The History of Alternative Test Methods in Toxicology

• Publisher: Academic Press
• Year: 2018
• ISBN:
  9780128136980
• Topic: Medicina
• Subtopic: Tossicologia

The History of Alternative Test Methods in Toxicology uses a chronological approach to demonstrate how the use of alternative methods has evolved from their conception as adjuncts to traditional animal toxicity tests to replacements for them. This volume in the History of Toxicology and Environmental Health series explores the history of alternative test development, validation, and use, with an emphasis on humanity and good science, in line with the Three Rs (Replacement, Reduction, Refinement) concept expounded by William Russell and Rex Burch in 1959 in their now classic volume, The Principles of Humane Experimental Technique.

The book describes the historical development of technologies that have influenced the application of alternatives in toxicology and safety testing. These range from single cell monocultures to sophisticated, miniaturised and microfluidic organism-on-a-chip devices, and also include molecular modelling, chemoinformatics and QSAR analysis, and the use of stem cells, tissue engineering and hollow fibre bioreactors. This has been facilitated by the wider availability of human tissues, advances in tissue culture, analytical and diagnostic methods, increases in computational processing, capabilities, and a greater understanding of cell biology and molecular mechanisms of toxicity.

These technological developments have enhanced the range and information content of the toxicity endpoints detected, and therefore the relevance of test systems and data interpretation, while new techniques for non-invasive diagnostic imaging and high resolution detection methods have permitted an increased role for human studies. Several key examples of how these technologies are being harnessed to meet 21st century safety assessment challenges are provided, including their deployment in integrated testing schemes in conjunction with kinetic modelling, and in specialized areas, such as inhalation toxicity studies.

The History of Alternative Test Methods in Toxicology uses a chronological approach to demonstrate how the use of alternative methods has evolved from their conception as adjuncts to traditional animal toxicity tests to replacements for them. This volume in the History of Toxicology and Environmental Health series explores the history of alternative test development, validation, and use, with an emphasis on humanity and good science, in line with the Three Rs (Replacement, Reduction, Refinement) concept expounded by William Russell and Rex Burch in 1959 in their now-classic volume, The Principles of Humane Experimental Technique.

The book describes the historical development of technologies that have influenced the application of alternatives in toxicology and safety testing. These range from single cell monocultures to sophisticated miniaturised and microfluidic organism-on-a-chip devices, and also include molecular modelling, chemoinformatics and QSAR analysis, and the use of stem cells, tissue engineering and hollow fibre bioreactors. This has been facilitated by the wider availability of human tissues, advances in tissue culture, analytical and diagnostic methods, increases in computational processing capabilities, and a greater understanding of cell biology and molecular mechanisms of toxicity.

These technological developments have enhanced the range and information content of the toxicity endpoints detected, and therefore the relevance of test systems and data interpretation, while new techniques for non-invasive diagnostic imaging and high resolution detection methods have permitted an increased role for human studies. Several key examples of how these technologies are being harnessed to meet 21st century safety assessment challenges are provided, including their deployment in integrated testing schemes in conjunction with kinetic modelling, and in specialised areas, such as inhalation toxicity studies.

Section 1

Setting the Scene

Outline

Chapter 1.1. The Introduction and Influence of the Concept of Humane Experimental Technique

Chapter 1.2. Types of Toxicity and Applications of Toxicity Testing

Chapter 1.3. The Key Technologies

Chapter 1.1

The Introduction and Influence of the Concept of Humane Experimental Technique

Michael Balls University of Nottingham, Nottingham, United Kingdom

Abstract

Since its publication in 1959, The Principles of Humane Experimental Technique, by Russell and Burch, which introduced the concept of the Three Rs (Reduction, Refinement and Replacement), has had a profound effect on attitudes, laws and practices related to the use of laboratory animals in education, research and testing. The search for more-relevant and more-reliable non-animal alternative procedures for predicting the potential hazards to humans represented by chemicals and cosmetics, pharmaceuticals and other products gained momentum in the 1970s, initially led by the Fund for the Replacement of Animals in Medical Experiments (FRAME) in the UK, later joined, in particular, by the Center for Alternatives to Animal Testing (CAAT) in the USA, the Center for Documentation and Evaluation of Alternative Methods to Animal Experiments (ZEBET) in Germany and the European Centre for the Validation of Alternative Methods (ECVAM) in Italy. The replacement of animal tests by alternative procedures and testing strategies is now the focus of scientific, political and administrative effort throughout the world.

Keywords

Alternatives; Animal experimentation; Directive 86/609/EEC; FRAME; High-fidelity fallacy; Human technique; Inhumanity; Replacement; Three Rs

Summary

Since its publication in 1959, The Principles of Humane Experimental Technique, by Russell and Burch, which introduced the concept of the Three Rs (Reduction, Refinement and Replacement), has had a profound effect on attitudes, laws and practices related to the use of laboratory animals in education, research and testing. The search for more-relevant and more-reliable non-animal alternative procedures for predicting the potential hazards to humans represented by chemicals and cosmetics, pharmaceuticals and other products gained momentum in the 1970s, initially led by the Fund for the Replacement of Animals in Medical Experiments (FRAME) in the UK, later joined, in particular, by the Center for Alternatives to Animal Testing (CAAT) in the USA, the Center for Documentation and Evaluation of Alternative Methods to Animal Experiments (ZEBET) in Germany and the European Centre for the Validation of Alternative Methods (ECVAM) in Italy. The replacement of animal tests by alternative procedures and testing strategies is now the focus of scientific, political and administrative effort throughout the world.

1. Inhumanity and the Three Rs Concept

The use of animals as surrogates for humans in potentially painful experiments had been the subject of controversy and unresolved conflict for hundreds of years before William Russell and Rex Burch proposed a way forward in 1959, in their outstanding book, The Principles of Humane Experimental Technique (The Principles; (1)). Much of what they said is no less applicable today than it was almost 60 years ago, but, as has been discussed in a series of articles in ATLA, the wisdom of Russell and Burch has all too often remained unrecognised or been ignored (see Ref. (2)).

Their underlying philosophy concerned the concept, sources and incidence of inhumanity and its diminution or removal through implementation of the Three Rs, Reduction, Refinement and Replacement, as a way of promoting humanity ‘without prejudice to scientific and medical aims’. They distinguished between direct inhumanity, ‘the infliction of distress as an unavoidable consequence of the procedure employed’, and contingent inhumanity, ‘the infliction of distress as an incidental and inadvertent by-product of the use of the procedure, which is not necessary for its success’. Inescapably, toxicity testing in animals involves direct inhumanity, and the way to avoid it is replacement, i.e. ‘any technique employing non-sentient material’ to ‘replace methods which use conscious living vertebrates’.

There are two main reasons for seeking to replace toxicity tests on animals: the insuperable inadequacy of animals as models for humans and the advantages afforded by the use of scientifically advanced, non-animal replacement tests and testing strategies.

As Russell and Burch pointed out, models must differ from what is being modelled, and the consequences of this difference depend on two major factors, fidelity (a high general similarity to what is being modelled) and discrimination (a high specificity to what is being modelled). They argued that a highly discriminative/poor fidelity model is more useful than a high-fidelity/poorly discriminative model, and warned of the dangers of the high-fidelity fallacy. This can be described as follows: humans are placental mammals, so members of other mammalian species are more likely to be useful as models of man that would be fish, or still more markedly, bacteria, nematodes or insects. However, the stubborn assumption that laboratory mammals are the best models for use in fundamental biomedical research, drug development or toxicity testing, where humans are the focus of concern, fails to recognise that the phenomena under investigation are not sufficiently well understood in the models or in humans (i.e. that what may be happening in one black box is used to predict what might happen in another black box), that there are fundamental differences between animals and humans (species differences) and that there are major differences within animal species and in humans (intra-species variation).

The second reason for seeking replacement is that although the non-animal procedures have limitations, which must be taken into account, they can be used to provide information of more-direct relevance to human situations. They can offer advantages of scale (number of replicates), speed and manageability, more-tightly defined and more-mechanistic approaches, talking advantage of progress in cell and molecular biology and the use of human material, from ‘normal’ controls or individuals with particular characteristics, susceptibilities or disease histories.

Replacement can be categorised as partial, where an animal is exposed to chemicals, then killed and its cells and tissues are subjected to further tests (ex vivo), or total, where all the experimental procedures are applied to cells and tissues in vitro. However, more useful is the distinction between direct and indirect replacement. An example of direct replacement is the application of potential irritants to isolated rabbit eyes instead of to the eyes of intact rabbits where the in vitro approach still has the disadvantages of the in vivo one. Seeking genuine indirect replacement is much more intelligent – it involves defining the information it was hoped to get from the animal, then obtaining it from a different experimental approach, e.g. testing drugs on human hepatocytes, instead of administering them to rats or dogs, which have different complements of drug metabolising enzymes. Direct replacement offers the possibility of direct relevance.

Russell and Burch recognised toxicity testing as a particular problem, ‘since this is one use [of animals] which is an urgent humanitarian problem, both numerically and in terms of severity, for it regularly involves a finite and large incidence of distress, which is often considerable and sometimes acute’. The problem that other mammals may only be superficially similar to humans ‘is met in practice by erring on the side of caution, and by using more than one mammalian species; it cannot be fully met, for there may always be metabolic peculiarities specific to man’. They also referred to the fact that a very large number of substances are toxic at high doses, so ‘the important concept is the therapeutic index of a drug, i.e. the ratio between its toxic doses and its therapeutically effective dosage: if this ratio is great, the use of the drug or preparation is prudent, since it allows for wide variations between human individual patients to sensitivity to the toxic effects’.

2. Progress Following the Publication of The Principles

An interesting chapter in The Principles deals with the factors governing progress. Russell and Burch divided them into individual personality factors, which they further subdivided into authoritarian and revolutionary factors, and sociological factors, which involved achieving a balance between efficiency or yield of information and humanity. They also warned against rationalisation, a mechanism of defence by which unacceptable actions are given acceptable reasons to justify them. Even today, despite all that happened since the publication of The Principles, there remains a struggle between authoritarian factors backed by rationalisation and revolutionary and sociological factors.

In the 1960s, very little attention was paid to the Three Rs concept, and Russell and Burch had gone off to further their own careers in different ways (3). However, in 1969, Dorothy Hegarty and her family and friends set up the Fund for the Replacement of Animals in Medical Experiments (FRAME), as a parallel development to the Three Rs concept, of which they seemed to be unaware (4). FRAME's aim was to benefit both man and animals by promoting ‘the use of more reliable, accurate and relevant methods in biology and medicine which at the same time help to reduce the need for laboratory animals’ and, in particular, by recognising that ‘the exchange of information, and its ready availability, are or paramount importance, if new techniques are to be adopted rapidly’. To achieve this, in 1973, they introduced ATLA Abstracts, containing abstracts of relevant articles, selected and written by staff located at the British Library, Boston Spa, Wetherby, Yorkshire, and arranged in 17 subject sections. This way of informing scientists may look rather naïve today, but it should not be forgotten that, at that time, there were few abstracting services, no personal computers, no Internet or websites and limited access to electric typewriters and photocopiers. Scientists kept themselves informed by going to the library and looking through hard copies of the scientific journals.

In 1976, Andrew Rowan arrived at FRAME, and during his short time as Scientific Director, he revolutionised the work of the charity. He increased the non-abstract coverage of the journal through news, reviews...