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ISSN 1757–6180
Bioanalysis (2010) 2(3), 393 –395
10.4155/BIO.09.168 © 2010 Future Science Ltd
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Human microdosing belongs to the group of
new, advanced experimental approaches that
meet 21st Century requirements. As suggested
by the Centre for Medicines Research, without
a new generation of product-development tools,
it will be difficult to improve the 20-year low in
the number of new medical therapies launched
onto the market [1], despite more investment
in biomedical research worldwide over the last
two decades.
As mentioned in the ‘critical path’ report
published by the US FDA [101], 92% of com-
pounds that pass animal tests fail in Phase I
clinical trials. A technique such as microdosing,
which provides early data about the behavior
of drugs in humans at very low doses, can only
improve predictions of drug toxicity and effi-
cacy, whilst also reducing the resources spent
and the number of animal tests carried out.
In streamlining the drug-development pro-
cess, human microdosing is also likely to reduce
animal testing, a responsibility shared by com-
panies, regulators and individual scientists
worldwide. Legislation in Europe requires the
Replacement, Reduction and Refinement (the
‘Three R s’) of animal experiments wherever
possible [2]. Similar policies are implemented in
many countries, including the USA.
In the EU, the Commission and the member
states also have a legislative duty to encourage
research into methods that could achieve equiva-
lent objectives, but using fewer animals or none
at all. The European Centre for the Validation of
Alternative Methods (ECVAM) was established
in 1992 for this reason and, since then, some 27
nonanimal models and assays have been scien-
tifically validated as full or partial replacements
for animal tests and 20 have gained regulatory
approval [102].
More than 12.1 million animals were used
in 2005 (latest available figures) in experiments
in the EU, of which 528,189 were used for
safety testing of pharmaceuticals [3]. As well as
their use in studies of absorption, distribution,
metabolism and excretion (ADME), animals are
used to assess longer-term toxicities of pharma-
ceuticals, such as subchronic and chronic toxic-
ity (67,651 animals in the EU in 2005), devel-
opmental and reproductive toxicity (49,026
animals) and carcinogenicity (26,589). The
wider use of human microdosing would mini-
mize all these animal tests, which are conducted
later in drug development, by providing human-
specific ADME data and, thus, identifying early
those compounds destined to fail later owing to
suboptimal pharmacokinetics or metabolism [4] .
A 10% improvement in identifying failing can-
didates before classic clinical trials could also
save US$100 million in development costs per
drug [5].
The British government’s Animal Procedures
Committee, which advises on animal experi-
ments, recommended in its 2002 report on
the use of primates that human microdosing
should be further developed and resourced. The
Committee recognised that microdosing has the
potential to limit the use of primates in repeat-
dose toxicity tests, pharmacokinetics and safety
pharmacology studies [6]. In 2008, repeat-dose
tests alone (subacute, subchronic and chronic)
involved 2346 primates in Britain [7].
In contrast to the fairly extensive animal
data required prior to a Phase I trial, single-
dose rodent tests can be the primary support
for microdose studies in humans and the
European Medicines Agency recognizes that
micro dosing, among other kinds of exploratory
clinical trials “can reduce overall animal use in
drug development” [103].
The use of subpharmacological and sub-
therapeutic microdoses in early clinical stud-
ies also offers better protection for Phase I trial
volunteers, without compromising the safety of
microdose subjects. It has been suggested that a
microdose study of TGN1412 might have pre-
vented the tragedy that occurred in 2006, when
Phase I trial subjects suffered life-threatening,
unexpected side effects despite extensive pre-
clinical tests on animals, including rhesus and
cynomolgus monkeys.
The wider use of human microdosing would minimize all these animal tests … by providing human-specic ADME data and,
thus, identifying early those compounds destined to fail later owing to suboptimal pharmacokinetics or metabolism.
Microdosing: safer clinical trials and fewer
animal tests
Gill Langley
Author for correspondence
Dr Hadwen Trust for
Humane Research, 18 Market
Place, Hitchin, Herts,
Tel.: +44 1462 436 819
Fax: +44 1462 436 844
E-mail: sciencesources@
Sebastien Farnaud
Dr Hadwen Trust for
Humane Research, 18 Market
Place, Hitchin, Herts,
Tel.: +44 1462 436 819
Fax: +44 1462 436 844
E-mail: sebastien@
394 future science group
news & AnAlysis | OpiniOn
TGN1412 targets the CD28 receptor on
T-cells, where there are differences between
the rhesus monkey and human sequences. A
microdose study with TGN1412, by systemic or
dermal application, could have determined the
amount of the antibody that bound to T cells
in the whole human body without risk to the
subjects, as well as providing pharmacokinetic
and metabolic data relevant to the human spe-
cies [8]. Safely obtained human microdose data
may have been able to improve the design of the
Phase I study.
In fact, the Duff report on the TGN1412 inci-
dent stated that “the view that higher-risk agents
should be given at specified low dose levels, for
example in the microgram or nanogram range
(‘microdosing’ and ‘nanodosing’) may have
value as a general guideline” [104].
The human relevance of microdosing applies
to a wider field than drug toxicology, as illus-
trated by its proposed uses in oncology. For
example, applying microdosing to measure
interindividual variability in drug disposition
caused by genetic and environmental factors is a
promising application [9]. In addition, microdos-
ing offers opportunities to conduct new study
designs and investigate previously unanswerable
questions in oncology [10].
The value of microdosing in other sectors
has already been recognized. As a case in point,
microdosing was considered by an ECVAM
expert group as a potential replacement for most
of the toxicokinetic and ADME data normally
obtained from animal tests for chemicals used
in cosmetics [11] .
The role of nonanimal techniques
Many researchers, governments and regula-
tors now agree that replacement techniques are
often more relevant, reliable, sensitive, cost-
effective and reproducible than experiments
on animals [105]. The FDA’s report stated that
a major problem is the failure to create and use
novel tools to deliver “fundamentally better
answers about how the safety and effectiveness
of new products can be demonstrated, in faster
time frames, with more certainty, and at lower
costs” [101].
The FDA called for a greater emphasis on
in vitro, clinical and computational tools. For
example, using human cell lines to character-
ize drug metabolic pathways can predict human
metabolism and help eliminate compounds with
unfavorable metabolic profiles. This approach
has meant that clinical failures due to drug-
interaction problems are now far less likely.
Computational modeling is increasingly being
applied to human disease simulation, to predict
novel drug pharmacokinetics and to conduct ‘vir-
tual’ clinical trials. In terms of ADME simula-
tion, the mathematical relationship between drug
doses, plasma concentrations, pharmacokinetics
and pharmacodynamics can be characterized
and patient covariates are included in the com-
putational model. The systematic application
of mathematical modeling could significantly
improve drug development.
Analytical techniques have already replaced
many animal tests and in so doing have improved
sensitivity, reliability and precision [12]. For exam-
ple, digitalis was once routinely tested for potency
on guinea pigs and pigeons using a lethal method.
In the late 1980s, this was replaced by a chemical
colorimetric assay which directly measured the
content of digitoxin. In 1980, insulin batch test-
ing used 600 mice per test, but was later replaced
by HPLC – a more rapid and precise method [13].
Where now?
The number of consortia and organizations work-
ing to replace animal tests in medicines develop-
ment is growing fast. The European Partnership
to Promote Alternative Approaches to Animal
Testing was established in 2005 as a joint ini-
tiative between the European Commission,
European trade associations from seven indus-
try sectors (including pharma ceuticals) and
individual companies. Its purpose is to promote
the development and implementation of mod-
ern Three R approaches, including non animal
methods, in the field of safety testing [106].
In 2009, an international memorandum of
co-operation was signed by four agencies, in the
USA, Japan, Canada and the EU, to co-ordinate
the adoption of nonanimal testing methods. The
agreement “will speed the adoption of new test
methods based on advances in science and tech-
nology. Animal welfare will also be improved
by the national and international acceptance of
alternative test methods that reduce, refine and
replace the use of animals”, according to William
Stokes, an assistant surgeon general in the US
Public Health Service and a leading scientist in
the Three Rs field [107].
Although nonlinear absorption or disposi-
tion characteristics could affect the validity
of microdose predictions of therapeutic-level
pharmaco kinetics, recent publications suggest
that a concordance of approximately 80% can
be anticipated [14]. Its direct and safe applica-
tions for humans, together with its economical
and ethical benefits, bring microdosing to the
Bioana lysis (2010) 2 (3)
www.fut ure- scie 395
future science group
OpiniOn | news & AnAlysis
forefront of 21st Century technology. Combining
human microdosing with other complementary
non animal approaches clearly offers ethical, sci-
entific and efficiency benefits for pharmaceutical
development. Wider recognition of this should
lead to faster progress towards that goal.
Financial & competing interests disclosure
Both authors are employed by the Dr Hadwen Trust for
Humane Research (Registered Charity No 261096), the
UK’s leading medical research charity exclusively funding
nonanimal research techniques to replace animal experi-
ments, benefiting people and animals. The authors have
no other relevant affiliations or financial involvement
with any organization or entity with a financial interest
in or financial conflict with the subject matter or materi-
als discussed in the manuscript. This includes employ-
ment, consultancies, honora ria, stock ownership or
options, expert t estimony, grants or patents received or
pending, or royalties.
No writing assistance was utilized in the production of
this manuscript.
1 Centre for Medicines Research International.
International Pharmaceutical R&D Factbook.
Thomson Reuters (2009).
2 Council Directive 86/609/EEC of
24 November 1986 on the approximation of
laws, reg ulations and administrative
provisions of the Member States regarding the
protection of anima ls used for experimental
and other purposes. Off. J. Eur. Comm. L358,
1–29 (1986).
3 Commission of the European
Communities, Brussels , 5.11.2007.
COM(2007) 675 fina l. Fifth Report on the
Statistics on the Number of Animals used for
Experimental and other Scientific Purposes in
the Member States of the European Union
4 Combes R D, Berridge T, Connelly J et al .
Early microdose studies in human volunteers
can minimise animal testing: proceedings of a
workshop organised by volunteers in resea rch
and testing. Eur. J. Pharmacol. Sci. 19, 1–11
5 Boston Consulting Group. A revolution in
R&D – how genomics and genetics will affect
drug development costs and times. In: Parexel
Pharmaceutical R &D Statistical Sourcebook.
6 Animal Procedures Committee. The use of
primates under the A nimals (Scientific
Procedures) Act (1986): analysis of current
trends with particular reference to regulatory
toxicolog y. (2002).
7 Home Office. Statistics of Scientific
Procedures on Living Animals Great Britain
2008. HC 800. The Stationery Office,
London, UK (2009).
8 Focus on Alternatives. Submission to the
Expert Working Group studying the
TGN1412 incident. Appendi x to Bhogal N
and Combes R . An update on TGN1412.
ATLA 34, 351–356 (2006).
9 Deeken JF, Figg WD, Bates SE,
Sparreboom A. Toward individualized
treatment: prediction of anticancer drug
disposition and toxicity with
pharmacogenetics. Anticancer Drugs 18,
111–126 (2007).
10 Sparreboom A . Unexplored pharmacokinetic
opportunities wit h microdosing in oncology.
Clin. Cancer Res. 13, 4033–4034 (2007).
11 Coecke S, Blaauboer BJ, Elaut G et al.
Toxicokinetics and metabolism. In:
Alternative (Nonanimal) Methods for
Cosmetics Testing: Current Status and Future
Prospects. A Report Prepared in the Context of
the 7th Amendment of the Cosmetics Directive.
ATLA. 33(Suppl. 1), 147–175 (2005).
12 Langley G, Evans T, Holgate ST, Jones A.
Replacing anima l experiments: choices,
chances and challenges. Bioessays 29, 918–926
13 Anonymous. Reduction of the use of
animals in the development and control of
biological products. Lancet 2, 900 –902
14 Lappin G, Garner C. The utility of
microdosing over the past 5 years. Exp. Opin.
Drug Metab. Toxicol. 4, 1499–1506 (2008).
101 US FDA, US Department of Health and
Human Services. Challenge and Opportunity
on the Critical Path to New Medical Products.
SpecialTopics/CriticalPathInitiative /
CriticalPathOpportunitiesReports /
102 Tracking system for a lternative test methods
review, validation and approval in the context
of EU regulations on chemicals
http: //
103 Europea n Medicines Agency. Note for
guida nce on non-clinica l safety studies for the
conduct of human clinical trials and
marketing authorization for pharmaceuticals
(CPMP/ICH/286/95) (2009)
104 Expert Scientific Group on Phase One
Clinical Trials, Final Report, 30 November
2006. The Stationery Office, London, UK,
(2006 )
105 The Government Reply to the Report of the
House of Lords Select Committee on Animals
in Scientific Procedures. Session 2001–2002.
Presented to Parliament by the Secretary of
State for the Home Department. 4, 7 January
2003 (Cm. 5729)
106 Europea n Partnership for A lternative
Approaches to Animal Testing
http: //
107 NIH. Countries unite to reduce animal use in
product toxicity testing worldwide. News
release. 27 April 2009
... There have been efforts by the European Bioanalysis Forum (EBF) and others encouraging the use of surrogate matrix to advance the 3R (replacement, reduction, refinement) approach to minimize the use of animals, without compromising scientific integrity (9)(10)(11)(12). These 3R strategies can be directly applied to mitigate the current matrix shortage caused by the COVID-19 pandemic. ...
The COVID-19 pandemic has strained the biological matrix supply chain. An upsurge in demand driven by numerous COVID-19 therapeutic and vaccine development programs to combat the pandemic, along with logistical challenges sourcing and transporting matrix, has led to increased lead times for multiple matrices. Biological matrix shortages can potentially cause significant delays in drug development programs across the pharmaceutical and biotechnology industry. Given the current circumstances, discussion is warranted around what will likely be increased use of surrogate matrices in support of pharmacokinetic (PK), immunogenicity, and biomarker assays for regulatory filings. Regulatory authorities permit the use of surrogate matrix in bioanalytical methods in instances where matrix is rare or difficult to obtain, as long as the surrogate is appropriately selected and scientifically justified. Herein, the scientific justification and possible regulatory implications of employing surrogate matrix in PK, immunogenicity, and biomarker assays are discussed. In addition, the unique challenges that cell and gene therapy (C&GT) and other innovative therapeutic modalities place on matrix supply chains are outlined. Matrix suppliers and contract research organizations (CROs) are actively implementing mitigation strategies to alleviate the current strain on the matrix supply chain and better prepare the industry for any future unexpected strains. To maintain ethical standards, these mitigation strategies include projecting matrix needs with suppliers at least 6 months in advance and writing or updating study protocols to allow for additional matrix draws from study subjects and/or re-purposing of subject matrix from one drug development program to another.
... To prepare the delegates, a survey of seven questions, all with the theme of sustainability in bioanalysis, was sent out a few weeks prior to the event. As part of this survey, the delegates were invited to also read reference papers for inspiration on the subjects [6][7][8][9][10]. ...
The 6th Young Scientist Symposium, a meeting organized by young scientists for young scientists under the umbrella of the European Bioanalysis Forum vzw and in collaboration with the Universities of Bologna and Ghent, included a variety of interesting presentations on cutting edge bioanalytical science and processes. Integrated in the meeting, an interactive round table session, the Science Café, discussed the challenges related to sustainability for bioanalytical lab activities. This manuscript reflects conclusions from these discussions. They can provide our community a compass for future business practices to embrace more sustainable laboratory activities considerate of smarter use of a wide array of resources and laboratory tools, resulting in increased wellbeing for our next generations and our planet.
Animal models have long served as a basis for scientific experimentation, biomedical research, drug development and testing, disease modelling and toxicity studies, as they are widely thought to provide meaningful, human-relevant predictions. However, many of these systems are resource intensive and time-consuming, have low predictive value and are associated with great social and ethical dilemmas. Often drugs appear to be effective and safe in these classical animal models, but later prove to be ineffective and/or unsafe in clinical trials. These issues have paved the way for a paradigm shift from the use of in vivo approaches, toward the ‘science of alternatives’. This has fuelled several research and regulatory initiatives, including the ban on the testing of cosmetics on animals. The new paradigm has been shifted toward increasing the relevance of the models for human predictivity and translational efficacy, and this has resulted in the recent development of many new methodologies, from 3-D bio-organoids to bioengineered ‘human-on-a-chip’ models. These improvements have the potential to significantly advance medical research globally. This paper offers a stance on the existing strategies and practices that utilise alternatives to animals, and outlines progress on the incorporation of these models into basic and applied research and education, specifically in India. It also seeks to provide a strategic roadmap to streamline the future directions for the country’s policy changes and investments. This strategic roadmap could be a useful resource to guide research institutions, industries, regulatory agencies, contract research organisations and other stakeholders in transitioning toward modern approaches to safety and risk assessment that could replace or reduce the use of animals without compromising the safety of humans or the environment.
Full-text available
The publication of the U.S. Food and Drug Administration's “Critical Path” document highlights the importance of translational research and the development of new concepts and tools that may help provide more confidence in the selection of drug candidates early in clinical development.[1][1] One
Microdosing studies (human Phase 0) are used to select drug candidates for Phase I clinical trials on the basis of their pharmacokinetic properties, using subpharmacologic doses (maximum 100 microg). There are questions as to whether pharmacokinetic data obtained at these low doses will predict those at the clinically relevant dose. To review the current literature on microdosing and assess how well microdose data have predicted the pharmacokinetics obtained at a therapeutic dose. All data published in the peer reviewed literature comparing pharmacokinetics at a microdose with a therapeutic dose were reviewed, excluding those studies aimed at imaging. Of the 18 drugs reported, 15 demonstrated linear pharmacokinetics within a factor of 2 between a microdose and a therapeutic dose. Therefore, data that support the utility of microdosing are beginning to emerge.
Testing the safety and efficacy of a successful human medicine involves many laboratory animals, which can sometimes be subjected to considerable suffering and distress. Also, it is necessary to extrapolate from the test species to humans. UK and European legislation requires that Replacement, Reduction and Refinement of animal procedures (the Three Rs) are implemented wherever possible. Over the last decade, there has been substantial progress with applying in vitro and in silico methods to both drug efficacy and safety testing. This paper is a report of the discussions and recommendations arising from a workshop on the role that might be played by human volunteer studies in the very early stages of drug development. The workshop was organised in November, 2001 by Volunteers in Research and Testing, a group of individuals in the UK which launched an initiative in 1994 to identify where and how human volunteers can participate safely in biomedical studies to replace laboratory animals. It was considered that conducting pre-Phase I very low dose human studies (sub-toxic and below the dose threshold for measurable pharmacological or clinical activity) could enable drug candidates to be assessed earlier for in vivo human pharmacokinetics and metabolism. Moreover, accelerator mass spectrometry (AMS), nuclear magnetic resonance (NMR) spectroscopy and positron emission tomography (PET) are potentially useful spectrometric and imaging methods that can be used in conjunction with such human studies. Some, limited animal tests would still be required before pre-Phase I microdose studies, to take account of the potential risk posed by completely novel chemicals. The workshop recommended that very early volunteer studies using microdoses should be introduced into the drug development process in a way that does not compromise volunteer safety or the scientific quality of the resulting safety data. This should improve the selection of drug candidates and also reduce the likelihood of later candidate failure, by providing in vivo human ADME data, especially for pharmacokinetics and metabolism, at an earlier stage in drug development than is currently the case.
A great deal of effort has been spent in defining the pharmacokinetics and pharmacodynamics of investigational and registered anticancer agents. Often, there is a marked variability in drug handling between individual patients, which contributes to variability in the pharmacodynamic effects of a given dose of a drug. A combination of physiological variables, genetic characteristics (pharmacogenetics) and environmental factors is known to alter the relationship between the absolute dose and the concentration-time profile in plasma. A variety of strategies are now being evaluated in patients with cancer to improve the therapeutic index of anticancer drugs by implementation of pharmacogenetic imprinting through genotyping or phenotyping individual patients. The efforts have mainly focused on variants in genes encoding the drug-metabolizing enzymes thiopurine S-methyltransferase, dihydropyrimidine dehydrogenase, members of the cytochrome P450 family, including the CYP2B, 2C, 2D and 3A subfamilies, members of the UDP glucuronosyltransferase family, as well as the ATP-binding cassette transporters ABCB1 (P-glycoprotein) and ABCG2 (breast cancer resistance protein). Several of these genotyping strategies have been shown to have substantial impact on therapeutic outcome and should eventually lead to improved anticancer chemotherapy.
Replacing animal procedures with methods such as cells and tissues in vitro, volunteer studies, physicochemical techniques and computer modelling, is driven by legislative, scientific and moral imperatives. Non-animal approaches are now considered as advanced methods that can overcome many of the limitations of animal experiments. In testing medicines and chemicals, in vitro assays have spared hundreds of thousands of animals. In contrast, academic animal use continues to rise and the concept of replacement seems less well accepted in university research. Even so, some animal procedures have been replaced in neurological, reproductive and dentistry research and progress is being made in fields such as respiratory illnesses, pain and sepsis. Systematic reviews of the transferability of animal data to the clinical setting may encourage a fresh look for novel non-animal methods and, as mainstream funding becomes available, more advances in replacement are expected.
13 Anonymous Reduction of the use of animals in the development and control of biological products
13 Anonymous. Reduction of the use of animals in the development and control of biological products. Lancet 2, 900–902 (1985).
A revolution in R&D - how genomics and genetics will affect drug development costs and times
  • Boston Consulting Group
Boston Consulting Group. A revolution in R&D – how genomics and genetics will affect drug development costs and times. In: Parexel Pharmaceutical R&D Statistical Sourcebook. (2002/2003).
Submission to the Expert Working Group studying the TGN1412 incident. Appendix to Bhogal N and Combes R. An update on TGN1412
  • Alternatives Focus On
Focus on Alternatives. Submission to the Expert Working Group studying the TGN1412 incident. Appendix to Bhogal N and Combes R. An update on TGN1412. ATLA 34, 351–356 (2006).
675 final. Fifth Report on the Statistics on the Number of Animals used for Experimental and other Scientific Purposes in the Member States of the European Union {SEC
COM(2007) 675 final. Fifth Report on the Statistics on the Number of Animals used for Experimental and other Scientific Purposes in the Member States of the European Union {SEC(2007)1455}.