Abstract
Objectives
The aim of this study was to compare the palatability of sunflower oil-based and aqueous verum formulations of molidustat in healthy cats in a one-bowl study with two groups in a parallel design.
Methods
A total of 16 adult cats (six males and 10 females) were divided into two groups of eight animals. Healthy cats trained on voluntary intake of a water/milk mixture offered in a bowl at daily volumes of 0.5 ml/animal over 4 study days (days –4 to –1) were included in the study. Daily general health observations were performed throughout the study. After the training, each cat was offered daily 0.2 ml/kg body weight of the same test formulation in a bowl for 7 consecutive days (days 0–6). The palatability of formulations was assessed individually by means of a visual analogue scale based on voluntary intake. The study ended on study day 7 for both groups.
Results
Voluntary intake scores were higher in cats in group 1 offered the sunflower oil-based suspension of molidustat. In contrast, almost no voluntary intake, even when cats were encouraged, was observed for the aqueous suspension of molidustat (group 2).
Conclusions and relevance
This study demonstrated that molidustat in a sunflower oil-based suspension was palatable to healthy cats. In contrast, the aqueous formulation was unpalatable.
Introduction
The development of oral drug formulations for cats relies on palatability to improve treatment compliance. Palatability refers to voluntary (free choice) acceptance or ingestion of a pharmaceutical formulation by the cat, as measured by a standard palatability test, such as an acceptance, preference or consumption test. 1 Acceptance means that the formulation is voluntarily taken into the mouth of the cat and consumed. The speed at which the cat accepts the product and the degree of interest or enthusiasm shown by the cat are also considered measures of palatability. Cats are more difficult to medicate than dogs, and some owners find liquids the easiest dosage form to administer. 2 It is ideal when the cat willingly and voluntarily consumes the medication, particularly when the medication needs to be administered chronically, such as molidustat. 1
Molidustat is an orally administered hypoxia-inducible factor prolyl hydroxylase inhibitor used to treat non-regenerative anaemia associated with chronic kidney disease (CKD) in cats. Given that cats with CKD often require long-term management of anaemia, the palatability of the molidustat oral suspension is crucial to ensure consistent administration and treatment compliance over extended periods. 3 A palatable formulation helps minimise stress for both cats and their owners, supporting adherence to the prescribed daily dosing regimen essential for achieving optimal therapeutic outcomes in this chronic condition.3 –5 The aim of this study was to compare the palatability of two verum formulations of molidustat in healthy cats in an exploratory study. Oral suspensions were either sunflower oil-based or aqueous. A verum formulation contains an active medicinal ingredient, such as molidustat in this study, whereas a placebo or control formulation contains solely the vehicle without any active ingredient.
Materials and methods
Ethical approval
The study was conducted in accordance with institutional Good Scientific Practice principles, although formal Good Laboratory Practice or Good Clinical Practice compliance was not required for this type of study. It also complied with all relevant national and European regulations on the protection of animals used for scientific purposes, including the German Animal Welfare Act (TierSchG) and EU Directive 2010/63/EU. The experimental protocol was reviewed and approved by the animal welfare officer and German authorities to ensure the highest standards of animal care and welfare throughout this laboratory study.
Animals
A total of 16 adult (aged 2.4–2.5 years) purpose-bred neutered domestic shorthair cats (six males and 10 females) were divided into two groups of eight animals each. The study was conducted at Bayer Animal Health Development, Clinical Development Animal Center, Monheim, Germany. Colony cats were acclimated to conditions of the test facility for at least 2 weeks before the study. Inclusion suitability was assessed on study day –4 based on normal findings from physical examination, haematology, serum chemistry and body weight. Body weight was measured again on study day 0. No concomitant medication was administered during the study. Cats were fasted overnight and then allowed access to normal food rations 1 h after being offered either test formulation. Daily general health observations such as general appearance, percentage of remaining feed and faeces consistency were performed for each group per cage (n = 4) throughout the study. The study ended on study day 7 for both groups, with a final physical examination and body weight measurement. At the end of the study, all cats were returned to the facility colony.
Study design and drug administration
Study cats were trained for voluntary intake of a water/milk liquid mixture offered in a bowl at 0.5 ml/cat q24h for 4 days before the study start (study days –4 to –1). The cats did not show any aversion behaviours when ingesting the water/milk mixture. Although all cats showed interest in the water/milk mixture, interest declined over the duration of the baseline period and sometimes cats had to be encouraged repeatedly to ingest the mixture.
To assess acceptability of the two verum formulations, a ‘one-bowl’ test was conducted with two groups of cats in a parallel design with sunflower oil-based (group 1; four males and four females) and aqueous (group 2; two males and six females) formulations containing 25 mg/ml of molidustat (2.5% (w/v). As this was an exploratory study, no formal randomisation was performed.
Voluntary intake
Palatability was assessed in individual cats by means of a visual analogue scale (VAS) based on the intake behaviour. The scale line was 10 cm in length with the following endpoints:
0 = worst possible voluntary intake (cat shows no interest and does not take up the test formulation)
10 = best possible voluntary intake (cat ingests the test formulation completely)
Beginning on study day 0, each cat was offered daily its assigned test formulation at 0.2 ml/kg body weight in a bowl for a maximum of 3 mins. The interest of the cat and the intake of the formulation could be encouraged (eg, via pushing the food bowl towards the cat). After offering the test formulation for voluntary intake, the assessor placed a vertical mark on the VAS line based on the cat’s behaviour (eg, no uptake/shows interest, smells at the test formulation/partial consumption of the test formulation/looks for more). All voluntary intake assessments were performed by the same individual who was not masked.
Statistical analysis
The individual animal was the experimental unit. For each VAS assessment, the distance from the end of the VAS line labelled 0 to the vertical mark was measured. Results were recorded as a figure between 0 and 10. Only descriptive statistical analyses were performed on the VAS scores of each group, including means, standard deviations, coefficients of variation, medians, and minimums and maximums. No statistical analysis could be performed for comparison between groups because of median VAS scores of zero for all study days for group 2.
Results
Demographic characteristics of the study population are provided in Table 1. The initial body weight of all cats was normal for age (3.4–5.3 kg) on study day –4. Changes in body weight (in kg and percentages) on study day 7 compared with study day −4 are also shown in Table 1. Results of the VAS scoring for the evaluation of palatability of both formulations are presented in Table 2. All cats offered the sunflower oil-based suspension (group 1) showed an interest in this formulation; however, interest declined over the duration of the study and all animals needed some encouragement to consume the formulation. Over the course of the 7-day study, median VAS scores were in the range of 6.5–10. The interindividual variability was low (CV% 5–23%), except on study day 3, where variability between subjects was high (39%) and the lowest median score was recorded (Table 2 and Figure 1). In contrast, on all but two offerings of the aqueous formulation, individual VAS scores were 0 – that is, cats showed no interest and did not voluntarily intake this test formulation.
Demographic characteristics of cats including body weight (BW) (kg) and change from study day –4 (baseline) in kg and percentage over time
All study cats were neutered
F = female; M = male
Evaluation of palatability of molidustat in sunflower oil-based or aqueous suspensions in healthy cats using a visual analogue scale scoring scheme*
Irrespective of how often the cats were encouraged to consume the liquid, whether the water/milk mixture (study days –4 to –1) or the respective oily/aqueous formulations (study days 0–6), scores of 8–10 were assigned as follows: score 10 = 100% intake; score 9 ≈ 95% intake; score 8 ≈ 80% intake
CV = coefficient of variation; NA = not applicable
Mean visual analogue scale (VAS) scores over time for group 1 (sunflower oil-based suspension of 25 mg/ml molidustat) and group 2 (aqueous suspension of 25 mg/ml molidustat). Error bars are standard deviations
An adverse event was reported in two cats from group 2. Vomit (undigested food from the previous day) was found at the bottom of the cage of these two cats (cats 7 and 8), which were housed together, on study day 2 during cage cleaning before the morning feeding. However, as no voluntary intake of the aqueous formulation had been observed in these two cats, this reported adverse event was not considered related to the test formulation. An additional physical examination was performed, and no clinical abnormalities were detected in either cat. No other clinical abnormalities were observed in the remaining cats during the study. All cats were assessed as normal during the physical examination on study day 7.
Discussion
The voluntary intake of a sunflower oil-based molidustat suspension compared with an aqueous verum formulation was evaluated in healthy cats over a period of 7 consecutive days. This study duration was in line with the European Medicines Agency (EMA) guideline on the demonstration of palatability of veterinary medicinal products for anticipated daily treatments lasting more than 14 days. 6 This study showed that cats preferred the sunflower oil-based suspension of molidustat over the aqueous formulation. Voluntary acceptance is defined in the EMA guidelines as the willingness of the target animal to consume voluntarily and spontaneously the veterinary medicinal product from the bowl/trough/ground or from the hand when offered as a treat by the animal’s owner. 6 In this study, the assessment of voluntary acceptance was based on a VAS scheme that showed all cats voluntarily accepted the oil-based suspension at all time points (56/56, 100%), whereas none of the cats from the aqueous group voluntarily accepted the verum formulation except at one time point for two cats (2/56, 3.6%). To be granted a palatability claim, the overall voluntary acceptance rate – calculated as the total number of successful administrations divided by the total number of administrations and multiplied by 100 – should reach at least 70% in cats. 6 The sunflower oil-based formulation met this threshold.
Over the 7-day evaluation period, a notable decrease in overall acceptance of the sunflower oil-based suspension occurred on study day 3, characterised by high inter-subject variability (39%) and the lowest median palatability score. Notably, 50% of the cats recorded VAS scores of 4 or 5 on this day. The transient dip in acceptance seen on study day 3 may be explained by short‑term factors such as flavour fatigue from repeated exposure, day‑to‑day variations in appetite or behaviour, or minor differences in testing conditions. Individual sensory preferences for flavour, aroma or texture may also have contributed. Such fluctuations are not uncommon in feline palatability studies and are generally considered temporary rather than indicative of a sustained decline in acceptance.
Domestic cats (Felis silvestris catus) are obligate carnivores that require meat-based diets. 4 The anatomical, physiological, chemical, genetic and behavioural aspects for taste and olfaction in cats have been previously researched. 4 The cat’s strong taste preference for umami (savoury or meaty taste) and amino acids, along with their preference for protein and fat, reflects their carnivorous diet. 5 Cats preferentially select high protein and fat diets but not diets with high carbohydrate content.7,8 Although taste and smell are both important in food selection, taste is the more dominant sense in influencing the food preference of cats as opposed to colour and orthonasal olfaction. 8 Studies revealed feline preferences for fish, specifically salmon, over commercial cat food (liver, chicken or beef flavoured) and rats. 9 Cats do not perceive sweet tastes because they lack a receptor for recognising this taste. 10 The aqueous formulation contained sorbitol, a sugar alcohol commonly used as a sweetener and commonly added to processed foods and medications for humans, but it had no positive effect on the palatability of the aqueous formulation for the cats in group 2.
Drug compliance in cats is the extent to which owners adhere to instructions when giving prescribed medications to their cats. In a survey of cat owners, owners were unable to give doses for one-quarter of the medications prescribed. 2 Most treatment failures were related to individual cat behaviour (eg, resistance to administration or spitting out the drug) or adverse effects, such as profuse salivation. Owners also reported that the unpleasant taste of the drug was an important factor for non-compliance. Solutions and suspensions have been shown to be significantly more acceptable than solid dosage forms. The palatability of oral suspensions approved for cats, administered either alone or mixed with food, has been demonstrated in previous studies.11,12 Therefore, the development of a palatable oral suspension for molidustat delivery may improve owner compliance.
The one-bowl test, also known as the single-bowl test, one-pan test or monadic test, involves presenting a predetermined formulation sample for a specific period of time.13,14 This is monitored under normal feeding parameters and is repeated for several days, typically 5 days or longer. The one-bowl test is low cost, and 8–10 animals are typically sufficient to detect a trend. The advantage of this acceptance test for drug formulations is that it tests consumption of the medication and therefore is a direct measure of compliance. 15 Test conditions replicate a typical home environment for cats, with one bowl provided per trial. Cats can be fed on their regular schedule and in their normal environment. Training is not necessary for participation. Results from group 2 cats, who were offered the aqueous-based formulation, illustrate that this test is effective in detecting unpalatable formulations, potentially resulting from undesirable flavour, aroma or texture.
Limitations of the acceptance test include interference from appetite factors. Appetite increases with time between offerings and decreases with quantity offered. Cats may prefer new foods (neophilia) or be hesitant to try new foods (neophobia), and this may affect the results of palatability tests. It is also difficult to control external factors such as noises and ambiance, which can affect results. Variability in preferences may be affected by previous exposure to different foods, feeding situation, family structure and individual factors such as sex, age, breed, health condition and season (cats eat less in the winter). 14 Although a crossover study design is generally preferred when comparing formulations to reduce interindividual variability, a parallel design was deemed acceptable in this study. This was because the sunflower oil-based formulation (group 1) showed a clear and pronounced preference among cats, with minimal interindividual variability; median scores were in the range of 6.5–10 for group 1 compared with 0 for group 2, making it unlikely that a crossover design would have altered the outcome.
Limitations of the study include its short duration and a decline in feline interest in the formulation over time, which is noteworthy given that molidustat is intended as a long-term treatment for non-regenerative anaemia associated with CKD in cats. Nevertheless, with minimal coaxing, cats continued to voluntarily consume the sunflower oil-based suspension, which was ultimately selected as the final commercial formulation for molidustat (Varenzin; Elanco Animal Health).
Extrapolation of palatability results for molidustat in a sunflower oil-based suspension from healthy cats to cats with CKD experiencing inappetence should be undertaken with caution. Although this study demonstrated the sunflower oil formulation was palatable to healthy cats, cats with CKD often have decreased appetite, nausea, altered taste perception, and concurrent illness that can affect their food and medication acceptance. However, given the clinical need for long-term oral administration of molidustat for the management of anaemia in cats with CKD, a palatable formulation such as the sunflower oil-based suspension may still improve voluntary intake compared with a less palatable aqueous version. This is particularly important because treatment compliance is critical for therapeutic success in this population. Supporting this, clinical studies of molidustat in anaemic cats with CKD (using the commercial sunflower oil-based suspension) have shown that it can be administered successfully over weeks with stable acceptance. 3 These real-world compliance observations in the target CKD population, despite variable appetite, strengthen confidence in extrapoliating the study results.
Conclusions
This study demonstrated that molidustat in a sunflower oil-based suspension was palatable to healthy cats, while the aqueous formulation was unpalatable.
Footnotes
Acknowledgements
The authors thank the study staff for their valuable assistance with the conduct of the study. The authors also thank Dr Chantal Lainesse (Integral Consulting Strategies) for providing medical writing assistance with this manuscript and submission to the journal on behalf of the authors. Dr Lainesse’s services were paid by Elanco Animal Health.
Conflict of interest
All authors were employees of Bayer Animal Health and are now employees of Elanco Animal Health.
Funding
The study was funded by the sponsor, Bayer Animal Health, acquired by Elanco Animal Health.
Ethical approval
The work described in this manuscript involved the use of experimental animals and the study therefore had prior ethical approval from an established (or ad hoc) committee as stated in the manuscript.
Informed consent
Informed consent (verbal or written) was obtained from the owner or legal custodian of all animal(s) described in this work (experimental or non-experimental animals, including cadavers, tissues and samples) for all procedure(s) undertaken (prospective or retrospective studies). No animals or people are identifiable within this publication, and therefore additional informed consent for publication was not required.