Development of Novel Therapeutics: From Small Molecules to Biologics

Abstract

The dynamic global health landscape demands innovative therapeutics to address multiple challenges. Broady, therapeutics fall into three main categories: small molecules, peptides and biologics. Small molecules continue to dominate the pharmaceutical landscape due to their ease of synthesis and use, while biologics are emerging as the next-generation tool for high precision therapy and diagnostics, and personalised medicine. Peptides lie at the interface of these two classes and combine the high specificity of biologics with the simplicity of small-molecule drugs, presenting a unique class of therapeutic diagnostics (theranostics). As the demand for targeted, efficient and adaptable medicine grows, the advancement of each class of therapeutics is essential in shaping the future of healthcare (Chapter One). Heparanase is the only known mammalian enzyme to catalyse the hydrolysis of heparan sulfate, a major component of the extracellular matrix. Also, it is upregulated in every human cancer studied to date where it contributes to larger tumour size and metastasis, and is implicated in several other chronic human diseases, such as Crohn's disease and diabetes. However, despite extensive research, no specific heparanase inhibitors have progressed through clinical trials. As such, it is clear that heparanase is an important drug target where traditional drug design methods have failed to produce effective inhibitors. Chapter two explores an emerging method of small molecule drug discovery, known as fragment-based drug discovery (FBDD) and its application to this prominent drug target, where several novel binders and inhibitors of heparanase were discovered and FBDD principles were used to decrease the IC50 of a selected fragment by more than seven-fold. Peptide-based drugs are rapidly emerging as next-generation therapeutics (e.g. Ozempic) due to their favourable characteristics compared to small molecules and biologics. However, many canonical peptides are restricted in their therapeutic applications due to inherent limitations caused by their flexible structures and susceptibility to proteolysis. Macrocyclisation is a commonly employed method to mitigate these limitations. Chapter three introduces oxime ligation, a biocompatible, selective, one-pot macrocyclisation technique that can be applied to a variety of amino acid sequences and is fully amenable to automation. Similarly, multicyclic peptides offer the opportunity to further stabilise peptide structures. Bismuth serves as a simple, single atom cyclisation scaffold that has therapeutic applications in chemotherapy, while also conferring superior structural stability to multicyclic peptides (Chapter Four). Biologics are the largest class of drugs and are becoming increasingly popular for use in personalised and targeted therapies, such as radiotherapy where they are conjugated to a therapeutic isotope. Nanobodies, the smallest naturally occurring antigen-binding fragments, have superior pharmacokinetic properties when compared to their larger counterpart, antibodies. Chapter Five describes the introduction of a single cysteine mutation adjacent to the canonical disulfide bond present in nanobodies, generating a metal binding site. Similarly, affibodies are incredibly small, engineered antigen-binding proteins derived from the antigen-binding domain of staphylococcal surface protein A that have excellent therapeutic properties. In Chapter Six, three cysteine residues are engineered into affibodies to generate a metal-binding site within the protein. These engineered metal-binding proteins form highly stable metal complexes and represent a new class of theranostics with promise for applications in imaging and radiotherapy.