Peptides from the venomous marine snail Conus victoriae: identification, characterisation and potential applications

Animal venoms represent a vast library of bioactive peptides and proteins with proven potential not only as research tools but also as drug leads and therapeutics. Several drugs based on venom peptides have been approved for the treatment of medical conditions including diabetes, hypertension and chronic pain. An additional ten in clinical trials, as well as several others in preclinical stages of development, have the potential to treat a wider range of conditions, including cancer, HIV, myocardial infarction, stroke, heart failure and multiple sclerosis. Components from the venoms of cone snails (genus Conus) show particular promise, with each Conus venom consisting of a mixture of hundreds of peptides (conotoxins) with a diverse array of molecular targets. Several conotoxins have found applications as important research tools, while some are being used or developed as therapeutics. Chapter 1 is structured as a review of conotoxins in the context of gene superfamily. The overall objective of the research described in this thesis has been the discovery, functional and structural characterisation, and optimisation (for a therapeutic endpoint) of conotoxins from an Australian species of cone snail, Conus victoriae. The initial discovery phase involved the generation of a C. victoriae venom gland transcriptome. To this end, a combination of state-of-the-art techniques in molecular biology and bioinformatics was utilized, including cDNA library normalization, high-throughput 454 sequencing, de novo transcriptome assembly and annotation with BLAST and profile hidden Markov models (pHMMs). Over 100 unique conotoxin sequences from 20 gene superfamilies were discovered (the largest and most diverse Conus venom gland library published to date), as described in Chapter 2. Subsequent matching of the transcriptome to a mass spectrometry profile of the crude venom was used to interrogate venom peptide composition and confirm post-translational modifications present in the mature venom peptides. Promising peptides were chosen for further functional and structural characterisation and produced synthetically. Chapter 3 describes the structural characterisation of one such peptide, contryphan_Vc1 and the subsequent discovery of a new elementary peptide fold and its potential application. Chapter 4 describes the unique method of discovery and interesting activity profile of another novel peptide, CNF-Vc1. One potential drawback of conotoxins as potential therapeutics is that they suffer from the generic problems of peptides in vivo. Significant efforts are being made in modifying peptides to improve their therapeutic potential. In the final chapter of this thesis, this step in venom peptide drug development is illustrated with the modification of α-conotoxin-Vc1.1, an analgesic peptide from the venom of C. victoriae. The approach used was the selective replacement of the native disulfides in the peptide with non-reducible dicarba bridges in an effort to improve its in vivo stability profile. Importantly, this modification resulted in a more favourable selectivity profile for the peptide, which may further improve its therapeutic utility. <div><br></div><div>Awards: Winner of the Mollie Holman Doctoral Medal for Excellence, Faculty of Pharmacy and Pharmaceutical Sciences, 2015.