WHO Expert Committee On Biological Standardization: WHO Technical Report Series

W H O Te c h n i c a l R e p o r t S e r i e s
1059
WHO Expert Committee

on Biological
Standardization
Seventy-ninth report
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W H O Te c h n i c a l R e p o r t S e r i e s
1 0 5 9
WHO Expert Committee

on Biological
Standardization
Seventy-ninth report
This report contains the collective views of an international group of experts and
does not necessarily represent the decisions or the stated policy of the World Health Organization
WHO Expert Committee on Biological Standardization: seventy-ninth report
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Contents
Abbreviations xi
1. Introduction 1
2. General 3
2.1 Strategic directions in biological standardization 3
2.1.1 Update on recent and planned WHO written standards for biological products 3
2.2 Cross-cutting activities of other WHO committees and groups 4
2.2.1 Report from the WHO Product Development for Vaccines Advisory Committee 4
2.3 Other matters 6
2.3.1 Statement made during the 154th session of the Executive Board regarding
the work of WHO expert committees 6
3. International Recommendations, Guidelines and other matters
related to the manufacture, quality control and evaluation of
biological products 7
3.1 General 7
3.1.1 Discontinuation of innocuity test 7
3.2 Biotherapeutics other than blood products 7
3.2.1 Nonclinical and clinical evaluation of monoclonal antibodies and related
products intended for the prevention or treatment of COVID-19 7
3.2.2 Development of WHO guidance on the nonclinical and clinical evaluation of
monoclonal antibodies and related products intended for the prevention of
respiratory syncytial virus disease 9
3.3 Blood products and related substances 10
3.3.1 Revision of the WHO guidelines on good manufacturing practices for blood
establishments 10
3.4 Vaccines and related substances 12
3.4.1 Revision of the WHO Guidelines to assure the quality, safety and efficacy of
live attenuated rotavirus vaccines (oral) 12
4. International reference materials – biotherapeutics other than
blood products 14
4.1 WHO international reference standards for biotherapeutics other than blood
products 14
4.1.1 First WHO International Standard for golimumab 14
4.2 Proposed new projects and updates – biotherapeutics other than blood products 15
4.2.1 Proposed Third WHO International Standard for interleukin-2 (human,
rDNA derived) 15
4.2.2 Proposed Fourth WHO International Standard for endotoxin 16
4.3 Proposed discontinuations and updates – biotherapeutics other than blood
products 17
4.3.1 Proposed discontinuation of the First WHO International Standard for
calcitonin ASU 1–7 eel calcitonin analogue (elcatonin) 17
5. International reference materials – blood products and related
substances 20
5.1 Proposed new projects and updates – blood products and related substances 20
5.1.1 Proposed WHO International Reference Reagent for autoantibodies to
ADAMTS13 (human, plasma) 20
6. International reference materials – in vitro diagnostics 22
6.1 WHO international reference standards for in vitro diagnostics 22
6.1.1 First WHO International Reference Panel for Lassa virus RNA for
NAT-based assays 22
6.1.2 First WHO International Standard for HIV-1 p24 antigen 23
6.2 Proposed new projects and updates – in vitro diagnostics 25
6.2.1 Proposed First WHO International Standard for carcinoembryonic antigen 25
6.2.2 Proposed First WHO International Standard for antibodies to Junin virus 27
7. International reference materials – standards for use in
high-throughput sequencing technologies 29
7.1 WHO international reference standards for use in high-throughput sequencing
technologies 29
7.1.1 First WHO International Reference Panel for adventitious virus detection
in biological products by high-throughput sequencing 29
8. International reference materials – vaccines and related substances 32
8.1 WHO international reference standards for vaccines and related substances 32
8.1.1 WHO International Reference Reagent for diphtheria antitoxin for use in
flocculation test (equine) 32
8.2 Proposed new projects and updates – vaccines and related substances 33
8.2.1 Proposed Fifth WHO International Standard for diphtheria toxoid (adsorbed) 33
8.2.2 Proposed Second WHO International Standard for Vi polysaccharide of
S. Typhi 33
8.2.3 Proposed First WHO International Standard for antibodies to group A
streptococcus antigens (human serum) 34
8.2.4 Proposed First WHO International Standard for antibodies to vaccinia virus 35
8.3 Proposed discontinuations and updates – vaccines and related substances 36
8.3.1 Proposal not to proceed with the development of a First WHO International
Standard for antibodies to influenza A(H7N9) virus 36
Annex 1
WHO Recommendations, Guidelines and other documents related to the manufacture,
quality control and evaluation of biological products 39
Annex 2
Nonclinical and clinical evaluation of monoclonal antibodies and related products
intended for the prevention or treatment of COVID-19
Addendum to Annex 2 of WHO Technical Report Series, No.1048 49
Annex 3
New and replacement WHO international reference standards for biological products 83
iv
WHO Expert Committee on Biological Standardization
Seventy-ninth meeting held virtually 11 to 14 March 2024
Committee members1
Dr S.S. Ben Amor, National Drug Control Laboratory, Tunis, Tunisia
Dr C. Burns, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
Professor K. Cichutek, Paul-Ehrlich-Institut, Langen, Germany (Co-chair)
Dr J.A. Dahlan, Directorate of Standardization of Drugs, Narcotics, Psychotropics, Precursors
and Addictive Substances, Jakarta, Indonesia
Dr S. Fakhrzadeh, Consultant, Tehran, Iran (Islamic Republic of )
Dr I. Feavers, Consultant, Nacton, United Kingdom (Rapporteur)
Professor S. Hindawi, King Abdulaziz University, Jeddah, Saudi Arabia (Co-chair)
Professor M.B.C. Koh, St George’s Hospital Medical School, London, United Kingdom; and
Health Sciences Authority, Singapore, Singapore
Dr Q. Meyer, South African National Control Laboratory for Biological Products,
Bloemfontein, South Africa
Dr A. Ramkishan, Ministry of Health and Family Welfare, Hyderabad, India
Dr S. Silveira, Agência Nacional de Vigilância Sanitária, Brasilia, Brazil
Dr J. Southern, Representative of the South African Health Products Regulatory Authority,
Simon’s Town, South Africa
Professor C.T. Tagny,2 University of Yaoundé; and University Teaching Hospital Yaoundé,
Yaoundé, Cameroon
Dr D. Teo, Visiting Consultant, Blood Services Group, Health Sciences Authority, Singapore,
Singapore (Co-rapporteur)
Dr J. Wang, National Institutes for Food and Drug Control, Beijing, China
Dr S. Wendel, Hospital Sirio-Libanês, São Paulo, Brazil
Dr T. Wisit, Ministry of Public Health, Nonthaburi, Thailand
Dr T. Wu, Biologic and Radiopharmaceutical Drugs Directorate, Health Canada, Ottawa,
Canada
1
The decisions of the Committee were taken in closed session with only members of the Committee and
WHO Secretariat present. Each Committee member had completed a Declaration of Interests form prior to
the meeting. These were assessed by the WHO Secretariat and no declared interests were considered to be
in conflict with full meeting participation.
2
Unable to attend.
v
WHO Expert Committee on Biological Standardization Seventy-ninth report
Temporary advisors
Dr M.H. Aldosari, Saudi Food and Drug Authority, Riyadh, Saudi Arabia
Dr A. Bisht, Ministry of Health and Family Welfare, Noida, India
Dr P. Boonprasirt, Food and Drug Administration, Nonthaburi, Thailand
Dr M. Buda, European Directorate for the Quality of Medicines & Healthcare, Strasbourg,
France
Dr A. Chia, Health Sciences Authority, Singapore, Singapore
Dr A. Fouad, Egyptian Drug Authority, Cairo, Egypt
Dr E. Griffiths, Consultant, Kingston upon Thames, United Kingdom
Dr Md. Harun-Or-Rashid, Directorate General of Drug Administration, Dhaka, Bangladesh
Dr A. Korovkin, Ministry of Health, Moscow, Russian Federation
Dr J. Lacroix, Health Canada, Ottawa, Canada
Dr M. Li, National Medical Products Administration, Beijing, China
Dr L. Mallet, European Directorate for the Quality of Medicines & Healthcare, Strasbourg,
France
Dr P. Minor, Consultant, St Albans, United Kingdom
Dr G.I. Parra, Center for Biologics Evaluation and Research, US Food and Drug Administration,
Silver Spring, MD, United States of America (USA)
Dr M. Powell, Health Products Regulatory Authority, Dublin, Ireland
Dr K. Quillen, Atrius Health, Boston, MA, USA
Dr Y. Sohn, Seoul National University, Seoul, Republic of Korea
Dr P. Stickings, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
WHO Technical Report Series, No. 1059, 2024
Dr Y. Sun, Paul-Ehrlich Institute, Langen, Germany

Dr A.L. Waddell, Consett, United Kingdom (Editor of the report of the Committee)
Dr M. Wierer, European Directorate for the Quality of Medicines & Healthcare, Strasbourg,
France
Dr C. Witten, Center for Biologics Evaluation and Research, US Food and Drug
Administration, Silver Spring, MD, USA
State actors
Dr N. Almond, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
Dr M. Bailey, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
vi
Dr E. Bentley, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
Dr B. Cowper, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
Ms T. Desai, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
Dr K. Dix, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
Dr A. Eder, Center for Biologics Evaluation and Research, US Food and Drug Administration,
Silver Spring, MD, USA
Dr O. Engelhardt, Medicines and Healthcare products Regulatory Agency, Potters Bar,
United Kingdom
Ms L. Hassall, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
Dr K. Ishii, National Institute of Infectious Diseases, Tokyo, Japan
Dr A. Ishii-Watabe, National Institute of Health Sciences, Kawasaki, Japan
Dr A. Khan, US Food and Drug Administration, Silver Spring, MD, USA
Dr D. Khokal, Consultant, Singapore, Singapore
Dr G. Mattiuzzo, Medicines and Healthcare products Regulatory Agency, Potters Bar,
United Kingdom
Dr F. Mawas, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
Dr M. Moore, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
Ms C. Morris, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
Dr M. Nübling, Paul-Ehrlich-Institut, Langen, Germany
Dr M. Ochiai, National Institute of Infectious Diseases, Tokyo, Japan
Ms K. Partridge, Medicines and Healthcare products Regulatory Agency, Potters Bar,
United Kingdom
Mr G. Prescott, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
Dr G. Raychaudhuri, Center for Biologics Evaluation and Research, US Food and Drug
Administration, Silver Spring, MD, USA
Dr N. Rose, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
vii
Dr M. Rosu-Myles, Biologic and Radiopharmaceutical Drugs Directorate, Health Canada,

Ottawa, Canada
Dr S-R. Ryu, Ministry of Food and Drug Safety, Chungcheongbuk-do, Republic of Korea
Dr C. Schärer, Swiss Agency for Therapeutic Products, Bern, Switzerland
Dr Y. Takahashi, National Institute of Infectious Diseases, Tokyo, Japan
Dr C. Thelwell, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
Mr R. Tierney, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
Ms S. Tierney, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
Dr G. Unger, Paul-Ehrlich-Institut, Langen, Germany
Ms B. Valente, Agência Nacional de Vigilância Sanitária, Brasilia, Brazil
Dr M. Vasheghani, Food and Drug Administration, Tehran, Iran (Islamic Republic of )
Dr M. Wadhwa, Medicines and Healthcare products Regulatory Agency, Potters Bar,
United Kingdom
Dr J. Weir, Center for Biologics Evaluation and Research, US Food and Drug Administration,
Silver Spring, MD, USA
Dr H. Wilmot, Medicines and Healthcare products Regulatory Agency, Potters Bar, United
Kingdom
Dr M. Xu, National Institutes for Food and Drug Control, Beijing, China
Observers from non-state actors in official relations

International Alliance for Biological Standardization
Dr R. Sheets
International Federation of Pharmaceutical Manufacturers & Associations

Ms S. Laghnimi-Hahn, Geneva, Switzerland
International Generic and Biosimilar Medicines Association
Dr T. Kirchlechner
International Society on Thrombosis and Haemostasis
Professor J. Meijers, Amsterdam, Netherlands (Kingdom of the)
United States Pharmacopeial Convention
Dr F. Atouf, Rockville, MD, USA
Representation from other entities

Biotechnology Innovation Organization
Dr E. Flores, Washington, DC, USA
viii
Pharmaceutical and Medical Device Regulatory Science Society of Japan

Dr Y. Nakagawa, Osaka, Japan
Plasma Protein Therapeutics Association
Dr H. Tuna
World Health Organization (WHO)

Access to Medicines and Health Products (MHP)
Dr Y. Nakatani, Assistant Director-General
Health Products Policy and Standards (MHP/HPS)
Mr D. Mubangizi, Director
WHO Secretariat
Technical Standards and Specifications (MHP/HPS/TSS)
Dr I. Knezevic (Secretary to the Committee; Lead for the vaccines and biotherapeutics track)
Dr Y. Maryuningsih (Lead for the blood products and in vitro diagnostics track)
Mr S. Chatzixiros
Ms S. Jenner
Dr E. Kim
Dr D. Lei
Dr T. Zhou
Other WHO staff

Product and Delivery Research
Dr E. Sparrow (IVB/PDR)
Representation from WHO regional offices3

WHO Regional Office for the Americas
Dr M. Beltran
Dr M.L. Pombo
WHO Regional Office for Europe
Dr D. Pirgari
WHO Regional Office for the Western Pacific
Dr J. Shin
3
Unable to attend: WHO Regional Office for Africa; WHO Regional Office for South-East Asia; and WHO
Regional Office for the Eastern Mediterranean.
ix
Abbreviations
ADCC antibody-dependent cellular cytotoxicity
AG-BRAS Advisory Group on Blood Regulation, Availability and Safety
BET bacterial endotoxin test
CEA carcinoembryonic antigen
CEACAM carcinoembryonic antigen-related cell adhesion molecule
CEPI Coalition for Epidemic Preparedness Innovations
COVID-19 coronavirus disease 2019
DNA deoxyribonucleic acid
EDQM European Directorate for the Quality of Medicines &
HealthCare
ELISA enzyme-linked immunosorbent assay
GAS group A streptococcus
GBS group B streptococcus
GMP good manufacturing practice(s)
HIV human immunodeficiency virus
HPLC high-performance liquid chromatography
HTS high-throughput sequencing
IFU instructions for use
IL-2 interleukin-2
IU International Unit(s)
JUNV Junin virus
LASV Lassa virus
Lf limit of flocculation
mAb monoclonal antibody
MAT monocyte activation test
NAT nucleic acid amplification technique
NC3Rs National Centre for the Replacement, Reduction and
Refinement of Animals in Research
xi
NCL national control laboratory

NRA national regulatory authority
PCR polymerase chain reaction
PDVAC Product Development for Vaccines Advisory Committee
rDNA recombinant deoxyribonucleic acid
RNA ribonucleic acid
RPT rabbit pyrogen test
RSV respiratory syncytial virus
SARS-CoV-2 severe acute respiratory syndrome coronavirus 2
TDM therapeutic drug monitoring
TNF tumour necrosis factor
TTP thrombotic thrombocytopenic purpura
VACV vaccinia virus
VLP virus-like particle
xii
1. Introduction
The seventy-ninth meeting of the WHO Expert Committee on Biological
Standardization was held virtually from 11 to 14 March 2024. The meeting was
opened on behalf of the Director-General of WHO by Dr Yukiko Nakatani,
Assistant Director-General, Access to Medicines and Health Products. Dr
Nakatani began by welcoming participants and thanking them for devoting their
time and expertise to the work of the Committee.
Dr Nakatani noted that the 18 members of the Committee represented
a good balance of expertise, regional representation and gender, and went on to
explain the ongoing efforts being made to broaden even further the expertise
and global representativeness of the WHO Expert Advisory Panel on Biological
Standardization from which Committee members were drawn.
Dr Nakatani then provided a brief update on WHO activities to ensure
broader access to medicines and health products, highlighting the WHO support
being provided in 120 countries to advance the goal of universal health coverage.
This support included the WHO prequalification of vaccines, diagnostics and other
biological products to promote more equitable access to such products worldwide.
In addition, several medicines had now been added to the WHO Model List of
Essential Medicines (EML), including new products for treating multiple sclerosis,
cancer and cardiovascular conditions. With ongoing support from the WHO
regulatory systems strengthening programme, a number of key improvements had
also been made to the national regulatory systems in several countries.
Dr Nakatani went on to note a statement that had been made on behalf of
the WHO African Region by Senegal at the recent Executive Board meeting (see
section 2.3.1 below) and highlighted the significance of this to the work of the
Committee. In addition, following the adoption of resolution WHA67.21 in 2014
on access to biotherapeutic products, WHO had continued to develop important
standards and tools to facilitate access to biosimilar products of assured quality,
safety and efficacy.
Dr Nakatani concluded by commenting on the ambitious agenda
of the current meeting, noting in particular the proposed addendum to the
WHO Guidelines on the nonclinical and clinical evaluation of monoclonal
antibodies and related products intended for the prevention or treatment of
infectious diseases. This addendum would provide guidance specifically on the
evaluation of monoclonal antibodies (mAbs) for use against coronavirus disease
2019 (COVID-19). Dr Nakatani also noted the proposed updating of the 2011
WHO guidelines on good manufacturing practices for blood establishments. In
these and other important areas, the expertise of the Committee will be key to
informing the provision of WHO support to countries and strengthening the
global health leadership provided by WHO.
1
Dr Ivana Knezevic, Secretary to the Committee, thanked Dr Nakatani

for her opening remarks. Dr Knezevic went on to note that as the directing
and coordinating authority on international health within the United Nations
system, WHO adheres to its organizational values of integrity, professionalism
and respect for diversity, as set out in its Constitution. Driven by its mission to
promote health worldwide, WHO, through its activities at headquarters, regional
office and country office level, provides leadership on global health matters, shapes
the health research agenda, sets norms and standards, articulates evidence-based
policy options and provides technical support to countries. The setting of norms
and standards for biological products, and promoting their implementation, is
therefore a core function of WHO.
Dr Knezevic then reminded meeting participants that the decision-making
body of WHO is the annual World Health Assembly attended by delegations
from all WHO Member States. The agenda of the World Health Assembly is set
in advance each year by the Executive Board, which consists of technical experts
elected for 3-year terms. All of the reports and recommendations of the current
Committee are submitted to the Executive Board for its consideration.
Dr Knezevic then outlined the procedures and working arrangements
of the present meeting. An open information-sharing session involving all
participants, including non-state actors, would be held on Monday 11 March
2024. Committee members, regulatory authority representatives and subject
matter experts from governmental organizations would then participate in the
main meeting from Monday 11 March to Wednesday 13 March 2024. All final
decisions and recommendations on the adoption of WHO written standards and
the establishment of WHO measurement standards would be made in a closed
session held on Thursday 14 March attended only by Committee members and
WHO staff. Dr Knezevic concluded by expressing her thanks to WHO staff
working in this area, colleagues from WHO collaborating centres and custodian
laboratories, and all of the individual experts present.
Following the conclusion of the open information-sharing session, Dr

Knezevic presented the declarations of interests that had been completed by
Committee members, WHO temporary advisers and other participants. After
evaluation, WHO had concluded that none of the interests declared constituted
a significant conflict of interest, and that the individuals concerned would be
allowed to participate fully in the meeting. In the absence of dissent, Professor
Klaus Cichutek and Dr Silvano Wendel were elected as Co-chairs, and Dr Ian
Feavers and Dr Diana Teo as Rapporteur and Co-rapporteur respectively.
The Committee then adopted the proposed agenda (WHO/
BS/2024.2473).
2
2. General
2.1 Strategic directions in biological standardization
2.1.1 Update on recent and planned WHO written
standards for biological products
Dr Knezevic summarized the WHO written standards for biological products
recently adopted on the advice of the Committee, and provided an overview
of the plans for new and revised such documents from 2024 onwards. The
adoption of the following four documents between 2020 and 2023 had been of
particular relevance during the COVID-19 pandemic: (a) the WHO Guidelines
on the quality, safety and efficacy of plasmid DNA vaccines; (b) Evaluation of
the quality, safety and efficacy of messenger RNA vaccines for the prevention of
infectious diseases: regulatory considerations; (c) the WHO Guidelines for the
production and quality control of monoclonal antibodies and related products
intended for medicinal use; and (d) the WHO Guidelines on the nonclinical and
clinical evaluation of monoclonal antibodies and related products intended for
the prevention or treatment of infectious diseases. Subject to the advice of the
Committee, the latter document will be supplemented by a series of addenda
covering considerations specific to mAbs against COVID-19, respiratory syncytial
virus (RSV) disease, rabies, malaria and human immunodeficiency virus (HIV).
Following the 2022 adoption of the WHO manual for the preparation
of reference standards for use as secondary standards in antibody testing, with
its focus on antibodies against severe acute respiratory syndrome coronavirus
2 (SARS-CoV-2), an implementation workshop had been held in November
2023. The report of the workshop had been published on the WHO website and
provided to the Committee. In addition, the WHO Guidelines on regulatory
preparedness for the oversight of pandemic or other emergency use vaccines in
importing countries had been adopted in 2023.
With regard to upcoming WHO written standards, Dr Knezevic
indicated that revised WHO Guidelines on rotavirus vaccines would be proposed
for adoption at the next meeting of the Committee in October 2024, with an
update on progress made to date scheduled for the current meeting (see section
3.4.1 below). It was further anticipated that revised WHO Guidelines on post-
approval changes for vaccines would be presented to the Committee in October
2025, and revised WHO Recommendations for the preparation, characterization
and establishment of international and other biological reference standards in
2025 or 2026. The revision of several other WHO written standards for vaccines
was expected to commence in 2026, and would include written standards on
yellow fever virus, dengue, measles, mumps and rubella, bacille Calmette-Guérin
and malaria vaccines, as well as more general WHO Guidelines on vaccine lot
release. In addition, and guided by the advice of the WHO Product Development
3
for Vaccines Advisory Committee (PDVAC) (see section 2.2.1 below) and by
stakeholder requests, the development of new WHO written standards on
chikungunya, paratyphoid, shigella and group B streptococcus (GBS) vaccines
was anticipated, along with WHO guidance on the evaluation of vaccines and
other biological products using high-throughput sequencing (HTS) technologies.
Dr Knezevic concluded by noting the completion in 2023 of the
independent, systematic review of WHO written standards by the National
Centre for the Replacement, Reduction and Refinement of Animals in Research
(NC3Rs) in the United Kingdom. During discussion of the full report of this
review, the Committee had recommended the establishment of a working group
to build on its findings and to help develop science-based guidance encouraging
the replacement of animal testing with non-animal testing for the quality control
of vaccines and biotherapeutic products. The recently formed working group was
expected to present such guidance for consideration by the Committee by 2026.
2.2 Cross-cutting activities of other WHO committees and groups

2.2.1 Report from the WHO Product Development for Vaccines
Advisory Committee
Dr Erin Sparrow summarized the outcomes of the most recent PDVAC meeting
held in December 2023 at which a range of cross-cutting and vaccine-specific
topics had been discussed. Dr Sparrow highlighted the provision of PDVAC
support to the Gavi 2024 Vaccine Investment Strategy process, noting that new
vaccines against tuberculosis, GBS, shigella and dengue had been shortlisted for
Gavi investment. A number of such vaccines, along with a range of other novel
vaccines and mAbs against infectious diseases, could be approved in the next
5–10 years.
Dr Sparrow went on to discuss the current tuberculosis vaccine pipeline
in detail, which mainly comprises vaccines intended to boost waning immune
responses to bacille Calmette-Guérin in adults and adolescents. Several different

types of candidate vaccines were now at an advanced stage of clinical development,
with PDVAC recommending that a WHO roadmap be developed setting out a
pathway for their introduction. PDVAC further advised that such novel vaccines
would require new written and measurement standards to support their WHO
prequalification.
Dr Sparrow then noted the potential application of controlled human
infection models in the development of vaccines against diarrhoeal and enteric
pathogens, with several such models now existing or in development. Noting
that the first bivalent typhoid conjugate vaccine and paratyphoid A vaccine
might be approved on the basis of data obtained using such a model, Dr Sparrow
highlighted the need for a regulatory consultation on the approval pathway of
such vaccines. Once again, specific WHO guidance in this area may be required
4
General
to support vaccine licensure and WHO prequalification. In addition, the recent

entry of several shigella candidate vaccines into the enteric vaccine pipeline would
necessitate the development of an antiserum reference standard to harmonize the
measurement of antibody responses and help identify correlates of protection.
Dr Sparrow further noted that GBS vaccines were also at an advanced
stage of clinical development. Such vaccines were unlikely to be approved on the
basis of traditional Phase III clinical studies as these were costly and logistically
challenging to perform. A licensure pathway based on immune correlates of
protection was therefore being explored with regulators and stakeholders to
ensure the ongoing commitment of vaccine developers. An approach based
on immune correlates of protection had already been used to approve the first
chikungunya vaccine. PDVAC recommendations in this area had included the
conducting of modelling, feasibility and other studies for a range of different
scenarios to inform chikungunya vaccine advocacy at national and regional level.
Noting the recent publication of the WHO Guidelines on the nonclinical
and clinical evaluation of monoclonal antibodies and related products intended
for the prevention or treatment of infectious diseases, and noting the disease-
specific addenda to this document due to be discussed at the current meeting,
Dr Sparrow concluded by summarizing the developmental status of a number
of such products. Related WHO activities included the development of general
preferred product characteristics for mAbs used within routine immunization
programmes, and of policy requirements for broadly neutralizing antibodies
used to prevent the vertical transmission of HIV to infants.
The Committee thanked Dr Sparrow for her review and enquired
whether there had been any progress towards the development of HIV-1 vaccines.
Dr Sparrow noted that despite more than 40 years of research, there were
currently no promising candidates in the pipeline. However, several promising
mAb products were in development, with particular interest in their potential
to prevent vertical transmission. The Committee also asked whether the recent
success of COVID-19 messenger RNA vaccines had led to the development of
other vaccines based on this technology. Dr Sparrow indicated that although
several such products were in development, their reactogenicity profile and cold
chain requirements meant that they were unlikely to replace existing vaccines
based on older technologies. Enquiring about the current status of Nipah virus
vaccine development, the Committee was informed that as such outbreak vaccines
were on the agendas of the WHO R&D Blueprint and the Coalition for Epidemic
Preparedness Innovations (CEPI) they did not come under the remit of PDVAC.
Responding to further questions from the Committee, Dr Sparrow
emphasized the importance of post-marketing surveillance to confirm the
efficacy of vaccines first licensed based on correlates of protection. Reflecting on
the lessons learned from the licensure of the chikungunya vaccine, the Committee
noted that although this was a pragmatic solution when efficacy studies were
5
impractical, it did rely on having a well-defined correlate of protection, ideally

defined in International Units (IU). In this regard and more generally, the
Committee encouraged the use of multinational randomized controlled studies
across endemic areas to expedite the approval of new vaccines.
2.3 Other matters

2.3.1 Statement made during the 154th session of the Executive
Board regarding the work of WHO expert committees
At its 154th session held in January 2024, the Executive Board was presented with
the main outcomes and recommendations of the 77th meeting of the Committee
held in March 2023, together with those of other WHO expert committees. At
that occasion, the following statement was made:
Chairman, Ministers, Distinguished Delegates
SENEGAL has the honour of delivering this statement on behalf
of the 47 Member States of the WHO African Region. The African
Region would first of all like to congratulate the expert committees on
the meetings they held during the biennium 2022–2023 from which
came important recommendations on biological standardization,
the selection and use of essential medicines and the evaluation of
certain food additives.
These recommendations are of crucial importance for public policy
and the programmes of the Organization and therefore we believe that:
1. The preparation of Guidelines on the nonclinical and clinical
evaluation of monoclonal antibodies and related products
intended for the prevention or treatment of infectious diseases is
to be promoted. These would make it possible to provide specific
updated guidance to national regulatory authorities and the

manufacturers of biological products in order to guarantee their
safety and efficacy.
2. The preparation of a regulatory framework for human cells and
tissue and for innovative medical treatments that would favour
regulatory harmonization of these products would also be useful.
3. We support the making available of WHO standards recognized
internationally to make it possible to extend universal health
coverage and the updating of the WHO Model List of Essential
Medicines, taking into account needs but also efficacy and
efficiency criteria so as to guarantee accessibility to all…
We urge the Executive Board to note the present report.
6
3. International Recommendations, Guidelines and
other matters related to the manufacture, quality
control and evaluation of biological products
3.1 General
3.1.1 Discontinuation of innocuity test
At its sixty-ninth meeting in 2018, the Committee had recommended the
immediate cessation of any requirement to perform the innocuity test (also
known as the abnormal toxicity or general safety test) in WHO written standards
for biological products. It had further recommended that any mention of
the test in WHO documents published before 2018 should be disregarded,
acknowledging that the requirement could only be removed from such documents
when they were revised or replaced. Despite these recommendations, a recent
review commissioned by WHO and carried out by the National Centre for the
Replacement, Reduction and Refinement of Animals in Research (NC3Rs) in the
United Kingdom had indicated continued widespread use of the innocuity test.
To further highlight the recommendations of the Committee in this
regard, it was agreed that an “important note” and a table listing all current
WHO written standards on biological products published prior to 2018 that
still mentioned the unneeded innocuity test would be added to Annex 1 of all
reports of the Committee from October 2023 onwards. There was a consensus
among Committee members that this would be an important addition that
should continue to be made until all the relevant documents had been updated
or withdrawn. Recognizing that many users of WHO written standards now
accessed WHO documents directly from the WHO website, the Committee
further suggested that the statement be reproduced on the relevant WHO
webpages, along with other web-based approaches, to inform users that they
should disregard any historic requirement to perform innocuity testing.
3.2 Biotherapeutics other than blood products

3.2.1 Nonclinical and clinical evaluation of monoclonal antibodies and
related products intended for the prevention or treatment of COVID-19
COVID-19 continues to pose a significant public health threat worldwide, with
SARS-CoV-2 causing severe disease and, in some cases, long COVID, including
among highly vulnerable groups. It is anticipated that as COVID-19 becomes
endemic, it will continue to cause severe illness, hospitalizations and deaths as
new variants emerge. In March 2023, the WHO Guidelines on the nonclinical
and clinical evaluation of monoclonal antibodies and related products intended
for the prevention or treatment of infectious diseases were adopted on the advice
of the Committee. This document provides guidance on the evaluation of mAbs
7
and related products regardless of the target pathogen or toxin. During the
adoption of these Guidelines, the Committee had recognized the need to also
develop a number of addenda on disease-specific regulatory considerations.
The Committee was provided with an overview of the development
and structure of a proposed addendum setting out a number of supplementary
considerations when evaluating the safety and efficacy of mAb products specifically
intended for the prevention or treatment of COVID-19. Following two rounds
of public consultation and subsequent revisions, the proposed addendum now
consisted of seven sections ranging from general considerations to more detailed
guidance on nonclinical and clinical evaluation. The Committee was reminded
that separate detailed guidance on the production and quality control of mAbs
was provided in the previously adopted WHO Guidelines for the production
and quality control of monoclonal antibodies and related products intended for
medicinal use.
The Committee sought clarification on a number of issues, including
on the guidance provided on the degree to which nonclinical and clinical data
obtained for specific mAb formats or for specific variants of concern could be
applied to other comparable mAb products. The Committee indicated that any
new mAb format or mAb targeting a newly emerging variant of concern should
be assessed as a new product, including in terms of their ability to neutralize the
target variant. Assurance was provided to the Committee that clear guidance on
these and related issues was provided in the parent Guidelines and the addendum
when these were read in conjunction.
The Committee also highlighted two issues with the proposed section
on international reference materials. While noting that such a section might
helpfully be included in all relevant WHO Recommendations, Guidelines and
guidance documents to promote the use of WHO measurement standards, the
Committee was concerned that the section would become out of date once
the cited standards were replaced or new variants emerged. Furthermore, the
international reference materials in question were based on antisera, and
although likely to be of utility, there was currently no evidence to support their
use in evaluating mAbs. In light of these comments, it was agreed that the text
would be modified to ensure that users check for the most up-to-date reference
materials, and that more cautionary language would be used regarding the use of
current international reference materials to characterize SARS-CoV-2 mAbs.
Despite a current lack of licensed products intended to be inhaled or
administered intranasally, the Committee noted that prospective such mAb
products against respiratory pathogens may be developed. The Committee agreed
that such products would require a number of specific additional pharmacokinetic
and pharmacodynamic considerations compared to parenterally administered
mAbs, including considerations arising from the potential use of a delivery device.
8
International Recommendations, Guidelines and other matters
Appropriate text was therefore added to the nonclinical and clinical sections of
the addendum to address this issue. The Committee also suggested that although
all currently approved neutralizing mAbs target the SARS-CoV-2 spike protein,
the text be broadened to include mAbs targeting other antigens that may be in
development. In addition, the Committee suggested some clarification of the
guidance be provided on the use of available safety data to support the inclusion
of pregnant women in clinical studies.
After due consideration of these and other minor modifications made to
the text, the Committee recommended that document WHO/BS/2024.2466 be
adopted and annexed to its report (Annex 2).
3.2.2 Development of WHO guidance on the nonclinical and clinical

evaluation of monoclonal antibodies and related products intended
for the prevention of respiratory syncytial virus disease
Worldwide, RSV is a leading cause of respiratory disease in all age groups,
frequently causing severe morbidity and mortality in infants, young children and
older adults. It is the second most common cause of infant mortality, with more
than 99% of childhood deaths due to RSV occurring in low- and middle-income
countries which typically lack safe and effective prevention or treatment options.
Two prophylactic mAb products for use against RSV disease have recently
been approved, with several others now in clinical development. Targeting
neutralization-sensitive epitopes on the virus, these mAbs provide immediate
protection against severe disease, especially in infants at particularly high risk.
The Committee was updated on progress made towards the development
of an addendum to the WHO Guidelines on the nonclinical and clinical
evaluation of monoclonal antibodies and related products intended for the
prevention or treatment of infectious diseases that would address specific issues
in the evaluation of mAbs used to prevent RSV disease. The addendum would be
based on a format similar to that used for the addendum on COVID-19 mAbs
recommended for adoption at the current meeting (see section 3.2.1 above). Its
purpose would be to provide supplementary considerations in the evaluation
of the safety and efficacy of prophylactic mAbs directed specifically against
RSV, and would apply primarily to parenterally administered mAb products. A
general considerations section will provide background on the pathology and
epidemiology of RSV disease, as well as on the mechanism of action of RSV mAbs
and their effectiveness. A section on nonclinical evaluation will provide guidance
on the use of in vitro and in vivo studies to assess the pharmacodynamics and
biological activities of candidate mAbs, with a section on clinical evaluation
focusing on assessment of their prophylactic effect.
A drafting group had now been established and the proposed addendum
was currently in preparation. Following two rounds of public consultation, it
9
was anticipated that the document would be submitted to the Committee for its
review at its next meeting in October 2024.
While reflecting on the potential advantages of vaccines over mAbs for
the prevention of RSV disease, the Committee also felt that access to both types
of products would be desirable at present, and that the addendum would be a
very important document in this context. The Committee looked forward to
seeing the resulting document in due course.
3.3 Blood products and related substances

3.3.1 Revision of the WHO guidelines on good manufacturing
practices for blood establishments
One of the strategic objectives of the WHO Action Framework to advance
universal access to safe, effective and quality-assured blood products 2020–2023
is the establishment of well-functioning and efficiently managed blood services,
with a functioning quality system as one of the specific outcomes. However, a lack
of knowledge in implementing and maintaining good manufacturing practices
(GMP) in blood establishments has been identified as a common barrier to
achieving this objective. To help address this issue, efforts were now under way
to update the 2011 WHO guidelines on good manufacturing practices for blood
establishments as part of the 2023–2024 workplan of the WHO Advisory Group
on Blood Regulation, Availability and Safety (AG-BRAS). As one of the planned
activities of the WHO Action Framework, it was intended that the revised
document would reflect new developments in the field and provide countries
with practical guidance on how to establish reliable quality assurance systems for
the whole chain of blood collection, testing, processing and distribution of blood
components in blood establishments and hospital blood banks.
Following an overview of the WHO procedures for developing guidance
documents, the Committee was provided with a summary of the objectives of
the revised document, and of the rationale and justification for its development.
The proposed revised document would set out the international principles and
standards of GMP for blood establishments and hospital blood banks that should
be implemented and enforced by the national regulatory authority (NRA) to
ensure the quality and safety of blood components for transfusion and as starting
material for further industrial manufacturing. The revised document would
also be aligned with existing international documents in the field to facilitate
harmonization in the global manufacturing and use of plasma products.
The Committee was informed that an expert working group comprising
AG-BRAS members had been established and was being supported by the WHO
Blood and Other Products of Human Origin team, with input from regional
advisers. Following a detailed summary of the structure and contents of the
proposed revised document, an update was provided on the progress made to
10
date. It was intended that a complete draft would be available in the first half of
2024 and that following a process of expert comment and public consultation,
the final version would be published in late 2024. Explanation was given that
although WHO planning clearance had been obtained based on developing
a normative and standard-setting publication, the proposal was also being
submitted to the Committee for its consideration as the 2011 WHO Guidelines
had been developed with its involvement and subsequently jointly published by
the WHO Expert Committee on Specifications for Pharmaceutical Preparations.
The Committee welcomed the efforts made by the working group
and AG-BRAS in revising this important document as this would directly
contribute to the consistent production of safe and quality-assured blood, blood
components and plasma products. Observing that the risks and challenges for
GMP implementation had also changed over the years, the Committee noted
that quality standards in hospital blood banks were often suboptimal, with
unsatisfactory clinical transfusion practices in many resource-limited countries.
Reflecting on how GMP and quality management concepts could best be
implemented in these areas, the Committee noted that GMP would only apply
to that which is necessary to maintain safety and quality during blood collection,
testing, processing, release, and distribution to, and storage in, hospitals. This
would not cover the key areas of clinical blood transfusion practices, which are
instead part of the activities of the hospital quality system. It was acknowledged
that the proposed document would thus cover quality system elements in blood
establishments that also applied to hospital blood banks rather than the full scope
of GMP in the latter setting. The Committee also suggested a number of additional
topics that might usefully be considered in the document, such as donor data
security/confidentiality when data is shared between blood establishments. It
was also recognized that the relationship between the blood establishments and
manufacturers of blood products was an important focus, as it was envisaged that
blood establishments compliant with the revised WHO Guidelines would achieve
a level of quality that would make plasma suitable for further manufacturing,
with manufacturers of plasma-derive medicinal products looking to only source
plasma from such GMP-compliant establishments.
The Committee concluded by discussing the proposed next steps
in developing and publishing the revised document. Given the significant
similarities between the WHO process for developing normative and standard-
setting publications and the Committee process, the parallel publication of the
proposed document appeared to be feasible. Under this parallel approach, the
completed draft would undergo public consultation, followed by presentation of
the final document to the Committee at its next meeting in October 2024. The
final document would also be published on the WHO website to expedite user
access and then independently published in the Technical Report Series, subject
to the recommendations of the Committee.
11
Recognizing the important and pressing need for WHO guidance to

countries in this area, the Committee expressed its support for the proposed
revision and parallel publication of the WHO guidelines on good manufacturing
practices for blood establishments.
3.4 Vaccines and related substances

3.4.1 Revision of the WHO Guidelines to assure the quality, safety
and efficacy of live attenuated rotavirus vaccines (oral)
Rotavirus is a highly infectious cause of severe dehydrating gastroenteritis in
children under the age of 5 years, and in the absence of antiviral treatment is best
prevented by vaccination. Universal rotavirus vaccination of infants has been
recommended by WHO for more than a decade and there is now considerable
experience of using the available live attenuated vaccines. Since the adoption of
the WHO Guidelines to assure the quality, safety and efficacy of live attenuated
rotavirus vaccines (oral) in 2005 there have been numerous developments. These
include the approval of the first two live attenuated rotavirus vaccines in Europe
and the USA and then in numerous countries, and their WHO prequalification.
Other live attenuated rotavirus vaccines have been licensed in India and
prequalified by WHO, while live attenuated rotavirus vaccines have also been
licensed nationally in China and Viet Nam. In addition, other candidate vaccines
were currently being developed, including non-replicating rotavirus vaccines.
Over the same period, several new and revised WHO guidance documents
with broad applicability to vaccine standardization have been published that
reflect technological advances in vaccine production, quality control and clinical
evaluation.
In light of such technological advances and other developments, it had
previously been proposed that the 2005 WHO Guidelines be updated, and the
Committee was updated on the progress made to date. Following consultations
with experts and stakeholders worldwide, a drafting group consisting of regulatory

experts from several countries had prepared a series of draft revised texts. Once
finalized, the revised document is intended for use by NRAs, national control
laboratories (NCLs) and vaccine manufacturers, and will primarily address
issues regarding live attenuated rotavirus vaccines for the prevention of disease.
However, the sections on the nonclinical and clinical evaluation of candidate
vaccines are intended to be applicable to all types of rotavirus vaccines.
The Committee was then provided with a detailed overview of the
various sections of the current draft document which follows a format similar
to recently adopted WHO vaccine Guidelines and Recommendations. Key
changes in the document reflect current practice in the production and control
of live attenuated rotavirus vaccines, provide guidance on the pharmacological
evaluation of candidate vaccines developed on different platforms, elaborate on
12
toxicological testing (including the risk of intussusception) and provide guidance

on the design of future trials (including in the context of available licensed
rotavirus vaccines, and for different types of such vaccines).
Comments received following a second round of public consultation on
the draft document had primarily focused on the requirement for sterility and
sterile filtration. A small number of respondents had expressed concerns with
regard to meeting sterile filtration requirements, and had advocated instead for
confirmation of low bioburden as an end-point. In response, it was noted that
whereas the original Guidelines required sterility at all production stages, the
revised document was already more flexible as it introduces the possibility of
using a bioburden test at an intermediate stage of production. However, such an
approach requires that a sterilizing filtration be performed later in the production
process. Conversely, and in line with long-established practice, such a filtration
step is not mandatory or required if the entire manufacturing process is aseptic,
and controlled by sterility testing and validated. This approach is consistent
with the regulatory requirements of other entities, including the European
Pharmacopoeia.
During discussion, a consensus was reached by the Committee that
orally administered vaccines such as oral poliomyelitis vaccines and oral
rotavirus vaccines should be sterile products given the target population and the
implications of microbial contamination for storage. The Committee supported
the revision of the wording in some places to provide more flexibility during
production processes but strongly emphasized that whichever approach was
used, the final product must in all cases be sterile.
Having agreed a further revised wording on sterility and sterile
filtration, and having suggested several other amendments to the current text,
the Committee recommended that the draft document be submitted for a third
round of public consultation. The Committee looked forward to reviewing the
final draft in more detail at its next meeting in October 2024, with a view to its
potential adoption.
13
4. International reference materials – biotherapeutics
other than blood products
4.1 WHO international reference standards for
biotherapeutics other than blood products
4.1.1 First WHO International Standard for golimumab
Golimumab is a human therapeutic mAb originally isolated from a hybridoma
clone generated from transgenic mice immunized with human tumour necrosis
factor (TNF). It is structurally well characterized and binds with high affinity
and specificity to both soluble and transmembrane forms of TNF thereby
neutralizing its biological activity. Golimumab is used in adults as a TNF
antagonist to treat moderate-to-severe rheumatoid arthritis, psoriatic arthritis,
ankylosing spondylitis, non-radiographic axial spondyloarthritis and moderate-
to-severe ulcerative colitis. It is also used to treat polyarticular juvenile idiopathic
arthritis in children. Variable efficacy among anti-TNF products has highlighted
a potential need for therapeutic drug monitoring (TDM), which measures the
levels of the therapeutic and of anti-drug antibodies. However, implementation
of this approach has been hampered by a lack of standardization of the analytical
methods used.
In order to develop a prospective reference standard for use in measuring
both the in vitro biological activity of golimumab and golimumab concentrations
in samples from treated patients, two international collaborative studies had been
conducted using a donated drug preparation. This preparation was formulated
similarly to WHO international standards for other TNF mAbs, and filled and
lyophilized in accordance with WHO guidelines.
In the first study, 16 laboratories in 11 countries had assessed the
suitability of the candidate material (NIBSC code 22/116) to standardize cell-
based bioassays and TNF binding assays. Results indicated that the candidate
material typically reduced the variability of potency estimates for tested

preparations compared with estimates expressed relative to in-house reference
standards. In the second study, using a panel of sera spiked with different amounts
of candidate material 22/116, eight laboratories had assessed the ability of the
candidate material to standardize the measurement of serum golimumab content
in clinical laboratories. Results indicated that use of the candidate material
reduced inter-laboratory variability thus harmonizing the clinical monitoring
tests used for TDM. Accelerated thermal degradation studies indicated that the
candidate material was stable during long-term storage at −20 °C, with additional
studies indicating that the reconstituted candidate material was stable for at least
1 week at 4 ºC or at room temperature. Freeze-thaw stability testing indicated
that the potency of the candidate material was retained for at least four freeze-
thawing cycles.
14
International reference materials – biotherapeutics other than blood products
Noting the importance of this class of mAb product, the Committee

considered this to have been an excellent study. However, concerns were expressed
that assigning both IU and SI unitages to the same material might lead some
users to erroneously relate activity to the mass of mAb. Reassurance was given
that the two distinct groups of users would either be measuring bioactivity (in
IU) or TDM (in µg/mL) but not both. In addition, the instructions for use (IFU)
would clearly caution against the potential misuse of the reference material.
Following further minor clarifications, the Committee considered the
report of the study (WHO/BS/2024.2467) and recommended that candidate
material 22/116 be established as the First WHO International Standard for
golimumab with assigned unitages of 500 IU/ampoule TNF neutralizing activity,
500 IU/ampoule TNF binding activity, 500 IU/ampoule FcγRIII binding activity
and 500 IU/ampoule antibody-dependent cellular cytotoxicity (ADCC) activity.
Noting that this material was intended to define IU of bioactivity for the purposes
of assay harmonization, the Committee emphasized that it was not intended
to define the specific activity of golimumab products or to serve as a reference
product for biosimilarity determination or dosing requirements. The Committee
further recommended that a unitage of 50 µg/ampoule also be assigned to the
First WHO International Standard for golimumab for the purpose of TDM.
4.2 Proposed new projects and updates –

4.2.1 Proposed Third WHO International Standard for
interleukin-2 (human, rDNA derived)
Interleukin-2 (IL-2) is approved for the treatment of metastatic renal cell
carcinoma and melanoma. Clinical studies are also ongoing on the therapeutic
use of IL-2 either on its own or as part of combination therapy for cancer
immunotherapy, autoimmune disease and potentially for amyotrophic lateral
sclerosis. The current WHO international standard (NIBSC code 86/500) had
been established in 2013 and is used in the potency labelling of IL-2-based clinical
products, and for calibrating commercial IL-2 reagents and immunoassay kits.
Compared to a rate of use of approximately 130 ampoules per year from
1986 to 2013 for the first WHO international standard (NIBSC code 86/504),
the current international standard is being used at a rate of approximately
300 ampoules per year. Despite close monitoring and imposition of restrictions
on distribution since late 2023, it was anticipated that stocks would likely be
exhausted in around 2 years, and a replacement reference standard was now
required.
It was proposed that a candidate replacement material would be prepared
using full human sequence IL-2 derived from recombinant DNA (HEK293
cells) formulated in 1% human serum albumin, 0.5% trehalose and PBS. With
15
the exception of the buffer used for the second WHO international standard,
this was essentially the formulation that had been used for both of the previous
two WHO international standards, and had proven to be stable. Although the
IL-2 in the preparation would be glycosylated, it had previously been shown
that glycosylation would not affect bioactivity in the assays used if the complete
human sequence was present. The Committee was informed that the source
material was now on site and that trial fills had been completed with a definitive
fill scheduled. To meet the increasing rate of demand, it was proposed that a
batch size of 8000 ampoules would be prepared – double that of the previous two
batches. An international multi-centre collaborative study would be conducted
in 2024–2025 to evaluate the performance of the candidate material and to assign
a unitage relative to the second WHO international standard. It was anticipated
that the results of the collaborative study would be submitted for consideration
by the Committee in 2026.
Noting the importance of the WHO international standard for IL-2 in the
development and production of IL-2-based products, and for the investigation
of novel therapeutic approaches, and the short time available for establishing
a replacement international standard, the Committee endorsed the proposal
(WHO/BS/2024.2472) to develop a Third WHO International Standard for
interleukin-2 (human, rDNA derived).
4.2.2 Proposed Fourth WHO International Standard for endotoxin

Testing parenteral medicinal products for pyrogens is essential to prevent serious
adverse events caused by contamination or inconsistent production, with bacterial
endotoxin being the most common pyrogen contaminating biological products.
The WHO international standard for endotoxin has been widely used to calibrate
critical reference materials used in three compendial methods: the rabbit pyrogen
test (RPT); the bacterial endotoxin test (BET); and the monocyte activation
test (MAT). Users of these international standards include pharmacopoeias,

regulatory agency laboratories, pharmaceutical companies and pyrogen testing
reagent manufacturers worldwide.
The current Third WHO International Standard for endotoxin (NIBSC
code 10/178) was established in 2012. To ensure a harmonized approach to
endotoxin standardization, a portion of this material had also been used to establish
EDQM and US Pharmacopoeia reference materials, with an understanding that
this harmonized approach would be continued. With the impending removal
of the RPT from the European Pharmacopoeia in 2026, and increasing use of
recombinant BET reagents as part of the move towards animal-free testing, it is
anticipated that the level of demand will increase. Therefore, although current
stocks were expected to last for a further 5–6 years, it was being proposed that
development of the replacement standard should begin immediately.
16
It was proposed that a donated lyophilized bulk endotoxin material

derived from Escherichia coli (Braude strain) would be used to develop a
candidate material. This bacterial endotoxin was the same source material that
had been used for the first and second WHO international standards, and for
previous lots of the companion reference materials. However, as stocks of the
source material itself were also running low, and would only be sufficient for the
development of one or two future replacement standards, it was proposed that
the replacement international standard be formulated at a lower concentration
(5000 IU/vial) to conserve stocks of the source material and ensure its efficient
use. Current methods for endotoxin testing were now very sensitive, and users
had been consulted on the proposed reduction. No responses or objections had
been received regarding this change and broader consultation was planned. The
source material was already on site and would be formulated in same way as
the previous WHO international standards, with a definitive fill in late 2024. An
international multi-centre collaborative study would then be conducted in 2025
to evaluate the performance of the candidate replacement material and to assign
potency. It was anticipated that the results of the collaborative study would be
submitted for consideration by the Committee by no later than October 2026.
Noting that only the BET would be included in the collaborative study
and that the RPT would no longer be included due to the move towards animal-
free pyrogen testing, the Committee reflected on the role and possible future
inclusion of the MAT. In response it was noted that, in general, the MAT was
considered to be more useful for detecting non-endotoxin pyrogenic substances
and not as suitable for endotoxin. Having also been assured that no complaints
had been received in response to the proposed reduction in potency, the
Committee endorsed the proposal (WHO/BS/2024.2472) to develop a Fourth
WHO International Standard for endotoxin.
4.3 Proposed discontinuations and updates –

4.3.1 Proposed discontinuation of the First WHO International Standard
for calcitonin ASU 1–7 eel calcitonin analogue (elcatonin)
Elcatonin, an eel analogue of calcitonin, inhibits the absorption and autolysis
of bone thereby lowering blood calcium levels. It is primarily used to reduce
pain associated with osteoporosis. The First WHO International Standard for
calcitonin, ASU 1–7 eel calcitonin analogue (elcatonin) (NIBSC code 84/614) was
developed in the 1980s for the calibration of therapeutic elcatonin preparations
using in vivo bioassays. Stocks of this international standard were expected
to be exhausted within 2–3 years. However, in light of its highly restricted
geographical use and associated low level of demand, any continuing need for
such a global international standard was likely to be minimal, with the regional
17
use of elcatonin potentially supported through available regional or national

standards. In addition, and in line with the increasing application of the 3Rs
principles (Replacement, Refinement, Reduction) regarding the use of animals
in research, there should now be a transition away from the assignment of IU to
elcatonin products based on the use of in vivo rat bioassays towards the dosing of
such products in mass units using animal-free methods. It was noted that other
calcitonin analogues were now being controlled using physicochemical methods
in both the European Pharmacopoeia and US Pharmacopoeia.
The Committee was therefore informed of the decision of the custodian
laboratory not to prepare a replacement international standard, and was presented
with two options going forward. The first option was to simply discontinue the
WHO international standard upon exhaustion of the current stocks. The second
option would be for another custodian laboratory to develop a replacement
WHO international standard.
During discussion, the potential implications of discontinuation of the
WHO international standard for users in China and Japan were considered, and
clarification sought on the availability of regional and national standards. The
monograph for elcatonin in the Japanese Pharmacopoeia includes an in vivo
bioassay supported by a national standard traceable to the WHO international
standard, and it was opined that the current batch of this national standard
would likely last for some time, with any future replacements calibrated against
it. The Committee was further informed that in China, elcatonin was marketed
as a chemical drug and was controlled by national standards based on in vivo
bioassay methods in Chinese Pharmacopoeia monographs.
Exploring the feasibility of using alternative assays such as high-
performance liquid chromatography (HPLC) to characterize the international
standard, the Committee was advised that although theoretically possible due
to the relatively small size and lack of complexity of the elcatonin peptide,
the presence of albumin in the preparation might be problematic, with issues

encountered in developing such physicochemical alternatives in Japan. One
possible long-term solution would be to calibrate and establish a reference
material using HPLC based on widely available EDQM and US Pharmacopoeia
standards for calcitonin, but this would depend on the acceptability of this
approach, as in some countries this would require changes to the way in which
products were controlled.
Reflecting on the importance of customer outreach efforts, the Committee
was informed that sales restrictions had been put in place to minimize the risk of
a large one-off order depleting the stock, with letters now being sent out to notify
customers that the future of the standard was uncertain and would be clarified.
Noting that customers requesting the material had not yet been informed of
the intention to discontinue supply, the Committee recommended that further
18
steps be taken to notify users and to ensure that any concerns are taken into
consideration. It was agreed that specific mention of this issue in the Executive
summary of the current meeting would be a useful first step in the broader
dissemination of the current intentions.
After due consideration, the Committee agreed that a decision on the
proposed discontinuation of the First WHO International Standard for calcitonin,
ASU 1–7 eel calcitonin analogue (elcatonin) would be taken at its next meeting in
October 2024 to provide sufficient time and opportunity for the information to
be disseminated and feedback obtained. The Committee also recommended that
any interest from other WHO collaborating centres or institutions in establishing
a replacement standard, including existing institutions in China and Japan
responsible for maintaining national standards, should be explored. Sufficient
material should be curated from current stocks to support the establishment of
such a replacement standard by another institution or to support any potential
regional standards that might be developed.
19
5. International reference materials – blood
products and related substances
5.1 Proposed new projects and updates – blood
products and related substances
5.1.1 Proposed WHO International Reference Reagent for
autoantibodies to ADAMTS13 (human, plasma)
Thrombotic thrombocytopenic purpura (TTP) is a rare disorder caused by
ADAMTS13 deficiency, with a reported annual incidence of around 6 per
million population. ADAMTS13 is a plasma protease enzyme responsible for the
cleavage of von Willebrand factor, which is involved in blood clotting. Prompt
and accurate diagnosis is essential, as the onset of clinical symptoms can be rapid
with 90% of cases fatal if left untreated. TTP may either be congenital due to an
inherited deficiency (cTTP) or immune mediated due to autoantibodies against
ADAMTS13 (iTTP). Differential diagnosis is based on Bethesda-type activity
assays or enzyme-linked immunosorbent assays (ELISA), and determines the
treatment modalities employed. Once recovered, patients must also be monitored
every 6–12 months in order to detect any onset of recurrence.
Several different activity assays and one ELISA-based binding assay are
currently available commercially, with significant discrepancies observed between
the different methods, and no common units of measurement exist between the
functional inhibitor and binding antibody methods. The Bethesda assay is also
inherently variable due to the use of different plasma pools by laboratories. The
standardization of assays and quantification of inhibitory activity will therefore
be important both for harmonization and for ensuring correct patient diagnosis,
monitoring and treatment.
It was proposed that a WHO international reference reagent for
ADAMTS13 autoantibodies for use by assay manufacturers and clinical
laboratories in hospitals managing TTP patients be developed using pools

of plasma collected from iTTP patients. An international collaborative study
involving 10–20 laboratories would be conducted to evaluate the suitability of a
candidate material using functional Bethesda-type assays to measure reduction
in ADAMTS13 activity or ELISA-based assays to detect anti-ADAMTS13
antibodies. As the project was still in its early stages, the source material had
not yet been acquired but a network of clinicians had indicated their willingness
to provide the source material. Based on the current indicative timeline, it was
anticipated that the results of the collaborative study would be submitted for
consideration by the Committee in 2027. It was noted that the high cost of assays
might limit study participation and thus geographical representation.
The Committee observed that in absolute terms, TTP was not that
uncommon, with several recent publications on case numbers. Access to source
20
International reference materials – blood products and related substances
material would not be a problem as large volumes of plasma were usually collected
from iTTP patients during plasma exchange and discarded as waste material.
Having been assured of the support of the clinician network in sourcing material
for this project, the Committee endorsed the proposal (WHO/BS/2024.2472)
to develop a WHO International Reference Reagent for autoantibodies to
ADAMTS13 (human, plasma).
21
6. International reference materials – in vitro diagnostics
6.1 WHO international reference standards for in vitro diagnostics
6.1.1 First WHO International Reference Panel for
Lassa virus RNA for NAT-based assays
The Committee was reminded that at its seventy-fifth meeting in April 2022, a
twin proposal had been made to establish both a WHO international standard
and WHO international reference panel for Lassa virus (LASV) RNA for use
in NAT-based assays. The Committee had subsequently recommended the
establishment of the First WHO International Standard for Lassa virus RNA for
NAT-based assays (NIBSC code 21/112). However, in the case of the international
reference panel, the Committee had recommended that additional panel member
characterization and performance studies be conducted prior to its establishment.
The Committee had further recommended that due to insufficient data at 3
months, along with a positive microbiological result obtained for two panel
members, additional stability testing be conducted for both the international
standard and international reference panel.
The Committee was informed that additional in-house characterization
testing had now been conducted on the international reference panel with the
data indicating that previously observed variability in potency estimates for
two panel members was not due to sample commutability issues but was assay
dependent. This further demonstrated the utility and fitness for purpose of all
reference panel members in evaluating NAT-based assay performance across
different LASV lineages.
The Committee was further informed that the recommended additional
stability testing had also been completed for both the international standard
and international reference panel for up to 1-year timepoints. In the case of the
international standard, minimal loss was demonstrated across all timepoints up
to 37 °C and only a slight loss of potency observed at 45 °C at 6 months and 1
year. Based on Arrhenius model calculations, only a negligible loss in potency

per year was predicted when stored at the recommended temperature of
−20 °C, and no change was necessary in the original recommendation to ship the
international standard at ambient temperature. For the international reference
panel, stability assessments completed at timepoints up to 1 year also indicated
minimal loss of potency in all samples for up to 3 months at temperatures up to
37 °C, and up to 1 year at 4 °C and 20 °C. However, some loss of potency after 1
year was observed at 37 °C and 45 °C. No clear differences were observed in the
stability of panel members found to have a higher residual moisture content, or
that had previously returned a positive microbiological test result. The data again
supported the recommendation for long-term storage at −20 °C and shipment at
ambient temperature. As it was not possible to predict long-term stability using
the Arrhenius model, periodic stability testing of the panel would be undertaken.
22
International reference materials – in vitro diagnostics
Noting that LASV lineages I and VI were not represented in the

international reference panel, the Committee was reassured that this was due
to their lower level of circulation, and that samples could be incorporated into
the panel if required. On the issue of preferred gene target, the Committee
was informed that the L-gene is not LASV specific and cross-reacts with other
arenaviruses. Most assay kits would therefore include both L-gene and GPC-gene
targets with a positive result for either target gene sufficient to suggest LASV
infection. Commenting that materials shipped at ambient temperature might be
affected by rising global temperatures, particularly in hot climates, the Committee
was assured that this would be guided by the results obtained at 37 °C and 45 °C
during stability testing, and that samples were also monitored during transit.
Having considered the above matters and the addendum (WHO/
BS/2024.2468) to the original report of the study, the Committee recommended
that the following four candidate materials be established without assigned
unitage as the First WHO International Reference Panel for Lassa virus RNA for
NAT-based assays (available under NIBSC code 22/108):
■ Candidate material 21/102 – LASV Lineage II
■ Candidate material 21/106 – LASV Lineage III
■ Candidate material 21/108 – LASV Lineage V
■ Candidate material 21/104 – LASV Lineage VII.
The Committee noted that the IFU would include a statement indicating
that the panel had been formulated to the same target potency based on
quantification of the common lentiviral gene, and that LASV-specific NAT-based
assays have detected discrepancies when targeting the S-segment that may be
assay dependent.
6.1.2 First WHO International Standard for HIV-1 p24 antigen

The core HIV-1 capsid protein p24 is present in abundance during the early
stages of HIV-1 infection and detectable during days 15–50 post infection, thus
making it a suitable target for the diagnosis of early-stage infection. Sensitive
fourth-generation serological tests utilize the detection of both antibodies and
HIV-1 p24, thereby narrowing the window period for diagnosis compared to
antibody-only testing.
The WHO International Reference Reagent for HIV-1 p24 antigen
(NIBSC code 90/636) was established in 1992 to support the development and
standardization of p24 assays. In 2009, the revised European Union common
technical specifications for in vitro diagnostic medical devices required HIV-1
p24 assays to have a limit of detection of 2 IU/mL based on calibration against
the WHO international reference reagent. In light of dwindling stocks of the
23
WHO international reference reagent, a proposal to develop a First WHO

International Standard for HIV-1 p24 antigen had been endorsed by the
Committee in October 2022.
Prior to this proposal, and due to the scarcity of high-titre HIV-1 stock
materials, a pilot study had been conducted to evaluate both a commercially
sourced recombinant p24 protein and an in-house preparation of virus-like
particles (VLPs) as alternative source materials. Although the recombinant
p24 protein had been found to be suitable for most assay systems, it was of an
unknown subtype and exhibited some discrepancy on Abbot platforms. The
VLPs had produced more consistent results in the study, appeared to be very
stable and would be easy to replace. The Committee had therefore agreed that the
VLP material would be suitable for further evaluation as a candidate material. It
had also been decided to formulate the proposed WHO international standard at
a lower concentration as the international reference reagent needed to be highly
diluted to evaluate current assays within their dynamic range.
An international collaborative study involving 15 laboratories in seven
countries had been conducted to evaluate the suitability of the candidate material
(NIBSC code 22/230) and to calibrate it against the current WHO international
reference reagent using a range of commercial qualitative HIV-1/2 fourth-
generation immunoassay systems. The candidate material was suitably diluted
in pooled negative plasma and lyophilized, with post-fill testing indicating that
oxygen content and residual moisture levels were within acceptable ranges.
Stability testing had been conducted at four timepoints (1, 3, 6 and 12 months),
with calculations based on the Arrhenius equation predicting a loss of 0.083% per
year when stored at −20°C. Using a wide range of different methods (including 3
prototype assays), the candidate material had been assessed as part of a blinded
panel of study samples which had also included the international reference
reagent, a liquid preparation of the candidate material, a recombinant p24 protein
preparation prepared from the same material used in the pilot study, a clinical
HIV-1 antigen- and antibody-positive sample identified as subtype B, and two
clinical HIV-1 antigen-positive window period samples identified as subtype C.
All samples were tested in duplicate on three occasions.
A total of 35 datasets were produced, with low intra-laboratory variability
indicating good precision for all of the assay types used. Potency estimates for the
candidate material were calculated relative to the international reference reagent.
Following exclusion of a number of outlier results and of the results obtained
using the not yet established prototype assays, a potency of 44.1 IU/ampoule was
established for the candidate material using a robust mean rather than geometric
mean to remove bias, and with results from laboratories using the same assay
combined to remove any over-representation of specific assay systems. Assessment
of the effect of lyophilization on the candidate material was assessed by evaluating
24
its relative potency against the liquid preparation, with currently unexplained
variations in estimated potency observed. Although a significant degree of non-
commutability was observed for the candidate material, the level observed was
equivalent to that of the current international reference reagent. The implications of
the revised common technical specifications for in vitro diagnostic medical devices
for manufacturers were also investigated by calculating the approximate limit of
detection for all laboratories and methods. Assuming a value of 44 IU/ampoule for
the candidate material, all assays produced values ranging from 0.14 IU to 1.99 IU
with the exception of the Abbott Determine HIV Early Detect assay.
The Committee expressed concern regarding the performance of the
Abbott assay and noted that although not marketed within the European Union,
it would still be marketed in other countries, and hence should be considered a
problem. Further noting that only subtypes B and C had been used for the study,
and that other subtypes might be more prevalent in some parts of the world, the
Committee was informed that this was because the WHO international reference
reagent was subtype B, while the inclusion of subtype C (the circulating variant)
had previously been recommended. It was also clarified that panels of p24 clades
have been produced and can be used to assess the performance of assays against
different subtypes. Recognizing the significant challenges involved in obtaining
window period samples for this project, the Committee offered its support with
regard to the sourcing of such donations in future projects.
Having considered the report of the study (WHO/BS/2024.2470), the
Committee recommended that candidate material 22/230 be established as the
First WHO International Standard for HIV-1 p24 antigen with an assigned
unitage of 44 IU/ampoule.
6.2 Proposed new projects and updates – in vitro diagnostics

6.2.1 Proposed First WHO International Standard
for carcinoembryonic antigen
Carcinoembryonic antigen (CEA) is a glycoprotein found at low levels (< 5
ng/mL) in the blood of healthy adults. CEA levels are raised in some cancers
(including bowel, lung, breast and pancreatic cancer), as well as during liver
disease and inflammatory bowel diseases such as Crohn’s disease and ulcerative
colitis. Raised CEA levels may also be predictive of metastatic disease in some
cancers, such as breast and colon cancers. CEA levels are measured using serum/
plasma-based immunoassays which are widely used as a diagnostic tool, and for
monitoring disease progression and response to treatment.
A WHO international reference preparation (NIBSC code 73/601) had
been prepared in 1976 using an extract of native CEA purified from liver cancer
tissue and formulated in 0.5% lactose, and was used predominantly to calibrate
test kits measuring CEA in patient samples. As stocks of this preparation were
25
now running low, and are expected to be exhausted by 2025, it was proposed
that a WHO international standard be developed to replace the it. The proposed
international standard would be prepared using a native material and formulation
similar to that used for the current international reference preparation.
As CEA contains a mixture of different carcinoembryonic antigen-
related cell adhesion molecules (CEACAMs), proteomic analysis of both the
current international reference preparation and trial-filled candidate materials
had been conducted to determine any impact of variable CEACAM distribution
on continuity. The analysis had demonstrated CEACAM5 to be present in
highest abundance, with some diversity noted in other less abundant CEACAMs.
As CEACAM5 is known to be the most prevalent and clinically relevant protein
targeted by commercial assays, it was concluded that any slight difference
in CEACAM distribution would not affect continuity. It was also proposed,
following discussion with kit manufacturers, to formulate the international
standard using 2 µg of material (compared to the 10 µg used in the current
international reference preparation) as this would be sufficient to cover the
dynamic ranges of current assays and would reduce waste. Furthermore, by using
isotope dilution mass spectrometry it may now be possible to assign SI units
to the international standard. This would help to address an observed issue of
uptake of the international reference preparation (calibrated in IU) since most
CEA immunoassays report results in ng/mL.
An international collaborative study involving 10–12 test kit
manufacturers and clinical laboratories performing CEA assays would be
conducted to evaluate the suitability of the candidate material. It was anticipated
that the results of the collaborative study would be submitted for consideration
by the Committee in 2025.
Noting that the current international reference preparation had
remained in use for 48 years, the Committee reflected that this had been due to a
combination of a large initial batch size and lower uptake rather than the use of
competing standards. The issue of low adoption would hopefully be addressed by

the use of SI units in the proposed international standard. The Committee also
considered the risk of discontinuity between the existing international reference
preparation (expressed in IU/mL) and the proposed international standard
(expressed in ng/mL). This was acknowledged to be an important issue that
would be taken into consideration during the proposed collaborative study. The
Committee was further reassured that a collaborative project currently under
way with the International Federation of Clinical Chemists on tumour marker
harmonization (including CEA) would provide an opportunity to discuss any
potential impact of this shift with clinicians and assay manufacturers.
Reflecting on the potential impact of variable CEACAM distribution, the
Committee opined that the candidate material should ideally be obtained from the
same type of tissue and be relevant to the diagnostic target of the immunoassays.
26
Clarification was given that the international reference preparation had been
derived from a liver biopsy of a metastatic tumour caused by colon cancer, and
that efforts would be made to ensure that a similar type of material was sourced.
While acknowledging that further detailed CEACAM analysis would not be
feasible or materially change the overall concept, the Committee recommended
that proposals for such future standards should underscore the need to obtain
source material from similar types of tissue to ensure continuity and relevance.
The Committee endorsed the proposal (WHO/BS/2024.2472) to develop
a First WHO International Standard for carcinoembryonic antigen.
6.2.2 Proposed First WHO International Standard for antibodies to Junin virus
Junin virus (JUNV) is the etiological agent of Argentine haemorrhagic fever,
which causes significant morbidity and has a mortality rate of 15–30%. The virus
is spread through a rodent reservoir that is so far confined to Argentina. Although
only 10–15 cases have been reported in recent years, approximately 5 million
people are considered to be at risk and live within the endemic area in which the
rodent reservoir is present. Post-exposure treatment options are limited to the
transfusion of immune plasma and the off-label use of ribavirin and favipiravir.
A live attenuated vaccine (Candid#1) has been licensed in Argentina since 1992
but its use is limited to adult populations at risk due to concerns regarding the
stability of the attenuated phenotype. Several alternative vaccine platforms are
now in preclinical development.
JUNV is a member of the family Arenaviridae – which is divided into
New World viruses (such as JUNV) and Old World viruses (such as LASV).
Global pandemic preparedness efforts are moving towards an approach based
on identifying prototype pathogens that could represent viruses within a whole
taxonomic group, rather than focusing on individual priority pathogens. In
this regard, LASV has been identified as a prototype pathogen for Old World
arenaviruses, with JUNV a likely suitable prototype pathogen for New World
arenaviruses.
Given the expected development of JUNV vaccines, therapeutics
and diagnostics, assay standardization and calibration will be vital for the
accurate evaluation and regulatory approval of such products. In-house ELISA,
immunofluorescence assays and neutralization assays are currently used to detect
antibodies to JUNV. The availability of a WHO international standard for use
by JUNV vaccine manufacturers, NCLs and other public health laboratories,
therapeutic antibody producers, assay kit manufacturers and research laboratories
would support the further development and standardization of such assays,
align with the prototype pathogen approach and complement the First WHO
International Standard for anti-Lassa virus immunoglobulin G established on the
advice of the Committee in 2021.
27
The candidate material would be prepared from human plasma or

serum obtained from convalescent individuals and Candid#1 vaccine recipients.
Following screening for JUNV and other bloodborne viruses, the material would
undergo virus inactivation treatment in accordance with protocols developed
by the Advisory Committee on Dangerous Pathogens to ensure safety during
broader distribution. The Committee was informed that financial and practical
support in sourcing and developing the candidate material would be provided by
CEPI, with support in securing and characterizing source materials also being
provided by the National Institute of Human Viral Diseases in Argentina.
An international collaborative study involving 15–20 laboratories would
then be conducted to evaluate the performance of the candidate material, assign
unitage and assess commutability. Participants would include NCLs, vaccine
manufacturers, and clinical and academic laboratories performing a range of
serological assays for the detection of JUNV antibodies. Potential difficulties were
anticipated in identifying such laboratories across all WHO regions given the highly
geographically restricted circulation of JUNV. It was provisionally anticipated that
the proposed collaborative study would be conducted in early 2025, with the results
submitted for consideration by the Committee in October 2025.
Recognizing that little was known regarding serological cross-reactivity
between Old World and New World arenaviruses, the Committee encouraged
further investigation into this aspect. In response to a query regarding the
use of pseudovirus or wild-type virus during the project, the Committee was
informed that only pseudovirus systems could be used for neutralization tests
in the custodian laboratory but that National Institute of Human Viral Diseases
laboratories were using a test based on wild-type virus, thus allowing for the use
of both systems during screening and characterization of the candidate material.
Acknowledging the need for this reference material, the Committee
endorsed the proposal (WHO/BS/2024.2472) to develop a First WHO
International Standard for antibodies to Junin virus.
28
7. International reference materials – standards for
use in high-throughput sequencing technologies
7.1 WHO international reference standards for use in
high-throughput sequencing technologies
7.1.1 First WHO International Reference Panel for adventitious virus
detection in biological products by high-throughput sequencing
HTS technologies have the ability to detect both known and novel adventitious
viruses in biological products. As an alternative to conventional adventitious virus
detection, which relies on in vivo animal testing and in vitro cell culture assays or
on polymerase chain reaction (PCR) assays, HTS has the potential to expand the
breadth of virus detection while also significantly shortening the time required
for the quality control testing of biological products. Importantly, the increasing
implementation of HTS for this purpose aligns with, and provides support for,
the shift towards reducing the use of animals in such testing. Encouraged by
regulatory guidance worldwide, the need for international viral standards for
HTS process qualification and validation to support the application of HTS to
biological product virus safety testing has long been recognized.
An international collaborative study involving nine laboratories in six
countries had therefore been conducted to evaluate the suitability of seven viruses
representing diverse virus families for use as a potential WHO international
reference panel for adventitious virus detection in biological products by HTS.
The following viruses had been selected based on their distinct physicochemical
and genome properties:
■ human betacoronavirus OC43 (hCoV)
■ porcine circovirus type 1 (PCV1)
■ mammalian orthoreovirus type 1 (REO)
■ feline leukemia virus (FeLV)
■ Epstein-Barr virus (EBV)
■ human respiratory syncytial virus (RSV)
■ minute virus of mice (MVM).
The viruses were spiked together at different spike levels but all laboratories
included testing at 104 genome copies/mL of each virus into 109 genome copies/
mL of adenovirus 5 to evaluate the breadth of virus detection using HTS in a
high-titre virus background mimicking a low-complexity biological sample
with reduced host cellular content – for example, a viral vaccine seed or virus
vector preparation. Participating laboratories followed a common protocol for
preparing the spiked samples and then used their own HTS workflow protocols.
29
All laboratories returning data detected the seven viruses between 104 and 105
genome copies/mL. Differences observed in virus detection between laboratories
reflected the different protocols used in the HTS workflow.
The Committee commended this excellent study and noted its timeliness
in the context of ongoing work to develop WHO guidance on the implementation
of 3Rs principles when testing biological products – which included the recent
establishment of a WHO working group on the implementation of 3Rs principles.
Further commenting on the related need for written guidance and training on
the application of HTS in ensuring the quality and safety of such products, the
Committee noted that several conferences on the application of HTS technologies
in biological product testing had been held by the International Alliance for
Biological Standardization.
Noting the urgent need for this reference panel and its relevance in
ensuring the safety of cell and gene therapy products, the Committee enquired
as to the anticipated demand and the likely effects of this on supplies. The
Committee was assured that the 1000 panels prepared should be sufficient for
about 5 years, with the expectation that panel titres would be sufficiently high to
last an individual organization for at least 1 year.
Enquiring about the potential impact of endogenous viruses in the cell
lines used to prepare the virus stocks, the Committee was informed that although
two retroviruses had been detected this did not affect the performance of the
panel. Similarly, contamination of the panel with residual host cell DNA did not
contribute significantly to the spike levels given the high titre of panel members.
However, the level of contamination with host cell DNA would be recorded in the
certificate of analysis.
Recognizing the importance of such a panel in helping to understand the
impact of bioinformatics on the sensitivity of HTS for detecting extraneous agents,
the Committee was informed that while the data to support establishment of the
panel had been based on targeted bioinformatics, collaborative study participants

had also returned data using non-targeted bioinformatics to reflect the real-life
situation, and these data would be published separately in the scientific press.
Reflecting on the prospect of HTS replacing in vivo and other in vitro tests for
the detection of adventitious viruses, the Committee concluded that although it
was impossible to generalize for all viruses, HTS was typically proving to be more
sensitive than animal-based methods and was not associated with the drawbacks
of PCR-based testing, such as mismatched primers. However, even though The
European Pharmacopoeia indicates that HTS may be used as a replacement
method, it would ultimately be the responsibility of the NRA to consider the
risks associated with specific products.
Committee recommended that the panel of seven viruses be established as the
30
International reference materials – standards for use in high-throughput sequencing technologies
First WHO International Reference Panel for adventitious virus detection in

biological products by high-throughput sequencing, with the following assigned
unitages:
CBER-FSCUST-90 (hCoV) 2.6 x 1010 genome copies/mL
CBER-FSCUST-91 (PCV1) 8.1 x 109 genome copies/mL
CBER-FSCUST-92 (REO) 1.5 x 1010 genome copies/mL
CBER-FSCUST-93 (FeLV) 4.0 x 1010 genome copies/mL
CBER-FSCUST-94 (EBV) 2.8 x 107 genome copies/mL
CBER-FSCUST-95 (RSV) 5.5 x 1010 genome copies/mL
CBER-FSCUST-96 (MVM) 1.2 x 1010 genome copies/mL.
The Committee further recommended that a set of five individual virus
preparations established in 2020 as WHO international reference reagents for
adventitious virus detection in biological products by HTS now be discontinued
in line with the proposed succession plan. These valuable resources should now
be solely used as CBER research reagents to support HTS development by new
users, with a clear indication that these reagents no longer have the status of
WHO international reference standards.
31
8. International reference materials – vaccines
and related substances
8.1 WHO international reference standards for
vaccines and related substances
8.1.1 WHO International Reference Reagent for diphtheria
antitoxin for use in flocculation test (equine)
Diphtheria toxoid is typically administered in combination with tetanus and
pertussis as DTP vaccines, and is also a carrier protein in several glycoconjugate
vaccines. Reliable evaluation of the concentration and quality of diphtheria
toxoid in combination vaccines depends upon the accuracy and robustness of the
routinely used flocculation test which requires the use of hyperimmune equine
diphtheria antitoxin as a critical reagent. The equine diphtheria antitoxin used
in the flocculation test is produced from hyperimmune horse serum, with a
lyophilized preparation of hyperimmune equine antitoxin (NIBSC code 63/007)
previously available as a non-WHO reference material. However, this reagent had
been widely used and stocks were now completely depleted. In 2023, the Committee
had therefore endorsed a proposal to develop a First WHO International Reference
Reagent for diphtheria antitoxin for use in flocculation test (equine).
The Committee was informed that a purified diphtheria equine
immunoglobulin preparation produced from a pool of equine serum had been
filled and lyophilized in ampoules to produce the candidate material (NIBSC code
23/102). The candidate material had an estimated diphtheria antitoxin potency
of 1032 IU/ampoule as assessed by toxin neutralization assay. As the proposed
international reference material would be used as a reagent (not a calibrant) and
as only a single method was involved, characterization of the candidate material
was undertaken in a small collaborative study involving seven laboratories in five
countries. Study participants calibrated the candidate material against the Third
WHO International Standard for diphtheria toxoid for use in flocculation test
(NIBSC code 13/212) using the Ramon flocculation method.
Study results (expressed in Lf-eq/mL against 13/212) gave an overall
geometric mean value of 956 Lf-eq/mL for the candidate material (range 890–
1032 Lf-eq/mL), with low intra- and inter-laboratory geometric coefficients of
variation observed. Initial data from an accelerated thermal degradation study
indicated that the candidate material would have good long-term stability, with
further testing to be performed. The 1831 ampoules produced would likely be
sufficient for at least 6–7 years.
Discussing the differences in results that had been noted between
different laboratories, the Committee was informed that this was most likely
attributable to the subjectivity of the flocculation test and difficulty in seeing
the end-point. The Committee also noted that sourcing sufficient material for
32
International reference materials – vaccines and related substances
this type of standard was challenging and advised that sufficient time be allowed
when sourcing future replacements.
Committee recommended that candidate material 23/102 be established as
the WHO International Reference Reagent for diphtheria antitoxin for use in
flocculation test (equine) with no assigned unitage.
8.2 Proposed new projects and updates – vaccines

and related substances
8.2.1 Proposed Fifth WHO International Standard
for diphtheria toxoid (adsorbed)
Diphtheria toxoid-containing vaccines are among the most commonly used
and successful human vaccines, with more than 30 different products approved
worldwide. As well as being an essential part of the primary immunization
programme for children, these vaccines are also used to boost immunity in
adolescents and adults. Ensuring the continuing supply of effective vaccines
depends on confirmation of their potency against a reference standard calibrated
in IU.
Following their establishment in 2009, both the Fourth WHO
International Standard for diphtheria toxoid (adsorbed) and European
Pharmacopoeia biological reference preparation (BRP batch 4) for diphtheria
toxoid have been widely used for potency testing by vaccine manufacturers and
control laboratories. As stocks of both materials were now low, replacement
materials were required. It was therefore proposed that an international
collaborative study involving laboratories in several WHO regions be conducted
to calibrate replacement WHO and European Pharmacopoeia standards in IU.
Two candidate materials would be prepared from adsorbed diphtheria toxoids
provided by vaccine manufacturers and evaluated in a study similar to that
used to calibrate the existing standards. It was anticipated that the proposed
collaborative study would be conducted in early 2025 and its results submitted
for consideration by the Committee in October 2025.
The Committee agreed that this was a straightforward and well-designed
project to replace the existing standards and endorsed the proposal (WHO/
BS/2024.2472) to develop a Fifth WHO International Standard for diphtheria
toxoid (adsorbed).
8.2.2 Proposed Second WHO International Standard

for Vi polysaccharide of S. Typhi
Typhoid fever is caused by infection with Salmonella enterica subspecies enterica
serovar Typhi (S. Typhi) expressing a Vi polysaccharide capsule, which is both a
virulence factor and protective antigen. As immunization is the most cost-effective
33
preventative strategy for controlling typhoid, especially in areas where multidrug-

resistant strains are endemic, licensed polysaccharide and glycoconjugate
vaccines are widely used in at-risk populations. The quality, safety and potency of
such vaccines are principally assessed using physicochemical methods to ensure
batch-to-batch consistency, with the WHO international standard being used to
calibrate secondary reference materials used in the different methods to quantify
the Vi polysaccharide content of bulk intermediates and final formulations. The
First WHO International Standard for Vi polysaccharide of S. Typhi (NIBSC
code 16/126) had been established in 2017 and due to low stocks a replacement
will be needed within 3 years.
A working freeze-dried material (previously filled under NIBSC code
17/260) had been identified as a suitable replacement, with sufficient ampoules
available to last for more than 10 years at the current rate of use. The candidate
material would be assessed for its suitability in an international collaborative
study involving approximately 20 laboratories. Based on the study results,
polysaccharide content would be assigned in milligrams using quantitative
nuclear magnetic resonance spectroscopy as an accepted primary method for
quantifying organic compounds in mass units. However, other assays would also
be included to assess the suitability of the candidate material across a range of
test methods. It was anticipated that the proposed collaborative study would be
conducted in 2024 and its results submitted for consideration by the Committee
in 2025.
Having been assured of the stability of the candidate Vi polysaccharide
preparation, the Committee endorsed the proposal (WHO/BS/2024.2472) to
develop a Second WHO International Standard for Vi polysaccharide of S. Typhi.
8.2.3 Proposed First WHO International Standard for antibodies

to group A streptococcus antigens (human serum)
Infection with group A streptococcus (GAS) is associated with a range of diseases

from mild pharyngitis to severe necrotizing fasciitis, as well as autoimmune
conditions such as rheumatic heart disease. Despite a high burden of disease
worldwide, GAS vaccine development has historically been hampered by a
number of scientific, regulatory and commercial challenges. Following concerted
global efforts to prioritize GAS vaccine development, particularly to reduce the
global burden of rheumatic heart disease, a number of candidate vaccines based
on M proteins and non-M protein antigens were now in preclinical or clinical
development.
The development of an international reference standard for antibodies
to GAS antigens will be an important step in standardizing the assays used to
compare antibody responses to prospective GAS vaccines, developing correlates
of protection and supporting the epidemiological surveillance studies that will
34
underpin vaccine implementation programmes. Such a standard may also be used

to standardize assays developed for the serodiagnosis of GAS infection. A multi-
centre international collaborative study was therefore being proposed to develop
a WHO international standard for antibodies to group A streptococcus antigens
using pooled serum obtained from either vaccinees or convalescent individuals.
A multiplex immunoassay measuring antibodies against target GAS antigens
would be used to assess the suitability and stability of the candidate materials. It
was anticipated that the proposed collaborative study would be conducted in late
2025 and its results submitted for consideration by the Committee in 2026.
Recognizing the challenge of obtaining serum from convalescent or
vaccinated subjects with reactivity to the relevant antigens, the Committee
was assured that this could be achieved by screening individual donations and
pooling only those exhibiting appropriate reactivities. Given the expected timing
of clinical trials, the project would likely begin by screening convalescent serum
donations. The Committee went on to query the current status of functional
assays and the potential impact of this on the development of the proposed
reference standard. Clarification was given that most advanced such assays were
currently based on antigen-binding platforms, which would therefore guide the
evaluation of the prospective standard.
Acknowledging the timeliness of this project, the Committee endorsed
the proposal (WHO/BS/2024.2472) to develop a First WHO International
Standard for antibodies to group A streptococcus antigens (human serum).
8.2.4 Proposed First WHO International Standard

for antibodies to vaccinia virus
Vaccinia virus (VACV) is a member of the Orthopoxvirus genus that includes
variola, cowpox and Mpox viruses. Antigenic similarity between these viruses
confers cross-reactivity and preclinical studies have shown that VACV vaccines
also provide a level of immunity against Mpox and are efficacious in reducing
Mpox disease severity. This led to the use of VACV-based vaccines for the control
of Mpox during the public health emergency of international concern declared
in 2022. Although this emergency had now been declared over, Mpox remains
a significant burden in endemic countries, with three licensed VACV vaccines
currently available and others under development.
Recognizing the need for serological assays to measure vaccine-induced
immune responses, evaluate new treatments and monitor disease epidemiology,
a proposal to develop a WHO international standard for antibodies to Mpox
virus to support assay development and harmonization had been endorsed by
the Committee in March 2023. During preparations for the collaborative study
to evaluate the candidate materials, it was recognized that several prospective
participants had developed assays capable of distinguishing between the antibody
35
responses to Mpox virus and VACV. Differentiating these responses in both

vaccinated and infected individuals would be important in understanding the
immune responses to vaccination versus infection, interpreting seroprevalence
studies and conducting clinical trials for new vaccines.
The Committee was informed that a candidate material (NIBSC code
05/124) had been prepared in 2005 from pooled defibrinated human plasma
obtained from volunteers vaccinated with the New York City Board of Health
strain of VACV, and then freeze-dried and stored. This material had subsequently
been evaluated in a 2010 collaborative study and proposed for establishment as a
WHO international standard to replace the previous standard for smallpox virus
antibodies. However, due to the limited number of assays used in the study, and
the differential performance of the candidate material compared with the anti-
smallpox serum, its establishment had not been supported. The committee was
informed that approximately 2600 ampoules were still available and although
originally intended for inclusion as one of the samples in the collaborative
study for the Mpox virus antibody standard, it was now proposed to instead
evaluate candidate material 05/124 in parallel for its suitability as a separate
WHO international standard for antibodies to VACV given the ability of some
participant laboratories to distinguish antibody responses to the two viruses.
The Committee was further informed that 16 collaborative study
participants had now been recruited and would analyse the candidate materials
using both neutralization and binding antibody detection methods. Although
the possibility of cross-reactivity between the different orthopoxviruses may
complicate the interpretation of the collaborative study results, it was nevertheless
anticipated that the study outcomes would be submitted for consideration by the
Committee in early 2025.
After due consideration, the Committee endorsed the proposal (WHO/
BS/2024.2472) to develop a First WHO International Standard for antibodies to
vaccinia virus.
8.3 Proposed discontinuations and updates –

vaccines and related substances
8.3.1 Proposal not to proceed with the development of a First WHO
International Standard for antibodies to influenza A(H7N9) virus
In 2014, a proposal to develop a First WHO International Standard for antibodies
to influenza A(H7N9) virus was endorsed by the Committee at a time when
this virus was causing severe respiratory disease in humans in China. Since
2018, changes in zoonotic influenza epidemiology have meant that pandemic
preparedness efforts were now focused on other influenza subtypes and the
Committee was informed that it would not now be possible to source the serum-
36
positive samples needed to prepare the candidate material. In addition, given the
dramatic decline in the number of human infections, and successful control of the
disease in poultry through immunization efforts, the need for this international
standard no longer existed.
While noting the potential value of such a reference material for
pre-pandemic preparedness for the re-emergence of influenza A(H7N9), the
Committee was nevertheless confident that in the event of such re-emergence
that a reference standard could be prepared quickly as there would then be
a ready supply of convalescent serum. Recognizing the challenge in sourcing
material at the current time, the Committee agreed that a final decision on
whether or not to proceed with the development of a First WHO International
Standard for antibodies to influenza A(H7N9) virus would be taken at its
meeting in October 2024.
37
Annex 1
WHO Recommendations, Guidelines and other documents
related to the manufacture, quality control and evaluation
of biological products
WHO Recommendations, Guidelines and other documents in the field of
biological product development and standardization are intended to be scientific
and advisory in nature. Each of these documents provides guidance for national
regulatory authorities (NRAs), developers and manufacturers of biological
products, and others who may have to decide upon appropriate methods for
ensuring that such products are safe, reliable and potent. In the case of WHO
Recommendations and Guidelines, the guidance provided may, if an NRA so
desires, be adopted as definitive national requirements or used as the basis of
such requirements.
WHO Recommendations, Guidelines and other guidance documents for
biological products are formulated by international groups of experts, and are
adopted on the recommendation of the WHO Expert Committee on Biological
Standardization. Following adoption, the documents are published in the WHO
Technical Report Series4 as part of the respective full report of the Committee,
and as listed in this annex. The full reports of the Committee are freely available
for download at https://iris.who.int/. Hard copies of the reports can also be
purchased from:
WHO Press
World Health Organization
20 avenue Appia, 1211 Geneva 27
Switzerland
Email: [email protected]
Website: www.who.int/bookorders
In all cases in which a previous version of a WHO Recommendations,
Guidelines or other guidance document has been revised and superseded by an
updated document, it is of paramount importance that only the latest version of
the document is used. All documents listed in this annex are current, with no
previous versions shown. The annex has also been arranged alphabetically either
by product type or regulatory topic to facilitate easy identification of the most
up-to-date document.
4
Abbreviated in the following pages to “TRS”.
39
Important note on the immediate discontinuation of the innocuity test

At its sixty-ninth meeting in 2018, the Committee recommended the immediate
discontinuation of any requirement to perform the innocuity test (also
known as the abnormal toxicity test or general safety test) in all future WHO
Recommendations, Guidelines and other guidance documents for biological
products published in the Technical Report Series.5 The Committee further
recommended that any mention of this test in any still current WHO document
published prior to 2018 be disregarded (Table 1).
These recommendations represent a significant step towards increasingly
science-based regulation and regulatory convergence at the global level, while
also promoting the application of the 3Rs principles (Replacement, Refinement,
Reduction) by reducing the use of animals in biological product quality control
and lot release testing.
Table 1
List of current WHO documents for biological products published prior to 2018 in
which any mention of the innocuity test (also known as the abnormal toxicity test or
general safety test) should be disregarded
Product WHO document Name of test as it

appears in document
Dengue fever vaccines (live, Annex 2: TRS 979 General safety
attenuated)
Diphtheria vaccines (adsorbed) Annex 4: TRS 980 Innocuity
DT-based combined vaccines Annex 6: TRS 980 [Control of final product]
Ebola vaccines Annex 2: TRS 1011 General safety (innocuity)
Hepatitis A vaccines (inactivated) Annex 2: TRS 858 General safety

Hepatitis B vaccines (recombinant) Annex 4: TRS 978 General safety (innocuity)
HFRS vaccines (inactivated) Annex 2: TRS 848 General safety
Haemophilus influenzae type b Annex 1: TRS 897 General safety (innocuity)
conjugate vaccines
Human papilloma virus VLP Annex 4: TRS 999 General safety (innocuity)
vaccines
5
Lei D, Schmidt H, Knezevic I, Zhou T, Kang H-N, Kopp S. Removal of the innocuity test from The International
Pharmacopoeia and WHO recommendations for vaccines and biological products. Biologicals. 2020;66:17–20
(https://www.sciencedirect.com/science/article/pii/S1045105620300610, accessed 27 December 2023).
40
Annex 1
Table 1 continued
appears in document
Human interferons Annex 3: TRS 786 Innocuity
Influenza vaccines (inactivated) Annex 3: TRS 927 General safety (innocuity)
Japanese encephalitis vaccines Annex 1: TRS 963 General safety (innocuity)
(inactivated)
Japanese encephalitis vaccines Annex 7: TRS 980 General safety
(live, attenuated)
Malaria vaccines (recombinant) Annex 3: TRS 980 General safety
Meningococcal A conjugate Annex 2: TRS 962 General safety (innocuity)
vaccines
Meningococcal C conjugate Annex 2: TRS 924 General safety (innocuity)
vaccines
Meningococcal polysaccharide Annex 2: TRS 594 Abnormal toxicity
vaccines (unconjugated) Annex 2 TRS 904
MMR and combined vaccines (live) Annex 3: TRS 840 General safety
Pertussis vaccines (acellular) Annex 4: TRS 979 Innocuity
Pertussis vaccines (whole cell) Annex 6: TRS 941 General safety (innocuity)
Pneumococcal conjugate vaccines Annex 3: TRS 977 General safety (innocuity)
Poliomyelitis vaccines (inactivated) Annex 3: TRS 993 General safety (innocuity)
Amendment (requirement Annex 3: TRS 1024
removed)
Rabies vaccines (inactivated) Annex 2: TRS 941 General safety (innocuity)
Rift Valley fever vaccines Annex 4: TRS 673 Innocuity
(inactivated)
Smallpox vaccines Annex 1: TRS 926 General safety (innocuity)
Snake antivenom immunoglobulins Annex 5: TRS 1004 Abnormal toxicity
Synthetic peptide vaccines Annex 1: TRS 889 Routine control
Tetanus vaccines (adsorbed) Annex 5: TRS 980 Innocuity
Tick-borne encephalitis vaccines Annex 2: TRS 889 General safety
(inactivated)
41
Table 1 continued
appears in document
Typhoid vaccines (live attenuated, Annex 3: TRS 700 Innocuity
Ty 21a, oral)
Typhoid vaccines (Vi polysaccharide) Annex 1: TRS 840 Abnormal toxicity
Vaccines (stability evaluation of ) Annex 3: TRS 962 General safety
Abnormal toxicity
Varicella vaccine (live) Annex 1: TRS 848 General safety
Yellow fever vaccines (live, Annex 5: TRS 978 General safety
attenuated)
DT = diphtheria and tetanus; HFRS = haemorrhagic fever with renal syndrome; VLP = virus-like particle;
MMR = measles, mumps and rubella.
42
Annex 1
List of all current WHO documents for biological products

Recommendations, Guidelines and other Reference
documents
Animal cells: use of as in vitro substrates for the Revised 2010, TRS 978 (2013)
production of biologicals
BCG vaccines (dried) Revised 2011, TRS 979 (2013)
Biological products: good manufacturing practices Revised 2015, TRS 999 (2016)
Biosimilars: evaluation of Revised 2022, TRS 1043 (2022)
Biotherapeutic products: changes to approved Adopted 2017, TRS 1011 (2018)
biotherapeutic products; procedures and data
requirements
Biotherapeutic protein products prepared by Revised 2013, TRS 987 (2014);
recombinant DNA technology Addendum 2015 on regulatory
assessment of approved rDNA-
derived biotherapeutics, TRS
999 (2016)
Blood, blood components and plasma derivatives: Revised 1992, TRS 840 (1994)
collection, processing and quality control
Blood and blood components: management as Adopted 2016, TRS 1004 (2017)
essential medicines
Blood components and plasma: estimation of Adopted 2016, TRS 1004 (2017)
residual risk of HIV, HBV or HCV infections
Blood establishments: good manufacturing Adopted 2010, TRS 961 (2011)
practices6
Blood plasma (human) for fractionation Adopted 2005, TRS 941 (2007)
Blood plasma products (human): viral inactivation Adopted 2001, TRS 924 (2004)
and removal procedures
Blood regulatory systems: assessment criteria for Adopted 2011, TRS 979 (2013)
national
Cholera vaccines (inactivated, oral) Adopted 2001, TRS 924 (2004)
Dengue tetravalent vaccines (live, attenuated) Revised 2011, TRS 979 (2013)
Diphtheria and tetanus based combined vaccines Revised 2012, TRS 980 (2014)
6
Adopted on the recommendation of the WHO Expert Committee on Specifications for Pharmaceutical
Preparations, with significant input from the WHO Expert Committee on Biological Standardization.
43

documents
Diphtheria vaccines (adsorbed) Revised 2012, TRS 980 (2014)
DNA vaccines: plasmid Revised 2020, TRS 1028 (2021)
Ebola vaccines Adopted 2017, TRS 1011 (2018)
Enterovirus 71 vaccines (inactivated) Adopted 2020, TRS 1030 (2021)
Haemophilus influenzae type b conjugate vaccines Revised 1998, TRS 897 (2000)
Haemorrhagic fever with renal syndrome vaccines Adopted 1993, TRS 848 (1994)
(inactivated)
Hepatitis A vaccines (inactivated) Adopted 1994, TRS 858 (1995)
Hepatitis B vaccines prepared from plasma Revised 1994, TRS 858 (1995)
Hepatitis B vaccines (recombinant) Revised 2010, TRS 978 (2013)
Hepatitis E vaccines (recombinant) Adopted 2018, TRS 1016 (2019)
Human cells and tissues and advanced therapy Adopted 2023, TRS 1048 (2023)
medicinal products: regulatory considerations
Human immunodeficiency virus rapid diagnostic Adopted 2017, TRS 1011 (2018)
tests for professional use and/or self-testing:
Technical Specifications Series for WHO
Prequalification – Diagnostic Assessment
Human interferons prepared from lymphoblastoid Adopted 1988, TRS 786 (1989)
cells
Influenza vaccines (inactivated) Revised 2003, TRS 927 (2005);
Addendum 2016 on labelling
information of inactivated
influenza vaccines for use in

pregnant women, TRS 1004
(2017)
Influenza vaccines (live) Revised 2009, TRS 977 (2013)
Influenza vaccines: human, pandemic; regulatory Adopted 2007, TRS 963 (2011)
preparedness
Influenza vaccines: human, pandemic; safe Adopted 2018, TRS 1016 (2019)
development and production of
International and other biological reference Revised 2004, TRS 932 (2006)
standards: preparation, characterization and
establishment of
44
Annex 1

documents
In vitro diagnostics (WHO-prequalified): Adopted 2020, TRS 1030 (2021)
collaborative procedure between WHO and
NRAs for assessment and accelerated national
registration
In vitro diagnostic medical devices: establishing Adopted 2017, TRS 1011 (2018)
stability of; Technical Guidance Series for WHO
Prequalification – Diagnostic Assessment
Japanese encephalitis vaccines (inactivated) for Revised 2007, TRS 963 (2011)
human use
Japanese encephalitis vaccines (live, attenuated) for Revised 2012, TRS 980 (2014)
human use
Louse-borne human typhus vaccines (live) Adopted 1982, TRS 687 (1983)
Malaria vaccines (recombinant) Adopted 2012, TRS 980 (2014)
Measles, mumps and rubella vaccines and Adopted 1992, TRS 840 (1994);
combined vaccines (live) Note 1993 TRS 848 (1994)
Medical devices including in vitro diagnostic Revised 2022, TRS 1045 (2023)
medical devices: WHO Global Model Regulatory
Framework; WHO Medical device technical series
Meningococcal A conjugate vaccines Adopted 2006, TRS 962 (2011)
Meningococcal C conjugate vaccines Adopted 2001, TRS 924 (2004);
Amendment 2007, TRS 963
(2011)
Meningococcal polysaccharide vaccines Adopted 1975, TRS 594 (1976);
Addendum 1980, TRS 658 (1981);
(2002)
Monoclonal antibodies as similar biotherapeutic Adopted 2016, TRS 1004 (2017)
products
Monoclonal antibodies and related products Adopted 2023, TRS 1048 (2023)
against infectious diseases: nonclinical and clinical Addendum 2024 on
evaluation monoclonal antibodies and
related products against
COVID-19, TRS 1059 (2024)
Monoclonal antibodies: production and quality Revised 2022, TRS 1043 (2022)
control
45

documents
Pandemic or other emergency use vaccines: Revised 2023, TRS 1054 (2024)
regulatory preparedness for oversight in importing
countries
Papillomavirus vaccines (human, recombinant, Revised 2015, TRS 999 (2016)
virus-like particle)
Pertussis vaccines (acellular) Revised 2011, TRS 979 (2013)
Pertussis vaccines (whole-cell) Revised 2005, TRS 941 (2007)
Pharmaceutical products: storage and transport of Adopted 2010, TRS 961 (2011)
time- and temperature-sensitive7
Pneumococcal conjugate vaccines Revised 2009, TRS 977 (2013)
Poliomyelitis vaccines (inactivated) Revised 2014, TRS 993 (2015);
(2020)
Poliomyelitis vaccines (oral) Revised 2022, TRS 1045 (2023)
Poliomyelitis vaccines: safe production and quality Revised 2018, TRS 1016 (2019)
control Amendment 2020, TRS 1028
(2021)
Quality assurance for biological products: Adopted 1991, TRS 822 (1992)
guidelines for national authorities
Rabies vaccines for human use (inactivated) Revised 2005, TRS 941 (2007)
produced in cell substrates and embryonated eggs
Reference materials: secondary; for antibody Adopted 2022, TRS 1043 (2022)
testing
Reference materials: secondary; for NAT-based Adopted 2016, TRS 1004 (2017)
and antigen assays; calibration against WHO
International Standards
Regulation and licensing of biological products Adopted 1994, TRS 858 (1995)
in countries with newly developing regulatory
authorities
Regulatory risk evaluation on finding an adventitious Adopted 2014, TRS 993 (2015)
agent in a marketed vaccine: scientific principles
7
Adopted on the recommendation of the WHO Expert Committee on Specifications for Pharmaceutical
Preparations, with significant input from the WHO Expert Committee on Biological Standardization.
46
Annex 1

documents
Respiratory syncytial virus vaccines Adopted 2019, TRS 1024 (2020)
RNA vaccines: messenger; for prevention of Adopted 2021, TRS 1039 (2022)
infectious diseases
Rotavirus vaccines (live, attenuated, oral) Adopted 2005, TRS 941 (2007)
Smallpox vaccines Revised 2003, TRS 926 (2004)
Snake antivenom immunoglobulins Revised 2016, TRS 1004 (2017)
Sterility of biological substances Revised 1973, TRS 530 (1973);
(1998)
Synthetic peptide vaccines Adopted 1997, TRS 889 (1999)
Tetanus vaccines (adsorbed) Revised 2012, TRS 980 (2014)
Thiomersal for vaccines: regulatory expectations for Adopted 2003, TRS 926 (2004)
elimination, reduction or replacement
Thromboplastins and plasma used to control oral Revised 2011, TRS 979 (2013)
anticoagulant therapy
Tick-borne encephalitis vaccines (inactivated) Adopted 1997, TRS 889 (1999)
Tuberculins Revised 1985, TRS 745 (1987)
Typhoid vaccines, conjugated Revised 2020, TRS 1030 (2021)
Typhoid vaccines (live, attenuated, Ty21a, oral) Adopted 1983, TRS 700 (1984)
Typhoid vaccines: Vi polysaccharide Adopted 1992, TRS 840 (1994)
Vaccines: changes to approved vaccines; Adopted 2014, TRS 993 (2015)
procedures and data requirements
Vaccines: clinical evaluation; regulatory Revised 2016, TRS 1004 (2017)
expectations
Vaccines: regulatory considerations; use of human Adopted 2016, TRS 1004 (2017)
challenge trials
Vaccines: lot release Adopted 2010, TRS 978 (2013)
Vaccines: nonclinical evaluation Adopted 2003, TRS 927 (2005)
Vaccines: nonclinical evaluation of vaccine Adopted 2013, TRS 987 (2014)
adjuvants and adjuvanted vaccines
47

documents
Vaccines: prequalification procedure Adopted 2010, TRS 978 (2013)
Vaccines: stability evaluation Adopted 2006, TRS 962 (2011)
Vaccines: stability evaluation for use under Adopted 2015, TRS 999 (2016)
extended controlled temperature conditions
Varicella vaccines (live) Revised 1993, TRS 848 (1994)
Yellow fever vaccines (live, attenuated) Revised 2010, TRS 978 (2013)
(2022)
Yellow fever virus: production and testing of WHO Adopted 1985, TRS 745 (1987)
primary seed lot 213-77 and reference batch 168-736
48
Annex 2
Nonclinical and clinical evaluation of monoclonal
antibodies and related products intended for the
prevention or treatment of COVID-19
Addendum to Annex 2 of WHO Technical Report Series, No.1048
1. Introduction 52
2. Purpose and scope 52
3. Terminology 53
4. General considerations 53
5. International reference materials 55
6. Nonclinical evaluation 55
6.1 Pharmacodynamics studies 56
6.2 In vivo studies 58
7. Clinical evaluation 62
7.1 Inclusion and exclusion criteria 64
7.2 Phase I studies 66
7.3 Clinical pharmacology 66
7.4 Phase II and III studies 67
Authors and acknowledgements 73
References 74
49
Guidelines and their addenda published by the World Health

Organization (WHO) are intended to be scientific and advisory
in nature. Each of the following sections constitutes guidance for
national regulatory authorities (NRAs) and for manufacturers
of biological products. If an NRA so desires, the parent WHO
Guidelines and this addendum may be adopted as definitive national
requirements, or modifications may be justified and made by the NRA.
It is recommended that modifications to the Guidelines and/or this
addendum are made only on condition that such modifications ensure
that the product is at least as safe and efficacious as that prepared in
accordance with the guidance set out.
50
Annex 2
Abbreviations
ACE2 angiotensin-converting enzyme 2 (receptor)
ADA anti-drug antibody
ADE antibody-dependent enhancement (of disease)
AE adverse event
AESI adverse event of special interest
COVID-19 coronavirus disease 2019
Fc fragment crystallizable (region)
FcγR Fc gamma receptor
GMT geometric mean titre
IMP investigational medicinal product
MAAE medically attended adverse event
mAb monoclonal antibody
PD pharmacodynamics
PK pharmacokinetics
NRA national regulatory authority
RBD receptor binding domain
RT-PCR reverse transcription-polymerase chain reaction
S spike (glycoprotein)
SAE serious adverse event
SARS-CoV-2 severe acute respiratory syndrome coronavirus 2
51
1. Introduction
Evaluating the safety and efficacy of monoclonal antibodies (mAbs) and related
products intended for the prevention or treatment of infectious diseases requires
different considerations than mAb products that target endogenous proteins,
such as those intended for the treatment of noncommunicable diseases. To help
address such differences, the WHO Guidelines on the nonclinical and clinical
evaluation of monoclonal antibodies and related products intended for the
prevention or treatment of infectious diseases (1) was adopted in 2023 on the
recommendation of the WHO Expert Committee on Biological Standardization.
These Guidelines outline the general principles applicable to the evaluation
of mAbs for use against infectious diseases. However, although the document
provides guidance on evaluating the safety and efficacy of mAb products
regardless of the targeted pathogen, it was recognized that pathogen-specific
considerations would potentially affect the interpretation and application of the
guidance provided.
2. Purpose and scope

The current addendum is intended to provide supplementary considerations
when evaluating the safety and efficacy of mAb products directed specifically
against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antigens.
This includes mAb products intended for pre- and post-exposure prophylaxis
as well as for the treatment of coronavirus disease 2019 (COVID-19). These
considerations are applicable to mAbs and related products, including single
and co-formulated mAbs against SARS-CoV-2. However, some mAb products
(for example, bispecific mAbs or those with a different mechanism of action)
may require additional nonclinical studies and the NRA should be consulted on
the need for such studies. Unless otherwise indicated, the guidance applies to
products that are administered parenterally.

It should be noted that mAbs and related products that target endogenous
human antigens (for example, those which block the angiotensin-converting
enzyme 2 (ACE2) receptor or cytokines) are not within the scope of this
addendum as these require different considerations for evaluating their safety
and efficacy.
Separate and detailed guidance on the production and quality control of
mAbs is provided in the WHO Guidelines for the production and quality control
of monoclonal antibodies and related products intended for medicinal use (2).
52
Annex 2
3. Terminology
The following definitions apply to the terms as used in this addendum. These
terms may have different meanings in other contexts. It should be noted that
additional terms relevant to this addendum are defined in the WHO Guidelines
on the nonclinical and clinical evaluation of monoclonal antibodies and related
products intended for the prevention or treatment of infectious diseases (1).
COVID-19: the disease caused by infection with SARS-CoV-2.
Long COVID: health problems that persist or develop after infection
with SARS-CoV-2, and which may last from several weeks to years.
SARS-CoV-2: the virus that causes COVID-19.
Variant: a virus that possesses mutations which may confer altered
transmissibility, receptor binding affinity, virulence, morbidity or mortality. A
number of SARS-CoV-2 variants have been labelled as “variants of interest”
(VOI), “variants of concern” (VOC) or variants under monitoring (VUM)
depending on their emerging dominance among actively circulating strains.
4. General considerations
SARS-CoV-2 is an enveloped positive-sense single-stranded RNA virus belonging
to the genus Betacoronavirus. The virus first emerged in Wuhan, China in 2019,
with sustained human-to-human transmission confirmed shortly afterwards,
followed by its rapid spread worldwide. Evidence of sustained global transmission
led WHO to declare COVID-19 a pandemic in March 2020. COVID-19 was
subsequently officially declared to no longer be a public health emergency of
international concern by WHO on 5 May 2023. Nevertheless, the disease remains
a major threat, with SARS-CoV-2 still in circulation in most regions of the world.
SARS-CoV-2 is transmitted primarily by the respiratory route producing
a mucosal infection after a short incubation period that results in a range of
disease symptoms and outcomes – from asymptomatic to severe disease leading to
hospitalization and death, as well as long COVID in some cases (3–7). Moreover,
there is a possibility that SARS-CoV-2 will become endemic and will continue
to cause substantial levels of hospitalization and death due to the emergence of
new variants. This is of particular concern among vulnerable groups such as the
immunocompromised and those with underlying comorbidities (8, 9).
Monoclonal antibodies, vaccines and other therapeutics against
SARS-CoV-2 were developed rapidly and authorized for use, initially under
emergency procedures. These early mAbs (10–12) and other products were all
based on the genetic sequence of the ancestral Wuhan strain and provided
protection against severe disease, hospitalization and death. However, to date,
no correlates of protection or threshold of protection have been established and
53
challenges remain in directly comparing the neutralizing antibody activity of

different products (13). It is clear that some of these early products now exhibit
reduced virus neutralization activity against SARS-CoV-2 variants, and a number
have been withdrawn from use, especially in regions with high levels of Omicron
circulation (10, 11, 14, 15). Although most early authorized mAbs have to varying
degrees lost their ability to neutralize variant strains, this finding is primarily based
on in vitro testing and does not necessarily reflect clinical experience (10, 13).
Protection against COVID-19 was initially provided both through SARS-CoV-2
neutralizing antibodies and through the induction of broader cellular immunity
following infection and/or vaccination (16). To date, although SARS-CoV-2
variants have evolved to evade neutralizing antibodies, with consequences for
infection, virus shedding and transmission, these have not significantly affected
the longer lasting and broader cellular immune responses (10, 11, 16).
Nevertheless, it is clear that the emergence of variant strains of
SARS-CoV-2 poses a major challenge to product development and evaluation
of efficacy in clinical use (9, 17–19). In response to this challenge, considerable
efforts are now under way to develop SARS-CoV-2 mAbs and vaccines that are
escape resistant (10, 11, 15, 20). For example, progress in B-cell technologies has
accelerated the identification and rapid isolation of candidate antibodies which
can overcome variants arising through antigenic shift (10, 11, 21).
The principal target of both mAb and vaccine development has been the
virus trimeric transmembrane spike (S) glycoprotein which protrudes from the
virus surface and mediates its entry into host cells by binding to the ACE2 receptor
(10, 22). Entry requires cleavage of the S transmembrane glycoprotein generating
S1 and S2 subunits to initiate fusion of the viral and cell membranes upon entry.
The SARS-CoV-2 S glycoprotein contains a furin-like cleavage site for host cell
proteases (23). Neutralizing mAbs targeting the receptor binding domain (RBD)
of the S glycoprotein, the S2 subunit or the S1/S2 proteolytic cleavage site have
been variously affected by the emergence of SARS-CoV-2 variants (10, 11, 14,
20, 24, 25). To expedite the development of new mAbs, consideration is being
given by some but not all regulatory authorities to immunobridging studies in
support of licensure (see section 7.4.2 below). However, it is important that such
an approach be discussed directly with the relevant NRA.
Clearly, the ongoing evolution of SARS-CoV-2 requires continuous
monitoring for significant changes in local circulating variant strains which
might impact the performance of mAbs, both in clinical studies and in use
(26). Similarly, careful attention needs to be given to the virus strains used in
nonclinical and clinical evaluation studies to ensure that the virus preparation
used is well characterized and standardized with respect to variant strains (27).
Although the emergence of resistant variants of SARS-CoV-2 is an issue
of concern with regard to efficacy, no major safety signals have been identified
regarding the use of mAbs to prevent or treat COVID-19. However, the potential
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for antibody-dependent enhancement (ADE) of disease is always a possibility, and

is an important aspect to consider as part of nonclinical and clinical evaluation
programmes (28, 29). The complexities of assessing and predicting mAb-induced
clinical ADE of disease in humans, including the poor predictability of both in
vitro systems and animal models, are discussed in detail elsewhere (28). Particular
attention should be given to the effects of any engineered modifications of the
fragment crystallizable (Fc)-mediated effector functions of mAbs that may, for
example, have been made to increase the half-life of the antibody (10, 11).
5. International reference materials

WHO international reference standards are the primary reference materials
used worldwide and such standards are available for SARS-CoV-2 antibodies to
support the development of serological assays and to increase the comparability
of results obtained by different laboratories. The WHO international reference
standards related to SARS-CoV-2 antibodies available at the time of publication
of the current document are:
■ Second WHO International Standard for anti-SARS-CoV-2
immunoglobulin (30);
■ First WHO International Standard for antibodies to SARS-CoV-2
variants of concern (31); and
■ First WHO International Reference Panel for antibodies to
SARS-CoV-2 variants of concern (32), and subsequent panel
expansion to include Gamma and Omicron variants (33).
Although the above standards are likely to be useful for laboratories
characterizing SARS-CoV-2 mAbs, further studies are needed to determine
whether these polyclonal plasma standards can effectively harmonize the
measurement of mAb neutralizing activity between laboratories.
Furthermore, it should be noted that when stocks become exhausted
or new variants emerge, new or replacement standards are established on the
recommendation of the WHO Expert Committee on Biological Standardization.
Users should take steps to ensure use of the most recent and appropriate WHO
international standard, international reference panel or other international
reference standard.
6. Nonclinical evaluation
There are several important factors to consider when designing nonclinical
studies for mAbs intended to prevent or treat SARS-CoV-2 infection. Such
studies should characterize the targeted SARS-CoV-2 binding site/epitope and
55
the ability of the mAb to neutralize virus variants. The primary pharmacological
effector functions of the mAb should be considered, especially if the Fc region
of the mAb has been engineered. Any potential risk of unwanted or unexpected
cross-reactivity with human cells or tissues, ADE or viral resistance should also
be explored.
For the assessment of inhaled or intranasally administered mAbs, the
selection of an animal model should take into consideration the differences in the
anatomy and physiology of human and animal respiratory systems. The animal
model selected should be justified when designing proof-of-concept studies for
demonstrating mAb antiviral activity. If a delivery device (for example, nebulizer
or dry powder inhaler) is required, its mechanism of delivery should be similar to
the device intended for clinical use in humans. In some cases, additional studies
may be required to ensure optimal conditions for mAb delivery in the animal
model to be used.
6.1 Pharmacodynamics studies

The pharmacodynamics (PD) of the mAb should be characterized using in vitro
assays.
6.1.1 Target antigen or epitope

To date, all neutralizing mAbs against COVID-19 target the SARS-CoV-2 S
protein and exhibit both antigen binding and neutralizing activity. The ability of
such a mAb to recognize the S protein should be demonstrated and its binding
affinity measured. In addition, inhibition of binding of the S protein RBD to the
human target ACE2 receptor should be demonstrated.
The epitope on the S protein targeted by the mAb should be identified for
single-formulated mAbs. In the case of co-formulated mAbs (where there are two
or more mAbs within a final product) or bispecific mAbs that target two binding
epitopes, each targeted binding epitope should be identified. This is to ensure

that the co-formulated or bispecific mAbs do not compete for the same epitope
or have overlapping epitopes that could lead to antagonism.
In future, mAbs that target relevant SARS-CoV-2 antigens other than the
S protein may be developed. In such cases, the ability of the mAb to recognize
the targeted virus epitope should be demonstrated. If the mAb is intended to
inhibit virus binding to a human cell then its ability to prevent virus binding and
subsequent infection should be demonstrated.
6.1.2 Virus neutralization assays

The primary antiviral mechanism of mAbs is direct virus neutralization. The in
vitro virus neutralization activity of the mAb should be assessed against historical,
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Annex 2
currently dominant and emerging variants. Virus neutralization activity can be

demonstrated using a live virus assay (for example, focus-reduction neutralization
test, plaque-reduction neutralization test or surrogate neutralization assay) and/
or a pseudovirus neutralization assay (34, 35).
For co-formulated mAbs, the virus neutralization activity of each
constituent mAb should be tested and any synergistic activity reported. For
bispecific mAbs, the virus neutralization activity of each independent targeted
epitope should be tested and reported.
6.1.3 Effector function assays

The need for effector function assays should be justified and is predominately
important in the assessment of immunoglobulin G1 products. The secondary
antiviral mechanism of mAbs is the effector functions driven by Fc gamma
receptor (FcγR) interactions. The effector properties of the mAb, such as
antibody-dependent cellular cytotoxicity (ADCC), antibody-dependent cellular
phagocytosis (ADCP) and complement-dependent cytotoxicity (CDC), should
be assessed.
If the Fc region of the mAb has been engineered, the engineered
pharmacological effects, such as extending mAb half-life or attenuation of Fc
binding activity to Fc receptors, should be assessed and reported.
6.1.4 Assessment of antibody-dependent enhancement of disease

Clinical evidence of ADE of COVID-19 is limited and this has not been
observed with currently approved mAb therapies or COVID-19 vaccines in
humans. However, mAbs can mediate ADE via FcγR interaction or complement
component C1q (36, 37), and so the potential risk of ADE cannot be ruled out.
Therefore, evaluation of the potential for ADE of disease should be carried out
and the results reported. The selection of an in vitro assay(s) for assessing ADE
should take into consideration current understanding of SARS-CoV-2 infection
and ADE, and the reliability of the assay(s) in predicting mAb-induced clinical
ADE of disease in humans. Although in vitro assays have limitations, they may
provide useful information relevant to the potential risk of ADE (for example,
information on virus neutralization, virus uptake and infectivity, or cytokine
production) (28).
6.1.5 Virus resistance assessment

The effectiveness of mAbs has been threatened by the emergence of resistant
SARS-CoV-2 variants, with reductions in the magnitude of neutralization
of such variants using existing treatments reported (38). Therefore, the
neutralization activity of the mAb against these variants should be evaluated in
57
virus neutralization assays using emerging variants from clinical surveillance,

experimentally derived viral escape mutants and/or modelled predicted
escape mutants. In addition, the risk of emergence of resistant viruses to the
investigational mAb under treatment due to the use of suboptimal doses should
be investigated in vitro before initiation of clinical studies. Where resistance
is observed, genotyping, phenotyping and cross-resistance analyses of the
potential escape mutants should be conducted.
6.2 In vivo studies

For animal proof-of-concept studies demonstrating antiviral activity, preference
should be given to animal models in which the SARS-CoV-2 infection is
reflective of the human infection and of the anticipated mechanism of action of
the mAb. The similarity of SARS-CoV-2 infection in the chosen animal model
to human infection and disease should be described and justified. With the
ongoing detection of SARS-CoV-2 variants, consideration should also be given
to assessing the similarity of infection across different variants in the animal
model and in humans.
Several animal models for COVID-19 have been developed for
the testing of mAbs and vaccines.7 Each of the following models are able to
reproduce some aspects of the clinical and pathological features of COVID-19
in humans (39, 40).
■ Syrian hamsters have been established as an animal model for
COVID-19 due to the similarity of hamster and human ACE2.
Viral replication is observed in the respiratory and gastrointestinal
tracts following infection. Laboured breathing and weight loss are
observed clinical symptoms. Severe interstitial pneumonia with
inflammation is observed more in aged or male hamsters compared
to young or female hamsters. The induction of serum neutralization
antibodies has been observed following infection. In addition,

hamsters have been shown to transmit SARS-CoV-2 by both close
contact and non-contact routes.
■ Normal mice are not a relevant model for COVID-19 as
SARS-CoV-2 does not bind effectively to mouse ACE2. However,
transgenic mice expressing human ACE2 or the use of mouse-
adapted SARS-CoV-2 strains have made the mouse a useful model.
Mice display a range of clinical symptoms (such as weight loss)
and pathological disease symptoms (such as lung inflammation)
7
Any animal species used for in vivo studies should be chosen carefully and thoroughly justified. For
scientific and ethical reasons, it is desirable to apply the 3Rs principles of “Replace, Reduce, Refine”.
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Annex 2
that vary in severity. The use of mouse-adapted virus models or

human ACE2 transgenic mice can also be a useful tool for studying
infection with SARS-CoV-2 variants.
■ Ferrets have long been a model for studying human respiratory
viruses such as influenza and respiratory syncytial virus, and are
now being used to investigate SARS-CoV-2 transmission and
COVID-19. Ferrets display mild clinical disease that includes fever,
wheezing and nasal discharge. Virus replication is observed in the
respiratory and gastrointestinal tracts. Histopathological studies
have shown pneumonia with lung inflammation. Transmission
studies have demonstrated virus transmission by both close contact
and non-contact routes, suggesting that airborne transmission of
SARS-CoV-2 among ferrets is possible, making them a useful model
for such studies.
■ Non-human primates have been infected with SARS-CoV-2 variants
and studies have shown high levels of viral replication in both the
upper and lower respiratory tract. Non-human primates display
mild clinical disease but with notable histopathology findings of
pneumonia. Severe disease has been observed in aged non-human
primates. The induction of natural protective immunity through
innate, humoral and cellular immune responses following infection
has also been observed. Non-human primates should only be
considered as a last resort option, and the selection of this model
should be extensively justified.
Based on the above differences in clinical and pathological aspects, the
selection of animal models for characterizing the potential clinical use of the
mAb (for prophylaxis or treatment, or both) should be justified. Furthermore,
the design of the proof-of-concept study should also reflect the intended clinical
use(s) of the mAb (that is, for pre-exposure prophylaxis and/or post-exposure
prophylaxis, and/or for treatment).
The characteristics of SARS-CoV-2 infection and COVID-19 outcomes
in the above animal models are summarized in Table 1. This summary table is
provided here for information only and the more detailed information available
in the scientific literature on the use of selected animal models for SARS-CoV-2
and COVID-19 (39–41) should be taken into consideration when designing
proof-of-concept studies.
59
Table 1
SARS-CoV-2 infection characteristics and disease outcomes in animal models
Relevant animal models Infection characteristics and disease outcome

Rodent
Syrian hamster • Susceptible to SARS-CoV-2 as hamster ACE2 is
similar to human ACE2
• Main clinical disease symptom is weight loss
• High levels of viral replication in lungs at early stage
after infection followed by rapid decline in virus
levels, and no virus detected 1 week after challenge
• Lung histopathological changes observed (for
example, pneumonia, inflammatory cell infiltration)
• Severe clinical disease observed more often in males
than in females and aged hamsters
• Naturally clear infection
Transgenic mouse • Expressed human ACE2 (hACE2) allows SARS-CoV-2
infection
• Main clinical disease symptom is weight loss but
some models may also show respiratory distress
• Respiratory tract infection following virus challenge
but other organs (for example, brain, heart) may be
infected due to secondary infection or expression of
hACE2
• Lung histopathological changes observed (for
example, pneumonia, diffuse alveolar damage,
inflammatory cell infiltration)
• Severe disease observed when challenged with
higher viral load
Mouse-adapted SARS-CoV-2 • Mutations (N501Y or Q498T and P499Y) in RBD
mouse region of SARS-CoV-2 makes it adaptive to mouse

ACE2
• Clinical signs typically limited to mild weight loss
but loss of pulmonary function also observed with
mouse-adapted virus carrying Q498T and P499Y
mutations
• Mild to moderate pneumonia observed with
mouse-adapted virus carrying N501Y mutation
• Respiratory tract infection observed
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Annex 2
Table 1 continued
Relevant animal models Infection characteristics and disease outcome
Other
Ferret • Naturally susceptible to SARS-CoV-2
• Clinical symptoms similar to humans (fever and mild
respiratory symptoms)
• Viral replication observed in respiratory tract (nasal
wash, lungs)
• Interstitial pneumonia observed
• No deaths from infection observed
Non-human primate – non-human primates should only be considered as a last
resort option, and the selection of this model should be extensively justified
Rhesus macaque • Mild clinical disease (for example, fever, weight loss)
• High levels of viral replication in respiratory tract
(detected by nasal swab, bronchoalveolar lavage,
lung tissue examination); lung histopathological
changes (for example, pneumonia, pulmonary
discoloration) similar to humans
• Severe disease not observed
• Naturally cleared infection
Cynomolgus macaque • Mild clinical disease (for example, fever, weight loss)
• High levels of viral replication in respiratory tract
(detected by nasal swab, bronchoalveolar lavage,
lung tissue examination); lung histopathological
changes (for example, diffuse alveolar damage,
pulmonary discoloration) similar to humans
• Severe disease not observed
• Naturally cleared infection
African green monkey • Clinical disease and histopathological changes
similar to rhesus or cynomolgus macaques
• Severe disease (acute respiratory distress syndrome)
observed in aged monkeys
Although the use of animal models has advanced the development of

several COVID-19 therapeutics, there is no specific animal model optimized
to mimic human SARS-CoV-2 infection and COVID-19. The selection of
appropriate animal models for proof-of-concept studies should take into
consideration the disease outcome of each animal model with regard to the
intended study end-points.
The design of proof-of-concept studies should also ensure the use of a
well-characterized virus challenge strain and acceptable route of inoculation.
61
The minimum anticipated biological effect level (MABEL) or biological effective

dose (BED) should be used to select dose levels and to optimize the anticipated
therapeutic effect.
7. Clinical evaluation
There are several factors to be considered in the clinical development programmes
of anti-SARS-CoV-2 mAbs as they impact the clinical trial design and end-points
to be used. One important consideration is whether the product is intended to
be used as a prophylactic (for pre-exposure prophylaxis and/or post-exposure
prophylaxis), as a therapeutic, or both.
For treatment indications, the timing of administration of the mAb is
especially relevant. Current anti-SARS-CoV-2 mAbs were generally found to be
more effective when administered early to patients with symptomatic COVID-19
and prior to hospitalization (42, 43). Some, but not all, studies suggest that such
mAbs may be associated with worse outcomes for patients requiring high-flow
oxygen or mechanical ventilation (44–46).
Inhaled or intranasally administered mAbs are currently under
development and may provide some advantages due to more-localized
administration and lower systemic exposure. Additional considerations may
be required for these alternative routes of administration, such as compatibility
of the delivery device with the mAb formulation, mAb distribution within the
airways and potential for systemic exposure. In addition, robust pharmacokinetic
and pharmacodynamic modelling should be performed. For efficacy evaluation,
consideration may be given to measuring the prevention of infection. Sponsors
should consult with the NRA to help ensure a comprehensive regulatory approach
for such products.
Because of the functionality of the mAb, healthy volunteers may not
be suitable candidates for therapeutic efficacy trials, but may be appropriate for
prophylactic studies. Healthy volunteers may also provide useful data on product
safety, preliminary pharmacokinetics (PK) and potential for anti-drug antibody
(ADA) induction in Phase I studies. PK parameters may require confirmation
in infected patients to highlight any differences compared to healthy volunteers.
As repeated administration of the mAb may alter its safety and activity profiles,
repeat-dosing studies should be conducted to support the use of additional
administrations.
Clinical trial duration can vary depending on the biological half-life of the
mAb. A number of anti-SARS-CoV-2 mAbs have been engineered for increased
half-lives of approximately 6 months. The duration of follow-up for participants
should be appropriate for the investigational product to provide information on
its long-term efficacy and safety, and should be discussed with the NRA.
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Annex 2
Participants in clinical trials should be representative of the population

targeted for eventual product use. This population should include individuals who
are unlikely to mount an adequate immune response to COVID‐19 vaccination
secondary to immunocompromised status, elderly people and/or subjects with
comorbidities such as:
■ obesity
■ cardiovascular disease, including hypertension
■ chronic lung disease, including asthma
■ type 1 or type 2 diabetes mellitus
■ chronic kidney disease, including those on dialysis
■ chronic liver disease.
All of these have been identified as groups at high risk of severe
COVID-19 and death (47). However, the immunocompromised population is also
quite heterogeneous and the risk of progression to severe disease, even in those
adequately vaccinated, can vary considerably between the different pathologies.
The COVAXID cohort study reported the 1-year follow-up immune
response of 356 subjects after COVID-19 messenger RNA vaccination in a real-
world setting. Subjects who had undergone solid organ transplant and who
had been treated with mycophenolate mofetil, those with common variable
immunodeficiency, with chronic lymphocytic leukaemia treated with ibrutinib, or
with X-linked agammaglobulinemia exhibited lower vaccine responses compared
to other groups of immunocompromised patients (48). Those with a higher risk
of disease progression even after vaccination thus require alternative therapies.
In this regard, several randomized trials and real-world studies have investigated
the role of mAbs in reducing hospitalization and preventing progression from
asymptomatic to symptomatic disease, and even death (49).
The complications that result from the inclusion of immunocompromised
individuals in clinical studies include ethical concerns, for example with regard to
the comparator used (that is, whether a placebo or active comparator is used). The
extrapolation of efficacy data in low-risk patients, based on neutralizing antibody
titres, may be reasonable, and should be discussed early in clinical development
with the NRA and ethics committee. In addition, in immunocompromised
patients, the virus can remain viable for a longer period of time which prolongs
the duration of potential spreading. Studies have shown that mAbs can reduce
the time needed to clear the replicating virus, which not only benefits infected
immunocompromised individuals through the avoidance of longer isolation
periods (50) but also reduces the risk of virus transmission to others.
Vaccines and mAb therapies are complementary approaches to
prophylaxis and treatment of COVID-19. However, due to the specificity of
63
immune support provided by mAbs, certain variants of SARS-CoV-2 can evade

the response. One related issue is that the early mAbs are no longer effective
against more recently circulating VOC, reinforcing the belief that new mAbs with
conserved efficacy across different VOC are needed. Therefore, the decision to
administer mAbs should be based on factors such as the regional prevalence of
resistant variants, and on individual patient health status (51, 52).
The epidemiological situation, including circulating variants, should
be monitored and noted in the study report as any change in circulating virus
variants may have a significant impact on the clinical efficacy of the mAb product.
The viral strain of infected patients should thus also be determined and recorded
during the clinical study. In addition, continued monitoring of emerging viral
variants and of the neutralization activity of the mAb against them is vital.
Furthermore, the risk of development of viruses resistant to the
investigational mAb should be evaluated in all clinical breakthrough cases, using
phenotypic, genotypic and cross-resistance analysis.
7.1 Inclusion and exclusion criteria

For considerations specific to the inclusion or exclusion of pregnant or
breastfeeding women see section 7.4.4 below.
7.1.1 Prophylaxis
Inclusion criteria
■ Participants who can benefit from passive immunization with

antibodies.
■ Medically stable participants.
■ Result from SARS-CoV-2 serology and RT-PCR testing at screening.
■ Able to understand and comply with study requirements/procedures
based on the assessment of the investigator.
Exclusion criteria
■ Significant infection or other acute illness, including fever > 37.8 °C

on the day prior to or day of randomization.
■ Known history of allergy or reaction to any component of the study
drug formulation.
■ Previous known hypersensitivity, infusion-related reaction or severe
adverse reaction following administration of a mAb.
■ Bleeding disorder or prior history of significant bleeding or bruising
following intramuscular injections or venepuncture.
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Annex 2
■ Any other significant disease, disorder or finding that may

significantly increase the risk to the participant because of
participation in the study, affect the ability of the participant to
participate in the study or impair interpretation of the study data.
■ Receipt of any investigational medicinal product (IMP) in a set
period (as defined by the sponsor) immediately prior to the study, or
expected receipt of an IMP during the period of study follow-up, or
concurrent participation in another interventional study.
7.1.2 Treatment
Inclusion criteria
■ Participant has a documented laboratory-confirmed SARS-CoV-2

infection.
■ WHO Clinical Progression Scale score > 1 and < 4 (53).
■ Participant must be dosed with the IMP within a set period (as
defined by the sponsor) following self-reported onset of COVID-19-
related symptoms (mild to moderate COVID-19).
■ One or more of the signs/symptoms relevant to COVID-19 infection
(for example, cough, sore throat, shortness of breath or difficulty
breathing at rest or with activity, body pain or muscle pain/aches,
fatigue, headache, chills, nasal obstruction or congestion, nasal
discharge, nausea or vomiting, diarrhea, new loss of taste or smell).
■ Oxygenation saturation of ≥ 92% obtained at rest by study staff
within 24 hours prior to Day 1 (unless participant regularly receives
chronic supplementary oxygen for an underlying lung condition).
Exclusion criteria
■ Current hospitalization for severe COVID-19, requiring oxygen

therapy or mechanical ventilation.
■ Previous known hypersensitivity, infusion-related reaction or
adverse reaction to any mAb, or known allergy to components of the
IMP or placebo.
■ Current requirement or anticipated impending need for mechanical
ventilation.
■ Any significant disease, disorder or finding that may increase risk
to the participant and that might affect their ability to participate in
the study.
65
■ Participant must not participate in another clinical trial for the

treatment of COVID-19 or SARS-CoV-2 during the study period
until reaching hospitalization or 28 days after entry into the study
(whichever is earliest).
■ Receipt of systemic steroids or inhaled steroids prior to study entry,
unless a stable dose is being used for a chronic condition.
■ Receipt of any other IMP or expected receipt of an IMP during
the study follow-up period, or concurrent participation in another
interventional study.
7.2 Phase I studies

Phase I and first-in-human studies are conducted to determine the initial safety
and tolerability of the IMP following completion of the essential nonclinical
studies. Clinical experience has demonstrated that most COVID-19 mAb
products are, in general, well tolerated.
The determination of starting dose, dose escalation steps and maximum
exposure for first-in-human studies should take into consideration all available
nonclinical information (for example, PD, PK, toxicokinetics and toxicological
profiles, and dose or exposure/effect relationships) as well as safety and toxicity
information derived from testing in a relevant animal model during nonclinical
evaluation. For additional information on animal models of SARS-CoV-2
infection see section 6.2 above.
Products with the same antibody scaffolding and manufacturing process
used for previously authorized anti-SARS-CoV-2 mAbs (that is, a product
that only differs from the authorized product in the epitope binding site) may
leverage the clinical development of the authorized product to expedite certain
aspects of their clinical development. However, this should be discussed with the
NRA, particularly if the mechanism of action has changed. Phase I trials may
be conducted in healthy volunteers to determine the mAb safety profile, PK and

potential physiological responses. If the product is intended to be administered
in the elderly, in children or in other specific groups, then safety and tolerability
data may be required for those specific groups.
7.3 Clinical pharmacology

7.3.1 Pharmacokinetics
Multiple-dose PK studies may not be required if the mAb is intended to be
given only in a single dose. However, if the product is intended to be repeatedly
administered, safety and tolerability data may be required to support the dosing
regimen.
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Annex 2
7.3.2 Pharmacodynamics
The PK, combined with nonclinical PD target levels, should guide the doses to
be evaluated. Such studies may involve the ex vivo assessment of the neutralizing
activity of the mAb in serum collected at different timepoints following
administration.
7.4 Phase II and III studies

7.4.1 Efficacy
The clinical trial design of Phase II and III studies for efficacy determination
will depend on whether the mAb is intended to be a prophylactic or therapeutic
product.
The efficacy of a prophylactic mAb should be evaluated in terms of its
ability to prevent the disease or progression to severe disease, but may also be
assessed in terms of its ability to eliminate the pathogen, reduce the viral load or
reduce virus shedding.
The efficacy of a therapeutic mAb should be evaluated in terms of its
ability to prevent disease progression (that is, prevent deterioration in overall
clinical status, hospitalization or death) and/or reduce clinically relevant end-
points, such as time to sustained alleviation of symptoms, following confirmation
of infection. The efficacy of a therapeutic mAb may also include the ability to
eliminate the pathogen, reduce the viral load or reduce virus shedding.
An emphasis should be placed on designing randomized controlled
trials that take into account the intended target population, the selected clinical
end-point(s) (Table 2) and case definitions (53).
The local epidemiology of circulating variants may also affect efficacy
outcomes, particularly if the mAb has different binding affinities to such variants.
For this reason, in vitro neutralizing assays against any identified new variant
should be conducted to ensure that the mAb retains activity against this new
variant and that the study can safely be continued.
The selection of an appropriate authorized product as a comparator for use
in efficacy trials will also require careful consideration and may vary depending
on intended use – that is, for prevention or treatment. A randomized controlled
double-blind trial design should be used in efficacy studies intended to prevent or
treat infections. A placebo control may be considered when there is no appropriate
comparator, no known therapeutic agent is effective, or when the natural history of
the untreated infectious disease is relatively benign or self-limiting (that is, of low
risk to patients) and where switching to an approved treatment is ensured in case
of progression to severe disease. Any other current standard of care practices for
the prevention or treatment of the infection must be provided to all participants
regardless of the treatment arm. It is recommended that in all cases, these issues
are discussed in advance with the NRA and ethics committee.
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7.4.2 Immunobridging
To accelerate initial approval of novel mAbs manufactured on the same platform
technology as already approved mAbs, an immunobridging approach could be an
acceptable pathway for mAbs intended for prevention (54). The immunobridging
should be based on a cross-variant comparison in a non-inferiority study with an
approved mAb with the same indication. The geometric mean titres (GMTs) of
neutralizing antibodies at Day 28 of an already approved mAb product against a
virus strain for which efficacy was shown (for example, Alpha) should be compared
to the GMTs achieved at the same timepoint with the new investigational mAb
product against circulating variants. The acceptable non-inferiority margin
when using a comparison of GMT values should be discussed with the NRA.
Following initial approval, post-marketing efficacy data (including data from the
investigation of breakthrough cases) should be collected, neutralizing antibody
concentrations monitored to determine the timing of antibody waning, and long-
term efficacy and safety data generated for at least 6 months.
Deciding upon the acceptability of an immunobridging approach,
particularly for bispecific mAbs and mAbs with a different mechanism of action,
will be in the remit of the NRA.
Presently, there are not sufficient data to derive a specific mAb
concentration or neutralizing threshold to derive a correlate of protection for
SARS-CoV-2.
7.4.3 Safety
The continual evaluation of mAb product safety is an important component
within all phases of clinical studies. Although mAbs generally have a very good
safety profile, each product is unique and should be considered independently.
Safety data should be obtained from a sufficient number of subjects
during the clinical trials to characterize and quantify the product safety profile,
which can include the type, frequency and severity of adverse drug reactions.
In some cases, it may be possible to consider safety data from multiple clinical
studies if both the products tested and the study conditions are sufficiently similar.
Evaluating the safety and tolerability of anti-SARS-CoV-2 mAbs should
include the recording of all adverse events (AEs), serious adverse events (SAEs),
medically attended adverse events (MAAEs) and adverse events of special interest
(AESIs) over the duration of the study (Table 2).
Product reactogenicity should also be clearly characterized by monitoring
immune responses to the mAb through ADA titres and immune system activity.
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Table 2
Objectives, estimands and clinical end-points
Objectives Estimand description/end-point

Primary
Estimate the efficacy of the mAb An appropriate time frame for the assessment
of efficacy should be provided to the NRA based
on the end-point being assessed (for example,
6 months for prophylaxis, Day 28 for treatment)
For pre-exposure prophylaxis, a binary
response whereby a participant is defined as a
COVID-19 case if a SARS-CoV-2 RT PCR-positivea
symptomatic illness occurs post dose(s) of the
mAb and prior to the specified time frame.
For post-exposure prophylaxis, a composite
outcome of either hospitalization or progression
of symptoms post dose(s) of the mAb during the
specified time frame.
For treatment, a composite outcome of medical
attendance visits, hospitalization or death from
any cause and/or time to sustained resolution of
symptoms post dose(s) of the mAb during the
specified time frame.b
Estimate the safety and AEs, SAEs, MAAEs and AESIs during the study
tolerability of the mAb period
Secondary
Estimate the efficacy of the mAb Incidence of SARS-CoV-2 RT-PCR-positive severe
in preventing severe or critical or critical symptomatic illness occurring after
symptomatic COVID-19 dosing with the mAb
Estimate the efficacy of the mAb The incidence of COVID-19-related emergency
in preventing COVID-19-related department visits occurring after dosing with
emergency department visits the mAb
Assess the PK of the mAb Serum concentrations
following administration of
an appropriate dose via an
appropriate route
Evaluate ADA response to the Incidence of ADA to the mAb in serum
mAb in serum
69
Table 2 continued
Objectives Estimand description/end-point
Exploratory
Estimate the efficacy of the mAb An appropriate time frame for assessment of
over a longer time frame efficacy for pre-exposure prophylaxis should be
provided to the NRA based on the end-point
being assessed. (for example, 12 months for
prophylaxis)
A binary response whereby a participant is
defined as a COVID-19 case if a SARS-CoV-2
RT-PCR-positive symptomatic illness occurs post
dose(s) of mAb and prior to the specified time
frame.
Determine anti-SARS-CoV-2 mAb Post-treatment GMT and geometric mean fold
levels in serum following the rise from baseline value through an extended
administration of the mAb time frame
Quantify SARS-CoV-2 viral loads in Viral genome copies in nasopharyngeal swabs
infected participants treated with at illness visits as determined by quantitative
the mAb RT-PCR
Quantify the duration of viral Duration of SARS-CoV-2 shedding in saliva
shedding in participants with
symptomatic COVID-19 treated
with the mAb
Characterize the risk of Genotypic analysis and biochemical and/or
development of resistance to the susceptibility analysis of SARS-CoV-2 variants
mAb in patients with virological
failure
Assess additional immune Other exploratory assays for humoral, mucosal

responses following and cellular immune responses may be
administration of the mAb performed based upon emerging safety, efficacy
and PD data.
Estimate the efficacy of the mAb The incidence of long COVID occurring after
in preventing long COVID dosing with the mAb
a
RT-PCR-positive = reverse transcription-polymerase chain reaction-positive.
b
As the rate of progression to severe COVID-19 has significantly decreased due to the increased levels
of immunization and seropositivity in the population, as well as the lower progression rates seen with
Omicron variants, the primary efficacy end-point of progression to severe disease or death may no
longer be appropriate. The use of an alternative end-point (such as sustained resolution of symptoms
or non-progression of the clinical status) should be considered. In all cases, consultation with the NRA is
recommended during trial design and end-point selection.
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Annex 2
7.4.4 During pregnancy and breastfeeding

Some studies have shown that COVID-19 infection during pregnancy was
associated with a greater probability of maternal, fetal and neonatal complications,
including pre eclampsia, increased risk of admission to an intensive care unit
for the mother, preterm birth and neonatal mortality compared to non-infected
pregnant women (55, 56). However, many studies related to SARS‐CoV‐2
infection in pregnancy were performed among hospitalized patients, which
may have led to overestimation of the risk of severe outcomes as not all cases
of SARS-CoV-2 infection in the pregnant population were included (57, 58).
Moreover, pregnant women who were obese and women with comorbidities were
more likely to develop severe disease or to present greater risk of complications
related to COVID-19 than pregnant women without such conditions (57, 59, 60).
The extensive physiological changes associated with pregnancy may
alter drug PK and PD, thus directly affecting the safety and efficacy of any
drug administered during pregnancy through alterations in drug absorption,
distribution, metabolism and excretion (61).
Currently, information on medicinal drug use in pregnancy and
breastfeeding generally is collected in the post-marketing setting, using data from
observational studies such as pregnancy exposure registries and other cohort
studies, case control studies and surveillance methods. However, this approach
commonly results in delayed access to new medicinal products for pregnant and
breastfeeding women. There are multiple reasons for considering the inclusion of
pregnant women in clinical trials, including:
■ Women need safe and effective treatment during pregnancy.
■ Failure to establish the dose/dosing regimen, safety and efficacy of
treatments during pregnancy may compromise the health of women
and/or the fetus.
■ In some settings, the enrolment of pregnant women in clinical trials
may offer the possibility of direct benefits to such women and/or
their fetus that are unavailable outside the research setting.
■ The development of accessible treatment options for pregnant
women is a significant public health issue.
Systematic consideration should be given to the possible use of any
new medicine by pregnant and breastfeeding women and, where warranted, to
planning formal investigations in these populations. Such planning should take
into account different variables, such as the benefit and risk perspective, as well
as the need for systematic and timely study of medicines likely to be used in this
population to support dosing, use rationale and other aspects (62). Data obtained
to date, mainly from clinical settings, suggest that COVID-19 mAb products seem
to be well tolerated and likely to be safe when used during pregnancy (63–68).
71
Sponsors should consult with the NRA early in the product development
phase on the requirements for specific nonclinical studies, and on the potential
inclusion of pregnant women in clinical studies to promote the health of pregnant
women and their fetus, and to inform prescribing decisions during pregnancy.
Proper follow-up of the mother-child pair should also be considered to fully
determine the impact of product administration on maternal and newborn
health (63–68).
7.4.5 Human challenge studies

Human challenge studies of SARS-CoV-2 infection have been conducted to
better understand COVID-19, especially during the early stages of infection, and
for the potential evaluation of candidate vaccines, antiviral drugs and antibodies
(69, 70). The first reported study (70) paid careful attention to the preparation of
the challenge stock and used highly characterized virus, including whole genome
sequencing to confirm that the challenge virus was unaltered compared to the
original isolate. Although not technically a clinical trial (no product was being
investigated), regulatory oversight of the challenge strain was provided by the
United Kingdom Medicines and Healthcare products Regulatory Authority,
which confirmed that its manufacture would be suitable for use in future efficacy
studies of an IMP. However, it may not be possible to undertake such studies in
some jurisdictions and the relevant NRA should always be consulted directly.
Such studies have not yet been used to assess the efficacy of mAbs but may be
used in the future.
7.4.6 Paediatric considerations

In children, severe COVID-19 is uncommon. However, those with certain
underlying conditions (such as cardiovascular, respiratory, neuromuscular or
malignant disease, or immunocompromised individuals) are prone to unfavourable
outcomes. Therefore, while most children infected with SARS-CoV-2 will recover
without therapy, treatment of mild or moderate infection should be considered in
paediatric patients at highest risk of progression to severe disease. This would be
in alignment with the current indication for the use of mAbs against SARS-CoV-2
in adults.
None of the currently licensed mAbs against SARS-CoV-2 are authorized
for use in children under 12 years of age. In addition, even for mAbs against
SARS-CoV-2 that have already been commercialized, safety and efficacy data
in paediatric patients are limited. Furthermore, the additional data available
from observational studies are associated with limitations. With regard to post-
authorization data, it is important to highlight that the generation of data in
children has been greatly hampered by the loss of effectiveness of early mAbs
against recently circulating VOC. Overall, the data generated so far do not suggest
72
Annex 2
an excessive risk of toxicity in children compared with adults, and mAbs seem to
be well tolerated. However, the lack of a comparator group in studies makes clear
estimation of the effectiveness of mAbs in preventing COVID-19 progression in
children difficult. Therefore, further studies are needed to fully define the safety
and efficacy of mAb therapy in the paediatric population (71–74).
The inclusion of children and adolescents in clinical trials should always
be considered when planning a study to avoid knowledge gaps and to facilitate
early access to new medicinal products (73–77). Sponsors are encouraged to
discuss paediatric drug development with the NRA early in clinical development,
including: (a) the potential for extrapolating efficacy data from studies in adults;
(b) appropriate PK trials in paediatric subjects to support dose selection; (c) the
recommended size of the pre-approval safety database in children; and (d) the
targeted age group(s) (78–80).
7.4.7 Post-authorization studies

The potential risk of treatment failure due to the development of SARS-CoV-2
variants resistant to the mAb, along with the potential risks associated with
any biological therapy (including mAbs) should continue to be assessed post-
authorization.
Data monitoring (including systematic and proactive review of the
emerging data) should be conducted using all available data sources, for example
by evaluating:
■ new and cumulative nonclinical data (antiviral activity and viral
resistance);
■ data on variants detected in clinical studies among patients who
received mAbs;
■ spontaneous reports related to lack of efficacy, including
information for variant lineages; and
■ literature reports or studies conducted by public health authorities.
The requirements for a risk-management plan, Phase IV studies and/or
use of real-world evidence and data should be discussed with the NRA.
Authors and acknowledgements

The first draft of this WHO addendum was prepared by Dr A. Chia (lead author
for the Nonclinical evaluation section), Health Sciences Authority, Singapore; Dr
E. Griffiths (lead author for the General considerations section), consultant, United
Kingdom; Dr R. Isbrucker (lead author for the Introduction, and Purpose and scope
sections) and Dr J. Lacroix (lead author for the Clinical evaluation section), Health
73
Canada, Canada. The draft document was then reviewed and revised by a drafting
group comprising Dr A. Chia, Dr E. Griffiths, Dr R. Isbrucker, Dr J. Lacroix,
and Dr S. Buchholz and Dr M. Gonzalez-Tome, European Medicines Agency,
Netherlands (Kingdom of the); and Dr B. Klug, Paul-Ehrlich-Institut, Germany;
and by Dr I. Knezevic and Dr E.K. Kim, World Health Organization, Switzerland.
The resulting draft document was posted on the WHO Biologicals website
from 1 November to 4 December 2023 for a first round of public consultation.
Comments were received from Dr S. Hufton and Dr G. Mattiuzzo, Medicines and
Healthcare products Regulatory Agency, United Kingdom; Dr J. Wang, National
Institutes for Food and Drug Control, China; Dr S. Tognarelli, Paul-Ehrlich-
Institut, Germany; Dr T. Cohen, AstraZeneca, USA; the Nonclinical working
party, 3Rs working party, and Pregnancy group, European Medicines Agency,
Netherlands (Kingdom of the); Dr J. Holst, Holst PharmaWorks, Norway; and Dr
R. Gupta, Vir Biotechnology, USA.
All comments received were collated and distributed to the drafting
group members for their consideration, and revisions to the text made
accordingly. The revised document (WHO/BS/2024.2466) was then posted on
the WHO Biologicals website from 11 January to 15 February 2024 for a second
round of public consultation. Comments were received from Dr S. Fakhrzadeh,
Consultant, Iran (Islamic Republic of); Dr I. Feavers, United Kingdom; Bharat
Biotech International Limited, India; Dr S. Silviera, Brazilian Health Regulatory
Agency, Brazil; Dr J. Southern, South African Health Products Regulatory
Authority, South Africa; European Medicines Agency, Netherlands (Kingdom of
the); and Dr T. Cohen, AstraZeneca, USA. All comments received were taken
into consideration and an updated document prepared.
Editorial review of the resulting document was then completed by Dr
T. Waddell, United Kingdom in accordance with WHO requirements for all
documents appearing in the WHO Technical Report Series.
Further changes were made to document WHO/BS/2024.2466 by the
WHO Expert Committee on Biological Standardization.
References
1. Guidelines on the nonclinical and clinical evaluation of monoclonal antibodies and related products
intended for the prevention or treatment of infectious diseases. In: WHO Expert Committee on
Biological Standardization: seventy-seventh report. Geneva: World Health Organization; 2023:
Annex 2 (WHO Technical Report Series, No. 1048; https://iris.who.int/handle/10665/373128,
accessed 20 December 2023).
2. Guidelines for the production and quality control of monoclonal antibodies and related products
intended for medicinal use. In: WHO Expert Committee on Biological Standardization: seventy-
fifth report. Geneva: World Health Organization; 2022: Annex 4 (WHO Technical Report Series, No.
1043; https://iris.who.int/handle/10665/362194, accessed 20 December 2023).
74
Annex 2
3. The COVID-19 pandemic in 2023: far from over. Editorial. Lancet. 2023;401(10371):79 (https://
www.thelancet.com/journals/lancet/article/PIIS0140-6736(23)00050-8/fulltext, accessed 20
December 2023).
4. Davis HE, McCorkell L, Vogel JM, Topol EJ. Long COVID: major findings, mechanisms and
recommendations. Nat Rev Microbiol. 2023;21:133–46 (https://www.nature.com/articles/s41579-
022-00846-2, accessed 20 December 2023).
5. COVID-19 Weekly Epidemiological Update. Edition 152. 20 July 2023. Geneva: World
Health Organization; 2023 (https://www.who.int/docs/default-source/coronaviruse/situation-
reports/20230720_weekly_epi_update_152.pdf, accessed 20 December 2023).
6. Sánchez FJM, Martínez-Sellés M, García JMM, Guillén SM, Rodríguez-Artalejo F, Ruiz-Galiana J et al.
Insights for COVID-19 in 2023. Rev Esp Quimioter. 2023;36(2):114–24 (https://seq.es/wp-content/
uploads/2022/12/martin13dec2022.pdf, accessed 20 December 2023).
7. Al-Aly Z, Agarwal A, Alwan N, Luyckx VA. Long COVID: long-term health outcomes and implications
for policy and research. Nat Rev Nephrol. 2023;19(1):1–2 (https://www.nature.com/articles/
s41581-022-00652-2, accessed 20 December 2023).
8. Shaman J, Galanti M. Will SARS-CoV-2 become endemic? Science. 2020;370(6516):527–9 (https://
www.science.org/doi/10.1126/science.abe5960?url_ver=Z39.88-2003&rfr_id=ori:rid:crossref.
org&rfr_dat=cr_pub%20%200pubmed, accessed 20 December 2023).
9. Cohen C, Pulliam J. COVID-19 infection, reinfection, and the transition to endemicity. Lancet.
2023;401(10379):798–800 (https://www.thelancet.com/journals/lancet/article/PIIS0140-
6736(22)02634-4/fulltext, accessed 20 December 2023).
10. Ling Z, Yi C, Sun X, Yang Z, Sun B. Broad strategies for neutralizing SARS-CoV-2 and other human
coronaviruses with monoclonal antibodies. Sci China Life Sci. 2023;66(4):658–78 (https://link.
springer.com/article/10.1007/s11427-022-2215-6, accessed 20 December 2023).
11. Focosi D, McConnell S, Casadevall A, Cappello E, Valdiserra G, Tuccori M. Monoclonal antibody
therapies against SARS-CoV-2. Lancet Infect Dis. 2022;22(11):e311–26 (https://www.thelancet.
com/journals/laninf/article/PIIS1473-3099(22)00311-5/fulltext, accessed 20 December 2023).
12. Jiang S, Hillyer C, Du L. Neutralizing antibodies against SARS-CoV-2 and other human coronaviruses.
Trends Immunol. 2020;41(5):355–9 (https://www.cell.com/trends/immunology/fulltext/S1471-
4906(20)30057-0?_returnURL=https%3A%2F%2Flinkinghub.elsevier.com%2Fretrieve%2Fpii%2
FS1471490620300570%3Fshowall%3Dtrue, accessed 20 December 2023). Erratum (see: https://
www.ncbi.nlm.nih.gov/pmc/articles/PMC7271084/, accessed 20 December 2023).
13. Knezevic I, Mattiuzzo G, Page M, Minor P, Griffiths E, Nuebling M et al. WHO International Standard
for evaluation of the antibody response to COVID-19 vaccines: call for urgent action by the
scientific community. Lancet Microbe. 2022;3(3):e235–40 (https://www.thelancet.com/journals/
lanmic/article/PIIS2666-5247(21)00266-4/fulltext, accessed 20 December 2023).
14. Wang Q, Guo Y, Iketani S, Nair MS, Li Z, Mohri H et al. Antibody evasion by SARS-CoV-2 Omicron
subvariants BA.2.12.1, BA.4 and BA.5. Nature. 2022;608(7923):603–8 (https://www.nature.com/
articles/s41586-022-05053-w, accessed 20 December 2023).
15. Gupta A, Gonzalez-Rojas Y, Juarez E, Crespo Casal M, Moya J, Falci DR et al. Early treatment for
COVID-19 with SARS-CoV-2 neutralizing antibody sotrovimab. N Engl J Med. 2021;385(21):1941–
50 (https://www.nejm.org/doi/full/10.1056/NEJMoa2107934, accessed 20 December 2023).
16. Da Silva MA, Nioche P, Soudaramourty C, Bull-Maurer A, Tiouajni M, Kong D et al. Repetitive mRNA
vaccination is required to improve the quality of broad-spectrum anti-SARS-CoV-2 antibodies in
the absence of CXCL13. Sci Adv. 2023;9(31):eadg2122 (https://www.science.org/doi/10.1126/
sciadv.adg2122, accessed 20 December 2023).
75
17. DeGrace MM, Ghedin E, Frieman MB, Krammer F, Grifoni A , Alisoltani A et al. Defining the risk
of SARS-CoV-2 variants on immune protection. Nature. 2022;605(7911):640–52 (https://www.
nature.com/articles/s41586-022-04690-5, accessed 20 December 2023).
18. Pulliam JRC, van Schalkwyk C, Govender N, von Gottberg A, Cohen C, Groome MJ et al. Increased
risk of SARS-CoV-2 reinfection associated with emergence of Omicron in South Africa. Science.
2022;376(6593):eabn4947 (https://www.science.org/doi/10.1126/science.abn4947, accessed 20
December 2023).
19. van Dorp L, Acman M, Richard D, Shaw LP, Ford CE, Ormond L et al. Emergence of genomic
diversity and recurrent mutations in SARS-CoV-2. Infect Genet Evol. 2020;83:104351 (https://
www.sciencedirect.com/science/article/pii/S1567134820301829, accessed 20 December 2023).
20. Wang Z, Schmidt F, Weisblum Y, Muecksch F, Barnes CO, Finkin S et al. mRNA vaccine-elicited
antibodies to SARS-CoV-2 and circulating variants. Nature. 2021;592(7855):616–22 (https://www.
nature.com/articles/s41586-021-03324-6, accessed 20 December 2023).
21. Dormeshkin D, Shapira M, Dubovik S, Kavaleuski A, Katsin M, Migas A et al. Isolation of an escape-
resistant SARS-COV-2 neutralizing nanobody from a novel synthetic nanobody library. Front
Immunol. 2022:13:965446 (https://www.frontiersin.org/articles/10.3389/fimmu.2022.965446/
full, accessed 23 December 2023).
22. Watanabe Y, Allen JD, Wrapp D, McLellan JS, Crispin M. Site-specific glycan analysis of the SARS-
CoV-2 spike. Science. 2020;369:330–3 (https://www.science.org/doi/10.1126/science.abb9983,
23. Coutard B, Valle C, de Lamballerie X, Canard B, Seidah NG, Decroly E. The spike glycoprotein of
the new coronavirus 2019-nCoV contains a furin-like cleavage site absent in CoV of the same
clade. Antiviral Res. 2020;176:104742 (https://www.sciencedirect.com/science/article/pii/
S0166354220300528, accessed 23 December 2023).
24. Wang Q, Bowen A, Valdez R, Gherasim C, Gordon A, Liu L et al. Antibody response to Omicron
BA.4–BA.5 bivalent booster. New Engl J Med. 2023;388(6):567–9 (https://www.nejm.org/doi/
full/10.1056/NEJMc2213907, accessed 23 December 2023).
25. Weisblum Y, Schmidt F, Zhang F, DaSilva J, Poston D, Lorenzi JCC et al. Escape from neutralizing
antibodies by SARS-CoV-2 spike protein variants. ELife. 2020;9:e61312 (https://elifesciences.org/
articles/61312, accessed 23 December 2023).
26. Xie X, Zou J, Fontes-Garfias CR, Xia H, Swanson KA, Cutler M et al. Neutralization of N501Y mutant
SARS-CoV-2 by BNT162b2 vaccine-elicited sera. bioRxiv. 2021 (https://www.biorxiv.org/content/1
0.1101/2021.01.07.425740v1, accessed 23 December 2023).

27. Funnell SGP, Afrough B, Baczenas JJ, Berry N, Bewley KR, Bradford R et al. A cautionary perspective
regarding the isolation and serial propagation of SARS-CoV-2 in Vero cells. NPJ Vaccines.
2021;6(1):83 (https://www.nature.com/articles/s41541-021-00346-z, accessed 23 December
2023).
28. Arvin AM, Fink K, Schmid MA, Cathcart A, Spreafico R, Havenar C et al. A perspective on potential
antibody-dependent enhancement of SARS-CoV-2. Nature. 2020;584(7821):353–63 (https://
www.nature.com/articles/s41586-020-2538-8, accessed 23 December 2023).
29. Ricke DO. Two different antibody-dependent enhancement (ADE) risks for SARS-CoV-2
antibodies. Front Immunol. 2021;12:640093 (https://www.frontiersin.org/articles/10.3389/
fimmu.2021.640093/full, accessed 23 December 2023).
76
Annex 2
30. Second WHO International Standard for anti-SARS-CoV-2 immunoglobulin. In: WHO Expert
Committee on Biological Standardization: seventy-sixth report. Geneva: World Health
Organization; 2023 (WHO Technical Report Series, No. 1045, section 8.1.2, pp. 43–46; https://iris.
who.int/handle/10665/368140, accessed 23 December 2023).
31. First WHO International Standard for antibodies to SARS-CoV-2 variants of concern. In: WHO
Expert Committee on Biological Standardization: seventy-sixth report. Geneva: World Health
32. First WHO International Reference Panel for antibodies to SARS-CoV-2 variants of concern. In: WHO
Expert Committee on Biological Standardization: seventy-sixth report. Geneva: World Health
33. Expansion of the First WHO International Reference Panel for antibodies to SARS-CoV-2 variants
of concern. In: WHO Expert Committee on Biological Standardization: seventy-seventh report.
Geneva: World Health Organization; 2023 (WHO Technical Report Series, No. 1048, section 8.1.1,
pp. 30–31; https://iris.who.int/handle/10665/373128, accessed 23 December 2023).
34. Mouna L, Razazian M, Duquesne S, Roque-Afonso AM, Vauloup-Fellous C. Validation of a SARS-
CoV-2 surrogate virus neutralization test in recovered and vaccinated healthcare workers. Viruses.
2023;15(2):426 (https://www.mdpi.com/1999-4915/15/2/426, accessed 24 December 2023).
35. Wisnewski AV, Liu J, Lucas C, Klein J, Iwasaki A, Cantley L et al. Development and utilization of
a surrogate SARS-CoV-2 viral neutralization assay to assess mRNA vaccine responses. PLoS
One. 2022;17(1):e0262657 (https://journals.plos.org/plosone/article?id=10.1371/journal.
pone.0262657, accessed 24 December 2023).
36. Sánchez-Zuno GA, Matuz-Flores MG, González-Estevez G, Nicoletti F, Turrubiates-Hernández FJ,
Mangano K et al. A review: antibody-dependent enhancement in COVID-19: the not so friendly
side of antibodies. Int Journal of Immunopathol Pharmacol. 2021;35 (https://journals.sagepub.
com/doi/full/10.1177/20587384211050199, accessed 24 December 2023).
37. Okuya K, Hattori T, Saito T, Takadate Y, Sasaki M, Furuyama W et al. Multiple routes of antibody-
dependent enhancement of SARS-CoV-2 infection. Microbiol Spectr. 2022;10(2):e0155321
(https://journals.asm.org/doi/full/10.1128/spectrum.01553-21, accessed 24 December 2023).
38. Cox M, Peacock TP, Harvey WT, Hughes J, Wright DW; COVID-19 Genomics UK (COG-UK) Consortium.
SARS-CoV-2 variant evasion of monoclonal antibodies based on in vitro studies. Nat Rev Microbiol.
2023;21(2):112–4 (https://www.nature.com/articles/s41579-022-00809-7, accessed 24 December
2023).
39. Muñoz-Fontela C, Dowling WE, Funnell SGP, Gsell PS, Riveros-Balta AX, Albrecht RA et al. Animal
models for COVID-19. Nature. 2020;586(7830):509–15 (https://www.nature.com/articles/s41586-
020-2787-6, accessed 24 December 2023).
40. Muñoz-Fontela C, Widerspick L, Albrecht RA, Beer M, Carroll MW, de Wit E et al. Advances and
gaps in SARS-CoV-2 infection models. PLoS Pathog. 2022;18(1)e1010161 (https://journals.plos.
org/plospathogens/article?id=10.1371/journal.ppat.1010161, accessed 24 December 2023).
41. Fan C, Wu Y, Rui X, Yang Y, Ling C, Liu S et al. Animal models for COVID-19: advances, gaps and
perspectives. Signal Transduct Target Ther. 2022;7(1):220 (https://www.nature.com/articles/
s41392-022-01087-8, accessed 24 December 2023).
42. Kadowaki T, Imajou S, Matsumoto N, Takao S, Yorifuji T. Timing of REGEN-COV administration and
progression to severe COVID-19. J Infect Chemother. 2022;28(11):1459–63 (https://www.jiac-j.
com/article/S1341-321X(22)00198-2/fulltext, accessed 24 December 2023).
77
43. Bruzzesi E, Ranzenigo M, Castagna A, Spagnuolo V. Neutralizing monoclonal antibodies for the
treatment and prophylaxis of SARS-CoV-2 infection. New Microbiol. 2021;44(3):135–44 (https://
www.newmicrobiologica.org/PUB/allegati_pdf/2021/3/135.pdf, accessed 24 December 2023).
44. Brobst B, Borger J. Benefits and risks of administering monoclonal antibody therapy for coronavirus
(COVID-19). In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2023 Jan; update 7
May 2023 (https://www.ncbi.nlm.nih.gov/books/NBK574507/, accessed 24 December 2023).
45. ACTIV-3–Therapeutics for Inpatients with COVID-19 (TICO) Study Group. Tixagevimab-cilgavimab
for treatment of patients hospitalised with COVID-19: a randomised, double-blind, phase 3 trial.
Lancet Respir Med. 2022;10(10):972–84 (https://www.thelancet.com/journals/lanres/article/
PIIS2213-2600(22)00215-6/fulltext, accessed 24 December 2023).
46. Hurt AC, Wheatley AK. Neutralizing antibody therapeutics for COVID-19. Viruses. 2021;13(4):628
(https://www.mdpi.com/1999-4915/13/4/628, accessed 24 December 2023).
47. Fung M, Babik JM. COVID-19 in immunocompromised hosts: what we know so far. Clin Infect
Dis. 2021;72(2):340–50 (https://academic.oup.com/cid/article/72/2/340/5864040, accessed 24
December 2023).
48. Chen P, Bergman P, Blennow O, Hansson L, Mielke S, Nowak P et al. Real-world assessment of
immunogenicity in immunocompromised individuals following SARS-CoV-2 mRNA vaccination:
a one-year follow-up of the prospective clinical trial COVAXID. EBioMedicine. 2023;94:104700
(https://www.thelancet.com/journals/ebiom/article/PIIS2352-3964(23)00265-7/fulltext,
49. Verderese JP, Stepanova M, Lam B, Racila A, Kolacevski A, Allen D et al. Neutralizing monoclonal
antibody treatment reduces hospitalization for mild and moderate coronavirus disease 2019
(COVID-19): a real-world experience. Clin Infect Dis. 2022;74(6):1063–9 (https://academic.oup.
com/cid/article/74/6/1063/6309036, accessed 24 December 2023).
50. Utzon AN, Johansen IS, Bang LL, Pedersen RM, Andersen TE, Madsen LW. Viral dynamics of SARS-
CoV-2 in immunocompromised patients. Clin Microbiol Infect. 2023;29(8):1087.e1–e3 (https://
www.clinicalmicrobiologyandinfection.com/article/S1198-743X(23)00237-9/fulltext, accessed 24
December 2023).
51. Cowan J, Amson A, Christofides A, Chagla Z. Monoclonal antibodies as COVID-19 prophylaxis
therapy in immunocompromised patient populations. Int J Infect Dis. 2023;134:228–38 (https://
www.ijidonline.com/article/S1201-9712(23)00645-8/fulltext, accessed 24 December 2023).
52. ETF warns that monoclonal antibodies may not be effective against emerging strains of SARS-
CoV-2. Amsterdam: European Medicines Agency; 2022 (https://www.ema.europa.eu/en/news/

etf-warns-monoclonal-antibodies-may-not-be-effective-against-emerging-strains-sars-cov-2,
53. WHO Working Group on the Clinical Characterisation and Management of COVID-19 infection.
A minimal common outcome measure set for COVID-19 clinical research. Lancet Infect
Dis. 2020;20(8):e192–e197 (https://www.thelancet.com/journals/laninf/article/PIIS1473-
3099(20)30483-7/fulltext, accessed 24 December 2023).
54. Summary report of the Joint EMA-FDA workshop on the efficacy of monoclonal antibodies in the
context of rapidly evolving SARS-CoV-2 variants. Amsterdam: European Medicines Agency; 2023
(Document EMA/67738/2023; https://www.ema.europa.eu/en/documents/report/summary-
report-joint-ema-fda-workshop-efficacy-monoclonal-antibodies-context-rapidly-evolving-sars_
en.pdf, accessed 24 December 2023).
78
Annex 2
55. Zambrano LD, Ellington S, Strid P, Galang RR, Oduyebo T, Tong VT et al. Update: characteristics
of symptomatic women of reproductive age with laboratory-confirmed SARS-CoV-2 infection by
pregnancy status – United States, January 22–October 3, 2020. MMWR Morb Mortal Wkly Rep.
2020;69(44):1641–7 (https://www.cdc.gov/mmwr/volumes/69/wr/mm6944e3.htm, accessed 24
December 2023).
56. Villar J, Ariff S, Gunier RB, Thiruvengadam R, Rauch S, Kholin A et al. Maternal and neonatal
morbidity and mortality among pregnant women with and without COVID-19 infection: the
INTERCOVID Multinational Cohort Study. JAMA Pediatr. 2021;175(8):817–26 (https://jamanetwork.
com/journals/jamapediatrics/fullarticle/2779182, accessed 24 December 2023).
57. Vousden N, Bunch K, Morris E, Simpson N, Gale C, O’Brien P et al. The incidence, characteristics and
outcomes of pregnant women hospitalized with symptomatic and asymptomatic SARS-CoV-2
infection in the UK from March to September 2020: a national cohort study using the UK Obstetric
Surveillance System (UKOSS). PLoS One. 2021;16(5):e0251123 (https://journals.plos.org/plosone/
article?id=10.1371/journal.pone.0251123, accessed 24 December 2023).
58. Aabakke AJM, Krebs L, Petersen TG, Kjeldsen FS, Corn G, Wøjdemann K et al. SARS-CoV-2
infection in pregnancy in Denmark – characteristics and outcomes after confirmed infection
in pregnancy: a nationwide, prospective, population-based cohort study. Acta Obstet Gynecol
Scand. 2021;100(11):2097–110 (https://obgyn.onlinelibrary.wiley.com/doi/10.1111/aogs.14252,
59. Allotey J, Stallings E, Bonet M, Yap M, Chatterjee S, Kew T et al. Clinical manifestations, risk factors,
and maternal and perinatal outcomes of coronavirus disease 2019 in pregnancy: living systematic
review and meta-analysis. BMJ. 2020;370:m3320 (https://www.bmj.com/content/370/bmj.
m3320.long, accessed 24 December 2023).
60. De Bruin O, Engjom H, Vousden N, Ramakrishnan R, Aabakke AJM, Äyräs O et al. Variations across
Europe in hospitalization and management of pregnant women with SARS-CoV-2 during the initial
phase of the pandemic: multi-national population-based cohort study using the International
Network of Obstetric Survey Systems (INOSS). Acta Obstet Gynecol Scand. 2023;102(11):1521–30
(https://obgyn.onlinelibrary.wiley.com/doi/10.1111/aogs.14643, accessed 24 December 2023).
61. Pregnant women: scientific and ethical considerations for inclusion in clinical trials. Guidance
for industry. Silver Spring (MD): U.S. Department of Health and Human Services, Food and Drug
Administration; 2018 (https://www.fda.gov/files/drugs/published/Pregnant-Women--Scientific-
and-Ethical-Considerations-for-Inclusion-in-Clinical-Trials.pdf, accessed 24 December 2024).
62. Nooney J, Thor S, de Vries C, Clements J, Sahin L, Hua W et al. Assuring access to safe medicines
in pregnancy and breastfeeding. Clin. Pharmacol. Ther. 2021;110(4):941–5 (https://ascpt.
onlinelibrary.wiley.com/doi/full/10.1002/cpt.2212, accessed 24 December 2024).
63. Crispino P, Marocco R, Di Trento D, Guarisco G, Kertusha B, Carraro A et al. Use of monoclonal
antibodies in pregnant women infected by COVID-19: a case series. Microorganisms.
2023:11(8):1953 (https://www.mdpi.com/2076-2607/11/8/1953, accessed 25 March 2024).
64. Chang MH, Cowman K, Guo Y, Bao H, Bernstein PS, Gendlina I et al. A real-world assessment
of tolerability and treatment outcomes of COVID-19 monoclonal antibodies administered in
pregnancy. Am J Obstet Gynecol. 2022;226(5):743–5 (https://www.ajog.org/article/S0002-
9378(22)00040-0/fulltext, accessed 25 March 2024).
65. Thilagar BP, Ghosh AK, Nguyen J, Theiler RN, Wick MJ, Hurt RT et al. Anti-spike monoclonal antibody
therapy in pregnant women with mild-to-moderate coronavirus disease 2019 (COVID-19). Obstet
Gynecol. 2022;139(4):616–8 (https://journals.lww.com/greenjournal/fulltext/2022/04000/anti_
spike_monoclonal_antibody_therapy_in_pregnant.18.aspx, accessed 25 March 2024).
79
66. Manciulli T, Modi G, Campolmi I, Borchi B, Trotta M, Spinicci M et al. Treatment with anti-SARS-
CoV-2 monoclonal antibodies in pregnant and postpartum women: first experiences in Florence,
Italy. Infection. 2022;50(5):1139–45 (https://link.springer.com/article/10.1007/s15010-022-
01777-z, accessed 25 March 2024).
67. Levey NH, Forrest AD, Spielman DW, Easley KA, Dude CM, Badell ML. Outcomes of pregnant
patients treated with REGEN-COV during the COVID-19 pandemic. Am. J. Obstet Gynecol. MFM.
2022;4(5):100673 (https://www.ajogmfm.org/article/S2589-9333(22)00108-2/fulltext, accessed
25 March 2024).
68. Tuan JJ, Sharma M, Kayani J, Davis MW, McManus D, Topal JE et al. Outcomes of pregnant
women exposed to Sotrovimab for the treatment of COVID-19 in the BA.1 Omicron predominant
era (PRESTO). BMC Infect Dis. 2023;23(1):258 (https://bmcinfectdis.biomedcentral.com/
articles/10.1186/s12879-023-08198-9, accessed 25 March 2024).
69. Edwards KM, Neuzil KM. Understanding COVID-19 through human challenge models. Nat Med.
2022;28(5):903–4 (https://www.nature.com/articles/s41591-022-01778-3, accessed 24 December
2024).
70. Killingley B, Mann AJ, Kalinova M, Boyers A, Goonawardane N, Zhou J et al. Safety, tolerability and
viral kinetics during SARS-CoV-2 human challenge in young adults. Nat Med. 2022;28(5):1031–41
(https://www.nature.com/articles/s41591-022-01780-9, accessed 24 December 2024).
71. Vora SB, Englund JA, Trehan I, Waghmare A, Kong A, Adler A et al. Monoclonal antibody and antiviral
therapy for mild-to-moderate COVID-19 in pediatric patients. Pediatr Infect Dis J. 2023:42(1):32–4
(https://journals.lww.com/pidj/fulltext/2023/01000/monoclonal_antibody_and_antiviral_
therapy_for.6.aspx, accessed 24 December 2024).
72. Mak G, Dassner AM, Hammer BM, Hanisch BR. Safety and tolerability of monoclonal antibody
therapies for treatment of COVID-19 in pediatric patients. Pediatr Infect Dis J. 2021;40(12):e507–
e509 (https://journals.lww.com/pidj/fulltext/2021/12000/safety_and_tolerability_of_monoclonal_
antibody.31.aspx, accessed 24 December 2024).
73. Bahakel H, Murphy C, Frenck RW Jr, Grimley MS, Marsh RA, Paulsen GC et al. Single site experience
of the use of monoclonal antibodies for the treatment of COVID-19 in high-risk pediatric and
young adult patients. Pediatr Infect Dis J. 2022;41(12):985–8 (https://journals.lww.com/pidj/
fulltext/2022/12000/single_site_experience_of_the_use_of_monoclonal.10.aspx, accessed 24
December 2024).
74. Sherman G, Lamb GS, Sharma TS, Lloyd EC, Nagel J, Dandam NN et al. Monoclonal antibody use for
coronavirus disease 2019 in pediatric patients: a multicenter retrospective study. J Pediatric Infect
Dis Soc. 2023:12(3):152–5 (https://academic.oup.com/jpids/article/12/3/152/7079700, accessed
24 December 2024).
75. Santos JL, Bhisitkul D, Carman M, Wilson K, Hasara S, Homa K et al. The use of monoclonal
antibody therapy in pediatric patients with COVID-19: a retrospective case series. Int J Emerg Med.
2022;15(1):9 (https://intjem.biomedcentral.com/articles/10.1186/s12245-022-00414-8, accessed
24 December 2024).
76. State of paediatric medicines in the EU. 10 years of the EU Paediatric Regulation. Report from
the Commission to the European Parliament and the Council. COM (2017) 626 (https://health.
ec.europa.eu/system/files/2017-11/2017_childrensmedicines_report_en_0.pdf, accessed 24
December 2023).
80
Annex 2
77. COVID-19: developing drugs and biological products for treatment or prevention. Guidance for
Industry. Silver Spring (MD): U.S. Department of Health and Human Services Food and Drug
Administration; 2021 (https://www.fda.gov/media/137926/download, accessed 24 December
2024).
78. Reflection paper on the use of extrapolation in the development of medicines for paediatrics.
London: European Medicines Agency; 2018 (Document EMA/189724/2018; https://www.ema.
europa.eu/en/documents/scientific-guideline/adopted-reflection-paper-use-extrapolation-
development-medicines-paediatrics-revision-1_en.pdf, accessed 24 December 2024).
79. Paediatric investigation plans [website]. European Medicines Agency (https://www.ema.europa.
eu/en/human-regulatory/research-development/paediatric-medicines/paediatric-investigation-
plans, accessed 24 December 2024).
80. Canada’s approach to drugs for children and youth [website]. Ottawa: Government of Canada
(https://www.canada.ca/en/health-canada/services/drugs-health-products/drug-products/
pediatrics.html, accessed 24 December 2024).
81
Annex 3
New and replacement WHO international reference
standards for biological products
The provision of global measurement standards is a core normative WHO activity.
WHO international reference standards are widely used by manufacturers,
regulatory authorities and academic researchers in the development and
evaluation of biological products. The timely development of new reference
standards is crucial in harnessing the benefits of scientific advances in new
biologicals and in vitro diagnosis. At the same time, management of the existing
inventory of WHO international reference standards requires an active and
carefully planned programme of work to replace established materials before
existing stocks are exhausted.
The considerations and guiding principles used to assign priorities
and develop the programme of work in this area have previously been set out
as WHO Recommendations.9 In order to facilitate and improve transparency
in the priority-setting process, a simple tool was developed as Appendix 1 of
these WHO Recommendations. This tool describes the key considerations taken
into account when assigning priorities, and allows stakeholders to review and
comment on any new proposals being considered for endorsement by the WHO
Expert Committee on Biological Standardization.
A list of current WHO international reference standards for biological
products is available at: https://www.who.int/teams/health-product-and-policy-
standards/standards-and-specifications/catalogue.
At its meeting held via video conference on 11–14 March 2024, the WHO
Expert Committee on Biological Standardization made the changes shown below
to the previous list. Each of the WHO international reference standards shown in
the table below should be used in accordance with its instructions for use (IFU).
9
Recommendations for the preparation, characterization and establishment of international and other
biological reference standards (revised 2004). In: WHO Expert Committee on Biological Standardization:
fifty-fifth report. Geneva: World Health Organization; 2006: Annex 2 (WHO Technical Report Series, No. 932;
https://www.who.int/publications/m/item/annex2-trs932, accessed 22 April 2024).
83
Additions10
Material Unitage Status
Biotherapeutics other than blood products
Golimumab 500 IU/ampoule TNF neutralizing activity First WHO
500 IU/ampoule TNF binding activity International
500 IU/ampoule FcγRIII binding activity Standard
500 IU/ampoule ADCC activity
50 μg/ampoule for therapeutic drug
monitoring
In vitro diagnostics
HIV-1 p24 antigen 44 IU/ampoule First WHO
International
Standard
Lassa virus RNA for No unitage assigned First WHO
NAT-based assays International
Lineages II, III, V and VII Reference Panel
Standards for use in high-throughput sequencing technologies

Adventitious virus CBER-FSCUST-90 (hCoV) First WHO
detection in biological 2.6 x 1010 genome copies/mL International
products by CBER-FSCUST-91 (PCV1) Reference Panel
high-throughput 8.1 x 109 genome copies/mL
sequencing11 CBER-FSCUST-92 (REO)
1.5 x 1010 genome copies/mL
CBER-FSCUST-93 (FeLV)
CBER-FSCUST-94 (EBV)
CBER-FSCUST-95 (RSV)

CBER-FSCUST-96 (MVM)
Vaccines and related substances
Diphtheria antitoxin for No unitage assigned WHO International
use in flocculation test Reference
(equine) Reagent
10
Unless otherwise indicated, all materials are held and distributed by the Medicines and Healthcare
products Regulatory Agency, Potters Bar, Herts, EN6 3QG, United Kingdom.
11
Developed by U.S. FDA/CBER and distributed by BEI Resources Repository. Catalogue number NR-59630;
First World Health Organization International Reference Panel (https://www.niaid.nih.gov/research/bei-
resources-repository).
84
SELECTED WHO PUBLICATIONS OF RELATED INTEREST

Seventy-eighth report.
WHO Technical Report Series, No. 1054, 2024 (xvii + 95 pages)

Seventy-seventh report.
WHO Technical Report Series, 1048, 2023 (xiv + 137 pages)

Seventy-sixth report.
WHO Technical Report Series, 1045, 2023 (xvi + 330 pages)

Seventy-fifth report.
WHO Technical Report Series, 1043, 2022 (xii + 252 pages)

Seventy-fourth report.
WHO Technical Report Series, No. 1039, 2022 (xv + 157 pages)

Report of the seventy-second and seventy-third meetings.
WHO Technical Report Series, No. 1030, 2021 (xvii + 269 pages)

Seventy-first report.
WHO Technical Report Series, 1028, 2021 (xii + 102 pages)

Seventieth report.
WHO Technical Report Series, No. 1024, 2020 (xvi + 227 pages)

Sixty-ninth report.
WHO Technical Report Series, No. 1016, 2019 (xv + 251 pages)
Website: https://www.who.int/health-topics/Biologicals#tab=tab_1
To purchase WHO publications, please contact:

WHO Press, World Health Organization, 20 Avenue Appia, 1211 Geneva 27, Switzerland;
email: [email protected]; order online: www.who.int/bookorders
This report presents the recommendations of a WHO Expert
Committee commissioned to coordinate activities leading
to the adoption of international recommendations for the
production and control of vaccines and other biological
products used in medicine, and the establishment of
international biological reference materials.
Following a brief introduction, the report summarizes a
number of issues brought to the attention of the Committee
at its meeting held virtually in March 2024. Of particular
relevance to manufacturers and national regulatory authorities
are the discussions held on the development and adoption of
new and revised WHO Recommendations, Guidelines and
guidance documents. Following these discussions, the WHO
document entitled Nonclinical and clinical evaluation of
monoclonal antibodies and related products intended for the
prevention or treatment of COVID-19 was adopted.
Subsequent sections of the report provide information on the
current status, proposed development and establishment of
international reference materials in the areas of: biotherapeutics
other than blood products; blood products and related
substances; in vitro diagnostics; standards for use in high-
throughput sequencing technologies; and vaccines and related
substances.
A series of annexes is then presented which includes an
updated list of all WHO Recommendations, Guidelines and
other documents related to the manufacture, quality control
and evaluation of biological products (Annex 1). The above
WHO document adopted on the advice of the Committee
is then presented as part of this report (Annex 2). Finally,
all new and replacement WHO international reference
standards for biological products established during the
March 2024 meeting are summarized in Annex 3. The
updated full online catalogue of WHO international
reference standards is available at: https://www.who.int/
teams/health-product-and-policy-standards/standards-and-
specifications/catalogue.
ISBN 9789240097971

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W H O Te c h n i c a l R e p o r t S e r i e s

WHO Expert Committee

To ensure the widest possible availability of authoritative information and guidance on

WHO Expert Committee

© World Health Organization 2024

Dr Y. Sun, Paul-Ehrlich Institute, Langen, Germany

Dr M. Rosu-Myles, Biologic and Radiopharmaceutical Drugs Directorate, Health Canada,

Observers from non-state actors in official relations

International Federation of Pharmaceutical Manufacturers & Associations

Representation from other entities

Pharmaceutical and Medical Device Regulatory Science Society of Japan

World Health Organization (WHO)

Other WHO staff

Representation from WHO regional offices3

NCL national control laboratory

Dr Ivana Knezevic, Secretary to the Committee, thanked Dr Nakatani

Following the conclusion of the open information-sharing session, Dr

2.2 Cross-cutting activities of other WHO committees and groups

responses to bacille Calmette-Guérin in adults and adolescents. Several different

to support vaccine licensure and WHO prequalification. In addition, the recent

impractical, it did rely on having a well-defined correlate of protection, ideally

2.3 Other matters

updated guidance to national regulatory authorities and the

3.2 Biotherapeutics other than blood products

3.2.2 Development of WHO guidance on the nonclinical and clinical

3.3 Blood products and related substances

Recognizing the important and pressing need for WHO guidance to

3.4 Vaccines and related substances

with experts and stakeholders worldwide, a drafting group consisting of regulatory

toxicological testing (including the risk of intussusception) and provide guidance

material typically reduced the variability of potency estimates for tested

Noting the importance of this class of mAb product, the Committee

4.2 Proposed new projects and updates –

4.2.2 Proposed Fourth WHO International Standard for endotoxin

test (MAT). Users of these international standards include pharmacopoeias,

It was proposed that a donated lyophilized bulk endotoxin material

4.3 Proposed discontinuations and updates –

use of elcatonin potentially supported through available regional or national

the presence of albumin in the preparation might be problematic, with issues

laboratories in hospitals managing TTP patients be developed using pools

year. Based on Arrhenius model calculations, only a negligible loss in potency

Noting that LASV lineages I and VI were not represented in the

6.1.2 First WHO International Standard for HIV-1 p24 antigen

WHO international reference reagent, a proposal to develop a First WHO

6.2 Proposed new projects and updates – in vitro diagnostics

competing standards. The issue of low adoption would hopefully be addressed by

The candidate material would be prepared from human plasma or

panel had been based on targeted bioinformatics, collaborative study participants

First WHO International Reference Panel for adventitious virus detection in

8.2 Proposed new projects and updates – vaccines

8.2.2 Proposed Second WHO International Standard

preventative strategy for controlling typhoid, especially in areas where multidrug-

8.2.3 Proposed First WHO International Standard for antibodies

Infection with group A streptococcus (GAS) is associated with a range of diseases

underpin vaccine implementation programmes. Such a standard may also be used

8.2.4 Proposed First WHO International Standard

responses to Mpox virus and VACV. Differentiating these responses in both

8.3 Proposed discontinuations and updates –

Important note on the immediate discontinuation of the innocuity test