CRANNME - The complete removal of animal use for neuromuscular effectors testing

Major advances in the care of pregnant women and children, public health campaigns, safer environments and the introduction of advanced surgical and medical techniques has led to the population of many global nations living longer. The downside of this is that conditions that tend to manifest later in life are creating many emotional and economic difficulties. Amongst these conditions, the shadow of neurological disease looms large. Conditions include motor neuron disease, most well known through the life of Professor Stephen Hawking and nerve injury and damage, as suffered by the Hollywood actor Christopher Reeve. In addition, neuromuscular blocking agents are used in a variety of situations (e.g. muscle relaxants during surgery) and are always in development. Therefore, although the research effort on this area is very high, a large proportion of this research occurs using animal models. Rodents are a major resource however there is an emerging tendency to return to primate use due to concerns over species specificity.

So there is a clear need to develop interventions to treat or prevent such conditions as mentioned above. The major testing models are animals however this type of experimentation is not amenable to wide-scale screening and is subject to various issues of expense and regulation e.g. how can multiple agents be screened? What about combinations of pharmacological and non-pharmacological therapies e.g. exercise?

Much of the research interest targets the interface between your muscles and your nerves and it is the complexity of this "junction" that has allowed animal-base research to persist. Through previous support by the NC3Rs, our group have sought to "grow muscles in a tube" and to try and recreate that link between these muscles and the nerve system. We can now "grow" human muscles and need a way to get hold of human nerve cells. Using the latest technology in stem cells (a single type of cells that can be used, theoretically, to produce all of the cell types (e.g. muscle, nerve, bone, liver, etc.) in the human body. NC3Rs has also, but independently to the muscle work, sponsored research in this area concerned with creation of motor neurons from stem cells.

There really could be no better time to combine these two areas in order to see if we can create nerve-muscle interfaces that can reduce or even replace the use of animals in this area and that can speed progress in the discovery of treatments for various disorders.

The proposal described herein represents the synergistic opportunity between two currently independently funded NC3Rs grants. Both of these NC3Rs funded projects have shown independent impact progress and we now propose to synergistically exploit this progress to refine a test bed that will completely replace animal testing for neuromuscular effectors. Using techniques and approaches from the world of tissue engineering, we have successfully created skeletal muscle in the laboratory that can be stimulated in a variety of different ways - human constructs have also been created. We have also been able to co-culture these constructs with motor neurons and have increasing evidence of interactions between the cell types. One of the ambitions for this work was to develop "...a situation where appropriate cell lines could be developed that would remove the need for animal tissues altogether; this proposal does not explicitly seek to achieve that aim although it could be a subject of future grants". The limit to this ambition was the fact that the motor neuron component required animal tissues but, through NC3Rs networking sessions, we have identified a fellow grant-holder who has been funded to generate neuronal cells from human stem cells. The current project therefore is a direct continuation of both projects where the use for animals is TOTALLY REMOVED.

The 3Rs impacts are we believe that this potential follow-on grant from two other NC3Rs funded projects moves beyond the REFINEMENT and REDUCTION arguments of the previous work and really gets to the heart of the REPLACEMENT agenda. As we describe in the Case for Support, the currently supported LEWIS work has looked to develop a novel way to investigate neuromuscular disease using cell-based, tissue engineered biomimetic models. The ultimate end-point is to REPLACE animal usage in this area however animal cells removed from organs where the animal was killed by a humane technique before collection of the material have been used up to now. This is especially true for the motor neurons although the muscle component has been completed with human-derived material. The NC3Rs funded work for Hill solves this issue - motor neurons derived from human embryonic stem cells becomes a reality. This can clearly lead to REPLACEMENT and if the nerve-muscle connections created with this combination are fully functional then they can be used to replace animals used in neural toxicity testing, research into neuromuscular diseases and nerve regeneration and in drug screening. If our grant is successful then the impact on other users should be high including both academic researchers (including our own laboratories and those of our collaborators) and industry. The "buy-in" required for wider take-up will certainly entail work but our communications strategy should help there. Economic and societal impacts will likely be in the generation and manufacture of animal-sparing 3D test kits, associated reagents, measuring instruments, training etc. Our group has expertise in this area as detailed in the Case for Support. Not only could the impact be made in manufacturing industries, it could also expand into other agencies that would use such kits e.g. Pharmaceuticals. There there would be impacts in terms of cost, quality and removal of ethical issues. Healthcare impacts could be found in the use of such test kits screening and diagnosis of disease. Impacts into public policy would be in understanding by the public of the reduction of use in animal models as well as the communication of the usefulness of the models/kits developed. This will interface with our stated communications plans. Academic impact will be demonstrate through the normal academic pathways such as publication in scholarly journals, conference attendances, networking meetings, one-to-one meetings with individual academic colleagues etc.