Oncogene database

2078

ERG

v-ets avian erythroblastosis virus E26 oncogene homolog;ERG;v-ets avian erythroblastosis virus E26 oncogene homolog

TMPRSS2/ERG fusion|ets-related|transcriptional regulator ERG|transcriptional regulator ERG (transforming protein ERG)|v-ets avian erythroblastosis virus E26 oncogene related|v-ets erythroblastosis virus E26 oncogene homolog|v-ets erythroblastosis virus E2

21q22.3

protein-coding

The oncogenic TLS-ERG fusion protein is found in human myeloid leukemia and Ewing s sarcoma as a result of specific chromosomal translocation. To unveil the potential mechanism(s) underlying cellular transformation, we have investigated the effects of TLS-ERG on both gene transcription and RNA splicing. Here we show that the TLS protein forms complexes with RNA polymerase II (Pol II) and the serine-arginine family of splicing factors in vivo. Deletion analysis of TLS-ERG in both mouse L-G myeloid progenitor cells and NIH 3T3 fibroblasts revealed that the RNA Pol II-interacting domain of TLS-ERG resides within the first 173 amino acids. While TLS-ERG repressed expression of the luciferase reporter gene driven by glycoprotein IX promoter in L-G cells but not in NIH 3T3 cells, the fusion protein was able to affect splicing of the E1A reporter in NIH 3T3 cells but not in L-G cells. To identify potential target genes of TLS-ERG, the fusion protein and its mutants were stably expressed in both L-G and NIH 3T3 cells through retroviral transduction. Microarray analysis of RNA samples from these cells showed that TLS-ERG activates two different sets of genes sharing little similarity in the two cell lines. Taken together, these results suggest that the oncogenic TLS-ERG fusion protein transforms hematopoietic cells and fibroblasts via different pathways.

Aneuploidy is one of the hallmarks of cancer. Acquired additions of chromosome 21 are a common finding in leukemias, suggesting a contributory role to leukemogenesis. About 10% of patients with a germ line trisomy 21 (Down syndrome) are born with transient megakaryoblastic leukemia. We and others have shown acquired mutations in the X chromosome gene GATA1 in ALL these cases. The gene or genes on chromosome 21 whose overexpression promote the megakaryoblastic phenotype are presently unknown. We propose that ERG, an Ets transcription factor situated on chromosome 21, is one such candidate. We show that ERG is expressed in hematopoietic stem cells, megakaryoblastic cell lines, and in primary leukemic cells from Down syndrome patients. ERG expression is induced upon megakaryocytic differentiation of the erythroleukemia cell lines K562 and UT-7, and forced expression of ERG in K562 cells induces erythroid to megakaryoblastic phenotypic switch. We also show that ERG activates the gpIb megakaryocytic promoter and binds the gpIIb promoter in vivo. Furthermore, both ERG and ETS2 bind in vivo the hematopoietic enhancer of SCL/TAL1, a key regulator of hematopoietic stem cell and megakaryocytic development. We propose that trisomy 21 facilitates the occurrence of megakaryoblastic leukemias through a shift toward the megakaryoblastic lineage caused by the excess expression of ERG, and possibly by other chromosome 21 genes, such as RUNX1 and ETS2, in hematopoietic progenitor cells, coupled with a differentiation arrest caused by the acquisition of mutations in GATA1.

TMPRSS2-ERG gene fusion leading to the androgenic induction of the ERG proto-oncogene expression is a highly prevalent oncogenic alteration in prostate tumor cells. Prostate cancer is a multi-focal disease, and the origins as well as biological contribution of multiple cancer foci remain unclear with respect to prostate cancer onset or progression. To assess the role of TMPRSS2-ERG alteration in prostate cancer onset and/or progression, we have evaluated the status of fusion transcripts in benign glands, prostatic intraepithelial neoplasia (PIN) and multiple cancer foci of each prostate. Quantitative expression of TMPRSS2-ERG fusion type A and C transcripts was analyzed in benign, tumor and PIN areas, selected from whole-mount radical prostatectomy slides. TMPRSS2-ERG expression was correlated with clinicopathological features. Overall, 30 of 45 (67%) patients exhibited TMPRSS2-ERG fusion transcripts in at least one tumor focus. Of 80 tumor foci analyzed, 39 had TMPRSS2-ERG fusion (type A only: 30, type C only: 2, both types A and C: 7), with predominant detection of the TMPRSS2-ERG fusion type A (27/30, 90%) in the index tumors. Of 14 PIN lesions, 2 were positive for type A fusion. Frequent presence of the TMPRSS2-ERG in index tumors suggests critical roles of ERG alterations in the onset and progression of a large subset of prostate cancer. However, heterogeneity of the TMPRSS2-ERG detection in the context of multiple cancer foci and its frequency in PIN also support the role of other genomic alterations in the origins of prostate cancer.

In a study of interactions between the raf-MEK-MAPK (ERK) and JNK-jun pathways, we found previously that JNK can induce phosphorylation of raf but not vice versa. In this study, we investigate the nature of the JNK-induced phosphorylation of raf. In in vitro experiments in which immunobead-bound raf is phosphorylated by activated JNK, we find strong phosphorylation signals at raf-Ser259 and Ser338. The Ser259 phosphorylation is surprising since it is associated with inhibition of migration of raf to the cell membrane where it can interact with ras-p21. We also find that in oocytes induced to mature with oncogenic ras-p21, which induces high levels of phosphorylated JNK and MAPK, the same pattern of phosphorylation of raf occurs. In contrast, in oocytes induced to mature with insulin, which requires activation of wild-type ras-p21, phosphorylation of raf-Ser338 but not raf-Ser259 occurs. In oncogenic ras-transformed human pancreatic cancer MIA-PaCa-2 cells, phosphorylation of both raf serines occurs. Treatment of these cells with the ras peptide, PNC-2 attached to a penetrating sequence that blocks JNK and MAPK phosphorylation and induces tumor cell necrosis, results in a marked decrease in phosphorylation of raf-Ser259, but not that of raf-Ser338. These results suggest that oncogenic ras-p21 induces phosphorylation of both raf-Ser259 and Ser338 and that raf-Ser 259 phosphorylation may be effected by activated JNK.

Ets-related gene (ERG) is a member of the ETS transcription factor gene family located on Hsa21. ERG is known to have a crucial role in establishing definitive hematopoiesis and is required for normal megakaryopoiesis. Truncated forms of ERG are associated with multiple cancers such as Ewing s sarcoma, prostate cancer, and leukemia as part of oncogenic fusion translocations. Increased expression of ERG is highly indicative of poor prognosis in acute myeloid leukemia and ERG is expressed in acute megakaryoblastic leukemia (AMKL); however, it is unclear if expression of ERG per se has a leukemogenic activity. We show that ectopic expression of ERG in fetal hematopoietic progenitors promotes megakaryopoiesis and that ERG alone acts as a potent oncogene in vivo leading to rapid onset of leukemia in mice. We observe that the endogenous ERG is required for the proliferation and maintenance of AMKL cell lines. ERG also strongly cooperates with the GATA1s mutated protein, found in Down syndrome AMKL, to immortalize megakaryocyte progenitors, suggesting that the additional copy of ERG in trisomy 21 may have a role in Down syndrome AMKL. These data suggest that ERG is a hematopoietic oncogene that may play a direct role in myeloid leukemia pathogenesis.

Gene fusions prevalent in prostate cancer (CaP) lead to the elevated expression of the ERG proto-oncogene. ERG activation present in 50-70% of prostate tumors underscores one of the most common oncogenic alterations in CaP. Despite numerous reports of gene fusions and mRNA expression, ERG oncoprotein status in CaP still remains to be defined. Furthermore, development of ERG protein-based assays may provide a new dimension to evaluation of gene fusions involving diverse androgen-regulated promoters and the ERG protein-coding sequence. Through exhaustive evaluations of 132 whole-mount prostates (261 tumor foci and over 200 000 benign glands) for the ERG oncoprotein nuclear expression, we demonstrated 99.9% specificity for detecting prostate tumor cells using a highly specific anti-ERG monoclonal antibody. The ERG oncoprotein expression correlated well with fusion transcript or gene fusion in randomly selected specimens. Strong concordance of ERG-positive foci of prostatic intraepithelial neoplasia (PIN) with ERG-positive carcinoma (82 out of 85 sections with PIN, 96.5%) affirms the biological role of ERG in clonal selection of prostate tumors in 65% (86 out of 132) of patients. Conversely, ERG negative PINs were associated with ERG-negative carcinoma. Taken together, the homogeneous and strong ERG expression detected in individual tumors establishes the potential for ERG oncoprotein-based stratification of CaP.

TMPRSS2-ERG and other gene fusions involving ETS factors and genes with strong promoter elements are common in prostate cancer. Although ERG activation has been linked to invasive properties of prostate cancers, the precise mechanisms and pathways of ERG-mediated oncogenesis remain poorly understood. Here, we show that ERG knockdown in VCaP prostate cancer cells causes an activation of cell adhesion, resulting in strongly induced active beta(1)-integrin and E-cadherin expression as well as changes in WNT signaling. These observations were corroborated by data from ERG-overexpressing nontransformed prostate epithelial cells as well as gene expression data from clinical prostate cancer samples, which both indicated a link between ERG and epithelial-to-mesenchymal transition (EMT). Upregulation of several WNT pathway members was seen in ERG-positive prostate cancers, with frizzled-4 (FZD4) showing the strongest overexpression as verified by both reverse transcription-PCR and immunostaining. Both ERG knockin and knockdown modulated the levels of FZD4 expression. FZD4 silencing could mimic the ERG knockdown phenotype by inducing active beta(1)-integrin and E-cadherin expression, whereas FZD4 overexpression reversed the phenotypic effects seen with ERG knockdown. Taken together, our results provide mechanistic insights to ERG oncogenesis in prostate cancer, involving activation of WNT signaling through FZD4, leading to cancer-promoting phenotypic effects, including EMT and loss of cell adhesion.

Androgen dependent induction of the ETS related gene (ERG) expression in more than half of ALL prostate cancers results from gene fusions involving regulatory sequence of androgen regulated genes (i.e. TMPRSS2, SLC45A3 and NDRG1) and protein coding sequence of the ERG. Emerging studies in experimental models underscore the functions of ERG in prostate tumorigenesis. However, biological and biochemical functions of ERG in prostate cancer (CaP) remain to be elucidated. This study suggests that ERG activation plays a role in prostaglandin signaling because knockdown of ERG expression in TMPRSS2-ERG fusion containing CaP cells leads to altered levels of the 15-hydroxy-prostaglandin dehydrogenase (HPGD), a tumor suppressor and prostaglandin catabolizing enzyme, and prostaglandin E2 (PGE2) . We demonstrate that HPGD expression is regulated by the binding of the ERG protein to the core promoter of this gene. Moreover, prostaglandin E2 dependent cell growth and urokinase-type plasminogen activator (uPA) expression are also affected by ERG knockdown. Together, these data imply that the ERG oncoprotein in CaP cells positively influence prostaglandin mediated signaling, which may contribute to tumor progression.

The development of prostate cancer and its progression to castrate-resistant prostate cancer (CRPC) after antiandrogen ablation therapy are driven by persistent biological activity of androgen receptor (AR) signaling. Moreover, studies have shown that more than 50% of human prostate cancers overexpress ERG (v-ets avian erythroblastosis virus E26 oncogene related gene) due to AR-regulated TMPRSS2-ERG fusion gene. However, the reported roles of TMPRSS2-ERG fusion in cancer progression are not clear. In this study, we investigated the signal transduction in the AR/TMPRSS2-ERG/Wnt signaling network for studying the aggressive behavior of prostate cancer cells and further assessed the effects of BR-DIM and CDF [natural agents-derived synthetic formulation and analogue of 3,3 -diindolylmethane (DIM) and curcumin, respectively, with improved bioavailability] on the regulation of AR/TMPRSS2-ERG/Wnt signaling. We found that activation of AR resulted in the induction of ERG expression through TMPRSS2-ERG fusion. Moreover, we found that ERG overexpression and nuclear translocation activated the activity of Wnt signaling. Furthermore, forced overexpression of ERG promoted invasive capacity of prostate cancer cells. More important, we found that BR-DIM and CDF inhibited the signal transduction in the AR/TMPRSS2-ERG/Wnt signaling network, leading to the inactivation of Wnt signaling consistent with inhibition of prostate cancer cell invasion. In addition, BR-DIM and CDF inhibited proliferation of prostate cancer cells and induced apoptotic cell death. On the basis of our findings, we conclude that because BR-DIM and CDF downregulate multiple signaling pathways including AR/TMPRSS2-ERG/Wnt signaling, these agents could be useful for designing novel strategies for the prevention and/or treatment of prostate cancer.

PIK3CA gain-of-function mutations are a common oncogenic event in human malignancy, making phosphatidylinositol 3-kinase (PI3K) a target for cancer therapy. Despite the promise of targeted therapy, resistance often develops, leading to treatment failure. To elucidate mechanisms of resistance to PI3K-targeted therapy, we constructed a mouse model of breast cancer conditionally expressing human PIK3CA(H1047R). Notably, most PIK3CA(H1047R)-driven mammary tumors recurred after PIK3CA(H1047R) inactivation. Genomic analyses of recurrent tumors revealed multiple lesions, including focal amplification of Met or Myc (also known as c-Met and c-Myc, respectively). Whereas Met amplification led to tumor survival dependent on activation of endogenous PI3K, tumors with Myc amplification became independent of the PI3K pathway. Functional analyses showed that Myc contributed to oncogene independence and resistance to PI3K inhibition. Notably, PIK3CA mutations and c-MYC elevation co-occur in a substantial fraction of human breast tumors. Together, these data suggest that c-MYC elevation represents a potential mechanism by which tumors develop resistance to current PI3K-targeted therapies.

TMPRSS2:ERG is a gene fusion resulting from the chromosomal rearrangement of the androgen-regulated TMPRSS2 gene and the ETS transcription factor ERG, leading to the over-expression of the oncogenic molecule ERG. This gene rearrangement has been found in approximately half of ALL prostate cancers and ERG overexpression is considered as a novel diagnostic marker for prostate carcinoma. However, little is known about the role of the TMPRSS2:ERG gene fusion in ovarian cancer. The purpose of this study was to test ERG expression in ovarian cancer and its potential as a diagnostic marker for ovarian carcinoma progression. A tissue microarray containing 180 ovarian cancer tissues of various pathological types and grades were examined by immunohistochemical analysis for expression of ERG. We also used 40 prostate carcinoma tissues and 40 normal tissues for comparison in parallel experiments. ERG-positive expression was detected in 40% of the prostate tumor cancer, as well as in internal positive control endothelial cells, confirming over-expression of ERG in prostate cancer at relatively the same rate observed by others. In contrast, ALL of the ovarian tumor patient tissues of varying histologic types were ERG-negative, despite some positivity in endothelial cells. These results suggest that the oncogenic gene fusion TMPRSS2:ERG does not occur in ovarian cancer relative to prostate cancer. Therefore, development of ERG expression profile would not be a useful diagnostic or prognostic marker for ovarian cancer patient screening.

Oncogenic activation of ERG resulting from gene fusion is present in over half of ALL patients with prostate cancer in Western countries. Although the underlying genetic mechanisms have been extensively studied, evaluation of the ERG oncoprotein--the translational product of ERG gene fusions--has just begun. The robust correlation between ERG oncoprotein detection and gene fusion status enables rapid characterization of this protein in large patient cohorts. Recent studies have focused on characterizing the ERG oncoprotein and determining its potential role in the diagnosis and biological stratification of prostate cancer.

Emerging evidence suggests that chromatin adopts a nonrandom 3D topology and that the organization of genes into structural hubs and domains affects their transcriptional status. How chromatin conformation changes in diseases such as cancer is poorly understood. Moreover, how oncogenic transcription factors, which bind to thousands of sites across the genome, influence gene regulation by globally altering the topology of chromatin requires further investigation. To address these questions, we performed unbiased high-resolution mapping of intra- and interchromosome interactions upon overexpression of ERG, an oncogenic transcription factor frequently overexpressed in prostate cancer as a result of a gene fusion. By integrating data from genome-wide chromosome conformation capture (Hi-C), ERG binding, and gene expression, we demonstrate that oncogenic transcription factor overexpression is associated with global, reproducible, and functionally coherent changes in chromatin organization. The results presented here have broader implications, as genomic alterations in other cancer types frequently give rise to aberrant transcription factor expression, e.g., EWS-FLI1, c-Myc, n-Myc, and PML-RARalpha.

The three-dimensional (3D) conformation of the genome is known to be structured and to affect gene transcription, but how chromatin conformation changes in diseases such as cancer is poorly understood. Similarly, oncogenic transcription factors bind to thousands of sites in the genome without a clear transcriptional role on nearby genes. Could these factors play a non-transcriptional role in promoting tumor progression by restructuring the shape of the genome? To address this question, we recently performed unbiased high-resolution mapping of intra- and inter-chromosome interactions upon overexpression of ERG, an oncogenic transcription factor frequently overexpressed in prostate cancer as a result of a gene fusion. By integrating data from genome-wide chromosome conformation capture (Hi-C), ERG binding and gene expression, we have demonstrated that oncogenic transcription factor overexpression is associated with global, reproducible and functionally coherent changes in chromatin organization. Perhaps more importantly, we have identified novel genomic alterations associated with ERG overexpression. These results suggest a yet unappreciated role for transcription factors in promoting genomic alterations through their effect on chromatin architecture.

OBJECTIVE: To systematically evaluate the ETS-related gene (ERG) alterations in the multifocal tumor context using whole-mount prostatectomy specimens from African and Caucasian American patients matched for age, pathologic grade and stage. Oncogenic activation of the ERG is the most common early genomic alteration in patients with prostate cancer (CaP) in Western countries. However, ERG alterations have not been systematically examined in African American patients with a known greater risk of CaP incidence and mortality. METHODS: ERG oncoprotein expression was analyzed in 91 Caucasian and 91 African American patients with CaP, who were matched for age, Gleason score, and pathologic stage. A unique aspect of the present study was the evaluation of ERG in whole-mount prostatectomy sections, minimizing sampling bias and allowing the careful assessment of the ERG in the multifocal tumor context of CaP. RESULTS: The frequency of ERG-positive prostate tumors was significantly greater among Caucasian Americans than among African Americans when assessed in ALL tumor foci (41.9% vs 23.9%, P < .0001). A markedly greater frequency of ERG oncoprotein expression was noted between the index tumors of the Caucasian Americans (63.3%) and those of the African Americans (28.6%). Also, in the African American patients, the higher grade index tumors were predominantly ERG negative. CONCLUSION: ERG typing of CaP established a major difference between the index tumors of Caucasian and African American patients. ERG-negative index tumors might indicate a less favorable outcome for African American patients. The results of the present study underscore that typing of CaP for the ERG could enhance our understanding of the biologic differences between the examined ethnic groups.

To investigate and characterize ERG oncoprotein expression in Japanese patients with prostate cancer (PCa), we evaluated 92 index tumors from Japanese patients who had undergone radical prostatectomy for PCa, using an antibody-based detection method to determine ERG expression. expression status was compared with clinicopathological findings including patient age, body mass index and preoperative prostate-specific antigen (PSA) levels, specimen Gleason score, pathological stage, positive surgical margin, size of index tumor, prostatic volume, zonal origin of index tumor and biochemical failure. Overall, ERG protein was expressed in 16.3% (15/92) of the index tumors, but not in benign glands. Younger patient age, lower Gleason score and negative surgical margins were found to be independently associated with its expression in the multivariate analysis (P= 0.015, 0.003 and 0.038, respectively). ERG expression was not associated with biochemical failure. Though not statistically significant, ERG expression was more frequently observed in peripheral zone than in transition zone cancers (28.2% vs 10%, respectively). In conclusion, ERG protein was less frequently expressed in this Japanese PCa cohort compared with Western reports. ERG expression was associated with a less aggressive tumor phenotype, and its biological significance needs to be further investigated.

BACKGROUND: ETS-related gene (ERG) protein is present in 40-70% of prostate cancer and is correlated with TMPRSS2-ERG gene rearrangements. This study evaluated ERG expression at radical prostatectomy to determine whether it was predictive of earlier relapse or prostate cancer-specific mortality (PCSM). METHODS: One hundred patients who underwent radical prostatectomy at Virginia Mason in Seattle between 1991 and 1997 were identified. Recurrence was confirmed by tissue diagnosis or radiographic signs. PCSM was confirmed by death certificates. Thirty-three patients with metastases or PCSM were matched to patients without recurrence at a 1:2 ratio. Paraffin embedded tissue was stained with two anti-ERG monoclonal antibodies, EPR3864 and 9FY. Nuclear expression intensity was evaluated as present/absent, on a 4-point relative intensity scale, and as a composite score (0-300). RESULTS: Mean follow-up was 10.26 years. The two antibodies were highly correlated (P < 0.0001). Patients with higher ERG expression intensity and composite scores were significantly more likely to develop biochemical relapse, metastases, and PCSM. Kaplan-Meier survival curve analysis for the composite score of ERG expression revealed a significant association between higher ERG expression (EPR3864) and shorter PCa-specific survival (P = 0.047). CONCLUSIONS: While the presence of ERG expression at the time of surgery was not predictive of earlier relapse or PCSM, the relative intensity and composite score for ERG expression was prognostic for the development of biochemical relapse, metastases, and PCSM. Quantitative ERG scoring may be useful to identify patients who would benefit from adjuvant treatment or closer follow-up, allowing more accurate individual patient treatment plans.

Prostate cancer is a leading cause of cancer mortality in men. One of the distinct characteristics of prostate cancer is over-expression of the ERG proto-oncogene. The TMPRSS2-ERG gene fusion, the most common gene fusion, is found in approximately 50% of prostate cancer cases. We show that certain microRNAs are extensively deregulated in prostate cancer cell lines and primary clinical cancer samples. MicroRNAs are capable of modulating post-transcriptional gene expression via inhibition of protein synthesis. Independent target prediction methods have indicated that the 3 untranslated region of the ERG mRNA is a potential target of miR-145. miR-145 is consistently down-regulated in prostate cancer. Here we show that the ERG 3 untranslated region is a regulative target of miR-145 in vitro. Ectopic expression of miR-145 led to a reduction in expression of the ERG protein. We analyzed 26 prostate cancer samples and corresponding normal tissue. ERG protein expression was found to be elevated in the tumor samples, together with increased expression of several ERG isoforms. We identified ERG proteins of 35 and 24 kDa, which may represent unknown ERG splice variants. Analyses of miR-145 and ERG mRNA expression revealed a general down-regulation of miR-145 irrespective of the presence or absence of translocations involving ERG. This observation indicates that down-regulation of miR-145 may contribute to the increased expression of most ERG splice variants sharing the miR-145 target sequence in their 3 untranslated region.

PURPOSE: To assess differentially methylated "landscapes" according to prostate cancer Gleason score (GS) and ERG oncogene expression status, and to determine the extent of polycomb group (PcG) target gene involvement, we sought to assess the genome-wide DNA methylation profile of prostate cancer according to Gleason score and ERG expression. EXPERIMENTAL DESIGN: Genomic DNA from 39 prostate cancer specimens was hybridized to CpG island microarrays through differential methylation hybridization. We compared methylation profiles between Gleason score and ERG expression status as well as Gleason score stratified by ERG expression status. In addition, we compared results from our dataset to publicly available datasets of histone modifications in benign prostate cells. RESULTS: We discovered hundreds of distinct differentially methylated regions (DMR) associated with increasing Gleason score and ERG. Furthermore, the number of DMRs associated with Gleason score was greatly expanded by stratifying samples into ERG-positive versus ERG-negative, with ERG-positive/GS-associated DMRs being primarily hypermethylated as opposed to hypomethylated. Finally, we found that there was a significant overlap between either Gleason score-related or ERG-hypermethylated DMRs and distinct regions in benign epithelial cells that have PcG signatures (H3K27me3, SUZ12) and lack active gene expression signatures (H3K4me3, RNA pol II). CONCLUSIONS: This work defines methylation landscapes of prostate cancer according to Gleason score, and suggests that initiating genetic events may influence the prostate cancer epigenome, which is further perturbed as prostate cancer progresses. Moreover, CpG islands with silent chromatin signatures in benign cells are particularly susceptible to prostate cancer-related hypermethylation.

Trefoil factor 3 (TFF3) is associated with various cancers and overexpressed in a subset of prostate cancers. Functional studies suggest that v-ets erythroblastosis virus E26 oncogene homolog (avian) (ERG) down-regulates TFF3 expression in hormone-naive prostate cancer. To characterize this inverse relationship, we developed a triple immunostain encompassing ERG, TFF3, and high-molecular-weight cytokeratin. Triple stain was performed on 96 tumors and 52 benign cases represented in tissue microarrays. Distinct ERG and TFF3 protein was expressed in 45% (43/96) and 36% (35/96) of prostate cancers, respectively. Coexpression was observed in 5% (5/96) of tumor cases, and 24% (23/96) did not express ERG or TFF3. The inverse expression of ERG and TFF3 was significant (P < .0001), with 57% (30/53) of ERG-negative tumors demonstrating TFF3 expression. Sensitivity and specificity of combined ERG and TFF3 expression in detecting prostate cancer were 76% and 96%, respectively. The feasibility of triple immunostain protocol was validated in a set of 76 needle biopsies. The application of this multiplex in situ biomarker for molecular characterization of prostate cancer and as a supplemental diagnostic and prognostic tool in prostate needle biopsies should be further explored.

INTRODUCTION AND OBJECTIVE: ERG oncogene fusions (predominantly TMPRSS2-ERG) represent the most common (50-70% frequency) and validated prostate cancer (CaP) genome alteration in the Western countries. A common TMPRSS2-ERG fusion type leads to the androgen dependent tumor cell specific expression of the TMPRSS2-ERG fusion transcript and amino terminally truncated ERG oncoprotein. CaP prevalence and aggressiveness, as well as genomic alterations vary in different geographic locations in the world. Recent studies from our group highlighted significantly lower frequency of ERG alterations in prostate index tumors of African American men (~30%) in comparison to Caucasian Americans (~60%). Further, much lower frequencies (10 -25%) of ERG alterations have been reported in studies from China and Japan. There is no study on ERG alterations in CaP patients from India, representing a significant portion of the world male population. This study focuses on the frequency of ERG oncoprotein expression in CaP patients from India. METHODS: De-identified formalin-fixed paraffin-embedded (FFPE) specimens from radical prostatectomy (RP) specimens of 51 patients from the Rajiv Gandhi cancer Institute and Research Centre (RGCI), New Delhi, India, were analyzed for ERG alterations. The ERG oncoprotein expression as a surrogate of ERG gene fusions was analyzed by using a highly specific ERG monoclonal antibody (9FY). TMPRSS2-ERG fusion was assessed by fluorescent in situ hybridization (FISH) assays using the break-apart ERG probes. RESULTS: Specimens reflecting prior hormonal treatment, or lacking any tumor content, were excluded from the analyses. Of the thirty evaluable specimens, ERG positive tumors were present in 8 cases (27%) and one tumor specimen exhibited rare ERG positive cells. None of the benign glands were positive for ERG supporting previous studies showing complete specificity of the ERG oncoprotein for detection of tumors cells in prostate. CONCLUSIONS: Frequency of ERG oncoprotein expression is much lower in CaP patients from India in comparison to higher frequency of ERG alterations noted in Western countries. ERG frequency in Indian CaP is similar to observations from Japan and China. Since ERG oncogenic activation is a promising biomarker and therapeutic target for CaP, careful evaluation of ERG is needed in CaP patients from different parts of the world.

The transcription factor E-twenty-six related gene (ERG), which is overexpressed through gene fusion with the androgen-responsive gene transmembrane protease, serine 2 (TMPRSS2) in approximately 40% of prostate tumors, is a key driver of prostate carcinogenesis. Ablation of ERG would disrupt a key oncogenic transcriptional circuit and could be a promising therapeutic strategy for prostate cancer treatment. Here, we show that ubiquitin-specific peptidase 9, X-linked (USP9X), a deubiquitinase enzyme, binds ERG in VCaP prostate cancer cells expressing TMPRSS2-ERG and deubiquitinates ERG in vitro. USP9X knockdown resulted in increased levels of ubiquitinated ERG and was coupled with depletion of ERG. Treatment with the USP9X inhibitor WP1130 resulted in ERG degradation both in vivo and in vitro, impaired the expression of genes enriched in ERG and prostate cancer relevant gene signatures in microarray analyses, and inhibited growth of ERG-positive tumors in three mouse xenograft models. Thus, we identified USP9X as a potential therapeutic target in prostate cancer cells and established WP1130 as a lead compound for the development of ERG-depleting drugs.

BACKGROUND: Filipinos with prostate cancer (CaP) are at increased risk of harboring advanced stages and lower survival rates compared to other Asians. This study aims to investigate prevalence of ETS-related gene (ERG) oncoprotein overexpression in Filipinos as surrogate of TMPRSS2-ERG gene fusions, using a highly specific monoclonal antibody (ERG-MAb), and conduct the first attempt to study the role of genetic alterations in the aggressive tumor biologic behaviour of CaP among Filipinos. METHODS: This case-matched, case-control retrospective study evaluated ERG expression in Filipino patients diagnosed with CaP and its effect on stage and Gleason grade of their disease. Men who underwent radical prostatectomy for organ-confined disease at the University of the Philippines-Philippine General Hospital (UP-PGH) comprised the organ-confined cohort. Age-matched adults who had trans-rectal ultrasound-guided prostate (TRUSP) biopsy or trans-urethral resection of the prostate (TURP) with bilateral orchiectomy for T4 or stage IV CaP composed the advanced disease cohort. RESULTS: Overall ERG expression frequency of 23.08% (N = 104) was demonstrated, with a higher rate observed in the advanced disease cohort (32.69%) compared to the organ-confined group (13.46%). Furthermore, ERG overexpression was only detected among intermediate and high-risk tumors. A high-specificity (98.08%) of the ERG-MAb for malignant prostatic cells was likewise demonstrated. CONCLUSIONS: In contrast to higher ERG frequency in Western countries, it is much lower in Filipino CaP, which is similar to lower rates noted from other Asian countries. The 98.08% specificity of ERG oncoprotein for prostate tumor cells combined with its increased association in advanced disease, suggests for prognostic potential of ERG that may aid clinicians in treatment decisions for Filipino CaP patients.

Overexpression of ERG in the prostate epithelium, due to chromosomal translocations, contributes to prostate tumorigenesis. Here, genomic analysis of ERG siRNA-treated prostate cells harboring the endogenous TMPRSS2-ERG fusion revealed an inverse relationship between ERG and Annexin A2 (ANXA2) expression at both the RNA and protein level. ANXA2, a Ca(2+)-dependent and phospholipid-binding protein, is involved in various cellular functions, including maintenance of epithelial cell polarity. Mechanistic studies defined the prostate-specific transcription start site of ANXA2 and showed that the recruitment of ERG to the ANXA2 promoter is required for transcriptional repression by ERG. Knockdown of ERG enhanced the apical localization of ANXA2, the bundling of actin filaments at cell-cell junctions and formation of a polarized epithelial phenotype. ERG overexpression disrupted ANXA2-mediated cell polarity and promoted epithelial-mesenchymal transition (EMT) by inhibiting CDC42 and RHOA, and by activating cofilin. Immunohistochemistry demonstrated a reciprocal relationship of ANXA2 and ERG expression in a large fraction of primary prostate cancer clinical specimens. ANXA2 was absent or markedly reduced in ERG(+) tumors, which were mostly well differentiated. ERG(-) tumors, meanwhile, expressed moderate to high levels of ANXA2, and were either poorly differentiated or displayed subsets of poorly differentiated cells. Taken together, the transcriptional repression of ANXA2 by ERG in prostate epithelial cells plays a critical role in abrogating differentiation, promoting EMT, and in the reciprocal correlation of ERG and ANXA2 expression observed in human prostate cancer. IMPLICATIONS: ANXA2 is a new component of the ERG network with potential to enhance biologic stratification and therapeutic targeting of ERG-stratified prostate cancers.

Overexpression of the erythroblast transformation-specific-related gene (ERG) oncoprotein due to transmembrane protease, serine 2 (TMPRSS2)-ERG fusion, the most prevalent genomic alteration in prostate cancer (CaP), is more frequently observed among Caucasian patients compared to patients of African or Asian descent. To the best of our knowledge, this is the first study to investigate the prevalence of ERG alterations in a multiethnic cohort of CaP patients. A total of 191 formalin-fixed paraffin-embedded sections of transrectal ultrasound-guided prostate biopsy specimens, collected from 120 patients treated at the Sime Darby Medical Centre, Subang Jaya, Malaysia, were analyzed for ERG protein expression by immunohistochemistry using the anti-ERG monoclonal antibody 9FY as a surrogate for the detection of ERG fusion events. The overall frequency of ERG protein expression in the population evaluated in this study was 39.2%. Although seemingly similar to rates reported in other Asian communities, the expression of ERG was distinct amongst different ethnic groups (P=0.004). Malaysian Indian (MI) patients exhibited exceedingly high expression of ERG in their tumors, almost doubling that of Malaysian Chinese (MC) patients, whereas ERG expression was very low amongst Malay patients (12.5%). When collectively analyzing data, we observed a significant correlation between younger patients and higher ERG expression (P=0.04). The prevalence of ERG expression was significantly different amongst CaP patients of different ethnicities. The higher number of ERG-expressing tumors among MI patients suggested that the TMPRSS2-ERG fusion may be particularly important in the pathogenesis of CaP amongst this group of patients. Furthermore, the more frequent expression of ERG among the younger patients analyzed suggested an involvement of ERG in the early onset of CaP. The results of this study underline the value of using ERG status to better understand the differences in the etiology of CaP initiation and progression between ethnic groups.

ETS-related gene (ERG) is a member of the E-26 transformation-specific (ETS) family of transcription factors with roles in development that include vasculogenesis, angiogenesis, haematopoiesis and bone development. ERG s oncogenic potential is well known because of its involvement in Ewing s sarcoma and leukaemia. However, in the past decade ERG has become highly associated with prostate cancer development, particularly as a result of a gene fusion with the promoter region of the androgen-induced TMPRRSS2 gene. We review ERG s structure and function, and its role in prostate cancer. We discuss potential new therapies that are based on targeting ERG.

The ERG gene is fused to TMPRSS2 in approximately 50% of prostate cancers (PrCa), resulting in its overexpression. However, whether this is the sole mechanism underlying ERG elevation in PrCa is currently unclear. Here we report that ERG ubiquitination and degradation are governed by the Cullin 3-based ubiquitin ligase SPOP and that deficiency in this pathway leads to aberrant elevation of the ERG oncoprotein. Specifically, we find that truncated ERG (DeltaERG), encoded by the ERG fusion gene, is stabilized by evading SPOP-mediated destruction, whereas prostate cancer-associated SPOP mutants are also deficient in promoting ERG ubiquitination. Furthermore, we show that the SPOP/ERG interaction is modulated by CKI-mediated phosphorylation. Importantly, we demonstrate that DNA damage drugs, topoisomerase inhibitors, can trigger CKI activation to restore the SPOP/DeltaERG interaction and its consequent degradation. Therefore, SPOP functions as a tumor suppressor to negatively regulate the stability of the ERG oncoprotein in prostate cancer.

We have isolated a cDNA clone representing the complete coding sequence of a human gene named erg, related to the ets oncogene. Nucleotide sequence analysis of this cDNA (4.6 kilobases long) revealed that this gene encodes a 363-residue protein whose predicted amino acid sequence showed a homology of approximately equal to 40% and approximately equal to 70% to two domains corresponding to the 5 and 3 regions of v-ets oncogene, respectively. A 3.2- to 3.6-kilobase and approximately equal to 5-kilobase transcript of the erg gene, which differ in size from those of the previously described Hu-ets 1 and Hu-ets 2 genes, were observed in different cells. These results suggest that the erg gene is a member of the ets oncogene family.