Establishing an Approach for the Selection and Design of Secondary Structure Mimetics to Antagonise Protein-protein Interactions.

Protein-protein interactions mediate most biological processes and are therefore important therapeutic targets. The biological activity of a protein usually stems from only a small localised region on its surface. At the molecular level such regions often correspond to key secondary structures known as alpha-helices or beta-sheets that reside within the protein. Creating molecules able to mimic these regions while retaining their structure are attractive options for drug design. However short regions of a protein are usually unable to adopt these structures in the absence of the rest of the protein. Rather, they populate random structures that are susceptible to degradation in addition to other shortcomings such as their inability to cross biological membranes and poor bioavailability.

To circumvent these issues we will collaborate with the Fairlie, a world leader in secondary structure mimetics, to create peptides that are able to form bioactive alpha-helices and beta-sheets in isolation. This will be achieved by introducing helix- or strand-inducing tethers into our growing collection of library derived peptides. Shorter constrained peptides can be derived from larger peptides known to bind with high affinity to their target. Our efforts will focus on two key areas in which we have track record:

i) creating peptides to antagonise the oncogenic transcriptional regulator, Activator Protein-1. We have previously used library screening assays to derive a range of peptides capable of antagonising function. We have already worked with Fairlie to demonstrate feasibility for this approach by targeting one AP-1 partner known as cFos and shedding over 40% of the peptide in the process. Using this approach we were able to derive stable helix-constrained peptides specific for their target protein that also resisted degradation (Rao et al, PLOS One 2013). We believe that much high affinity interactions can be achieved by targeting another AP-1 component, known as cJun, where many more hydrophobic interactions required for high binding affinity can be formed. Previous related work has demonstrated that this approach can yield tethered peptides as short as five amino acids (Harrison et al, PNAS, 2010) that are able to meet many of the requirements necessary for a drug, such as high stability and resistance to biological breakdown.

ii) Creating peptides capable of modulating amyloid formation. We have used library screening to derive small beta-strand peptides that bind to the Alzheimer's beta-amyloid peptide (Acerra et al, Protein Eng Des Sel 2013). We now seek to collaborate with Fairlie in creating mimetics of these short peptides that result in improved compounds that are able to circumvent many of the above issues.

To achieve these goals Mason will travel to the Institute for Molecule Bioscience (IMB) at the University of Queenland on three visits over three years to further develop our collaboration with the Fairlie group. Fairlie is internationally known as a research and opinion leader in chemistry, biochemistry, pharmacology, and drug discovery. The award will permit Mason to gain new skills and techniques that can be brought back to Essex and further developed in the UK, in addition to the exchange of ideas and the further development of the collaboration. Having developed methods for stabilising alpha-helices and beta-strands in general there will be considerable scope to apply these techniques, and consequent rules for peptide and peptide mimetic design, to other peptide systems. Finally while at Queensland there will also be ample opportunity to hold seminars and meet and discuss research plans with other members of the IMB (e.g. Professors Glenn King and David Craik) who have similar interests in developing peptide-based drugs.

In this proposal we seek to travel to Queensland and learn how to create a number of different constraints within peptides using solid-phase peptide synthesis. These skills can be brought back to the UK. In addition the visits will allow us to collaborate more closely with Fairlie in creating small potent inhibitors of the cJun component of the AP-1 protein and beta-amyloid systems. This will involve rational design based on previous scaffolds, comparison of constrained/unconstrained sequences and screening of constrained helical libraries using an intracellular screening process called PCA. These initial peptides and mimetics will deliver the project to the preclinical stage by providing a scaffold from which new antagonising compounds can be developed. This is a standard approach to developing new treatments. There is clearly enormous potential benefit to the public sector health provision costs. This research offers the potential to derive such a treatment, which as well as benefiting the public sector, will potentially transform the approach used to derive new inhibitors, by providing a pre-defined intracellular screen for efficacy.

We also expect that our research will lead to considerable benefits for academics attempting to generate inhibitors of protein interactions both for therapeutics and non-therapeutics such as novel functional self-assembling structures for application in medical and material sciences. Furthermore the acceleration of the drug design process gained by generating novel helix-constrained and strand-constrained peptides can serve as the starting point for therapeutics against other disease processes involving a huge variety of alpha-helix/beta-strand based PPIs. This has considerable commercial benefit as these advances from the basic level of understanding are exploited at the commercial level, carrying with it considerable benefits to the knowledge economy of the UK.

All findings will be published in high impact publications to disseminate with the wider scientific community. In addition, seminars will be hosted for scientists (invited seminars and meetings) and the general public (e.g. cafe scientifique, schools days at Essex, Essex 'Open House'). In addition impact can be delivered via our Wellcome Trust People Award which is developing an electronic game for mobile devices (iOS and Android) to educate about the pathology of Alzheimer's Disease and could be later applied to the AP-1 system. This is being accompanied by a series of talks and presentations (e.g. Bristol Watershed, Bigbang@Essex 2013, BigBang 2014) as well as featured on the Wellcome Trust website.

Communications outside the university will be aided by the Communications Office at Essex who liaise with journalists for radio/television and other media pieces, and who also produce their own monthly publication 'Wyvern'. A dedicated website will report on project developments and be fully accessible to readers of all levels. Relevant media pieces including podcasts and vodcasts will be placed onto the site.