REVIEW OF PLASMA PRODUCTS FOR TRANSFUSION

1. REVIEW OF PLASMA PRODUCTS FOR TRANSFUSION March 17, 2005 Mark Weinstein, Ph.D. Office of Blood Research and Review CBER, FDA

2. Issue FDA seeks advice from the Committee on the extent to which the available scientific data may support: • Changes to further standardize processing of plasma products for transfusion • The need for additional scientific studies that would be helpful to resolve current areas of uncertainty.

3. Background • Currently, plasma products for transfusion are poorly defined and characterized in the CFR and the AABB Circular of Information. • Scientific uncertainty exists on the extent to which conditions of plasma product preparation may affect the final products. • We therefore are seeking to evaluate data acquired from all available sources to consider the possible development of minimal standards for these products that would further ensure their clinically relevant safety, purity and potency. • 2004 FDA workshop on plasma standards identified multiple parameters that can affect the composition and potentially the quality of plasma products for transfusion • time and temperature of separation of plasma from cells • the anticoagulant used • freezing, storage, thawing, and post thawing conditions • This session will consist of a review of the key literature on plasma processing and a presentation on the clinical use of plasma products.

4. Topics • Issue • Safety and quality of products • Plasma products and their uses • Potential areas of improvement • Focus on temperature-related issues of preparing plasma transfusion products

5. What’s the Problem? • Current standards may not reflect current scientific knowledge. • Side effects and hazards of plasma for transfusion products (AABB circular) • Immediate immunologic complications:febrile nonhemolytic reactions, allergic reactions, anaphylactoid reactions, transfusion-related acute lung injury (TRALI) • Delayed immunologic complications:alloimmunization to antigens of red cells, white cells, platelets, or plasma proteins. • Transmission of infectious agents: virus, bacteria, prions • Circulatory overload, hypothermia, or metabolic complications. • Plasma products may not be made in a way that will minimize risks and exposure to product.

6. What’s the Current Situation?

7. Product Definitions (CFR, AABB Circular): Fresh Frozen Plasma • [21 CFR 640.32 (a)] When whole blood is intended for Plasma, Fresh Frozen Plasma, and Liquid Plasma, it shall be maintained at a temperature between 1 and 6 oC until the plasma is removed. [Note: FFP can also be made from plasma separated within 8 h of collection from Whole Blood that has been maintained at room temperature“Eight-Hour Hold; Blood and Plasma Inspectors – memo 11/13/89” ] • [21 CFR 640.34 (b)] The plasma shall be separated from the red blood cells, and placed in a freezer within 8 hrs or within the timeframe specified in the directions for use for the blood collecting, processing, and storage system, and stored at <-18 oC • [AABB Circular] Consists of the fluid portion of blood that is separated and placed at <-18 oC or below within 8 h of collection of whole blood if the anticoagulant is CPD….Plasma collected in ACD … must be placed at <-18 oC within 6 h. Plasma components may be prepared from whole blood collection or by apheresis

8. Factors that Could Affect Plasma Quality • Time, temperature of plasma contact with cells • Anticoagulant • Residual cellular content • Rate of freezing (affected by the following) • Volume of plasma to be frozen • Shape of container • Freezing device, e.g., freezer, cold room, blast freezer

9. Definition of Other Plasma Transfusable Products (AABB Circular) • Cryoprecipitate: Prepared by thawing FFP between 1-6 oC and recovering the precipitate. Each unit should contain > 80 IU FVIII and >150 mg fibrinogen in approximately 15ml plasma • 21 CFR 640.54 (a) (3) (b) “…AHF shall be separated from the plasma by a procedure that has been shown to produce an average of no less than 80 units of antihemophilic factor per final container” • Plasma, Cryoprecipitate Reduced: is prepared from FFP that is thawed and centrifuged, with the cryoprecipitate removed by centrifugation. • Thawed Plasma is derived from FFP prepared in a way that ensures sterility (closed system), thawed at 30-37 oC, and maintained at 1-6 oC for 1-5 days. A product not in the CFR, not a licensed product. • Liquid Plasma is separated no later than 5 days after the expiration date of the Whole Blood. Plasma may be stored at <-18 oC. Liquid plasma is stored at refrigerator temperature 1-6 oC. • Plasma frozen within 24 hours after phlebotomy: must be separated and placed at <-18 oC within 24 hours of whole blood collection.

10. Labeled Uses of FFP (AABB) • Management of preoperative or bleeding patients who require replacement of multiple plasma coagulation factors (e.g., liver disease). • Patients on warfarin who are bleeding or need to undergo an invasive procedure… • Patients with massive transfusion who have clinically significant coagulation deficiencies • For transfusion or plasma exchange in patients with thrombotic thrombocytopenic purpura (TTP) • Manage coagulation factor deficiencies, congenital or acquired, for which no specific coagulation concentrates are available • Manage rare specific plasma protein deficiencies, such as C-1-esterase inhibitor and FV. • Source material for Cryoprecipitate, Cryopoor plasma, and thawed plasma

11. Labeled Uses of Other Transfusable Plasma Products (AABB circular) • Cryoprecipitate: Provides FVIII, fibrinogen, vWF, FXIII. Used as second-line therapy for vWD and hemophilia A. Control of bleeding associated with fibrinogen deficiency, and to treat FXIII deficiency • Plasma, Cryoprecipitate Reduced: Provides for defective or deficient plasma proteins except fibrinogen, FVIII, vWF, FXIII. Used for TTP refractory to FFP. • 24 h Plasma; thawed plasma; Liquid plasma:Serve as a source of defective or deficient plasma proteins except for FV and FVIII. Indications same as for FFP except not to be a source of FV or FVIII.

12. Factors Affecting Quality of Transfusable Plasma Products Blood drawing technique; adequate mixing of anticoagulant with plasma Appropriate technique is needed to avoid activation of coagulation and other proteins Anticoagulant: CPD or ACD Literature is mixed on whether collection in ACD results in less FVIII than CPD. Anticoagulant-related differences in plasma protein activity occur over time in plasma stored at 6 oC. Residual cells in plasmaCells and cellular components in plasma have the potential of causing immunological reactions and release of proteolytic enzymes, that could lead to some side effects seen in plasma product transfusions. There are no standards on cellular content of plasma products, but plasma products must be ABO compatible. Plasma separation techniques vary widely.

13. Factors Affecting Quality of Transfusable Plasma Products Time, Temperature from draw to freezing Allowable times and temperatures vary among different regulatory bodies with varying degrees of scientific justification. Conditions optimal for plasma quality may not be optimal for cellular preservation. Plasma contact with cellular components – Temperature effectsCold activation of the coagulation and plasma kinin system is of concern. FVIII and vWF can be reduced by significant amounts Rate of freezing plasma, thaw and post thaw conditions

14. Plasma Contact with Cellular Components:Cold Activation [Favaloro, 2004] • Experiment: 39 donors, Whole Blood into 3.2% NaCit tubes • Aliquot centrifuged immediately, plasma frozen –80 oC • Aliquot whole blood held at 4oC, 3.5 h • Aliquot whole blood held at 22oC, 3.5 h • Separate plasma; test for FII, V, VII, VIII, IX, X, XI, XII, vWF:Ag, vWF:CB • Result: No effect of temperature on FII, V, VII, IX, X, XI, XII • For plasma from WB kept at 4oC, 3.5 h: • 25% loss of FVIII • 50% loss of vWF:Ag (Non-O blood) • 60% loss of vWF:CB (Non-O blood) • True for plasma from blood collected in CPD or ACD? • Should the time and temperature of WB before plasma separation be more narrowly defined?

15. Anticoagulant Affects Activity of Proteins in Stored Plasma Blomback, 1984

16. Focus on Freezing Plasma • FDA sponsored a workshop on August 31-September 1, 2004 to • review scientific data, • regulatory requirements, • current industry practices regarding the freezing, storage, and shipping of plasma. • Purposes: • help develop regulatory standards for recovered plasma • examine the potential of harmonizing regulations with Europe

18. Comparison of US and COE Standards and Recommendations: Issues to Consider • What is the scientific basis for differences in freezing conditions between US and COE? • What are the advantages and disadvantages of rapid freezing? • Product use? • Product need? • Practical implications?

19. Parameters Affecting Plasma Quality Plasma Separation from WB Method Time Temperature Cell content Whole Blood ACD, CPD? Rate of Freezing Plasma unit Storage time, temperature Thaw Method Cryoppt Cryosup FFP recovered plasma “thawed plasma”

20. Time to Freeze 700 ml, 1500 ml Plasma Under Varying Conditions Carlebjork, 1986

21. Factor VIII Recovery in FFP Affected by Freezing Rate Carlebjork, 1986

22. FVIII Activity in FFP After “Slow” and “Rapid” Freeze Akerblom, 1992 Samples: 200ml, N = 10, prepared within 2 hr of blood draw Rapid freeze: -40 oC, <40 min Slow freeze: place in -40 oC or -20 oC cabinet freezer; time to freeze not reported

23. Effect of Freezing of on FFPAkerblom, et al, 1992 • Rapid freeze(-40 oC, <40 min): loss of about 8% FVIII from pre to post freeze; 9% increase in beta-TG (platelet lysis) but not statistically significant • Slow freeze, (-20 oC): 20% loss FVIII; 8% loss FV; 14% increase in beta-TG • Slow freeze, (-40 oC): 14% loss FVIII; freeze rate more important than final temperature • No difference between fast and slow freeze in FVII, vWF, soluble fibrin, TAT complex, C1-esterase inhibitor

24. Parameters Affecting Plasma Quality Plasma Separation from WB Method Time Temperature Cell content Whole Blood ACD, CPD? Rate of Freezing Plasma unit Thaw Method Storage time, temperature Cryoppt Cryopoor FFP recovered plasma “thawed plasma”

25. Effect of Freezing Rate of FFP on Cryoprecipitate/supernatant Production Samples: 200 ml, N=6, prepared within 3 hr of donation, thaw siphon process to make cryoppt Slow freeze: -40 oC freezer, -0.33 oC/min Rapid freeze: - 70 oC Liquid N2/ ETOH, -5 oC/min Note: VIIIR:Ag = vWF Farrugia, 1985

26. Effect of Freezing of on FFP, CryopptFarrugia, et al, 1985 • Total recoverable FVIII activity, (cryosupernatant plus cryoprecipitate) not significantly different between slow and fast freeze. This suggests that the FVIII activity in FFP might not differ significantly between the two freezing rates. • More FVIII (25%), vWF (24%), and fibrinogen (13%) were in cryoprecipitate made from fast frozen than from slow frozen plasma, using the thaw siphon methodology to prepare cryoprecipitate. • Potentially, under these fast freeze conditions, less cryoppt and cryopoor plasma would be needed to achieve the desired outcome

27. FVIII in FFP: Fast and Slow FreezeInterpret Data with Caution! Slow freeze Fast freeze Sample: 200ml plasma, n = 10 Fast: -30oC, fluorocarbon bath Slow: -30oC, cold room Thaw 37 oC, store 22 oC Farrugia, 1992

28. Under Certain Conditions, “Fast” and “Slow” Freeze of FFP has Little Effect on FVIII, vWF, and Fibrinogen in Cryoprecipitate Farrugia, 1992

29. Considerations • Need to define “fast” and “slow” freeze conditions. • Increasing the rate of freezing plasma decreases the loss of FVIII activity up to a point, beyond which the rate has little effect. • The amount of FVIII, vWF, and fibrinogen in cryoprecipitate is highly dependent on the thaw method as well as the freezing rate. • Under defined conditions there is little advantage in freezing to -30 oC within 1 h compared to 2 h.

30. Parameters Affecting Plasma Quality Plasma Separation from WB Method Time Temperature Cell content Whole Blood ACD, CPD? Plasma unit Rate of Freezing Storage time, temperature Thaw Method Cryoppt Cryopoor FFP recovered plasma “thawed plasma”

31. No Change in FVIII and Other Proteins in FFP Stored at –20 oC vs – 40 oC for 36 Months Kotitschke, 2000

32. Effect of Storage Temperatures on FVIII -74oC 5 ml aliquots of plasmapheresis plasma Frozen and stored at -74oC (x) Frozen and stored at - 24 oC (o) - 24 oC Woodhams, 2001

33. Effect of Temperature Cycling of Plasma on Fibrinogen Content in Cryoprecipitate Plasma temperature cycling Farrugia, 1985

34. Considerations • Most, but not all studies suggest that rapidly frozen FFP can be stored at –20 oC for more than one year without significant changes in the activity of certain proteins, including FVIII. Current standards specify one year storage for FFP. • Temperature fluctuations in frozen FFP can affect cryoprecipitate composition. Current regulations for FFP do not permit deviations above –18 oC. • Should these standards be reconsidered?

35. Challenges in Evaluating Data • Comparison of study results is complicated because of differences in measured parameters, and assay techniques • Laboratory studies may not reflect practically achievable conditions • Conditions optimal for production of one product may not be optimal for others

36. Requests to the Committee • Please discuss the extent to which the available literature on plasma processing may support changes to improve the clinically relevant safety, purity, potency or consistency of various plasma products for use in transfusion, e.g. time to plasma separation, time from collection or separation to freezing, freezing rate and target temperature, storage temperature, allowed temperature excursions, cellular content. • What additional scientific studies are needed? • What recommendations do you have for the next steps forward?