Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
Comparative Study
, 154 (9), 3219-27

Growth Hormone Potentiates 17β-estradiol-dependent Breast Cancer Cell Proliferation Independently of IGF-I Receptor Signaling

Affiliations
Comparative Study

Growth Hormone Potentiates 17β-estradiol-dependent Breast Cancer Cell Proliferation Independently of IGF-I Receptor Signaling

Dana L Felice et al. Endocrinology.

Abstract

Estrogen action in mammary gland development and breast cancer progression is tightly linked to the GH/IGF-I axis. Although many of the effects of GH on mammary gland growth and development require IGF-I, the extent to which GH action in breast cancer depends on IGF-I is not known. We examined GH action in a panel of estrogen receptor-positive breast cancer cell lines and found that T47D cells express significant levels of GH receptor and that GH significantly enhances 17β-estradiol (E2)-stimulated proliferation in these cells. GH action in the T47D cells was independent of changes in IGF-I and IGF-I receptor (IGF-IR) expression and IGF-IR signaling, suggesting that GH can exert direct effects on breast cancer cells. Although E2-dependent proliferation required IGF-IR signaling, the combination of GH+E2 overcame inhibition of IGF-IR activity to restore proliferation. In contrast, GH required both Janus kinase 2 and epidermal growth factor receptor signaling for subsequent ERK activation and potentiation of E2-dependent proliferation. Downstream of these pathways, we identified a number of immediate early-response genes associated with proliferation that are rapidly and robustly up-regulated by GH. These findings demonstrate that GH can have important effects in breast cancer cells that are distinct from IGF-IR activity, suggesting that novel drugs or improved combination therapies targeting estrogen receptor and the GH/IGF axis may be beneficial for breast cancer patients.

Figures

Figure 1.
Figure 1.
GH potentiates E2-stimulated proliferation in T47D human breast cancer cells. A, GHR mRNA expression was examined in a panel of ER+ human breast cancer cell lines by QPCR using the ΔΔcycle threshold method with 36B4 as an internal control. B, T47D cells were treated with 500 ng/mL GH, 10 nM E2, or both for 5 days. Cell numbers were determined using the methylene blue assay and are presented as percent of untreated cells on day 0. C, BrdU incorporation was assessed in T47D cells after 72 hours of treatment with GH, E2, or both. D, T47D cells were treated for 5 days as described in B in the absence or presence of 1 μM of the ER antagonist ICI 182,780. Data points or bars with different letters are significantly different (P < .05).
Figure 2.
Figure 2.
GH potentiates E2-stimulated T47D cell proliferation independently of IGF-I expression or IGF-IR activation. A, T47D cells were treated with 500 ng/mL GH for 24 hours. IGF-I mRNA expression was determined by QPCR. B, T47D cells were treated as in A, and conditioned media were collected. Levels of IGF-I protein were measured by ELISA. IGF-I (5 ng/mL) added to media served as a positive control. C, T47D cells were treated with hormones for 24 hours, after which total levels of IGF-IR protein were determined by Western blot. D, T47D cells were pretreated for 2 hours with 2 μM AEW541, an inhibitor of IGF-IR tyrosine kinase activity, followed by 30 minutes of treatment with GH+E2 or 50 ng/mL IGF-I. Phospho- and total IGF-IR levels were examined by Western blot to demonstrate the effectiveness of the inhibitor. E, T47D cells were treated with E2, E2+GH, or E2+IGF-I in the presence of 1 μM AEW541 or vehicle [dimethylsulfoxide (DMSO)] for 5 days. Proliferation was measured by the methylene blue assay. Similar results were found using the DNA assay (data not shown). Bars with different letters are significantly different (P < .05).
Figure 3.
Figure 3.
GH activates STAT5 and ERK in breast cancer cells. A, T47D cells were treated for 30 minutes with GH, E2, or both. Phospho- and total STAT5 and ERK levels were assessed by Western blot. B, T47D cells were pretreated for 2 hours with vehicle (dimethylsulfoxide) or inhibitors for EGFR (AG1478, 15 μM) or JAK2 (AZD1480, 1 μM), followed by treatment with GH+E2 for 30 minutes.
Figure 4.
Figure 4.
Roles of JAK2, ERK, and EGFR in E2- and E2+GH-stimulated proliferation. T47D cells were treated for 5 days with E2, GH, or both in the presence of vehicle [dimethylsulfoxide (DMSO)] or inhibitors for JAK2 (AZD1480, 1 μM) (A), MEK (U0126, 10 μM) (B), or EGFR (AG1478, 15 μM) (C). Proliferation was determined by the DNA assay. Bars with different letters are significantly different (P < .05).
Figure 5.
Figure 5.
GH rapidly up-regulates IEGs in a JAK2-, EGFR-, and ERK-dependent manner. A, T47D cells were treated with GH for up to 6 hours. Expression of IEGs was examined by QPCR. Data are presented as a heat map with the highest levels of expression for each gene in red and the lowest levels in green. B, GH-induced expression of IEGs was examined after pretreatment with inhibitors for EGFR (AG1478), MEK (U0126), or JAK2 (AZD1480).

Similar articles

See all similar articles

Cited by 6 PubMed Central articles

See all "Cited by" articles

Publication types

MeSH terms

Feedback