Elsevier

Gynecologic Oncology

Volume 116, Issue 1, January 2010, Pages 109-116
Gynecologic Oncology

Liver X receptor agonist inhibits proliferation of ovarian carcinoma cells stimulated by oxidized low density lipoprotein

https://doi.org/10.1016/j.ygyno.2009.09.034Get rights and content

Abstract

Objectives

We previously observed an association between ovarian cancer outcome and statin use and hypothesized lipoproteins have direct effects on ovarian cancer proliferation. Here we investigate the direct effects of low density lipoprotein (LDL) and oxidized LDL (oxLDL) on proliferation and the inhibitory effects of fluvastatin and a liver X receptor (LXR) agonist.

Methods

The effects of LDL, oxLDL, the LXR agonist TO901317, fluvastatin and cisplatin on cellular proliferation were determined using MTT assays. LXR pathway proteins were assayed by immunoblotting. Cytokine expression was determined by antibody array.

Results

Concentrations of oxLDL as small as 0.1 μg/ml stimulated CAOV3 and SKOV3 proliferation, while LDL had no effect. TO901317 inhibited the proliferation of CAOV3, OVCAR3 and SKOV3 cells stimulated by oxLDL. Fluvastatin inhibited oxLDL mediated proliferation of CAOV3 and SKOV3. Cardiotrophin 1 (CT-1) was mitogenic to CAOV3 and SKOV3, was induced by oxLDL, and was reversed by TO901317. OxLDL increased cisplatin IC50s by 3.8 μM and  >  60 μM for CAOV3 and SKOV3 cells, respectively. The LXR pathway proteins CD36, LXR, and ABCA1 were expressed in eight ovarian carcinoma cell lines (A2780, CAOV3, CP70, CSOC882, ES2, OVCAR3, SKOV3).

Conclusions

OxLDL reduced ovarian carcinoma cell chemosensitivity and stimulated proliferation. These effects were reversed by LXR agonist or fluvastatin. The LXR agonist also inhibited expression of the ovarian cancer mitogen CT-1. These observations suggest a biologic mechanism for our clinical finding that ovarian cancer survival is associated with statin use. Targeting LXR and statin use may have a therapeutic role in ovarian cancer.

Introduction

The association between serum cholesterol and cancer risk and survival is unclear. Attempts to correlate risk or outcome of numerous cancers with serum low density lipoprotein (LDL) cholesterol have yielded somewhat controversial results with evidence pointing towards improved outcome in patients with high serum cholesterol [1], [2], [3]. One study also demonstrated that women on a diet high in cholesterol had no increased risk of epithelial ovarian cancer [4]. However, recently it was reported that the levels of oxidized LDL (oxLDL) in the serum of ovarian cancer patients were positively associated with patient outcome [5]. Our group has also recently reported that epithelial ovarian cancer patients on statins had improved survival [6].

Levels of oxLDL are in part regulated by the available oxidizable LDL precursor and may be altered by cholesterol lowering drugs such as statins. Cholesterol homeostatis is maintained in part by cells expressing scavenger receptors (SRA and CD36) that internalize oxLDL which are then converted to oxysterol ligands of the nuclear liver X receptors α and β (LXRα and LXRβ), heterodimers of the retinoid X receptor (RXR) [7]. Activated LXR/RXR heterodimers activate target genes possessing the LXR element (LXRE) including the ATP binding cassette transporters ABCA1 and ABCG1 leading to cholesterol efflux to HDL or cholesterol excretion via bilial and intestinal cells [7]. LXR activation is also associated with increases in the expression of numerous proinflammatory cytokines which also promote cellular proliferation [7]. While statin therapy can lower serum oxLDL [8], [9], [10], [11], statins also have lipid independent functions on oxidative pathways including regulation of scavenger receptors. In monocytes and foam cells, statins can alter scavenger receptor expression via regulation of PPAR, NF-κB, Rho and Ras [12], but these statin functions have not been studied in cancer cells.

One potential means for treating oxLDL related diseases is modulation of LXR by synthetic LXR agonists. Presently three such synthetic LXR ligands have been developed including TO901317, GW3965 and N,N-dimethyl-3β-hydroxycholenamide (DMHCA). LXR agonists upregulate expression of the ATP binding cassette transporters and cholesterol efflux while simultaneously downregulating oxysterol mediated activation of proinflammatory cytokines [13], [14]. In mouse models of atherosclerosis LXR agonists reduce both inflammation in atherosclerotic plaques and serum cholesterol [15]. LXR agonists also inhibit the proliferation of endothelial cells [16]. oxLDL and LXR agonists have been little studied on cells other than endothelial, monocytes and macrophage foam cells in the context of cardiovascular diseases, but one study demonstrated oxLDL is a mitogen to cultured human fibroblasts [17].

In the present study we determined that ovarian carcinoma cells possess CD36 scavenger receptor and are stimulated to proliferate by oxLDL. The LXR agonist TO901317 and fluvastatin reversed oxLDL mediated proliferation. We also demonstrated that oxLDL reduced the sensitivity of ovarian carcinoma cells to cisplatin. Our study demonstrated that increases in oxidized LDL cholesterol may negatively impact ovarian cancer outcome and suggests that LXR ligands and statins may be an effective strategy for treating ovarian cancer patients.

Section snippets

Tissue culture

CAOV3, ES2, OVCAR3, PA1, and SKOV3 were cultured as recommended by American Type Culture Collection. CSOC882 and CSOC909 were cultured as previously described [18]. A2780 and CP70 were cultured in RPMI 1640 and 10% fetal calf serum (FCS) supplemented with 2 mM l-glutamine and 0.2 U/ml insulin. OVCA432 was cultured in modified Eagle's medium (MEM) and 10% FBS + 2 mM l-glutamine. All reagents were purchased from GIBCO.

RNA interference

RNA interference (RNAi) was accomplished by suspension transfection of CAOV3

oxLDL stimulated proliferation of ovarian carcinoma cell lines, but LDL had no effect

We determined the effects of LDL and ox-LDL on proliferation of SKOV3 and CAOV3 ovarian carcinoma cell lines by MTT assays. Increasing doses of LDL did not significantly alter the growth of either of these cell lines (Fig. 1A). However, the proliferation of each of these cell lines was significantly increased upon treatment with oxLDL (Fig. 1B). Increased proliferation was also evident visually by examination of cells treated for 24 h with oxLDL by standard phase microscopy (Fig. 1C).

LXR pathway proteins are expressed in ovarian carcinoma cell lines

We

Discussion

Ovarian cancer outcome may be influenced by genetic, epigenetic, and clinical determinates as well as metabolic conditions that alter cancer proliferation. Recently we demonstrated that ovarian cancer patients taking statins had improved survival [6]. In addition, a recent meta-analysis of 37,248 individuals found reduced cancer incidence for statin users [20]. While not all studies have demonstrated positive associations between serum cholesterol levels and outcome in solid malignancies [1],

Conflict of interest statement

The authors have no conflicts of interest to declare.

Acknowledgments

This work was supported by contributions from the L&S Milken Foundation and an American Cancer Society California Division Early Detection Professorship to BYK.

References (33)

  • D. Pennica et al.

    Cardiotrophin-1: a multifunctional cytokine that signals via LIF receptor-gp 130 dependent pathways

    Cytokine Growth Factor Rev.

    (1996)
  • M. Fritzenwanger et al.

    Cardiotrophin-1 induces interleukin-6 synthesis in human monocytes

    Cytokine

    (2007)
  • S. Zvonic et al.

    Effects of cardiotrophin on adipocytes

    J. Biol. Chem.

    (2004)
  • J.H. Tanne

    Meta-analysis says low LDL cholesterol may be associated with greater risk of cancer

    BMJ

    (2007)
  • A.M. Fiorenza et al.

    Serum lipoprotein profile in patients with cancer. A comparison with non-cancer subjects

    Int. J. Clin. Lab. Res.

    (2000)
  • J.M. Genkinger et al.

    A pooled analysis of 12 cohort studies of dietary fat, cholesterol and egg intake and ovarian cancer

    Cancer Causes Control

    (2006)
  • Cited by (0)

    View full text