Metrics & MoreArticle RecommendationsCONSPECTUS: Insulin has remained indispensable in the treatment of diabetes since it was first discovered in 1921. Unlike small molecular drugs, insulin and other protein drugs are prone to degradation when exposed to elevated temperatures, mechanical agitation during transportation, and prolonged storage periods. Therefore, strict cold-chain management is crucial for the insulin supply, requiring significant resources, which can limit the access to insulin, particularly in lowincome areas. Moreover, although insulin formulations have advanced tremendously in the last century, insulin treatment still imposes a challenging regimen and provides suboptimal outcomes for the majority of patients. There is an increasing focus on pursuing improved pharmacology, specifically on safer, more user-friendly insulin therapies that minimize the self-management burden. These challenges underscore the need for developing novel insulin formulations with improved stability that are compatible with advanced insulin therapy. Insulin stabilization can be achieved through either chemical modification of insulin or formulation component design. Inspired by insulin-like peptides from invertebrates, we have developed novel stable insulin analogs based on a fundamental understanding of the insulin receptor engagement for insulin bioactivity. We created a novel four-disulfide insulin analog with high aggregation stability and potency by introducing a fourth disulfide bond between a C-terminal extended insulin A-chain and residues near the C-terminus of the B-chain. In an effort to stabilize insulin in its monomeric state to develop ultrafast-acting insulin with rapid absorption upon injection, we have developed a series of structurally miniaturized yet fully active insulin analogs that do not form dimers due to the lack of the canonical B-chain C-terminal octapeptide. Additionally, our study provided strategies for expanding the scope of cucurbit[7]uril (CB[7])-assisted insulin stabilization by engineering safe and biodegradable CB[7]-zwitterionic polypeptide excipients. We also explored insulin N-terminal substitution methods to achieve pH-dependent insulin stabilization without prolonging the duration of action. This Account describes our exploration of engineering stable insulin analogs and formulation design strategies for stabilizing insulin in aqueous solutions. Beyond conventional stabilization strategies for insulin injections, the unmet challenges and recent innovations in insulin stabilization are discussed, addressing the growing demand for alternative, less invasive routes of insulin administration. Additionally, we aim to provide a thorough overview of insulin stabilization from the perspective of commercially available insulin drugs and common pharmaceutical engineering practices in the industry. We also highlight unresolved insulin stabilization challenges and ongoing research strategies. We anticipate that further emphasis on collective efforts of protein engineering, pharmaceutical form...
