Pathology Molecular Diagnostics in the Netherlands Past, Present, Future

Winand N.M. Dinjens Clinical Scientist in Molecular Pathology (CSMP)

Head Molecular Diagnostics

Department of Pathology

Erasmus MC Cancer Institute, Rotterdam, the Netherlands

w.dinjens@erasmusmc.nl

Norwegian Society of Molecular Pathology

March 6, 2019

Trondheim, Norway

August 24, 2017

Rosenborg – Ajax: 4-2

August 24, 2017

Rosenborg – Ajax: 4-2

March 5, 2019

Real Madrid – Ajax: 3-5

Disclosures

Translational research fees: AstraZeneca

Bristol-Myers Squibb

AbbVie

Consultancy: Amgen

Bayer

Bristol-Myers Squibb

Novartis

Roche

the Netherlands: 42.500 km2

the Netherlands

Population: 17.1 million

the Netherlands

Hospital Health Care

UMC: 8 (7)

Netherlands Cancer Institute Antoni van Leeuwenhoek, Amsterdam

Peripheral hospitals: 70 (?)

the Netherlands

Pathology

Pathology labs: 42 (?)

Practising Pathologists: +/- 350

the Netherlands

Pathology

Dutch Pathological Society (NVVP)

>600 members: General Pathologist

Neuropathologist

Veterinairy Pathologist

Toxicological Pathologist

Forensic Pathologist

Clinical Scientist in Molecular Pathology

https://pathology.nl/

https://www.vapweb.nl/

Society Pathology Technicians (VAP)

>1250 members

Since 2015

Joint meeting Dutch

Pathological Society and

Society Pathology Technicians

the Netherlands

Pathology Molecular Diagnostics

Past

Start 1995 – 2000: Group Molecular Biologists and Pathologists (<15)

Exchange ideas, assays, controls, information,

QC, reporting, education, meetings

Allelotyping (tissue admixture)

T- and B-lymphocyte clonality

Solid tumor clonality analyses (mutation, LOH)

HPV

Working group Molecular Diagnostics in Pathology

(within the Dutch Pathological Society, NVVP)

1. Stimulating molecular diagnostics in pathology in NL

2. QC and improving quality

3. Renew molecualr diagnostics

4. Inventarising and adjustment

5. Advice

6. Sharing information and experience

7. Education and training

the Netherlands

Pathology Molecular Diagnostics

Past

2000 – 2012: Group Molecular Biologists, Pathologists and

Molecular Technicians (+/- 100)

Allelotyping (tissue admixture)

T- and B-lymphocyte clonality

Solid tumor clonality

Mutation, Translocation

HPV

Klinisch Moleculair Bioloog in de Pathologie (KMBP)

Clinical Molecular Biologist in the Pathology

Clinical Scientist in Molecular Pathology (CSMP)

1. 2010 : Educational program finalised

2. 2011: Transitional arrangement

3. 2012: call for registration

4. 2013: 22 registered (4 pathologists), 5 CSMP in training

5. 2014: call for registration of educational program

instructor, deputy instructor, educational program,

site visits, registration committee

Clinical Scientist in Molecular Pathology (CSMP)

Klinisch Moleculair Bioloog in de Pathologie (KMBP)

• PhD in Molecular Biology subject (biol, chem, med doctors, etc: 29, 3 PA)

• Educational program, two year fulltime training (currently 12, 1 PA)

• Embedded within the Dutch Pathological Society (NVVP)

• Guideline: involvement CSMP is necessary when performing

molecular diagnostics

• Yearly Pathology Molecular Diagnostics Day (13)

Educational program Clinical Scientist in Molecular Pathology

General competences: Molecular diagnostic action

Communication

Collaboration

Knowledge and Science

Social acting

Organisation

Professionalism

the Netherlands

Pathology Molecular Diagnostics

Past - Present

2012 – 2019: Group Molecular Biologists, Pathologists and

Molecular Technicians (>300)

Recognition and establishment of discipline

Clinical Scientist in Molecular Pathology, July 2013

2013: NGS era

Astrid Eijkelenboom, Ph.D.; Bastiaan B.J. Tops; Anke van den Berg; Adrianus J.C. van den Brule; Winand

N.M. Dinjens; Hendrikus J. Dubbink; Arja ter Elst; Willemina R.R. Geurts-Giele; Patricia J. Groenen; Floris

H. Groenendijk; Daniëlle A.M. Heideman; Manon M.H. Huibers; Cornelis J.J. Huijsmans; Judith W.M.

Jeuken; Léon C. van Kempen; Esther Korpershoek; Leonie I. Kroeze; Wendy W.J. de Leng; Carel J.M.

van Noesel; Ernst-Jan M. Speel; Maartje J. Vogel; Tom van Wezel; Petra M. Nederlof; Marjolijn J.L.

Ligtenberg

Virchows Archiv, 2019, accepted

Recommendations for the Clinical Interpretation and Reporting of Copy

Number Gains using gene panel NGS analysis in Routine Diagnostics

13th yearly day

Molecular Diagnostics

in Pathology

the Netherlands

Pathology Molecular Diagnostics

Present

2019: 23/42 Pathology labs perform Molecular Diagnostics

14 labs perform NGS

Nation wide project collaborations

Erasmus MC, Rotterdam, Department of Pathology

Laboratory for Molecular Diagnostics

(ISO 15189, 2012 accredited)

DNA:

1. Sanger sequencing 2. Mutation-specific PCR

3. SNAPshot analyses 4. Fragment analyses

5. CNV-MLPA 6. MS-MLPA

7. MS-PCR 8. Methylation-array

9. MSI 10. ISH

11. Next Generation Sequencing (NGS) 12. ctDNA (liquid biopsy)

13. Tumor mutation load/burden

(T- and B-lymphocyte clonality, HPV)

DNA isolation from routine Pathology (FFPE) specimens

Mix of tumor and normal cells, highly degraded DNA

Paraffin block Immuno stained section

Paraffin section (stained) cytology preparation

H&E stained section

“Massive parallel”

“Single molecule”

100s-1000s fragments / analysis

Output 50 – >1000 x 106 bases

Short amplicons (<200bp)

Lab developed panels

Enriched with SNP amplicons Dubbink et al., J Mol Diagn. 2016.

doi: 10.1016/j.jmoldx.2016.06.002

Low amount of input DNA (<<10 ng)

High sensitivity (<5%)

Mean coverage 500-1500x

>Semi-quantitative

Pooling of samples

Bio-informatics support

Next Generation Sequencing (NGS)

Ion GeneStudio S5 Prime System

Erasmus MC, Rotterdam, Department of Pathology

Laboratory for Molecular Diagnostics

(ISO 15189, 2012 accredited)

Next Generation Sequencing (NGS):

1. Standard diagnostics NGS panel (CMA):

NSCLC, CRC, melanoma, GIST, tumor clonality

2. Brain tumor (CMA)

3. Lynch syndrome (CMA)

4. Thyroid cancer (CMA)

5. BRCA1/2, Homologous Recombination Deficiency (HRD) (CMA)

6. Pheochromocytoma / Paraganglioma (CMA)

7. TML, Oncomine TML assay

8. ctDNA, Oncomine ctDNA Lung (CRC) assay

Erasmus MC, Rotterdam, Department of Pathology

Laboratory for Molecular Diagnostics

(ISO 15189, 2012 accredited)

RNA

1. CMA RT-PCR

2. ISH

3. Archer

4. Nanostring (u.d.)

5. RNAscope

Protein:

1. IHC

MMR-proteins

(MLH1, MSH2, MSH6, PMS2),

ALK, ROS1, pan-NTRK, P53, IDH1,

BRAF, P16, β-Catenin, E-Cadherin,

SDHA, SDHB

the Netherlands

Pathology Molecular Diagnostics

Present

2019: Nation wide project collaborations:

Predictive Analysis for THerapy:

PATH to optimising Access to

Personalised Cancer Therapy in

the Netherlands (36 months, 1.1 million €)

Prof. Dr. M.J.L. Ligtenberg

CSMP

Radboudumc

Nijmegen

the Netherlands

Pathology Molecular Diagnostics

Present

2019: Nation wide project collaborations:

CtDNA On the way to Implementation in the Netherlands (COIN)

(48 months, 3.7 million €)

Prof. Dr. G.A. Meijer, Pathologist

Netherlands Cancer Institute Antoni van Leeuwenhoek

Amsterdam

The Netherlands Organisation for

Health Research and Development

CZ

Health Insurance

the Netherlands

Pathology Molecular Diagnostics

Future

Future 1: tissue / Pathology

2020-…..: Less but larger Pathology laboratories

centralisation, sub-specialisation

Molecular Diagnostics: larger panels (WES, RNA seq)

more centralisation

Future 2: tissue / Pathology

Future 3: tissue / Pathology

Future 4: tissue / Pathology

Immune Checkpoint Blockade related

predictive en prognostic tissue biomarkers:

1. TILs (Tumeh et al, Nature 2014)

2. T-cell Inflamed Immune Gene Expression Profile (GEP)

(Cristescu et al., Science 2018)

3. Immune Gene Expression Signature (Jiang et al., Nat Med 2018)

Cohen et al., Science 2018

Gandara et al., Nat Med 2018

Hogan et al., Cancer Immunol Res, 2018

Peripheral blood TCR repertoire profiling may facilitate patient stratification for immunotherapy against melanoma.

Future 5: Blood / Liquid biopsy

NEXTSEQ

6000

Molecular Diagnostics

Future 1, 2, 3, 4: Pathology

NEXTSEQ

6000

Molecular Diagnostics

Future 1, 2, 3, 4: Pathology

Future 5: Pathology?

NEXTSEQ

6000

Molecular Diagnostics

Future 1, 2, 3, 4: Pathology

Future 5: Pathology?

Future 6: Pathology?

Tumor profiling

exhaled air

Youssef et el.,

Oncotarget, 2017

Composition Microbiome

(Gopalakrishnan et al.,

Science 2018)

(Routy et al., Science 2018)

NEXTSEQ

6000

Molecular Diagnostics

Future 1, 2, 3, 4: Pathology

Future 6: Pathology?

Future 5: Pathology?

Tumor profiling

exhaled air

Youssef et l.,

Oncotarget, 2017

Future 7: Pathology?

www.amp.org Association for Molecular Pathology (AMP)

TWO-DAY MOLECULAR PATHOLOGY DIAGNOSTICS SYMPOSIUM (MD)

Monday 9 and Tuesday 10 September 2019

Erik Jan

Dubbink

Niels

Krol

Ronald

van Marion

Ina

Geurts-Giele

Peggy

Atmodimedjo

Ludo

Uytdewilligen

Lotte

Berger

Hein

Sleddens

CSMP CSMP

in training Senior

Techs

Techs

NGS

Techs

ISH

Bio-

Inf. Secr.

Isabelle

Meijssen

Albertina

Dirkx-van der

Velden

Dorine

den Toom

Carolina

Valente

Marit

de Haan

Laura

Moonen

Esther

Korpershoek

Vera

Martens

Techs

NGS

Margot

van den

Akker

Jan

von der Thüsen

Pathol.

Shelly

Bierhuizen

Floris

Groenendijk

Walter

Voogt

Thank you for your attention