A framework for the in vitro evaluation of cancer-relevant molecular characteristics and mitogenic potency of insulin analogues.

Abstract

Epidemiological and laboratory studies raised the possibility of a link between clinically prescribed insulin analogues and increased cancer risk. Accordingly, there is a regulatory mandate for cancer-related pre-clinical safety evaluation during insulin analogue development, but currently, there is no standardised framework for such in vitro evaluation. We tested human insulin; the super-mitogenic insulin, X10; and IGF-I, in four cancer cell lines with a range of IGF-IR/IR ratios (HCT 116, HT-29, COLO 205, MCF7), and related these to IGF-IR and IR expression in 17 human adenocarcinomas. All cell types were IRA isoform dominant. We determined IGF-IR/IR signalling pathway endpoints in dose- and time-varying experiments, and performed mitogenic dose-response equivalent assays to derive EC50 values, and correlated these with IGF-IR/IR ratios. We superimposed relative EC50 values onto data from the literature in a meta-analysis. The IGF-IR/IR ratios varied from < 1 to 12 in the selected cell lines; similar pattern ranges were observed in human adenocarcinomas. The three ligands demonstrated differential IR/IGF-IR and Akt phosphorylation, which correlated with cell-specific IGF-IR/IR ratios. Mitogenic profiles of X10 mimicked those for IGF-I, and correlated with IGF-IR/IR ratios. The meta-analysis, adding data from 5 additional studies, supported the hypothesis that ligand mitogenic potency, relative to human insulin, increases with increasing cell-specific IGF-IR/IR ratio. This study established a framework for the in vitro evaluation of cancer-relevant bio-assays for comparisons of insulin analogues, and specifically consolidated earlier studies that determination of the cell-specific IGF-IR/IR ratio is crucial for the interpretation of ranking relative biological activities.