From Feasibility to Utilityin Precision Cancer Medicine –

Experiences from the First Norwegian Studyof NGS-Based Therapy Decisions in Advanced Cancer

Anne Hansen Ree, Professor of OncologyAkershus University Hospital, University of Oslo

Hege G. Russnes, Molecular Pathology Senior Consultant and Researcher

Oslo University Hospital, Norwegian Radium Hospital

Precision Cancer Medicine

What is PCM?

Technological advances in molecular biology,functional imaging, and informatics

New therapies within medical, radiation, and surgical remedies

Updated education curricula, clinical structures,and regulatory approval pathways

Resources that reside in the industrial, regulatory, academic,and clinical practice sectors

Precision Cancer Medicine

Identified and integrated existing hospital and research facilitiesand expertise at the two study centers

established a PCM diagnostic pipeline within the existing practice of the public health services

educated multidisciplinary teams essential to a PCM program

offered molecularly matched therapy in progressing end-stage cancerobtained long-lasting responses to the NGS-based therapy decisions

The Project Group

Management TeamGunhild M. Mælandsmo – Molecular BiologistAnne-Lise Børresen-Dale – Molecular BiologistKjersti Flatmark – Oncology SurgeonAnne Hansen Ree – Med.-Rad. OncologistHege G. Russnes – Molecular PathologistOle Christian Lingjærde – BioinformaticianGry A. Geitvik – Lab. Engineer

Eivind Hovig – BioinformaticianVivi A. Flørenes – Molecular BiologistVessela N. Kristensen – Molecular BiologistInger Riise Bergheim – Lab. EngineerVeronica Skarpeteig – Lab. EngineerMenaka Sathermugathevan – Lab. EngineerMarius Lund-Iversen – PathologistTorill Sauer – PathologistKlaus Beiske – PathologistSalah Nasser – RadiologistEspen A. Ruud – RadiologistLars Julsrud – RadiologistClaudius H. Reisse – RadiologistKirsten T. Hagene – Study Protocol Designer

Clinical Trial TeamDaniel Heinrich – Med.-Rad. OncologistSvein Dueland – Med.-Rad. OncologistKjetil Boye – Med.-Rad. OncologistVigdis Nygaard – Molecular BiologistLaxmi Silwal-Pandit – Molecular BiologistOlga Østrup – Molecular BiologistVegard Nygaard – BioinformaticianEinar A. Rødland – BioinformaticianSigve Nakken – BioinformaticianJanne-Merete Torset Øien – Study NurseChristin Johansen – Study NurseEline Aas – Health EconometricianHilde Lurås – Health Econometrician

Study idea– End-stage metastatic cancer from any origin– Disease progression according to RECIST

– A metastatic site available for biopsy – detailed molecular analyses– Administration of drug targeting an identified molecular disruption

– Radiological evaluation intervals of 6–12 weeks– Rebiopsy during treatment and at end-of-study facilitating studies of tumor evolution

MetAction hypothesis

Study design

Patient eligibility– End-stage metastatic cancer from any origin– Disease progression according to RECIST

– ECOG performance status 0–1– Life expectancy of more than 3 months

– Previous line of systemic therapyfor 6 or more weeks

– Radiological evaluation intervals of 6–12 weeks

Primary end point: PFS (Period-B/Period-A ≥ 1.3)

Secondary end points: OS, ATI rate, CTCAE grade 3–5 adverse events

Ree, Russnes et al.,ESMO Open 2017

Targeted sequencing – Ion AmpliSeq Cancer Hotspot Panel

• IonTorrent PGM– Cancer Hotspot Panel

» 50 genes/210 primer pairs

– Coverage/Read depth: 2,000x

– Fast method

– Small amount of input material (10 ng DNA)

• Validation at Section of Molecular Diagnostics

• Development of bioinformatics pipeline

• FISH for selected CNA (HER2) and translocations (ALK and BCR-ABL)

Re-run and/or validation

Prelim. pathology report (cytology)

Final pathology report

OncologistRadiologist

Cytologist/PathologistStudy nurse

Lab.engineerMol.biol.

Mol.pathol.

Bioinfo.Lab.engineer

Mol.biol.Mol.pathol.

Prelim. variant report Final variant report

Ree, Russnes et al.,ESMO Open 2017

The diagnostic pipeline

Laboratory pipeline

A booklet including all personnel contact info, logistics and SOPs were created for a secure and rapid diagnostic pipeline

Key personnel: constantly updated Sample-kits included

premarked and barcoded slides, cytospin tubes,

and freezing vials

Collect sequencing- and meta-datafrom IonTorrent;

Run variant calling

Annotation of variants

Technical/QC assessment of runand of variant calling

Generate an “initial” variant reportMainly technical information,interactive links to databases

Interpretation of variants;Potential clinical impact

Molecular Tumor Board

Finalizing the variant report;Technical, molecular, and functional information

as well as a summary

Bioinformatician

Bioinformatician -> Lab.engineer

Bioinformatician

Bioinformatician, molecular biologist, pathologist, oncologist

Bioengineer, bioinformatician,molecular biologist, pathologist, oncologist

Bioinformatician, molecular biologist(pathologist, oncologist)

Lab.engineer, bioinformatician

Bioinformatics pipeline

Collect sequencing- and meta-datafrom IonTorrent;

Run variant calling

Annotation of variants

Technical/QC assessment of runand of variant calling

Generate an “initial” variant reportMainly technical information,interactive links to databases

Interpretation of variants;Potential clinical impact

Molecular Tumor Board

Finalizing the variant report;Technical, molecular, and functional information

as well as a summary

Bioinformatics pipeline

One pathology report was issued integrating cytology, FISH, and molecular analyses.

Data and report presented at tumor board (videolink OUS-AHUS) but completed and signed after consensus.

Treatment recommendation reported in clinical journal.

1st study phase: median 18.0 (range, 8–39) days

Reporting results

Study cases – 1st phase (enrolment: 9 May 2014 – 26 Aug 2015)

SexAge

(years)

Primary

tumor siteHistological diagnosis Sampling site(s)

Adverse

eventGene mutation(s)

Male 43 Parotid gland Neuroendocrine carcinoma Lung None None detected

Male 69 Liver Hepatocellular carcinoma Liver transplant None None detected

Female 59 Pancreas Adenocarcinoma Liver None KRAS p.G12D

Male 62 Pancreas Adenocarcinoma Liver None KRAS p.G12D, TP53 p.R306X

Female 66 Pancreas ND Liver None KRAS p.G12D, GNAS p.R201H

Female 46 Right colon Adenocarcinoma Liver None KRAS p.G12D, SMAD4 p.R361C, TP53 p.C176F

Male 54 Right colon Adenocarcinoma Liver None KRAS p.G12D, PIK3CA p.E545K, SMAD4 p.G419R

Male 62 Right colon Adenocarcinoma Liver NoneKRAS p.G12D, SMAD4 p.Y353C, APC p.Q1444X, TP53 c.304_307

ACCT>ACT

Male 52 Left colon Adenocarcinoma Liver None KRAS p.G12V, PIK3CA p.E545Q

Male 59 Left colon Adenocarcinoma Liver None TP53 p.R273H, TP53 p.L350fs

Male 65 Left colon Mucinous adenocarcinoma Lung None KRAS p.A146T, PIK3CA p.H1047R

Female 68 Left colon Adenocarcinoma Liver transplant None KRAS p.G12A, APC p.Q1291fs

Male 70 Left colon Adenocarcinoma Liver None KRAS p.G13D, APC p.E1306X, APC p.K889fs, TP53 p.R248W

Male 54 Rectum Adenocarcinoma Lung None KRAS p.G12S, APC p.E1317X, TP53 p.R175G

Male 61 Rectum Adenocarcinoma Lung Pneumothorax KRAS p.G12S, APC p.Q1378X, FBXW7 p.R505C

Male 61 Rectum Mucinous adenocarcinoma Lung None BRAF p.V600E, SMAD4 p.W524C, TP53 p.R306X

Male 68 Rectum Adenocarcinoma Lung None KRAS p.G12D, PIK3CA p.E542K

Male 63 Kidney Clear cell renal cell carcinoma Thoracic wall None VHL p.L158P

Male 65 KidneyRenal Xp11.2 translocation

carcinomaLiver None None detected

Male 61 Urinary bladder Urothelial carcinomaInguinal lymph node

and peritoneumNone None detected

Female 48 Ovary Small-cell sarcoma Peritoneum NoneKRAS p.T58I, PIK3CA p.H1047Y, ERBB2 p. V842I, CTNNB1 p.G34R,

TP53 p. R306X

Male 65 Prostate Adenocarcinoma Axillary lymph node None TP53 p.D184fs

ATI rate: 0/22Ree, Russnes et al.,ESMO Open 2017

1st study phase – conclusions

• 24 patient enrolled

– actionable targets detected

– none treated (as per protocol)

• Limitations

– few actionable targets on the screening panel

– only single-agent therapy: one biomarker, one drug

• Comprehensive diagnostic procedure established

– expedite and safe mutation profiling of metastatic tumors

– data interpretation at multidisciplinary tumor boards

Study amendments (September 2015 – February 2016)

Diagnostic gene mutation panel– from the Ion AmpliSeq™ Cancer Hotspot Panel

to the Ion Oncomine™ Comprehensive Panel

Data interpretation– extended liberty for the MTB to interpret the mutation data

Therapy– opportunity for the CTB to conclude on combination regimens

(with established safety data)

Targeted sequencing – Ion Oncomine Comprehensive Panel

8

Oncomine® Cancer Comprehensive Panel* Gene List

DNA Panel RNA Panel

Hotspot genes, n=73 (hotspot coverage)

ABL1

AKT1

ALK

AR

ARAF

BRAF

BTK

CBL

CDK4

CHEK2

CSF1R

CTNNB1

DDR2

DNMT3A

EGFR

ERBB2

ERBB3

ERBB4

ESR1

EZH2

FGFR1

FGFR2

FGFR3

FLT3

FOXL2

GATA2

GNA11

GNAQ

GNAS

HNF1A

HRAS

IDH1

IDH2

IFITM1

IFITM3

JAK1

JAK2

JAK3

KDR

KIT

KNSTRN

KRAS

MAGOH

MAP2K1

MAP2K2

MAPK1

MAX

MED12

MET

MLH1

MPL

MTOR

MYD88

NFE2L2

NPM1

NRAS

PAX5

PDGFRA

PIK3CA

PPP2R1A

PTPN11

RAC1

RAF1

RET

RHEB

RHOA

SF3B1

SMO

SPOP

SRC

STAT3

U2AF1

XPO1

Copy gain, n=49

ACVRL1

AKT1

APEX1

AR

ATP11B

BCL2L1

BCL9

BIRC2

BIRC3

CCND1

CCNE1

CD274

CD44

CDK4

CDK6

CSNK2A1

DCUN1D1

EGFR

ERBB2

FGFR1

FGFR2

FGFR3

FGFR4

FLT3

GAS6

IGF1R

IL6

KIT

KRAS

MCL1

MDM2

MDM4

MET

MYC

MYCL

MYCN

MYO18A

NKX2-1

NKX2-8

PDCD1LG2

PDGFRA

PIK3CA

PNP

PPARG

RPS6KB1

SOX2

TERT

TIAF1

ZNF217

CDS, n=26 (full gene)

APC

ATM

BAP1

BRCA1

BRCA2

CDH1

CDKN2A

FBXW7

GATA3

MSH2

NF1

NF2

NOTCH1

PIK3R1

PTCH1

PTEN

RB1

SMAD4

SMARCB1

STK11

TET2

TP53

TSC1

TSC2

VHL

WT1

Fusion drivers, n=22

(183 assays)

Copy Number Variants

ALK

RET

ROS1

NTRK1

ABL1

AKT3

AXL

BRAF

CDK4

EGFR

ERBB2

ERG

ETV1

ETV1

ETV5

FGFR1

FGFR2

FGFR3

NTRK3

PDGFRA

PPARG

RAF1

*For Research Use Only . Not f or use in diagnostic procedures.

Current Status: 143 Regions • Covers 143 regions: Hotspot mutations, CNV, and expression of selected fusion genes• FDA-approved platform for NGS-based therapy selection (NCI-MATCH trial)

1st study phase: median 18.0 (range, 8–39) days2nd study phase: median 17.5 (range, 9–57) days

Study cases – 1st phase (censoring date: 9 May 2016)

Ree, Russnes et al.,ESMO Open 2017

Study amendments (September 2015 – February 2016)

Diagnostic gene mutation panel– from the Ion AmpliSeq™ Cancer Hotspot Panel

to the Ion Oncomine™ Comprehensive Panel

Data interpretation– extended liberty for the MTB to interpret the mutation data

Therapy– opportunity for the CTB to conclude on combination regimens

(with established safety data)

Study cases – 2nd phase (enrolment: 14 Mar 2016 – 8 Mar 2017)

Study cases – 2nd phase (enrolment: 14 Mar 2016 – 8 Mar 2017)

Study cases – 2nd phase (enrolment: 14 Mar 2016 – 8 Mar 2017)

Study cases – 2nd phase (enrolment: 14 Mar 2016 – 8 Mar 2017)

Case 1: right-sided colon adenocarcinoma with liver metastases

Case 2: right-sided colon adenocarcinoma with liver metastases

Case 2: right-sided colon adenocarcinoma with liver metastases

conclusions

• NGS-based therapy decisions in advanced cancer

– feasible within the public health services of Norway

– a PCM diagnostic pipeline within the existing clinical practice, with operational multidisciplinary teams

• The MetAction diagnostic pipeline and patient treatment

– safe !

– reversed the disease course in end-stage cancer

• Remaining challenges in PCM

– drug availability and high costs

– limited knowledge about the functional outcome of gene mutations in the real-world oncology practice

perspectives

British Columbia

Cancer AgencyNCI-MATCH MetAction

MetAction 2

(planning by Jan 2018)

Catchment

population4.6M

>1,100 sites across

the USA5.2M 5.2M

Sequencing

strategy

WGS/WTS

(at the Genome

Science Centre)

Targeted sequencing

(Oncomine

Comprehensive

Panel)

Targeted sequencing

(Oncomine

Comprehensive

Panel)

Targeted sequencing ?

WGS/WTS ?

Bioinformatics

pipelineGenomic report MATCHBox Targeted report Targeted report

Multidisciplinary

board(s)MTB by videolink MATCHBox

MTB, then CTB by

videolink

MTB, then CTB by

videolink

Clinical actionOngoing early-phase

trials, off-label access

30 treatment arms

(by Mar 2017)

with RP2D

Designated MA trial

Designated MA trial with

defined treatment arms

(mini NCI-MATCH)

Total timeline Approx. 5 weeks Median of 37 days Median of 17.5 days Aiming to maintain

Partnership Not known FDA and NCIUnsuccessful

with pharma

Will try to

engage pharma

Funding BC Cancer Foundation NCI RCN Anyone?

acknowledgments