Methylation status of Vitamin D receptor gene promoter in benign and malignant adrenal tumors

We previously showed a decreased expression of vitamin D receptor (VDR) mRNA/protein in a small group of adrenocortical carcinoma (ACC) tissues, suggesting the loss of a protective role of VDR against malignant cell growth in this cancer type. Downregulation of VDR gene expression may result from epigenetics events, that is, methylation of cytosine nucleotide of CpG islands in VDR gene promoter. We analyzed methylation of CpG sites in the VDR gene promoter in normal adrenals and adrenocortical tumor samples. Methylation of CpG-rich 5' regions was assessed by bisulfite sequencing PCR using bisulfite-treated DNA from archival microdissected paraffin-embedded adrenocortical tissues. Three normal adrenals and 23 various adrenocortical tumor samples (15 adenomas and 8 carcinomas) were studied. Methylation in the promoter region of VDR gene was found in 3/8 ACCs, while no VDR gene methylation was observed in normal adrenals and adrenocortical adenomas. VDR mRNA and protein levels were lower in ACCs than in benign tumors, and VDR immunostaining was weak or negative in ACCs, including all 3 methylated tissue samples. The association between VDR gene promoter methylation and reduced VDR gene expression is not a rare event in ACC, suggesting that VDR epigenetic inactivation may have a role in adrenocortical carcinogenesi

Epitope Characterization of an Aromatase Monoclonal Antibody Suitable for the Assessment of Intratumoral Aromatase Activity

Immunohistochemistry is one of the most suitable methods for the detection of intratumoral aromatase in order to identify patients who may respond to aromatase inhibitor therapy in hormone-dependent breast cancer. Previous studies showed statistically significant correlation between results of immnuohistochemistry and biochemical analysis in carcinoma components stained by aromatase monoclonal antibody 677. In this study, determination of the antigenic peptides recognized by aromatase antibodies through epitope mapping, combined with the new knowledge on aromatase-reductase interaction, provide insights for understanding various immunostaining patterns using different aromatase antibodies. Our studies on aromatase-reductase interaction also provided critical information on how aromatase and reductase interact with each other on the endoplasmic reticulum membrane, and identified key residues, including K108 of aromatase, that are involved in the interaction with reductase. Through epitope mapping and taking into consideration the interference with aromatase immunohistochemical staining by NADPH-cytochrome P450 reductase, we demonstrated that monoclonal antibody 677 is a suitable antibody for an assessment of intratumoral aromatase activity in breast cancer patients for making clinical management decisions. These results also provide valuable information to identify new aromatase antibodies for immunohistochemical diagnosis of hormone-dependent breast cancer in future

Aromatase expression and outcomes in the P024 neoadjuvant endocrine therapy trial

Background Expression of aromatase by malignant breast epithelial cells and/or the surrounding stroma implies local estrogen production that could influence the outcome of endocrine therapy for breast cancer. Methods A validated immunohistochemical assay for aromatase was applied to samples from the P024 neoadjuvant endocrine therapy trial that compared tamoxifen and letrozole. The presence of aromatase expression by tumor or stromal cells was correlated with tumor response, treatment induced changes in proliferation index (Ki67), relapse-free survival (RFS) and breast cancer-specific survival (BCSS). Results Tumor and stromal aromatase expression were highly correlated (P = 0.0001). Tumor cell aromatase, as a semi-continuous score, also correlated with smaller tumor size at presentation (P = 0.01) higher baseline ER Allred score (P = 0.006) and lower Ki67 levels (P = 0.003). There was no significant relationship with clinical response or treatment-induced changes in Ki67. However, in a Cox multivariable model that incorporated a post-treatment tumor profile (pathological T stage, N stage, Ki67 and ER status of the surgical specimen), the presence of tumor aromatase expression at baseline sample remained a favorable independent prognostic biomarker for both RFS (P = 0.01, HR 2.3, 95% CI 1.2–4.6 for absent expression) and BCSS (P = 0.008, HR 3.76, 95% CI 1.4–10.0). Conclusions Autocrine estrogen synthesis may be most characteristic of smaller, more indolent and ER-rich breast cancers with lower baseline growth rates. However, response to endocrine treatment may not depend on whether the estrogenic stimulus has a local versus systemic source

Therapeutic advances in hormone-dependent cancers: focus on prostate, breast and ovarian cancers

Hormonal cancers affect over 400,000 men and women and contribute collectively to over 100,000 deaths in the United States alone. Thanks to advances in the understanding of these cancers at the molecular level and to the discovery of several disease-modifying therapeutics, the last decade has seen a plateauing or even a decreasing trend in the number of deaths from these cancers. These advanced therapeutics not only effectively slow the growth of hormonal cancers, but also provide an insight on how these cancers become refractory and evolve as an altogether distinct subset. This review summarizes the current therapeutic trends in hormonal cancers, with focus on prostate, breast and ovarian cancers. The review discusses the clinical drugs being used now, promising molecules that are going through various stages of development and makes some predictions on how the therapeutic landscape will shift in the next decade

Accelerated telomere shortening in adrenal zona reticularis in patients with prolonged critical illness

BackgroundThe number of patients with prolonged critical illness (PCI) has been increasing in many countries, and the adrenal gland plays an important role in maintaining homeostasis during PCI. Chronic disease burden is reportedly associated with shorter telomere lengths in human tissues. Telomere shortening in human somatic cells is largely dependent on cell divisions, and critically short telomeres lead to cellular dysfunction and aging. However, the association between PCI and telomere lengths in human adrenal cells is poorly understood. In this study, we investigated this association to assess whether the burden of PCI could accelerate the aging process in adrenal cells.MethodsAdrenocortical tissues from patients who died after PCI usually show a diffuse pattern of intracellular cholesterol ester depletion (i.e., lipid depletion). This study examined near-normal adrenal glands obtained from autopsied patients who died suddenly (control group) and lipid-depleted adrenal glands obtained from autopsied patients who died after PCI (PCI group). The control group included 7 men aged 80 to 94 years (mean age: 85.3 years) and 7 women aged 84 to 94 years (mean age: 87.7 years). The PCI group included 10 men aged 71 to 88 years (mean age: 78.8 years) and 8 women aged 77 to 95 years (mean age: 85.6 years). By using quantitative fluorescence in situ hybridization, relative telomere lengths (RTLs) were determined in the parenchymal cells of the three adrenocortical zones (zona glomerulosa, zona fasciculata, and zona reticularis [ZR]) and in the chromaffin cells of the medulla. The number of adrenal parenchymal cells was determined by immunohistochemistry and digital image analysis.ResultsRTLs in ZR cells were significantly shorter in the PCI group than in the control group for both men and women (P = 0.0001 for men and P = 0.0012 for women). However, RTLs in the remaining three types of adrenal cells did not differ between the control and PCI groups for both men and women. The number of ZR cells was higher in the PCI group than in the control group for both men and women (P < 0.0001 for both men and women). The proportion of the number of ZR cells to the total number of adrenocortical parenchymal cells was also higher in the PCI group than in the control group (P < 0.0001 for both men and women). The Ki-67 proliferation index in ZR cells was higher in the PCI group than in the control group (P = 0.0039 for men and P = 0.0063 for women).ConclusionsThis study demonstrated ZR cell-specific telomere shortening in patients with adrenal lipid depletion who died after PCI. Our results suggest that the reactive proliferation of ZR cells accelerates the telomere shortening and aging process in ZR cells in these patients. The results of our study may contribute to the understanding of adrenal aging during PCI

In situ aromatase expression in primary tumor is associated with estrogen receptor expression but is not predictive of response to endocrine therapy in advanced breast cancer

<p>Abstract</p> <p>Background</p> <p>New, third-generation aromatase inhibitors (AIs) have proven comparable or superior to the anti-estrogen tamoxifen for treatment of estrogen receptor (ER) and/or progesterone receptor (PR) positive breast cancer. AIs suppress total body and intratumoral estrogen levels. It is unclear whether <it>in situ </it>carcinoma cell aromatization is the primary source of estrogen production for tumor growth and whether the aromatase expression is predictive of response to endocrine therapy. Due to methodological difficulties in the determination of the aromatase protein, COX-2, an enzyme involved in the synthesis of aromatase, has been suggested as a surrogate marker for aromatase expression.</p> <p>Methods</p> <p>Primary tumor material was retrospectively collected from 88 patients who participated in a randomized clinical trial comparing the AI letrozole to the anti-estrogen tamoxifen for first-line treatment of advanced breast cancer. Semi-quantitative immunohistochemical (IHC) analysis was performed for ER, PR, COX-2 and aromatase using Tissue Microarrays (TMAs). Aromatase was also analyzed using whole sections (WS). Kappa analysis was applied to compare association of protein expression levels. Univariate Wilcoxon analysis and the Cox-analysis were performed to evaluate time to progression (TTP) in relation to marker expression.</p> <p>Results</p> <p>Aromatase expression was associated with ER, but not with PR or COX-2 expression in carcinoma cells. Measurements of aromatase in WS were not comparable to results from TMAs. Expression of COX-2 and aromatase did not predict response to endocrine therapy. Aromatase in combination with high PR expression may select letrozole treated patients with a longer TTP.</p> <p>Conclusion</p> <p>TMAs are not suitable for IHC analysis of <it>in situ </it>aromatase expression and we did not find COX-2 expression in carcinoma cells to be a surrogate marker for aromatase. <it>In situ </it>aromatase expression in tumor cells is associated with ER expression and may thus point towards good prognosis. Aromatase expression in cancer cells is not predictive of response to endocrine therapy, indicating that <it>in situ </it>estrogen synthesis may not be the major source of intratumoral estrogen. However, aromatase expression in combination with high PR expression may select letrozole treated patients with longer TTP.</p> <p>Trial registration</p> <p>Sub-study of trial P025 for advanced breast cancer.</p