Whole Genome Sequencing in Cancer Diagnostics (research) Nederlandse Pathologiedagen 19 & 20 November 2015

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1 Whole Genome Sequencing in Cancer Diagnostics (research) Nederlandse Pathologiedagen 19 & 20 November 2015 Dr. I.J. Nijman

2 Disclosure slide (Potential) conflict of interest None For this meeting relevant financial relationship(s) and relevant nonfinancial relationship(s) Sponsorship or research money Fee or other (financial) compensation Stock holder Other relationship: Hartwig Medical Foundation (advisor) None None None None

3 Where is the challenge? Changes in DNA are the basis for cancer But also make every cancer patient unique

4 Personalized treatment

5 Decades of research has yielded unprecedented insight in tumorigenesis - Drugs have been or are being designed to target various biological pocesses - Many drugs only work in part of patients - No biomarkers are available for most drug sensitivity or resistance

6 Biomarker discovery requires large cohorts and systematic integration of large scale genetic and treatment data

7 1990 ~3,000 bp/day January 2015: 1,000 dollar genome ,000 bp/day million bp/day billion bp/day Disease gene identification Reference genome Personal genomes

8 From one test per gene to one test for all genes - Genetics first approach: - Decrease diagnostics - More comprehensive genetic screening - Faster diagnosis

9 BioInformatics: common steps in analyzing NGS data Read mapping (alignment): fit single reads on the reference genome Variant calling: detect deviations from the reference genome (and discriminate from sequencing errors) Copy Number detection: detect gains or losses of pieces of DNA. Structural Variation: detect rearrangement of DNA.

10 Applications of Personal Genomes in Clinical Care From Cradle to Grave >50 60 >100 Birth planning - Screening carriership Disease prevention - BRCA, CFTR Aging - Understanding healthy aging Pregnancy - NIPT (trisomy 13/18/21, gender, carriership) Cancer - Personalized treatment Newborn - Replacing heel prick: detection rare congenital disease Diagnostics congenital disease - De novo mutation screening - Whole genome scan Pharmacogenetics -drug/dose choice Death - Genetic autopsy unexplained disease cause

11 National initiatives Genomics England, with the consent of participants and the support of the public, is creating a lasting legacy for patients, the NHS and the UK economy, through the sequencing of 100,000 genomes. US$215 million investment in precision medicine research, Goal: to collect gene, biochemical, lifestyle and other data from one million volunteers.

12 Center for Personalized Cancer Treatment Bottom-up initiative founded in 2010 UMCU, EUR, NKI UMC Groningen AMC Amsterdam VuMC Amsterdam LUMC Leiden Meander Radboud Nijmegen MUMC Maastricht

13 Two Weeks Center for Personalized Cancer Treatment Patient with Metastatic Disease 2-4 Biopsies (fresh frozen) Pathological Analysis ng DNA Isolation ng Patient Stratification Research IonTorrent PGM MiSeq Illumina HiSeq X ten or + Biomarker Discovery Actionable Mutations & Amplification >50 genes Profiling Cancer Pathways and Processes Start Targeted Therapy Allocation Fase1 Clinical Trial Systems Biology Whole Genome Sequencing Response monitoring Bioinformatic analysis Resistance / Progression Remission / Cure Databanking Mutations, INDELs, Copy Number Variations in vitro / in vivo Modeling of Hypotheses Response monitoring in ecrf

14 National scaling: Hartwig Medical Foundation Value chain Including patients Collecting clinical data Taking biopsies 1 Preprocessing biopsies Sequencing DNA of biopsies Value chain Managing sequence Analyzing database and clinical data in database, for new treatment and and reporting non-treatment options Conducting clinical trials Storage of tissue 1 (biobank) Centralized Facility in a Foundation setup - Made possible through philanthropy (2 to 3 years) - Whole genome sequencing using Illumina Xten setup - Integrate clinical and genetic data - Provide input for individual patient reporting - Provide access to cohort information for research to benefit future patient care Location: Matrix VI, Amsterdam Science Park Start: April 2015 Operational: Summer 2015

15 HUB organoids Test specific drugs on tumor organoids to confirm sensitivity Treat patient with selected drug(s) until disease progression Bioinformatics and Systems Biology to identify pathways Obtain patient biopsy

16 Big data Sequencing 50 genomes per day.. ~4.5 Terabyte of raw data Alignment time required: 5000 hrs on a single core Variant calling time required: 3000 hrs on a single core additional analyses (somatics, copy number, structural variation): ~100 hrs on a single core Large compute cluster or cloud compute (Google, Amazon). Local/cloud based storage? Store what for how long? Distribution and data sharing?

17 Challenges Somatic / low frequency calling on WGS: Additional coverage needed: 90x tumor samples vs 30x reference sample Reduces capacity on sequencer from 16 to 4 samples per run of 3 days. Increased computational load: analysis pipelines run ~5-6 days on limited hardware IT problem: scalability of infrastructure. Meeting desired turn around time of 7 days. Integration of sequencing results with clinical and other data & interpretation. Translation to clinical application & diagnostic certification

18 Systematic data generation and integration

19 Summary & conclusions Affordable, routine WGS with reasonable turn-around-time is possible Still in research setting, but diagnostic certification process started. Structural collection of clinical, sequencing and follow-up data started in CPCT for metastatic patients with evaluable data for ~targeted treatments IT challenges (infrastructure, scalability, databasing, integration) not to be underestimated Biological (and clinical) interpretation and actionability will need continuous scientific input.

20 Acknowledgements All CPCT contributors Patients

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22 Nextbio Integrates genomic & clinical data Powerful annotation and filter options Both population and single patient based views, analyses and reporting.

23 Patient reporting and cohort mining Illumina/Nextbio: data integration platform and diagnostic reporting software Validated Clinically- Actionable Markers Off-label drug program (drug repositioning) in collaboration with all large pharma - CPCT: DRUP - ASCO: TAPUR Additional Markers with Potential Clinical Benefit

24 Treatment Z Gene X wild type Systematic biomarker discovery Identify DNA changes associated with good or poor response Gene Z mutated

25 Timeline view of treatment