Review of evolving technologies in CUP - PowerPoint Presentation

Slide1

Review of evolving technologies in CUP

Dr

Natalie Cook

Senior Clinical Lecturer in Experimental Cancer Medicine, University of Manchester

Honorary Consultant in Medical Oncology

Christie NHS Foundation Trust

Talk overview

Liquid biopsy technologies

to be discussed;

Circulating tumours cells (CTCs)

Circulating free DNA (

cfDNA

)Tumour educated platelets (TEPs)Current research

in Manchester, UK

Liquid biopsy

Clinical Application

Resistance

Relapse

ctDNA

CTC

Circulating microRNA

Exosomes

Microvesicles

TEPs

Blood

Plasma

RNA expression

Protein expression

CTC derived

organoids

Point mutations

Methylation patterns

Chromosomal aberrations

Translocations

Amplifications/deletions

Early detection

Diagnosis

/

prognosis

Minimal residual disease

Response to therapy

Therapy selection

Diagnostic challenges of CUP

Delays in diagnosis

& multiple (invasive) investigations to find primary site

Limited

robust evidence

that treating patients based on

molecularly-predicted primary improves outcomesSeveral research & commercial tissue of origin classifiers; not recommended for diagnosis of CUP

Without tissue of origin, no access to novel therapeutics or immunotherapy10-20% failure rate of molecularly profiling tumour tissue

How representative is a single small biopsy of 1 metastatic lesion?

Alternative = liquid biopsies

Tissue

of origin from liquid biopsy?

Discovery of methylated circulating DNA biomarkers for comprehensive non-invasive monitoring of treatment response in metastatic colorectal cancer

Barault

et al 2017, Gut, 67(11

):1995

Identification of methylation haplotype blocks aids in deconvolution of heterogeneous tissue samples & tumor tissue-of-origin mapping from plasma DNAGuo

et al 2017, Nat Genetics, 49(4):635

Plasma DNA tissue mapping by genome-wide methylation sequencing for non-invasive prenatal, cancer, and transplantation assessments

Sun

et al

2015, PNAS, 112(40):E5503

Detection and localization of surgically resectable cancers with a multi-analyte blood test

Cohen

et al

2018, Science, 359(6378

):

926

RNA-

Seq

of tumor-educated platelets enables blood-based pan-cancer, multiclass and

m

olecular pathway cancer diagnostics

Best

et al

2015, Cancer Cell, 28(5):666

Where could liquid biopsies

fit?

Tissue of origin and biomarker status

Treat as equivocal tumour type

Palliative care

Immuno-therapy

Chemo-therapy

Prognostic indicators of appropriateness of therapy

Targeted therapy

Prognostic information

Quicker diagnosis to direct investigations

Determine

Primary site

Molecular subtype to stratify to treatment

Early indicators or response/resistance

Understand the

biology of CUP

Diagnosis

Management decisions

Treatment response or resistance

Circulating Tumour Cells

(CTCs)

Circulating tumour cells are very rare

CTC

1 ml of blood is about 1/5

th

of

a teaspoon

5,000,000,000 red blood cells/ml blood

7,000,000

white blood cells/ml blood

<1-15

circulating tumour cells/ml

blood

<1 CTC per 500,000 WBC

What does 1 in

500,000 look like?

What about

CTCs in CUP?

Very limited published research

(total of 21 patients evaluated across 2 studies)

Only CellSearch

technology used (only identifies

EpCam positive CTCs)No cellular or molecular characterisation

Allard et al 2004. Clin Cancer Res, 10(20):6897

Utility of measuring

CTC counts

to assess the efficacy of treatment for

CUP

Komine

et al

2014.

Anticancer Res, 34(6):

3165

Summary of CTC counts in 7.5 mL of blood from patients with various types of carcinomas

The Manchester approach;

the CUP BioBank

Parsortix

method

HDSCA

slide preparation

Cell pellet

Double spun plasma

Cells stored in glycerol

Total nucleic acids extracted

HDSCA

method

‘no

cell left

behind’

12 x slides

~

3 million cells per slide

Prospective recruitment of new CUP patients pre-treatment

Characterise CTCs & evaluate

heterogeneity

Consent

to:

Use of surplus biopsy

2 additional blood samples for CTC banking and nucleic acid extraction

Clinical

information

HDSCA;

High

Definition Single Cell

Assay

Circulating Tumour

DNA

(

ctDNA

)

Circulating tumour DNA

Multiple potentials for molecular profiling from blood

& tissue

(

none

funded in the UK)

FoundationOne; tissue, >300 genesFoundationOne Liquid

, blood based, 70 genesGuardant 360; 73 genes, blood based

Oncologica; tissue, 505 genes

Local Academic Initiatives;

eg TARGET

Looks for

Targetable

driver &

actionable

mutations

Copy number translocation

Tumour mutational burden in some cases

All solid tumours

ctDNA

&

germline controls

DNA

360

gene panel

650 somatic mutations

Molecular Tumour Board

Stratified to treatment (Phase I)

Next generation sequencing

Determine tissue of origin

Tissue of origin directed therapy

Copy number analysis

Development and optimisation of

cfDNA

methylation assay

Combine mutational profile, methylation profile and copy number variation to

evaluate prognostic and predictive

biomarkers

Improved predictive power of mutational profile

Manchester approach

Within the TARGET

ctDNA

trial

TARGET CUP patients so far

Gene

7

13

43

57

62

68

69

70

71

80

94

122

169

182

242

243

272

TP53

●

●

●

●

●

●

TTN

MAP2K2

NOTCH1

●

KRAS

●

●

ABCC1

ABL2

●

AKAP9

ALK

●

ALK

ARID4A

ATIC

ATM

BAP1

●

●

BUB1B

CDH1

CHD5

CYLD

●

DDR2

EPHA5

ERBB3

●

ERCC2

●

FBXW7

●

FGFR1

FLT4

HOXA11

IGF2R

KDM5A

KIF1B

KMT2A

●

KMT2C

LIFR

MLK1

MLLT6

MUC16

NOTCH3

NF1

NSD1

●

NTRK2

NTRK3

NUP214

NUTM2B

P2RY8

PCDHGB1

PRRX1

●

RAP1GDS1

SOX2

SPEN

●

SRGAP3

●

STK38

SYNE1

TCF12

TCF7L2

TRIP11

Total

17

2

5

2

4

2

2

0

5

3

6

3

2

5

5

1

6

17 CUP TARGET patients recruited to date

>10% VAF

1-9% VAF

●

Predicted driver mutation in CGI

Mutation data alone insufficient to determine tissue of origin

We can determine presence of tumour DNA

Majority of patients have at least 1 mutation (16/17)

13/17 patients have VAF of >5%

Tumour Educated Platelets

(TEPs)

Tumour Educated Platelets

Best

et al

2015.

Cancer Cell 28(5):

666-676

Manchester CUP project overview

Cells

&

Proteins

team

Tissue Biomarker team

Nucleic acid biomarker team

Preclinical

Team

TARGET trial

Phase 1

ECMT

Clinical and Experimental Pharmacology

TIIML

CTC team

Advanced cultures

CUP biology

& heterogeneity

Immune markers

MSI/

dMMR

CTC enumeration

CTC biology

Longitudinal

cfDNA

analysis of patients on

treatment

Mutation & Copy Number analysis

Methylation patterns

MCRC CUP Biobank

Archival tissue

CUP project

Conclusions

Lots of

new technologies in the field of liquid biomarkers

None routinely used;

lack evidence of improved outcomes

Mostly research

initiatives, but slowly being incorporated into trialsFuture standard of care?

Acknowledgements

STR

Supported by

The Systemic Therapy Research Group

@ The Christie NHS Foundation Trust

The patients and their families

Alicia Marie Conway

Claire Mitchell

Caroline

Dive

Ged Brady

Experimental Cancer Medicine Team, Christie NHS Foundation Trust

Clinical and Experimental Pharmacology Group, Cancer Research UK Manchester

Institute