Using a 'Designer Receptor Exclusively Activated by Designer Drug' to define the role of short chain fatty acids in metabolic disease and inflammation

Food is often considered simply as a source of fuel for the body. However, in recent times it has become clear that components of what we eat either also act directly to regulate cellular and tissue functions or are converted into molecules that do so. The concept of a 'healthy diet' and how this might be orchestrated and defined by the vast number of bacteria that populate the intestine is not new but, in recent times, knowledge about and public interest in the relationship between diet, well-being and human disease has grown substantially. For example, it is widely recognised that 'fibre' is an important component of a healthy diet. Microbes in the gut convert 'fibre' into short chain fatty acids (SCFAs) and high levels of these are present in the gut and are transported into the body. In both of those locations such SCFAs are believed to produce many of their important roles by binding to and activating transmembrane receptors called free fatty acid receptor (FFAR)2 and FFAR3. Both FFAR2 and FFAR3 are members of the G protein-coupled receptor (GPCR) family and developing drugs that either activate or block other GPCRs has proved to be a very effective approach to tackling a wide range of human diseases. As these receptors are present on a number of tissues including various white blood cells that control inflammation and immunity, adipocytes that store fat, and various cells of the lower gut that release hormones that in turn modify release of insulin from the pancreas and have effects on the desire to eat, they are being considered as possible receptors to target in diseases that range from inflammatory bowel disease to diabetes. However, prior to making long term decisions that targeting any receptor is likely to be a productive approach, pharmaceutical companies seek 'validation', to generate as much confidence in the idea as possible. There are a number of issues in relation to validation of either FFAR2 or FFAR3. Firstly these two receptors are activated by the same SCFAs. This means that in cells and tissues in which both are present it is difficult to be certain which is most important. Previous efforts to overcome this have involved making mice that lack either FFAR2 or FFAR3. Although useful, the absence of one of the two results in change in amount of the other. Secondly, a number of key effects may require both to be present. To overcome this we have made a version of FFAR2 that instead of being activated by SCFAs is instead activated by a molecule called sorbic acid. A key plan proposed is to generate mice in which the normal copy of FFAR2 is replaced with this engineered variant. In these animals both FFAR2 and FFAR3 will be present but now they can be activated by different molecules and this will allow us to tease out their individual contributions to function. This will provide validation of either FFAR2 or FFAR3 in different therapeutic indications. Of course this is a means to an end and the long term plan is to develop selective medicines that target FFAR2 and FFAR3. The applicants have begun to develop assays and approaches to do this and have generated a number of pharmacological 'tool' compounds to help further define the function of FFAR2 and FFAR3. However, pharmaceutical companies are better able to bring the skills required to convert such 'tool' compounds into molecules that have 'drug-like' properties and, therefore, might be used to test ideas developed in either simple cell systems and in mice into humans. As such, the applicants have agreed to collaborate in this project with scientists from the pharmaceutical company AstraZeneca. In these studies we will be provided with a range of novel 'tool' compounds for FFAR2 and FFAR3 to provide further insight and the outcomes will assist the company in making key decisions on whether FFAR2 and/or FFAR3 become 'validated' as realistic targets to develop effective medicines against.

The relationship between diet, well-being and human disease is of considerable public interest and the subject of government health policy. Traditionally this has centred on the correct balance of nutrients where food was simply viewed as fuel or providing essential factors. It is now clear, however, that components of our diet can act as bioactive molecules in their own right, having a direct impact on biological processes. Prominent among the nutrients that act as bioactive molecules are short-chain free fatty acids (SCFAs). These are generated by the fermentation of non-digestible carbohydrates by the gut microbiota; and are proposed to control biological responses such as glucose homeostasis and inflammation. Thus, the SCFAs exemplify the relationship between diet, the microbiota and well-being and as such are of considerable interest to the public, government and pharmaceutical companies targeting obesity, type II diabetes, as well as metabolic and inflammatory diseases. The issue is, however, that we know very little about the modes of action of the SCFAs that act through two G-protein coupled receptors, FFAR2 and FFAR3. This is due to the fact that there are very few pharmacological tool compounds available to probe the in vivo function of these receptors together with the fact animal knockout models are inadequate. In collaboration with our industrial partner, AstraZeneca, we address these key issues by not only generating an array of novel tool compounds to FFAR2/3 receptors but also by generating a novel chemical genetic mouse model (FFAR2-DREADD) that will be used to probe the function of these receptors in glucose homeostasis, metabolism and inflammation. Thus, using the FFAR2-DREADD mice together with FFAR2-KO and FFAR3-KO mice and in conjunction with the novel FFAR2/3 ligands this study aims to not only define the physiological role of SCFAs in mouse but will also establish the therapeutic potential of targeting the FFAR2 and FFAR3 receptors.

Who will benefit from this research?

This programme addresses the relationship between diet, the microbiota and well-being; an area that is currently of general interest to the public where there is an understanding that controlling the microbiota with probiotics has an impact on health. Miller, Milligan, and Tobin are are experienced in public engagement (press releases/popular media), hence dissemination of the results to the general public will facilitate the broad impact of our studies. Associated with this is the possibility that the data generated here might have impact on government health policy particularly in the area of diet and well-being. The most immediate beneficiaries from the research, however, will be our industrial partner AstraZeneca. Although the work detailed in the proposal will be made publically available, AstraZeneca will have immediate access to the results, in advance of public presentation. This is likely to provide AstraZeneca benefits commensurate with their contribution to the project both via direct funding and 'in-kind' contributions. Once the work has been made publically available then the next set of beneficiaries will be the wider pharmaceutical industry. There is considerable interest in whether acute or more sustained pharmacological manipulation of FFAR2 and/or FFAR3 might result in effective control of both gut inflammatory and metabolic diseases and the studies that will be performed are likely to shed considerable light on these topics. The third group of beneficiaries from this research are the neutraceutical and food industries. This reflects their interest in the potential effectiveness of both pre- and pro-biotic strategies to maintain or improve health and the developing concept of 'functional foods'.

How will they benefit from this research?

The general public and government agencies will certainly benefit from a clearer understanding of the impact of diet on well-being. This might affect public dietary habits and government policy on good nutrition. The results will certainly benefit the pharmaceutical industry as a whole, many of who are targeting free fatty acid receptors in the control of obesity, type II diabetes and inflammation. Thus, validation of FFAR2 and FFAR3 as targets in these indications will benefit these companies. The simple fact AstraZeneca have provided support to allow for an Industrial Partnership Award highlights the strategic decision of the company to invest in fully defining the therapeutic potential of FFAR2 and FFAR3. Rapid access to the research findings will allow more informed decisions at a corporate level about investment in efforts to identify lead and candidate molecules to target FFAR2/3. Other pharmaceutical companies will benefit in a similar way, although without such immediate access to the results. Any drug discovery programme targeting chronic diseases such as diabetes and inflammation of the lower gut is inherently a long term endeavour (8-12 years) but the work proposed certainly has the potential to impact on quality of life and healthy aging. It is generally easier to obtain regulatory clearance in some aspects of 'food and nutrition', so it is possible, therefore, that the research here would impact decisions in the food and nutrition industries in a 3-5 year time scale.
The project also has great potential in terms of staff training in that the PDRAs will benefit from opportunities to perform cutting edge research in a broad swathe of areas relevant to modern pharmacological studies, to enhance team working via the need to integrate work from two sites and three laboratories, and to interact directly with staff within a major international pharmaceutical company. This training is likely to be optimal to ensure the greatest range of subsequent career opportunities.