Next-Generation Sequencing is a Valuable

Tool in an Oncologist’s Practice -

ABSOLUTELY

Kimberly Blackwell MD

Professor of Medicine

Duke Cancer Institute

Miami Breast Meeting

March 8, 2014

Overview

Evolutionary Strategies Endocrine Therapy

HER2 Based Therapy

Next steps: Revolutionary? Identifying potential clinical trials

Next Generation Sequencing Trials\

Identifying Prognostic Mutations

Reason #1: Identifying Potential Trial

Candidates

HER2 mutation identified in HER2- breast cancer in

ACOSOG Z1031 by next generation

massively parallel sequencing

SURGERY

Postmenopausal • Clinical Stage II or III • ER+ (Allred 6-8) • HER2-

Exemestane

Letrozole

Anastrozole

R

BIOPSY

HER2 mutation Stage ER PR HER2 V777L IIB + + Negative

del.755-759 IIB + + Negative

25 Patients with HER2 Somatic

Mutations

• Each blue circle represents a patient.

• From 8 publications with a total of 1,499 patients.

• 20% of patients have mutations at amino acids 309 or 310.

• 68% of patients have mutations at amino acids 755-780.

Bose, R., et al., Cancer Discov, 2013. 3(2): p. 224-37.

A Phase II Trial of Neratinib in Metastatic

HER2 Non-amplified but HER2 Mutant Breast Cancer

CLIA Tumor HER2 Sequencing

HER2 Negative Metastatic Breast Cancer

*Neratinib 240 mg P.O. daily

days 1-28 each cycle #

** May continue neratinib at progression if trastuzumab is added in the most recent amendment

Continue therapy until

disease progression** or unacceptable toxicity

*Prophylactic loperamide

during 1st cycle. May

escalate to 320 mg in the

most recent amendment

# Each cycle is 28

days

HER2 Mutation

Negative

Not eligible for

study treatment

Screening Consent & Pre-registration

HER2 Mutation Positive

Consent to treatment & Registration

Tumor measurement every 2 cycles

Reason #2: Identifying Potential Compounds

that Have a Higher Likelihood of Benefitting

Patients (AKA the National Cancer Institute

thinks it is time!)

Treatment MBC according to genomic alterations SAFIR01

423 patients consented

CGH arrays: 287 patients

Biopsy of metastasis: 404 patients

194 targetable genomic

alterations 25%

treatment driven by

genomics in 48 patients

+ 4 patients ERBB2 amp CGH array

12%

Andre, F, et al. ASCO 2013

Therapies matched to the genomic alterations

Rapalogs

FGFR FGFR/VEGFR

EGFR/mTOR

AKT

TORC1/2

PI3K

16 regimens

CDK4 Raf

MET

MDM2

MGMT CHEK1

EGFR

FAK

Targets of the drugs

Efficacy data on 48 patients treated with therapy matched to genomic analysis

Efficacy n (%)

Objective response 4 (9%)

SD>16 weeks 8 (19%)

OR + SD>16 weeks 12 (28%)

Progression within 16 weeks 32 (72%)

Ongoing therapy SD <16 weeks 4

Erbb2 conversion (n=4) 1 OR

1 long term SD

(10 months)

Targets picked-up: EGFR amplification AKT gene alteration

FGF-amplified BC IGF1R amplification

12 out of 404 (3%) patients got some benefit from the biopsy procedure and genomic profiling

MATCH NCI Study

• Umbrella protocol multiple, single-arm phase II

• CTEP-IND for protocol template

• Initially focused on single-agents

– Combinations will be considered for targets that have validated combination targeted therapy

• Tumor biopsies & sequencing at progression to illuminate resistance mechanisms

• Screen 1500 to 3000 patients that progressed following standard therapy to enroll 500 – 1000

• Limit enrollment to 1000 eligible patients – Restrict ratio of common vs. uncommon histologies screened – Adjust screening ratio to achieve adequate representation of

rare tumors among those enrolled on treatment • 75% “common”: breast, NSCLC, colon, prostate, (lymphoma) • 25% “rare” tumors

SCHEMA

Study agent

Complete or partial

response (CR+PR)1

Actionable

mutation detected

No additional actionable mutations, or withdraw consent

Genetic sequencing

Progressive

disease (PD)1

Stable disease

(SD)1 for 6 months

Drug

holiday PD

Study agent

Off study

PD

Check for additional actionable mutations3,4

Stable disease or

better2

Continue on

study agent

until

progression

Continue on

study agent

until

progression

Course 1

Course 2

1CR, PR, SD, and PD as defined by RECIST

2Stable disease is assessed relative to tumor status at re-initiation of study agent 3

4Rebiopsy; if additional mutations, offer new targeted therapy

Reason #3: Defining Prognosis and

Potentially Actionable Mutations in Metastatic

Breast Cancer

Extraordinary Responders to lapatinib with trastuzumab

primary-refractory IBC share common cancer genotype

PIK3CA H1047R TP53 R273H

ERBB2 amplification TP53 E204

PIK3CA H1047R

ERBB2 amplification

TP53 Y205fs*3

CDKN2A loss

CDKN2B loss BRCA2 mutation

CDKN2A loss

CDKN2B loss PALB2 mutation

TBX3 splice

BLM R643H

PTEN C296fs*1

Patient 3 Patient 2 Patient 1

O’Shaughnessy, J. SABCS 2013

ERBB2 amplification

original core biopsy

4th patient with identical phenotype and genotype recently identified

Patients 1 and 2 with multple amplicons: FGFR1, MYST3, ZNF703, CDK6

All Rights Reserved, Duke Medicine 2011

ABC Metastatic TNBC Clinical Trial/ Tissue Procurement (Clinical Breast Cancer, 2013. 13:6, 416-20)

1. Metastatic

Biopsy

2. Germline DNA

3. Primary

Tumor Block

San Antonio Breast Cancer Symposium – Cancer Therapy and Research Center at UT

Health Science Center – December 10-14, 2013.

This presentation is the intellectual property of the author/presenter. Contact them at black034@mc.duke.edu for permission to reprint and/or distribute.

Total Population (38) Tissue Population (34)

Age 50 (30-76) 50 (30-76)

PFS (mo.) 9.2 (7.8-25.1) 14.6 (6.7-28.8)

OS (mo.) 21.6 21.9

All Rights Reserved, Duke Medicine 2011

Most Frequently Occurring Mutations

• 122 genes somatically mutated in >1 patient – Mean 10 mutations/sample (Range 0-42)

– 74 genes had mutations occurring ≥3 times.

San Antonio Breast Cancer Symposium – Cancer Therapy and Research Center at UT

Health Science Center – December 10-14, 2013.

This presentation is the intellectual property of the author/presenter. Contact them at black034@mc.duke.edu for permission to reprint and/or distribute.

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Identified as Potential Cancer Target

All Rights Reserved, Duke Medicine 2011

Genes With Mutations Associated with Progression Free Survival

Gene (# of Patients

Mutations)

Mutation (mo.) Wild Type (mo.) P-value

WNK1 (3) 1.4 7.3 0.03

TP53 (10) 4.7 8.2 0.04

JAK1 (3) 1.6 7.3 0.04

DCHS2 (5) 2.8 7.5 0.04

ATXN7 (3) 10.8 6.6 0.03

MST1 (3) 11.4 6.2 0.04

WNK1 TP53 JAK1 DCHS2

All Rights Reserved, Duke Medicine 2011

Genes with Mutations Associated with Disease Free Interval

Gene (# of

Mutations)

Mutation

(mo.)

No Mutation

(mo.)

P-value

HGF (3) 107.4 19.4 0.009

PLXNA3 (4) 37.9 26.0 0.02

CSDE1 (3) 39.6 25.4 0.02

ZNF710 (4) 86.2 19.3 0.03

CNN2 (3) 103.9 22.3 0.03

PAPLN (3) 53.8 23.6 0.03

SETBP1 (2) 81.2 24.0 0.04

MTOR (4) 59.3 22.8 0.05

This presentation is the intellectual property of the author/presenter. Contact them at black034@mc.duke.edu for permission to reprint and/or distribute.

All Rights Reserved, Duke Medicine 2011

Genes with Mutations Associated with Overall Survival

Gene (# of

Mutations)

Mutation (mo.) Wild Type (mo.) P-value

TP53 (10) 17.5 25.9 0.02

ITSN2 (3) 7.6 27.6 0.03

ALDH8A1 (2) 6.5 27.1 0.05

SPHKAP (2) 66.6 23.5 0.03

TP53 ITSN2 ALDH8A1 SPHKAP

This presentation is the intellectual property of the author/presenter. Contact them at black034@mc.duke.edu for permission to reprint and/or distribute.

All Rights Reserved, Duke Medicine 2011

Actionable Mutations

• Studied in Breast Cancer

– p53: Vaccine, Gene therapy, Wee-1 inhibitors, Kevetrin

– PARP: PARP Inhibitors

– ESR (ER) : Alternative Endocrine Therapies

– JAK1: JAK1 Inhibitors

– mTOR: mTOR Inhibitors

• Not Studied in Breast Cancer (yet)

– Dynein: hsp90 Inhibitors, HDAC inhibitors

– MST1: Anti-MST1 receptor (Ron) antibodies

– ROS-1: Inhibitors

– HGF: Antibodies against c-met; Inhibitors against c-met

– ALDH8A1: Disulfiram

Source: www.clinicaltrials.gov

This presentation is the intellectual property of the author/presenter. Contact them at black034@mc.duke.edu for permission to reprint and/or distribute.

There has been a revolution of Science but the

clinical application has lagged behind….next-gen

sequencing will catch the science up!

Where the Generator is

Busted:

Bcr-Abl CML (4,500)

C-KIT mutation (1,000)

EGFR Mutation (1-2,000

patients)

HER2 Driven Breast

Cancer (40,000 patients)

Everything Else Where

Several Sockets are

Busted (191 CAN-

genes): 1.4 Million

patients

Hanahan and Weinburg, Cell, 2000

Wood LD, et al. Science, 2007.

WNT

Cell

ECM

Growth factors (e.g. HRG, EGF, TGF a )

Nuclear receptors (e.g. estrogen)

Survival factors (e.g. IGF1)

Cytokines (e.g. ILs , IFNs )

Death factors

(e.g. FasL )

Anti - growth factors (e.g. TGF b )

GPCR ligands

Frizzled Dishevelled

GSK - 3 b

APC

Tubulin

TCF

Integrins

b - Catenin b - Catenin:TCF E - Cadherin CdC42 PI3K Rac

Fak Cas Crk Src

Fyn Shc

NF1

Ras RTK Grb2 SOS Ral MEK MAPK MAPK

MEKK

PLC

PKC Mos MKKs JNKs

ELK

Myc:Max

Max:Max Fos

JUN

Abl

7 - TMR

CdC42 Rac Rho

G - Prol Ad Cycl PKA CREB

PKC NF k B

NHR (e.g. ER)

NF k B

P13K Akt Akka IKB

PTEN ?

Stat 3.5

Stat 3.5

Stat 3.5

Bcl XL

Caspase 9

Cytochrome C

Jaks

Bad Bid Mitochondria

Bim , etc. Abnormal sensor

Bcl - 2

Cell Death (Apoptosis) Caspase 8

Fap

FADD Bcl - 2

Bax

ARF

p53

Mitochondria

MDM2

DNA damage sensor Cell

Proliferation (cell cycle)

Changes in Gene

Expression

Cycl E:CDK2 p21

p27

E2Fs

Rb

p16

Cycl D:CDK+ p15 Smads

RTK

Cytokine R

Decoy R

Fas

Surface Ag

TGF b R

HPVE7

IAP

Don’t be an Evolutionist, Be a

Revolutionist when taking care of

your breast cancer patients!

THANKS