Cell Permeable Cyclic Peptide Library Generation for Target-Based Screening

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Most drugs on the market are small molecules. Whilst these typically have good oral bioavailability, small molecule drugs do not always have a high selectivity for their targets. This can lead to increased toxicity from off-target effects. Likewise, small molecules are unable to selectively target proteins with large binding surfaces, such as those that are involved in protein-protein interactions. Large biologics such as proteins and antibodies exhibit very high selectivity for these targets but are not cell permeable and can thus only bind onto extracellular proteins. Furthermore, biologics are costly to manufacture compared to synthetic molecules. Peptides are promising therapeutics as they lie between the spectrum of small molecules and biologics whereby they have high selectivity for their targets, are able to probe a larger chemical space by targeting proteins with large binding surfaces and have relatively low production costs. However, the issue of low cell permeability, rapid metabolism and thus low oral bioavailability still persists for peptides and is a major limiting factor in their use as therapeutics.It has been observed that cyclisation of peptides has several structural benefits, such as eliminating the N- and C- termini to prevent proteolytic degradation, increasing rigidity and reducing the polar surface area to improve cell permeability and metabolic stability, leading to increased oral bioavailability.Over 1000 naturally occurring cyclic peptides have been discovered and some have been developed into widely used clinically available drugs, such as vancomycin, cyclosporin A and actinomycin D. However, the physicochemical properties underlying cyclic peptide cell permeability are unclear. The design of small molecule drugs is guided by Lipinski's Rule of 5 which defines the physicochemical properties that a drug should have to be orally bioavailable. Such rules include having a molecular weight <500, logP <5, number of hydrogen bond donors <5 and number of hydrogen bond acceptors <10. Due to the larger size of peptides, they do not conform to these rules and thus they cannot be used for the design of orally bioavailable cyclic peptides.This project aims to elucidate the physicochemical properties underlying the cell permeability of cyclic peptides. By ascertaining these properties, they can be used as screening filters in mRNA-display based cyclic peptide libraries to efficiently identify cell permeable cyclic peptide hits against target proteins of interest. This has the potential of vastly improving the landscape for cyclic peptide drug development by eliminating the need for modulating cell permeability of hit peptides, which can be highly time-consuming and difficult. Following the generation of these libraries, they can undergo screening against targets for the discovery of cell permeable peptides with therapeutic potential.