The insulin-like growth factor 1 pathway is a potential therapeutic target for low-grade serous ovarian carcinoma

Abstract

Objective: To validate the overexpression of insulin-like growth factor 1 (IGF-1) and its receptor (IGF-1R) in low-grade serous ovarian carcinoma (SOC), and to investigate whether the IGF-1 pathway is a potential therapeutic target for low-grade SOC.

Methods: Gene expression profiling was performed on serous borderline ovarian tumors (SBOTs) and low-grade SOC, and overexpression of IGF-1 in low-grade SOC was validated by RT-PCR and immunohistochemistry. The effect of exogenous IGF-1 on cell proliferation was determined in cell lines by cell proliferation assays, cell migration assays, and Western blot. Signaling pathways downstream of IGF-1 and the effects of the AKT inhibitor MK-2206 were investigated by Western blot analysis and by generating IGF-1R short hairpin RNA stable knockdown cell lines. Low- and high-grade cell lines were treated with the dual IGF-1R- and insulin receptor-directed tyrosine kinase inhibitor OSI-906, and cellular proliferation was measured.

Results: mRNA analysis and immunostaining revealed significantly higher IGF-1 expression in low-grade SOCs than in SBOTs or high-grade SOCs. In response to exogenous treatment with IGF-1, low-grade cell lines exhibited more intense upregulation of phosphorylated AKT than did high-grade cell lines, an effect that was diminished with IGF-1R knockdown and MK-2206 treatment. Low-grade SOC cell lines were more sensitive to growth inhibition with OSI-906 than were high-grade cell lines.

Conclusions: IGF-1 is overexpressed in low-grade SOCs compared with SBOTs and high-grade SOCs. Additionally, low-grade SOC cell lines were more responsive to IGF-1 stimulation and IGF-1R inhibition than were high-grade lines. The IGF-1 pathway is therefore a potential therapeutic target in low-grade SOC.

Figure 1

Box plot of IGF-1 mRNA expression in 9 serous borderline tumors and 18 low-grade and 15 high-grade SOC samples compared to human ovarian surface epithelium (HOSE). The box is bounded by the 25^th and 75^th percentiles, with the median expression level depicted by the line in the box. Outlying values are drawn individually. Expression of IGF-1 was significantly higher in low-grade SOC than in SBOT (p < 0.0005) or high-grade SOC (p = 0.033).

Figure 2

A-C. Representative examples of IGF-1 immunohistochemical staining of individual paraffin sections from SBOTs (2A), low-grade SOCs (2B), and high-grade SOCs (2C). D. Representative example of pAKT immunohistochemical staining of individual paraffin sections from low-grade SOCs (magnification: 200×).

Figure 3

A. Western blot analysis of pAKT activation in ovarian cancer cell lines treated with exogenous IGF-1. B. Activation of pAKT by IGF-1 in low-grade ovarian cancer cell lines was blocked by the AKT inhibitor MK-2206 in a dose-dependent manner.

Figure 4

IGF-1 no longer activated pAKT after shRNA IGF-1R knockdown. We performed five independent shRNA knockdowns and a nontarget shRNA control knockdown using the low-grade SOC cell line HOC-7. pAKT expression correlated with the extent of IGF-1R knockdown.

Figure 5

Cell proliferation assays in two low-grade SOCs cell lines (HOC-7, MPSC1), two high-grade SOCs cell lines (SKOV3, 2774), and one SBOT (ML46) cell line exposed to exogenous IGF-1 versus controls. Proliferation was higher in treated cells than in control cells for all cell lines, but the difference between control and treatment was most pronounced in the low-grade SOC cell lines.

Figure 6

Cell proliferation assays in two low-grade (HOC-7, MPSC1) and three high-grade (SKOV3, HEYA8, OVCA420) SOC cell lines after treatment with various concentrations of OSI-906 expressed as a percentage of the proliferation in the control cells. Inhibition was most profound in the low-grade SOC cell lines.