1. WHO Guidelines on Tissue Infectivity Distribution in Transmissible Spongiform Encephalopathies World Health Organization 2006 Proceedings of a Consultation Convened by WHO Geneva 14-16 September 2005 Summary David M. Asher, MD Laboratory of Bacterial, Parasitic & Unconventional Agents Division of Emerging & Transfusion-Transmitted Diseases Office of Blood Research & Review Center for Biologics Evaluation & Research US Food & Drug Administration

2. WHO Guidelines on Tissue Infectivity Distribution in Transmissible Spongiform Encephalopathies http://www.who.int/bloodproducts/TSEREPORT-LoRes.pdf World Health Organization 2006 Proceedings of WHO Consultation Geneva 14-16 September 2005 http://www.who.int/bloodproducts/TSEREPORT-LoRes.pdf Quality and Safety of Plasma Derivatives and Related Substances Department of Medicines Policy and Standards Health Technology and Pharmaceuticals Cluster World Health Organization -------------------------  ------------------------- WHO Secretariat A Padilla (project leader) S Groth L Rago D Wood FX Meslin N Dhingra

3. Proceedings of a Consultation Convened by WHO Geneva 14-16 September 2005 Goals of Consultation Revise WHO 2003 Consultation advice on TSE safety of medicinal products, especially biological products  Vaccines, etc., prepared with ruminant-derived materials  Products derived from human blood (components, plasma derivatives)  Other human cells and tissues and products derived from them Review scientific information on TSEs relevant to safety of medicinal products Compare TSE risk assessments for various countries Provide general advice to “national regulatory authorities with limited resources” about TSE risks associated with medical products and suggest approaches to assess and, if indicated, attempt to reduce risk Special goal: Summarize and evaluate reliability of available information about distribution of infectivity and disease-related prion protein in tissues and body fluids of TSEs of humans and ruminants

4. WHO TSE Consultation 14-16 September 2005: Report Issued 2006 Executive summary Recent scientific developments  Epidemiology, clinical features, Dx criteria of CJD  BSE and scrapie  Risk of transmission of CJD/vCJD by human blood, blood products Recommendations  Tissue infectivity  Reducing risk to humans from biological and pharmaceutical products made with ruminant-derived materials  Tissue source  Tissue removal/processing  Production: vaccines, recDNA (from banked cells), other  Reducing risk to humans from biological and pharmaceutical products made with human-derived materials  Risk of transmitting vCJD by blood, blood products  Risk assessment  Risk reduction: product retrieval/market withdrawal, donor deferral, plasma products, &c.  Risk from HCT/Ps Conclusions Annexes 1.TSE infectivity (selected): humans, BSE cattle, scrapie sheep/goats 2.Summary of presentations

5. 26 Countries with BSE in Native Cattle [yr first reported & approx. total cases reported to OIE thro’ 07 Sept 2006] l UK 1986 (>184,431) [1202 ’01; 1144 ’02; 611 ’03; 343 ’04; 225 ’05; >61 ’06] l Ireland 1989 (1579) l Switzerland 1990 (461) l France 1991 (971) l Portugal 1994 (990) l Belgium 1997 (131) l Netherlands 1997 (80) l Luxembourg 1997 (1) l Liechtenstein 1998 (3) l Denmark 2000 (14) l Germany 2000 (389) l Spain 2000 (654) l Italy 2000 (132) l Greece 2001 (1) l Czech Repub 2001 (24) l Slovakia 2001 (23) l Japan 2001 (28) l Slovenia 2001 (6) l Finland 2001 (1) l Austria 2001 (5) l Poland 2002 (48) l Israel 2002 (1) l Canada 2003 (1UK+8) l USA 2004 (1 Canada+2) l Croatia 2006 (1) l Sweden 2006 (1)

6. Total Reported UK Exports of MBM 1986 –1995 ( unconfirmed review by UK authorities of export records) No data 0 - < 5 5 - < 10 10 - < 20 20 - < 100 100 - < 1.000 1.000 - < 10.000 > 10.000 Legend: (in tonnes)

9. Unique Pathology of vCJD (Chazot G & al. Lancet 1996;347:1181. Will RG & al. Lancet 1996:347:921-5. Hill AF & al. Lancet 1999;353:183-9)

10. BSE and vCJD deaths in UK Feed ban Transfusion Cases Reported 12/03 7/04 2/06

11. vCJD Non-UK cases = 34 ( Aug 2006 UK =162) mod from R. Knight/CJD SU Edinburgh FRANCE 20 (3?UK) ITALY1 NETHERLANDS2 PORTUGAL1 SPAIN1 REPUBLIC of IRELAND2  REPUBLIC of IRELAND2  USA2  CANADA1  JAPAN1  SAUDI ARABIA (origin uncertain) 1  Probably acquired in UK Probably non-UK origin

12. > 360 Iatrogenic Transmissions of CJD/vCJD by Classes of FDA-regulated Medical Products of Human Origin RBC: the only new class of medical product CJD-implicated past 10 yr ProductCases Incub’n Average Incub’n Range Cornea  3 3 ? 16 mo (excluding outlier) 13, 18, 320 mo Dura mater> 16812 yr1.3-22 yr Pit hormones Growth Gonadotropin > 180 4 12 yr 13 yr 5-39 yr 12-16 yr Neurosurgical instruments (includes electrode) 6 (? 7)19 mo15 – 28 mo RBC (UK vCJD)3> 6 yr7.8 yr, 6.5 yr, (> 5 yr)

13. WHO Consultation Suggested Nomenclature: PrP TSE for All Abnormal Forms of Prion Protein (PrP) Associated with Transmissible Spongiform Encephalopathies PrP TSE  PrP Sc scrapie-type PrP, or PrP res proteinase-K-resistant PrP, or PrP d disease-associated PrP Solubility in Detergent-Salt SolubleSedimented PK- sensitive PrP C PrP sen sPrP Sc PK- resistant ? rPrP Sc PrP res PrP d

14. Proceedings of a Consultation Convened by WHO Geneva 14-16 September 2005 Selected Highlights … [W]hen feasible, tissues or body fluids of ruminant origin should be avoided in the preparation of biological and pharmaceutical products. When bovine materials must be used, they should be obtained from sources assessed to have negligible risk from the infectious agent of BSE. Most bovine tissues, including bovine muscle, used to manufacture biologicals, if carefully selected by taking into account the geographical distribution of BSE and collected according to guidelines, have little risk of contamination with BSE agent … No country except UK and Ireland has reported more than 1500 BSE cases. Recent findings of disease-associated proteins in muscles of sheep with scrapie (… not known to infect humans) and the recognition of BSE itself in a goat, reinforce the need for manufacturers of biologicals to maintain … precautionary safety measures …

15. Proceedings of a Consultation Convened by WHO Geneva 14-16 September 2005 Selected Highlights In naturally affected cattle, BSE infectivity, detected by assay in mice, has been demonstrated only in brain, spinal cord and retina … [and] in a pool of nictitating membranes but not in pools of lymph nodes or spleen. Recently infectivity was detected in some peripheral nerves and a solitary muscle, of a single case of BSE in a German cow … In cattle experimentally exposed by the oral route, BSE infectivity has been detected by mouse assay in the distal ileum through much of the disease course from six months post exposure and in the … CNS … and sensory ganglia of the peripheral nervous system from late in the incubation period. Infectivity has also been found in palatine tonsil, at a single time point … only by assay in cattle and not by the mouse assay. BSE has been experimentally transmitted via the oral route to sheep and goats. [T]here is recent evidence that one goat [and perhaps another] has been naturally infected with BSE.

16. Proceedings of a Consultation Convened by WHO Geneva 14-16 September 2005 Selected Highlights [U]nder specific experimental conditions, the brains of some TSE- affected rodents may be infectious by bioassay while PrP TSE remains undetected. [I]mmunoassays have detected PrP TSE in the brains of BSE-infected cattle at least three months before onset of clinical illness. However, no immunological method has yet been validated to be sufficiently sensitive to detect PrP TSE in the blood of infected animals or humans, though promising initial results have been reported by several groups of investigators … Transfusion experiments have not been conducted in cattle … [however, s]tudies using small amounts of blood components or spleens of cattle with BSE assayed in mice and cattle injected by the most effective routes failed to detect infectivity. A conservative regulatory approach would assume that bovine serum might potentially contain TSE infectivity—presumably in small amounts.

17. Proceedings of a Consultation Convened by WHO Geneva 14-16 September 2005 Selected Highlights Ruminant blood and blood derivatives, such as fetal calf serum in cell cultures media and bovine serum albumin stabilizers … [have] not been identified as a source of infection, and properly collected fetal bovine serum has a negligible risk. However, blood of sheep with experimental BSE or natural scrapie can be infectious and, because scrapie and BSE agents behave similarly in sheep and goats, the blood of small ruminants should either be avoided in preparing biologicals or selected very carefully from sources known to be free of TSEs. There is a continuing need to ensure that all national regulatory authorities with limited resources have ready access to reliable and up- to-date information when assessing TSE risks and evaluating [medicinal] product safety.

18. Proceedings of a Consultation Convened by WHO Geneva 14-16 September 2005 Results of National Risk Assessments for vCJD and Blood UK (M Turner, P Bennett)  Labile components: 1/120,000 transfusions may be from donors incubating vCJD, but—if strict indications for transfusion observed—benefits clearly exceed risk  Plasma-derived products: minimal risk  Surgical/dental instruments, HCT/Ps: highly uncertain risk France (J-H Trouvin): Independent assessment yielded results very similar to those for UK (for both labile components and plasma-derived products) Australia (A Farrugia): FVIII has highest risk among PDs Canada (S ElSaadany): Due to uncertainties, risk communication is difficult Germany (J Löwer)  Under realistic conditions for Germany, vCJD should not become an endemic infection maintained only by blood transfusion.  Deferring transfused donors only slightly reduces vCJD risk in Germany.  German conclusions should not be applied to countries with different BSE/vCJD risks. USA (D Scott): Due to uncertainties in assumptions, risk assessments to date allow no confident predictions regarding probability of vCJD infections and clinical illnesses in individuals exposed to various blood components and plasma derivatives.

19. WHO TSE Consultation Report: Annex 1 TSE Infectivity and PrP TSE Detected in Human and Animal Tissues, Other Materials (WHO tables converted to tile table by OA Maximova, CBER, FDA) IA. High Infectivity  CNS tissues with high titers of infectivity in late stages of all TSEs  Certain tissues anatomically associated with CNS IB. Lower Infectivity  Peripheral tissues with infectivity and/or PrP TSE in at least one TSE IC. No Detected Infectivity Table Entries

20. Table IA: High-infectivity Tissues CNS tissues with high titres of infectivity in TSEs and Certain tissues anatomically associated with CNS

21. Table IB: Lower-infectivity Tissues Peripheral tissues testing positive for infectivity and/or PrP TSE in at least one form of TSE

22. Table IB: Lower-infectivity Tissues (Cont.) Peripheral tissues testing positive for infectivity and/or PrP TSE in at least one form of TSE

23. Six Candidate PrP TSE Blood Screening Tests: Early Results WHO TSE Consultation Sept 2005 Sensitivity = results with samples from known infected animals or human CJD cases Specificity = results with samples from uninfected animals or negligible-CJD-risk pts Spike LOD determined from last brain or spleen dilution detected and known infect titer NoNo Test DescriptionModel Sens (+/total) Spec (F+/total) Spike LOD 1 Palindromic peptide binding to PrP TSE (no PK) Scr sheep52/520/451 ID 50 /ml Scr hamster43/430/23 BSE cow36/401/40 CJD mouse5/5 CJD monkey8/80/4 CJD human14/140/53

24. Six Candidate PrP TSE Blood Screening Tests:Early Results WHO TSE Consultation Sept 2005 Sensitivity = results with samples from known infected animals or human CJD cases Specificity = results with samples from uninfected animals or negligible-CJD-risk pts Spike LOD determined from last brain or spleen dilution detected and known infect titer NoNo Test DescriptionModel Sens (+/total) Spec (F+/total) Spike LOD 2 Conc PrP TSE, PK, ppt w/ streptomycin capture w/calix arene, detect w/ sandwich ELISA CJD human7/100/500 3 Peptides on mag beads bind to PrP TSE, no PK, detect w/ sandwich ELISA Scr sheep3/83 ID 50 /ml 4 PMCA: ampl of PrP TSE by repeated add’ns of nl tissue extracts, sonications, PK, Western blot Scr hamster 20 da pi 100 da pi 3/6 16/18 0/12 5 Proprietary ligand on mag bead, no PK, elute, denature, ELISA Scr sheep2/30/26100 ID 50 /ml 6 Capt w/ IgM mAb to PrP TSE, sand ELISA or FACS Scr sheep BSE cow 6/6 3/12 0/16 0/10

25. Table IC: Tissues with No Detected Infectivity Tissues examined for infectivity and/or PrP TSE with negative results

26. Limitations of Negative TSE Infectivity and PrP TSE Studies Small numbers of human cases and animals studied Small numbers of bioassays attempted Small volumes of materials sampled Limits of detection usually unknown  Bioassay animal species: some relatively insensitive  PrP TSE assays: some relatively insensitive Very limited numbers of infected animals studied during incubation periods Infected humans rarely if ever identified and studied before onset of overt disease Uncertain relevance of TSE models to each other

27. WHO Guidelines on Tissue Infectivity Distribution in TSEs Partial Credits Consultation Chairman WG van Aken, Netherlands Draft Report DM Asher, USA J-P Deslys, France E Griffiths, Canada A Padilla, WHO R Bradley UK M Pocchiari, Italy RG Rohwer, USA J-H Trouvin, France GAH Wells, UK RG Will, UK Annex 1 Working Group P Brown, USA (chairman) R Bradley, GAH Wells, UK (animal TSEs)