Update on the Replacement of the HCV RNA International Standard

1. Update on the Replacement of the HCV RNA International Standard Sally Baylis & Alan Heath, NIBSC SoGAT XX, Warsaw 12-13 June 2007

2. Replacement of the 2nd International Standard for HCV RNA (96/798) • Proposal made at SoGAT 2005 to replace the HCV RNA International Standard as requested by WHO • Agreement that HCV 1a genotype would be sourced & would be anti-HCV negative and diluted in plasma rather than cryosupernatant

3. Candidate Standards for Evaluation • 3 anti-HCV negative window period genotype 1a donations have been obtained • The genotype of each has been confirmed by LiPA & DNA sequencing • Absence of other viral markers confirmed in these stocks • Material has been freeze-dried in two batches • Batch 1 (NIBSC Code 06/100), 2085 vials, fill CV = 0.62% • Batch 2 (NIBSC code 06/102), 2100 vials, fill CV = 1.82%

4. Panel for Collaborative study • Sample 1 – 2nd International Standard for HCV RNA (96/798) • Sample 2 - new freeze-dried preparation 06/100 • Sample 3 - new freeze-dried preparation 06/102 • Sample 4 – liquid/frozen bulk material (NIBSC code 06/118) that was used to prepare Samples 2 & 3

5. Collaborative Study • Collaborative study commenced in November 2006 • Participants requested to test samples in four independent assays • 33 laboratories from 14 countries participated in the study • 40 data sets received; 25 from quantitative assays & 15 from qualitative assays • NIBSC collated & analysed data

6. Types of Quantitative Assays Used by Participants

7. Types of Qualitative Assays Used by Participants

8. Estimated IU/ml (log10) from Quantitative Assays N - Number of laboratory estimates SD - Standard Deviation of log10 estimates across laboratories

13. Estimated PCR Detectable Units/ml (log10) from Qualitative Assays N - Number of laboratory estimates SD - Standard Deviation of log10 estimates across laboratories

17. Overall Mean Potencies (log10 IU/ml) Relative to Sample 1 N - Number of laboratory estimates SD - Standard Deviation of log10 estimates across laboratories

21. Accelerated Degradation Studies Sample 2 • Relative potencies of accelerated degradation samples with respect to Sample 2 stored at -20ºC (log10 drop), qPCR analysis • Stability appears similar to 96/790 and 96/798; studies on-going • 96/798 from this study has not lost potency after >10 years storage at -20 ºC

22. Proposal for 3rd International Standard for HCV RNA • Real-time & accelerated degradation data indicate that Sample 2 (06/100) is stable & suitable for long term use • Propose that 06/100 be established as the 3rd International Standard for HCV RNA with a unitage of 5.19 log10 IU/ml (4.89 log10 IU/vial) • Report will be submitted to ECBS by July

23. Acknowledgements Nita Shah & David Padley NIBSC Professor Ewa Brojer, Institute of Hematology & Blood Transfusion, Warsaw Dr Richard Smith, National Genetics Institute, Los Angeles Dr Vivienne James, HPA, Colindale Dr Michael Chudy, WHO, Geneva Collaborative study participants

24. HCV RNA Collaborative Study Participants • S Baylis, N Shah South Mimms, UK • T Cuypers M Koppelman Amsterdam, The Netherlands • C Defer Lille, France • L Fanning Cork, Eire • G Gesu Milan, Italy • A Goglio Bergamo, Italy • P Grant London, UK • P Iudicone Rome, Italy • L Jarvis, A Cleland Edinburgh, UK • C Jork Springe, Germany • S Kerby Bethesda, USA • A Klotz, M Gessner Vienna, Austria • R Madej Pleasanton, USA • B Miller Karlsruhe, Germany • S Mizusawa Tokyo, Japan • C Negru Timişoara, Romania • M Nübling Langen, Germany

25. HCV RNA Collaborative Study Participants • JM Pawlotsky Créteil, France • M Pecorari Modena, Italy • G Pisani Rome, Italy • A Portela Moreira Madrid, Spain • S Sauleda Barcelona, Spain • H Schindl Vienna, Austria • G Schneider Des Plaines, USA • A da Silva Lisbon, Portugal • R Smith Los Angeles, USA • S Tassen Brentwood, UK • P Taylor London, UK • A van der Zanden Apeldoorn, The Netherlands • YC Yang Taipei, Taiwan • S Yerly Geneva, Switzerland • M Zaccanelli Treviglio, Italy

26. Update on the Stability of the HAV RNA International Standard Sally Baylis, Nita Shah & Alan Heath, NIBSC SoGAT XX, Warsaw 12-13 June 2007

27. Background – Hepatitis A Virus RNA IS • The 1st International Standard for HAV RNA (00/560) and a second batch (00/562) were prepared in Q1, 2001 • 00/560 was evaluated in an international collaborative study, with data returned from 16 laboratories • The mean log10 equivalents/ml were 5.29 (00/560) and 5.07 (00/562) • 1st International Standard (00/560) was established in 2003 with 50,000 IU/vial • ECBS noted that 00/562 may be a replacement for 00/560

28. Stability Data 00/560 & 00/562 • Accelerated thermal degradation samples for 00/560 and 00/562, stored at +4ºC for ~6 years were tested in parallel with samples of 00/560 stored at -20 ºC • 00/560 – a drop on 0.1 log10 • 00/562 – a drop on 1.0 log10 • Conclude that 00/560 is highly stable

29. Proposal to Replace the Parvovirus B19 DNA International Standard Sally Baylis, NIBSC SoGAT XX, Warsaw 12-13 June 2007

30. Replacement of the Parvovirus B19 DNA IS 99/800 • The 1st International Standard for B19V DNA was established by the WHO ECBS in October 2000 • Estimated date of exhaustion of the IS will be 2008/9 at current rate of usage • Materials coded AA (99/800) & BB (99/802) showed no significant difference in potency in the collaborative study • ECBS noted that BB (made from the same stock as AA) could be reserved for potential future use as a replacement standard

31. Candidate 2nd International Standard • Materials coded AA (99/800) & BB (99/802) showed no significant difference in potency in the original collaborative study • ECBS noted that BB (prepared from the same stock as AA) could be reserved for potential future use as a replacement standard • Current study designed to demonstrate the equivalence of the candidate replacement (BB) to AA • Real-time data on samples AA & BB • Accelerated degradation data for samples AA & BB