Insulin analogues differently activate insulin receptor isoforms and post-receptor signalling

Abstract

Aims/hypothesis Five insulin analogues, with modified insulin-like molecular structures, are currently approved for treating diabetic patients. They activate cell signalling and biological responses via insulin receptor isoforms (IR-A and IR-B), each having specific characteristics for eliciting cell responses. The molecular and biological effects of these analogues on receptor isoforms in comparison to native insulin are not well defined, and their effects on the IGF1 receptor (IGF1R) are controversial. The characterisation of these effects was the aim of the present study. Methods Short-acting (insulin lispro [B28Lys,B29Pro human insulin], insulin aspart [B28Asp human insulin], insulin glulisine [B3Lys,B29Glu human insulin]) and long-acting (insulin glargine [A21Gly,B31Arg,B32Arg human insulin], insulin detemir [B29Lys(ε-tetradecanoyl),desB30 human insulin]) insulin analogues were studied in three engineered cell models (R − , IGF1R-deprived mouse fibroblasts transfected with either only human IR-A or IR-B or IGF1R). Receptor binding and phosphorylation, AKT and extracellular signal-regulated kinase (ERK) activation, cell proliferation and colony formation were evaluated after exposing the cells to each analogue and were compared with insulin, IGF1 and the carcinogenic analogue B10Asp.Results All short-acting insulin analogues produced molecular and biological effects similar but not identical to those of insulin. Relative to insulin, long-acting analogues more strongly activated the ERK pathway via both IR-A and IGF1R as well as increased cell proliferation. At the concentration tested, no analogue (except B10Asp via IR-A) had increased transforming activity. Conclusions/interpretation Cell models that permit comparisons of the activity of insulin to that of insulin analogues via each receptor individually indicate that only minor differences exist between insulin and short-acting analogues. By contrast, long-acting analogues activate the mitogenic signalling pathway more effectively than insulin and cause increased cell proliferation.

Insulin X10 revisited: a super-mitogenic insulin analogue

“…An asymmetrical preferential phosphatidylinositol 3-kinase (PI3K) pathway activation and, specifically, stimulation of protein translation, was seen after stimulation with insulin X10 [29]. Other studies have also found an important role of the PI3K pathway [25], whereas yet more results seem to favour a more pronounced stimulation of the extracellular signal-regulated kinase (ERK) pathway [30]. Thus, while it is apparent that insulin X10 and other ligands can induce a signalling pattern different from that of insulin, the exact pattern is not entirely obvious and, clearly, more studies comparing the relative activation of the different pathways are needed.…”

“…2, but in addition the issue of different IR isoforms and IR/IGF-1R hybrids must be taken into account. It has been speculated that an increase in the mitogenic properties of an insulin analogue could alternatively (or additionally) reflect a binding preference for the shorter IR-A isoform of IR relative to the longer IR-B isoform [30,33]. This hypothesis is derived from the observation that IR-A has high affinity for binding IGF-2 and is extensively expressed in fetal tissue, where it mediates growth responses.…”

“…This raises the possibility that this IR isoform may be relevant for the mitogenesis of cancer cells [33]. However, studies have shown that both the binding affinity and the activation of the two IR isoforms are very similar after stimulation with insulin X10 [22,30,34,35]. Cells with both IR and IGF-1R will also express heterodimer hybrid receptors [36,37].…”

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“…IGF-1R knockout substantially reduced the proliferative responses of these cells to insulin glargine, which elicited responses equipotent to those of human insulin [37] . Conversely, insulin glargine and insulin detemir were both able to stimulate the growth of mouse fi broblasts to greater extent than native insulin in the presence of either IR-A, IR-B or IGF-1R; IGF-1 stimulated proliferation only in cells containing IGF-1R [34] . However, the fi ndings of Shukla et al are inconsistent with those reported by Staiger et al [38] , who compared the mitogenic potency ( 3 H-thymidine incorporation) of insulin glargine, human insulin and a negative control in MCF-7 and MCF-10 cells.…”

“…Consequently, Asp B10 insulin could potentially stimulate proliferation of IR-A-expressing tumor cells. A direct test of this questions was provided by Sciacca et al [34] , who studied the binding and action of multiple insulin analogues in mouse fi broblasts engineered to express either IR-A, IR-B, or IGF-1R exclusively. While Asp B10 insulin bound to both IR-A and IR-B with a higher affi nity than human insulin, the long-acting analogues insulin glargine and insulin detemir displayed lower affi nities than human insulin for IR-A; both bound to IGF-1R with affi nities ~ 100-fold lower than IGF-1.…”