D-amino acids as modulators of the genotoxic polyketide, colibactin.

"You are what you eat" is a commonly used phrase that everyone should take on board. For example, information from over 4 million patients has shown that obesity, type 2 diabetes, and poor metabolic health are three significant risk factors for worse COVID-19 outcomes. These risk factors are heavily influenced by the food we eat. The challenge is that our diet is complex and it can be hard to know what is "good" or "bad" for you. Moreover, there is an increasing appreciation of the importance of our "gut health", the complex mix of different microbes that help us break down food and protect against harmful bacteria. Many people try and supplement their gut microbes with so called "good bacteria" or probiotics that claim to provide, and in some cases are even shown, to have clear positive impacts.

But do we know these probiotics are actually safe? Recent work has shown that strains of E. coli that have been used for almost 100 years as probiotics, can actually produce a toxin that induces colo-rectal cancer. This is really worrying for us. For example, bowel cancer is the 3rd most common cancer (10% of all new female cancer cases/ 13% of all new male cancer cases) in the UK and highly related to diet. By chance, we have found a simple supplement that helps reduce the production of the toxin made by E. coli when it is grown in the lab. We now have 3 important questions: (1) can we make even more effective supplements to block the toxin. (2) how does the supplement affect toxin production and (3) can it work in animals? These are the three aims of this work.

In addition, our work also aims to understand if the supplement itself affects the normal bacteria within the gut. We will test this by looking at the different species that live in the gut before and after the treatment. This is key to knowing that the treatment itself does not cause problems.

We have previously shown that D-amino acids (AA), particularly D-serine (D-ser), can act as a powerful signal to E. coli, suppressing the virulence of the enterohaemorrhagic E. coli (EHEC) pathotype. We recently observed that D-ser also markedly reduces expression of the potent genotoxin colibactin, which is carried by many strains of E. coli including a probiotic strain E. coli Nissle. Our preliminary data show that E. coli strains that carry the pks cluster produce the genotoxin colibactin that induces both double stranded DNA breaks and morphological changes such as megalocytosis in eukaryotic cells. Addition of D-ser reduces expression of colibactin and significantly affects the development of these cancer-associated changes. Based on these findings, our hypothesis is that supplementing D-AAs in the diet can affect production of colibactin in the GI tract and reduce the negative effects this genotoxin can induce, including colo-rectal cancer.

Our previous work led us to focus on D-ser, yet numerous D-AAs have been reported to affect regulatory processes. Therefore, our first objective will be to address if other D-AAs, either singly or in combinations, can reduce colibactin production. Secondly, we want to understand the molecular mechanism that of how D-AAs interfere with colibactin production. We have excellent data supporting the role of specific regulators in this process and will tease apart this molecular mechanism. These two objectives will help us optimise the most potent combination of D-AAs before moving to our third objective, which will be to test if dietary D-AAs alter the effects of colibactin in vivo. Our research is important because it provides a direct and tangible way to reduce colibactin levels with obvious downstream health benefits. The work aligns perfectly with the BBSRC strategic priority "Food, Nutrition and Health".