CSER Clinical Sequencing Exploratory Research

1. CSERClinical Sequencing Exploratory Research Accomplishments, Challenges and Opportunities 19 May 2014 Jim Evans MD, Ph.D University of North Carolina at Chapel Hill

2. Green & Guyer. Nature; 470:204.2011

3. Medical Science = Clinical Care • Basic science is the foundation of medical practice but the endeavors demand different approaches • Theory alone is insufficient to guide action in clinical practice • The balance between premature translation and inappropriate delay in translation is difficult to achieve • The stakes are high • Misapplication regularly results in profound harm • To patients • To society through increased cost • Through missed opportunities • Medical practice ultimately involves individual values • Which ultimately can’t be gainsaid

4. A Long History of Premature Application of Medical Intervention • Therapeutically • Reflexic HRT after menopause • Anti-arrhythmics for PVCs • Excessively strict glucose control in diabetes • In Screening • Routine use of PSA screening • Whole body MRIs

5. The Stakes are High in the Clinical Application of Genomics • Patients (& families) make serious decisions when we apply genomic knowledge • False positives lead to: • Unnecessary surgery; years of unnecessary screening • Premature end to diagnostic pursuit, forgoing the true answer • False negatives lead to: • Forgoing necessary preventive/therapeutic modalities • Amplified by misclassification of family members as at-risk or not • Family planning & abortion • The psychological damage of misinformation ? ? ? ? ?

6. Our Values Regarding Genetic Information Vary If you carry a mutation that essentially guarantees that you will develop a severe, unpreventable & untreatable neurological disease by age 60 would you wish to be told? • Yes • No • I’m not sure A few really bad genetic diseases… • Alzheimer Disease • Fatal Familial Insomnia • Spinocerebellar Ataxia • Huntington Disease • CADASIL We need to understand how heterogeneous values can be accommodated

7. Underlying Rationale of CSER &Related Clinical Sequencing Programs Lots of sequencing is already being performed; it’s getting really cheap, why not just introduce it into clinical care and learn that way? • Unsystematic introduction of new modalities into clinical care generates poor and misleading data regarding its utility & appropriate clinical applications • Shifts costs to an already constrained medical system • Nothing is free in the world of clinical medicine • A free but inappropriate clinical test is very expensive • Makes patients de facto research subjects • Privacy concerns are especially resonant in genomics • Undermines systematic collection & sharing of genomic & phenotypic information - critical to understanding the medical implications of the genome

8. To establish best practices for translation and implementation of genomic medicine

9. CSER is Seeking Data Regarding: • Patient characteristics that signal potential utility of WG/ES, e.g. • Diagnostic “hit” rates in different clinical populations • When is somatic analysis of tumors most informative for guiding Rx? • How to analyze large data sets in a clinical environment? • Quality control • Timeliness • Confidentiality • Integration with EMR • etc. • Special considerations in different populations • e.g. minorities and traditionally underserved groups • Different values • Different allele frequencies • How should we deal with the plethora of highly heterogeneous “non-target” data generated when performing genomic sequencing? Gene discovery is important but incidental to the main goals of CSER

10. Aggregate CSER Accomplishments • Implemented and integrated genome-scale sequencing into the clinical arena with successful sequencing, analysis, reporting and return of results • Designed & implemented a variety of (relatively) efficient analytic systems and workflows • Established numerous “gene lists” to facilitate analysis • Progress towards open-source analytic workflows • Implementation of facile reporting mechanisms and formats to providers and patients • Demonstration that WES from FFPE tumors is equivalent to that from frozen samples

11. Aggregate CSER DataReported 3/12/2014 • Total Enrollment: 1,532 • 1,051 Adults (69%) / 481 Children (31%) • 796 Female (52%) / 736 Male (48%) • Racial/Ethnic breakdown*: • White 74% • African American 7% • Hispanic 8% • Asian 3% • American Indian 2% • Not Reported 7% • Total Sequenced: 1049 • 232 Tumor-focused / 817 Germline-focused • 109 Presentations and Posters / 61 Publications

12. Tumor WES results HIGHEST category of mutation PER PATIENT Mutations of “established clinical utility” Cat. 1 Cat. 2 Mutations of “potential clinical utility” Cat. 4 Only mutations in “non-cancer genes” Cat. 3 Mutations in other “cancer genes”

13. Return of Results & Clinical Relevance • Germline analysis of 817 individuals • “Positive”for diagnostic finding 241 (29%) • Incidental Findings per ACMG list 24 (2.9%) • Incidental Findings by relaxed criteria 46 (5.6%)

14. Diagnostic Yield by Patient Category 175 cases with overall 41% positive/possible; 59% negative Cardiology – 21 cases 3 positive 5 possible 13 negative Retinal - 33 cases 7 positive 11 possible 15 negative Dysmorphology – 24 cases 5 positive 3 possible 16 negative • BLSNHL – 24 cases • 3 positive • 8 possible • 13 negative • Neuro – 49 cases • 9 positive • 12 possible • 28 negative • Cancer - 24 cases • 3 positive • 2 possible • 19 negative 46% 38% 54% 43% 21% 33%

15. ELSI–Related Results • A conscious and planned focus on ELSI is part of CSER’s core mission • Early results in this sphere: • Informed consent in the context of genome-scale sequencing • CSER-wide study systematically analyzed the IC documents from the 9 sites & formulated specific recommendations for best practices • Defining specific challenges related to clinical genomics • Reliable measurement tools • Needs/opportunities for instrument development • Factors influencing patient understanding & outcomes • Applications of qualitative, quantitative and mixed-methods research

16. ELSI–Related Results • Incidental Findings are an active focus of CSER • Frequency of “medically actionable” IFs: 3-6% • Degree of choice provided to patients has been an issue • CSER sites have been instrumental in shaping debate, revealing discrepancies between current policies and patient preferences • Led to shift in ACMG’s policy for ROR • Beginning to accrue data addressing what patients wish to know regarding “off target” findings • Majority of patients “want everything” when asked • But even minimal “real world” barriers (e.g. need to make a phone call) dramatically decreases desired return • CSER is collecting data on measures of patient satisfaction/distress with regard to both diagnostic findings and off-target results

17. Future Challenges & Opportunities Outcomes • CSER has erected an integrated infrastructure to implement clinical genomic medicine • The central challenge now is to assess whether the application of genomic medicine actually improves patient outcomes • Health • Morbidity & mortality • Patient satisfaction, values, and well being • Economics • Do we save money? • How much do improved outcomes cost? • Explore outcomes in diverse settings, including ethnic, socioeconomic, clinical status A role for CSER/NHGRI since the “market” does not focus on outcomes

18. Outcomes • Assessing genomic outcomes demands new approaches • Qualitatively different approaches • New cohorts of patients with a range of diagnoses • Comparisons between those with application of various approaches & those with standard care (e.g. CER) • e.g WES vs. WGS vs. multigene panels vs. combined approaches • Will feed into efforts of IGNITE and EMERGE • Systematic, quantitative extension of existing cohorts • Large numbers of patients • Longitudinal follow up of those with positive results • Impact on care, dissemination to family members, etc. • Longitudinal follow up of those with negative results • Optimal approaches to dynamic reinterpretation as knowledge / technology evolve • Outcomes measurement must continue to include ELSI-oriented outcomes

19. Future Needs, Challenges & Opportunities Analytical Approaches • CSER sites have established a variety of inventive analytic workflows for analysis of clinical genomic data • Need for controlled investigations to determine optimal approaches in order to maximize efficiency • How to best make difficult trade-offs between sensitivity and specificity • e. g. phenotypically-driven “gene lists” that maximize specificity vs. “gene agnostic” approach, maximizing sensitivity • Likely to differ by clinical context • Cancer vs. ID • Especially important given time & resource constraints of analysis in the clinical setting

20. Analytical Approaches Variants of Uncertain Significance • Adjudication of a variant’s impact is the single most significant obstacle to the application of genomic medicine • We are evolutionarily predisposed to tolerate false positives & therefore over-interpret data • The genome is big & thus coincidences are common • Our protocols for adjudication are wholly inadequate • The stakes are high • There rarely exist “gold standards” for adjudication • Need for statistical approaches like “mutational burden” & thus large numbers • Planned overlap • ClinGen (aggregation of multiple sources of variation, ranging from commercial labs to more basic LSS efforts) • CSER (investigating optimal processes and mechanisms for curation in the clinical setting) • EMERGE (large scale integration with EMRs)

21. Future Needs, Challenges & Opportunities Integration & Sharing • Progress will rely on integration and sharing • Need to develop formats that facilitate data sharing amongst sites/labs/vendors/clinical settings • Incorporate data science to meet the expected problems of scale • While protecting privacy and complying with HIPPA • The market is poorly equipped (indeed, often antithetical) to integration and sharing data • Well adapted to a UO1 consortium model • Integration of genomic data with other “omic” data • Where clinically promising • e.g. RNAseq in tumor analysis • Integration with training programs

22. Future Needs, Challenges & Opportunities Rational & Targeted Expansion • Expansion of groups sequenced • New diagnostic categories of patients • Need to consider whether genomic approaches can improve health in the general population • Carrier screening for interested individuals • Targeted sequencing of selected genes in general population that, when mutated, strongly predispose to severe but preventable disease • Facilitate longitudinal use of genomic information throughout lifespan • Expansion regarding venues & populations: • Ensure broad diversity of patients • Beyond the academic setting where warranted • Natural progression from CSER to IGNITE • e.g. coordination with PBRNs • Investigate cascade impact on family members • We must carefully consider what we mean by “genomic approaches”

23. Future Needs, Challenges & Opportunities Clinical Genomics = WGS • Focused testing is usually optimal clinically • Unnecessarily broad testing can lead to waste, false positives and downstream harm with increased cost • Genomic corollary is determining the proper role of WGS vs. panels (virtual or otherwise) • CSER has (appropriately) focused on WG/ES • Is now well equipped to also determine when WG/ES vs. more targeted sequencing is optimal • Comparisons of WGS vs. WES vs. targeted panels in differing clinical contexts

24. We’re Wasting Data • We don’t know the true penetrance or new mutation prevalence for almost any Mendelian condition • Ascertainment bias has likely over-estimated penetrance and underestimated new mutation rate for most conditions • Accurate figures will be critical for both clinical and public health genomic applications • Boot-strap on existing sequencing efforts • Create registry for reporting of “off target” mutations in critical disease genes to define true prevalence & mutation rates • Must include ability to obtain: • Personal medical history • Family history • Long-term f//u • Where possible, parental genotypes • CSER & EMERGE are logical places to start

25. Thank You jpevans@med.unc.edu

26. Spectrum of Genomic Medicine Implementation: Intensity vs. Breadth Breadth of Implementation IGNITE eMERGE Depth of Patient Characterization CSER Evidence Generation System-Wide Impact UDN NSIGHT Testing Multiple Models Individual Patient Focus

27. Intensity of Phenotypic/Genomic Characterization vs. Breadth of Implementation IGNITE 51,000 NSIGHT 1000 UDN 1400 eMERGE 25,000 CSER 3000 Testing Multiple Models Individual Patient Focus Evidence Generation System-Wide Impact

28. Summary of Clinical Programs of MI-ONCOSEQ

29. Categories of Mutations Identified Through MI-ONCOSEQ 8% Failed Biopsy/tumor content 13% Non-informative n=402

32. A Long History of Premature Application of Medical Intervention • Therapeutically • Reflexic HRT after menopause • Anti-arrhythmics for PVCs • Excessively strict glucose control in diabetes • Beta-carotene supplementation • In Screening • Routine use of PSA screening • Whole body MRIs

33. ELSI–Related Results • The Outcomes & Measures Working Group identified priority areas for investigation of psychosocial and behavioral impact of integrating sequencing into clinical process • Identified specific challenges related to clinical genomic sequencing, including: • Reliable measurement tools • Needs/opportunities for instrument development • Factors influencing patient understanding & outcomes • Importance of qualitative, quantitative and mixed-methods research

34. Under-sequenced Medically-Relevant Genes 1. KCNQ1: potassium voltage-gated channel, KQT-like subfamily, member 1 Familial atrial fibrillation; Short QT syndrome; Acquired Long QT syndrome ~50% of Exon 1 poorly covered 2. PMS2: postmeiotic segregation increased 2 (S. cerevisiae) Lynch syndrome; colorectal cancer, endometrial cancer Multiple exons with poor coverage at 3’ end; 5’ portion of last exon segmental duplications

36. Results by Diagnostic Categories 43% • By clinical category 21% 38% 54% 54%

37. PediSeq Project First 39 cases(prospective and unknown retrospective) 3 positive (~13%) 8 possible (~33%) 13 negative (~54%) 46% • By clinical category • BLSNHL = 24 cases • Sudden Cardiac = 9 cases • Mitochondrial = 1 case • Intellectual Disability = 5 cases 2 positive (~22%) 2 possible (~22%) 5 negative (~56%) 0 positive (0%) 0 possible (0%) 1 negative (100%) 1 positive (20%) 1 possible (20%) 3 negative (60%) 47 validation (known) Cases – all positive (100%)

38. Results by Diagnostic CategoriesFirst 145+ cases 8 positive (~18%) 11 possible (~25%) 25 negative (~57%) 43% • By clinical category • Neuro = 44 cases • Cancer = 24 cases • Cardiology = 12 cases • Retinal = 33 cases • Dysmorphology= 24 cases 3 positive (~13%) 2 possible (~8%) 19 possible (~79%) 21% 1 positive (~8%) 3 possible (~25%) 8 negative (~67%) 33% 7 positive (~21%) 11 possible (~33%) 15 negative (~45%) Total = 145 cases 25 positive (17%) 33 possible (23%) 87 negative (60%) 7 positive (~21%) 11 possible (~33%) 15 negative (~45%) 54% 54% (40% positive /possible)

39. Next two slides should be distributed, along with some general CSER information, to Council ahead of meeting so they can review on their own and I don’t need to waste time on it.

40. Lucia – check on publications/presentations for update Scope of CSER at 9 Sites • Clinicians Involved: 154 • Non-Health Care Professionals: 180 • Number of Institutions 26 • Peer reviewed publications 56 • Non-peer reviewed publications 5 • Posters at regional/Ntl/Interntl venues 14 • Presentations 95

41. Patient Focus of Individual CSER Sites Somatic tumor sequencing Germline only analysis • Baylor • WES of tumor and germline in childhood cancers • Brigham & Women / Harvard • Randomized trial of WGS in healthy adults & those with suspected hereditary cardiomyopathy • Children’s Hospital of Philadelphia • WES and WGS in four cohorts of children with pediatric disease • SNHL, ID, mitochondrial, sudden cardiac death • Dana Farber / Broad Institute • WES of tumor and germline in assorted cancers • University of North Carolina at Chapel Hill • Randomized trial of WES in adults and children with wide variety of likely genetic diseases • Cardiac disease, neurodevelopmental disorders, dymorphology, cancer, ophthalmology diseases • University of Washington • Randomized trial of tumor and germline WES to compare usual care vs. provision of genomic information in adults with CRC • Hudson Alpha (new) • Children with intellectual dysfunction • Kaiser (new) • Adult couples in context of preconception carrier screening • University of Michigan (new) • WES of tumors in adults and children with advanced cancer

45. Early tumor report data NUMBER OF SUBJECTS with mutations in each category Proven or potential clinical utility Any “cancer gene” mutation Sept 2013: 58 tumor exomes

46. Case of Interest I Challenges of IFs Incidental Variants on WES • MSH6 c.1170delT (p.Phe391fs) • BRCA2 c.2857G>T (p.Glu953*) • Diagnostic Variants • ABCA4c.1622T>C (p.Leu541Pro) • ABCA4 c.1805G>A (p.Arg602Gln) Chance? Lower penetrance in unselected populations? New mutation(s)?

47. Case of Interest II Occasional Therapeutic Implications • Subject dxd at age 6 with “hereditary spastic paraplegia” • Confined to crutches and wheelchair • Painful spasticity on daily basis • At 36 no alternative dx had been found in spite of multiple referrals and evaluations nationally • On WES: GCH1 [p.Arg216*] mutation, consistent with a Diagnosis of dopa-responsive dystonia • Rx with oral L-dopa often effective • Dramatic response

48. Case of Interest III Expansion of our understanding of phenotypes • Child with non-syndromic cone-rod dystrophy • Two likely deleterious mutations in BBS9 • c.1255C>G (p.Pro419Ala) • c.2153A>T (p.Glu748Val) • BBS9 has not been associated with non-syndromic CRD, but CRD is a feature of BBS • There are other BBS genes associated with non-syndromic retinitis pigmentosa

49. Assorted Challenges Highlighted by CSER Sites in Last Update • Timely analysis • Especially in setting of diseases with poor prognosis (Baylor, Dana Farber) • When Rx/EOL decisions hinge on analysis (Baylor, Dana Farber) • Disappointment among some participants when not randomized to receive maximal results (BW) • Construction of user-friendly reports for full range of users (providers to patients) • Curation and signing out variants • Complex and time consuming (everyone) • Inaccurate public databases of variants (everyone) • Analytic approach (gene lists vs. gene-agnostic approach (CHOP and UNC) • Difficulty of providing informed consent given inherent clinical time constraints • Difficulty of / need for independent CLIA confirmation • Eventual expansion of phenotypes associated with mutations • Proliferation of available commercial tests of large panels – how to navigate? • Insufficient knowledge about penetrance of mutations, especially in context of non-target information

50. Maintaining a Clinical Focus • MPS is (just another) highly complex medical test • Claims that “soon everyone will have their whole genome sequenced” are typically predicated only upon its low cost • Even if “free”, perceived low cost is an illusion • Indiscriminate application of medical tests is very expensive • Morbidity/mortality to individuals (e.g. PSA screening) • Expense to individuals & society • It should be applied as are other medical tests: • When and if the situation warrants • With targeted genomic analysis in most settings • Targeting could be actual (e.g. capture) or virtual (at the analytic stage) • WGS/WES useful in selected diagnostic settings and in research • Think before burdening the system with a flood of extraneous information that begs for misinterpretation • CSER is generating data to guide its optimal use