title: A framework for the in vitro evaluation of cancer-relevant molecular characteristics and mitogenic potency of insulin analogues.
abstract: Epidemiological and laboratory studies raise 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 standardized framework for such in vitro evaluation. We tested human insulin; the super-mitogenic insulin, X10 and insulin-like growth factor I, in four cancer cell lines with a range of insulin-like growth factor-I receptor (IGF-IR)/IR (insulin receptor) ratios (HCT 116, HT-29, COLO 205 and MCF7) and related these to IGF-IR and IR expression in 17 human adenocarcinomas. All cell types were IR-A 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 insulin-like growth factor I (IGF-I) and correlated with IGF-IR/IR ratios. The meta-analysis, adding data from five 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 bioassays 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.
authors.affiliation: Faculty Institute of Cancer Sciences, Manchester Academic Health Science Centre, The Christie NHS Foundation Trust, University of Manchester, Wilmslow Road, Manchester M20 4BX, UK, Inositide Laboratory, Cancer Research UK Manchester Institute, Paterson Building, Manchester M20 4BX, UK and Diabetes Research Unit, Novo Nordisk A/S, Måløv, Denmark.
authors.forename: Ivona
authors.initials: I
authors.lastname: Baricevic
authors.affiliation: Inositide Laboratory, Cancer Research UK Manchester Institute, Paterson Building, Manchester M20 4BX, UK and.
authors.forename: David R
authors.initials: DR
authors.lastname: Jones
authors.affiliation: Faculty Institute of Cancer Sciences, Manchester Academic Health Science Centre, The Christie NHS Foundation Trust, University of Manchester, Wilmslow Road, Manchester M20 4BX, UK, Inositide Laboratory, Cancer Research UK Manchester Institute, Paterson Building, Manchester M20 4BX, UK and Diabetes Research Unit, Novo Nordisk A/S, Måløv, Denmark.
authors.forename: Darren L
authors.initials: DL
authors.lastname: Roberts
authors.affiliation: Diabetes Research Unit, Novo Nordisk A/S, Måløv, Denmark.
authors.forename: Anne
authors.initials: A
authors.lastname: Lutzen
authors.affiliation: Diabetes Research Unit, Novo Nordisk A/S, Måløv, Denmark.
authors.forename: Anders
authors.initials: A
authors.lastname: Lundby
authors.affiliation: Diabetes Research Unit, Novo Nordisk A/S, Måløv, Denmark.
authors.forename: Jesper
authors.initials: J
authors.lastname: Worm
authors.affiliation: Diabetes Research Unit, Novo Nordisk A/S, Måløv, Denmark.
authors.forename: Bo F
authors.initials: BF
authors.lastname: Hansen
authors.affiliation: Faculty Institute of Cancer Sciences, Manchester Academic Health Science Centre, The Christie NHS Foundation Trust, University of Manchester, Wilmslow Road, Manchester M20 4BX, UK, Inositide Laboratory, Cancer Research UK Manchester Institute, Paterson Building, Manchester M20 4BX, UK and Diabetes Research Unit, Novo Nordisk A/S, Måløv, Denmark andrew.renehan@ics.manchester.ac.uk.
authors.forename: Andrew G
authors.initials: AG
authors.lastname: Renehan
mesh_terms: D000230:Adenocarcinoma / Q000473:pathology; D017209:Apoptosis / Q000235:genetics; D001943:Breast Neoplasms / Q000453:epidemiology / Q000235:genetics / Q000473:pathology*; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000453:epidemiology / Q000235:genetics / Q000473:pathology*; D005260:Female; D045325:HCT116 Cells; D019073:HT29 Cells; D006801:Humans; D007328:Insulin / Q000031:analogs & derivatives*; D007334:Insulin-Like Growth Factor I / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D061986:MCF-7 Cells; D010766:Phosphorylation; D020033:Protein Isoforms / Q000096:biosynthesis / Q000235:genetics; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D017526:Receptor, IGF Type 1 / Q000235:genetics; D011972:Receptor, Insulin / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction
chemical_list: C583557:IGF1 protein, human; D007328:Insulin; D020033:Protein Isoforms; D007334:Insulin-Like Growth Factor I; D017526:Receptor, IGF Type 1; D011972:Receptor, Insulin; D051057:Proto-Oncogene Proteins c-akt