Content uploaded by Ana C Zenclussen
Author content
All content in this area was uploaded by Ana C Zenclussen on Feb 21, 2014
Content may be subject to copyright.
A preview of the PDF is not available
GPER functions as a tumor suppressor in MCF-7 and SK-BR-3 breast cancer cells
Abstract and Figures
The orphan, membrane-bound estrogen receptor (GPER) is expressed at high levels in a large fraction of breast cancer patients, and its expression is favorable for patients' survival. We investigated the role of GPER as a potential tumor suppressor in MCF-7 and SK-BR-3 breast cancer cells.
The effect of GPER agonist G-1 in cell culture was used to determine whether GPER inhibit cell growth. The methylation status of GPER promoter was investigated by methylation-specific PCR.
GPER-specific agonist G-1 inhibited breast cancer cell proliferation in concentration-dependent manner via induction of the cell cycle arrest in M-phase, enhanced phosphorylation of histone 3 and cell apoptosis. Analysis of the methylation status of the GPER promoter in MCF-7 and SK-BR-3 cells revealed that GPER expression is regulated by epigenetic mechanisms and GPER expression is inactivated by promoter methylation. Overall, our results are consistent with our recent findings in triple-negative breast cancer cells, and the cell surface expression of GPER makes it an excellent potential therapeutic target for non-triple-negative breast cancer.
Figures - uploaded by Ana C Zenclussen
Author content
All figure content in this area was uploaded by Ana C Zenclussen
Content may be subject to copyright.
... On a different note, research by Sathya et al. [19] indicated that under low oxygen conditions, estrogen suppresses breast cancer growth via the GPER1/ROS/p38 MAPK/p21 signaling pathway. Weißenborn's findings [34,35] demonstrated that the GPER1-specific agonist G-1 activates GPER1 in a concentration-dependent manner, effectively inhibiting breast cancer cell growth. This suggests that cell surfaceexpressed GPER1 holds promise as a potential therapeutic target for non-triple-negative and triplenegative breast cancers. ...
Background:
The G protein-coupled oestrogen receptor (GPER) 1 mediates non-genomic oestrogen-related signalling and plays an important role in the regulation of cell growth and programmed cell death through multiple downstream pathways. Despite the increasing interest in the role of GPER1 in cancer development, no pan-cancer analysis has been available for GPER1.
Methods:
In this study we performed a comprehensive analysis of the role of GPER1 in pan-cancer via Human Protein Atlas (HPA), The Cancer Genome Atlas (TCGA), University of California, Santa Cruz Xena (UCSC XENA), Genotype-Tissue Expression (GTEx), MethSurv, The University of Alabama at Birmingham CANcer data analysis Portal (UALCAN), cBioPortal, STRING and TISIDB detabases, followed by enrichment analysis using R software.
Results:
GPER1 was widely expressed in tissues and organs and differed in expression from normal tissue in a variety of cancers. In diagnostic assessment, it's Area Under the Curve (AUC) surpassed 0.9 in nine cancer types. Survival analysis showed that GPER1 was correlated with the prognosis of 11 cancer types. Moreover, GPER1 expression was associated with immune infiltration in multiple cancers.
Conclusions:
In summary, GPER1 has good diagnostic or prognostic value across various malignancies. Together with its extensive correlation with immune components, the aforementioned results suggests that GPER1 shows promise in tumour diagnosis and prognosis, providing new ideas for precise and personalised anti-tumour strategies.
... As shown in Figure 2B, LTED cells exhibited more aggressive proliferative features than MCF-7 cells. The reduced expression of GPER1 favors cell aggressiveness, such as stimulated cell proliferation, particularly in LTED cells, as indicated in the reports suggesting that GPER1 can act as a tumor suppressor in different types of cancers, including breast cancer [33][34][35][36]. Therefore, the possible negative (antiproliferative) effects of GPER1 agonists (G-1 and 2-MeO-E2) [8,17] on the aggressive behavior of LTED cells were not observed or weakened. ...
To identify effective treatment modalities for breast cancer with acquired resistance, we first compared the responsiveness of estrogen receptor-positive breast cancer MCF-7 cells and long-term estrogen-deprived (LTED) cells (a cell model of endocrine therapy-resistant breast cancer) derived from MCF-7 cells to G-1 and 2-methoxyestradiol (2-MeO-E2), which are microtubule-destabilizing agents and agonists of the G protein-coupled estrogen receptor 1 (GPER1). The expression of GPER1 in LTED cells was low (~0.44-fold), and LTED cells displayed approximately 1.5-fold faster proliferation than MCF-7 cells. Although G-1 induced comparable antiproliferative effects on both MCF-7 and LTED cells (IC50 values of >10 µM), 2-MeO-E2 exerted antiproliferative effects selective for LTED cells with an IC50 value of 0.93 μM (vs. 6.79 μM for MCF-7 cells) and induced G2/M cell cycle arrest. Moreover, we detected higher amounts of β-tubulin proteins in LTED cells than in MCF-7 cells. Among the β-tubulin (TUBB) isotype genes, the highest expression of TUBB2B (~3.2-fold) was detected in LTED cells compared to that in MCF-7 cells. Additionally, siTUBB2B restores 2-MeO-E2-mediated inhibition of LTED cell proliferation. Other microtubule-targeting agents, i.e., paclitaxel, nocodazole, and colchicine, were not selective for LTED cells. Therefore, 2-MeO-E2 can be an antiproliferative agent to suppress LTED cell proliferation.
... several cancer types [21][22][23][24][25] . GPER is distinct from classical nuclear estrogen receptors and has been shown to signal through cAMP and calcium in multiple cellular contexts [26][27][28][29] . ...
Despite recent therapeutic advances, the 5-year survival rate for adults with acute myeloid leukemia (AML) is poor and standard-of-care chemotherapy is associated with significant toxicity, highlighting the need for new therapeutic approaches. Recent work from our group and others established that the G protein-coupled estrogen receptor (GPER) is tumor suppressive in melanoma and other solid tumors. We performed a preliminary screen of human cancer cell lines from multiple malignancies and found that LNS8801, a synthetic pharmacologic agonist of GPER currently in early phase clinical trials, promoted apoptosis in human AML cells. Using human AML cell lines and primary cells, we show that LNS8801 inhibits human AML in preclinical in vitro models, while not affecting normal mononuclear cells. Although GPER is broadly expressed in normal and malignant myeloid cells, this cancer-specific LNS8801-induced inhibition appeared to be independent of GPER signaling. LNS8801 induced AML cell death primarily through a caspase-dependent apoptosis pathway. This was independent of secreted classical death receptor ligands, and instead required induction of reactive oxygen species (ROS) and activation of endoplasmic reticulum (ER) stress response pathways including IRE1α. These studies demonstrate a novel activity of LNS8801 in AML cells and show that targeting ER stress with LNS8801 may be a useful therapeutic approach for AML.
Significance
Previous work demonstrated that LNS8801 inhibits cancer via GPER activation, especially in solid tumors. Here we show that LNS8801 inhibits AML via GPER-independent mechanisms that include ROS induction and ER activation.
... The observation that GPER protein is rarely expressed in human breast cancer cases is noteworthy, given the substantial number of reports discussing the role of GPER in preclinical models of breast cancer (16). Although GPER signaling was originally reported to be tumor promoting in some breast cancer models (17), subsequent reports show that GPER signaling inhibits breast cancer (18)(19)(20). Additionally, several studies have documented correlations between GPER expression and prognosis in breast cancer, with contradictory results (21,22). ...
GPER (G protein-coupled estrogen receptor) has been reported to play roles in several areas of physiology including cancer, metabolic disorders, and cardiovascular disease. However, the understanding of where this receptor is expressed in human tissue is limited due to limited available tools and methodologies that can reliably detect GPER protein. Recently, a highly specific monoclonal antibody against GPER (20H15L21) was developed and is suitable for immunohistochemistry. Using this antibody, we show that GPER protein expression varies markedly between normal human tissue, and also among cancer tissue. As GPER is an emerging therapeutic target for cancer and other diseases, this new understanding of GPER distribution will likely be helpful in design and interpretation of ongoing and future GPER research.
... GPER1 functions as a tumor suppressor and promotes apoptosis and antiproliferation signaling in ER-positive BC cells. 12,36 However, GPER1 expression is reportedly correlated with pro-metastasis pathways and promotes migration and invasion of ER-negative BC cells. 37 Several studies have noted a different prognostic effect of GPER1 in ER-positive versus ER-negative BC. 8,12 Hence, subgroup analysis of the gene expression data was performed to explore the prognostic effect of GPER1 based on ER status. ...
Purpose: To determine whether G protein-coupled estrogen receptor 1 (GPER1) is a suitable biomarker to predict the treatment outcome of breast cancer (BC). Methods: A meta-analysis of the literature was performed to clarify the correlation between GPER1 protein expression and BC outcome. The relationship between GPER1 mRNA expression and survival was analyzed using Breast Cancer Gene-Expression Miner (bc-GenExMiner) v4.6 software. Results: Six studies involving 2697 patients were included in the meta-analysis. Four studies reported the correlation between GPER1 protein expression and relapse-free survival (RFS) and 4 others reported the impact of GPER1 protein expression on overall survival (OS). The results showed that high GPER1 protein expression was not associated with RFS (hazard ratio [HR] = 1.58; 95% confidence interval [CI] = 0.71-3.48; P = .26) or OS (HR = 1.18; 95% CI = 0.64-2.18; P = .60). Subgroup analysis suggested that nuclear expression of GPER1 was not associated with OS (HR = 0.91; 95% CI = 0.77-1.08; P = .30), but high expression of cytoplasmic GPER1 was significantly associated with longer OS (HR = 0.69; 95% CI = 0.55-0.86; P = .001). Furthermore, the association of GPER1 mRNA and OS of BC patients was analyzed using bc-GenExMiner v4.6. Two data sets involving 4016 patients were included in the analysis. The targeted prognostic analysis results showed that high mRNA expression of GPER1 was predictive of better OS in BC patients (HR = 0.71; 95% CI = 0.59-0.86; P = .0005), which was remarkably similar to the result of cytoplasmic GPER1. Further subgroup analysis demonstrated that high mRNA expression of GPER1 was predictive of better OS in estrogen receptor (ER)-positive, but not ER-negative or triple-negative BC patients. Conclusions: High mRNA and cytoplasmic protein expression of GPER1 were predictive of better OS of BC patients.
... Some subgroups of ERα-negative BRCA cells, such as SKBR-3 cells, are membrane G protein-coupled estrogen receptor (GPER) positive (Steiman et al., 2013). In vitro research claimed that GPER might function as a tumor suppressor in BRCA cells (Ariazi et al., 2010;Weißenborn et al., 2014). Recent studies revealed that GPER and its mediated signaling pathway [phosphatidylinositide 3-kinase (PI3K)/AKT] have a vital role in the proliferation of BRCA cells (Molina et al., 2017;Hsu et al., 2019). ...
Breast cancer has recently been known as the first lethal malignancy in women worldwide. Despite the existing treatments that have improved the patients’ prognosis, some types of breast cancer are serious challenges to treat. Therefore, efforts are underway to provide more efficient therapy. Cryptotanshinone (CPT) is a liposoluble diterpenoid derivation of a traditional Chinese herbal medicine called Salvia miltiorrhiza Bunge. It has been considered in the past decades due to its vast therapeutic properties, including anti-tumor, anti-inflammatory, and anti-fibrosis. Recently, studies have found that CPT showed a significant anti-breast cancer effect in vivo and in vitro through different physiological and immunological mechanisms. This study summarized the latest research findings on the antitumor effect of CPT in breast cancer. Further, the main molecular mechanisms based on breast cancer types and combination with other drugs were reviewed to provide essential evidence for future longitudinal research and its clinical application in breast cancer treatment.
Naringenin is a flavanone able to suppress the growth of various cancer cells including estrogen receptor-alpha-66 (ERα66)-positive breast cancer cells, but the anti-tumorigenic roles of naringenin in ERα66-negative breast cancer remain unclear. This study aims to determine the potential effects of naringenin on two ERα66-negative breast cancer cells (SKBR3 and MDA-MB-231) and to define their mechanisms of action. Herein, several cellular biological behaviors, including cell viability, migration, cycle, apoptosis, reactive oxygen species (ROS) generation, and oxidative stress (e.g., LDH, SOD and GSH), were evaluated in the present study. Then the gene expression levels involved in cell migration, cycle and apoptosis, as well as two membrane estrogen receptors (ERα36 and GPR30) were analyzed with real-time quantitative polymerase chain reaction (RT-qPCR) assay and western blotting. Results showed that naringenin exerted a concentration-dependent response on the growth of both ERα66-negative cell lines. The cell growth regulation was accompanied by modulation of cell migration, cycle and apoptosis, which were dependent on the ROS-regulated p53 signaling cascade. Importantly, ERα36 and GPR30 were involved in the (anti)-proliferative effects. These findings indicate naringenin possesses contrasting effects on ERα66-negative breast cancer cell growth depending on its dose
The high reoperation rate in pelvic organ prolapse (POP) surgery is probably due to impaired wound healing, and therefore, novel strategies that promote tissue regeneration are urgently needed. Estrogen therapy shows beneficial effects for vaginal wound healing: it reduces the inflammatory response and improves vaginal tissue strength and quality. Earlier, scaffolds that release the most potent estrogen, β‐estradiol (E2) have been described. This work describes the results of conjugating E2 covalently to the synthetic, but highly mimetic, polyisocyanide hydrogel (PIC‐E2). As adipose‐derived stem cells (ASCs) possess regenerative capabilities, the combination of PIC‐E2 and ASCs may have a synergistic effect on pelvic floor tissue regeneration. The results show that the PIC‐E2 bioactivity is not hampered upon covalent E2 conjugation and upregulates various extracellular matrix (ECM) genes in ASCs. Moreover, PIC‐E2 exerts similar effects regarding ECM metabolism compared to other E2‐releasing biomaterials, but at much lower doses, which is economically and therapeutically favorable. Based on these in vitro findings, the combination of the PIC‐E2 hydrogel and ASCs may have the potential to stimulate tissue regeneration in vivo, and therefore, feasibly improve the surgical outcomes for POP in the future.
Over the past decades, designing of privileged structures has emerged as a useful approach to the discovery and optimisation of novel biologically active molecules, and many have been successfully exploited across and within different target families. Examples include indole, quinolone, isoquinoline, benzofuran and chromone, etc. In the current study, we focus on synthesising a novel hybrid scaffold constituting naturally occurring benzophenone (14) and indanone (22) ring systems, leading to a general structure of 2-(diphenylmethylene)-2,3-dihydro-1H-inden-1-one (23). It was hypothesised this new hybrid system would provide enhanced anti-cancer activity owing to the presence of the common features associated with the tubulin binding small molecule indanocine (10) and the estrogen receptor (ER) antagonist tamoxifen (24). Key hybrid molecules were successfully synthesised and characterised, and the in vitro cytotoxicity assays were performed against cancer cell lines: MCF7 (breast) and SKBR3 (breast), DU145 (prostate) and A549 (lung). The methyl-, chloro- and methoxy-, para-substituted benzophenone hybrids displayed the greatest degree of cytotoxicity and the E-configuration derivatives 45, 47 and 49 being significantly most potent. We further verified that the second benzyl moiety of this novel hybrid scaffold is fundamental to enhance the cytotoxicity, especially in the SKBR3 (HER2+) by the E-methyl lead molecule 47, MCF7 (ER+) by 45 and 49, and A549 (NSCLC) cell lines by 49. These hybrid molecules also showed a significant accumulation of SKBR3 cells at S-phase of the cell cycle after 72 hrs, which demonstrates besides of being cytotoxic in vitro against SKBR3 cells, 47 disturbs the replication and development of this type of cancer causing a dose-dependent cell cycle arrest at S-phase. Our results suggest that DNA damage might be involved in the induction of SKBR3 cell death caused by the hybrid molecules, and therefore, this novel system may be an effective suppressor of HER2+/Neu-driven cancer growth and progression. The present study points to potential structural optimisation of the series and encourages further focussed investigation of analogues of this scaffold series toward their applications in cancer chemoprevention or chemotherapy.
Menopausal women often face long-term estrogen treatment. G protein-coupled estrogen receptor (GPER) expressed in intestinal crypt was activated by estrogen therapy, but it was unclear whether chronic GPER activation during menopause had an effect on intestinal stem cells (ISCs). We tested the effect of chronic GPER activation on ISCs of ovariectomized (OVX) mice by injection of the selective GPER agonist G-1 for 28 days, or G-1 stimulation of organoids derived from crypts of OVX mice. G-1 up-regulated crypt depth, the number of Ki67+, bromodeoxyuridine+ cells and Olfm4+ ISCs, and the expression of ISCs marker genes (Lgr5, Olfm4 and Axin2). G-1 administration promoted organoid growth, increased the number of EdU+ cells per organoid and protein expression of Cyclin D1 and cyclin B1 in organoids. After G-1 treatment in vivo or in vitro, Paneth cell derived Wnt3, Wnt3 effector β-catenin and Wnt target genes c-Myc and Cyclin D1 increased in ileum or organoids. Once blocking the secretion of Wnt3 from Paneth cells, the effects of G-1 on organoids growth, ISCs marker genes and Wnt/β-catenin signaling were abolished. G-1 did not affect the number of Paneth cells in ex vivo organoids, while activated Mmp7/cryptdin program in Paneth cells, promoted their maturation, and increased the expression of lysozyme protein. G-1 pretreatment in OVX mice inhibited radiation-induced ISCs proliferation injury and enhanced the resistance of mice to intestinal injury. In conclusion, chronic GPER activation prompted the Wnt3 synthesis in Paneth cells, thus increased the proliferation of ISCs via activation of Wnt3/β-catenin signaling in OVX mice.
We have previously shown that estrogens binding to estrogen receptor (ER) α increase proliferation of Leydig tumor cells. Estrogens can also bind to G protein-coupled ER (GPER) and activation of this receptor can either increase or decrease cell proliferation of several tumor types. The aim of this study was to investigate GPER expression in R2C rat tumor Leydig cells, evaluate effects of its activation on Leydig tumor cell proliferation and define the molecular mechanisms triggered in response to its activation. R2C cells express GPER and its activation, using the specific ligand G-1, is associated with decreased cell proliferation and initiation of apoptosis. Apoptosis after G-1 treatment was asserted by appearance of DNA condensation and fragmentation, decrease in Bcl-2 and increase in Bax expression, cytochrome c release, caspase and poly (ADP-ribose) polymerase-1 (PARP-1) activation. These effects were dependent on GPER activation because after silencing of the gene, using a specific small interfering RNA, cyt c release, PARP-1 activation and decrease in cell proliferation were abrogated. These events required a rapid, however, sustained extracellular regulated kinase 1/2 activation. G-1 was able to decrease the growth of R2C xenograft tumors in CD1 nude mice while increasing the number of apoptotic cells. In addition, in vivo administration of G-1 to male CD1 mice did not cause any alteration in testicular morphology, while cisplatin, the cytotoxic drug currently used for the therapy of Leydig tumors, severely damaged testicular structure, an event associated with infertility in cisplatin-treated patients. These observations indicate that GPER targeting for the therapy of Leydig cell tumor may represent a good alternative to cisplatin to preserve fertility in Leydig tumor patients.
It is known that the new membrane-bound estrogen receptor GPER-1 acts suppressive in breast cancer cells and its expression decreases during disease progression. This study was conducted to evaluate the GPER-1 expression in ovarian cancer and its correlation with progression. Its function was tested in vitro in ovarian cancer cells.Patients and methods: GPER-1 expression was analyzed by immunohistochemistry in 35 benign ovarian tumors, 35 tumors of low-malignant potential and in 124 ovarian cancers. GPER-1 expression was correlated to the prospectively evaluated disease-free survival of ovarian cancer patients. We also tested GPER-1 expression in ovarian cancer cells and the effect of GPER-1 stimulation on cell growth.
GPER-1 expression was significantly lower in ovarian cancer tissue than in benign and low-malignant ovarian tumors. GPER-1 expression was observed in 83.1% of malignant tumors and was higher in early stage cancers and tumors with high histological differentiation. GPER-1 expression was associated with favourable clinical outcome. The difference in 2-year disease-free survival by GPER-1 expression was significant, 28.6% for GPER-1 negative and 59.2% for GPER-1 positive cases (p = 0.002). GPER-1 expression was observed in SKOV-3 and OVCAR-3 ovarian cancer cell lines. G-1, a selective GPER-1 agonist, suppressed proliferation of the two cell types via inhibition of cell cycle progression in G2/M phase and stimulation of caspase-dependent apoptosis. The blockade in G2/M phase was associated with increased expression of cyclin B1 and Cdc2 and phosphorylation of histone 3.
GPER-1 emerges as a new tumor suppressor with unsuspected therapeutic potential for ovarian cancer.
We are in an era where the potential exists for deriving comprehensive profiles of DNA alterations characterizing each form of human cancer. Such profiles would provide invaluable insight into mechanisms underly- ing the evolution of each tumor type and will provide molecular markers, which could radically improve cancer detection. To date, no one type of DNA change has been defined which accomplishes this purpose. Herein, by using a candidate gene approach, we show that one category of DNA alteration, aberrant methylation of gene promoter regions, can enor- mously contribute to the above goals. We have now analyzed a series of promoter hypermethylation changes in 12 genes (p16INK4a, p15INK4b, p14ARF, p73, APC,5 BRCA1, hMLH1, GSTP1, MGMT, CDH1, TIMP3, and DAPK), each rigorously characterized for association with abnormal gene silencing in cancer, in DNA from over 600 primary tumor samples rep- resenting 15 major tumor types. The genes play known important roles in processes encompassing tumor suppression, cell cycle regulation, apopto- sis, DNA repair, and metastastic potential. A unique profile of promoter hypermethylation exists for each human cancer in which some gene changes are shared and others are cancer-type specific. The hypermethy- lation of the genes occurs independently to the extent that a panel of three to four markers defines an abnormality in 70 -90% of each cancer type. Our results provide an unusual view of the pervasiveness of DNA alter- ations, in this case an epigenetic change, in human cancer and a powerful set of markers to outline the disruption of critical pathways in tumori- genesis and for derivation of sensitive molecular detection strategies for virtually every human tumor type.
GPER-1 protein expression was immunohistochemically examined in 164 primary breast cancer specimens and their matched normal breast epithelium. GPER-1 down-regulation correlated significantly with increased histological grading (p = .015), lymph node metastases (p = .032), and negative estrogen receptor status (p = .018). The decrease of GPER-1 expression in breast cancer tissue, relative to normal tissue, was associated with poor overall survival (p = .043) and disease-free survival (p = .037) and remained a significant unfavorable factor in multivariate analysis for DFS (HR = 1.569; 95% CI, 1.024-2.797; p = .041) and OS (HR = 2.082; 95% CI, 1.248-4.773; p = .039). Thus aberrant GPER-1 expression seems to be an important factor in breast cancer progression.
G-protein coupled estrogen receptor 1 (GPER) plays an important role in mediating estrogen action in many different tissues under both physiological and pathological conditions. G-1 (1-[4-(6-bromobenzo[1,3]dioxol-5yl)-3a,4,5,9b-tetrahydro-3H-cyclopenta [c]quinolin-8-yl]-ethanone) has been developed as a selective GPER agonist to distinguish estrogen actions mediated by GPER from those mediated by classic estrogen receptors. In the present study, we surprisingly found that G-1 suppressed proliferation and induced apoptosis of KGN cells (a human ovarian granulosa cell tumor cell line), actions that were not blocked by a selective GPER antagonist G15 or siRNA knockdown of GPER. G-1 also suppressed proliferation and induced cell apoptosis in GPER-negative HEK-293 cells and MDA-MB 231 breast cancer cells. Our results demonstrate that G-1 suppresses proliferation of ovarian and breast cancer cells in a GPER-independent manner. G-1 may be a candidate for the development of drugs against ovarian and breast cancer.
Learning Objectives
After completing this course, the reader will be able to:
Differentiate between the actions of oncogenes and tumor suppressor genes in the development of breast cancer. Describe the results of studies with antibodies and small molecule drugs that target growth factor receptors. Evaluate the current and potential roles of molecular and protein profiles of breast tumors in prognosis and in predicting response to therapy.
Access and take the CME test online and receive 1 hour of AMA PRA category 1 credit at CME.TheOncologist.com
Carcinogenesis is a multistep process characterized by genetic alterations that influence key cellular pathways involved in growth and development. Oncogenes refer to those genes whose alterations cause gain-of-function effects, while tumor suppressor genes cause loss-of-function effects that contribute to the malignant phenotype. The effects of these alterations are complex due to the high number of changes in a typical case of breast cancer and the interactions of the biological pathways involved. This review focuses on the more common abnormalities in oncogenes and tumor suppressor genes in human breast cancer and their known associations with clinical outcome in terms of tumor classification, prognosis, and response to specific therapies. A better understanding of these relationships has led to new therapeutic applications. Agents that target oncogenes and their associated pathways are now in clinical use, with many more undergoing preclinical and clinical testing. The availability of antibodies, small synthetic molecules, cyotokines, gene therapy techniques, and even natural compounds that are screened for specific biological properties has greatly increased the number of candidate drugs. Nevertheless, clinical successes have been limited because of the redundancy of many cancer-related pathways as well as the high degree of variability in genotype and phenotype among individual tumors. Likewise, strategies to replace tumor suppressor gene functions face numerous technical hurdles. This review summarizes the current achievements and future prospects for the therapeutic targeting of oncogenes and tumor suppressor genes and new technology to better classify tumors and accurately predict responses to standard and novel agents.
The cooperation and communication between different cell signaling transduction pathways are considered critical in the development of various types of cancer as well as drug resistance. There is evidence of crosstalk between the G protein-coupled receptor 30 (GPR30), the newly discovered estrogen receptor (ER), and the ErbB family. Heregulin (HRG)-β1, the ligand for ErbB3 and ErbB4, upregulates GPR30 expression in MCF-7, T-47D and BT-474 breast cancer cell lines that express ERα. In the present study, recombinant human HRG-β1 was used to investigate the upregulation of GPR30 expression by HRGs in MCF-7 breast cancer cells which were ERα-positive. In MCF-7 cells, the ErbB2 inhibitor, AG825, the MAPK inhibitor, PD98059, and the MEK1/2 inhibitor, U0126, blocked the HRG-β1-induced GPR30 expression. 17-β-estradiol (E2) boosted the HRG-β1-induced proliferation, migration and invasion of MCF-7 cells. Similar to E2, the specific GPR30 agonist, G-1, promoted HRG-β1-induced migration and invasion, but inhibited growth. Using the specific GPR30 antagonist, G-15, or the small interfering RNA for GPR30, the functions of GPR30 after treatment with HRG-β1 were further investegated. The results from our study indicate that the interruption between GPR30 signaling and the ErbB family system may serve as a promising therapeutic strategy for breast cancer.
G protein-coupled receptor 30/G protein estrogen receptor-1 (GPR30/GPER-1) is a novel membrane receptor for estrogen whose mRNA is expressed at high levels in estrogen-dependent cells such as breast cancer cell lines. However, mutations in GRP30 related to diseases remain unreported. To detect unknown mutations in the GPR30 open reading frame (ORF) quickly, the experimental conditions for high-resolution melting (HRM) analysis were examined for PCR primers, Taq polymerases, saturation DNA binding dyes, Mg(2+) concentration, and normalized temperatures. Nine known SNPs and 13 artificial point mutations within the GPR30 ORF, as well as single nucleotide variants in DNA extracted from subjects with breast cancers were tested under the optimal experimental conditions. The combination of Expand High Fidelity(PLUS) and SYTO9 in the presence of 2.0 mM MgCl(2) produced the best separation in melting curves of mutations in all regions of the GPR30 ORF. Under these experimental conditions, the mutations were clearly detected in both heterozygotes and homozygotes. HRM analysis of GPR30 using genomic DNA from subjects with breast cancers showed a novel single nucleotide variant, 111C>T in GPR30 and 4 known SNPs. The experimental conditions determined in this study for HRM analysis are useful for high throughput assays to detect unknown mutations within the GPR30 ORF.