World Health Organization International Biological Standards Elwyn Griffiths Biologics and Genetic Therapies Directorate

WHO’S BIOLOGICAL STANDARDIZATION PROGRAMME Develops Recommendations (Requirements) and Guidelines on the production and control of specific biologicals (Written standards) Develops and establishes International Biological Standards and Reference Reagents (Physical standards)

WHO’S BIOLOGICAL STANDARDIZATION PROGRAMME Work part of Constitution of WHO (1946) Inherited from League of Nations International Biological Standardization initiated under League of Nations in 1920s – Commission on Biological Standardization

FIRST BIOLOGICAL 1890’s Diphtheria antitoxin (Germany) Tremendous achievement France successful/ England not Attributed to “weak sera” Paul Ehrlich found answer – STANDARD ANTITOXIN PREPARATION - Units Standard used to calibrate future batches

INSULIN STANDARD Henry Dale (London) applied concept to insulin and other biologics (1920s) Banting & Best discovered insulin in Toronto but STANDARDIZATION CRITICAL for clinical usefulness Need for INTERNATIONAL oversight INTERNATIONAL STANDARD / INTERNATIONAL UNITS (IU) as measure of “strength” or “activity”

WHO Standard Setting Process Expert Committee on Biological Standardization (1 st meeting 1947) Biologicals Unit (Quality & Safety of Biologicals/Quality & Safety of Plasma Derivatives) ( Secretariat) WHO International Laboratories for Biological Standards/ Collaborating Centres

WHO Laboratories and Collaborating Centres *National Institute for Biological Standards and Control, Potters Bar, UK ( NIBSC) *Sanquin-Central Laboratory Netherlands Red Cross Blood Transfusion Service ( CLB) Center for Biologics Evaluation and Research FDA, Bethesda, USA (CBER) * Hold and distribute international standards

Collaboration Other standard setting bodies eg Council of Europe/European Department for the Quality of Medicines: USP/International Standards Organization (ISO) National Regulatory Authorities/National Control Laboratories Scientific Societies/Associations Manufacturers Associations ( International Federation of Pharmaceutical Manufacturers Associations)

Types of WHO biological reference materials International Standards enable the activity of biological preparations to be expressed in the same way globally mostly in (IUs) International Units Reference Reagents differ from International Standards in the extent of characterisation and intended use not assigned IU’s Interim

Types of WHO biological reference materials International Reference Panels Group of reference materials established to collectively aid evaluation of assays or diagnostic tests. Comply with requirements for WHO reference standards/reagents

Examples of WHO Standards relevant to in vitro diagnostics HBsAg, subtype adw2 genotype A (33 IU/vial) 2 nd IS 2003 Hep B virus DNA (500,000 IU/vial) 1 st IS 1999 Hep C RNA ( 50,000 IU/vial) 1 st IS 1997 HIV-1 p24 Ag ( 1000 IU/ampoule) 1 st Int Ref Reagent 1992 HIV-1 RNA ( 100,000 IU/vial) 1 st IS 1999 HIV-1 genotype Reference Panel 2003

Biologicals - What’s the Problem? Special consideration/ challenges Inherent variability of biological systems, including biological and immunological assays Potential for microbial contamination Complex macromolecules/systems

Biologicals – What’s the Problem? Cannot be adequately characterised by chemical and/or physical means alone Used in prophylaxis, therapy or diagnosis of human diseases (in vitro diagnostics)

WHO biological reference materials Play a vital role in facilitating transfer of laboratory science into worldwide clinical practice Contribute to development and on going use of safe and effective biologicals and reliable diagnostics

Use of WHO International Reference Materials Form basis of quality control, regulation and clinical dosing for biological medicines Form basis of quality control and regulation of in vitro diagnostic devices

Biotechnology Derived Products Past 20 years seen explosion in molecular biology/novel bioproduction methods Opened new possibilities for disease diagnosis/treatment /prevention Cutting - edge of biomedical research Economically fastest growing sector in pharmaceuticals

Regulatory Developments in standardisation International Standards Organisation developed written standards for establishment of reference materials European Commission adopted ISO standards for in vitro diagnostic devices Implications for WHO biological reference preparations

Regulatory Developments in standardisation Issue - extent to which principles for characterization of reference materials in other fields can be applied to biological reference materials Much debated.

ISO requirements for reference materials Apply to ALL reference materials, chemical and biological, Following ISO/WHO discussions recognized biologicals “different” Biological standards were not considered “primary” standards because could not meet all metrological principles – confusion

ISO requirements for reference materials Metrological principles - primary standards established in SI units Single method Measurement uncertainty Commutability Traceability to previous standard DOES NOT MAKE SENSE FOR ALL BIOLOGICAL SITUATIONS

Biological Situation Many biologicals exist in both active and inactive states in plasma Activity (IU) rather than content (mol) may better reflect the clinical situation Calibration in less precise biological units (IU) more appropriate than calibration in more precise, but clinically irrelevant SI units

Biological Situation Conversely, situations exists where measurement of inactive or total (active plus inactive) analyte (mol) may be more clinically relevant than activity (IU)

ISO requirements for reference materials Placed WHO standardization activities outside accepted metrological principles Made compliance with some external requirements (eg EU in vitro diagnostic devices directive) problematic

International Consultations – on in vitro diagnostics WHO worked with ISO, other standard setting bodies, regulatory authorities, scientific community and users Through series of consultations reviewed scientific basis for the preparation and characterization of biological reference materials.

International Consultations – on in vitro diagnostics WHO Consultation on Global Measurement Standards and their use in the in vitro Biological Diagnostic Field June 2004

International Consultations – on in vitro diagnostics Re-affirmation that concepts used by WHO for biological standardization still appropriate Re-affirmation of the continued need and usefulness of this class of reference materials Need for improved clarity in explaining principles used to establish WHO International Standards

Preparation, characterisation and establishment of WHO biological reference preparations Guidelines in WHO Technical Report Series,1978, revised in 1986,1990. Have now been updated taking into consideration recent developments and need for improved clarity Adopted as Recommendations by Expert Committee on Biological Standardization, November 2004

Recommendations not Guidelines For Preparation, Characterization and Establishment of International and other Biological Reference Standards Comprehensive document As usual considerable consultation in their development

New Recommendations Choice of unit ( IU/SI /none ) - should be based on the biological, medical and physicochemical information available on a case-by- case basis Where it is appropriate for WHO biological standard to be calibrated in SI units, principles of ISO to be followed

New Recommendations Deal with number of Issues/principles such as: Methods – single or multiple Measurement uncertainty Commutability in vitro diagnostics field (deals with matrix issues) Traceability Definitions

New Recommendations Sections include - Quality Assurance Treatment of bulk liquid Quality of final containers Freeze drying Characterization / Stability International collaborative studies Use

New Recommendations WHO biological standards cover a broad range of uses range of options should continue to be used in their characterisation.

New Recommendations Essential to define the intended use of a standard prior to initiation of studies - aids in design the studies to characterise the material and in the eventual value assignment to the material.

Assessment of need formalized Not all requests for development of international standards are appropriate. Need to assess priority in establishing International Standards/Reference materials Decision tree developed ( Appendix 1) ECBS is the decision making body, but with advice from other bodies and consultations eg SoGAT

European Commission – Common Technical Document Clarification of process and background to development and establishment of WHO International Standards Enabled EC to adopt WHO International Standard for Hepatitis B ( calibrated in IU) as the standard required to fulfil Common Technical Document

European Commission – Common Technical Document Legally binding in EC Further collaboration with the EC on going regarding adoption of other WHO International Standards for in vitro diagnostics.

Keeping Pace with Developments WHO needs to keep pace with developments in all biologicals fields SoGAT valuable venue to discuss scientific developments in NAT assays, to look to the future and to support the work of the ECBS

WHO Biologicals Field Information re WHO activities in biologicals field found on internet : / biologicals www. who.int / bloodproducts/en/ Catalogue of International Standards and Reference Reagent on line

WHO Consultation June 2003 Review the scientific basis for preparation and characterisation of WHO biological reference materials review a draft revision of the WHO guidelines make recommendations to WHO to ensure that WHO biological reference materials retain the widest acceptance of fitness of purpose

Calibration of standards (prEN/ISO 17511) requires: 1Single method studies (conventionally agreed or, where possible reference method) 2SI units rather than IU (where possible) 2 Traceability to previous standard, with defined uncertainty WHO guidelines produce standards: 1calibrated in a multi-method study (rather than a single, reference method) 2With values assigned in International units (rather than mg/ml) With no imprecision assigned to the ampoule content prEN/ISO WHO

Calibration of the current International Reference Preparation for TSH Deviation of any assay result from the mean is composed of two elements; the assay imprecision, and the bias: The WHO multi-method approach, by including all assays, seeks to average out, and therefore eliminate the bias effect. The WHO approach will provide an estimate which is “accurate” but not “precise” The “reference-method approach will provide an estimate which may be “precise”, but not “accurate” Method bias, and single method vs multi method calibration

Single or multi-method calibration studies June 2003 consultation: “the choice of method depends on whether the most important consideration is metrological to minimise imprecision, in which case a single method should be used, or is biological to achieve a “true” overall value, in which case multiple methods should be used”.

To measure in activity (IU) or content (SI units)? Many biologicals exist in both active and inactive states in plasma, where the activity (IU) rather than content (mol) reflects the clinical situation of the patient. Calibration in less precise biological units would hence be more appropriate than calibration in more precise, clinically irrelevant SI units 2Conversely, situations exists where measurement of inactive or total (active plus inactive) analyte (mol) may be more clinically relevant that activity (IU)

The case of the drifting hepatitis B nanogram The 1st HBsAg IS was assigned an IU value Used to calibrate immonoassays Some users also assigned a ng value A recent WHO collaborative study showed differences between ng assignments of some secondary standards

Drift in the HBsAg “ng”

Choice of units June 2003 consultation “the choice of unit should be based on biological, medical and physico- chemical criteria and not by perceived metrological status”

Assignment of uncertainty WHO standards are established either: - as the first International Standard for any given analyte, in which case in which case the international unit is arbitrarily established for the first time - or as replacement international standards, in which case it is necessary to ensure continuity of the value of the unit; two approaches - calibration or value assignment

Calibration approach the second standard is calibrated in terms of the first standard, preserving a line of metrological traceability of the international unit. This approach requires assignment of uncertainty, and the minimization of that uncertainty through the use of defined or even reference methods

Calibration approach Advantages compliance with the metrological principles established in various ISO standards Disadvantages a significant range of uncertainty would provide difficulties for the users for some standards (eg NAT standards) the uncertainties would run into orders of magnitudes for frequently replaced standards (eg factor VIII), the accumulated uncertainty would soon render the standard useless

Value assignment approach the second standard is arbitrarily assigned a value intended to preserve as closely as possible the value of the unit, but where traceability to the first IS is discontinued and re-established to the second. This approach requires an arbitrary assignment without uncertainty, as wide a range of methods as possible, and sometimes reference to additional factors outside the collaborative study (eg standard plasma pools for factor VIII)

Value assignment approach Advantages it works the observed variation in a study is verified statistically to fall within acceptable ranges and confidence limits for the assay methods used very little evidence in significant drift in the value of the IU Disadvantages places WHO standardization activities outside currently accepted metrological principles makes compliance with external requirements (eg EU in vitro diagnostic devices directive) problematic

Next steps concerning uncertainty WHO will convene further meetings - with the in vitro diagnostics regulators - a WHO working group on uncertainty - with ISO The next ECBS will be asked to consider stating some elements of uncertainty (imprecision of fill; predicted loss of activity on storage) in the memoranda that accompany standards

Other recommendations from June 2003 meeting WHO should make further use of experts in different specialities to plan the study design and evaluation of study results prior to submission of studies to the ECBS Additional guidance should be provided by WHO on the stability testing of international standards, but it is not appropriate to assign expiry dates to biological reference materials.

Conclusions from June meeting The continued usefulness of WHO biological standards shows that the approaches taken by WHO to biological standardisation have served well over a long period of time. The current two classes of reference preparation, the International Standard and WHO Reference Reagents should be maintianed.

Conclusions from June meeting As WHO biological standards cover a broad range of uses a range of options should continue to be used in their characterisation. It is essential to define the intended use of a standard prior to initiation of studies. This is to aid in the design of the studies to characterise the material and in the eventual value assignment to the material.