The effect of statin treatment on intratumoral cholesterol levels and LDL receptor expression: a window-of-opportunity breast cancer trial

Abstract

Background: Deregulated lipid metabolism is common in cancer cells and the mevalonate pathway, which synthesizes cholesterol, is central in lipid metabolism. This study aimed to assess statin-induced changes of the intratumoral levels of cholesterol and the expression of the low-density lipoprotein receptor (LDLR) to enhance our understanding of the role of the mevalonate pathway in cancer cholesterol metabolism.

Methods: This study is based on a phase II clinical trial designed as a window-of-opportunity trial including 50 breast cancer patients treated with 80 mg of atorvastatin/day for 2 weeks, between the time of diagnosis and breast surgery. Lipids were extracted from frozen tumor tissue sampled pre- and post-atorvastatin treatment. Intratumoral cholesterol levels were measured using a fluorometric quantitation assay. LDLR expression was evaluated by immunohistochemistry on formalin-fixed paraffin-embedded tumor tissue. Paired blood samples pre- and post-atorvastatin were analyzed for circulating low-density lipoprotein (LDL), high-density lipoprotein (HDL), apolipoprotein A1, and apolipoprotein B. In vitro experiments on MCF-7 breast cancer cells treated with atorvastatin were performed for comparison on the cellular level.

Results: In the trial, 42 patients completed all study parts. From the paired tumor tissue samples, assessment of the cholesterol levels was achievable for 14 tumors, and for the LDLR expression in 24 tumors. Following atorvastatin treatment, the expression of LDLR was significantly increased (P = 0.004), while the intratumoral levels of total cholesterol remained stable. A positive association between intratumoral cholesterol levels and tumor proliferation measured by Ki-67 expression was found. In agreement with the clinical findings, results from in vitro experiments showed no significant changes of the intracellular cholesterol levels after atorvastatin treatment while increased expression of the LDLR was found, although not reaching statistical significance.

Conclusions: This study shows an upregulation of LDLR and preserved intratumoral cholesterol levels in breast cancer patients treated with statins. Together with previous findings on the anti-proliferative effect of statins in breast cancer, the present data suggest a potential role for LDLR in the statin-induced regulation of breast cancer cell proliferation.

Trial registration: The study has been registered at ClinicalTrials.gov (i.e., ID number: NCT00816244 , NIH), December 30, 2008.

Keywords: Breast cancer; Cholesterol; LDL receptor; Statin.

Conflict of interest statement

No conflicts of interest were disclosed by the other authors.

Figures

Fig. 1

Intracellular cholesterol homeostasis. When intracellular cholesterol levels are low, SREBP-2 is delivered to the Golgi where the active, N-terminal fragment is released and translocated to the nucleus where it activates the expression of cholesterol-related genes, such as HMGCR and the LDL receptor. The transcriptional activation of HGMCR leads to the de novo synthesis of cholesterol via the mevalonate pathway. The activation of the transcription of the LDLR leads to an increase in cellular cholesterol uptake through receptor-mediated endocytosis of LDL. When cholesterol levels are high, SREBP-2 is retained in the ER. In order to prevent over-accumulation of free cholesterol in the plasma and intracellular membranes, cholesterol is converted to cholesteryl esters primarily by the enzyme ACAT. Cholesteryl esters are stored as cytosolic lipid droplets. Excess cholesterol also generates oxysterols, natural ligands for LXRs. Their binding to LXRs activates the transcription of genes involved in cholesterol efflux, including ABCA1, ABCG1, and ABCG5/8. This figure was drawn by the author M. Feldt using the image bank of Servier Medical Art. URL to the images are https://smart.servier.com/category/cellular-biology/intracellular-components. Servier Medical Art by Servier is licensed under a Creative Commons Attribution 3.0 Unported License. https://creativecommons.org/licenses/by/3.0/

Fig. 2

Paired samples total cholesterol levels. Total cholesterol levels in tumor tissue were measured using a cholesterol assay before and after 2 weeks of treatment with 80 mg atorvastatin daily. The tumor tissue total cholesterol content was higher in 11 of the 14 paired samples after 2 weeks of atorvastatin treatment lower than before treatment in the remaining three cases. No statistically significant differences in the levels of total cholesterol were observed [P = 0.11 (Wilcoxon signed-rank test)]

Fig. 3

Correlation between tumor tissue total cholesterol and Ki-67. a Between pre-treatment tumoral total cholesterol and pre-treatment Ki-67, a non-significant positive correlation was found. b Between post-treatment tumoral total cholesterol and post-treatment Ki-67, a significant positive correlation was found

Fig. 4

Change in tumor tissue LDLR score. Change in tumor expression of LDLR from baseline (i.e., before atorvastatin treatment) to time of surgery (i.e., after atorvastatin treatment). A significant increase in the expression of the LDLR was found (P = 0.004, Wilcoxon matched-pairs signed-rank test)

Fig. 5

Lipid droplets in MCF-7 cells. Lipid droplets in MCF-7 cells treated with 5 or 10 μM atorvastatin for 24, 48, or 72 h, respectively, compared to untreated control. A concentration- and time-dependent increase in the abundance of intracellular LDs was observed. Values are expressed as the geometric mean ± 95% confidence interval of the geometric mean of three independent experiments