Dr Catherine Alix-Panabières, MCU-PHUniversity Medical Centre Montpellier

FRANCEFEDER

Clinical relevance of Liquid Biopsy in cancer patients

2013

2010

March 2018

Complementary circulatingbiomarkers

22 337 articles

4 000 articles 3 288 articles

LIQUID BIOPSYNovember 2018

Clinical Trials - CTCs and ctDNA in ClinicalTrial.Gov

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50

100

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200

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completed active

CTC ctDNA

Nu

mb

er

of

Clin

ical

Tri

als

TNM 2010: CTCs in new cM0(i+) Classification

Epithelial-to-mesenchymal transitionAnoikis resistance

Homing & metastasis-initiating potential

Genomic heterogeneity

Invasion/Intravasation ability (single CTCs and/or clusters)

Phenotypic diversity

... ...

Liquid phase of tumor progression

Alix-Panabieres & Pantel, Nature Rev. Cancer 2014

The technical challenge:

Finding one tumor cell in 106 – 108 normal blood cells

Principle of CTC assays:

CTC enrichment followed by CTC detection

CTC Enrichment

Alix-Panabières & Pantel

Nat Rev Cancer

2014

Detection of CTCsAlix-Panabières & Pantel, Nat Rev Cancer 2014

EPISPOT assay

DNA

mRNA

secreted proteins by

VIABLE cells

Detection of Functional CTC/DTC

2002

2005

1rst

2016

2017

EpiDrop

EPIDROP technology

EpiDrop

LNCaP : PSA secretion and EpCAM staining

EpCAM+/-

+/- PSA Secretion

=> PSA release in 1h

Prostate Cancer Patients (N = 100)

Healthy Controls (N=50)

EpiDrop

Clinical Trial

CTCs & ctDNA (cfmiRNA, platelets and

exosomes) provide

complementary information for

Liquid Biopsy

Assays need to be validated by

independent expert groups (Ratner, Nature Biotech 2018)

Interventional clinical studies are

required to demonstrate clinical

utility of Liquid Biopsy(Merker et al. JCO 2018)

! Therapy must work !

Alix-Panabieres & Pantel, Cancer Discovery 2016

CLINICAL RELEVANCE IN EARLY STAGE

CANCER PATIENTS

(TNM-STAGE M0)

CTCs in Neoadjuvant Breast Cancer Therapy

CTC counts provide prognostic information independent from pathological complete response (pCR)

Validated by an international meta-analysis of 2,156 individual patients “21 studies & 16 centers” (Bidard, Pantel et al. JNCI 2018)

2017

CTCs before Neoadjuvant Cancer Therapy & Overall Survival in Breast Cancer

Stratified p value <.0001

N pts % events Hazard Ratio

0 CTC 1175 9.8% 1

1 CTC 199 10.6% 1.09 [0.65-1.69]

2 CTCs 59 23.7% 2.63 [1.42-4.54]

3-4 CTCs 47 29.8% 3.84 [2.08-6.66]

≥ 5 CTCs 93 46.2% 6.25 [4.34-9.09]

months

Surv

ivalfr

act

ion

Bidard, Pantel et al. JNCI 2018

<1 CTC/sample

> 5 CTCs/sample

DETECTION OF MINIMAL RESIDUAL DISEASE

(POST-TREATMENT SURVEILLANCE)

CTCs in high-risk early breast cancer patients during follow-up

Trapp, Janni, Pantel et al. (SUCCESS study group), JNCI 2018

CTCs detected 2 years after adjuvant chemotherapy

CLINICAL RELEVANCE IN METASTATIC PATIENTS

DETECTION & MONITORING OF CTCS

CTC CHARACTERIZATION (DNA, RNA, PROTEINS)

THERAPEUTIC TARGETS (HER2, ER, PD-L1)

RESISTANCE MECHANISMS (AR-V7)

2015

Deactivation

Activation

Antibodies targeting the PD-1/PD-L1

checkpoint have shown dynamic and durable

tumor regressions

→ suggesting a rebalancing of the host–

tumor interaction.

Cancer uses a sort of “cellular camouflage”

Mazel et al., Mol Oncol 2015

(Editorial by R. David in Lancet Oncol. 2015)

Metastatic Breast Cancer Patients

PD-L1PE

William Jacot

PD-L1 is frequently expressed on CTCs (> 60% of patients) in metastatic breast cancer

patients

2017

2014

- 17 centres provided data for 1944 eligible patients from 20 studies

- Meta-analysis on raw data.

Clinical Validity !

CTCs vs. conventional tumor markers (PFS, p values) in

metastatic breast cancer patients receiving chemotherapy

Model

used as

referen-

ce

(

baseline 3-5 weeks 6-8 weeks

CTCBL CA15-3BL CEABL CTC3-5

CA15-3

BL +

CA15-3 3-

5

CEABL +

CEA 3-5CTC6-8

CA15-3

BL +

CA15-3 6-

8

CEABL +

CEA 6-8

N patients 1193 914 593 436 357 289 279 215 170

CP 6 E-10 .10 .04

CP

+CTCBL.32 .12 5 E -05 .25 .35 9 E-05 .40

Few

events

CP

+CTCBL

+ CTC3-5

.26 .41

CP

+CTCBL

+ CTC6-8

.36Few

events

Bidard, Pierga, Michels, Pantel et al, Lancet Oncology 2014, European Pooled

Analysis of CTCs in metastatic BC (n=1944)

Androgen Receptor Variant 7 (AR-V7) Expression in CTCs: Predictive marker for Abiraterone or Enzalutamide Therapy in Prostate Cancer

Antonarakis et al. J. Clin. Oncol. 2017

Good Response Moderate Response Poor Response

INTERVENTIONAL STUDIES BASED ON LIQUID BIOPSY

→ CLINICAL UTILITY

STIC CTC METABREAST

A randomized study to assess

« Circulating Tumor Cells » count at baseline

as a tool to choose between

HormonoT vs ChemoT

in HR(+) M(+) breast cancer patients

J-Y PiergaF-C Bidart

Clinical Utility !

Standard arm N=497 CTC arm N=497

≥5 CTC/7.5ml<5 CTC/7.5ml

CTC-driven decision

Hormone T. ChemoT

Clinician choice

unfavorablefavorable

M+ HR+ breast cancer patients before any treatmentwho can be treated either by HormoneT or chemoT.

Randomizationstratified on center & PS and metastasis-free interval

Hormone T. ChemoT

Endpoints

Medical : PFS (non-inferiority), quality of life, toxicity, OS

Economics : differential costs per life year without disease progression, global costs

Baseline CTC count

blinded

Baseline CTC count

disclosed

Tumor evaluation until progression

Oral Presentation

San Antonio

Breast Cancer

Symposium

12 / 2018

Personalized Treatment of Metastatic castrate-resistant prostate

cancer according to CTC kinetic during chemotherapy:

TACTIK Project

Chemotherapy will be

given or not according

to the investigator’s

decision

N=792 mCRPC patients

to ensure that 396

patients will present >= 5

CTC before initiation of

chemotherapy.

S. Culine

Circulating Cell-free DNA in Blood

Schwarzenbach, Hoon, Pantel, Nature Rev. Cancer 2011

LOW CTDNA CONCENTRATION AS ANALYTICAL CHALLENGE

ctDNA analyses require sensitive technologiesCAPP-Seq: 0.00025 % (Newman et al. Nature Biotech. 2016)

Tumor-associated mutations in ageing individuals

(clonal hematopoiesis, benign tumors)

Low concentration of ctDNA and high background of non-tumor cfDNA

-> need for very sensitive methods

with changes from Cabel L. et al. EJSO 2017

Liquid Biopsy to Stratify NSCLC Patients for EGFR-targeted therapy

Douillard et al. Br J Cancer 2014

N Rate (%) 95% CI

Concordance 652 94.3 92.3-96.0

Sensitivity 105 65.7 55.8-74.7

Specificity 547 99.8 99.0-100

PPV 70 98.6 92.3-100

NPV 582 93.8 91.5-95.6

FDA Approval: June 2016

“… if such mutations are not detected in the

blood, then a tumor biopsy should be

performed to determine if the NSCLC

mutations are present….”

Phallen et al. Science TM 2017: 155 / 216 (72%) alterations were identical in both

plasma and tumor samples (100 pats, various tumor types)

EGFR mutations in tumors and blood plasma:

L858R, exon 19 deletions [19 different mutations] and T790M

• High specificity and sensitivity for ovarian cancer

• Detection of early cancer stages and some tumor types (breast cancer) is still a challenge

In vitro expansion of colon cancer CTCs

Thibaut

Mazard

Laure

Cayrefourcq

2015

AACR AWARD 2017

Cell line from human circulating colon cancer cells- SUMMARY

- Stable for 5 years

- Shares important features of the tumor cells in the patient with colon cancer (primary

tumor/metastasis: KRAS wt; BRAF mutated)

- Showed an intermediate epithelial/mesenchymal phenotype (EM Plasticity) with stem cell–like

characteristics (e.g., growth as microspheres and expression of cancer stem cell markers)

- Induced in vitro angiogenesis

- Induced tumors in immunodeficient mice.

CTC-MCC-41

We have established a cell line with important properties relevant for the development and

progression of metastatic disease.

2017

CTC-MCC LINES

D1

CTC-MCC-41 No colosphere No colosphereCTC-MCC-41.5

A, B, C, D, E, F, G

D30 D60

C1

D15 D45DIAGNOSTIC

in vitro CTC

culture

Patient 044

1rst Line 2nd Line

No treatment

RELAPSE

D75 D90 D105 D165

CTC-MCC-41.4

D158

DEATH

C2 C3 C4 C5 C1 C2 C3

RELAPSE RECHUTE

Blood samples

EDTA tubes

30295 ND

3278

286

CTC-MCC-41 CTC-MCC-41.2 CTC-MCC-41.3 CTC-MCC-41.4 CTC-MCC.41.5

CTC

count

CellSearch

Soler et al. Sci Rep. 2018

Cayrefourcq et al, Cancer Res 2015

Alix-Panabières et al, Clin Chem 2017

2018

Comprehensive outlook on the molecular events involved in colon

cancer progression and provides potential CTC biomarkers that may

help developing new therapies

…to specifically target CTCs with stem cell properties that cause

metastases and tumor relapse in patients with colon cancer.

2017

Platelets

Platelets

KEY AIM: COMBINED ANALYSIS OF CTCS AND CFDNA

Prostate Cancer

(CTCs)

PHRC

Merkel Carcinoma

(CTCs, circulating nucleic acids)

Prostate Cancer

(CTCs, ctDNA & exosomes)

Transcan ‘PROLIPSY’

Lung Cancer

(CTCs, ctDNA, miRNA, exosomes, platelets)

Marie Curie

Breast & pancreas Cancer: AXL

(CTCs, exosomes, plasma proteins)

FUI ‘PANTHER’

Breast & lung Cancers

(CTCs, ctDNA, exosomes)

IMI CALL

CURRENT PROJECTS

Head & Neck Cancer

(CTCs)

CIRCUTEC

Colon Cancer

(CTCs)

COLOSPOT

Cancer-ID is a project funded by the Innovative Medicines Initiative Joint Undertaking (IMI JU) under grant agreement nr. 115749.

Academia, EFPIA

European

Liquid

Biopsy

Society (ELBS)

…European Liquid Biopsy Society (ELBS)

46

Academia, EFPIA, patient organisations, health insurance and

reimbursementorganisations

Coordination

UKE, Prof. Klaus Pantel

Executive Board

Individual Partners

Individual Partners

Individual Partners

Individual Partners

Individual Partners

Individual Partners

Individual Partners

Individual Partners

Individual Partners

ELBS structure

Key Objective:

Foster the

introduction of

liquid biopsy into

clinical practice

Laboratory „Detection Rare Human Circulating Cells“

A big thank you to the patients for donating blood

Magali Boyer

Alexe

Soler

Françoise GarimaMartine MazelLaure CayrefourcqCatherine

Alix-Panabières

Zahra Eslamisamarin

Luis Cortez Hernandez

Léa Sinoquet

Sarah Francini

Renaud Garrel

Jean-Michel Fabre

Eric Assenat

Olivier Dereure

Vincent Foulongne

Valérie Costes

Jean-Louis pujol

Pascal Pujol

Rodolphe Thuret

Delphine Topart

MONTPELLIER

COLLABORATIONS

Laurent Meunier

Stéphane Droupy

NIMES

Xavier Rebillard

MONTPELLIER

William jacot

David Azria

Thibault Mazard

Marc Ychou

Amélie Darlix

Didier Cupissol

Pierre-E. Colombo

Céline Bourgier

Xavier Quentin

Philippe Rouanet

MONTPELLIER

Pierre-Jean Lamy

MONTPELLIER

Sandrine Moutel

Franck Perez

Jean-Yves Pierga

François-Clément Bidard

PARIS

Fabienne Thomas

Etienne Chatelut

Jean-Pierre Delord

TOULOUSE

Paul Hofman

NICE

Jean-Marc Lacorte

PARIS

Philippe Clézardin

Edith Bonnelye

Martine Crozet

LYON

Dominique Heymann

NANTES

Klaus Pantel

Anna Babayan

Simon Joosse

HAMBURG

Klaus Eyer

Guilhem Chenon

Andrew Griffiths

Jérôme Bibette

Jean Baudry

PARIS

Evi Lianidou

ATHENS

Maciej Zabel

Joanna Budna

POZNAN

Angel Carracedo

Ceres Fernadez Rozadilla

Jesus Hita Millan

Angel Diaz-Lagares

SANTIAGO DEL COMPOSTELLA

Hugh Fan GAINSVILLE

Bruce Zetter BOSTON

Hans Lilja NEW-YORK

Sendurai Mani HOUSTON

SUPPORTS

FP6 EU STREP ‘DISMAL’TRANSCAN ‘CTCSCAN’ 2012 TRANSCAN ‘PROLIPSY’ 2016IMI ‘CANCER-ID’ 2015Marie Curie ‘ELBA’ 2017

FUI PANTHER 2016INCa – ARC - La Ligue Fondation de France – MINEFI –SIRIC

MENARINI/ROCHE

Sandrine Dabernat

Etienne Buscail

BORDEAUX

Tom Würdinger

AMSTERDAM

FEDER

Thank you foryour attention !