Addressing potential ethical issues regarding the supply of human-derived products or reagents in in vitro OECD Test Guidelines 1

The number and scope of Organisation for Economic Cooperation and Development (OECD) in vitro test guidelines (TGs) are increasing, in an effort to both improve human relevance and replace in vivo animal testing. In vitro test methods being developed for TG use are increasing the use of human based reagents, in combination with, or replacing animal derived reagents, and demand for human reagents is likely to grow in the near future. There are a range of issues associated with the ethical use of human reagents, particularly human serum, in the adaptation and development of in vitro TGs, especially to ensure that there is no human exploitation, legal requirements are adhered to, and that the origin of the reagent is assured. To address these concerns, the OECD has instigated a workshop on ethics, sources, availability and traceability of human based reagents for TG purposes, to be held in March, 2019. The focus is to provide guidance on acceptable sources of human serum for use in in vitro TGs, in terms of donor ethics and informed consent regarding commercial use and Quality Control for safety and consistent performance, with a view to providing guidance to support the adaptation and/or development of in vitro TGs using human reagents, and to ensure that in reporting the test results to regulators, clearly defined ethical and traceability aspects are adequately addressed, for the Mutual Acceptance of Data principle to be accepted in all OECD member countries. This thought-starter provides a discussion basis to achieve those objectives.


Introduction 2
The OECD Test Guideline Programme agrees upon test guidelines, for the hazard testing of chemicals that can be accepted by any member country, under the Mutual Acceptance of Data (MAD) agreement, -this saves both costs to industry, not having to redo tests for different regulatory authorities, but also reduces animal use in chemical testing greatly.Indeed, the TGP ha a strong commitment to the 3Rs, -to Replace, Reduce and Refine animals in scientific research and toxicity testing, and the replacement of in vivo animal testing, with in vitro testing has grown greatly in recent years, and is increasingly being incorporated into regulatory legislation.
Fetal bovine (or calf) serum (FBS/FCS) is most commonly used at present for the maintenance and growth of the cells used in many of the in vitro TGs.However, some cosmetics companies including Lush 3 and the Body Shop 4 have developed policies to avoid all animal generated products, and where in vitro methods are used, there is a preference to use xeno-free materials in the chemical testing, sourced solely from humans.(Xeno-free conditions are defined as those not containing any elements that are not from the same species as the cells used, in this case, human (OECD, 2018a)).In line

Current situation
At this time, while there are no global or harmonised regulations specially addressing the use of human serum for OECD test method purposes, there are regulations in individual countries that describe both quality and regulatory requirements with respect to the use of human tissues.
In Europe, human serum use within test method development and for in vitro test guidelines is considered a medical device, as it is used as part of a test method where the purpose is to diagnose and prevent disease (definition provided by the UK National Health Service, Health Research Authority7 ).Whilst there are no specific legislation examples for the purposes of TGs, there are examples of relevant legislation for the US and UK, and these are provided in Table 1.
Methodologically, for time and resource reasons, the authors utilised selected reviews and followed up directly with the OECD National Coordinators and their contacts.This paper does not therefore provide a comprehensive global review, but does however show that for several OECD member countries, whilst there are some outstanding issues to address, it should be possible in the future to satisfactorily address concerns regarding the ethical supply and use of these materials.

What are the needs and concerns of the relevant parties?
There is a need to ensure that the commercial use of human serum, for example by CROs, in Test Guideline protocols meets the varying ethical legislative requirements and guidance specifications in place in OECD member countries.There appear to be six core elements to the ethical discussion.These relate to: (i) Any health-related aspects of the donation procedure; (ii) permission from the donor for commercial use; (iii) payment of the donors and the potential for exploitation of low-income populations; (iv) overarching data protection of the donors; (v) a practical concern of sufficient supply of human based reagents from more than one commercial source, so that there is both no 'monopoly' in supply, but also so that supply can be ensured, whilst protecting the clinical supply chain, and not competing with medical needs; (vi) Cultural perceptions in relation to human organ (including blood) donation, and in vitro chemical testing.The fifth point is a broader question that also applies to the supply of key elements in other TGs, but several sources of human serum are provided in Table 2. hPL is produced from out of date blood products, and is easily available due to the short shelf life (5 days) for transfusion purposes.
An extensive global survey on hPL for cell propagation, has been conducted and published (Strunk et al., 2018a,b).Currently unknown whether the use of human serum is permitted in this institute.
Small number of batches available and more expensive than FBS.

Switzerland / Large Company
Human serum available -from the blood bank & probably other sources Current understanding is that there are no restrictions on blood donation for life science applications, or any customs restrictions on the import of commercial blood products of human origin.Import permits can easily be obtained from the Government.
Expired donations from the blood bank has been used in the past and was easy to obtain.Not aware of any cultural barriers or restrictions.

Tab. 2: Examples of responses received regarding how the ethical considerations in the use of human serum are currently addressed in selected OECD member countries
Questions arising from the information supplied are indicated in bold italics.

Country/ Government
Donor consent and relevant ethical permission considerations/legislation References UK/ Government In the UK, the Human Tissue Act 2004 sets out the legal requirements for consent for the removal, storage and use of human organs and tissue for scheduled purposes in England, Wales and Northern Ireland.Legal requirements for Scotland are set out in the Human Tissue (Scotland) Act 2006.The Human Tissue Authority (HTA) is the regulatory body that oversees the consent and licensing requirements set out in the Act.A number of factors are relevant when considering whether consent is legally required for the removal and use of human material: Firstly, the purpose for which the material is removed or donated, as only scheduled purposes as defined in the Act are within scope.For example, when material is donated by the living for performance assessment or quality assurance purposes, the consent requirements do not apply as these activities are not considered as scheduled purposes under the Act when the material is donated by the living.
A further consideration is whether the material in question is considered as 'relevant material' as defined in the Act -'material other than gametes, which consists of, or includes, human cells'.Human serum is normally not considered as relevant material, as centrifugation is expected to remove intact human cells from the resulting serum.Further information on consent for the removal of relevant material is available in the HTA Code of Practice A on Consent.
Another consideration is that the consent provisions of the Act do not apply to imported material.However, the HTA considers it good practice to ensure mechanisms are in place in the source country for obtaining consent.Further information on imported material is available in the HTA Code of Practice E on Research, paragraphs 98-114.
There are also a number of consent exceptions when relevant material is used for research purposes, these are set out in paragraphs 56-69 of Code E. it is one way to deliver transparency and foster trust between researchers and participants; and so should be sought in most situations.
Whenever seeking consent, the information provided to potential donors should be proportionate and understandable, supporting the participant to make the decision to donate or not.Appropriate consent is based on the principle that competent individuals are entitled to choose freely whether to participate in research and should be given appropriate information to be able to make this choice.As an additional safeguard, it should be complemented by independent ethical review of the consent process and proposed protocol.
It is important that there is clarity of arrangements for allowing commercial access to human biological material originally donated for research projects… Where possible, participants should know when their sample or products derived from it may be used by the commercial sector, and the potential benefits of this access.It is also important to let the participant know they will not be entitled to a share of any profits that might ensue, as is also the case for IP rights generated from sample use in the academic sector.Medical uses of human blood donations include well known blood transfusion needs, and the derivation of therapeutic blood plasma to manufacture albumin and gamma globulin, both of which are in high demand for hospital usage.These donations are also critical for the cell therapy market, where for human treatment purposes, tests need to be developed and propagated using fresh human blood products.
There is a responsibility on Contract Research Organisations (CROs) to ensure compliance by using human serum and other human-derived reagents from a reputable source.Currently, there is need for a clear framework to guide test laboratories through this process.This work is in progress in regard to animal derived materials, and is being initiated for human serum products, by the International Serum Industry Association (ISIA).
With respect to vi), key examples of global variability in the cultural aspects of human organ/blood donation can be summarized as follows: a) Countries that have predominantly western belief systems that hold to the sanctity of life before birth such as Italy (where Catholicism is the state religion) do not hold with the use of human fetal stem cells b) Animal experimentation and animal welfare concerns which espouse no or minimal exploitation of animals, prioritize human derived materials for scientific research and test method development c) Shintoism, has integral notions of cleanliness and purity of the human body, the pollution of death and the deceased and respect for the ancestors, this reduces the availability of human derived materials in clinical and scientific research, and translational applications in chemical regulation.
(i) Health-related aspects of the donation procedure: tissue screening and need for post manufacturing treatment of human serum Human products are screened for a certain panel of viruses in line with the regulations pertaining in the country of collection, but this applies only if material is collected from a registered donor centre.If collected elsewhere then there could well be no viral testing carried out.Generally, all commercial human serum, HSA and hPL products are screened for viruses including HBsAG (Hepatitis B Surface Antigen), HIV-1/2 (Human Immunodeficiency Virus Type 1 and 2), and HCV (Hepatitis C Virus) and syphilis using FDA approved methods.The final pool is also tested for HCV RNA, HBV (Hepatitis B Virus) DNA, HTLV-1/2 (Human T-Lymphotropic Virus) and HIV-1RNA.They are also screened for mycoplasma infection, etc.Typical examples of certificates of analysis are provided by reputable suppliers.Laboratory work with commercial human serum, HSA and hPL products requires strict use of biosafety hazard cell culture hoods, where FBS does not.
It also needs to be clarified that material is required to be tested for only certain viruses.A pool of donor material, such as that used to produce HSA, pooled serum and plasma products etc, can be made up from donations from many hundreds of donors and each of these can be carrying viruses that are not on the test panel.
The ISIA have conducted extensive work on the gamma radiation of serum to gamma irradiation of serum intended to mitigate the risk of introducing adventitious contaminants into cell cultures and thereby improve safety (Brown et al., 2018;Croonenborghs et al., 2016;Versteegen et al., 2016).
Follow up: Develop an obligatory checklist based upon the information from ISIA to include the above screening tests.

(ii) Permission from the donor for commercial use
Major global human serum suppliers generally source in the US, where the donors may be paid, and provide consent forms, and in Europe, where the donors are generally not paid, and consent forms are provided to and signed by the donor.With respect to the UK Human Tissues Act 2004, the factors considered relevant when considering whether consent is legally required for the removal and use of human material, only scheduled purposes defined in the Act are within scope.
Where material is collected from paid donors, such donors are often on a low income, and in need of the income.This drives them to donate in several donor centres, and their health status is not always of the highest.In the US and the UK, coordination of donation registries do monitor individual donations within specific time limits, and alert the blood collection centres where these are being exceeded.
Where material such as human serum and human platelets for the manufacture of hPL is donated by the living for performance assessment or quality assurance purposes, the consent requirements do NOT apply, as these are not considered to be scheduled activities under the Act.
It must be noted that human serum is normally not considered as 'relevant material' as centrifugation removes intact human cells from the resulting medium.
While it is considered good practise to ensure mechanisms are in place in the source country for obtaining consent, consent exceptions do exist when relevant material is used for research purposes, and not for clinical purposes, as shown in the examples in Table 2.
Follow up: Work on this issue might need to include direct approaches to collection centres and further examination of consent forms.Development of explanatory information for the donors will be required.
(iii) Payment of the donors and the potential for exploitation of low income populations Donors are commonly paid in the US, but not in Europe.Whilst globally there are serious concerns and legislation in place in an attempt to prevent human organ trafficking and exploitation of vulnerable populations, the legislation in place in the US and UK does not consider human extracted blood products to fall within the legal requirements (Table 1).However the customer service guide for the use of blood products for non-clinical purposes provided by UK National Health Service Blood Transfusion (NHSBT) for organisations seeking access to donated human material for non-therapeutic use, specifically stipulates that currently the NCI supply chain does not permit the issue of donated material for use as a tissue culture medium supplement (NHSBT, 2018).However, there are good indications that this could be revised (See (v) below).
Follow up: Develop an understanding of global procedures for payment of donors (iv) Data protection of the donors Data protection is also an increasingly important issue, for example, the General Data Protection Regulation (GDPR) came into force in the UK on 25 th May 2018 and significantly increased expectations in this area.In the US, the HIPAA regulations, addressing health data protection, would be expected to have relevance to the serum donation process.Follow up: Understand global data protection requirements

(v) Competition with altruistic therapeutic donation
With increasing use of human serum/products for OECD Test Guideline Programme purposes, there could be increased competition in the availability of these resources.For example, as indicated in iii) above, in the general guidance provided by the UK NHSBT for organisations seeking access to donated human material for non-therapeutic use, it is stated that currently the NCI supply chain does not permit the issue of donated material for use as a tissue culture medium supplement.
(NHSBT, 2018).However, there are instances where blood donations are not adequate for therapeutic uses, and where blood is classified as medical waste.This waste off-the-clot whole blood could be reclassified and potentially become a good source for serum extraction, for non-clinical uses particularly with respect to the OECD Test Guideline Programme.
In Japan, blood donations are lower than other geographies at less than 1% of the population, compared to 3.5 % in other parts of the world, according to WHO statistics for 2010.Hence domestic utilization of blood donation products results in a shortage of available material for therapeutic uses, and the likelihood of sufficient waste material being available for production of human serum is likely to be low to non-existent in Japan.
In the UK, 2% of whole blood donated becomes medical waste.Indicative figures for volume and frequency of use are needed to ensure availability of supply (NHSBT, 2018).
Follow up: Work to ensure that wherever possible blood donations that would otherwise be considered waste and disposed of, should be the priority source for the production of human serum, assuming that they meet all the requirements as a raw material.There should however be systems put in place to ensure that there is no competition with clinical and therapeutic applications.

(vi) Cultural perceptions in relation to human organ (including blood) donation, and in vitro chemical
testing At its best, 'Science' is all too often considered to be an objective, not subjective way, to gather evidence which transcends most cultural and religious differences.To avoid subjective bias, good practise in scientific lines of inquiry has given rise to gold standards in well controlled study designs, informed consent and appropriate statistics.While objectivity is the goal of scientific investigation, cultural bias can cause unexpected and unconscious subjectivity to encroach on scientific routes of enquiry.
Within Western belief systems, aside from a very small number of minority religious groups, it is assumed that it is ethically acceptable and indeed scientifically preferable to use human tissues, so long as there is no detriment to the human source, or in the case of organ donation, the person is deceased (excepting kidney donation, but this is outside the scope of this paper).
We are also seeing increased public concern regarding the use of animals in science, and the growth of animal welfare considerations has led to and been integrated into Directives and also legislative requirements in the EU, particularly Directive 2010/63/EU revising Directive 86/609/EEC on the protection of animals used for scientific purposes which was adopted on 22 September 2010.The Directive is firmly based on the principle of the 3Rs, to replace, reduce and refine the use of animals used for scientific purposes.The 3Rs does not specifically call out material derived from animals that is a byproduct of another industry, such as the meat industry.Whilst it is recognised that we have a duty to use all by-products from the slaughter process to the maximum, to reduce waste, and to add value to those animals that are slaughtered for meat purposes, it is also recognised that suffering of the fetus should be avoided (More et al., 2017, Nielsen and Hawkes, 2019, van der Valk et al., 2018).
In the EU, the Cosmetics sector is the first manufacturing sector, where in vivo animal testing is specifically prohibited.The EC Cosmetics Directive, Chapter V Article 18 specifies, without prejudice, that animal in vivo testing is prohibited, and only alternative validated methods should be used prior to placing the final formulation on the market. 9uch alternative test methods include in vitro test methods that may include materials of animal origin, such as fetal bovine serum, and animal derived antibodies.They also include in silico based analysis of existing data and 'read across'10 (As stated previously, some companies have developed policies to avoid all animal generated products, and where in vitro methods are used, they are only prepared to use xeno-free materials in the chemical testing. In Japan, the Shinto and Buddhist faiths predominate.Over 53% of the population adhere to Shinto beliefs of purity and the wholeness of the physical body.The human body and tissues thereof are sacred.Anything connected with death or the dead is considered particularly polluting.Whilst legal in Japanese law since 1997, and eased in 2010 by the Organ Transplant Act 11 , according to Shinto tradition organ transplantation is not acceptable after someone has died, and this is very much reflected in practise in the World Health Organisation deceased organ donation statistics in Japan, which are far lower than other developed economies (overall 0.83% in 2010 compared to 3.52% globally12 .)Shintoism impacts also upon living donations, such that blood donation in Japan comes from less than 1% of the population 13 .
Thus in Japan, with a preference for the use of domestically produced reagents in chemical testing, and with blood donations in very short supply, competition with non-therapeutic uses of human blood and serum will be detrimental to clinical uses, and animal sourced products are ethically far more acceptable than human sourced products.
In Japan, numbers of animals used in in vivo testing are minimised as far as statistically reasonable, and are generally harmonised with OECD and ISO standards.The sacrifice of life by the test animals is highly respected: commonly animal in vivo laboratories and organisations running in vivo studies have Shinto shrines and traditional ceremonial days to mark respect and reverence.
4 To what extent have these issues been addressed in the OECD Good In Vitro Method Practices document (OECD, 2018b) The use of human serum for cell culture applications is already referenced in the Guidance Document on Good In Vitro Method Practices (GIVIMP) (OECD, 2018b).This discussion does not go into great detail on the sources and ethics, but most major points are briefly addressed.Additional relevant references particularly regarding hPL, are provided therein and in Table 2.
The GIVIMP states "Furthermore, it is recommended to develop new in vitro methods with a serum-free, chemically-defined medium, to avoid potential sources of uncertainty that may be introduced by using animal serum."While it is agreed that this is an ideal approach for the longer term, it is important to recognise that, for many cell types, serum-free or chemically defined media do not yet exist, and their development is extremely time-consuming and expensive: and the GIVIMP document acknowledges this (OECD, 2018b).Commercial organisations are not willing to spend years investing in the development of a defined medium, only to then give away the composition.While there are also issues with components of chemically-defined medium giving rise to batch-to-batch variability and performance, these are minimal if sourced from the same supplier, compared with serum.Furthermore, the development of these types of media is a relatively low priority for funding bodies.While such media may provide the potential for optimal reproducibility between experiments, it could also be argued that synthetic, defined media may not be able to match the realistic level of representation of a human population provided by the use of pooled serum from a large number of human donors.In the context of regulatory safety testing, variability between donors may be viewed as a realistic reflection of the human population and a pooled product reflects this variation.
The GIVIMP states, that in the context of human serum, "The same critical points, e.g.batch-to-batch variability, as for any serum-derived products hold true.These issues are compounded by the reduced availability of large-scale batches and high variability of batches due to the diversities of human donors, which should be taken into consideration when using human serum."It is worth noting, in this context, that pooled human serum is freely available in lots containing material from multiple donors.As already mentioned, such sources may be assumed to provide a good "average" indication of the human populationa major scientific advantage in terms of regulatory safety testing.However, it is acknowledged that this could complicate the ethical clarity as all of these donors must be shown to have given appropriate consent.Thus a batch of 500 litres of human serum may have donations from as many as 250 donors, with each donor giving a maximum of 500mL blood, which translates into 200 -250mL serum.CROs currently depend on a single statement from the supplier to cover the whole batch, so it is vital that appropriate traceability measures can be clearly demonstrated by the serum suppliers.
In some applications, the use of human serum in cell culture may be expected to provide a superior in vitro model of human physiology compared with animal sera.For example, as cited in the GIVIMP (OECD, 2018b), "FCS compared to autologous (human) serum has been found to induce a more differentiated and less stable transcriptional profile in human bone marrow mesenchymal stem cells, particularly at late passages, as shown by analysis of genome-wide microarray analysis (Shahdadfar et al., 2005).

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To what extent have these issues been addressed through other information sources?
With respect to ensuring consistent commercial sources of human serum, and the need to avoid reliance on a single supplier, discussions were initially focussed on the FDA-approved supply from Sigma Aldrich (now Merck).Follow up investigations have confirmed a large number of other commercial sources, as shown in Table 1.Many of these suppliers are also certified members of the International Serum Industry Association (ISIA) (see Section 5).While most suppliers originate in the USA, many have a network of global distributors, ensuring easy access to the products from wide geographical territories across Europe, Asia and beyond.Human serum is relatively simple to ship around the world, because there is currently no customs requirement for an equivalent to the veterinary certificate required for FBS.This makes human serum easier to transport but also more difficult to control in this context.Greater control is needed on the importation of human serum to ensure future traceability.
CROs require access to a consistent commercial supply of human serum, with full ethical and Quality Control approval, especially for regulatory work.The dominance of the US market in terms of global supply raises the particular importance of understanding the details of the ethical process in the US.Certificates of Origin from suppliers may include a statement such as "All donor units are collected in donor centers in the US which are licensed by the FDA" (example taken from Sigma/Merck).It is important to understand what such statements mean in terms of the detail of the informed consent process.It is currently understood that the FDA is not directly involved in the ethics of donation and informed consent during its inspection and approval of collection centres.Furthermore, preliminary investigations suggest that the responsibility for the specific content of consent forms rests with individual collection centres in the US.
In Europe, there is a standardised approach to informed consent.In France there are examples of collection as part of medical waste, and in the UK, Sweden, and Austria products are generated from out of date blood donations.A cluster of collection centres exists in Germany, for which one consent form, which is known to comply to the European standard, states about the donated blood: "its components are used in the context of patient care; in rare cases the blood or the components are used for standardization purposes as well as for quality control or for research and development projects."(Personal communication).
CROs have a responsibility to ensure that the sources of human serum used in their laboratories are sourced in an ethical way.Certified members of the International Serum Industry Association (ISIA) (www.serumindustry.org)must adhere to a strict code covering both full traceability and consistent product quality.As stated in the GIVIMP document (OECD, 2018b), "Ideally, sera should be obtained from vendors that can provide traceability certification from industry bodies such as the International Serum Industry Association (ISIA)."ISIA is also US-based but has a global membership and global influence.However, a traceability scheme specifically for human serum is not yet available through this organisation (see note below in Section 5).

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Issues requiring further elucidation Instigation of traceability audit scheme for human serum Whilst a traceability audit scheme for FBS has been developed by ISIA and implemented over the last 10 years, this is not yet the case for human serum.This is currently under discussion and investigation, and would apparently be straight forward to instigate, following the model established for FBS.This is being discussed with the ISIA.This expansion of the protocol would involve expanding the front end of the existing system to include informed consent verification and other, as yet unclear, details.Alternate possibilities need to be made available until this programme becomes available, such as purchasing the serum from a reputable commercial source with an appropriate Certificate of Origin that includes an ethical statement, and specifically addressing donor consent to be provided.This could also be requested from suppliers of human serum made from out of date batches, and from material generated from medical treatments.A commitment from the ISIA to institute such an audit scheme will include fair, reasonable, and non-discriminatory ("FRAND") terms (as recommended in OECD draft guidance 2018, currently unpublished, revised in February 201914 ), in their membership/associate membership subscription fees.
Informed consent: The inclusion of specific wording relating to the potential commercial use of the donated blood Follow up: Should this be sought across all member countries where human serum is collected for commercial / research applications?Follow up : Clarification with the FDA Center for Biologics and Research regarding the approach to regulation/ standardisation of the informed donor consent process in the US, and also the role of the US Health Insurance Portability and Accountability (HIPAA) regulations Follow up: Request further additional detailed input from OECD member countries Whilst Table 2 provides a selected and preliminary overview of common supply, it would be useful to gain a better understanding of the similarities and differences between expectations in North America, Asia and Europe.It is noted that the two dominant geographical sources of human serum for research and commercial use worldwide are the US and Europe.A structured approach to inviting further detailed input from member countries regarding local issues would be valuable.Information on local sources of human serum, how it is regulated to ensure ethics and performance, customs restrictions on import and cultural expectations would be very useful to elucidate further, to gain a truly international picture of any challenges faced.
Figure 1 provides a simplified overview of the production of human serum and the necessary traceability, auditing, geography, and import/export checks necessary for each stage of the production line.

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Discussion of issues raised to consider before initiating further work at the OECD It should be noted that the use of human serum is largely optional within the OECD Test Guideline Programme-there are currently no TG's exclusively demanding the use of human serum.Therefore, the option to use human serum allows member countries to do so when ethical sources are available, but such an approach does not exclude others from using the TG methods, as they can revert to the use of animal sera if required, and this will be the case for Japan for instance.Further development of the suggestions within this thought starter will be facilitated by a targeted 'Ethics and the use of human reagents in OECD test guidelines' workshop to adequately address ethical and regulatory considerations in the sourcing of materials, together with examination of appropriate approaches for global origin identification auditing, and

Fig. 2: Overview of the International Serum Industry Association Global Network
Reprinted with permission from ISIA standardisation measures, as has been done for hPL (Strunk et al., 2018a,b).The participation of the International Serum Industry Association (ISIA) (Fig. 2) will be key to the workshop, as the organisation works promote standards of excellence and ethics in the animal serum and animal derived products industry, representing and advising the suppliers of 90% of the animal serum used in life science applications worldwide, utilizing programs that have immediate relevance to human serum market.
In April 2018, the OECD Working Group of National Coordinators to the Test Guideline Programme (WNT) requested that the (forthcoming) workshop in March 2019 will be in accordance with, and expanding upon the OECD GIVIMP (OECD, 2018b), on the use of human serum within the OECD Test Guideline Programme, such that evidence of ethical considerations focusing on sourcing of the human blood derived products such as serum, should be included with the test report.Failure to adequately and satisfactorily address and meet the criteria will result in non-acceptance (noncompliance) of the test method results by OECD member regulatory authorities.Essential components will be: − Demonstrated membership of traceability audit scheme − Provision of certificate of origin of human serum specifying: − Donor permission freely given − Any compensation received by the donor (e.g.payment) − Source (i.e.preference for: out of date blood product/ medical waste not fresh, that has been sourced primarily for medical needs) Building upon, the overview of the production of human serum and the necessary traceability, auditing, geography, and import/export checks necessary for each stage of the production line, Table 3 provides a more detailed overview of likely key data requirements for these essential components concerning the origins of human reagents used in in vitro cell culture for OECD test guideline purposes to be submitted with the test results, to regulatory authorities.For further investigation and refinement in collaboration with OECD nominated experts, serum suppliers, the ISIA, and the WNT).

+
It is therefore proposed that where versions of 'me-too' or 'xeno-free' modified test guidelines specify the use of human serum, they should state that users may only obtain human serum from certified companies i.e.ISIA members.(There is an related precedent to this as some TGs already stipulate the use of products from particular suppliers, e.g. the validated tissue models for skin and eye irritation in TGs 439 and 492 respectively).While there is currently no formal traceability geographic standard for human serum in the same way as that built for animal serum, the ISIA have initiated and contracted work on this, and the contractor has proven concept for the building of a reference database for human serum.This is necessary to underpin a traceability audit scheme (see figure 1).Preliminary results have demonstrated initial feasibility, and will be the basis for planning further work.The work underway does not yet address the issues of other human derived material such as HSA, hPL etc.
Ultimately, there is also a responsibility on the test laboratory / CRO to ensure ethical compliance, and they will need to be adequately provided with this information, by their suppliers.Ideally it would be audited, but feasibility needs investigation.

Proposed way forward
Although not a comprehensive review, on the basis of the breadth of the evidence provided in 2018, the OECD WNT were assured that multiple sources of human blood products are available, some of which are provided by consenting donors that do not require a (e.g.HTA) license; and some of which no longer require consent (e.g.out of date blood products, medical waste products), and that these products are available for use in the in vitro test methods.The literature evidence for the performance of these human derived products is as good as FBS in certain applications, but there need to be substantial improvements in the rigour of documentation.However, whilst there is currently a plentiful supply that can be distributed globally, increasing use/adaptation in TGs will potentially increase demand, and we need to ensure that the provision of such materials will not threaten donor and/or wider public support for altruistic therapeutic donation.This is an area where Japan is particularly vulnerable, with a low level of domestic blood donations.Given cultural differences with test methods that exclusively use human serum, if alternative protocols including FBS, chemically defined serum, serum free medium etc., are not included in the TG, it is unlikely that such TGs will be able to be harmonised at the OECD level at the current time.Thus, the standard in vitro TG methods using FBS are available to all OECD member regulatory jurisdictions, but test method users wishing to use xenofree TG adaptations where developed, will be able to do so, so long as the ethical and QC concerns have been adequately and demonstrably addressed.
In April 2018, the WNT agreed to undertake the following next steps, to include; i) Instigating a workshop on sources, availability, traceability of human based reagents for TG purposes, this will be held at the OECD on March 18-19, 2019.ii) A focus on providing CROs with guidance on acceptable sources of human serum for use in in vitro TGs, in terms of QC for safety and consistent performance, donor ethics and informed consent regarding commercial use.This may best be achieved through dialogue and collaboration with the serum industry (via ISIA), regulators, users and other stakeholders across member countries as part of the workshop preparation and outputs.

Fig. 1 :
Fig. 1: Overview of the production of human serum and the necessary traceability, auditing, geography, and import/export checks necessary for each stage of the production line Note that testing is carried out on individual donor units after collection, and throughout the process.

Tab. 1: Examples of how human serum is sourced by different test method developers and users, summary of OECD National Coordinator survey responses conducted in 2017
Questions arising from the information supplied are indicated in bold italics Registration with the US FDA: The FDA does not give "approval" for facilities but they have registrations.The site manufacturing this material is registered with the FDA.Testing and guidelines: Both H4522 and H6914 are tested under FDA requirements and are collected under AABB guidelines.Batch-to-batch consistency of the product specifications: Batch to batch consistency is good, but different donors in different batches can always cause differences.Long term supply (>5 years)?Sigma: We have used the current collection centers for >15 years.

Products are class 1 medical devices intended for research use only. Not for therapeutic applications. Does research use include commercial use also?
As freely supplied to CROs, it is assumed so, but would be useful to confirm.Human AB serum is collected from healthy volunteer male donors of the AB serotype at FDA-licensed facilities in the United States and collected in compliance with health requirements established by 21CFR 640, subpart G.All approved donor units are also tested according to testing requirements for communicable disease agents as stated in 21 CFR 610.40.
All units are negative (non-reactive) for HBsAG (Hepatitis B Surface Antigen), HIV-1/2 (Human Immunodeficiency Virus Type 1 and 2), and HCV (Hepatitis C Virus) and syphilis using FDA approved methods.The final pool is also tested for HCV RNA, HBV (Hepatitis B Virus) DNA, HTLV-1/2 (Human T-Lymphotropic Virus) and HIV-1 RNA.Converted serum is routinely screened for the presence of animal contaminants.This is performed by DNA analysis.

similar global survey would be useful for human serum.
Human blood products, from which human serum is obtained, are past a 'use by' date, cannot be used for human use, and would be incinerated as waste, if not repurposed.

It does not appear to state a requirement to provide information on the potential commercial or research use of the donated blood or its components.
Within the US Code of Federal Regulations, 21 CFR 640 covers the requirements for informed consent from donors of human blood products.The text of the regulation covers a requirement for donors to be fully aware of any health implications of the donation procedure.
Within the FDA, the Center for Biologics Evaluation and Research (CBER) is responsible for regulating blood products for transfusion and the manufacture of pharmaceuticals.It is not currently clear whether they also cover life science applications.