Personal Genome Project (PGP)

1. Personal Genome Project (PGP) • Harvard Medical School IRB Human Subjects protocol • Approved Aug-2005. • Highly-informed individuals consenting to potentially non-anonymous genomes & extensive phenotypes (medical records, imaging, omics).Volunteer waiting list • ** http://pgen.us ** • Cell lines in Coriell NIGMS Repository • (B-cells, keratinocytes, fibroblasts) • G M Church GM (2005) The Personal Genome Project • Nature Molecular Systems Biology doi:10.1038/msb4100040 • Kohane IS, Altman RB. (2005) Health-information altruists--a potentially critical resource.N Engl J Med. 353:2074-7. • McGuire AL, Gibbs RA (2006). Genetics. No longer de-identified. Science.312:370-1.

2. PGP: 1 million Genome/Phenomes via electronic medical records • Why? • Genealogy, forensics, ‘obvious’ traits • Causative mutations: new/old ; germ/soma • Allergens, microbes, viruses (bioweather map) • Immune T & B-cell receptors • Pathology/cancer RNA/DNA • How? $300M in 2003, $3K (free) in 2007 • ‘Next-generation’ sequencing technology • Polymerase colonies (Polonies) 1% cost • Selecting regions (1% : exons, enhancers, etc.)

3. Hair: Baldness [alopecia](minoxidil) Eyes: [Near/Far-sightedness](glasses) Iris color [ARMD] (glasses) Face: [Developmental syndromes, Wrinkles] (Botox) Brain: ADHD (Ritalin); Depression (Prozac) Sleep & Circadian (caffeine, amphetamine, modafinil) Motion sickness (Dramamine, and Scopolamine) Allergies (antihistamines, cortisone, epinephrine, theophylline) Headache (analgesics) Lip: [Cleft palate] (surgery); [Hirsutism] (calcium thioglycolate) Ears: Sensitivity (hearing aids) Nose: Size [breathing disorders] Mouth: Halitosis, throat exams; aerosols [airborne pathogens] Skin: Body odor, Perspiration, Pheromones, Surface texture [psoriasis], Immune components (acne treatments) [acne] Skin color [vitamin D & sunburn] Hands: Dermatoglyphics [syndromes], [Arthritis](corticosteroids) Internal sensors: Proprioceptor, Repetitive stress syndrome Body: Height [short stature] (hGH) [Marfan] Weight [obese](phenethylamine); [anorexia] Metabolic polymorphisms (vitamins, minerals, insulin) Back: Strain sensitivity [IDD] (analgesics) Feet: Plantar fasciitis (orthotic shoes) Athlete’s foot (miconazole, itraconazole, terbinafine, salicylate) Obvious Trait Genetics:Lower Risk Medical Systems Biology

4. Status quo“de-identification” International HapMap Project, reconsent form: “It will be very hard for anyone to learn anything about you personally from any of this research because none of the samples, the database, or the HapMap will include your name or any other information that could identify you or your family.” Genome Wide Association Studies: The proposed GWAS Policy calls for investigators funded by the NIH for GWAS 1) to submit de-identified genotypic and phenotypic data to a centralized NIH repository; and, 2) to submit documentation that describes how the investigators will protect privacy and confidentiality of research participants.

5. Problems with Status quo“de-identification” • Less integrated, holistic, comprehensive • Less enabling of system-wide medicine • Subjects not informed enough to “opt-out” • Life-saving info can’t be shared with subjects • False sense of anonymity (next slide)

6. Is “de-identification” realistic? 1) Re-identification after “de-identification” using other public data. Group Insurance Commission list of birth date, gender, and zip code was sufficient to re-identify medical records of Governor Weld & family via voter-registration records (1998) (2) Hacking. “Drug Records, Confidential Data vulnerable via Harvard ID number & PharmaCare loophole” (2005). A hacker gained access to confidential medical info at the U. Washington Medical Center -- 4000 files (names, conditions, etc, 2000) (3) Combination of surnames from genotype with geographical info An anonymous sperm donor was traced on the internet 2005 by his 15 year old son who used his own Y chromosome genealogy to access surname relations. (4) Inferring phenotype from genotypeMarkers for eye, skin, and hair color, height, weight, racial features, dysmorphologies, etc. are known & the list is growing. (5) Unexpected self-identification. An example of this at Celera undermined confidence in the investigators. Kennedy D. Science. 2002 297:1237. Not wicked, perhaps, but tacky. (6) A tiny amount of DNA data in the public domain with a name leverages the rest. This would allow the vast amount of DNA data in the HapMap (or other study) to be identified. This can happen for example in court cases even if the suspect is acquitted. (7) 26 million Veterans’ medical records including SSN and disabilities stolen Jun 2006. (8)Unauthorized access to DNA bearing samples (9) Identification by phenotype.If CT or MR imaging data is part of a study, one could reconstruct a person’s appearance .

7. Ethical, Legal Social (ELSI) Advisors Misha Angrist Duke Inst. Genome Sciences & Policy Terry Bard HMS IRB, BIDMC Chaplain Dan Brock Harvard Program in Ethics & Health Ruth Chadwick CESAGen, Cardiff Univ. Mildred Cho Stanford Ethics Robert Cook-Deegan Duke Center for Genome Ethics, Law, & Policy Lisa Geller Wilmer-Hale IP Dept. Eric Juengst CWRU Center for Biomedical Ethics Jeantine Lunshof EMGO Institute, Amsterdam Amy McGuire Baylor Bioethics Paul Rabinow UC Berkeley Anthropology John Robertson Univ.of Texas School of Law Peter Singer Univ. of Toronto Joint Centre for Bioethics Daniel Vorhaus Harvard Law School Laurie Zoloth Director, Bioethics, Northwestern Univ

8. ‘Next Generation’ Sequencing Status fL =1E-15 liters (femto) Multi-molecule Reaction Volume AB/APG Ligase beads 1 fL 454/Roche Pol beads 100,000 fL Solexa Pol term 1 fL CGI Ligase 1 fL Affymetrix Hybr array 100 fL Single molecules Helicos Biosci Pol <1fL Visigen Biotech Pol FRET <1fL Pacific Biosci Pol <1fL Agilent Nanopores <1fL

9. Next generation sequencing: Polonies Beads or not, Ligase or Polymerase A G C T Shendure, Porreca, et al. (2005) Science 309:1728

10. Polony Sequencing EquipmentHMS/AB/APG CCD camera microscope with xyz controls HPLC autosampler (96 wells) flow-cell syringe pump temperature control

11. In vitro paired tag libraries Monolayer gel immobilization SOFTWARE Images → Tag Sequences Tag Sequences → Genome Integrated Polony Sequencing Pipeline(open source hardware, software, wetware) Enrich amplified beads Bead polonies via emulsion PCR SBE or SBL sequencing Epifluorescence & Flow Cell Shendure, Porreca, Reppas, Lin, McCutcheon, Rosenbaum, Wang, Zhang, Mitra, Church (2005) Science 309:1728.

12. Why low error rates? Goal of genotyping & resequencing  Discovery of variants E.g. cancer somatic mutations ~1E-6 (or lab evolved cells) Consensus error rateTotal errors(E.coli)(Human) 1E-4 Bermuda/Hapmap 500 600,000 4E-5 454 @40X 200 240,000 3E-7 Polony-SbL @6X 0 1800 1E-8 Goal for 2006 0 60 Also, effectively reduce (sub)genome target size by enrichment for exons or common SNPs to reduce cost & # false positives.

13. Monitoring resistance to BCR-ABL-kinase inhibitors with polonies during CML patient therapy Nardi, Raz, Chao, Wu, Stone, Cortes, Deininger, Church, Zhu, Daley (submitted) M244V T315I E255K