Chemical synthesis and biological characterization of insulin prodrugs

Insulin is a miraculous substance that has addressed the needs of millions of insulin-dependent diabetics, but it is an imprecise medicine with sizable potential for life-threatening overdose. There is tremendous need for insulin therapy that can normalize optimal glycemic control and minimize the adverse effects of chronic hyperglycemia. This thesis explores the application of novel peptide-based prodrug chemistry to the discovery of novel insulin analogs with improved therapeutic index. Prodrug chemical synthesis of insulin explored the use of a newly discovered "1+2" synthetic method that avoids the challenges in preparation of insulin analogs by conventional methods. This synthetic approach once refined was employed to identify the preferred site for prodrug attachment. Specific dipeptides were prepared, characterized and demonstrated variable rates in conversion to biologically active insulin analogs, when exposed to physiological buffers. The restoration of native insulin activity was observed by in vitro, in vivo and ex vivo analysis. To extend the time action of select peptide-based insulin prodrugs the respective additional chemical modification represented by site-specific fatty acylation and pegylation was explored. Pegylation proved superior to acylation in its ability to provide a basal insulin action profile when tested in mice. It was concluded that a releasable PEG-prodrug holds the potential for quantitatively delivering insulin in a safely controlled manner that could support enhanced glycemic control and patient convenience.