Laboratory of Cellular Hematology

1. Laboratory of Cellular Hematology Platelet efficacy and safety section Jaro Vostal, M.D., Ph.D.

2. red cells Product Review in the Laboratory of Cellular Hematology additive solutions anticoagulants storage bags platelet substitutes platelet testing devices platelets granulocytes platelet-derived products apheresis devices leukoreduction filters sterile connecting devices

3. Current projects in the platelet efficacy and safety section • Development of an animal model to assess efficacy of human platelets • Modeling of risk from transfusion transmitted spongiform encephalopathy diseases (TSE)

4. Development of an animal model to replace human volunteers in evaluation of novel platelet products • In vivo recovery and survival of radiolabeled platelets in human volunteers is the current “gold standard” in evaluating platelet efficacy • Damaged platelets are quickly cleared from circulation • These studies are cost-prohibitive, logistically difficult and pose a risk to the volunteers • CBER Mission Relevance: An alternate method of detecting damage to platelets would reduce the time and cost of product development

5. Human platelet clearance from SCID mouse circulation • Severe Combined Immunodeficiency (SCID) mice • Lack T and B cells • Do not reject allogeneic tissue • Natural killer cells and splenic macrophages function normally

6. Mouse model to evaluate efficacy of human platelets Storage under inadequate conditions Day 1 Wild type mice SCID mice Day 7

7. Transmissible spongiform encephalopathies (TSE) • Neurodegenerative disease with long asymptomatic phase and rapid terminal course • Asymptomatic but infected donors could donate transfusion products • Concerns about transfusion transmission • 3 out of 66 recipients of transfusion products from donors that later developed vCJD were diagnosed with vCJD (all 3 received RBCs) • No therapy and no diagnostic test available • Normal prion protein (PrPc) conversion to a pathogenic form (PrPscrapie)

8. GPIIbIIIa PrPc CTRL GPIIbIIIa CTRL PrPc Fluorescence intensity Lack of TSE infectivity and prion protein expression in hamster platelets PrPc on human platelets PrPc GPIIbIIIa PrPc PrPc on hamster platelets

9. PrPc expression on blood cells of different species *

10. Expression of Surface PrPc on Blood Cells of Primates ( MAb 6H4) Molecules per cell Species RBC PLTS LYMPHO MONO GRANULO Human (6)* 200 600 11,000 10,000 600 Chimpanzee (3) 3600 600 1000 9000 100 Cynomolgus (5) 30 10 4000 6500 30 Rhesus monkey (4) 4000 100 1900 3500 20 Squirrel monkey (3) 1100 5 1000 4000 300 Microcebus (4) 0 10 2000 n.d. n.d.

11. Relevance to FDA/CBER mission • Defining the blood cells associated with TSE infectivity in transfusion products will help in focusing detection method development • Distribution of cell PrPc expression may correlate with infectivity distribution and could identify the appropriate animal for modeling transfusion risk

12. Future Plans • SCID mouse model • Further develop and validate for evaluation of human platelet efficacy • Compare survival of normal and pathogen reduction treated platelets, cold stored platelets, platelet substitutes • TSE disease transmission by blood transfusion • Infectivity study in mice to identify role of red cells as carriers

13. Acknowledgements LCH Staff Collaborators Jay Lozier, MD, PhD CBER, FDA Robert Rohwer, PhD Baltimore VA Paul Brown, MD NIMH David Asher, MD CBER, FDA Neal Young, MD NHLBI Platelet in vivo clearance Nat Wolins, PhD John Piper, PhD Prion Studies Karel Holada, PhD Jan Simak, PhD Jan Zivny, MD, PhD Monique Gelderman, PhD