General principles of preclinical study design.

Preclinical studies using animals to study the potential of a therapeutic drug or strategy are important steps before translation to clinical trials. However, evidence has shown that poor quality in the design and conduct of these studies has not only impeded clinical translation but also led to significant waste of valuable research resources. It is clear that experimental biases are related to the poor quality seen with preclinical studies. In this chapter, we will focus on hypothesis testing type of preclinical studies and explain general concepts and principles in relation to the design of in vivo experiments, provide definitions of experimental biases and how to avoid them, and discuss major sources contributing to experimental biases and how to mitigate these sources. We will also explore the differences between confirmatory and exploratory studies, and discuss available guidelines on preclinical studies and how to use them. This chapter, together with relevant information in other chapters in the handbook, provides a powerful tool to enhance scientific rigour for preclinical studies without restricting creativity

Anti-nausea effects and pharmacokinetics of ondansetron, maropitant and metoclopramide in a low-dose cisplatin model of nausea and vomiting in the dog: a blinded crossover study

Nausea is a subjective sensation which is difficult to measure in non-verbal species. The aims of this study were to determine the efficacy of three classes of antiemetic drugs in a novel low dose cisplatin model of nausea and vomiting and measure change in potential nausea biomarkers arginine vasopressin (AVP) and cortisol. A four period cross-over blinded study was conducted in eight healthy beagle dogs of both genders. Dogs were administered 18 mg/m2 cisplatin intravenously, followed 45 min later by a 15 min infusion of either placebo (saline) or antiemetic treatment with ondansetron (0.5 mg/kg; 5-HT3 antagonist), maropitant (1 mg/kg; NK1 antagonist) or metoclopramide (0.5 mg/kg; D2 antagonist). The number of vomits and nausea associated behaviours, scored on a visual analogue scale, were recorded every 15 min for 8 h following cisplatin administration. Plasma samples were collected to measure AVP, cortisol and antiemetic drug concentrations

Differential hypoglycaemic, anorectic, autonomic and emetic effects of the glucagon-like peptide receptor agonist, exendin-4, in the conscious telemetered ferret.

Background: Rodents are incapable of emesis and consequently the emetic potential of glucagon-like peptide-1 receptor (GLP-1R) agonists in studies designed to assess a potential blood glucose lowering action of the compound was missed. Therefore, we investigated if the ferret, a carnivore with demonstrated translation capability in emesis research, would identify the emetic potential of the GLP-1R agonist, exendin-4, and any associated effects on gastric motor function, appetite and cardiovascular homeostasis. Methods: The biological activity of the GLP-1R ligands was investigated in vivo using a glucose tolerance test in pentobarbitone-anesthetised ferrets and in vitro using organ bath studies. Radiotelemetry was used to investigate the effect of exendin-4 on gastric myoelectric activity (GMA) and cardiovascular function in conscious ferrets; behaviour was also simultaneously assessed. Western blot was used to characterize GLP-1R distribution in the gastrointestinal and brain tissues. Results: In anesthetised ferrets, exendin-4 (30 nmol/kg, s.c.) reduced experimentally elevated blood glucose levels by 36.3%, whereas the GLP-1R antagonist, exendin (9–39) (300 nmol/kg, s.c.) antagonised the effect and increased AUC0–120 by 31.0% when injected alone (P < 0.05). In animals with radiotelemetry devices, exendin-4 (100 nmol/kg, s.c.) induced emesis in 1/9 ferrets, but inhibited food intake and decreased heart rate variability (HRV) in all animals (P < 0.05). In the animals not exhibiting emesis, there was no effect on GMA, mean arterial blood pressure, heart rate, or core body temperature. In the ferret exhibiting emesis, there was a shift in the GMA towards bradygastria with a decrease in power, and a concomitant decrease in HRV. Western blot revealed GLP-1R throughout the gastrointestinal tract but exendin-4 (up to 300 nM) and exendin (9–39), failed to contract or relax isolated ferret gut tissues. GLP-1R were found in all major brain regions and the levels were comparable those in the vagus nerve. Conclusions: Peripherally administered exendin-4 reduced blood glucose and inhibited feeding with a low emetic potential similar to that in humans (11% vs 12.8%). A disrupted GMA only occurred in the animal exhibiting emesis raising the possibility that disruption of the GMA may influence the probability of emesis occurring in response to treatment with GLP-1R agonists

Assessment of low-dose cisplatin as a model of nausea and emesis in beagle dogs, potential for repeated administration

Cisplatin is a highly emetogenic cancer chemotherapy agent, which is often used to induce nausea and emesis in animal models. The cytotoxic properties of cisplatin also cause adverse events that negatively impact on animal welfare preventing repeated administration of cisplatin. In this study, we assessed whether a low (subclinical) dose of cisplatin could be utilized as a model of nausea and emesis in the dog while decreasing the severity of adverse events to allow repeated administration. The emetic, nausea-like behavior and potential biomarker response to both the clinical dose (70 mg/m2) and low dose (15 mg/m2) of cisplatin was assessed. Plasma creatinine concentrations and granulocyte counts were used to assess adverse effects on the kidneys and bone marrow, respectively. Nausea-like behavior and emesis was induced by both doses of cisplatin, but the latency to onset was greater in the low-dose group. No significant change in plasma creatinine was detected for either dose groups. Granulocytes were significantly reduced compared with baseline (P = 0.000) following the clinical, but not the low-dose cisplatin group. Tolerability of repeated administration was assessed with 4 administrations of an 18 mg/m2 dose cisplatin. Plasma creatinine did not change significantly. Cumulative effects on the granulocytes occurred, they were significantly decreased (P = 0.03) from baseline at 3 weeks following cisplatin for the 4th administration only. Our results suggest that subclinical doses (15 and 18 mg/m2) of cisplatin induce nausea-like behavior and emesis but have reduced adverse effects compared with the clinical dose allowing for repeated administration in crossover studies

Retrospective harm benefit analysis of pre-clinical animal research for six treatment interventions

The harm benefit analysis (HBA) is the cornerstone of animal research regulation and is considered to be a key ethical safeguard for animals. The HBA involves weighing the anticipated benefits of animal research against its predicted harms to animals but there are doubts about how objective and accountable this process is.i. To explore the harms to animals involved in pre-clinical animal studies and to assess these against the benefits for humans accruing from these studies; ii. To test the feasibility of conducting this type of retrospective HBA.Data on harms were systematically extracted from a sample of pre-clinical animal studies whose clinical relevance had already been investigated by comparing systematic reviews of the animal studies with systematic reviews of human studies for the same interventions (antifibrinolytics for haemorrhage, bisphosphonates for osteoporosis, corticosteroids for brain injury, Tirilazad for stroke, antenatal corticosteroids for neonatal respiratory distress and thrombolytics for stroke). Clinical relevance was also explored in terms of current clinical practice. Harms were categorised for severity using an expert panel. The quality of the research and its impact were considered. Bateson's Cube was used to conduct the HBA.The most common assessment of animal harms by the expert panel was 'severe'. Reported use of analgesia was rare and some animals (including most neonates) endured significant procedures with no, or only light, anaesthesia reported. Some animals suffered iatrogenic harms. Many were kept alive for long periods post-experimentally but only 1% of studies reported post-operative care. A third of studies reported that some animals died prior to endpoints. All the studies were of poor quality. Having weighed the actual harms to animals against the actual clinical benefits accruing from these studies, and taking into account the quality of the research and its impact, less than 7% of the studies were permissible according to Bateson's Cube: only the moderate bisphosphonate studies appeared to minimise harms to animals whilst being associated with benefit for humans.This is the first time the accountability of the HBA has been systematically explored across a range of pre-clinical animal studies. The regulatory systems in place when these studies were conducted failed to safeguard animals from severe suffering or to ensure that only beneficial, scientifically rigorous research was conducted. Our findings indicate a pressing need to: i. review regulations, particularly those that permit animals to suffer severe harms; ii. reform the processes of prospectively assessing pre-clinical animal studies to make them fit for purpose; and iii. systematically evaluate the benefits of pre-clinical animal research to permit a more realistic assessment of its likely future benefits

Cisplatin-induced emesis: systematic review and meta-analysis of the ferret model and the effects of 5-HT3 receptor antagonists

PURPOSE: The ferret cisplatin emesis model has been used for ~30 years and enabled identification of clinically used anti-emetics. We provide an objective assessment of this model including efficacy of 5-HT(3) receptor antagonists to assess its translational validity. METHODS: A systematic review identified available evidence and was used to perform meta-analyses. RESULTS: Of 182 potentially relevant publications, 115 reported cisplatin-induced emesis in ferrets and 68 were included in the analysis. The majority (n = 53) used a 10 mg kg(−1) dose to induce acute emesis, which peaked after 2 h. More recent studies (n = 11) also used 5 mg kg(−1), which induced a biphasic response peaking at 12 h and 48 h. Overall, 5-HT(3) receptor antagonists reduced cisplatin (5 mg kg(−1)) emesis by 68% (45–91%) during the acute phase (day 1) and by 67% (48–86%) and 53% (38–68%, all P < 0.001), during the delayed phase (days 2, 3). In an analysis focused on the acute phase, the efficacy of ondansetron was dependent on the dosage and observation period but not on the dose of cisplatin. CONCLUSION: Our analysis enabled novel findings to be extracted from the literature including factors which may impact on the applicability of preclinical results to humans. It reveals that the efficacy of ondansetron is similar against low and high doses of cisplatin. Additionally, we showed that 5-HT(3) receptor antagonists have a similar efficacy during acute and delayed emesis, which provides a novel insight into the pharmacology of delayed emesis in the ferret

General Principles of Preclinical Study Design

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