Characterization of a Novel Primary Mammary Tumor Cell Line Reveals that Cyclin D1 Is Regulated by the Type I Insulin-Like Growth Factor Receptor

Abstract

The importance of type I insulin-like growth factor receptor (IGF-IR) overexpression in mammary tumorigenesis was recently shown in two separate transgenic models. One of these models, the MTB-IGFIR transgenics, was generated in our lab to overexpress IGF-IR in mammary epithelial cells in a doxycycline (Dox)-inducible manner. To complement this transgenic model, primary cells that retained Dox-inducible expression of IGF-IR were isolated from a transgenic mammary tumor. This cell line, RM11A, expressed high levels of IGF-IR, phosphorylated Akt, and phosphorylated extracellular signal -regulated kinase 1/2 in the presence of Dox. IGF-IR overexpression provided the primary tumor cells with a survival advantage in serum-free media and seemed to induce ligand-independent activation of the IGF-IR because RM11A cells cultured in the presence of Dox were largely nonresponsive to exogenous IGFs. IGF-IR overexpression also augmented the growth of RM11A cells in vivo because injection of these cells into mammary glands of wild-type mice produced palpable tumors in 15.8 F 3.4 days when the mice were administered Dox, compared with 57.8 F 6.3 days in the absence of Dox. DNA microarray analysis revealed a number of genes regulated by IGF-IR, one of which was cyclin D1. Suppression of IGF-IR expression in vitro or in vivo was associated with a decrease in cyclin D1 protein, suggesting that at least some of the proliferative actions of IGF-IR are mediated through cyclin D1. Therefore, this article characterizes the first primary murine mammary tumor cell line with inducible IGF-IR expression. These cells provide a powerful in vitro/in vivo model to examine the function of IGF-IR in mammary tumorigenesis.

Reversibility and recurrence of IGF-IR-induced mammary tumors

“…This supports our earlier findings that short-term doxycycline withdrawal (24-72 h) results not only in the rapid downregulation of the IGF-IR transgene but also leads to reduction in the levels of cyclin D1 and activated Akt (Jones et al, 2008). Thus, the early events of tumor regression after IGF-IR suppression involves downregulation of the cell-cycle regulator cyclin D1 and the loss of the pro-survival effects of Akt leading to reduced proliferation and increased apoptosis of tumor cells.…”

“…Generation of MTB-IGFIR transgenic mice and the primary MTB-IGFIR mammary tumor cell line, RM11A, has been described earlier (Jones et al, 2007(Jones et al, , 2008. To induce transgene expression, standard rodent chow was replaced with rodent chow containing 2 g/kg of doxycycline (Bio-Serv, Frenchtown, NJ, USA).…”

Inhibition of proliferation and migration of luminal and claudin-low breast cancer cells by PDGFR inhibitors

“…RJ345 and RJ348 cells were generated in our lab and have previously been described [30]. RJ345 and RJ348 cells were cultured in DMEM containing 10% FBS, 1 mmol/L sodium pyruvate, 10 mmol/L HEPES, 4 mmol/L glutamine, 2 mmol/L hydrocortisone, 5 μg/mL estrogen, 5 μg/mL prolactin, 10 μg/mL epidermal growth factor, 10 μg/mL insulin and 1% antibiotic/antimycotic.…”

Molecular Signatures of the Insulin-Like Growth Factor 1-Mediated Epithelial-Mesenchymal Transition in Breast, Lung and Gastric Cancers

“…This mechanism of action is exerted by pp60 src [57], c-myb [58,59] and hepatitis B virus X oncogenes [60]. Jones et al suggested that in transformed cells overexpressing IGF-1R, the latter’s proliferative/survival signaling can be ligand-independent above a certain expression level, whereas in normal cells in which the receptor is under expressed, IGF stimulation triggers a robust response [61]. Such ligand-independent/dependent signaling may play an instrumental role when IGF-1 is used at supraphysiological doses as a performance-enhancing drug [15,62,63].…”