1. FDA’s Current Considerations of Parvovirus B19 Nucleic Acid Testing (NAT) Mei-ying W. Yu, PhD Division of Hematology CBER/FDA Extraordinary SoGAT Meeting on Parvovirus B19 NIBSC, UK; March 2, 2007

2. FDA’s Discussion of Parvovirus B19 NAT (I) Sep 1999 Blood Products Advisory Committee (BPAC) –Safety of Pooled Plasma S/D Treated correlated with those lots having <10 4 geq/mL of B19 DNA in manufacturing pool (a phase 4 study). –For plasma for further manufacturing, B19 NAT screening was recommended as an in-process test. –In-date (+) blood components were recommended to be quarantined and destroyed when possible. Dec 1999 FDA NAT Workshop –The possibility of limiting <10 4 geq/mL of B19 DNA in all manufacturing pools was discussed.

3. FDA’s Discussion of Parvovirus B19 NAT (II) Dec 1999 NHLBI Parvovirus B19 Workshop –FDA’s in-process B19 NAT standard for plasma for further manufacturing was presented. Dec 2001 FDA NAT Workshop –Proposed limit: <10 4 IU/mL of B19 DNA in all manufacturing pools due to the WHO Std availability Mar 2002 BPAC –FDA’s current thinking on B19 NAT for Blood and Plasma Jul 2002 Ad Hoc PHS Panel –Medical benefits to donors and close contacts

4. FDA’s Discussion of Parvovirus B19 NAT (III) Dec 2002 BPAC –B19 NAT testing of Whole Blood (WB) donations would reduce the risk to transfusion recipients if high-titer positive units (≥10 6 geq/mL) of WB and its components were withheld from use. Based upon current limitations, a threshold value was however considered not yet sufficiently established to warrant screening of WB donors. –Temporary deferral of high-titer positive donors is warranted only for apheresis donations to make transfusion components but not WB or Source Plasma donations (due to either donation frequency or resolution time) –Potential medical benefits to close contacts of B19 infected donors warrant notification of high-titer donors. But only if donor notification can be completed within several weeks of donations.

5. In-Process B19 NAT Testing of Plasma for Further Manufacturing (I) Minipool NAT testing (Pool size: 96 – 512) –Most Source Plasma fractionators are performing high- titer B19 minipool NAT screening by in-house methods. –Sensitivities of NAT assays differ, but most tests will exclude original donations exceeding a B19 DNA level of 10 5 to 10 7 IU/mL. –Reactive minipools are resolved to single reactive donations, which are rejected. –Blood collection establishments voluntarily retrieve and discard in-date components to prevent their use in transfusion when a donor of plasma is found to have high titer B19. Little is known about the minimal infectious dose for B19. However, we reported that infusion of 2 X10 4 IU of B19 DNA in a FVIII product devoid of anti-B19 IgG infected a seronegative individual (Wu et al, Transfusion 2005).

6. In-Process B19 NAT Testing of Plasma for Further Manufacturing (II) Manufacturing pool NAT testing –To ensure that high-titer plasma donations are removed as a result of minipool NAT screening, the viral load in manufacturing pools destined for plasma derivatives is measured directly and can be limited to ≤10 4 IU/mL of B19 DNA, the standard proposed by the FDA. Anti-B19 in large pools complexes/neutralizes virus Some manufacturing procedures clear 4 logs of the virus when validated using a model parvovirus.

7. In-Process B19 NAT Testing of Plasma for Further Manufacturing (III) FDA has reviewed and approved some in-house B19 NAT procedures for minipools and manufacturing pools under Biologics Licensing Applications (BLAs) or their supplements for plasma derivatives. –B19 NAT testing is validated as an analytical procedure with respect to sensitivity, specificity, and reproducibility. –Primers and probes used for B19 NAT testing should be chosen so that all B19 genotypes can be detected.

8. Issues To Be Considered Because of known risks to those individuals who are pregnant, chronically anemic, and immuno-compromised, B19 NAT testing may be changed to donor screening testing if commercial tests are available as kits and the resolution time to confirmation (i.e., minipool single (+) donation) is short, similar to NAT for HCV/HIV/HBV. –Currently the mean resolution time by industry is long (25 to 60 days, PPTA presentation at Dec 2002 BPAC). –Unresolved issues “Up-front” donor screening Lookback –retrieval of in-date products; notification of recipients