 650

Number at risk

1.0

0.8

0.6

0.4

0.2

0.0

4340

4135

3627

3520

2818

2515

1413

66

65

24

13 2 1


Janne PA et al, Lancet Oncol 2014

Trametinib and pemetrexed for KRAS mutated NSCLC


Mazieres J et al. WCLC 2013

Tivantinib: Study DesignRandomized, placebo-controlled, double-blind clinical trial

RANDOMIZE

Erlotinib 150 mg PO QD+ Placebo

28-day cycle

Erlotinib 150 mg PO QD+ ARQ 197 360 mg PO BID

28-day cycle

Endpoints • 1° PFS• 2° ORR, OS• Subset analyses• Crossover: ORR

NSCLC• Inoperable locally adv/

metastatic dz.• ≥1 prior chemo

(no prior EGFR TKI)

• 33 sites in 6 countries

• Study accrual over 11 months (10/08-9/09)

• Randomization stratified by prognostic factors incl. sex, age, smoking, histology, performance status, prior therapy and best response, and geography (U.S. vs. ex-U.S.)

PD


5.02.0

Tivantinib: PFS in Histologic and Molecular Subgroups

ARQ197/erlotinib Placebo/erlotinib

Unadjusted HR (95% CI)N Median PFS (95% CI, weeks)

Squamous Cell 26 / 24 13.7 (8.0‒18.1) 8.4 (7.9‒21.0)

Non-Squamous Cell 58 / 59 18.9 (15.0‒31.1) 9.7 (8.0‒16.0)

c-MET FISH >4 19 / 18 15.4 (8.1‒24.4) 15.3 (7.1‒16.3)

c-MET FISH >5 8 / 11 24.1 (16.3‒NE) 15.6 (7.9‒31.4)

EGFR mutant 6 / 11 24.1 (8.0‒32.1) 21.0 (8.1‒36.0)

EGFR wt 51 / 48 13.7 (8.1‒18.1) 8.1 (7.9‒9.9)

KRAS mutant 10 / 5 9.7 (7.9‒NE) 4.3 (1.1‒8.0)

KRAS wt 49 / 45 15.4 (8.1‒18.1) 9.9 (8.0‒16.0)

1.00Favors

ARQ 197/ErlotinibFavors

Erlotinib/placebo

HR=0.70

HR=0.18

0.5 1.5

HR=1.01

HR=1.23

HR=0.45

HR=0.71

HR=1.05

HR=0.71


MARQUEE phase III study design

Primary end-point: OSIstituto Toscano Tumori – Livorno, Italy

MARQUEE: Forest plot for OS in key subgroups


Conclusions• KRAS mutations are not prognostic in early nor advanced

NSCLC• KRAS mutations are not predictive for OS in:

– Early NSCLC exposed to adjuvant chemotherapy– Early NSCLC exposed to EGFR-TKIs– Advanced NSCLC exposed to EGFR-TKIs

• KRAS mutations predict no response to EGFR-TKIs• KRAS mutations cannot be used for precluding a treatment

with– Adjuvant Chemotherapy– EGFR-TKIs in second or subsequent lines– In patients candidate for checkpoint inhibitors

• No drugs are currently available in clinical practice• Several compounds are under investigation


Documents

Targeting KRAS and down streaming pathways Federico Cappuzzo Istituto Toscano Tumori Ospedale Civile Livorno-Italy