title: Surface-expressed insulin receptors as well as IGF-I receptors both contribute to the mitogenic effects of human insulin and its analogues.
abstract: There is a medical need for new insulin analogues. Yet, molecular alterations to the insulin molecule can theoretically result in analogues with carcinogenic effects. Preclinical carcinogenicity risk assessment for insulin analogues rests to a large extent on mitogenicity assays in cell lines. We therefore optimized mitogenicity assay conditions for a panel of five cell lines. All cell lines expressed insulin receptors (IR), IGF-I receptors (IGF-IR) and hybrid receptors, and in all cell lines, insulin as well as the comparator compounds X10 and IGF-I caused phosphorylation of the IR as well as IGF-IR. Insulin exhibited mitogenicity EC(50) values in the single-digit nanomolar to picomolar range. We observed correlations across cell types between (i) mitogenic potency of insulin and IGF-IR/IR ratio, (ii) Akt phosphorylation and mitogenic potency and (iii) Akt phosphorylation and IR phosphorylation. Using siRNA-mediated knockdown of IR and IGF-IR, we observed that in HCT 116 cells the IR appeared dominant in driving the mitogenic response to insulin, whereas in MCF7 cells the IGF-IR appeared dominant in driving the mitogenic response to insulin. Together, our results show that the IR as well as IGF-IR may contribute to the mitogenic potency of insulin. While insulin was a more potent mitogen than IGF-I in cells expressing more IR than IGF-IR, the hyper-mitogenic insulin analogue X10 was a more potent mitogen than insulin across all cell types, supporting that the hyper-mitogenic effect of X10 involves the IR as well as the IGF-IR. These results are relevant for preclinical safety assessment of developmental insulin analogues.
authors.affiliation: Department of Biomedical Sciences, University of Copenhagen, Blegdamsvej 3, 2200, Copenhagen N, Denmark.
authors.forename: Anders
authors.initials: A
authors.lastname: Lundby
authors.affiliation: Diabetes Research Unit, Novo Nordisk A/S, Novo Nordisk Park, 2760, Maaloev, Denmark.
authors.forename: Pernille
authors.initials: P
authors.lastname: Bolvig
authors.affiliation: Diabetes Research Unit, Novo Nordisk A/S, Novo Nordisk Park, 2760, Maaloev, Denmark.
authors.forename: Anne Charlotte
authors.initials: AC
authors.lastname: Hegelund
authors.affiliation: Diabetes Research Unit, Novo Nordisk A/S, Novo Nordisk Park, 2760, Maaloev, Denmark.
authors.forename: Bo F
authors.initials: BF
authors.lastname: Hansen
authors.affiliation: Diabetes Research Unit, Novo Nordisk A/S, Novo Nordisk Park, 2760, Maaloev, Denmark.
authors.forename: Jesper
authors.initials: J
authors.lastname: Worm
authors.affiliation: Diabetes Research Unit, Novo Nordisk A/S, Novo Nordisk Park, 2760, Maaloev, Denmark.
authors.forename: Anne
authors.initials: A
authors.lastname: Lützen
authors.affiliation: Department of Biomedical Sciences, University of Copenhagen, Blegdamsvej 3, 2200, Copenhagen N, Denmark.
authors.forename: Nils
authors.initials: N
authors.lastname: Billestrup
authors.affiliation: Diabetes Research Unit, Novo Nordisk A/S, Novo Nordisk Park, 2760, Maaloev, Denmark.
authors.forename: Christine
authors.initials: C
authors.lastname: Bonnesen
authors.affiliation: Centre for Biosecurity and Biopreparedness, Statens Serum Institute, Artillerivej 5, 2300, Copenhagen S, Denmark.
authors.forename: Martin B
authors.initials: MB
authors.lastname: Oleksiewicz
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004797:Enzyme-Linked Immunosorbent Assay; D045325:HCT116 Cells / Q000187:drug effects; D056945:Hep G2 Cells / Q000187:drug effects; D006801:Humans; D007328:Insulin / Q000031:analogs & derivatives / Q000494:pharmacology*; D061986:MCF-7 Cells / Q000187:drug effects; D008938:Mitosis / Q000187:drug effects*; D051381:Rats; D017526:Receptor, IGF Type 1 / Q000378:metabolism*; D011972:Receptor, Insulin / Q000378:metabolism*
chemical_list: D007328:Insulin; D017526:Receptor, IGF Type 1; D011972:Receptor, Insulin
keywords: IGF-I; IGF-I receptor; hybrid receptor; insulin; insulin analogues; insulin receptor; mitogenicity