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Labeling Claims for TSE Reduction Studies with Blood Processing Filters

Jaro Vostal, M.D., Ph.D.

Division of Hematology, OBRR

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Normal prion protein (PrPc)

Conversion of PrPc to PrPsc

Pathologic prion protein (PrPsc) Protease resistantProtease resistantless solubleless soluble

Accumulated PrPscand neurotoxicity

Altered conformations of PrPc

TSE infectivity present

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spleen

stomach

CNS neur

on

CNS

Transport of TSE from peripheral inoculum to the CNS

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AsymptomaticBSE infected sheep Healthy sheep

BSE infected sheep

Blood transfusion

BSE transmission by blood transfusionHouston, F. et al. Lancet 2000

Hunter, N. et al. J. Gen Virology, 2002

XX

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Epidemiological evidence for vCJD transmission by blood transfusion in humans

Llewelyn, C.A. et al, Lancet 363: 417-421, 2004Peden, A.H. et al. Lancet 364: 527-529, 2004

• The UK National CJD surveillance system identified individuals (48 total, 17 alive) who received blood products from 15 donors who later became diagnosed as vCJD cases

• 2 living recipients were subsequently diagnosed with vCJD– 1 developed and died from symptoms of vCJD– 1 died for unrelated cause and was unsymptomatic

for vCJD but had PrPres identified in spleen

• Both received non-leukoreduced red cells

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Issues to consider for validation of devices and processes used to reduce TSE

infectivity in human blood • Distribution of TSE infectivity in blood• Cell-associated

– Intracellular vs extracellular

• Free floating in plasma– Physical attributes of infectivity (aggregates, fibrils,

microvesicles)

• Interaction of the individual units of infectivity with the devices

• Distribution during and after processing– (microparticle generation)

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Steps to validate TSE reduction efficacy claims

• In vitro spiking studies of TSE infectivity into human blood– Detection of infectivity by surrogate

markers (PrPres) or by bioassay

• Endogenous TSE infectivity in an animal model– Detection of infectivity by bioassay

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Predictive value of animal models for human situation

• Comparability of animal blood to human blood– Cell type number, size and physical properties of

blood cells– Interaction of animal and human blood cells with

different materials

• Transmissibility or infectivity of a TSE agent may be influenced by– Strain of agent used – The dose of agent – Distribution of infectivity in blood – Distribution of normal prion ?

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Hematologic values for various species

Species RBC count x 106/ uL

RBC MCV (fL)

WBC

Count x 103/ uL

Platelet x 103/ uL

Sheep 10-13 35 4-12 500

Golden Hamster

6-7 60-70 5-10 500-800

Mouse 9-11 50 7-12 250-500

Human 4-5 86-98 4-11 150-450

Schalm, O.W. Veterinary Hematology, 2nd ed. Lea & Febiger, Philadelphia 1965Harrison, Principles of Internal Medicine, 17th ed. McGraw-Hill, 1994

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PrPc expression on blood cells of different speciesVostal, J.G. and Holada, K. Transf Med Rev. 15: 268-281, 2001

Species Platelets Erythrocytes Lymphocytes Monocytes Granulocytes

Human ++ + +++ +++ +/- Cat ++ - ++ ++ +/- Cow +/- - +++ + - Sheep - - ++ + - Hamster - - - - - Mouse - + +/- +/- +/-

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Validation of TSE infectivity reduction is only one part of the evaluation process

• TSE reduction devices also need to be evaluated for their impact on transfusion product quality

• Evaluation of red cells, platelets or plasma • FDA follows a standardized evaluation

approach to each transfusion product based on previous experience with devices that process transfusion products (e.g. leukoreduction filters)

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Leukoreduction: the process of reducing the total number of leukocytes in a transfusion

component

• Methods:filtration of blood products or collection of products by apheresis

• Use of leukoreduced blood products has been associated with reduction of – febrile non hemolytic transfusion reactions– Alloimmunization– Cytomegalovirus (CMV)

• No claims by manufacturers for their devices beyond meeting the criteria for leukoreduction– US <5x106 leukocytes per transfusion product– Europe <1x106 leukocytes per transfusion product

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Validation of Leukoreduction Filters-Efficacy

• Quantitation of leukocytes in particular blood product before and after filtration– Whole blood, red cells, platelets, plasma

• Define timing of leukoreduction from time of collection

• Explore effects of temperature on filtration efficacy (room vs cold temperatures)

• Validation specific for particular anticoagulant

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Validation of Leukoreduction Filters-Safety

• Biocompatibility and integrity of materials• Effect on cellular products

– In vitro recovery (85%)– Hemolysis at end of storage (<1%)– In vivo recovery of radiolabelled cells (platelets

and red cells)

• Plasma – – Levels and function of plasma proteins– Complement activation

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FDA proposal for validating a claim of reducing TSE infectivity in human blood products

• Demonstrate reduction of endogenous TSE infectivity by bioassay in two animal models (rodent and sheep)

• Use full scale blood unit and leukoreduction filter• TSE infectivity from BSE or vCJD strain• Reduction of PrPsc in blood product will be considered

supportive but not sufficient for a claim• Study performed at two separate sites to minimize issues

of cross contamination and differences in laboratory practice

• Study size sufficient to support statistically valid conclusion

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Question 1:Are the FDA’s proposed minimal criteria for validation of TSE infectivity reduction by filtration

adequate and appropriate?

Please comment on the following points • A) Rationale for the use of specific animal models to

study the properties of blood-borne TSE infectivity (Are experiments in rodents sufficient, or should experiments also be done in sheep?)

• B) Is it necessary that each experiment should be done at two separate laboratory sites (i.e. to ensure reproducibility, and accuracy of clearance)?

• C) General description of informative scaled-down processes for reducing TSE infectivity in blood

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Question 1:Are the FDA’s proposed minimal criteria for validation of TSE infectivity reduction

by filtration adequate and appropriate?Please comment on the following points

• D) Levels of clearance acceptable for claims of reduced TSE infectivity in blood components as used in clinical settings

• E) Estimated logs of clearance of TSE infectivity required to conclude that blood filters have effectively removed infectivity from blood components

• F) Methodology appropriate to use in evaluating TSE agent clearance (bioassays for infectivity, Western blot or other assay for prion proteins, other methods)

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Question 2: Does the FDA’s proposed labeling for a filter meet the appropriate criteria for a claim of reduction of TSE

infectivity in blood or blood components?

A) This filter (device) has been shown to reduce TSE infectivity in blood from an infected animal model.

orB) This filter (device) has been shown to reduce

transmission of TSE infectivity by transfusion in an animal model.

and (A+C or B+C) C) Due to lack of feasibility, studies have not been

performed to validate this claim in a human population.