Torben Ørntoft Prof. Chief Physician Dept . Molecular Medicine Aarhus University Hospital

1. Risk screening and personalizedtherapy in cancer and otherdiseases: a personalisedmedicineexample Torben Ørntoft Prof. ChiefPhysician Dept. MolecularMedicine Aarhus University Hospital

2. Staff of 70 20 in Diagnostics 50 in R&D NGS core–center Microarray core-center Biobank core-center Research Areas: Prostate Cancer Colorectal Cancer Bladder Cancer Bioinformatics DENMARK

3. Definition • Genomicmedicine- usinggenomic information in the clinic to enable a more precisestratification of patients and citizens, for the purpose of surveillance, prevention, diagnosis and treatment

4. AUH –Vision -Det Molekylær medicinske hospital Forskning DIAGNOSE Cloud Tolkning/ validering Patient Blodprøve DNA sekventering (HiSeq 2000) DNA isolering Tolkning/ Validering BEHANDLING

5. Clinicaluse of sequencing • Inheritedsyndromes and diseases • Identification of citizens with a highgeneticrisk of disease, with respect to e.g. screening • Develop a novel and more precisestratification of diseasesbased on cellbiology • Enablechosing the right treatment • More exactdiagnosis • Including patient specificmetabolism of dugs

6. Request form

7. The application of NGS at Aarhus University hospital • Inheriteddiseases • Cancer, Heart, Liver, Skeleton, Kidney, Syndromes, Immune deficiencies, Endocrinediseases • Screening of males with highrisk of prostate cancer • Bacteria • Eg. Cysticfibrosis; Microbiome, Nosocomialinfections • Cancer Tissue • CUP - treatmentselection

8. The use of moleculardiagnosticswithin cancer diseases • Prevention and screening for cancer • Inherited cancer diseases • High risk of cancer • Diagnosing cancer • Earlydiagnosiscures • Precisediagnosis (subgroups and aggressive VS non -aggressive) • Treatment of cancer • Selectingtherapybased on mutations in cancer tissue

9. Kræft panel med >70 gener der kan give arvelig kræft Analysers for de klinisk genetiske afdelinger i Ålborg. AUH, Vejle og Odense Analysen laves i MOMA Kommenteret svar til Klin. Genetisk Afd. (KGA) KGA svarer patienterne

10. The use of moleculardiagnosticswithin cancer diseases • Prevention and screening for cancer • Inherited cancer diseases • High risk of cancer • Diagnosing cancer • Earlydiagnosiscures • Precisediagnosis (subgroups and aggressive VS non -aggressive) • Treatment of cancer • Selectingtherapybased on mutations in cancer tissue

11. Prostate Cancer - Hits 12% of all males

12. Clinical Challenges: Overdiagnosis and Overtreatment • PSA does not distuinguish aggressive/non-aggressive PC • Increased use of PSA testing (overdiagnosis) • Lack of accurate prognostic tools (overtreatment) - Gleason score (grade) and TNM stage • PSA-based screening saves one life for every 48 PC patients treated by surgery ERSCP, European Randomized Study of Screening for Prostate Cancer Aarhus county Age-standardized PC incidence & mortality (DK) Mukai et al. 2010 Storm et al. 2010

13. Background >30% PC risk SNPs identified by GWAS studies Typical OR: range 1.1-1.2 Cumulative effects have been reported: Zhang et al. NEJM 2008 5 risk SNPs +/- family history: -10-fold increased PC risk Xu et al. The Prostate 2009: 14 risk SNPs +/- family history: -absolute risk prediction -distinguish 52% vs. 8% PC risk (20-years) -withouth this knowledge all men have background risk (10-13%) Easton & Eeles 2010 More than 30% of familial likelihood identified, manuscript accepted in nature genetics april 2011

14. Screened – treated - saved

15. Screened – treated - saved Low geneticriskneedsno PSA test Takesplace in general practice

16. Prostate cancer challenge

17. Aggressive or non-aggressive?

18. Vi forudsiger hvilke prostata cancere der er aggressive vs non-aggressive

19. Screened – treated - saved Low geneticrisk non -aggressive

21. The use of moleculardiagnosticswithin cancer diseases • Prevention and screening for cancer • Inherited cancer diseases • High risk of cancer • Diagnosing cancer • Earlydiagnosiscures • Precisediagnosis (subgroups and aggressive VS non -aggressive) • Treatment of cancer • Selectingtherapybased on mutations in cancer tissue

23. DiagnosticYield • 23 heart patients previously tested for few genes by Sanger sequencing with no positive findings • In 10 of these, we identified likely pathogenic mutations • In 3 additional patients, we identified possibly disease causing mutations • Huge gain in diagnostic yield for heart patients

24. Colon CancerNew taxonomy • Cancer in the bowel

25. Unsupervised NMF clustering of trancriptome data from 314 Danish CRC patients identifies 5 natural sample subtypes Goblet Stroma Serrated RNA decay CIN (n=73) (n=35) (n=53)(n=31) (n=106) HE-stain, Goblet HE-stain, Stroma HE-stain, serrated HE-stain, CIN NMF: Nonneg. Matrix Factoring Jesper and Trine – unpublished

27. IMMUNE - Therapy Unsupervised NMF clustering of trancriptome data from 314 Danish CRC patients identifies 5 natural sample subtypes Goblet Stroma Serrated RNA decay CIN (n=73) (n=35) (n=53)(n=31) (n=106) HE-stain, Goblet HE-stain, Stroma HE-stain, serrated HE-stain, CIN NMF: Nonneg. Matrix Factoring Jesper and Trine – unpublished

28. The use of moleculardiagnosticswithin cancer diseases • Prevention and screening for cancer • Inherited cancer diseases • High risk of cancer • Diagnosing cancer • Earlydiagnosiscures • Precisediagnosis (subgroups and aggressive VS non -aggressive) • Treatment of cancer • Selectingtherapybased on mutations in cancer tissue

29. Liquid Biopsy • Plasma cfDNA

30. Liquid Biopsy • Plasma cfDNA Sekventering af cirkulerende DNA

31. Strategic research alliance grant NOCRC, Novel CRC Screening Tools Improve Survival and Cost-effectiveness20 mio. d.kr for 2014-18 The Danish Council for Strategic Research, Programme Commission on Individuals, Disease and Society

32. Solid tumors release DNA to the circulation Cell free DNA, incl. ctDNA, isolated from plasma Circulating tumor DNA, ctDNA

33. Monitoring recurrence using ctDNA Stage II rectal cancer Marker 12 mth molecular lead time Method: Digital PCR Plasma input: 1 ml ctDNA markers: Somatic mutations (structural genomic rearrangements)

34. Monitoring recurrence using ctDNA Stage II rectal cancer Marker 12 mth molecular lead time Ready for clinicaltesting Method: Digital PCR Plasma input: 1 ml ctDNA markers: Somatic mutations (structural genomic rearrangements)

35. Individualiseret / skræddersyet behandling

39. Workinggroupbased in US and UK US 18 groups

42. Molecular diagnostics in patients with unknown primary carcinoma (CUP) Workflow/Methods Targetedsequecing of tumor DNA/RNA Illumina NGS VARIANT CALLING: SomaticSNPs and indels: MuTecht 2 Copynumber variations: Sequenza Structural variants: Delly 2 TUMOR BOARD Oncologist, pathologist, molecularbiologist, clinicalpharmacologist

45. Conclusions • Much has beenachievedbased on exomes (2%) • Much is in clinicaluse / or on the way to clinicaluse • BUT: We still only find mutations in 50% of thosethat have a seriousinherited cancer disease, the rest is to bediscovered in the remaining 98% non- codinggenome Wecanonly match pharmaceuticals to mutations in <20% of patients We have difficulties in findingdisease markers for the polygenecommondiseasese.g. Diabetes, Metabolic Sdr., Hypertension, Arthritis, Allergyetc We have toofewpharmaceuticalsdirectedtowards the non-Coding part of the genome Future: Weneedwholegenomesequencing of welldefineddiseasegroups to stratify patients in relevant sub-groups.

46. Spørgsmål?