RESUMEN
Breast cancer etiology is associated with both proliferation and DNA damage induced by estrogens. Breast cancer risk factors (BCRF) such as body mass index (BMI), smoking, and intake of estrogen-active drugs were recently shown to influence intratissue estrogen levels. Thus, the aim of the present study was to investigate the influence of BCRF on estrogen-induced proliferation and DNA damage in 41 well-characterized breast glandular tissues derived from women without breast cancer. Influence of intramammary estrogen levels and BCRF on estrogen receptor (ESR) activation, ESR-related proliferation (indicated by levels of marker transcripts), oxidative stress (indicated by levels of GCLC transcript and oxidative derivatives of cholesterol), and levels of transcripts encoding enzymes involved in estrogen biotransformation was identified by multiple linear regression models. Metabolic fluxes to adducts of estrogens with DNA (E-DNA) were assessed by a metabolic network model (MNM) which was validated by comparison of calculated fluxes with data on methoxylated and glucuronidated estrogens determined by GC- and UHPLC-MS/MS. Intratissue estrogen levels significantly influenced ESR activation and fluxes to E-DNA within the MNM. Likewise, all BCRF directly and/or indirectly influenced ESR activation, proliferation, and key flux constraints influencing E-DNA (i.e., levels of estrogens, CYP1B1, SULT1A1, SULT1A2, and GSTP1). However, no unambiguous total effect of BCRF on proliferation became apparent. Furthermore, BMI was the only BCRF to indeed influence fluxes to E-DNA (via congruent adverse influence on levels of estrogens, CYP1B1 and SULT1A2).
Asunto(s)
Neoplasias de la Mama/metabolismo , Daño del ADN , Estrógenos/metabolismo , Glándulas Mamarias Humanas/metabolismo , Adulto , Arilsulfotransferasa/metabolismo , Índice de Masa Corporal , Neoplasias de la Mama/etiología , Proliferación Celular/fisiología , Cromatografía Líquida de Alta Presión , Citocromo P-450 CYP1B1/metabolismo , Femenino , Humanos , Glándulas Mamarias Humanas/patología , Estrés Oxidativo/fisiología , Factores de Riesgo , Espectrometría de Masas en TándemRESUMEN
Understanding intramammary estrogen homeostasis constitutes the basis of understanding the role of lifestyle factors in breast cancer etiology. Thus, the aim of the present study was to identify variables influencing levels of the estrogens present in normal breast glandular and adipose tissues (GLT and ADT, i.e., 17ß-estradiol, estrone, estrone-3-sulfate, and 2-methoxy-estrone) by multiple linear regression models. Explanatory variables (exVARs) considered were (a) levels of metabolic precursors as well as levels of transcripts encoding proteins involved in estrogen (biotrans)formation, (b) data on breast cancer risk factors (i.e., body mass index, BMI, intake of estrogen-active drugs, and smoking) collected by questionnaire, and (c) tissue characteristics (i.e., mass percentage of oil, oil%, and lobule type of the GLT). Levels of estrogens in GLT and ADT were influenced by both extramammary production (menopausal status, intake of estrogen-active drugs, and BMI) thus showing that variables known to affect levels of circulating estrogens influence estrogen levels in breast tissues as well for the first time. Moreover, intratissue (biotrans)formation (by aromatase, hydroxysteroid-17beta-dehydrogenase 2, and beta-glucuronidase) influenced intratissue estrogen levels, as well. Distinct differences were observed between the exVARs exhibiting significant influence on (a) levels of specific estrogens and (b) the same dependent variables in GLT and ADT. Since oil% and lobule type of GLT influenced levels of some estrogens, these variables may be included in tissue characterization to prevent sample bias. In conclusion, evidence for the intracrine activity of the human breast supports biotransformation-based strategies for breast cancer prevention. The susceptibility of estrogen homeostasis to systemic and tissue-specific modulation renders both beneficial and adverse effects of further variables associated with lifestyle and the environment possible.
Asunto(s)
Biotransformación/fisiología , Neoplasias de la Mama , Mama/metabolismo , Estrógenos/metabolismo , 17-Hidroxiesteroide Deshidrogenasas , Aromatasa/metabolismo , Estradiol , Estrona/análogos & derivados , Estrona/metabolismo , Homeostasis , Humanos , Factores de RiesgoRESUMEN
Despite being widely used to investigate 17ß-estradiol (E2)-induced mammary gland (MG) carcinogenesis and prevention thereof, estrogen homeostasis and its significance in the female August Copenhagen Irish (ACI) rat model is unknown. Thus, levels of 12 estrogens including metabolites and conjugates were determined mass spectrometrically in 38 plasmas and 52 tissues exhibiting phenotypes ranging from normal to palpable tumor derived from a representative ACI study using two different diets. In tissues, 40 transcripts encoding proteins involved in estrogen (biotrans)formation, ESR1-mediated signaling, proliferation and oxidative stress were analyzed (TaqMan PCR). Influence of histo(patho)logic phenotypes and diet on estrogen and transcript levels was analyzed by 2-way ANOVA and explanatory variables influencing levels and bioactivity of estrogens in tissues were identified by multiple linear regression models. Estrogen profiles in tissue and plasma and the influence of Hsd17b1 levels on intra-tissue levels of E2 and E1 conclusively indicated intra-mammary formation of E2 in ACI tumors by HSD17B1-mediated conversion of E1. Proliferation in ACI tumors was influenced by Egfr, Igf1r, Hgf and Met levels. 2-MeO-E1, the only oxidative estrogen metabolite detected above 28-42 fmol/g, was predominately observed in hyperplastic tissues and intra-tissue conversion of E1 seemed to contribute to its levels. The association of the occurrence of 2-MeO-E1 with higher levels of oxidative stress observed in hyperplastic and tumor tissues remained equivocal. Thus, the present study provides mechanistic explanation for previous and future results observed in the ACI model.
Asunto(s)
Estradiol/toxicidad , Estrógenos/toxicidad , Neoplasias Mamarias Experimentales/patología , Estrés Oxidativo/efectos de los fármacos , Animales , Proliferación Celular/efectos de los fármacos , Dieta , Estradiol/metabolismo , Receptor alfa de Estrógeno/metabolismo , Estrógenos/metabolismo , Femenino , Espectrometría de Masas , Ratas , Ratas Endogámicas ACIRESUMEN
Because of its assumed role in breast cancer etiology, estrogen biotransformation (and interaction of compounds therewith) has been investigated in human biospecimens for decades. However, little attention has been paid to the well-known fact that large inter-individual variations exist in the proportion of breast glandular (GLT) and adipose (ADT) tissues and less to adequate tissue characterization. To assess the relevance of this, the present study compares estrogen biotransformation in GLT and ADT. GLT and ADT were isolated from 47 reduction mammoplasty specimens derived from women without breast cancer and were characterized histologically and by their percentages of oil. Levels of 12 unconjugated and five conjugated estrogens were analyzed by GC- and UHPLC-MS/MS, respectively, and levels of 27 transcripts encoding proteins involved in estrogen biotransformation by Taqman® probe-based PCR. Unexpectedly, one-third of specimens provided neat GLT only after cryosection. Whereas 17ß-estradiol, estrone, and estrone-3-sulfate were detected in both tissues, estrone-3-glucuronide and 2-methoxy-estrone were detected predominately in GLT and ADT, respectively. Estrogen levels as well as ratios 17ß-estradiol/estrone and estrone-3-sulfate/estrone differed significantly between GLT and ADT, yet less than between individuals. Furthermore, estrogen levels in GLT and ADT correlated significantly with each other. In contrast, levels of most transcripts encoding enzymes involved in biotransformation differed more than between individuals and did not correlate between ADT and GLT. Thus, mixed breast tissues (and plasma) will not provide meaningful information on local estrogen biotransformation (and interaction of compounds therewith) whereas relative changes in 17ß-estradiol levels may be investigated in the more abundant ADT.
Asunto(s)
Tejido Adiposo/metabolismo , Mama/metabolismo , Estradiol/metabolismo , Estrógenos/metabolismo , Adolescente , Adulto , Anciano , Cromatografía de Gases , Cromatografía Líquida de Alta Presión , Femenino , Humanos , Persona de Mediana Edad , Espectrometría de Masas en Tándem , Adulto JovenRESUMEN
There is an ongoing debate whether the intake of soy-derived isoflavones (sISO) mediates beneficial or adverse effects with regard to breast cancer risk. Therefore, we investigated whether nutritional exposure to a sISO-enriched diet from conception until adulthood impacts on 17ß-estradiol (E2)-induced carcinogenesis in the rat mammary gland (MG). August-Copenhagen-Irish (ACI) rats were exposed to dietary sISO from conception until postnatal day 285. Silastic tubes containing E2 were used to induce MG tumorigenesis. Body weight, food intake, and tumor growth were recorded weekly. At necropsy, the number, position, size, and weight of each tumor were determined. Plasma samples underwent sISO analysis, and the morphology of MG was analyzed. Tumor incidence and multiplicity were reduced by 20 and 56 %, respectively, in the sISO-exposed rats compared to the control rats. Time-to-tumor onset was shortened from 25 to 20 weeks, and larger tumors developed in the sISO-exposed rats. The histological phenotype of the MG tumors was independent of the sISO diet received, and it included both comedo and cribriform phenotypes. Morphological analyses of the whole-mounted MGs also showed no diet-dependent differences. Lifelong exposure to sISO reduced the overall incidence of MG carcinomas in ACI rats, although the time-to-tumor was significantly shortened.
Asunto(s)
Estradiol/toxicidad , Glycine max/química , Isoflavonas/toxicidad , Neoplasias Mamarias Experimentales/inducido químicamente , Carga Tumoral/efectos de los fármacos , Animales , Dieta , Femenino , Isoflavonas/aislamiento & purificación , Isoflavonas/farmacología , Neoplasias Mamarias Experimentales/metabolismo , Neoplasias Mamarias Experimentales/patología , Ratas Endogámicas ACI , Factores de TiempoRESUMEN
Using rimonabant, a potent inverse agonist for cannabinoid receptor type 1 (CB1R), as parent ligand, a series of novel univalent and bivalent ligands were designed by variation of spacer length and its chemical structure. The ligands synthesized were evaluated for affinity and selectivity by radioligand displacement and a functional steady-state GTPase assay. The results showed the nature of the spacer influences the biological readout. Albeit all compounds show significantly lower affinities than rimonabant, this fact could be used to demonstrate that affinities and selectivity are influenced by the chemical structure and length of the spacer and might be helpful for designing bivalent probes for other GPCR receptors.
Asunto(s)
Amidas/farmacología , Aminas/farmacología , Agonistas de Receptores de Cannabinoides/química , Agonistas de Receptores de Cannabinoides/farmacología , Ácidos Carboxílicos/farmacología , Diseño de Fármacos , Receptor Cannabinoide CB1/agonistas , Receptor Cannabinoide CB1/metabolismo , Amidas/síntesis química , Amidas/química , Aminas/síntesis química , Aminas/química , Agonistas de Receptores de Cannabinoides/síntesis química , Ácidos Carboxílicos/síntesis química , Ácidos Carboxílicos/química , Relación Dosis-Respuesta a Droga , Humanos , Ligandos , Estructura Molecular , Relación Estructura-ActividadRESUMEN
The design of bivalent ligands targeting G protein-coupled receptors (GPCRs) often leads to the development of new, highly selective and potent compounds. To date, no bivalent ligands for the human cannabinoid receptor type 2 (hCB2R) of the endocannabinoid system (ECS) are described. Therefore, two sets of homobivalent ligands containing as parent structure the hCB2R selective agonist 13a and coupled at different attachment positions were synthesized. Changes of the parent structure at these positions have a crucial effect on the potency and efficacy of the ligands. However, we discovered that bivalency has an influence on the effect at both cannabinoid receptors. Moreover, we found out that the spacer length and the attachment position altered the efficacy of the bivalent ligands at the receptors by turning agonists into antagonists and inverse agonists.
Asunto(s)
Bencimidazoles/química , Receptor Cannabinoide CB2/agonistas , Bencimidazoles/síntesis química , Bencimidazoles/metabolismo , Diseño de Fármacos , Humanos , Cinética , Ligandos , Unión Proteica , Receptor Cannabinoide CB1/agonistas , Receptor Cannabinoide CB1/antagonistas & inhibidores , Receptor Cannabinoide CB1/metabolismo , Receptor Cannabinoide CB2/antagonistas & inhibidores , Receptor Cannabinoide CB2/metabolismoRESUMEN
In epidemiological studies, blood levels of 17ß-estradiol (E2) are associated with hormone-dependent diseases. The lack of specific methods impedes studies on the role of E2 metabolites and their conjugates in the etiology of hormone-dependent diseases. Stable-isotope dilution tandem mass spectrometry methods (coupled to gas chromatography and liquid chromatography systems) for the analysis of 22 endogenous estrogens, including both oxidative metabolites, as well as sulfates and glucuronides, was validated and the method applied to plasma of women with no breast cancer. No changes in estrogen profile during sample cleanup were observed and values for limit of detection (7fmol/ml - 2 pmol/ml), accuracies (80-122%) as well as intra- and inter-day precision (below 28%) at levels near the limit of quantification were acceptable. In human plasma only seven estrogens were detected and estrone conjugates contributed most to the estrogen profile.