CJD 2007 November 5-6, Vancouver, BC

1. CJD 2007November 5-6, Vancouver, BC Presented in Partnership by:

2. CJD 2007: Creutzfeldt-Jakob and other Prion Diseases Surveillance, Diagnosis and Treatment • Held at the Fairmont Waterfront • Inaugural year • Approx. 50 delegates • 14 Speakers • 5 Posters

3. Aims and Objectives • Theme = Crossing Borders • PrioNet Canada provides $35 million / 7 years for use in everything from basic to applied prion research • APRI, Public Health Agencies, Health Canada, Food Inspection Agencies all have an investment • Aim is to bring them together in a research consortium

4. A Little Bit About Creutzfeldt-Jakob Disease • Most Common type of TSE • Rare • Believed to have long incubation period • Currently untreatable / incurable • Fatal

5. Variants of CJD • sCJD - Sporadic • iCJD - Iatrogenic • fCJD - Familial - PRNP • vCJD - Variant • Occurs in younger people • Associated with BSE • Type 1 vs. Type 2 • Proposed: PrPDs

6. Other Prion Diseases in Humans • Gerstmann- Straussler-Scheinker Syndrome (GSS) • Fatal Familial Insomnia (FFI) • Kuru (extinct)

7. Prion Diseases in Animals • Bovine Spongiform Encephalopathy • Scrapie • Mink Spongiform Encephalopathy • Chronic Wasting Disease

8. Presentations • Surveillance • Europe • Mexico • USA • Canada • Diagnosis • Treatment • Amyloid treatments

9. National CJD Surveillance Unit Robert Will • Case Ascertainment • Difficult to Diagnose / Heterogenous Symptom Set • Based on referral, death certificates • Diagnostic Tools • 14-3-3 - PRNP • EEG - Neuropathology • MRI • Rate of Occurrence • Criteria • Use of 14-3-3 (and problems) • Neuropathology • May be amended to include use of MRI

10. Interesting Facts / Implications / Issues • PrP genotype is obtained in 30% of cases, but this is not necessarily representative • Changing Criteria • Is CJD Surveillance Ethically Justifiable?

11. Instituto Mexicano Del Seguro SocialVictor Sanchez • Project is just starting out • Mexico has 105 mill, 65 mill • Complete unknown about the rate

12. Interesting Facts / Implications/ Issues • There is still very little knowledge & understanding of CJD in neurologists and pathologists in Mexico: some do not even believe in the prion protein • Difficult to convince pathologists to leave brains or some portion of the brains out of formalin • Family consent for post-mortems • Lack of precautionary measures in place to help reduce possibility of transmission by equipment, etc (reuse of surgical tools, etc)

13. National Prion Disease Pathology Surveillance Centre Pierluigi Gambetti • Goals: • Observe & categorize (origin, phenotype, etiology) as many cases as possible • Analyze tissues • Store tissues • Examine by Western Blot, Immunochemistry, Histology and PrP gene sequencing • Promptly identify and report cases • 33 States Actively participating • Types of Human Prion Diseases

14. Interesting Facts / Implications • 14-3-3 results in many cases being classified as ambiguous • Other projects: • MRI • Blood Donation Look-back • Study of new mutations, atypical cases, co-occurances of PrPsc types 1 & 2 • Proposed new type: Abnormal prion protein, sensitive to Proteases (PrPDs)

15. Public Health Agency of Canada Mike Coulthart • Prevention --> precaution --> preparedness • National Blood Systems Surveillance • Incidence • 22 Cases, P-102-L, E-200K, codon 129 • Genotypes are increasingly M/M • Rate of Autopsies and therefore found cases is fairly high, but additional case-finding may be needed

16. Interesting Facts / Implications • 14-3-3 is band ranked 1-4 so contains some error and subjectivity • Trade-off depending on placement of ‘positive’ and ‘negative’ • Implications are different for different uses • Practicality issues as compared to CSF • pre-emptive work: identifying BSE can affect trade and yet research money depends on this sacrifice as funding is difficult to obtain without a crisis

17. Summary of Surveillance • Facts • Standard rate of 1/million across countries • Various diagnostic measures used • Need for Surveillance • Issues • Early and accurate diagnosis • Criteria (WHO) • Finding ways to increase rates of neuropathology • Question Marks • 14-3-3?

18. CJD in Canada – Detection of Cases & Survival (Gerard Jansen, Public Health Agency of Canada) • 1473 suspected cases referred, many are pending • Typical survival = 4 months • 323 confirmed, 1/10 identified only coincidentally by pathologist

19. Interesting Facts / Implications • By the time reporting to Canadian CJD system occurs, mean survival = 20 days • Why are they missed? • Lack of experience • Similar age of onset and symptoms to many other neurodegenerative disorders • Dependence on certain clinical signs • This makes research into therapies difficult

20. Prions in Tissues and Body Fluids: Implications for Diagnosis and Transmission (Jiri Safar, Institute for Neurodegenerative Diseases, UCSF) • Based on model of CWD, want to determine means of transmission • Uninoculated SHa cohabitated with orally infected Sha or exposed to their bedding • After incubation of 140 days, 80-100% infection rate • Prion concentration in feces was measured at approx 60 ng/g of PrP(Sc) and prion titers of 106.6 units/g • Tested levels of PrP(Sc) in human sCJD patients and generally found low levels in plasma, PrP(Sc) was only found in about half of muscle tissue samples

21. Interesting Facts / Implications • Transmission of CWD most likely happens through natural coprophagy • Concentration of prions in feces has implications for safe disposal of waste from infected animals, especially considering the long-term survival of prions in soil • Human findings suggest possible blood test for CJD and imply importance of plasma lipoprotein removal or blood donor screening as leukodepletion is insufficient

22. Detection of Prions in Sheep Scrapie Blood by Epitope Protection Technology (Marty Lehto, Amorfix Life Sciences) • Development of EP-vCJD test intended to screen human blood for prions • Essentially masks the surface of normal prion proteins so that PrPSc can be detected in an excess of normal proteins • Magnetic beads are then attached and collect proteins magnetically • Assay uses 96-well plate and takes 3.5 hours • Can reliably detect vCJD after diluting 10% homogenate from brain and spleen as much as 10^5 fold • Screened more than 1000 normal samples • A similar test for Scrapie has shown success with naturally and artificially infected animals with various genotypes

23. Interesting Facts / Implications • Good sensitivity- 9.4% false negative • Fresh plasma better than frozen • Criticism that this finding was advertised too soon and false information provided • Issues related to commerce and medical testing

24. Summary of Diagnosis • Facts • Early diagnosis difficult due to variability of symptoms • CJD presents differently according to connectivity and where the disease starts to act, based on where prions are accumulating, based on where they are acquired (peripherally or otherwise). this is in line with the pattern of MRI involvement • Issues • Many cases lost as they can be confused with other diseases or the patient simply dies before CJD is suspected • Commerce and medical testing = conflict of interest • Question Marks • Validity of EP-vCJD test?

25. Amyloidophilic Chemicals for Therapeutics of Prion Diseases (Katsumi Doh-Ura, Tohoku University Graduate School of Medicine • Amyloid-imaging probes developed for Alzheimers disease diagnosis previously appeared to be effective anti-prion chemicals when administered by IV • New chemicals derived from styrylbenzoaole show better penetration through blood-brain barrier and can detect prion protein amyloid in the brain • Demonstration: orally administered compB is effective against all tested prion strains (though effectiveness varies) - it protects neuronal cells from beta-amyloid toxicity and inhibits beta-amyloid formation

26. Interesting Facts / Implications • Appears to be promising lead in drug treatment development • Safety and pharmacokinetics are not yet established • May not be highly effective in some strains • Less effective at later disease stages • may be additionally effective for treatment of Alzheimer’s patients

27. Immunomodulation to Prevent Prion Infection (Thomas Wisniewski, New York University School of Medicine) • Developed as a strategy for treating Alzheimers: passive and active immunization has shown to be highly effective in Humans • Anti-PrP antibody (passive) developed with tissue culture model appears to be effective in mice • Active vaccine also developed for mucosal immunization

28. Interesting Facts / Implications • Potential for complications demonstrates the need to balance humoral immune response with auto-immune toxicity • Varying mechanisms and applications are interesting but the need to identify exposure compromises usefulness: may mean that passive form used for at risk-populations has more utility

29. Prion Disease Therapeutics : Tetracyclic compounds in in vitro models, animal models, and patients (Fabrizio Taliavini, Neurological Institute Carlo Besta) • Iodoxorubicin and tetracyclines • Reduce & revert protease-resistance of PrPSc extracted from brain tissue • Reduce infectivity titer in contaminated material & delays PrPrccs accumulation (ex vivo) • Prevent aggregation by binding to PrP peptide aggregates, abolish neurotoxicity of synthetic amyloid peptile, abolish astroglial proliferation, & prolong survival (in vitro) • Based on inhibition of amyloid formation in vitro • Doxycycline was tested over five years: well tolerated, low toxicity, permeates well • Patients showed very significant difference in longevity even when pair-matched (13.+- 4 vs. 6.0 +- 0.7 months, p<0.01)

30. Interesting Facts / Implications • Currently being verified by in double-blind study by AIFA. • May have implications for Alzheimers research as results appear to be related to direct interaction with misfolded proteins

31. New Approaches to Diagnosis and Treatment of sCJD (Michael Geschwind, UCSF Memory & Aging Centre) • Quinacrine identified as effective in eliminating prions from cell culture (Prusiner) • Randomized, double-blind, placebo-controlled • 34 enrolled, 13 ineligible due to iCJD, vCJD, non-CJD or non-randomization • Quinacrine was generally well tolerated: • Drug induced behavioural changes in some patients: hypomania. 2 had to be taken off. • Some had GI effects • Some required dose adjustments

32. Interesting Facts/Implications • A relational database with clinical, imaging, EEG, laboratory (serological and CSF) and pathological data was developed to assist with diagnosis and differentiation: emphasis on information approach • Out of 976 patients referred to the program, only 3/4 had possible, probable or definite prion diseases. 18% of patients were identified as having treatable non-prion RPD

33. Summary of Treatments • Facts • Certain Amyloid-affecting compounds have shown effectiveness • Some prospective treatments are novel applications of inexpensive antibiotics • Vaccination approach may be alternative to IV/PO therapies • Issues • Commerce & Research: Pharmaceutical disinterest in Prion-only treatments .: drugs with multiple applications may be more viable treatment • Difficulties with diagnosis and limited survival compromises the ability to conduct accurate clinical trials • Question Marks • Potentially interesting to investigate the relationship between Alzheimer’s and TSE?

34. Posters • Diagnosing Genetic Prion Disease (Geshwind, Johnson, et al.) • Review of medical records of referred patients with suspected prion disease • 15% had prion disease, most indicated by family history, but present even in cases without any family history • Genetic testing should be undertaken for any one with prominent CJD-like symptoms or who has suspected sCJD • Interrelationships between knowledge of vCJD & beef consumption decisions • Consumer confidence stufy conducted in Calgary found that 34% of respondents will stop beef consumption if vCJD is found in Canada. • 27% will continue to eat beef until there are multiple cases • 34% would not change beef consumption

35. Posters • Diagnostic Utility of CSF Biomarkers in a sCJD Cohort (Geschwind et al.) • Examined sensitivity and specificity of biomarkers for sCJD (14-3-3, NSE, AB42, Tau, etc.) • Sensitivity, Specificity • 14-3-3 = 51%, 71% • NSE = 63%, 78% • T-Tau= 65%, 100% (but small n) • Quantifying the species barrier in CWD by a novel in vitro conversion assay (Li Li, et al.) • Prions incubated with normal brain homogenates • Readily converts normal cervid molecules, but less of an effect on human, bovine, hamster and mouse • When homogenates are partially denatured (pH 3.5), conversion is greatly enhanced in all species

36. Poster #5 • Mathmatical Modeling of Prion Propogation (Yousefi et al., UBC) • Dizzy Software • Nucleated polymerization model is used in simulating molecular reactions • Gillespie-Direct discrete event simulator based on ordinary differential equations • Modified previously reported estimations of conversion rates (Rubenstein et al.) to reflect newly established [] of 50,000 molecules per cell in normal tissue (Cashman et al.) • Monte Carlo sim. Shows rapid decrease of PrPc in first two days (to 600 molecules) and then a smooth slope that steadies after 70 days • Found that prions converted sooner than expected, with a 9x greater rate of conversion and a much higher [] of PrPsc • Findings of model have shown to be consistent with experimental evidence

37. Closing RemarksFlorence Krantz • Applications: The Research YOU are doing is not only working toward tangible improvements in understanding treatment, but giving hope to families • For medical professionals: the importance of incorporating patients and families in an active role • How individuals who are interested can contribute on a variety of levels, not just academic roles

38. Some Worthwhile Comments about the Practice of Science from The TSE Lightbox : Reflections on Transmission, Refraction and Polarization by Paul Brown • Good Science doesn’t always speak for itself • Learn to write: “We live and die on the basis of publications” • Authorship criteria • Be generous • Identify those who contribute (e.g. physicians who make referrals) • Acknowledge and attempt to explain contradictory results • Cite the best and most important research, not just the oldest or newest • Para-research activities (I.e., Politics) • Do not abuse power relationships • Bring honesty and integrity to your work • Take leadership: sponsor meetings, associations, achieve a presence in the community • Use the media, knowing that you may get burned!

39. Upcoming Event: PrP Canada 2008Canada’s Prion Research Conference • “Making Tracks” to Share, Network and Discover • February 3-5, 2008 • To be held at the Fairmont Royal York in Toronto, Ontario, Canada • Registration: $500 CAN/ $175 for Students • Deadlines • January 4th - for Poster Abstracts • January 4th - for Registration / Hotel • See www.prionmeeting.ca for more details and a list of confirmed speakers

40. Thank Yous & Acknowledgements Dr. David Wishart The Prion Project APRI & PrioNet Dr. Neil Cashman & the members of his lab Speakers & organizers Contact me:Connie Sobsey - csobsey@ualberta.ca