Dataset Information

Cystatin M loss is associated with the losses of estrogen receptor, progesterone receptor, and HER4 in invasive breast cancer.

ABSTRACT: INTRODUCTION: This study was aimed at understanding the clinicopathological significance of cystatin M loss, and investigating possible factors responsible for cystatin M loss in breast cancer. METHODS: The expression of estrogen receptor (ER), progesterone receptor (PR), HER2, HER4, and cystatin M was retrospectively analyzed using immunohistochemistry in 117 patients with ductal carcinoma in situ (DCIS) and in 175 patients with invasive breast cancer (IBC). The methylation status of CST6 gene encoding cystatin M was evaluated using methylation-specific polymerase chain reaction (PCR) in formalin-fixed paraffin-embedded tissues from 292 participants and using pyrosequencing in fresh-frozen tumor and matched normal tissues from 51 IBC patients. RESULTS: Cystatin M loss was found in 9 (8%) of 117 patients with DCIS and in 99 (57%) of 175 with invasive breast cancer (IBC) (P < 0.0001). Cystatin M loss was found in 58 (57%) of 101 HER2-negative IBCs and in 41 (55%) of 74 HER2-positive IBCs, and this difference was not statistically significant (P = 0.97). However, cystatin M loss was significantly associated with the loss of ER (P = 0.01), PR (P = 0.002), and HER4 (P = 0.003) in IBCs. Cystatin M loss occurred in 34 (76%) of the 45 HER4-negative IBCs and in 65 (50%) of the 130 HER4-positive IBCs. Multivariate analysis showed that cystatin M loss occurred at a 3.57 times (95% CI = 1.28 to 9.98; P = 0.01) higher prevalence in the triple-negative IBCs of ER, PR, and HER4 than in other subtypes, after adjusting for age. The quantity of CST6 methylation was associated with ER loss (P = 0.0002) in IBCs but not with the loss of PR (P = 0.64) or HER4 (P = 0.87). CONCLUSIONS: The present study suggests that cystatin M loss may be associated with the losses of ER, PR, and HER4 in IBC.

SUBMITTER: Ko E

PROVIDER: S-EPMC3046445 | biostudies-literature | 2010

REPOSITORIES: biostudies-literature

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Cystatin M loss is associated with the losses of estrogen receptor, progesterone receptor, and HER4 in invasive breast cancer.

Ko Eunkyung E Park Seong-Eun SE Cho Eun Yoon EY Kim Yujin Y Hwang Jung-Ah JA Lee Yeon-Su YS Nam Seok Jin SJ Bang Saik S Park Joobae J Kim Duk-Hwan DH

Breast cancer research : BCR 20101123 6

<h4>Introduction</h4>This study was aimed at understanding the clinicopathological significance of cystatin M loss, and investigating possible factors responsible for cystatin M loss in breast cancer.<h4>Methods</h4>The expression of estrogen receptor (ER), progesterone receptor (PR), HER2, HER4, and cystatin M was retrospectively analyzed using immunohistochemistry in 117 patients with ductal carcinoma in situ (DCIS) and in 175 patients with invasive breast cancer (IBC). The methylation status ...[more]

PMID: 21092257

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Project description:Simple Summary Most breast cancers co-express estrogen receptor α (ERα) and progesterone receptor (PgR). These cancers are sensitive to endocrine therapy and, in general, have superior outcomes. However, a subset of tumors expresses ERα but loses expression of PgR in various mechanisms. The processes driving the loss of PgR may cause resistance to hormonal treatment and a more aggressive clinical course. The current review summarizes current knowledge on the biology of ERα-positive PgR(−)negative breast cancer and discusses the associations between molecular mechanisms and clinical characteristics. Abstract Estrogen receptor α (ERα) and progesterone receptor (PgR) are crucial prognostic and predictive biomarkers that are usually co-expressed in breast cancer (BC). However, 12–24% of BCs present ERα(+)/PgR(−) phenotype at immunohistochemical evaluation. In fact, BC may either show primary PgR(−) status (in chemonaïve tumor sample), lose PgR expression during neoadjuvant treatment, or acquire PgR(−) phenotype in local relapse or metastasis. The loss of PgR expression in ERα(+) breast cancer may signify resistance to endocrine therapy and poorer outcomes. On the other hand, ERα(+)/PgR(−) BCs may have a better response to neoadjuvant chemotherapy than double-positive tumors. Loss of PgR expression may be a result of pre-transcriptional alterations (copy number loss, mutation, epigenetic modifications), decreased transcription of the PGR gene (e.g., by microRNAs), and post-translational modifications (e.g., phosphorylation, sumoylation). Various processes involved in the down-regulation of PgR have distinct consequences on the biology of cancer cells. Occasionally, negative PgR status detected by immunohistochemical analysis is paradoxically associated with enhanced transcriptional activity of PgR that might be inhibited by antiprogestin treatment. Identification of the mechanism of PgR loss in each patient seems challenging, yet it may provide important information on the biology of the tumor and predict its responsiveness to the therapy.

Project description:The EGFR-family member HER4 undergoes regulated intramembrane proteolysis (RIP) to generate an intracellular domain (4ICD) that functions as a transcriptional coactivator. Accordingly, 4ICD coactivates the estrogen receptor (ER) and associates with ER at target gene promoters in breast tumor cells. However, the extent of 4ICD coactivation of ER and the functional significance of the 4ICD/ER transcriptional complex is unclear. To identify 4ICD coactivated genes we performed a microarray gene expression analysis of β-estradiol treated cells comparing control MCF-7 breast cancer cells to MCF-7 cells where HER4 expression was stably suppressed using a shRNA. In the MCF-7 cell line, β-estradiol significantly stimulated or repressed by 2-fold or more 726 or 53 genes, respectively. Significantly, HER4/4ICD was an obligate coactivator for 277 or 38% of the β-estradiol stimulated genes. Ingenuity Pathway Analysis of β-estradiol regulated genes identified significant associations with multiple cellular functions regulating cellular growth and proliferation, cell cycle progression, cancer metastasis, decreased hypoplasia, tumor cell migration, apoptotic resistance of tumor cells, and increased transcription. Genes coactivated by 4ICD displayed functional specificity by only significantly contributing to cellular growth and proliferation, cell cycle progression, and decreased hypoplasia. In direct concordance with these in situ results we show that HER4 knockdown in MCF-7 cells results in a loss of estrogen stimulated tumor cell proliferation and cell cycle progression, whereas, estrogen stimulated tumor cell migration was unaffected by loss of HER4 expression. In summary, we demonstrate for the first time that a cell surface receptor functions as an obligate ER coactivator with functional specificity associated with breast tumor cell proliferation and cell cycle progression. Nearly 90% of ER positive tumors coexpress HER4, therefore we predict that the majority of breast cancer patients would benefit from a strategy to therapeutic disengage ER/4ICD coregulated tumor cell proliferation.

Project description:The HER4 intracellular domain (4ICD) is a potent estrogen receptor (ERalpha) coactivator with activities in breast cancer and the developing mammary gland that appear to overlap with progesterone receptor (PgR). In fact, 4ICD has recently emerged as an important regulator and predictor of tamoxifen response, a role previously thought to be fulfilled by PgR. Here we investigated the possibility that the 4ICD coactivator regulates PgR expression thereby providing a mechanistic explanation for their partially overlapping activities in breast cancer. We show that 4ICD is both sufficient and necessary to potentiate estrogen stimulation of gene expression. Suppression of HER4/4ICD expression in the MCF-7 breast tumor cell line completely eliminated estrogen stimulated expression of PgR. In addition, the HER4/4ICD negative MCF-7 variant, TamR, failed to express PgR in response to estrogen. Reintroduction of wild-type HER4 but not the gamma-secretase processing mutant HER4V673I into the TamR cell line restored PgR expression indicating that 4ICD is an essential PgR coactivator in breast tumor cells. These results were substantiated in vivo using two different physiologically relevant experimental systems. In the mouse mammary gland estrogen regulates expression of PgR-A whereas expression of PgR-B is estrogen independent. Consistent with a role for 4ICD in estrogen regulated PgR expression in vivo, PgR-A, but not PgR-B, expression was abolished in HER4-null mouse mammary glands during pregnancy. Coexpression of PgR and 4ICD is also commonly observed in ERalpha positive breast carcinomas. Using quantitative AQUA IHC technology we found that 4ICD potentiated PgR expression in primary breast tumors and the highest levels of PgR expression required coexpression of ERalpha and the 4ICD coactivator. In summary, our results provide compelling evidence that 4ICD is a physiologically important ERalpha coactivator and 4ICD cooperates with ERalpha to potentiate PgR expression in the normal and malignant breast. We propose that direct coupling of these signaling pathways may have important implications for mammary development, breast carcinogenesis, and patient response to endocrine therapy.

Project description:IntroductionEstrogen receptor (ER) and progesterone receptor (PR) testing are performed in the evaluation of breast cancer. While the clinical utility of ER as a predictive biomarker to identify patients likely to benefit from hormonal therapy is well-established, the added value of PR is less well-defined. The primary goals of our study were to assess the distribution, inter-assay reproducibility, and prognostic significance of breast cancer subtypes defined by patterns of ER and PR expression.MethodsWe integrated gene expression microarray (GEM) and clinico-pathologic data from 20 published studies to determine the frequency (n = 4,111) and inter-assay reproducibility (n = 1,752) of ER/PR subtypes (ER+/PR+, ER+/PR-, ER-/PR-, ER-/PR+). To extend our findings, we utilized a cohort of patients from the Nurses' Health Study (NHS) with ER/PR data recorded in the medical record and assessed on tissue microarrays (n = 2,011). In both datasets, we assessed the association of ER and PR expression with survival.ResultsIn a genome-wide analysis, progesterone receptor was among the least variable genes in ER- breast cancer. The ER-/PR+ subtype was rare (approximately 1 to 4%) and showed no significant reproducibility (Kappa = 0.02 and 0.06, in the GEM and NHS datasets, respectively). The vast majority of patients classified as ER-/PR+ in the medical record (97% and 94%, in the GEM and NHS datasets) were re-classified by a second method. In the GEM dataset (n = 2,731), progesterone receptor mRNA expression was associated with prognosis in ER+ breast cancer (adjusted P <0.001), but not in ER- breast cancer (adjusted P = 0.21). PR protein expression did not contribute significant prognostic information to multivariate models considering ER and other standard clinico-pathologic features in the GEM or NHS datasets.ConclusionER-/PR+ breast cancer is not a reproducible subtype. PR expression is not associated with prognosis in ER- breast cancer, and PR does not contribute significant independent prognostic information to multivariate models considering ER and other standard clinico-pathologic factors. Given that PR provides no clinically actionable information in ER+ breast cancer, these findings question the utility of routine PR testing in breast cancer.

Project description:IntroductionOf the nearly 1.4 million new cases of breast cancer diagnosed each year, a large proportion is characterized as hormone receptor negative, lacking estrogen receptors (ER) and/or progesterone receptors (PR). Patients with receptor-negative tumors do not respond to current steroid hormone-based therapies and generally have significantly higher risk of recurrence and mortality compared with patients with tumors that are ER- and/or PR-positive. Previous in vitro studies had shown that the progesterone metabolites, 5α-dihydroprogesterone (5αP) and 3α-dihydroprogesterone (3αHP), respectively, exhibit procancer and anticancer effects on receptor-negative human breast cell lines. Here in vivo studies were conducted to investigate the ability of 5αP and 3αHP to control initiation, growth, and regression of ER/PR-negative human breast cell tumors.MethodsER/PR-negative human breast cells (MDA-MB-231) were implanted into mammary fat pads of immunosuppressed mice, and the effects of 5αP and 3αHP treatments on tumor initiation, growth, suppression/regression, and histopathology were assessed in five separate experiments. Specific radioimmunoassays and gas chromatography-mass spectrometry were used to measure 5αP, 3αHP, and progesterone in mouse serum and tumors.ResultsOnset and growth of ER/PR-negative human breast cell tumors were significantly stimulated by 5αP and inhibited by 3αHP. When both hormones were applied simultaneously, the stimulatory effects of 5αP were abrogated by the inhibitory effects of 3αHP and vice versa. Treatment with 3αHP subsequent to 5αP-induced tumor initiation resulted in suppression of further tumorigenesis and regression of existing tumors. The levels of 5αP in tumors, regardless of treatment, were about 10-fold higher than the levels of 3αHP, and the 5αP:3αHP ratios were about fivefold higher than in serum, indicating significant changes in endogenous synthesis of these hormones in tumorous breast tissues.ConclusionsThe studies showed that estrogen/progesterone-insensitive breast tumors are sensitive to, and controlled by, the progesterone metabolites 5αP and 3αHP. Tumorigenesis of ER/PR-negative breast cells is significantly enhanced by 5αP and suppressed by 3αHP, the outcome depending on the relative concentrations of these two hormones in the microenvironment in the breast regions. The findings show that the production of 5αP greatly exceeds that of 3αHP in ER/PR-negative tumors and that treatment with 3αHP can effectively block tumorigenesis and cause existing tumors to regress. The results provide the first hormonal theory to explain tumorigenesis of ER/PR-negative breast tissues and support the hypothesis that a high 3αHP-to-5αP concentration ratio in the microenvironment may foster normalcy in noncancerous breast regions. The findings suggest new diagnostics based on the relative levels of these hormones and new approaches to prevention and treatment of breast cancers based on regulating the levels and action mechanisms of anti- and pro-cancer progesterone metabolites.

Dataset Information

Cystatin M loss is associated with the losses of estrogen receptor, progesterone receptor, and HER4 in invasive breast cancer.

Publications

Cystatin M loss is associated with the losses of estrogen receptor, progesterone receptor, and HER4 in invasive breast cancer.

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