When talking about animal testing, there is usually only one point where people agree: that it is a controversial subject. It is often condemned for being cruel and “unnecessary”, and that humans have no right to abuse animals for their own good. Often it is said in vitro testing could already replace animal studies. But has animal testing really become obsolete?
Animal Testing in Drug Development
Despite the usually common consensus that “animal testing is cruel”, hardly anyone living in an industrialized country has never in their life taken any drugs – most popular probably pain killers. Probably only few people give that little pill they swallow a second thought: Where does a drug come from? How is it developed? Drug development is a stringently controlled process from the discovery of a promising substance to the packed and ready-to-use product in the end.
As seen in the process chart (click to enlarge), the pre-clinical in vivo testing (animal testing) is a bridge between in vitro testing and testing directly in humans. The key questions answered in this phase of testing address the Pharmacodynamics and Pharmacokinetics of the potential drug candidate.
Pharmacokinetics describes the “fate” of the substance in within an organism. Pharmacodynamics describe the properties of a drug substance with respect to “what it does to the body” and generally deals with the effect of the drug on the target organism, such as the the body or a microbe or parasite on or within the body.
Pharmacodynamics describes the mechanism in which the drug acts. In order to understand this, usually a set of tests are performed. The interaction of the drug with its target site, such as a ligand binding to a receptor, induce a so called primary response which in turn may cause a series of secondary responses. ( In order to distinguish and define these, drugs are commonly at first investigated at molecular, cellular and tissue level in vitro, and then animal tests are used to better understand how the drug acts in complex organisms.)
Another important topic of pharmacodynamic studies is the relationship between concentration and effect, which is commonly determined in toxicity studies. The general aim of pharmacodynamic studies is to describe the affinity, efficacy, potency and sensitivity with numerical values, such as NOAEL (no observed adverse effect level) or LD50/100 (absolute/median lethal dose). 1
All of the above mentioned can be observed on a small scale in cell and tissue culture, however ultimately questions such as for example organ-specific toxicity, lethal dose in relation to mass and chronic toxicity can only sufficiently be answered when testing in multicellular organisms – such as animals. Leaving out the animal tests in this process would mean to make a direct jump from in vitro to in vivo testing in humans i.e. being against animal testing would mean to be for testing in humans without any knowledge about safety of dosage and possible side effects due to secondary responses (e.g. from the immune system). From an ethical point of view, this is a very critical point.
Animal Models in Basic Research
But not only in drug development, also in basic research the role of the animal model is of great importance. Complex signaling pathways that affect multiple sites of the body can be analyzed in animal models, knock-out and knock-in models serve for example as disease models and help us to understand the effect of certain genes on the metabolism. Certain studies simply can – as of now – only rely on animal models, for example obesity studies, or research in the field of behavioral science. This article talks about an interesting outcome of a paraplegia study, where the researchers managed to get paralyzed rats to walk again. While it is clear that such studies would not work without the contribution of model animals, it is still a highly debated topic (e.g. the highly controversial research in humans’ relatives, the primates)
A knockout mouse (left) with a knocked-out gene affecting fur growth compared to a normal mouse (right). Picture via
The problematic of the current methods of toxicity testing
Sometimes it seems that the public image of a scientist describes him or her as “lunatic who likes to experiment on cute and fluffy little bunnies”. But it is in fact a major research goal of our time to find suitable alternatives to animal testing. Motivation is not only fueled by the ethical aspect of animal testing, but also by the fact that animal testing is time consuming and expensive (think about the handling and upkeep of the animals, which often require employment of special workers), so an efficient and reliable alternative is highly desired.
Furthermore, while being as close to a human organism as you can possibly get, even the most reliable model organism is simply not entirely the same as a human. Considering now the drug development process, potential drug candidates might get eliminated in an earlier phase of in vivo studies due to complications in the chosen test animal, that maybe would not have occurred in humans – whereas other substances might not show certain side effects in the model organism, that only occur later in human test subjects of clinical trials, rendering it unsuitable for the market. And the later a drug candidate gets eliminated from the development process, the more money has been “wasted” on it before. 2
It is obvious that a lot of arguments stand against animal testing, and a broad variety of alternatives is heavily researched in order to find novel methods to predict pharmacodynamics in vitro before clinical trials, circumventing the undesired animal testing. However, these novel in vitro cell culture models are still very restricted in their reliability. Considering that toxicity testing is a dynamic process that is often organ. or even species specific, that involves mechanisms of transport in and out of cells via transporters and diffusion, is influenced by enzymes and proteins of different tissues and the immune system and communication between cells, it becomes evident that a generalized or simplified in vivo model, such as 2D cell culture, is greatly oversimplified and not capable to supply sufficiently reliable information on toxicity in the whole body.2
Outlook: Critical Factors for in vitro Cell Culture Toxicity Models
While currently, animal models are the best we can do to predict how a drug substance will work in a body, different screening methods will ultimately replace these tests if we are able to close the gap between in vitro tissue and cell culture models and the human body.
In order to achieve this goal it is necessary to understand the molecular mechanisms involved in toxicity. A. Astashinka et al. put the problem down to a single sentence: “Most current methods assess cellular damage in cultured cell lines instead of trying to measure indicators that better inform toxicity in humans”. This is the key factor to help in vitro cell culture models to success – first, we have to understand the mechanisms of toxicity in order to reliably predict it. For example, organotypic cell culture models might help to bridge this gap, as toxicity testing in these cell cultures can give information about tissue toxicity, which can be then compared to known data from previous animal experiments to ultimately allow a better in vitro to in vivo comparison. So even when we find methods to replace animal testing, we first have to correlate the data we obtain form these methods to the data obtained in vivo.2
In the end, in vitro cell culture systems could greatly improve the quality, accuracy and specificity of toxicity testing, and efficiently enable us to translate the results of in vitro studies to clinical studies in humans. However, as for now, animal testing cannot be fully replaced without compromising the saftey of humans involved in clinical trials during pharmaceutical research, and cannot be replaced in basic research if we strive to understand the basic mechanisms of disease and life in general. Nevertheless, it goes without question that suitable alternatives for animal testing are highly desirable for a variety of reasons, such as cost efficiency, higher quality of data and of course the ethical aspect of the whole subject. It is therefore necessary to continue research in this field and improve the quality of in vitro testing, so that one day, animal testing does indeed become obsolete.
by Stephanie G.