title: MCF-7 human mammary adenocarcinoma cells exhibit augmented responses to human insulin on a collagen IV surface.
abstract: Human mammary cell lines are extensively used for preclinical safety assessment of insulin analogs. However, it is essentially unknown how mitogenic responses can be optimized in mammary cell-based systems. We developed an insulin mitogenicity assay in MCF-7 human mammary adenocarcinoma cells, under low serum (0.1% FCS) and phenol red-free conditions, with 3H thymidine incorporation as endpoint. Based on EC50 values determined from 10-fold dilution series, beta-estradiol was the most potent mitogen, followed by human IGF-1, human AspB10 insulin and native human insulin. AspB10 insulin was significantly more mitogenic than native insulin, validating the ability of the assay to identify hypermitogenic human insulin analogs. With MCF-7 cells on a collagen IV surface, the ranking of mitogens was maintained, but fold mitogenic responses and dynamic range and steepness of dose-response curves were increased. Also, PI3K pathway activation by insulin was enhanced on a collagen IV surface. This study provided the first determination and ranking of the mitogenic potencies of standard reference compounds in an optimized MCF-7 assay. The optimized MCF-7 assay described here is of relevance for in vitro toxicological testing and carcinogenicity safety assessment of new insulin compounds.
authors.affiliation: Copenhagen University, Institute for Pharmacology and Pharmacotherapy, Universitetsparken 2, 2100 København Ø, Denmark.
authors.forename: Nicolai
authors.initials: N
authors.lastname: Listov-Saabye
authors.forename: Marianne Blirup
authors.initials: MB
authors.lastname: Jensen
authors.forename: Benedicte
authors.initials: B
authors.lastname: Kiehr
authors.forename: Erik W
authors.initials: EW
authors.lastname: Hansen
authors.forename: Jette E
authors.initials: JE
authors.lastname: Svendsen
authors.forename: Anders
authors.initials: A
authors.lastname: Lundby
authors.forename: Gitte-Mai Nelander
authors.initials: GM
authors.lastname: Holm
authors.forename: Martin B
authors.initials: MB
authors.lastname: Oleksiewicz