A Physiologist’s Perspective
No doubt about it, metabolism (both synthesis and degradation) of 17β-estradiol is complicated! For nonbiochemists, just thinking about oxidation, hydroxylation (>30 identified products), glucuronidation, sulfonation, and O-methylation1 is enough to make our heads spin. Yet, it is just this complex series of regulatory pathways that enables the endogenous hormone to regulate physiological changes of pregnancy, regulate skeletal development, modulate neurotransmission, promote some types of cancer, and affect development of cardiovascular disease. It is important, therefore, to better understand the biochemical and physiological effects of the various products of 17β-estradiol metabolism.
Genomic, biochemical, and physiological effects of 2-methoxyestradiol (2-ME), a methylation product of catechol estrogens by catechol-O-methyltransferase (COMT), are highlighted in this volume (Figure).2 This thorough study by Barchiesi et al2 extends our knowledge of mechanistic effects of 2-ME by defining gene expression in response to 2-ME in cultured human (female) vascular smooth muscle cells. Functional outcomes of gene expression are confirmed by validating changes in protein expression, enzyme activity, and functional assays of cell proliferation, viability, and apoptosis. Similar to mechanistic pathways identified in some cancer cell lines, 2-ME reduces smooth muscle cell proliferation through inhibition of genes involved in cell division, including cyclin D1, cyclin B1, cdk6, cdk4, and tubulin polymerization.3 Thus, it appears that cellular pathways activated by 2-ME are common to several cell types, including some cancers, and that these genomic effects are independent of the sex of the cells (ie, of male or female origin). Indeed, 2-ME was tested as a chemo-intervention to treat hormone-resistant prostate cancer in men.4 In experimental animals, administration of 2-ME reduced atherosclerotic lesion formation in female apolipoprotein E–deficient mice and increased renal blood flow and glomerular filtration without affecting systolic blood pressure in aged, obese, and diabetic male Zucker rats.5,6
Both intravenous and subcutaneous delivery of 2-ME to either male or female rodents decreased serum cholesterol. Based on results of the study by Barchiesi et al,2 this response is consistent with the ability of 2-ME to downregulate 3-hydroxy-3-methylglutaryl-coenzyme A reductase, which is routinely targeted by statin therapy to reduce cardiovascular risk. Because a significant portion of estrogen metabolism occurs in the liver, regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase by 2-ME may explain why oral exogenous estrogenic products (including conjugated equine estrogen) have a greater lipid-lowering effect than transdermal products in postmenopausal women.
An important observation of the study by Barchiesi et al2 that may provide a link between estrogen metabolism and ischemic heart disease is that 2-ME upregulated genes for peroxisome proliferator activated receptor (PPAR) α and PPARγ. However, this activation was not attributed to direct ligand binding to PPAR receptors but rather by prostaglandins generated through increased-activity cyclooxygenase 2. Similar to rosiglitazone, a ligand for PPAR receptors, 2-ME downregulated the transcription factor hypoxia-inducible factor 1α. Hypoxia-inducible factor 1α is required for angiogenesis. Although reducing angiogenesis would be beneficial in reducing nutrient blood supply to solid tumors, it may be deleterious in nontumor tissues like the heart. Increases in adverse cardiac ischemic events and myocardial infarction are reported from clinical trails evaluating rosiglitazone for treatment of type 2 diabetes mellitus.7 It remains to be determined whether the presentation of myocardial microvascular disease in women could be related to metabolic products of endogenous estrogen.
2-ME also upregulated expression of matrix metalloprotease 1, an enzyme implicated in destabilization of plaques. Therefore, whereas 2-ME may prevent progression of a vascular lesion, existing plaques may be destabilized, as was observed in the Heart and Estrogen/Progestin Replacement Study. The ability of estrogenic products to reduce progression of vascular lesions is being tested in the ongoing Kronos Early Estrogen Prevention Study.8 Genetic analysis of enzymes involved in the metabolism of estrogen in conjunction with a profile of estrogen metabolites in Kronos Early Estrogen Prevention Study participants will provide complementary information of how an individual’s ability to metabolize estrogen affects various cardiovascular risk factors.
Collectively, data reported by Barchiesi et al2 contribute to the accumulating evidence that 2-ME may be a promising therapeutic compound to treat certain proliferative diseases, such as cancers and formation of vascular lesions. However, several challenges remain before the use of this estrogen metabolite reaches therapeutic reality in humans. One challenge is to better define the contribution of 2-ME in normal physiology. Such studies would be facilitated by the identification of specific receptors for 2-ME and the subsequent development of selective agonists and antagonists. Additional experimental animal studies are needed to delineate the physiological consequences of administration of the compound to female diabetic, hypertensive, and aged animals and perhaps the contribution of 2-ME, which is in high concentrations in the placenta to preeclampsia.
Evaluation of the estrogen metabolites in humans is difficult because of metabolism of estrogen in some target tissues. Thus, measurement of unmetabolized estrogen and estrogen metabolites in the plasma or urine may not provide an accurate assessment of estrogenic activity in the target tissue. Nonetheless, 2-ME has not been evaluated in studies of menopausal hormone treatment most likely because of expense and availability of sensitive assays by liquid chromatography/tandem mass spectrometry. Better understanding of the relative concentration of the metabolite in women using various hormonal treatments may provide insight into the susceptibility to menopause-associated increases in cardiovascular risk factors (increased cholesterol, hypertension, and insulin resistance).
As 2-ME decreased expression of hypoxia-inducible factor 1α, evaluation of myocardial microvasculature in experimental animals treated with 2-ME may provide information about factors contributing to myocardial ischemic microvascular disease in women. COMT is key to degradation of norepinephrine, and much attention has been paid to genetic variation and activity of COMT in relationship to central neuronal function associated with addiction, Parkinson disease, and Alzheimer disease. More work is needed to better understand how genetic polymorphisms or copy number of COMT influence development of hypertension, because the kidney contains large amounts of the enzyme.1
Improved insight into the ability of individuals to metabolize 17β-estradiol9 would be useful to identify the most effective type, dose, and mode of delivery of estrogenic hormonal treatments. For example, in the study by Barchiesi et al,2 intravenous delivery of 2-ME to sexually mature male rats decreased serum cholesterol, progesterone, and testosterone2; in female ovariectomized apolipoprotein E knockout mice, subcutaneous administration of 2-ME decreased total serum cholesterol but to a lesser extent than 17β-estradiol.5 Development of an oral analog of 2-ME with appropriate solubility and circulating half-life awaits further study.4 Encapsulation of 2-ME within polyelectrolyte multilayers is being investigated as a potential delivery mode, which could be applied to tumors or implantable devices, such as vascular stents.5,10 An important fact needed in developing delivery products for 2-ME is that, for vascular smooth muscle cells, a treatment window of ≥30 hours is needed for modulation of gene transcription.2
Cumulative intracellular effects of estrogenic compounds will reflect direct activation of estrogen receptors and actions of estrogenic metabolites, like 2-ME, that might be independent of the classical estrogen receptors. The article by Barchiesi et al2 provides a wealth of information about potential gene and enzyme pathways affected by 2-ME (Figure). These pathways provide insight into the regulation and interconnection of hormonal status with major risk factors for cardiovascular disease, such as increases in blood cholesterol, hypertension, insulin insensitivity, vascular occlusion, and ischemia. The antiproliferating and nonfeminizing actions of 2-ME make it an attractive therapeutic agent for some cancers and vascular occlusive disease. However, before this objective is realized, more integrated physiological studies in both male and female animals of various ages and disease models are needed to better understand the overall benefits and risks of this and other specific estrogen metabolites.
Sources of Funding
V.M.M.’s salary is supported by grants from the National Institutes of Health (HL90639, AG29624, and NS066147), the Kronos Longevity Research Institute, and the Mayo Clinic.
V.M.M. is president of the Organization for the Study of Sex Differences and serves on the board of directors for the Society of Women’s Health Research.
The opinions expressed in this editorial are not necessarily those of the editors or of the American Heart Association.
Zhu BT, Conney AH. Functional role of estrogen metabolism in target cells: review and perspectives. Carcinogenesis. 1998; 19: 1–27.
Barchiesi F, Lucchinetti E, Zaugg M, Ogunshola OO, Wright M, Meyer M, Rosselli M, Schaufelberger S, Gillespie DG, Jackson EK, Dubey RK. Candidate genes and mechanisms for 2-methoxyestradiol–mediated vasoprotection. Hypertension. 2010; 56: 964–972.
Sweeney C, Liu G, Yiannoutsos C, Kolesar J, Horvath D, Staab MJ, Fife K, Armstrong V, Treston A, Sidor C, Wilding G. A phase II multicenter, randomized, double-blind, safety trial assessing the pharmacokinetics, pharmacodynamics, and efficacy of oral 2-methoxyestradiol capsules in hormone-refractory prostate cancer. Clin Cancer Res. 2005; 11: 6625–6633.