Investigating the role of endogenous estrogens, hormone replacement therapy, and blockade of estrogen receptor-α activity on breast metabolic signaling

Abstract

Purpose: Menopause is associated with an increased risk of estrogen receptor-positive (ER +) breast cancer. To characterize the metabolic shifts associated with reduced estrogen bioavailability on breast tissue, metabolomics was performed from ovary-intact and ovariectomized (OVX) female non-human primates (NHP). The effects of exogenous estrogen administration or estrogen receptor blockade (tamoxifen treatment) on menopause-induced metabolic changes were also investigated.

Methods: Bilateral ovariectomies were performed on female cynomolgus macaques (Macaca fascicularis) to model menopause. OVX NHP were then divided into untreated (n = 13), conjugated equine estrogen (CEE)-treated (n= 13), or tamoxifen-treated (n = 13) subgroups and followed for 3 years. Aged-matched ovary-intact female NHP (n = 12) were used as a premenopausal comparison group. Metabolomics was performed on snap-frozen breast tissue.

Results: Changes in several different metabolic biochemicals were noted, particularly in glucose and fatty acid metabolism. Specifically, glycolytic, Krebs cycle, acylcarnitines, and phospholipid metabolites were elevated in breast tissue from ovary-intact NHP and OVX + CEE in relation to the OVX and OVX + tamoxifen group. In contrast, treatment with CEE and tamoxifen decreased several cholesterol metabolites, compared to the ovary-intact and OVX NHP. These changes were accompanied by elevated bile acid metabolites in the ovary-intact group.

Conclusion: Alterations in estrogen bioavailability are associated with changes in the mammary tissue metabolome, particularly in glucose and fatty acid metabolism. Changes in these pathways may represent a bioenergetic shift in gland metabolism at menopause that may affect breast cancer risk.

Keywords: Breast cancer; Conjugated equine estrogen; Estrogen; Hormone replacement therapies; Non-targeted metabolomics; Post-menopausal; Tamoxifen.

Fig. 1

Volcano plots showing the distribution of metabolites by group. a Volcano map showing the distribution of metabolites in the ovary-intact (non-ovariectomized; no-OVX) vs ovariectomized (OVX) groups. b Volcano map showing the distribution of metabolites in the OVX + CEE-treated group vs. the OVX group. c Volcano map showing the distribution of metabolites in the OVX + TAM-treated group vs. OVX group

Fig. 2

Changes in glucose metabolites associated with estrogen availability. a Glycolysis pathway. b Heat map showing effects of estrogens on glycolysis metabolites in mammary glands. c Differences in glucose metabolite across groups. d Differences in pyruvate metabolite across groups. e Differences in 3-phosphoglycerate across groups. f Differences in phosphoenolpyruvate across groups. g Differences in glycerate across groups. h Differences in glucose-6-phosphate across groups (p ≤ 0.05)

Fig. 3

Changes in nucleotide metabolism mediated by estrogen. a Nucleotide metabolism pathway. b Heat map showing effects of estrogens on nucleotide sugar metabolites in mammary glands. c Differences in UDP-glucose across groups. d Differences in UDP-galactose across groups. e Differences in UDP-glucuronate across groups. f Differences in N-acetylglucosamine/galactosamine across groups (p ≤ 0.05)

Fig. 4

Changes in central energy metabolism associated with estrogen status. a Tricarboxylic acid cycle (TCA) pathway. b Heat map showing effects of estrogens on TCA metabolites in mammary glands. c Differences in isocitrate across groups. d Differences in α-ketoglutarate across groups. e Differences in succinate across groups. f Changes in malate across groups. g Changes in fumarate across groups. (p ≤ 0.05)

Fig. 5

Changes in bile acid metabolism associated with estrogen bioavailability. a Bile acid synthesis pathway. b Heat map showing effects of estrogens on bile acid metabolism in mammary glands. c Changes in primary bile acid glycocholate metabolism across groups. d Changes in secondary bile acid glycodeoxycholate metabolism across groups. e Changes in secondary bile acid glycochenodeoxycholate metabolism across groups. f Changes in taurocholate across groups. (p < 0.05). (GCA glycocholic acid, TCA taurocholic acid, GCDCA glycochenodeoxycholic acid, TCDCA taurochenodeoxycholic acid, DCA deoxycholic acid, LCA lithocholic acid, GDCA glycodeoxycholic acid, TDCA taurodeoxycholic acid, GLCA glycolithocholic acid, TLCA taurolithocholic acid, GUDCA glycoursodeoxycholic acid, TUDCA tauroursodeoxycholic acid)

Fig. 6

Changes in oxidative stress metabolites associated with estrogen status. a Heat map showing effects of estrogens on oxidative stress metabolites. a Changes in NAD + across groups. c Changes in NADH metabolism across groups. d Ratio of NAD + to NADH across groups. e Changes in FAD + across groups. f Changes in prostaglandin F2α across groups. g Changes in glutathione across groups. h Ratio of reduced to oxidized glutathione across groups