Guidelines for preclinical animal research in ALS/MND: A consensus meeting

Figures & data

Table I.  Proposals to improve clinical understanding and impact of a preclinical study.

Develop principles equivalent to the CONSORT guidelines for clinical trials Provide clear statement and its methodological foundation in each published paper regarding the type of research (‘proof of concept’, ‘preclinical drug testing trial’) Clearly state whether the pharmacological intervention is meant to be ‘preventive’ or ‘disease-modifying’ Clearly state whether and how a potential biomarker can be transferred to humans Replicate ‘positive’ studies, report negative studies, perform meta-analyses Report conflicts of interests and financial resources

Table II.  Table II gives the minimum requirements for any proof of concept or preclinical study and the additional requirements specific to preclinical studies. We suggest this table be used as a check list for both investigators and reviewers.

Standard considerations for proof of concept and preclinical studies
1	Genetic background	Clearly stated, including breeding protocol and source of animals and gender. Gender should be specified for each experimental group
2	Potential sources of variability	Understood and addressed (e.g. gender, litter, copy number)
3	Numbers per group	Justified (power analysis, dependent on measure)
4	Select key read-outs from phenotype	Select key read-outs from an overt phenotype being predictive for disease and corresponding treatment
5	Proof of mechanism/exposure	Evidence that the target has been modulated or target tissue concentrations are adequate, ideally as a first step and in the disease model
6	Experimental design
(a)	Blinding	Placebo or drug treatment should be blinded to observer
(b)	Onset of clinical signs	Include measure of onset, well validated, robust, reproducible e.g. weight loss, behavioural signs
(c)	Disease progression	Quantitative measure of disease progression (weight loss, grip strength, wire hanging, gait analysis, neurological scoring)
(d)	Survival	For survival studies in G93A animals (high copy), each group should consist of a minimum of 12 animals of a single gender, unless justified by power analysis
(d)	Histology	Mandatory for positive outcomes. Should include motor neuron counts in gender specific groups of n = 5–6 minimum. Methods of analysis clearly described
(e)	Statistical analysis	Justified for each type of analysis. If potential confounding variables not controlled for in experimental design then appropriate statistical tests used (e.g. Cox proportional hazards regression)
7	External validity of data	Control group data should be compared to published data using the same background strain. Consider inter-laboratory comparison
8	Claims about therapeutic value	Must not be made in proof of concept studies
Additional considerations for preclinical studies
9	Replication of effect	In an independent laboratory or in a different mammalian model (e.g. different strain, different mutation)
10	Post-symptomatic efficacy	Highly recommended (effect on disease progression or survival)
11	Efficacy linked to exposure or biomarker of drug action	Facilitates translation to the clinic
(a)	Biomarker of drug action	Highly recommended. Link efficacy with biomarker of drug action, which has the potential to be replicated in the clinic.
(b)	Exposure	Highly recommended. Link efficacy to concentration of agent achieved in the target tissue and plasma (plasma concentration easily measured in the clinic)
12	Additional outcome measures	More specialized tests of disease progression would strengthen pre-clinical studies (e.g. MRI, physiology, comprehensive histopathology, gene profiling, proteomics)

Table III.  Recommendations for the use of animals.

the high-copy transgenic G93A-SOD animals remain the gold standard other rodent models should be systematically compared (other SOD mutations, lower-copy animals) advance and compare new animal models (Fus, TDP-43, high-copy dynactin G59S) try to develop drug-specific biomarkers use and develop new and fast in vivo drug screening tools (zebrafish, Drosophila)

Table IV.  Recommendations for pharmacological and toxicological studies in a preclinical drug testing study. We recommend that most of the information will be obtained by industrial partners.

preliminary qualitative understanding of the drug metabolism is mandatory (PK studies) ADME testing should be performed in the relevant species analysis of pharmacokinetics may also include tissue distribution (such as the central nervous system) Quantitative assessment of drug metabolism and distribution may be necessary to facilitate dose finding obtain data for NOAEL (no adverse effect level) and MTD (maximal tolerated dose) in the relevant species Thoroughly choose the route of application (also in proof of principle studies) obtain dose-response curves develop a drug-specific disease marker to facilitate translation

Table V.  Summary of new technologies providing a high likelihood of fulfilling these needs.

Technology	Deliverable
MRI based imaging	High anatomical and structural (DTI) resolution, potential of assessment of functional changes (e.g. MRI spectroscopy)
PET based imaging	High sensitivity for disease-specific functional changes
Automated home cage behavioural systems (e.g. Intellicage, PhenoMaster, PhenoTyper)	Stress-independent, ethologically-based behaviours with multidimensional recording of parameters, specific behavioural task within the home cages, multifactorial statistics and datamining
Automated home cage physiological systems for calorimetry and biopotentials (e.g. PhenoMaster, other telemetric systems)	Stress-independent, ethologically-based physiological read-outs at baseline with multidimensional recording of parameters and in combination with behaviour, specific physiological responses within the home cages, multifactorial statistics and datamining
Comprehensive set-ups within a dedicated facility/centre spanning/covering all dimensions/techniques of phenotyping (morphological, physiological, behavioural)	Multidimensional, highly standardized phenotyping/drug screening within a single centre increasing the likelihood of detecting a phenotyping/treatment effect