Minimizing Mitogenic Potency of Insulin Analogues Through Modification of a Disulfide Bond.

The mechanisms by which insulin activates the insulin receptor to promote metabolic processes and cellular growth are still not clear. Significant advances have been gained from recent structural studies in understanding how insulin binds to its receptor. However, the way in which specific interactions lead to either metabolic or mitogenic signalling remains unknown. Currently there are only a few examples of insulin receptor agonists that have biased signalling properties. Here we use novel insulin analogues that differ only in the chemical composition at the A6-A11 bond, as it has been changed to a rigid, non-reducible C=C linkage (dicarba bond), to reveal mechanisms underlying signaling bias. We show that introduction of an A6-A11 cis-dicarba bond into either native insulin or the basal/long acting insulin glargine results in biased signalling analogues with low mitogenic potency. This can be attributed to reduced insulin receptor activation that prevents effective receptor internalization and mitogenic signalling. Insight gained into the receptor interactions affected by insertion of an A6-A11 cis-dicarba bond will ultimately assist in the development of new insulin analogues for the treatment of diabetes that confer low mitogenic activity and therefore pose minimal risk of promoting cancer with long term use.

Author Correction: A structurally minimized yet fully active insulin based on cone-snail venom insulin principles.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

A structurally minimized yet fully active insulin based on cone-snail venom insulin principles.

Human insulin and its current therapeutic analogs all show propensity, albeit varyingly, to self-associate into dimers and hexamers, which delays their onset of action and makes blood glucose management difficult for people with diabetes. Recently, we described a monomeric, insulin-like peptide in cone-snail venom with moderate human insulin-like bioactivity. Here, with insights from structural biology studies, we report the development of mini-Ins-a human des-octapeptide insulin analog-as a structurally minimal, full-potency insulin. Mini-Ins is monomeric and, despite the lack of the canonical B-chain C-terminal octapeptide, has similar receptor binding affinity to human insulin. Four mutations compensate for the lack of contacts normally made by the octapeptide. Mini-Ins also has similar in vitro insulin signaling and in vivo bioactivities to human insulin. The full bioactivity of mini-Ins demonstrates the dispensability of the PheB24-PheB25-TyrB26 aromatic triplet and opens a new direction for therapeutic insulin development.

How IGF-II Binds to the Human Type 1 Insulin-like Growth Factor Receptor.

Human type 1 insulin-like growth factor receptor (IGF-1R) signals chiefly in response to the binding of insulin-like growth factor I. Relatively little is known about the role of insulin-like growth factor II signaling via IGF-1R, despite the affinity of insulin-like growth factor II for IGF-1R being within an order of magnitude of that of insulin-like growth factor I. Here, we describe the cryoelectron microscopy structure of insulin-like growth factor II bound to a leucine-zipper-stabilized IGF-1R ectodomain, determined in two conformations to a maximum average resolution of 3.2 Å. The two conformations differ in the relative separation of their respective points of membrane entry, and comparison with the structure of insulin-like growth factor I bound to IGF-1R reveals long-suspected differences in the way in which the critical C domain of the respective growth factors interact with IGF-1R.

How ligand binds to the type 1 insulin-like growth factor receptor.

Human type 1 insulin-like growth factor receptor is a homodimeric receptor tyrosine kinase that signals into pathways directing normal cellular growth, differentiation and proliferation, with aberrant signalling implicated in cancer. Insulin-like growth factor binding is understood to relax conformational restraints within the homodimer, initiating transphosphorylation of the tyrosine kinase domains. However, no three-dimensional structures exist for the receptor ectodomain to inform atomic-level understanding of these events. Here, we present crystal structures of the ectodomain in apo form and in complex with insulin-like growth factor I, the latter obtained by crystal soaking. These structures not only provide a wealth of detail of the growth factor interaction with the receptor's primary ligand-binding site but also indicate that ligand binding separates receptor domains by a mechanism of induced fit. Our findings are of importance to the design of agents targeting IGF-1R and its partner protein, the human insulin receptor.