Collaboration on the road to better preclinical research

In this blog post Adrian Smith from Norecopa discusses the role that the PREPARE Guidelines and Website play in improving the robustness and translatability of animal studies.

Criticism of animal research has come from an unexpected quarter in recent years: scientists themselves have expressed concerns about the poor reproducibility and translatability of preclinical studies [1,2]. The criticism includes, among other things, concerns about underpowered experiments, bias caused by a lack of randomisation and blinding, and incorrect use of statistical analyses in attempts to demonstrate significance.

Working inside animal facilities has given me insight into some additional causes of this state of play. From that viewpoint, it quickly becomes clear that high-quality animal research is utterly dependent upon collaboration, from the earliest possible stage, between scientists and animal care staff. While scientists have the insight and expertise to plan relevant experiments, they rely on those with intimate knowledge of the animals to translate these plans into valid and robust in vivo studies. Even simple events such as a scientist’s request for a blood sample trigger questions about a range of issues, including factors that affect the quality and shelf-life of the sample, and the physiological effects of the blood loss on the animal [3]. Similar practical questions arise for every aspect of a study, many of which will never be reported in the publications ensuing from the work. These include assessment of the facility’s standard and competence, staffing levels, refinement and standardisation of procedures, distribution of costs, and health and safety concerns. Early dialogue with animal care staff, who are often proficient at lateral thinking, may reveal better ways of conducting a procedure, thanks to their previous experience. If these questions are inadequately addressed, we risk that animal-related issues become the greatest sources of variability or poor validity in a study. Mutual respect for the skills and knowledge of scientists and lab animal staff alike is therefore paramount if we are to improve reproducibility and translatability.

Although better reporting is often promoted in connection with the reproducibility crisis, this is only part of the solution. There is no doubt that improved reporting is needed, both to aid evaluation of the quality of experiments and to enable them to be repeated. Inadequacies that have been demonstrated include faulty study design, poor descriptions of the animals, and insufficient detail about their environment and peri-operative care [4-6]. Reporting guidelines have been published for many years to address these issues [7], but without better planning we are merely trying to improve the description of a burnt cake: we need to go back to the kitchen and change the recipe. Lack of compliance with reporting guidelines, or even knowledge of them, despite journal endorsement, is also a major problem [8-9].

In 2016, the EU Commission was faced with a European Citizen’s Initiative (ECI) to ban animal experimentation [10]. Public opinion plays an important role in defining animal research, and we have both legal and ethical obligations not to waste animal lives. Part of the Commission’s response to the ECI was to hold a meeting entitled Non-Animal Approaches – The Way Forward in Brussels in December 2016 [11]. Here, almost for the first time, I heard participants discussing the need for planning guidelines. Spurred on by this, we published the PREPARE guidelines in 2017 [12]. PREPARE consists of a checklist covering 15 main areas (Figure 1) and a website with supplementary information and references to resources for each of the 40 topics on the checklist (norecopa.no/PREPARE).

PREPARE guides scientists through all the steps of planning animal experiments. The principles embodied in PREPARE apply to all in vivo studies, both in dedicated facilities and in the field, regardless of species used and the level of their legal protection. The PREPARE checklist is divided into three sections: formulation of the study, dialogue between the scientists and animal care staff, and quality control of the components in the study. Some of these topics will be the responsibility of the animal facility rather than of the research team, but it is important that all have been considered. The first section gives advice on literature searching (including systematic reviews), legal aspects, harm-benefit assessment, humane endpoints and other ethical issues, experimental design and statistical analysis. The second section covers important practicalities such as the distribution of labour and responsibilities, facility evaluation and competence, as well as health and safety issues. The third section provides tips on quality control of all the stages of a preclinical study, including the test substances, procedures, the animals and their environment, husbandry methods, humane killing and necropsy routines. PREPARE aims to give practical advice which will ensure both scientific rigour and optimal animal welfare.

Although only recently published, the principles in PREPARE were developed over a 20-year period, during which earlier versions were discussed with scientists on courses in Laboratory Animal Science. PREPARE is also full of lessons learned by animal facilities when applying for international accreditation. The checklist has proved popular, it has been translated into over 20 languages, and the website is updated regularly as new resources are published. A 3-minute cartoon film, with optional subtitles in many languages, has been produced to illustrate the main principles embodied in PREPARE (norecopa.no/PREPARE/film).

We are now working to encourage uptake of PREPARE by scientists on a voluntary basis. This is being done by arranging workshops and webinars, through newsletters and social media, and by contacting research animal facilities. PREPARE is not intended to be yet another hurdle on the road to publication. On the contrary, it is our hope that PREPARE will be seen as a means of ensuring that all the issues likely to be raised by reviewers will have been addressed before it is too late. PREPARE should also prove helpful to those who evaluate proposals for animal studies, including funding bodies, ethical review boards and regulatory authorities.

To complete this cycle of increased quality, I would also like to see more emphasis in scientific papers on the efforts the authors have made to replace, reduce or refine animal use (“the three Rs”) [13]. This emphasis is unfortunately not a clear feature of reporting guidelines. We owe it to the general public, to our funders and to the animals. We must do this prominently, since many bibliographic databases index only the title and abstract of a paper. My 3-step recipe for better science is therefore:

1. Be PREPAREd: plan in collaboration with animal care staff from day 1
2. Demonstrate that you have ARRIVEd: submit a manuscript which documents how the potential causes of irreproducibility have been tackled
3. Flag the 3Rs: highlight efforts to refine, reduce or replace animal use

I am optimistic. In the words of Edward Everett Hale [14]:

Coming together is a beginning, Keeping together is progress, Working together is success.

About the Author

Adrian Smith studied Veterinary Medicine at Cambridge University, graduating in 1979. After working in clinical veterinary practice in the UK, he emigrated to Norway and held positions related to animal research at the Norwegian School of Veterinary Science for 30 years. He was awarded his PhD in 1988 for his work on seasonal variations in the reproductive activity of the Arctic fox (Vulpes lagopus). He held the Chair in Laboratory Animal Science from 1988 to 2011, and led the work of obtaining and renewing international accreditation of the School’s laboratory animal facilities. During this period he also arranged over 50 courses in Laboratory Animal Science for scientists and technicians, and served on the Norwegian Animal Research Authority, during which time he was involved in writing the draft of new national legislation on animal research.
   Adrian has been Secretary of the Norwegian platform for the replacement, reduction and refinement of animal experiments (Norecopa) since its foundation in 2007. He has led the work of developing Norecopa’s website (norecopa.no) into a source of international resources for scientists and the laboratory animal community. The website currently has over 9,000 pages and a quarter of a million hits annually. He led the work of publishing the PREPARE guidelines for planning animal experiments. Collaboration with international colleagues is an important part of this work, and the PREPARE checklist has been translated into over 20 languages. Other tasks for Norecopa have included the organisation of international consensus meetings on the care and use of animals in research, the production of position statements and guidelines about animal procedures, and the regular issue of newsletters in English.

References

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2. Begley CG, Ioannidis JP (2015) Reproducibility in science: improving the standard for basic and preclinical research. Circ Res. 116(1):116-126. doi:10.1161/CIRCRESAHA.114.303819
3. Blood sampling. https://www.nc3rs.org.uk/3rs-resources/blood-sampling Accessed 30 September 2020
4. Baker, M (2016) 1,500 scientists lift the lid on reproducibility. Nature 533:452–454. doi:10.1038/533452a
5. Avey MT, Moher D, Sullivan KJ, Fergusson D, Griffin G, Grimshaw JM, Hutton B, Lalu MM, Macleod M, Marshall J, Mei SHJ, Rudnicki M, Stewart DJ, Turgeon AF, McIntyre L (2016) The Devil Is in the Details: Incomplete Reporting in Preclinical Animal Research. PLoSONE 11:e0166733. doi:doi:10.1371/journal.pone.0166733
6. Bradbury AG, Eddleston M, Clutton RE (2016): Pain management in pigs undergoing experimental surgery; a literature review (2012-4). Br. J. Anaesth. 116: 37-45. https://doi.org/10.1093/bja/aev301
7. Norecopa (2020) Reporting guidelines. https://norecopa.no/more-resources/reporting-guidelines Accessed 30 September 2020
8. Reichlin TS, Vogt L, Wurbel H (2016): The Researchers’ View of Scientific Rigor-Survey on the Conduct and Reporting of In Vivo Research. PLoS One 11: e0165999. 2016/12/03. DOI: 10.1371/journal.pone.0165999.
9. Percie du Sert N, Hurst V, Ahluwalia A, Alam S, Avey MT, Baker M, Browne WJ, Clark A, Cuthill IC, Dirnagl U, Emerson M, Garner P, Holgate ST, Howells DW, Karp NA, Lazic SE, Lidster K, MacCallum CJ, Macleod M, Pearl EJ, Petersen O, Rawle F, Peynolds P, Rooney K, Sena ES, Silberberg SD, Steckler T, Wurbel H (2020): The ARRIVE guidelines 2.0: updated guidelines for reporting animal research. PLoS Biol. 18(7): e3000410. doi: 10.1371/journal.pbio.3000410
10. Stop Vivisection. European Citizens’ Initiative. https://europa.eu/citizens-initiative/stop-vivisection_en Accessed 30 September 2020
11. Non-Animal Approaches – the Way Forward. Report on a European Commission Scientific Conference held on 6-7 December at The Egg, Brussels, Belgium. ISBN 978-92-79-65840-2. doi: 10.2779/373944 https://ec.europa.eu/environment/chemicals/lab_animals/3r/pdf/scientific_conference/non_animal_approaches_conference_report.pdf Accessed 30 September 2020
12. Smith AJ, Clutton RE, Lilley E, Hansen KEAa, Brattelid T (2018) PREPARE: Guidelines for planning animal research and testing. Lab. Anim. 52(2): 135-141. doi: 10.1177/0023677217724823 Accessed 30 September 2020
13. Norecopa (2020) The Three R’s. https://norecopa.no/alternatives/the-three-rs Accessed 30 September 2020
14. Edward Everett Hale. https://en.wikiquote.org/wiki/Edward_Everett_Hale Accessed 30 September 2020

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