The aim of the proposed research is to find new enzymes that have potential uses in industry by searching for the genes for these enzymes in the DNA extracted directly from soil, compost or other environments. Enzymes are very useful in biocatalysis which is a sustainable method of making chemicals in industry. If enzymes are used the eventual industrial process can be cleaner and greener as it avoids the use of toxic reagents such as metals needed for many chemical catalysis steps, and often uses water-based systems. Biocatalysis can also replace several steps in a chemical process with one enzyme step due to their selectivity and this has a major effect of saving money and time in the overall process for making high value chemicals such as bioactive compounds in the fine chemical and pharmaceutical industry. We will use a technique called metagenomics to find new enzymes for biocatalysis. Many enzymes are derived from microbial sources and these would normally be found by growing bacteria on agar plates and analysing the enzymes they contain using special assays. However, several years ago scientists studying soil microorganisms found that there was a very large difference between the numbers of bacteria they could grow from a soil sample compared with the numbers they could identify by analysing the DNA from the same quantity of soil. These DNA techniques showed that there were over 1,000 times more bacteria in the soil than can be grown on agar plates. So by using plating and growth techniques to find bacteria for biocatalytic enzymes were are missing over 99.9% of the potential enzymes. A technique called metagenomics was developed by several researchers which started with the extraction of DNA directly from a soil sample and this DNA would potentially contain all the genes of the bacteria including the genes from bacteria that cannot be grown in the laboratory. We will use this metagenomic technique to isolate DNA from soils and other environmental samples. The metagenomic DNA will be sequenced and potential genes for biocatalysis will be searched for using computer based techniques to analyse the metagenome. When we find what could be useful genes we will amplify the gene from a sample of the metagenomic DNA and put the amplified gene into a laboratory bacterium that we can grow in large amounts and test the activity of the new biocatalytic enzyme. We call this overall method Functional Metagenomics. The new biocatalysts will be tested in collaboration with researchers at Almac who use enzymes and chemistry to make pharmaceutical compounds. We will test the range of reactions the new biocatalysts can perform and test the chemicals made. A new concept called enrichment metagenomics will also be investigated where we will enrich for bacteria able to use a specific compound before doing the metagenomics. This has the potential to increase the number of bacteria with the desired biocatalytic enzyme. Another new concept called cDNA metagenomics will be tested where we extract messenger RNA from the sample and convert this into what is known as cDNA. This technique will allow us to look for genes from the microorganisms such as soil fungi that have introns in their DNA. This could enable us to find a hitherto unaccessed pool of new enzymes for biocatalysis.

Colorectal cancer (CRC) is the 3rd most common cancer in the UK, with >40,000 new cases in 2011. While there have been improvements in CRC treatment, it remains a significant killer, with 16,000 deaths in 2011. Research by ourselves/others has revealed that a "one size fits all approach" will not work, as genetic changes in their CRC cells can cause treatments to fail in particular patients. This increased understanding has given rise to the concept of "stratified medicine", where testing a patient's sample prior to treatment can indicate which therapy works in this particular patient. This "stratified" approach also allows patients who will not respond to be spared the often toxic side effects. Recognising the need to provide treatments leading to better survival/Quality of Life (Qol), a group of researchers, clinicians, patient groups and industry have formed a consortium (S-CORT), harnessing its members expertise to develop new approaches to stratify patients to improve outcomes, thus delivering real benefit for CRC patients. S-CORTs objectives are to: 1. Create a consortium united in the common goal to employ stratified medicine to yield better survival and QoL for CRC patients 2. Build on discoveries by S-CORT researchers to identify particular stratification approaches for patients receiving different therapies for CRC. Three priorities have been established a. While the drug Oxaliplatin has increased our options for treating CRC, approximately 50% of patients don't respond and develop side effects that can affect their nervous system and reduce their QoL. Being able to decide in advance which patients respond, allows those patients to receive the drug while sparing non-responders the toxic side effects b. ChemoRadiotherapy (CRT) is used in the treatment of rectal cancer, but 40% of patients with locally advanced disease gain no benefit. A stratification approach may not only indicate which patients to treat, but also allow design of new approaches to make RT more effective c. In early disease, some patients can have aggressive cancer which invades other parts of the body. Identifying these patients in advance of treatment would allow them to receive more extensive surgery/RT while those with less aggressive disease can be treated with local rectal preserving treatment 3. Establish a more complete understanding of the precise changes that occur in the genes and proteins of CRC cells and use this information to provide novel therapies for patients 4. Develop our best candidates into clinical tests that select patients for the therapies that have the greatest chance of success and/or with the fewest side effects in their particular disease 5. Bring together all our research into a database that will be a vital resource for future research, within and outside this consortium 6. Ensure that the patient is at the centre of all activities in S-CORT, helping with the design of studies, participating in focus groups, meetings and conferences and contributing to the communication of the activities of S-CORT to healthcare and research professionals, patient groups and the public at large 7. Publish our research findings in the best scientific journals and present our results at national and international conferences, thus demonstrating the quality of S-CORT's research 8. Examine how tests that we are developing will perform in the hospital for CRC patients and evaluate the health, economic and societal benefits of this approach 9. Ensure S-CORT's long term sustainability, thus driving implementation of new stratification approaches for CRC patients over the next decade, both in the UK and globally Delivering these ambitious objectives will allow development of new clinical tests to predict success/ failure of new therapies which, coupled with our increased knowledge of CRC biology will drive a new treatment vision where stratified medicine approaches can significantly benefit our patients.

KEYWORDS: chemistry, computer vision, image processing, manufacturing, productivity, process monitoring. Chemical and biochemical manufacturing are dominated by colour changes, both subtle and stark. Such phenomena are often reported by-eye but not routinely quantified, especially over time. This renewal of a research and leadership programme aims to empower any chemist with any camera to capture any visible trend from any high-value chemical process, all without having to disturb the process under study. Most industrial chemists are accustomed to extracting chemical monitoring information using invasive, probe-based technologies. These technologies are robust and trusted. However, no current technologies are seamlessly applicable to monitoring chemical processes in real-time on BOTH the high throughput lab scale (the 'teacup') AND process/plant scales (the 'swimming pool'). Instead, current process analytical technologies are oftentimes tied to one specific hardware platform, and each example of such probe-based hardware can only monitor one process at a time. Ultimately, this can produce bottlenecks in analysis, slowing chemical product development and deployment. To address this productivity and chemical data throughput challenge, there is a real drive from R&D budget holders to invest in digital-ready analytical technologies. Computer Vision is the science of digitally quantifying real-world colours and objects using cameras. With cameras and computer vision, and further development through this fellowship renewal, the hardware and software needed for more time-, cost-, and safety-effective monitoring of high-value chemical processes can be realised in an accessible and globally adoptable manner. The global investment for digitalisation of process analytical technology (PAT) in the chemical industry is expected to reach $31 billion by 2028, representing an annual growth rate of approximately 6% from the present $23.5 billion market (sources: Made Smarter Review, 2017. Frost & Sullivan, 2017, and 2022). Underpinning this trend, R&D managers across chemical manufacturing are driving the streamlined adoption of new digital-ready chemical technology, to improve productivity, process safety, and ability to exploit the adjacent evolution of artificial intelligence.

Chiral amines are prevalent in natural products, which often display potent biological activity. Such chiral amine motifs are also frequently found in pharmaceutical drug compounds and chemical building blocks meaning that the development of environmentally benign and sustainable routes to produce these important motifs is extremely desirable. Nature synthesizes these complex and valuable molecules through the action of highly specialized enzymes. These natural catalysts enable an extremely efficient biosynthesis from simple starting materials, installing functional groups with exceptional levels of selectivity. Chemical catalysts are frequently designed to mimic the action of enzymes and are often capable of achieving impressive selectivity. However, unlike enzymes, processes involving these catalysts usually involve high temperatures, sub-optimal pH, organic solvent and complex purification methods. Enzymes called omega-transaminases (TAs) catalyze the conversion of commercially available or easily accessible starting materials to high-value amines. These biocatalysts require an additional donor molecule to provide the amine functional group. This donor is ultimately converted to a by-product and the desired amine product is formed. However, the reaction is freely reversible and unless this by-product is removed from the reaction, low yields of the desired amine will be isolated, as the enzyme will more readily catalyse the reverse reaction to regenerate starting materials. A number of elegant approaches have been reported which remove this ketone by-product and allow access to appreciable quantities of the chiral amine. These strategies include the addition of expensive enzymes or the use of extremely large quantities of the amine donor in combination with the technically challenging removal of ketone by-products. One such approach, which relies on an extensively modified TA, is currently used for the industrial synthesis of the antidiabetic drug compound, sitagliptin. However, the approach is far from efficient and the development of this heavily modified TA biocatalyst was enormously challenging, highlighting an immediate need for more sustainable strategies for performing these biotransformations and for developing suitable enzyme catalysts. This research will build upon recent work reported in our laboratory that describes arguably the most efficient approach to date for performing biotransformations involving TAs. The success of the approach is due to spontaneous precipitation of the by-product, which cannot regenerate starting materials. This polymer is also highly colored and has allowed the development of an effective high-throughput screening strategy that enables the rapid identification of active enzymes. Our focus now is to optimize the process further and make it more suitable for industrial application. Specifically, low cost amine donor molecules will be used that are spontaneously removed from the reaction in a similar way to our previously reported method. We will also apply a simple high-throughput screening strategy to assist in the genetic engineering of natural enzymes in order to increase the scope of the reactions that they can catalyze and make them suitable for industrial scale synthesis. The enzymes developed in this study will enable cost-effective, sustainable and environmentally neutral methods for the small/medium and industrial scale production of one of the most important compound classes.

Currently, most of the manufacturing the high-value chemicals such as agrochemicals and pharmaceuticals, are performed in 'batch' reactors, where the chemical feedstocks (largely petrochemicals based) are converted into the product through a sequence of 'units of operations', which includes several chemical transformations, and purification steps. As the volume of each reactor is fixed, some of these operations, if not the entire sequence, have to be repeated, in order to meet the market demand. Very often, batch-to-batch variation in quality can result, which has to be monitored closely at each stage of the process in order to meet stringent regulatory requirements for product purity. Conversely, in a continuous flow process, the individual units of operation are integrated to enable an uninterrupted flow of material and product. Inline analytics (sensors and detectors) can also be implemented to monitor the quality of the produced product in real-time. As the entire process operates non-stop ('steady state'), the volume of production is no longer limited by the reactor size. Potentially, a continuous process is more efficient in saving costs, energy, and time, without comprising product quality. Traditionally, high-value chemical products, such as agrochemicals and pharmaceuticals, are produced using batch reactors, as they are usually required in small volumes. In more recent years, there are significant economical and sustainability drivers for the chemical industry to adopt the use of continuous flow processes. However, their implementation is not easy; as continuous reactors tend to be less flexible, in terms of modifying them to produce different products. The ambition of the IConIC Partnership is to redesign the continuous process: from a fully-integrated, single-purpose unit, towards a flexible 'plug-and-play' system, where each unit of operation ('module') can be replaced or substituted easily without affecting the overall performance of the continuous process. This will require a better understanding of how the interplay between molecular properties, timescales of reactions (reaction kinetics), and process parameters. For industrial implementation, additional factors (e.g. costs, sustainability and regulatory requirements) also need to be taken into consideration to justify the capital investment needed to switch from batch to flow production. Over the past 5 years, BASF has been working with ICL to foster an active 'Flow Chemistry' community involving 50 researchers at both institutions. The IConIC partnership will not cement the relationship by initiating a programme of exciting and ambition research projects to translate the benefits of Flow Chemistry from the R&D lab into industrial practice. An important aspect is an emphasis on a seamless data flow and translation process across the WPs, including decision-making under uncertainty, multi-fidelity design of experiments, transfer learning, and proof-of-concept demonstration for scale-up. A key feature of IConIC is the inclusion of a number of other UK-based industrial partners to form a 'vertical consortium' along the value chain. Over the period of the grant, the Partnership will be expanded to include additional academic and industrial partners at the appropriate junctures, to leverage synergistic values. Ultimately this will enable the UK to take leadership in continuous flow manufacturing.