ERG-positive Tumors Research Articles

Abstract 1183: Structure-activity studies and biological evaluations of ERGi-USU, a highly selective inhibitor for ERG-positive prostate cancer cells

Abstract Introduction and objectives: While new prostate cancer (CaP) treatments (Abiraterone and Enzalutamide) have improved survival in castration-resistant prostate cancer (CRPC), their benefits are short-lived and drug resistance develops likely due to numerous adaptive mutations. Therefore it is a critical to develop effective novel inhibitors to target other signaling pathways that promote or contribute to prostate tumorigenesis. Accumulating evidence has established the androgen regulated TMPRSS2-ERG fusion as a common oncogenic driver that contributes to the early development and progression of over half of CaP. Therefore, ERG oncoprotein and ERG dependent pathways are promising targets for CaP therapy in early stages when cancer is most responsive to treatment. We previously identified a small molecule inhibitor, ERGi-USU, which selectively inhibits ERG protein and cell growth in ERG positive tumor cell lines and mouse xenograft models. In an effort to further develop ERGi-USU with enhanced efficacy we performed detailed structure-activity relationship (SAR) evaluation of ERGi-USU core structure and developed new derivatives. Methods: Based on SAR of the core structure of ERGi-USU, 48 new derivatives were designed and synthesized by substitutions with alkyl, alkoxy, cycloalkyl, heterocycloalkyl, aryl, heteroaryl or hydroxyl groups. The new ERGi-USU derivatives were evaluated for inhibition of cell growth and ERG protein levels in the TMPRSS2-ERG fusion harboring CaP cell line, VCaP. Four of these compounds have been selected for evaluation of ERG selectivity by defining IC50 in ERG positive malignant cells (VCaP, KG1, MOLT-4 and COLO320), ERG negative CaP cell line (LNCaP) or ERG positive normal primary endothelium-derived cells (HUVEC). Result: Like parental compound, four new ERGi-USU derivatives exhibited inhibition of cell growth and ERG protein levels in ERG positive VCaP, KG1, MOLT-4 and COLO320 cell lines, with no or minimal effects on LNCaP and HUVEC cells. One of the new derivatives (ERGi-USU#6) showed increased efficacy for cell growth inhibition (IC50=0.074µM) compared to the parental ERGi-USU (IC50=0.200µM). Other three new compounds showed similar IC50 as the ERGi-USU. Conclusion: Comprehensive evaluation of ERGi-USU derivatives along with parental compound has continued to underscore selective inhibition of ERG positive tumor cells by these small molecules. Citation Format: Ahmed A. Mohamed, Charles P. Xavier, Gauthaman Sukumar, Samuel D. Banister, Vineet Kumar, Shyh-Han Tan, Shilpa Katta, Lakshmi Ravindranath, Muhammad Jamal, Taduru Sreenath, David G. McLeod, Gyorgy Petrovics, Albert Dobi, Meera Srivastava, Sanjay Malhotra, Clifton Dalgard, Shiv Srivastava. Structure-activity studies and biological evaluations of ERGi-USU, a highly selective inhibitor for ERG-positive prostate cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1183. doi:10.1158/1538-7445.AM2017-1183

MP87-14 STRUCTURE-ACTIVITY RELATIONSHIP STUDIES OF ERGI-USU, A HIGHLY SELECTIVE INHIBITOR FOR ERG POSITIVE PROSTATE CANCER CELLS

You have accessJournal of UrologyProstate Cancer: Basic Research & Pathophysiology III1 Apr 2017MP87-14 STRUCTURE-ACTIVITY RELATIONSHIP STUDIES OF ERGI-USU, A HIGHLY SELECTIVE INHIBITOR FOR ERG POSITIVE PROSTATE CANCER CELLS Ahmed Mohamed, Charles Xavier, Gauthaman Sukumar, Samuel Banister, Vineet Kumar, Shyh-Han Tan, Shilpa Katta, Lakshmi Ravindranath, Muhammad Jamal, Taduru Sreenath, David McLeod, Gyorgy Petrovics, Albert Dobi, Meera Srivastava, Sanjay Malhotra, Clifton Dalgard, and Shiv Srivastava Ahmed MohamedAhmed Mohamed More articles by this author , Charles XavierCharles Xavier More articles by this author , Gauthaman SukumarGauthaman Sukumar More articles by this author , Samuel BanisterSamuel Banister More articles by this author , Vineet KumarVineet Kumar More articles by this author , Shyh-Han TanShyh-Han Tan More articles by this author , Shilpa KattaShilpa Katta More articles by this author , Lakshmi RavindranathLakshmi Ravindranath More articles by this author , Muhammad JamalMuhammad Jamal More articles by this author , Taduru SreenathTaduru Sreenath More articles by this author , David McLeodDavid McLeod More articles by this author , Gyorgy PetrovicsGyorgy Petrovics More articles by this author , Albert DobiAlbert Dobi More articles by this author , Meera SrivastavaMeera Srivastava More articles by this author , Sanjay MalhotraSanjay Malhotra More articles by this author , Clifton DalgardClifton Dalgard More articles by this author , and Shiv SrivastavaShiv Srivastava More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2017.02.2718AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES While new prostate cancer (CaP) treatments (Abiraterone and Enzalutamide) have improved survival in castration resistant prostate cancer (CRPC), their benefits are short-lived and drug resistance develops likely due to numerous adaptive mutations. Accumulating evidence has established the androgen regulated TMPRSS2-ERG fusion as a common oncogenic driver that contributes to the early development and progression of over half of CaP. Therefore, ERG oncoprotein and ERG dependent pathways are promising targets for CaP therapy in early stages when cancer is most responsive to treatment. We previously identified a small molecule inhibitor, ERGi-USU, which selectively inhibits ERG protein and cell growth in ERG positive tumor cell lines and mouse xenograft models. In an effort to further develop ERGi-USU with enhanced efficacy we performed detailed structure-activity relationship (SAR) evaluation of ERGi-USU core structure and developed new derivatives. METHODS Based on SAR of the core structure of ERGi-USU, 48 new derivatives were designed and synthesized by substitutions with alkyl, alkoxy, cycloalkyl, heterocycloalkyl, aryl, heteroaryl or hydroxyl groups. The new ERGi-USU derivatives were evaluated for inhibition of cell growth and ERG protein levels in the TMPRSS2-ERG fusion harboring CaP cell line, VCaP. Four of these compounds have been selected for evaluation of ERG selectivity by defining IC50 in ERG positive malignant cells (VCaP, KG1, MOLT-4 and COLO320), ERG negative CaP cell line (LNCaP) or ERG positive normal primary endothelium-derived cells (HUVEC). RESULTS Like parental compound, four new ERGi-USU derivatives exhibited inhibition of cell growth and ERG protein levels in ERG positive VCaP, KG1, MOLT-4 and COLO320 cell lines, with no or minimal effects on LNCaP and HUVEC cells. One of the new derivatives (ERGi-USU#6) showed increased efficacy for cell growth inhibition (IC50=0.074μM) compared to the parental ERGi-USU (IC50=0.200μM). Other three new compounds showed similar IC50 as the ERGi-USU. CONCLUSIONS Comprehensive evaluation of ERGi-USU derivatives along with parental compound has continued to underscore selective inhibition of ERG positive tumor cells by these small molecules. © 2017FiguresReferencesRelatedDetails Volume 197Issue 4SApril 2017Page: e1171 Advertisement Copyright & Permissions© 2017MetricsAuthor Information Ahmed Mohamed More articles by this author Charles Xavier More articles by this author Gauthaman Sukumar More articles by this author Samuel Banister More articles by this author Vineet Kumar More articles by this author Shyh-Han Tan More articles by this author Shilpa Katta More articles by this author Lakshmi Ravindranath More articles by this author Muhammad Jamal More articles by this author Taduru Sreenath More articles by this author David McLeod More articles by this author Gyorgy Petrovics More articles by this author Albert Dobi More articles by this author Meera Srivastava More articles by this author Sanjay Malhotra More articles by this author Clifton Dalgard More articles by this author Shiv Srivastava More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Abstract 4504: Developing a PTEN-ETS signature to improve molecular risk stratification in prostate cancer

Abstract Prostate cancer (PCa) is a heterogeneous disease and the development of a molecular classification is critical to distinguish lethal from indolent tumors and reduce overtreatment. Genomic alterations of the PTEN and ETS genes (e.g., ERG) are among the most common in PCa and there is a huge interest in exploiting these alterations for risk assessment. We have found that PTEN loss is associated with PCa death most strongly in patients whose tumors do not carry an ERG rearrangement, contradicting findings from mouse models. Despite technological advances have enabled extraordinary insight into PCa molecular alterations, our understanding of the interaction between PTEN and ETS genes during PCa progression remains very limited. Hence, we have analyzed a large collection of multi-omics datasets to characterize the molecular underpinnings of PTEN loss in the context of ETS-negative versus ETS-positive tumors and further investigate their role in driving PCa prognosis. First we have developed a classification approach for PCa stratification based on dichotomized gene expression of PTEN and ETS family members. We have then validated this approach using TCGA DNA copy number variation and rearrangement status information. Our results confirmed that transcriptome analysis can be used for detecting PTEN and ETS alterations, and that in the absence of ERG rearrangements, tumors with PTEN loss frequently overexpress alternative ETS genes. Next, we examined whether somatic mutation profiles might differ by PTEN/ETS status, providing an potential explanation for the lethal behavior of ERG-negative/PTEN-negative tumors. As expected SPOP mutation was found to be mutually exclusive with ERG expression, but it did not vary significantly by PTEN status. Most interestingly, TP53 mutation - the most frequently altered gene in lethal metastatic PCa - was significantly enriched among PTEN-negative tumors irrespective to ERG- status. Collectively these findings suggest that known prognosis-driving somatic mutations do not explain the aggressive behavior of ERG-negative/PTEN-negative tumors. Finally, we analyzed differential gene expression associated with PTEN loss in the presence or absence of ERG gene expression and identified the associated biological processes and signaling pathways. This analysis failed to confirm the correlation between ERG/ETV1 expression and higher androgen signaling output previously reported in murine models. Furthermore, our findings revealed that MYC targets and the genes involved in cell cycle progression are the most up-regulated by PTEN loss in ETS-negative tumors, while genes involved in cell differentiation, response to DNA damage, and TNF signaling are the most induced by PTEN loss in ERG-positive tumors. Overall, our analysis clarified some important aspects of the role of PTEN/ETS network in PCa progression, suggesting a number of promising targets for the development of novel therapeutic interventions. Citation Format: Luigi Marchionni, Tamara L. Lotan. Developing a PTEN-ETS signature to improve molecular risk stratification in prostate cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4504.

Abstract LB-152: EZH2-induced lysine methylation and ERG-EZH2 genomic co-occupancy set the basis for extensive transcriptome reprogramming and prostate cancer progression

Abstract The TMPRSS2-ERG gene fusion is found in about half of prostate tumors and represents one of the most frequent genetic rearrangements in human cancers. The gene fusion provides a mechanism for androgen-stimulated ERG over-expression and broad transcriptional reprogramming in prostate cancer. However, the biological impact of aberrant ERG expression on tumor initiation and progression is still unclear. ERG apparently requires additional co-factors to fully exert its oncogenic effects but the molecular details of these interactions are not defined yet. Understanding these mechanisms could have a huge impact on the clinical management of this disease. Enhancer of zest homolog 2 (EZH2), the catalytic subunit of the Polycomb repressive complex 2 (PRC2) catalyzing histone H3 lysine 27 tri-methylation, is over-expressed in many human cancers and is associated with prostate cancer progression. In primary and metastatic prostate tumors ERG and EZH2 are frequently and concomitantly up-regulated. In this study we tested the hypothesis that EZH2 could act as a co-factor of ERG enhancing its transcriptional and oncogenic activity and identified a novel mechanism driving ERG activation and prostate cancer progression. We found that EZH2 physically interacts with ERG in ERG fusion positive cell lines and human tumors. Moreover, ERG/EZH2 co-occupied multiple genomic sites forming co-activator/co-repressor complexes and enabling massive transcriptional reprogramming. Expression of ERG/EZH2 co-occupied genes reflected the level of ERG activation, was preferentially deregulated in ERG-positive tumors and predicted clinical outcome. Furthermore, EZH2 catalyzed the methylation of ERG at a highly conserved lysine (K362) residue, which resulted in increased chromatin binding and transcriptional activity of ERG. PTEN deficiency and AKT activation promoted ERG methylation and ERG/EZH2 genomic co-occupancy along with a more aggressive and metastatic phenotype in ERG fusion positive cancer cells. Thus, this study identifies the ERG/EZH2 interaction and EZH2-induced ERG methylation as important elements promoting prostate tumorigenesis and at the center of cross-talks between the ERG gene fusion and PTEN deficiency in prostate cancer. Notably, these events were blocked effectively by pharmacological inhibitors of EZH2 providing the rationale for novel context-dependent therapeutic strategies in ERG positive prostate cancer. Citation Format: Giuseppina M. R. Carbone, Laura Curti, Marita Zoma, Abhishek Mitra, Dheeraj Shinde, Domenico Albino, Simona Rossi, Gianluca Civenni, George N. Thalmann, Giovanna Chiorino, Carlo Catapano. EZH2-induced lysine methylation and ERG-EZH2 genomic co-occupancy set the basis for extensive transcriptome reprogramming and prostate cancer progression. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr LB-152.

The Germ Cell Gene TDRD1 as an ERG Target Gene and a Novel Prostate Cancer Biomarker

TMPRSS2-ERG fusion occurs in about half of prostate cancers and results in over-expression of the oncogenic ERG protein in the prostate. The mechanism by which ERG contributes to prostate cancer initiation and progression remains largely unknown. Because ERG is a transcriptional activator, we reasoned that the target genes regulated by ERG could contribute to prostate cancer development. In a search for ERG target genes, we took advantage of published datasets from the MSKCC Prostate Oncogene Project, in which a comprehensive analysis was applied to define transcriptomes in 150 prostate tumors. We retrieved the mRNA expression dataset, split them based on ERG expression, and identified genes whose expression levels are associated with ERG mRNA levels. mRNA expression levels of 21 genes were found to be significantly increased, while for one gene it was decreased in ERG-positive prostate tumors. Among them, the expression of TDRD1 was the most significantly increased in ERG-positive tumors. Among 131 primary prostate tumors which were primarily from European American patients, TDRD1 is over-expressed in 68% of samples, while ERG is overexpressed in 48% of samples, suggesting an additional ERG-independent mechanism of TDRD1 overexpression. In African American prostate tumors, TDRD1 mRNA is expressed in 44%, while ERG is expressed in 24% of samples. In normal tissues, TDRD1 mRNA is exclusively expressed in germ cells and its protein is also known as cancer/testis antigen 41.1 (CT41.1). We generated a mouse monoclonal antibody that recognizes human TDRD1 protein with high specificity and sensitivity. By Western blot analysis and immunohistochemistry (IHC) staining, we demonstrate that TDRD1 protein is expressed in the majority of human prostate tumors, but not in normal prostate tissue. Finally, TDRD1 is not induced in the prostate of ERG overexpression transgenic mice, suggesting that such model does not fully recapitulate the TMPRSS2/ERG fusion-dependent human prostate cancer development. Our results suggest TDRD1 as a novel prostate cancer biomarker. As an ERG target gene, TDRD1 might play an important role in human prostate cancer development, and as a cancer/testis antigen, TDRD1 might have long-term potential to be a therapeutic target for prostate cancer immunotherapy. Prostate 76:1271-1284, 2016. © 2016 Wiley Periodicals, Inc.

High alpha-methylacyl-CoA racemase (AMACR) is associated with ERG expression and with adverse clinical outcome in patients with localized prostate cancer.

Alpha-methylacyl-CoA racemase (AMACR) is a well-characterized marker extensively utilized in prostate cancer (PCA) diagnosis. However, the prognostic value of AMACR expression and its relation to TMPRSS2-ERG gene rearrangement as one of the most common molecular alterations in PCA is not fully explored. AMACR expression was investigated in a cohort of 218 men with localized PCA treated by radical prostatectomy and correlated with ERG and various clinical and pathological parameters. In vitro studies assessed AMACR changes to ERG knockdown and other related genes. In addition, bioinformatics validated the significance of AMACR/ERG expression and assessed relevant genetic signatures in relation to AMACR/ERG expression. AMACR expression was significantly associated with disease progression and with ERG (p ∼0). Seventeen percent of cancer foci showed negative/weak AMACR expression while being ERG positive. High AMACR expression was significantly associated with positive surgical margins (p = 0.01), specifically in tumors with lower Gleason score <7, with ∼95% exhibiting positive surgical margin (p = 0.008). High AMACR showed marginal association with PSA biochemical recurrence (BCR) (p = 0.06) which was slightly more pronounced in ERG-positive tumors (p = 0.04). This was validated in other public cohorts. However, in this cohort, the association with BCR was not statistically significant in multivariate analysis (p = 0.09). Using in vitro cellular models, AMACR messenger RNA (mRNA) expression, but not protein levels, showed an association with ERG expression. We report for the first time a significant association between AMACR and ERG with prognostic implication. Patients with high AMACR/ERG-positive PCA may be at higher risk for disease progression, and additional studies in larger cohorts are needed to confirm the above findings. Functional studies investigating the molecular pathways connecting AMACR and ERG may provide an additional insight into PCA progression pathways.

The tumor suppressive miR-200b subfamily is an ERG target gene in human prostate tumors.

The TMPRSS2-ERG fusion occurs in approximately 50% of prostate cancer (PCa), resulting in expression of the oncogenic ERG in the prostate. Because ERG is a transcriptional activator, we hypothesized that ERG-regulated genes contribute to PCa development. Since microRNA (miRNA) has crucial functions in cancer, we searched for miRNAs regulated by ERG in PCas. We mined published datasets based on the MSKCC Prostate Oncogene Project, in which a comprehensive analysis defined the miRNA transcriptomes in 113 PCas. We retrieved the miRNA expression datasets, and identified miRNAs differentially expressed between ERG-positive and ERG-negative samples. Out of 369 miRNAs, miR-200a, −200b, −429 and −205 are the only miRNAs significantly increased in ERG-positive tumors. Strikingly, miR-200a, −200b and −429 are transcribed as a single polycistronic transcript, suggesting they are regulated at the transcriptional level. With ChIP-qPCR and in vitro binding assay, we identified two functional ETS motifs in the miR-200b/a/429 gene promoter. Knockdown of ERG in PCa cells reduced expression of these three miRNAs. In agreement with the well-established tumor suppressor function, overexpression of the miR-200b/a/429 gene inhibited PCa cell growth and invasion. In summary, our study reveals that miR-200b/a/429 is an ERG target gene, which implicates an important role in TMPRSS2/ERG-dependent PCa development. Although induction of the tumor suppressive miR-200b subfamily by oncogenic ERG appears to be counterintuitive, it is consistent with the observation that the vast majority of primary prostate cancers are slow-growing and indolent.

MP90-14 DISTINCT BIOLOGICAL FEATURES OF PROSTATE TUMORS IN THE CONTEXT OF RACE/ETHNICITY AND ERG STATUS

You have accessJournal of UrologyProstate Cancer: Basic Research & Pathophysiology IV1 Apr 2016MP90-14 DISTINCT BIOLOGICAL FEATURES OF PROSTATE TUMORS IN THE CONTEXT OF RACE/ETHNICITY AND ERG STATUS Jennifer Cullen, Denise Young, Yongmei Chen, Michael Degon, Wagner Baptist, James Farrell, Jason Sedarsky, Claire Huai-Ching Kuo, Jacob Kagan, Sudhir Srivastava, Inger Rosner, Gyorgy Petrovics, Albert Dobi, David McLeod, Shiv Srivastava, and Isabell Sesterhenn Jennifer CullenJennifer Cullen More articles by this author , Denise YoungDenise Young More articles by this author , Yongmei ChenYongmei Chen More articles by this author , Michael DegonMichael Degon More articles by this author , Wagner BaptistWagner Baptist More articles by this author , James FarrellJames Farrell More articles by this author , Jason SedarskyJason Sedarsky More articles by this author , Claire Huai-Ching KuoClaire Huai-Ching Kuo More articles by this author , Jacob KaganJacob Kagan More articles by this author , Sudhir SrivastavaSudhir Srivastava More articles by this author , Inger RosnerInger Rosner More articles by this author , Gyorgy PetrovicsGyorgy Petrovics More articles by this author , Albert DobiAlbert Dobi More articles by this author , David McLeodDavid McLeod More articles by this author , Shiv SrivastavaShiv Srivastava More articles by this author , and Isabell SesterhennIsabell Sesterhenn More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2016.02.2558AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Oncogenic activation of the ERG in prostate cancer (CaP) results from tumor cell specific gene fusions (predominantly TMPRSS2-ERG) that encode truncated ERG oncoprotein. Despite the prevalence of ERG alterations in CaP, emerging reports have underscored remarkable differences in the frequency of ERG alterations among different racial/ethnic groups. In this study, ERG oncoprotein status is compared in a large cohort of African American (AA) and Caucasian American (CA) men within an equal-access military healthcare system and examined for its association with CaP progression. METHODS Representative sections of whole mounted prostatectomy specimens from AA (N=373) and CA (N=684) patients were evaluated for ERG oncoprotein status by previously established immuno-histochemistry assays that showed very high concordance between ERG oncoprotein expression and ERG genomic fusions. ERG positive or negative status of index prostate tumors was analyzed for associations with self reported race/ethnicity, BMI, tumor grade, stage, multi-focality, and biochemical recurrence (BCR) and distant metastasis. RESULTS Comparison of ERG frequencies in index tumors of AA and CA men confirmed the significantly lower rate of ERG alterations in this large cohort of AA CaP. Another striking finding was the significant association between negative ERG index tumor status with greater odds of BCR and metastasis over time for CA men, controlling for key clinical covariates. CONCLUSIONS Distinct biological behavior of ERG positive or negative tumors in the context of race was evident in this study. These data are consistent with some of the previous reports including ours. This large cohort study underscores that ERG negative index status in CA men is predictive of disease progression. © 2016FiguresReferencesRelatedDetails Volume 195Issue 4SApril 2016Page: e1150 Advertisement Copyright & Permissions© 2016MetricsAuthor Information Jennifer Cullen More articles by this author Denise Young More articles by this author Yongmei Chen More articles by this author Michael Degon More articles by this author Wagner Baptist More articles by this author James Farrell More articles by this author Jason Sedarsky More articles by this author Claire Huai-Ching Kuo More articles by this author Jacob Kagan More articles by this author Sudhir Srivastava More articles by this author Inger Rosner More articles by this author Gyorgy Petrovics More articles by this author Albert Dobi More articles by this author David McLeod More articles by this author Shiv Srivastava More articles by this author Isabell Sesterhenn More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Characterizing the molecular features of ERG-positive tumors in primary and castration resistant prostate cancer.

The TMPRSS2-ERG gene fusion is detected in approximately half of primary prostate cancers (PCa) yet the prognostic significance remains unclear. We hypothesized that ERG promotes the expression of common genes in primary PCa and metastatic castration-resistant PCa (CRPC), with the objective of identifying ERG-associated pathways, which may promote the transition from primary PCa to CRPC. We constructed tissue microarrays (TMA) from 127 radical prostatectomy specimens, 20 LuCaP patient-derived xenografts (PDX), and 152 CRPC metastases obtained immediately at time of death. Nuclear ERG was assessed by immunohistochemistry (IHC). To characterize the molecular features of ERG-expressing PCa, a subset of IHC confirmed ERG+ or ERG- specimens including 11 radical prostatectomies, 20 LuCaP PDXs, and 45 CRPC metastases underwent gene expression analysis. Genes were ranked based on expression in primary PCa and CRPC. Common genes of interest were targeted for IHC analysis and expression compared with biochemical recurrence (BCR) status. IHC revealed that 43% of primary PCa, 35% of the LuCaP PDXs, and 18% of the CRPC metastases were ERG+ (12 of 48 patients [25%] had at least one ERG+ metastasis). Based on gene expression data and previous literature, two proteins involved in calcium signaling (NCALD, CACNA1D), a protein involved in inflammation (HLA-DMB), CD3 positive immune cells, and a novel ERG-associated protein, DCLK1 were evaluated in primary PCa and CRPC metastases. In ERG+ primary PCa, a weak association was seen with NCALD and CACNA1D protein expression. HLA-DMB association with ERG was decreased and CD3 cell number association with ERG was changed from positive to negative in CRPC metastases compared to primary PCa. DCLK1 was upregulated at the protein level in unpaired ERG+ primary PCa and CRPC metastases (P = 0.0013 and P < 0.0001, respectively). In primary PCa, ERG status or expression of targeted proteins was not associated with BCR-free survival. However, for primary PCa, ERG+DCLK1+ patients exhibited shorter time to BCR (P = 0.06) compared with ERG+DCLK1- patients. This study examined ERG expression in primary PCa and CRPC. We have identified altered levels of inflammatory mediators associated with ERG expression. We determined expression of DCLK1 correlates with ERG expression and may play a role in primary PCa progression to metastatic CPRC. Prostate 76:810-822, 2016. © 2016 Wiley Periodicals, Inc.

Loss of miR-449a in ERG-associated prostate cancer promotes the invasive phenotype by inducing SIRT1.

Epigenetic regulation by SIRT1, a multifaceted NAD+-dependent protein deacetylase, is one of the most common factors modulating cellular processes in a broad range of diseases, including prostate cancer (CaP). SIRT1 is over-expressed in CaP cells, however the associated mechanism is not well understood. To identify whether specific microRNAs might mediate this linkage, we have screened a miRNA library for differential expression in ERG-associated CaP tissues. Of 20 differentially and significantly expressed miRNAs that distinguish ERG-positive tumors from ERG-negative tumors, we find miR-449a is highly suppressed in ERG-positive tumors. We establish that SIRT1 is a direct target of miR-449a and is also induced by ERG in ERG-associated CaP. Our data suggest that attenuation of miR-449a promotes the invasive phenotype of the ERG-positive CaP in part by inducing the expression of SIRT1 in prostate cancer cells. Furthermore, we also find that suppression of SIRT1 results in a significant reduction in ERG expression in ERG-positive CaP cells, indicating a feed-back regulatory loop associated with ERG, miR-449a and SIRT1. We also report that ERG suppresses p53 acetylation perhaps through miR-449a-SIRT1 axis in CaP cells. Our findings provide new insight into the function of miRNAs in regulating ERG-associated CaP. Thus, miR-449a activation or SIRT1 suppression may represent new therapeutic opportunity for ERG-associated CaP.

Heterogeneity and chronology of 6q15 deletion and ERG-fusion in prostate cancer

Prostate cancer is notorious for its heterogeneity, which poses a problem for the applicability of diagnostic molecular markers. However, heterogeneity analysis can provide valuable information on the chronology in which molecular alterations arise. Here, we constructed a heterogeneity tissue microarray (TMA) comprising samples from 10 different tumor areas of 189 prostate cancers each in order to study the sequence of two frequent molecular alterations, i.e. 6q15 deletion and TMPRSS2:ERG fusion. Previous work shows a marked inverse relationship between these alterations, suggesting that presence of one of these alterations might impact development of the other. 6q15 deletion was analyzed by fluorescence in situ hybridization and ERG-expression by immunohistochemistry. Only 6.6% of 334 ERG-positive but 28.4% of 440 ERG-negative TMA spots showed 6q15 deletions (p < 0.0001). A breakdown of these data to the level of tumor foci revealed 6q deletions in 138 tumor foci that were large enough to have at least 3 analyzable TMA spots. These included 42 tumor foci with homogeneous ERG positivity and 16 with homogeneous 6q15 deletions. Remarkably, six of the 42 homogeneously ERG-positive tumor foci (14.3%) harbored small 6q15-deleted areas, but none of the 34 6q15-deleted foci showed areas of ERG positivity (p = 0.022). In conclusion, our data suggest that ERG-fusion can precede 6q15 deletion, but not vice versa. The complete absence of ERG-positive tumor areas in 6q15-deleted tumor foci further suggest that the functional consequences of 6q15 deletions may prevent the development of TMPRSS2:ERG fusions.

PTEN Loss as Determined by Clinical-grade Immunohistochemistry Assay Is Associated with Worse Recurrence-free Survival in Prostate Cancer

BackgroundPTEN is the most commonly deleted tumor suppressor gene in primary prostate cancer (PCa) and its loss is associated with poor clinical outcomes and ERG gene rearrangement. ObjectiveWe tested whether PTEN loss is associated with shorter recurrence-free survival (RFS) in surgically treated PCa patients with known ERG status. Design, setting, and participantsA genetically validated, automated PTEN immunohistochemistry (IHC) protocol was used for 1275 primary prostate tumors from the Canary Foundation retrospective PCa tissue microarray cohort to assess homogeneous (in all tumor tissue sampled) or heterogeneous (in a subset of tumor tissue sampled) PTEN loss. ERG status as determined by a genetically validated IHC assay was available for a subset of 938 tumors. Outcome measurements and statistical analysisAssociations between PTEN and ERG status were assessed using Fisher's exact test. Kaplan-Meier and multivariate weighted Cox proportional models for RFS were constructed. Results and limitationsWhen compared to intact PTEN, homogeneous (hazard ratio [HR] 1.66, p=0.001) but not heterogeneous (HR 1.24, p=0.14) PTEN loss was significantly associated with shorter RFS in multivariate models. Among ERG-positive tumors, homogeneous (HR 3.07, p<0.0001) but not heterogeneous (HR 1.46, p=0.10) PTEN loss was significantly associated with shorter RFS. Among ERG-negative tumors, PTEN did not reach significance for inclusion in the final multivariate models. The interaction term for PTEN and ERG status with respect to RFS did not reach statistical significance (p=0.11) for the current sample size. ConclusionsThese data suggest that PTEN is a useful prognostic biomarker and that there is no statistically significant interaction between PTEN and ERG status for RFS. Patient summaryWe found that loss of the PTEN tumor suppressor gene in prostate tumors as assessed by tissue staining is correlated with shorter time to prostate cancer recurrence after radical prostatectomy.

Abstract 5277: ERG-based stratification of prostate cancer highlights ethnicity associated biological differences

Abstract Introduction: The molecular basis of racial/ethnic differences in the detection and outcomes of the carcinoma of the prostate (CaP) needs to be better defined to improve biomarkers and identify appropriate therapeutic targets. ETS-related gene fusions in proto-oncogenes (ERG, ETV1) are among the most common early oncogenic alterations described in CaP. Studies have highlighted the racial/ethnic variation in ERG fusion and ERG oncoprotein expression frequencies showing: the highest frequencies in Caucasian (CA) men, the lowest frequencies in Asian men, and intermediate frequencies in African American (AA) men. These differences persisted when patients were carefully matched for age and clinico-pathologic features. Building on our and other's previous findings, the current study was conducted to evaluate ERG expression frequencies in a large cohort of AA and CA CaP patients. Methods: Patients enrolled at the Walter Reed National Military Medical Center who consented to provide radical prostatectomy (RP) specimens to the CPDR Tissue Bank and Clinical Database were evaluated. The current study included 443 AA and 696 CA patients. ERG oncoprotein expression was detected by immunohistochemistry using the 9FY mouse monoclonal anti-ERG antibody and was correlated with clinico-pathologic features and CaP outcomes, including time to biochemical recurrence.&amp;lt;br Results: The result revealed lower frequencies of ERG in the index tumors of AA men as compared to CA men (28.4% vs. 51.5%, p &amp;lt; 0.0001). Furthermore, AA men with higher grade CaP (Gleason 8-10) had even lower ERG frequency (16%), suggesting distinct molecular biologic features in aggressive CaP for AA men. A trend with age was observed for both racial groups, with younger CA showing higher ERG frequencies. At least one focus of ERG positive PIN was present in prostates with ERG positive tumors. In both CA and AA patients, a very low frequency (8%) of ERG expression was noted in anterior index tumors compared to posterior tumors. Conclusions: This study lends increasing support to differences of the most common early genomic alteration (ERG) between AA and CA CaP patients. The study also underscores the importance of identifying new or improving existing biomarkers with consideration of patient race/ethnicity. Citation Format: Albert Dobi, Michael Degon, James Farrell, Wagner Baptiste, Denise Young, Yongmei Chen, Gyorgy Petrovics, Jennifer Cullen, Jacob Kagan, Sudir Srivastava, Inger Rosner, David G. McLeod, Shiv Srivastava, Isabell Sesterhenn. ERG-based stratification of prostate cancer highlights ethnicity associated biological differences. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5277. doi:10.1158/1538-7445.AM2015-5277

Abstract 2016: Evaluation of prostate cancer biomarkers and therapeutic targets in FFPE specimens using the NanoString platform

Abstract Introduction: Identification of prognostic indicators of disease progression is one of the highest priorities in prostate cancer (CaP) research. The majority of CaP tissue specimens available for biomarker discovery and validation are formalin fixed paraffin embedded (FFPE) specimens. The RNA quality of these tissues for traditional gene expression analysis approaches is not adequate, especially of cases with long term- follow up necessary for prognostic marker studies. We systematically evaluated and optimized the NanoString platform for CaP gene expression analysis in FFPE whole mounted prostate specimens with up to 15 years follow up. Methods: NanoString readouts were compared between a series of input total RNA prepared through different tissue dissection methods. We then performed NanoString analysis of 151-CaP probe set on 63 archived CaP samples to evaluate this modified protocol. ERG gene expression was validated by immunohistochemistry (IHC) assay using CPDR ERG-MAb (9FY). Results: We have performed NanoString analysis of CPDR designed 151- CaP gene panel on whole-mounted prostate tissue specimens using our optimized protocol. The prostate epithelial cell markers KLK3, MSMB and PAP had the highest signals in all samples reflecting the prostate epithelial origin of the specimens. Established CaP specific genes, such as AMACR, PCA3 and PSGR were most highly over-expressed in tumors compared to matched benign prostate epithelium. ERG positive tumors had strong expression of both TMPRSS2-ERG fusion probes and probes for several ERG splice forms (ERG 1,2,3 and 8), with no signals in the matched benign samples. The ERG gene expression status was consistent with blinded ERG IHC results. Conclusions: The feasibility of NanoString profiling in archived whole mounted FFPE prostate specimens was demonstrated. This approach is well suited for a medium throughput evaluation and validation of prognostic biomarker and therapeutic target candidates in CaP. Citation Format: Jocelyn Lee, Wusheng Yan, Denise Young, Yingjie Song, Yongmei Chen, Shilpa Katta, Ahmed Mohamed, Lakshmi Ravindranath, Alagarsamy Srinivasan, Jennifer Cullen, Jacob Kagan, Sudhir Srivastava, Albert Dobi, Inger Rosner, David G. McLeod, Isabell A. Sesterhenn, Shiv Srivastava, Gyorgy Petrovics. Evaluation of prostate cancer biomarkers and therapeutic targets in FFPE specimens using the NanoString platform. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2016. doi:10.1158/1538-7445.AM2015-2016

Abstract 939: A prospective investigation of PTEN loss and ERG expression in lethal prostate cancer

Abstract Introduction: PTEN is a tumor suppressor frequently deleted in prostate cancer that may be a useful prognostic biomarker. However, the association of PTEN loss with lethal disease has not been tested in a large, predominantly surgically-treated cohort. Moreover, PTEN loss is more common in TMPRSS2:ERG gene fusion-positive tumors, compared to fusion-negative tumors. Thus, presence of TMPRSS2:ERG gene fusion may modify the effects of PTEN loss on disease progression. We conducted a large patho-epidemiology investigation among prostate cancer patients in the Health Professional Follow-up Study and the Physicians’ Health Study addressing: 1) the association of PTEN loss, assessed by a validated IHC protocol, with lethal progression, and 2) the potential for TMPRSS2:ERG gene fusion, detected by IHC, to modify the role of PTEN loss in lethal disease progression. Methods: In the Health Professionals Follow-Up Study and Physicians’ Health Study, we followed 1045 incident prostate cancer cases diagnosed between 1986 - 2009 for cancer and all-cause mortality. A genetically validated PTEN immunohistochemistry (IHC) assay was performed on tissue microarrays (TMA). TMPRSS2:ERG status was previously assessed in a subset of cases by a genetically validated IHC assay for ERG. Cox proportional hazards models adjusting for age and BMI at diagnosis, Gleason grade, and clinical or pathologic TNM stage were used to estimate hazard ratios (HR) and 95% confidence intervals (CI) of the association with lethal disease. Results: On average men were followed 11.6 years, during which there were 82 lethal events. 16% of cases had complete PTEN loss in all TMA cores and 9% had heterogeneous PTEN loss in a subset of cores. Men whose tumors showed complete PTEN loss had a significantly higher risk for lethal progression compared to men whose tumors showed PTEN intact or heterogeneous loss, even after adjusting for clinical-pathologic characteristics (HR = 1.9; 95% CI = 1.2-3.0). The association of PTEN loss (complete or heterogeneous) with lethal progression was only among men with ERG negative (HR = 2.8; 95% CI = 1.5-5.2), but not ERG positive (HR = 1.2; 95% CI = 0.6-2.2) tumors. Conclusions: PTEN loss is independently associated with increased risk of lethal progression, particularly in the ERG fusion negative subgroup. These validated and inexpensive IHC assays may be useful for risk stratification in prostate cancer. Citation Format: Thomas U. Ahearn, Andreas Pettersson, Ericka M. Ebot, Travis Gerke, Carlos De Morais, Jessica Hicks, Kathryn M. Wilson, Jennifer R. Rider, Michelangelo Fiorentino, Stephen Finn, Edward L. Giovannucci, Massimo Loda, Meir J. Stampfer, Angelo M. De Marzo, Lorelei A. Mucci, Tamara L. Lotan. A prospective investigation of PTEN loss and ERG expression in lethal prostate cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 939. doi:10.1158/1538-7445.AM2015-939

Loss of SOX9 Expression Is Associated with PSA Recurrence in ERG-Positive and PTEN Deleted Prostate Cancers.

The transcription factor SOX9 plays a crucial role in normal prostate development and has been suggested to drive prostate carcinogenesis in concert with PTEN inactivation. To evaluate the clinical impact of SOX9 and its relationship with key genomic alterations in prostate cancer, SOX9 expression was analyzed by immunohistochemistry on a tissue microarray containing 11,152 prostate cancers. Data on ERG status and deletions of PTEN, 3p13, 5q21 and 6q15 were available from earlier studies. SOX9 expression levels were comparable in luminal cells of normal prostate glands (50% SOX9 positive) and 3,671 cancers lacking TMPRSS2:ERG fusion (55% SOX9 positive), but was markedly increased in 3,116 ERG-fusion positive cancers (81% SOX9 positive, p<0.0001). While no unequivocal changes in the SOX9 expression levels were found in different stages of ERG-negative cancers, a gradual decrease of SOX9 paralleled progression to advanced stage, high Gleason grade, metastatic growth, and presence of PTEN deletions in ERG-positive cancers (p<0.0001 each). SOX9 levels were unrelated to deletions of 3p, 5q, and 6q. Down-regulation of SOX9 expression was particularly strongly associated with PSA recurrence in ERG-positive tumors harboring PTEN deletions (p=0.001), but had no significant effect in ERG-negative cancers or in tumors with normal PTEN copy numbers. In summary, the results of our study argue against a tumor-promoting role of SOX9 in prostate cancer, but demonstrate that loss of SOX9 expression characterizes a particularly aggressive subset of ERG positive cancers harboring PTEN deletions.

Weaker ERG expression in patients with ERG-positive prostate cancer is associated with advanced disease and weaker androgen receptor expression: An Indian outlook

IntroductionGene fusion between TMPRSS2 and ERG has a causal role in prostate cancer initiation. However, studies evaluating its role clinically have shown conflicting results. We investigated simultaneously multiple aspects of ERG, namely, “presence” and “strength” of ERG expression and “correlation” of ERG with other common clinicopathological parameters. Materials and methodsFrom January 2012 to November 2013, the status of ERG, androgen receptor (AR), and p53 was prospectively determined by immunohistochemistry unselectively in all types of specimens positive for prostate cancer. “Strength” of expression was measured in terms of “intensity” as well as “percentage positivity,” with each parameter given a score from 0 to 3 based on fixed protocol, which was tested for interrater variability as well as test-retest reliability. Data were collected for age, Gleason score, prostate specific antigen levels, presence of perineural invasion and lymphovascular invasion, high-grade prostatic intraepithelial neoplasia, and cancer stage. ResultsIn total, 100 specimens were analyzed, and overall 51 patients had ERG-positive immunostaining. ERG-positive tumors had lower presence of high-grade prostatic intraepithelial neoplasia and p53 positivity, with no significant difference in prostate specific antigen levels, Gleason scores, and presence of lymphovascular invasion. Moreover, 54 patients had complete stage information, and the absolute number of patients with ERG positivity increased with increasing clinical stage. Among these, 30 patients were ERG positive, and ERG score had strong positive correlation with AR expression (Spearman correlation coefficient 0.677). However, median ERG scores showed a significant decline (consistent across percentage positivity and intensity) in patients with stage 4 disease, and score≤2 had 88.2% specificity in identifying patient with stage 4 disease. Cohen׳s κ = 0.81, whereas intraclass correlation coefficient was 0.95, indicating substantial agreement and near-perfect reproducibility of scoring scheme for immunohistochemistry. ConclusionERG-positive tumors increase in proportion with increasing stage of disease, but strength of ERG expression in ERG-positive patients shows a significant decline, or “loss,” in patients with stage 4 disease. This may have potential therapeutic implications as ERG expression score showed strong positive correlation with AR expression score.

MP6-13 PTEN LOSS BY IMMUNOHISTOCHEMISTRY IS ASSOCIATED WITH BIOCHEMICAL RECURRENCE IN ERG-NEGATIVE PROSTATE TUMORS: A STUDY OF THE CANARY COHORT

You have accessJournal of UrologyProstate Cancer: Markers II1 Apr 2015MP6-13 PTEN LOSS BY IMMUNOHISTOCHEMISTRY IS ASSOCIATED WITH BIOCHEMICAL RECURRENCE IN ERG-NEGATIVE PROSTATE TUMORS: A STUDY OF THE CANARY COHORT Tamara L. Lotan, Wei Wei, Carlos L. Morais, Ziding Feng, Sarah Hawley, Ladan Fazli, Antonio Hurtado-Coll, Dean Troyer, Jesse McKenney, Jeff Simko, Lawrence True, James D. Brooks, and CAnary Pathology Team Investigating Tissue Biomarkers (CAPIT) Tamara L. LotanTamara L. Lotan More articles by this author , Wei WeiWei Wei More articles by this author , Carlos L. MoraisCarlos L. Morais More articles by this author , Ziding FengZiding Feng More articles by this author , Sarah HawleySarah Hawley More articles by this author , Ladan FazliLadan Fazli More articles by this author , Antonio Hurtado-CollAntonio Hurtado-Coll More articles by this author , Dean TroyerDean Troyer More articles by this author , Jesse McKenneyJesse McKenney More articles by this author , Jeff SimkoJeff Simko More articles by this author , Lawrence TrueLawrence True More articles by this author , James D. BrooksJames D. Brooks More articles by this author , and CAnary Pathology Team Investigating Tissue Biomarkers (CAPIT)CAnary Pathology Team Investigating Tissue Biomarkers (CAPIT) More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2015.02.260AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES PTEN is the most commonly lost tumor suppressor gene in primary prostate cancer, and its loss is associated with poor clinical outcomes. Though ERG gene rearrangements occur in nearly half of prostate cancers, most studies have shown that ERG status alone is not prognostic. Here, we tested whether ERG status may modify the association of PTEN loss with biochemical recurrence (BCR) in a large cohort of surgically-treated patients. METHODS 1275 primary prostate tumors from the Canary Retrospective Prostate Cancer Tissue Microarray (TMA) cohort were analyzed for PTEN loss by immunohistochemistry (IHC) using a genetically validated automated staining protocol and scoring system to assess homogeneous (in all sampled tumor glands) or heterogeneous (in a subset of sampled tumor glands) PTEN loss. IHC for ERG was performed on 954 tumors and scored dichotomously. Associations of PTEN loss and ERG status were assessed by Fisher's exact test, and Kaplan-Meier and multivariate weighted Cox proportional models for time to BCR were constructed for all patients and the ERG-positive and ERG-negative subsets separately. RESULTS PTEN loss (homogeneous or heterogeneous) was enriched in the ERG-positive tumors, with 33% (132/401) of ERG-positive tumors showing any PTEN loss compared to 18% (99/537) of ERG-negative tumors (p<0.0001). Homogeneous PTEN loss was also associated with increased Gleason grade and pathologic stage (p<0.0001 for each), however, PTEN loss was not associated with pre-operative PSA or patient age. In the whole cohort, homogeneous PTEN loss was significantly associated with decreased time to BCR in multivariate models compared to PTEN intact or heterogeneous loss (HR=1.39 and 1.64, p = 0.02 and 0.01, respectively). However, the association of PTEN loss with time to BCR differed by ERG status. Homogeneous PTEN loss was significantly associated with decreased time to BCR compared with intact or heterogeneous loss in the ERG-negative tumors after adjusting for clinical-pathologic factors (HR= 2.22 and 2.33, p < 0.0001 and p = 0.003, respectively) but not in the ERG-positive tumors (HR = 1.22 and 1.47, p = 0.54 and 0.30). CONCLUSIONS Homogeneous PTEN loss by validated IHC assay is associated with decreased time to BCR; this effect was much stronger in ERG-negative tumors than in ERG-positive tumors. If validated in additional cohorts, these data suggest that the PTEN and ERG biomarkers should be assessed together. © 2015 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 193Issue 4SApril 2015Page: e59 Advertisement Copyright & Permissions© 2015 by American Urological Association Education and Research, Inc.MetricsAuthor Information Tamara L. Lotan More articles by this author Wei Wei More articles by this author Carlos L. Morais More articles by this author Ziding Feng More articles by this author Sarah Hawley More articles by this author Ladan Fazli More articles by this author Antonio Hurtado-Coll More articles by this author Dean Troyer More articles by this author Jesse McKenney More articles by this author Jeff Simko More articles by this author Lawrence True More articles by this author James D. Brooks More articles by this author CAnary Pathology Team Investigating Tissue Biomarkers (CAPIT) More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

MP66-06 OPTIMIZATION OF NANOSTRING PLATFORM FOR EVALUATION OF PROSTATE CANCER BIOMARKERS AND THERAPEUTIC TARGETS IN FFPE SPECIMENS

INTRODUCTION AND OBJECTIVES: Identification of prognostic indicators of disease progression is one of the highest priorities in prostate cancer (CaP) research. The majority of CaP tissue specimens available for biomarker discovery and validation are formalin fixed paraffin embedded (FFPE) specimens. The RNA quality of these tissues for traditional gene expression analysis approaches is not adequate, especially of cases with long termfollow up necessary for prognostic marker studies. We systematically evaluated and optimized the NanoString platform for CaP gene expression analysis in FFPE whole mounted prostate specimens with up to 15 years follow up. METHODS: NanoString readouts were compared between a series of input total RNA prepared through different tissue dissection methods. We then performed NanoString analysis of 151-CaP probe set on 63 archived CaP samples to evaluate this modified protocol. ERG gene expression was validated by immunohistochemistry (IHC) assay using CPDR ERG-MAb (9FY). RESULTS: We have performed NanoString analysis of CPDR designed 151CaP gene panel on whole-mounted prostate tissue specimens using our optimized protocol. The prostate epithelial cell markers KLK3, MSMB and PAP had the highest signals in all samples reflecting the prostate epithelial origin of the specimens. Established CaP specific genes, such as AMACR, PCA3 and PSGR were most highly over-expressed in tumors compared to matched benign prostate epithelium. ERG positive tumors had strong expression of both TMPRSS2-ERG fusion probes and probes for several ERG splice forms (ERG 1,2,3 and 8), with no signals in the matched benign samples. The ERG gene expression status was consistent with blinded ERG IHC results. CONCLUSIONS: The feasibility of NanoString profiling in archived whole mounted FFPE prostate specimens was demonstrated. This approach is well suited for a medium throughput evaluation and validation of prognostic biomarker and therapeutic target candidates in CaP.

MP6-15 PTEN LOSS AND ERG EXPRESSION IN PROSTATE CANCER SURVIVAL

You have accessJournal of UrologyProstate Cancer: Markers II1 Apr 2015MP6-15 PTEN LOSS AND ERG EXPRESSION IN PROSTATE CANCER SURVIVAL Thomas Ahearn, Andreas Pettersson, Ericka Ebot, Travis Gerke, Carlos De Morais, Jessica Hicks, Kathryn Wilson, Jennifer Rider, Michelangelo Fiorentino, Stephen Finn, Edward Giovannucci, Massimo Loda, Meir Stampfer, Angelo De Marzo, Lorelei Mucci, and Tamara Lotan Thomas AhearnThomas Ahearn More articles by this author , Andreas PetterssonAndreas Pettersson More articles by this author , Ericka EbotEricka Ebot More articles by this author , Travis GerkeTravis Gerke More articles by this author , Carlos De MoraisCarlos De Morais More articles by this author , Jessica HicksJessica Hicks More articles by this author , Kathryn WilsonKathryn Wilson More articles by this author , Jennifer RiderJennifer Rider More articles by this author , Michelangelo FiorentinoMichelangelo Fiorentino More articles by this author , Stephen FinnStephen Finn More articles by this author , Edward GiovannucciEdward Giovannucci More articles by this author , Massimo LodaMassimo Loda More articles by this author , Meir StampferMeir Stampfer More articles by this author , Angelo De MarzoAngelo De Marzo More articles by this author , Lorelei MucciLorelei Mucci More articles by this author , and Tamara LotanTamara Lotan More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2015.02.262AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES PTEN is a tumor suppressor frequently deleted in prostate cancer that may be a useful prognostic biomarker. However, the association of PTEN loss with lethal disease has not been tested in a large, predominantly surgically-treated cohort. Moreover, PTEN loss is more common in TMPRSS2:ERG gene fusion-positive tumors, compared to fusion-negative tumors. Thus, presence of TMPRSS2:ERG gene fusion may modify the effects of PTEN loss on disease progression. We conducted a large patho-epidemiology investigation among prostate cancer patients in the Health Professional Follow-up Study and the Physicians' Health Study addressing: 1) the association of PTEN loss, assessed by a validated IHC protocol, with lethal progression, and 2) the potential for TMPRSS2:ERG gene fusion, detected by IHC, to modify the role of PTEN loss in lethal disease progression. METHODS In the Health Professionals Follow-Up Study and Physicians' Health Study, we followed 1045 incident prostate cancer cases diagnosed between 1986 – 2009 for cancer and all-cause mortality. A genetically validated PTEN immunohistochemistry (IHC) assay was performed on tissue microarrays (TMA). TMPRSS2:ERG status was previously assessed in a subset of cases by a genetically validated IHC assay for ERG. Cox proportional hazards models adjusting for age and BMI at diagnosis, Gleason grade, and clinical or pathologic TNM stage were used to estimate hazard ratios (HR) and 95% confidence intervals (CI) of the association with lethal disease. RESULTS On average men were followed 11.6 years, during which there were 81 lethal events. 16% of cases had complete PTEN loss in all TMA cores and 9% had heterogeneous PTEN loss in a subset of cores. After adjustment for clinical-pathologic variables, complete PTEN loss was associated with lethal progression (HR=1.9; 95% CI=1.2-3.0). The association of PTEN loss (complete or heterogeneous) with lethal progression was only among men with ERG negative (HR=2.8; 95% CI=1.5-5.2), but not ERG positive (HR=1.2; 95% CI=0.6-2.2) tumors. CONCLUSIONS PTEN loss is independently associated with increased risk of lethal progression, particularly in the ERG fusion negative subgroup. These validated and inexpensive IHC assays may be useful for risk stratification in prostate cancer. © 2015 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 193Issue 4SApril 2015Page: e59-e60 Peer Review Report Advertisement Copyright & Permissions© 2015 by American Urological Association Education and Research, Inc.MetricsAuthor Information Thomas Ahearn More articles by this author Andreas Pettersson More articles by this author Ericka Ebot More articles by this author Travis Gerke More articles by this author Carlos De Morais More articles by this author Jessica Hicks More articles by this author Kathryn Wilson More articles by this author Jennifer Rider More articles by this author Michelangelo Fiorentino More articles by this author Stephen Finn More articles by this author Edward Giovannucci More articles by this author Massimo Loda More articles by this author Meir Stampfer More articles by this author Angelo De Marzo More articles by this author Lorelei Mucci More articles by this author Tamara Lotan More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...