High Levels Of Progesterone Receptors Research Articles

Abstract 3994: Differential regulation of hormone receptors and Ki67 in luminal breast carcinomas and tumor-adjacent mammary glands by mifepristone treatment

Abstract Since antiprogestins inhibit the growth of preclinical mammary tumor models expressing higher levels of progesterone receptor (PR) isoform A (PRA) than isoform B (PRB), we designed a window-of-opportunity trial (MIPRA; NCT02651844) to study the benefits of mifepristone (MFP) in luminal breast carcinomas from postmenopausal patients selected by their PR+ expression (>50%) and their PR isoform ratio (PRA/PRB>1.5; PRA-H). A decrease in the cell proliferation marker Ki67 was registered after treatment in 14 out of the 20 tumors evaluated. This inhibition was associated with a decrease in PR, estrogen receptor (ER) and pSer118ER evaluated by immunohistochemistry while no changes in pSer167ER expression were observed (PMID: 36269797). In selected tissues from these samples we observed that the staining intensity in trapped glands within the MFP-treated tumors did not follow a similar trend as that of tumor cells. Thus, the aim of this study was to evaluate the expression of hormone receptors and Ki67 in non-neoplastic human mammary glands (nnMG) adjacent to the PRA-H tumor tissue of postmenopausal patients, treated (n=8) or not (n=9) with MFP. No differences in PR, ER and pSer118ER expression were observed between MFP-treated nnMG and those from untreated patients. Notably, contrarily to what occurred in tumors, there was a significant decrease of pSer167ER expression (p=0.008) in the MFP-treated nnMG compared to the untreated glands, suggesting a selective modulation in the AKT-mediated activation of ER. Regarding the proliferative state of the nnMGs, there was a slight but significant increase (p=0.02) in Ki67 expression in MFP-treated vs. non-treated nnMG. Data regarding the effect of antiprogestins in normal mammary glands is limited and points, in premenopausal women, to a decrease in the Ki67 index after treatment. The basal quiescent status of nnMG in postmenopausal women may explain this slight stimulatory effect. In conclusion, our results show that MFP exerts a specific regulatory effect in PRA-H tumors that is not observed in the nnMG, probably due to the fact that nnMG have presumably equimolar levels of PRA and PRB. In addition, to counteract the possible stimulatory effect that MFP may induce on nnMG, the combination of MFP and tamoxifen treatment is suggested. Citation Format: Leo Saldain, Silvia Vanzulli, Paula Martinez Vazquez, Javier Burruchaga, Eunice Spengler, Claudia Lanari, Paola Rojas. Differential regulation of hormone receptors and Ki67 in luminal breast carcinomas and tumor-adjacent mammary glands by mifepristone treatment. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3994.

Expression of Sex Hormone Receptors in Canine Osteosarcoma.

Simple SummaryOsteosarcoma is a bone tumor that arises from normal bone cells that become malignant. This is a disease that is relatively common in dogs and also affects humans. Dogs are a useful model for human osteosarcoma as they share many similarities including genetic changes, manner of spread and how they respond to treatment. In dogs, the tumors are frequently seen in large breeds and may be caused by environmental and genetic factors. Sex steroids have been shown to regulate bone metabolism directly and indirectly through receptors on bone. The aim of this study was to investigate the expression levels of sex hormone receptors in canine osteosarcoma tissue and cell lines. Our results demonstrate varying levels of receptors in tissue and a high expression of progesterone receptors in the canine osteosarcoma cell lines. Low oxygen levels further increased progesterone receptor expression. Lastly, estradiol decreased the expression of progesterone receptor in one of the cell lines, and progesterone decreased cell proliferation in the same cell line. Further investigation is recommended to determine whether or not sex steroid receptor levels and sensitivity to hormones are related.Sex steroids regulate bone metabolism directly and indirectly through receptors on bone. Estrogen receptors (ER-∝, ER-β), progesterone receptor (PR), and androgen receptor (AR), have been previously identified on human osteosarcoma (OSA) cells, and are considered to influence tumor growth, but their expression and role in canine OSA is unknown. The aim of this study was to characterize sex hormone receptor expression levels in naturally occurring OSA tissue and in three canine OSA cell lines. The expression of ER-α, ER-β, PR, and AR was investigated using RT-PCR. PR expression levels were also quantified in OSA cells cultured under hypoxic conditions or in the presence of estradiol. The effects of progesterone on cell proliferation were quantified. Results demonstrated varying expression levels of these receptors in five OSA subtypes. OSA cell lines demonstrated high gene expression levels of PR and low gene expression levels of ER-α and ER-β and no gene expression of AR. PR expression was increased in OSA cells cultured under hypoxic conditions in a HIF-∝ independent manner. Interestingly, one cell line expressed very high levels of PR, expression of which decreased in response to estradiol. In addition, progesterone decreased OSA cell proliferation in this particular cell line. Further investigation of the role of sex steroids, particularly PR and its ligands, in regulation of canine OSA is recommended.

High Levels of Progesterone Receptor B in MCF-7 Cells Enable Radical Anti-Tumoral and Anti-Estrogenic Effect of Progestin.

The widely reported conflicting effects of progestin on breast cancer suggest that the progesterone receptor (PR) has dual functions depending on the cellular context. Cell models that enable PR to fully express anti-tumoral properties are valuable for the understanding of molecular determinant(s) of the anti-tumoral property. This study evaluated whether the expression of high levels of PR in MCF-7 cells enabled a strong anti-tumoral response to progestin. MCF-7 cells were engineered to overexpress PRB by stable transfection. A single dose of Promegestone (R5020) induced an irreversible cell growth arrest and senescence-associated secretory phenotype in MCF-7 cells with PRB overexpression (MCF-7PRB cells) but had no effect on MCF-7 cells with PRA overexpression. The growth-arresting effect was associated with downregulations of cyclin A2 and B1, CDK2, and CDK4 despite an initial upregulation of cyclin A2 and B1. R5020 also induced an evident activation of Nuclear Factor κB (NF-κB) and upregulation of interleukins IL-1α, IL-1β, and IL-8. Although R5020 caused a significant increase of CD24+CD44+ cell population, R5020-treated MCF-7PRB cells were unable to form tumorspheres and underwent massive apoptosis, which is paradoxically associated with marked downregulations of the pro-apoptotic proteins BID, BAX, PARP, and Caspases 7 and 8, as well as diminution of anti-apoptotic protein BCL-2. Importantly, R5020-activated PRB abolished the effect of estrogen. This intense anti-estrogenic effect was mediated by marked downregulation of ERα and pioneer factor FOXA1, leading to diminished chromatin-associated ERα and FOXA1 and estrogen-induced target gene expression. In conclusion, high levels of agonist-activated PRB in breast cancer cells can be strongly anti-tumoral and anti-estrogenic despite the initial unproductive cell cycle acceleration. Repression of ERα and FOXA1 expression is a major mechanism for the strong anti-estrogenic effect.

55P Morphological changes and regulation of hormone receptor-related proteins in breast cancer patients with higher levels of progesterone receptor A than B treated with mifepristone

Twenty breast cancer patients from Magdalena V. de Martínez Hospital, Buenos Aires, selected by their higher levels of progesterone receptor isoform A (PRA) than isoform B (PRB) were treated for 14 days with 200 mg mifepristone (MFP) p.o. before surgery (MIPRA; NCT02651844). A decrease in the proliferation index, greater than 30% was observed in 70% of the MFP-treated patients [primary endpoint (e.p.)]. RNA-Seq studies performed in 8 paired samples supported these findings and suggested that MFP increased tissue remodeling and immune-related pathways (secondary e.p.).

101P Proteome and RNA-Seq changes in breast cancer patients with higher levels of progesterone receptor A than B after mifepristone treatment

Antiprogestins inhibit experimental breast cancer growth when progesterone receptor isoform A (PRA) levels are higher than those of isoform B (PRB). Thus, we conducted a window of opportunity trial to determine the therapeutic effects of oral mifepristone (MFP, 200mg) in 20 breast cancer patients selected by their high PRA/PRB isoform ratio (MIPRA; NCT02651844). We identified 14/20 patients that achieved the pre-specified response parameter (30% relative reduction) showing a 49.62% decrease in Ki67 staining after MFP treatment (p = 0.0003).

Abstract P5-16-09: Transcriptome modulation by mifepristone treatment in breast cancer patients with higher levels of progesterone receptor A than B

Abstract Background: Preclinical data suggests that antiprogestins inhibit the growth of luminal breast carcinomas expressing higher levels of progesterone receptor isoform A (PRA) than isoform B (PRB). Thus, we designed a pre-surgical window of opportunity trial to determine the therapeutic effects of oral mifepristone (MFP) in 20 breast cancer patients selected by their high PRA/PRB isoform ratio (MIPRA; NCT02651844).Methods. MIPRA is an open-label, one-arm, prospective interventional study. After the selection process, 20 patients that met the inclusion criteria, with ER+, PRA/PRB>1.5 determined by Western blots, and total PR ≥50% determined by immunohistochemistry (IHC), were included for daily MFP treatment (200 mg/day p.o., 14 days). Core needle biopsies and surgical samples were formalin-fixed for IHC studies, and others were snap-frozen for further molecular studies. Besides, plasma samples were obtained for MFP dosing by LC-MS/MS. RNA was extracted from frozen tissue with a column-based method. The library for sequencing was constructed using SMART-Seq v4 Ultra Low Input RN from 8 paired samples in which tissue was available and passed the RNA quality control. Sequencing data was aligned with STAR and processed in R/Bioconductor. The counts matrix was obtained with featureCounts and differential expression paired analysis was performed with DESeq2. Gene set enrichment analysis (GSEA) was executed with the MSigDB collection.Results: A 49.62% decrease in Ki-67 staining was registered in all surgical specimens compared to baseline (p = 0.0003). Using the pre-specified response parameter (30% relative reduction) we identified 14/20 responders. Mifepristone was detected in all available plasma samples and the mean concentration was 308.33 ±57.91 ng/ml (716.71 ±134.62 nM). We conducted an RNA-seq analysis to explore changes at the transcript level. First, we conducted the study with all pairs of samples, without considering the response to MFP in Ki-67 studies. Unsupervised analysis showed the paired samples clustered together but neither the principal component analysis, nor the hierarchical clustering, showed any relevant cluster. The differential expression analysis identified 11 and 76 genes down- and up-regulated, respectively. We performed a GSEA based on KEGG databases and determined enriched pathways related to modulation of cell cycle and apoptosis. When the analysis was conducted with the REACTOME up-modulation of immune bioprocess and extracellular matrix re-modeling pathways such as degradation of extracellular matrix, activation of matrix metalloproteinases and collagen formation, were observed in the latter. Additionally, we observed down-modulation of pathways related to cell cycle such as DNA replication and synthesis, APC/C, and phase transition pathways. Interestingly, when we considered the tumors that responded to MFP according to Ki-67 data (n=4) and those that did not respond (n=4), we found that the non-responsive group shared some of the up and down modulated pathways as the responsive group.Conclusion: The results obtained in RNA-Seq data support the findings obtained by IHC indicating that MFP inhibits cell proliferation in luminal breast carcinomas with higher levels of PRA than PRB. The fact that MFP increased immune-related pathways is in line with previous preclinical data from our laboratory suggesting that MFP may prime this group of tumors for. further immune therapy. Interestingly, we found that some relevant pathways regarding inhibition of cell proliferation were also modulated in the non-responsive group suggesting that tumors in which a decrease in Ki-67 was not evident, may be still responding to MFP treatment. Ongoing analysis will determine changes in other markers that may help to further define MFP-responsive patients. Citation Format: Andres Elia, Martin Abba, Hugo Gass, Caroline A Lamb, Victoria T Fabris, Paula Martinez Vazquez, Javier Burruchaga, Eunice Spengler, Ines Caillet Bois, Alejandra Castets, Silvia Lovisi, Marcos Liguori, Gabriela Pataccini, Maria Florencia Abascal, Virginia Novaro, Alfredo Molinolo, Silvia I Vanzulli, Paola Rojas, Claudia Lanari. Transcriptome modulation by mifepristone treatment in breast cancer patients with higher levels of progesterone receptor A than B [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-16-09.

Single-cell transcriptome profiling of the human endometrium of patients with recurrent implantation failure.

Introduction: Despite great advances in assisted reproductive technology (ART), recurrent implantation failure (RIF) cannot be effectively avoided. Notably, cellular characteristics and communication that regulate endometrial receptivity and differentiation, and its disorders in RIF at window of implantation (WOI) remain rudimentary.Objectives: In this study, we profiled the endometrial cells present at the WOI timing in RIF patients and healthy controls using single-cell RNA sequencing (scRNA-seq) and provided a detailed molecular and cellular map of a healthy and RIF endometrium at the WOI.Method: In the current study, the endometrium from RIF patient (n = 6; age range, 32 - 35 years) and control (Ctrl) (n = 3; age range, 29 - 35 years) groups were studied at a single-cell resolution. single-cell RNA-seq and analysis were performed on the endometrium of patients with RIF and Ctrl. Immunofluorescence, flow cytometry assays, and quantitative real-time polymerase chain reaction (qRT-PCR) were performed to verify cellular identity and function.Results: We profiled the transcriptomes of 60222 primary human endometrial cells isolated from control and RIF patients at a single-cell resolution. We discovered dramatic differential expression of endometrial receptivity-related genes in four major endometrial fibroblast-like cells from RIF patients compared to the control endometrium. We observed that CD49a+CXCR4+NK cells were diminished in proportion with RIF. The decrease in subset of CD63highPGRhigh endometrial epithelial cells with high levels of progesterone receptor, autophagy and exosomes should contribute to the decrease in subset of NK cells. Additionally, we characterized aberrant molecular and cellular characteristics and endometrial cell-cell communication disorders in RIF patients.Conclusion: Our study provides deeper insights into endometrial microenvironment disorder of RIF that are potentially applicable to improving the etiological diagnosis and therapeutics of unexplained RIF.

Abstract B36: Progesterone receptor signaling in estrogen receptor-positive breast cancer

Abstract Approximately 65% of breast cancers are estrogen receptor alpha (ERα)-positive and depend on ERα signaling for growth. Clinical data indicate that ERα-positive breast cancers have a better prognosis when another hormone receptor, the progesterone receptor (PR), is coexpressed. Recent studies show that in the presence of its ligand progesterone, PR redirects ERα chromatin binding and transcriptional activity, thereby impacting prognosis and therapeutic response. To further examine PR function, we knocked out the PGR gene in the ERα-positive breast cancer cell lines MCF-7 and T47D using CRISPR-mediated gene editing. Knockout clones from each model were analyzed for ER- and PR-mediated transcriptional activity and gene expression profile, as well as response to selective estrogen receptor degraders (SERDs). The T47D cell line expresses high levels of PR in the presence or absence of estrogen, and stimulation with progesterone induces robust PR phosphorylation and modulation of PR target gene expression. Furthermore, progesterone attenuates estradiol-induced cell proliferation in a dose-dependent manner. Knockout of PR in this model abrogates the ability of progesterone to induce gene expression changes or attenuate estradiol-induced cell proliferation. In contrast, the MCF-7 cell line expresses low basal levels of PR, and is refractory to stimulation with progesterone. Interestingly, in estrogenic growth media, knockout of PR in this model results in basal gene expression changes that resemble a gene signature associated with tamoxifen resistance. Consistent with these findings, knockout of PR reduces the response to SERDs in a 5-day cell proliferation assay. ChIP-seq and RNA-seq analyses are ongoing to examine how loss of PR impacts ERα chromatin binding and transcriptional output. Citation Format: Amy Young, Jane Guan, Anneleen Daemen, Lori Friedman, Kui Lin. Progesterone receptor signaling in estrogen receptor-positive breast cancer [abstract]. In: Proceedings of the AACR Special Conference: Advances in Breast Cancer Research; 2017 Oct 7-10; Hollywood, CA. Philadelphia (PA): AACR; Mol Cancer Res 2018;16(8_Suppl):Abstract nr B36.

The unique transcriptional response produced by concurrent estrogen and progesterone treatment in breast cancer cells results in upregulation of growth factor pathways and switching from a Luminal A to a Basal-like subtype.

BackgroundIn breast cancer, progesterone receptor (PR) positivity or abundance is positively associated with survival and treatment response. It was initially believed that PR was a useful diagnostic marker of estrogen receptor activity, but increasingly PR has been recognised to play an important biological role in breast homeostasis, carcinogenesis and metastasis. Although PR expression is almost exclusively observed in estrogen receptor positive tumors, few studies have investigated the cellular mechanisms of PR action in the context of ongoing estrogen signalling.MethodsIn this study, we contrast PR function in estrogen pretreated ZR-75-1 breast cancer cells with vehicle treated ZR-75-1 and T-47D breast cancer cells using expression microarrays and chromatin immunoprecipitation-sequencing.ResultsEstrogen cotreatment caused a dramatic increase in the number of genes regulated by progesterone in ZR-75-1 cells. In T-47D cells that have naturally high levels of PR, estrogen and progesterone cotreatment resulted in a reduction in the number of regulated genes in comparison to treatment with either hormone alone. At a genome level, estrogen pretreatment of ZR-75-1 cells led to a 10-fold increase in the number of PR DNA binding sites detected using ChIP-sequencing. Time course assessment of progesterone regulated genes in the context of estrogen pretreatment highlighted a series of important regulatory pathways, including those driven by epithelial growth factor receptor (EGFR). Importantly, progesterone applied to cells pretreated with estradiol resulted in switching of the PAM50-determined intrinsic breast cancer subtype from Luminal A to Basal-like, and increased the Oncotype DX® Unscaled Recurrence Score.ConclusionEstrogen pretreatment of breast cancer cells increases PR steady state levels, resulting in an unequivocal progesterone response that upregulates key members of growth factor pathways. The transformative changes progesterone exerts on the breast cancer subtype suggest that these subtyping tools should be used with caution in premenopausal women.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-015-1819-3) contains supplementary material, which is available to authorized users.

Characterization of LHY-821, a novel moderately differentiated endometrial carcinoma cell line

Endometrial cancer is a major problem for women but only a small number of comprehensively characterized cell models are available for studies. Here, we established a new cell line derived from a Stage IIIc(1) Grade 2 endometrial adenocarcinoma. The cell line, designated LHY-821, was characterized using growth curve, karyotyping, immunohistochemical staining, immunoblotting, drug sensitivity assay, invasion assay, and xenografting in nude mice. LHY-821 has a doubling time of about 46h and a colony-forming efficiency of approximately 71%. These cells expresse high levels of progesterone receptor but not estrogen receptor and are sensitive to medroxyprogesterone acetate (MPA). LHY-821 also expresses pan-cytokeratin, PTEN, p53, β-catenin, IGF-1, and IGF-2. In addition, karyotype analysis revealed that LHY-821 possessed a near diploid karyotype including 6q-, 10p-, Xq-, 13q+, 17p+, and Triplo-12. LHY-821 showed highly tumorigenicity in nude mice (100%) and weak invasiveness. Chemosensitivity tests showed that LHY-821 was sensitive to both carboplatin and paclitaxel. LHY-821 is an immortalized cell line which had survived more than 80 serial passages; it may provide a novel tool to study the molecular mechanism and potential treatment for endometrial cancer.

Abstract C239: Progesterone receptor B through Sp1 induces the phosphatase and tensin homologue deleted from chromosome 10 (PTEN) gene promoter activity and cell death in breast cancer cell lines.

Abstract Phosphatase and tensin homolog on chromosome 10 (PTEN) is a tumor suppressor attracting considerable attention as new target for effective therapeutic strategies in breast cancer. While recent studies suggest the down-regulation of PTEN by Estrogen Receptor alpha, the role of Progesterone Receptor (PR) is not clearly established. In the present study we report that PR produces a significant inhibition of MCF-7 breast cancer cell survival with a concomitant increase of PTEN protein, mRNA and gene promoter activity. On the contrary PRs failed to enhance the transcriptional activity of the PTEN promoter lacking the region between −1398 bp to −1118 bp. By analyzing this DNA sequence to search potential binding sites for transcription factors, we observed that this region contains several DNA motifs known to bind the transcription factors Sp1that has been shown to interact with PRs, activating specific gene transcription. In our experimental conditions mithramycin, prevented the PTEN promoter transactivation on OHPg treatment. ChIP analyses showed specific recruitment of PR, Sp1 and RNA polymerase II upon progesterone treatment at PTEN promoter. The persistence of such effects in T47 cells which express high PR levels, addresses how PR/PTEN signalling may be considered as useful pharmacological tool to be exploited for innovative adjuvant strategies in breast cancer treatment. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr C239.

A Steroid-Conjugated Magnetic Resonance Probe Enhances Contrast in Progesterone Receptor Expressing Organs and Tumors in Vivo

Progesterone receptor (PR) is a significant biomarker in diseases such as endometriosis and breast, ovarian, and uterine cancers that is associated with disease prognosis and therapeutic efficacy. While receptor status is currently determined by immunohistochemistry assays, the development of noninvasive PR imaging agents could improve molecular characterization, treatment decisions, and disease monitoring. ProGlo, a progesterone-conjugated magnetic resonance imaging (MRI) contrast agent, was evaluated in vivo to determine whether it targets and enhances signal intensity in organs and tumors that express high PR levels. A tissue distribution study indicated that ProGlo accumulates in the PR-rich uterus, which was confirmed by in vivo imaging studies. Ex vivo images of these organs revealed that ProGlo was distributed in the substructures that express high PR levels. In xenograft tumor models, ProGlo was taken up to a greater extent than the nonfunctionalized Gd-DO3A in tumors, particularly in PR(+) tumors. The ability to accumulate and enhance signal intensity in PR(+) organs and tumors suggests that ProGlo may be a promising MRI probe for PR(+) diseases.

Abstract 1315: DNMT and HDAC inhibitors resensitize resistant tumors to antiprogestin therapy in a mouse breast cancer model

Abstract Mouse mammary carcinomas induced by medroxyprogesterone acetate (MPA) are maintained by syngeneic transplantation and tumor variants with different hormone responsiveness have been generated. Tumors which grow without the MPA supply were defined as hormone independent (HI). Most of these tumors regress after antiprogestin (mifepristone, MIF) administration, while others show constitutive resistance to hormone therapy. We have previously demonstrated that responsive tumors show higher levels of progesterone receptor A (PRA) than PRB, while the opposite occurs in resistant tumors, indicating that the PR isoform ratio might be a predictor of antiprogestin responsiveness. We have also demonstrated that in constitutive resistant tumors PRA is silenced by methylation of its promoter, and that treatment with 5azadC (demethylating agent) induces PRA expression and tumors partially recover antiprogestin responsiveness. The aim of this study was to investigate whether the co-treatment with DNA methyltransferase (DNMT) and histone deacetylase (HDAC) inhibitors would enhance even more the responsiveness to MIF. For this purpose we investigated the expression of HDAC1 in constitutive resistant tumors and whether treatment of these tumors with 5azadC and trichostatin A (TSA; HDAC inhibitor) restored MIF responsiveness better than 5azadC alone. The constitutive resistant tumor 59-HI showed a higher expression of HDAC1 as compared to the antiprogestin sensititive tumor 59-2-HI as determined by Western blot (p<0,001) and immunohistochemistry, suggesting that these tumors might be candidates to respond to an HDAC inhibitor. 59-HI tumors were implanted sc in BALB/c mice and when tumors were palpable they were treated with a) 5azadC (1 mg/kg/every other day; ip), b) MIF (12 mg/kg/daily; sc), c) 5azadC plus MIF or d) TSA (1 mg/kg/every other day; sc), e) TSA plus 5azadC, f) TSA, 5azadC and MIF, or g) vehicle. Interestingly the co-treatment of 5azadC, TSA and MIF induced a significant inhibitory effect that was even greater than the one induced by 5azadC plus MIF (p<0.001). No inhibitory effects were observed when each compound was used alone or with TSA and MIF. The expression of PR was evaluated in control, 5azadC- and TSA plus 5azadC- treated tumors. An increase in PRA expression was observed by western blot and immunohistochemistry. The increase of PRA was higher with TSA plus 5azadC than with 5azadC alone (p<0,001). In conclusion, we report for the first time that a) constitutive resistant mammary carcinomas show high levels of HDAC1 which might be responsible for histone acetylation within the methylated PRA promoter, b) the combined treatment of demethylating agents and HDAC inhibitors increases the efficiency in both, PR re-expression and antiprogestin sensitivity. These results support our hypothesis that tumors with high levels of PRA are those which respond to antiprogestin treatment. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1315. doi:10.1158/1538-7445.AM2011-1315

Extrapleural Solitary Fibrous Tumor: A Clinicopathologic Study of 19 Cases

This study reports a series of 19 extrapleural solitary fibrous tumors. The patients included 6 men and 13 women with age ranging from 27 to 86 years. Three patients showed local recurrence. In 2 tumors, a diagnosis of malignancy was made. All of the tumors were strongly positive for CD34, and 3 of them expressed high levels of progesterone receptor. Solitary fibrous tumors are fairly rare, occurring in many parts of the body, and their behavior is unpredictable.

Regulation of Progesterone Receptor Expression by Estradiol Is Dependent on Age, Sex and Region in the Rat Brain

Progesterone receptor (PR) expression is highly dependent on estradiol in the medial preoptic nucleus (MPN) and the ventromedial nucleus (VMN) of the adult rat brain. During development, males express high levels of PR in the MPN, whereas females have virtually no PR, a sex difference resulting entirely from differential exposure to estradiol. Because PR is also estradiol dependent in the adult VMN, the present study examined the regulation of PR immunoreactivity (PRir) in the developing VMN. Surprisingly, PRir was present at high levels in the VMN of both neonatal males and females. In the neonatal VMN, PR expression was dependent on gonadal hormones in males but not females. When females were ovariectomized and exposed to estradiol at various ages from neonatal to adulthood, estradiol reliably induced PRir in the MPN at postnatal d 7 but failed to induce PRir in the VMN of the same animals. Only later in development, around postnatal d 14, did estradiol increase PRir in the female VMN. There appears to be a developmental switch in the VMN when PR expression changes from estradiol independent to estradiol dependent. Furthermore, this switch is anatomically specific and does not exist in the MPN. The present results indicate that the regulation of PR expression by estradiol is dependent on age, sex, and brain region, suggesting that PR may play a critical but specific role in the normal development of these reproductively important brain areas. In addition, the neonatal female VMN may provide a unique model in which to examine the mechanisms underlying the specificity of steroid-induced gene expression.

BORIS, a paralogue of the transcription factor, CTCF, is aberrantly expressed in breast tumours

BORIS (for brother of the regulator of imprinted sites), a paralogue of the transcription factor, CTCF, is a novel member of the cancer-testis antigen family. The aims of the present study were as follows: (1) to investigate BORIS expression in breast cells and tumours using immunohistochemical staining, western and real-time RT–PCR analyses and (2) assess potential correlation between BORIS levels in tumours with clinical/pathological parameters. BORIS was detected in all 18 inspected breast cell lines, but not in a primary normal breast cell culture. In 70.7% (41 of 58 cases) BORIS was observed in breast tumours. High levels of BORIS correlated with high levels of progesterone receptor (PR) and oestrogen receptor (ER). The link between BORIS and PR/ER was further confirmed by the ability of BORIS to activate the promoters of the PR and ER genes in the reporter assays. Detection of BORIS in a high proportion of breast cancer patients implies potential practical applications of BORIS as a molecular biomarker of breast cancer. This may be important for diagnosis of the condition and for the therapeutic use of BORIS. The ability of BORIS to activate promoters of the RP and ER genes points towards possible involvement of BORIS in the establishment, progression and maintenance of breast tumours.

Progesterone and Breast Cancer

Mutations in the breast cancer susceptibility gene BRCA1 greatly increase a woman’s risk of developing breast and ovarian cancers. Why do these mutations predominantly affect hormone-responsive tissues when the mutant gene is widely expressed throughout the body? Poole et al . (see the news story by Marx) suggest that this tissue specificity is caused in part by BRCA1-mediated effects on signaling by the hormone progesterone. Mammary epithelial cells (MECs) of Brca-1/p53 -deficient mice accumulated high levels of progesterone receptors, probably through defective degradation by the proteasome, and developed aberrant proliferation of the MECs. Treatment with the progesterone antagonist mifepristone (RU 486) prevented or delayed mammary tumor development in the mice. A. J. Poole, Y. Li, Y. Kim, S.-C. J. Lin, W.-H. Lee, E. Y.-H. P. Lee, Prevention of Brca1 -mediated mammary tumorigenesis in mice by a progesterone antagonist. Science 314 , 1467-1470 (2006). [Abstract] [Full Text] J. Marx, Squelching progesterone’s signal may prevent breast cancer. Science 314 , 1370 (2006). [Summary] [Full Text]

Progestin-induced caveolin-1 expression mediates breast cancer cell proliferation

Progestin regulation of gene expression was assessed in the progestin-dependent murine tumor line C4HD which requires MPA, a synthetic progestin, for in vivo growth and expresses high levels of progesterone receptor (PR). By using suppressive subtractive hybridization, caveolin-1 was identified as a gene whose expression was increased with in vivo MPA treatment. By Northern and Western blot analysis, we further confirmed that caveolin-1 mRNA and protein expression increased in MPA-treated tumors as compared with untreated tumors. When primary cultures of C4HD cells were treated in vitro with MPA, caveolin-1 levels also increased, effect that was abolished by pre-treatment with progestin antagonist RU486. In addition, MPA promoted strong caveolin-1 promoter transcriptional activation both in mouse and human breast cancer cells. We also showed that MPA regulation of caveolin-1 expression involved in activation of two signaling pathways: MAPK and PI-3K. Short-term MPA treatment of C4HD cells led to tyrosine phosphorylation of caveolin-1 protein, where Src was the kinase involved. Additionally, we showed that MPA-induced association of caveolin-1 and PR, which was detected by coimmunoprecipitation and by confocal microscopy. Finally, we proved that MPA-induced proliferation of C4HD cells was inhibited by suppression of caveolin-1 expression with antisense oligodeoxynucleotides to caveolin-1 mRNA. Furthermore, we observed that inhibition of caveolin-1 expression abrogated PR capacity to induced luciferase activity from a progesterone response element-driven reporter plasmid. Comprehensively, our results demonstrated for the first time that caveolin-1 expression is upregulated by progestin in breast cancer. We also demonstrated that caveolin-1 is a downstream effector of MPA that is partially responsible for the stimulation of growth of breast cancer cells.

Complex agonist-like properties of ICI 182,780 (Faslodex) in human breast cancer cells that predominantly express progesterone receptor-B: implications for treatment resistance.

ICI 182,780 (Faslodex), considered a pure anti-estrogen, is approved for treatment of post-menopausal breast cancer patients who fail to respond to tamoxifen therapy. We recently reported that, like mifepristone, ICI 182,780 exhibits anti-progestin activity, blocking the progestin-dependent increase in endogenous vascular endothelial growth factor (VEGF) mRNA and protein release. Some anti-progestins have partial agonist-like activity in breast cancer cells expressing high levels of progesterone receptor B (PRB). Our results show that ICI 182,780 can also induce reporter activity from a plasmid containing a simple progestin responsive element (PRE) in these cells. Using small interfering RNA, we determined that induction is dependent on the presence of PR, estrogen receptor and SRC-1. Regulation of more complex progestin-responsive promoters was context-dependent; induction was observed from the MMTV promoter but not from the VEGF promoter. In contrast, ICI 182,780 increased the release of angiogenically active VEGF from cells expressing elevated levels of PRB. This effect was dependent on the phosphatidylinositol-3 kinase and ERK/MAPK signaling pathways. We hypothesize that these agonist-like properties of ICI 182,780 (one genomic and one non-genomic) may contribute to the acquisition of drug resistance, suggesting that both anti-hormonal and anti-angiogenic treatment may be appropriate in these patients.

Immunohistochemical studies on the progesterone receptor (PR) in the sow uterus during the oestrous cycle and in inseminated sows at oestrus and early pregnancy

Physiological changes in the sow uterus involve the regulation by progesterone and its receptor proteins (PR). Therefore, the aim of the present study was to investigate the localization of PR during different stages of the oestrous cycle and in inseminated sows during early pregnancy by use of immunohistochemistry. Uterine samples were collected from cyclic and inseminated sows at different stages of the oestrous cycle and early pregnancy. The samples were fixed in 10% formaldehyde and embedded in paraffin. Immunohistochemistry was done by use of a mouse monoclonal antibody to PR. The highest PR immunostaining in the surface epithelium was observed at oestrus/5-6 h after artificial insemination (AI) and early dioestrus/70 h after AI. In the glandular epithelium, the highest level of PR was found at oestrus with the lowest at late dioestrus/d 19. Higher levels of PR were observed in inseminated groups compared with cyclic sows. In the myometrium, a high level of PR was found at oestrus, while stromal PR cells were constantly present throughout the oestrous cycle and at different stages of early pregnancy. In conclusion, this study shows that the immunopresence of PR in the sow uterus differed between uterine compartments at the same reproductive stage. Differences were also found for some uterine compartments between cyclic and inseminated/early pregnant sows. The relatively consistent immunostaining of PR in the stroma strengthens a stromal role in the regulation of physiological activities in the sow uterus during the oestrous cycle as well as early pregnancy.