Exploration and Elaboration of Reported Insulin Superagonism through Site-Selective Replacement at TyrB26

Abstract

Introduction Insulin has virtually the universal ability to lower blood glucose and is currently used in multiple forms by millions of patients in the treatment of diabetes. DNA technology has facilitated the biosynthesis of insulin and various related analogs in unlimited quantity. Nonetheless, the relatively low potency of insulin renders it a unique commercial challenge where yearly production is measured in tonnage. The low inherent potency of insulin is a significant obstacle in the development of noninvasive methods for insulin administration since low bioavailability is commonly reported. A recent report [1] outlined replacements for TyrB26 in a C-terminally shortened insulin analog that yielded unprecedented increases in in vitro potency. Bioactivity constituted measurement of glucose transport in isolated primary rat adipocytes and insulin receptor binding in plasma membranes. N-methylHisB26, des(27-30) insulin prepared via semi-synthesis was reported to possess increased potency relative to native insulin, in excess of fifty-fold in receptor binding and tenfold in stimulating glucose uptake. We have explored this observation through synthesis of the same insulin analog by chain combination with totally synthetic A and B chains. Bioactivity was measured relative to insulin in engineered cells that over-express the human insulin receptor. Additionally, we have prepared a number of more acidic B26 insulin analogs to further elaborate the molecular basis for the superagonism.