Non-neoplastic Urothelium Research Articles

Artificial Intelligence to Predict the BRAF V595E Mutation in Canine Urinary Bladder Urothelial Carcinomas.

In dogs, the BRAF mutation (V595E) is common in bladder and prostate cancer and represents a specific diagnostic marker. Recent advantages in artificial intelligence (AI) offer new opportunities in the field of tumour marker detection. While AI histology studies have been conducted in humans to detect BRAF mutation in cancer, comparable studies in animals are lacking. In this study, we used commercially available AI histology software to predict BRAF mutation in whole slide images (WSI) of bladder urothelial carcinomas (UC) stained with haematoxylin and eosin (HE), based on a training (n = 81) and a validation set (n = 96). Among 96 WSI, 57 showed identical PCR and AI-based BRAF predictions, resulting in a sensitivity of 58% and a specificity of 63%. The sensitivity increased substantially to 89% when excluding small or poor-quality tissue sections. Test reliability depended on tumour differentiation (p < 0.01), presence of inflammation (p < 0.01), slide quality (p < 0.02) and sample size (p < 0.02). Based on a small subset of cases with available adjacent non-neoplastic urothelium, AI was able to distinguish malignant from benign epithelium. This is the first study to demonstrate the use of AI histology to predict BRAF mutation status in canine UC. Despite certain limitations, the results highlight the potential of AI in predicting molecular alterations in routine tissue sections.

CD10 expression in urinary bladder urothelial carcinoma is associated with high-tumor grade and stage.

Primary urinary bladder carcinoma is a common cancer worldwide. There is limited published data about CD10 immunoexpression pattern in urothelial bladder carcinoma (UBC). This study aims to examine CD10 immunoexpression in UBC and evaluate its relationship with clinicopathological parameters. The retrospective study examined 130 samples of UBC tissue and 30 samples of non-neoplastic urothelial bladder tissue, which were obtained from the Anatomic Pathology Department, King Abdulaziz University, Jeddah, Saudi Arabia. The project started in June 2019 and completed in February 2021. Tissue microarrays (TMA) were prepared from paraffin blocks and tissue sections prepared from the recipient blocks were used for immunohistochemistry studies utilizing CD10 antibody. The immunostaining results were recorded and analyzed. Positive staining of CD10 was observed in 64 (49%) cases of UBC and was not detected in any non-neoplastic urothelium samples. CD10-positive staining was identified in 36.7% and 66.7% of low and high-grade tumors, respectively. There was an association between positive CD10 immunostaining and high tumor grade (p=0.006) and muscularis propria invasion (p=0.007). There was no association between CD10 immunoexpression and age, gender, nodal and distant metastasis, lymphovascular invasion, and tumor recurrence. CD10 immunoexpression was not associated with the probabilities of overall survival (log rank 1.663, p=0.197) or disease-free survival (log rank 1.637, p=0.201). In UBC, CD10 immunoexpression is associated with higher tumor grade and muscle invasion, but it is not associated with patient survival or other clinicopathological parameters. CD10 immunoexpression cannot be used as a biomarker for poor prognosis in UBC.

Clinicopathological significance of claspin overexpression and its efficacy as a novel biomarker for the diagnosis of urothelial carcinoma.

We previously reported that claspin is a key regulator in the progression of gastric cancer and renal cell carcinoma. However, the clinicopathological significance of claspin in urothelial carcinoma (UC) has not been investigated. We analyzed the expression and distribution of claspin in UC cases by immunohistochemistry. In the non-neoplastic urothelium, the expression of claspin was either weak or absent, whereas UC tissues showed nuclear staining. The expression of claspin was detected in 58 (42%) of a total of 138 upper tract UC cases treated by radical nephroureterectomy without neoadjuvant chemotherapy. Claspin-positive UC cases were associated with nodular/flat morphology, variant histology, high tumor grade, high pathological T grade, and lymphatic and venous invasion. The expression of claspin was significantly associated with decreased progression-free survival and cancer-specific survival. In addition, claspin was co-expressed with Ki-67, PD-L1, HER2, EGFR, and p53 in consecutive tumor sections of UC. An immunohistochemical analysis of claspin in biopsy specimens revealed that strong to moderate claspin staining was more frequently observed in carcinoma in situ in comparison to dysplasia or the benign urothelium. Furthermore, immunocytochemistry for claspin on urine cytology slides demonstrated that the proportion of claspin-positive cells was significantly greater in high-grade UC than in benign cases. These results suggest that claspin may be a novel prognostic marker and a possible therapeutic target molecule for UC. Moreover, claspin could be a useful diagnostic biomarker of urothelial neoplasia.

PD42-07 INDEPENDENT EXPRESSION OF PROGRAMMED DEATH LIGAND 1 FROM NECTIN-4 IN UPPER TRACT UROTHELIAL CARCINOMA

You have accessJournal of UrologyBladder Cancer: Basic Research & Pathophysiology II (PD42)1 Sep 2021PD42-07 INDEPENDENT EXPRESSION OF PROGRAMMED DEATH LIGAND 1 FROM NECTIN-4 IN UPPER TRACT UROTHELIAL CARCINOMA Kazutoshi Fujita, Eisuke Tomiyama, Diana Taheri, Maria Rodriguez Pena, Eri Banno, Taigo Kato, Koji Hatano, Atsunari Kawashima, Takeshi Ujike, Motohide Uemura, Tetsuya Takao, Seiji Yamaguchi, Hiroaki Fushimi, Kazuhiro Yoshimura, Hirotsugu Uemura, George Netto, and Norio Nonomura Kazutoshi FujitaKazutoshi Fujita More articles by this author , Eisuke TomiyamaEisuke Tomiyama More articles by this author , Diana TaheriDiana Taheri More articles by this author , Maria Rodriguez PenaMaria Rodriguez Pena More articles by this author , Eri BannoEri Banno More articles by this author , Taigo KatoTaigo Kato More articles by this author , Koji HatanoKoji Hatano More articles by this author , Atsunari KawashimaAtsunari Kawashima More articles by this author , Takeshi UjikeTakeshi Ujike More articles by this author , Motohide UemuraMotohide Uemura More articles by this author , Tetsuya TakaoTetsuya Takao More articles by this author , Seiji YamaguchiSeiji Yamaguchi More articles by this author , Hiroaki FushimiHiroaki Fushimi More articles by this author , Kazuhiro YoshimuraKazuhiro Yoshimura More articles by this author , Hirotsugu UemuraHirotsugu Uemura More articles by this author , George NettoGeorge Netto More articles by this author , and Norio NonomuraNorio Nonomura More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000002056.07AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Locally advanced or metastatic upper tract urothelial carcinoma (UTUC) is a lethal disease with limited treatment options for patients who do not positively respond to platinum chemotherapy and/or immune checkpoint inhibitor. Enfortumab vedotin is a novel antibody–drug conjugate targeting Nectin-4, which is highly expressed in urothelial carcinoma. However, the expression status of Nectin-4 in UTUC remains unclear. The relationship between Nectin-4 and Programmed Death Ligand 1 (PD-L1) in UTUC is also unclear. In the present study, we investigated the expression of Nectin-4 in UTUC using tissue microarray (TMA) specimens of patients with UTUC as well as the association between Nectin-4 protein expression and clinicopathological features or outcomes. We also examined PD-L1 expression and its correlation with that of Nectin-4. METHODS: UTUC TMA was constructed with spotted triplicate urothelial tumour samples (from dominant tumours/invasive components if present) and paired normal-appearing urothelial tissues (from the renal pelvis and ureter) obtained from 99 patients with non-metastatic UTUC, who underwent radical nephroureterectomy performed at Osaka General Medical Center. We performed immunohistochemical analysis of 99 UTUC tissue microarray with anti-PDL1 and anti-Nectin-4 antibody. RESULTS: Nectin-4-positivity was detected in 65 (65.7%) samples, which was significantly higher than that in non-neoplastic urothelium (p<0.001). Expression of PD-L1 was detected in 24 (24.2%) samples, and there was no correlation between the expression of Nectin-4 and PD-L1. Patients with strong Nectin-4 expressing tumours had a significantly higher risk of tumour progression (p=0.031) and cancer-specific mortality (p=0.036). Strong Nectin-4 expression was also an independent predictor of tumour progression in the high-risk group (pT3 ≤ or presence of lymphovascular invasion or lymph node metastasis) (Hazard ratio, 3.32 [95% confidence interval, 1.20–7.98; p=0.027]). CONCLUSIONS: We demonstrated that Nectin-4 expression rate in UTUC was 65.7% and independent of PD-L1 expression. Strong expression of Nectin-4 was associated with poor prognosis in UTUC. These findings suggested that enfortumab vedotin may be effective in a broad range of patients with UTUC regardless of the PD-L1 expression level. Source of Funding: None © 2021 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 206Issue Supplement 3September 2021Page: e727-e727 Advertisement Copyright & Permissions© 2021 by American Urological Association Education and Research, Inc.MetricsAuthor Information Kazutoshi Fujita More articles by this author Eisuke Tomiyama More articles by this author Diana Taheri More articles by this author Maria Rodriguez Pena More articles by this author Eri Banno More articles by this author Taigo Kato More articles by this author Koji Hatano More articles by this author Atsunari Kawashima More articles by this author Takeshi Ujike More articles by this author Motohide Uemura More articles by this author Tetsuya Takao More articles by this author Seiji Yamaguchi More articles by this author Hiroaki Fushimi More articles by this author Kazuhiro Yoshimura More articles by this author Hirotsugu Uemura More articles by this author George Netto More articles by this author Norio Nonomura More articles by this author Expand All Advertisement Loading ...

Identification of BXDC2 as a Key Downstream Effector of the Androgen Receptor in Modulating Cisplatin Sensitivity in Bladder Cancer.

Simple SummaryIt remains unclear why chemotherapy is often ineffective in patients with bladder cancer. Meanwhile, we previously reported that male sex hormones (i.e., androgens) could considerably reduce the efficacy of cisplatin, an anti-cancer drug used as the first-line treatment against advanced bladder cancer. The present study aimed to investigate how androgen receptor signaling, which is activated by binding of androgenic hormones, modulates sensitivity to cisplatin treatment in bladder cancer, using cell line models and surgical specimens. We found that the expression levels of the androgen receptor and a molecule (BXDC2) were inversely correlated and that loss of BXDC2 was associated with cisplatin resistance. We thus provide evidence to suggest an underlying molecular mechanism responsible for androgen receptor-induced chemoresistance in bladder cancer.Underlying mechanisms for resistance to cisplatin-based chemotherapy in bladder cancer patients are largely unknown, although androgen receptor (AR) activity, as well as extracellular signal-regulated kinase (ERK) signaling, has been indicated to correlate with chemosensitivity. We also previously showed ERK activation by androgen treatment in AR-positive bladder cancer cells. Because our DNA microarray analysis in control vs. AR-knockdown bladder cancer lines identified BXDC2 as a potential downstream target of AR, we herein assessed its functional role in cisplatin sensitivity, using bladder cancer lines and surgical specimens. BXDC2 protein expression was considerably downregulated in AR-positive or cisplatin-resistant cells. BXDC2-knockdown sublines were significantly more resistant to cisplatin, compared with respective controls. Without cisplatin treatment, BXDC2-knockdown resulted in significant increases/decreases in cell proliferation/apoptosis, respectively. An ERK activator was also found to reduce BXDC2 expression. Immunohistochemistry showed downregulation of BXDC2 expression in tumor (vs. non-neoplastic urothelium), higher grade/stage tumor (vs. lower grade/stage), and AR-positive tumor (vs. AR-negative). Patients with BXDC2-positive/AR-negative muscle-invasive bladder cancer had a significantly lower risk of disease-specific mortality, compared to those with a BXDC2-negative/AR-positive tumor. Additionally, in those undergoing cisplatin-based chemotherapy, BXDC2 positivity alone (p = 0.083) or together with AR negativity (p = 0.047) was associated with favorable response. We identified BXDC2 as a key molecule in enhancing cisplatin sensitivity. AR-ERK activation may thus be associated with chemoresistance via downregulating BXDC2 expression in bladder cancer.

Expression of cancer stem cells markers in urinary bladder urothelial carcinoma and its precursor lesions

Cancer stem cells (CSC) and their role in tumorigenesis of various solid tumors have been studied in past decades. Urothelial CSC were first identified 10 years ago and subsequent studies have been performed with the aim to identify reliable markers of CSC. So far, a few studies have investigated a relationship between CSC markers expression in urothelial carcinoma tissue and histopathological characteristics of the tumor. In our study, we evaluated an immunoexpression of the CSC markers CD24, CD44, CD66 and CD133 in tissue sections of urothelial carcinoma (all tumor grades and stages were included), urothelial carcinoma in situ and non-neoplastic urothelium, totally 218 specimens were enrolled. All studied molecules were expressed either in tumor tissue and non-neoplastic urothelium. Urothelial carcinomas of higher tumor grade and stage expressed molecules CD24 and CD133 significantly more frequently whereas molecules CD44 and CD66 did not show significant association with tumor histopathological features. Our results showed that studied molecules are not suitable for direct detection of CSC in urothelial carcinoma tissue sections, but an expression of molecules CD24 and CD133 is significantly related to urothelial carcinoma grade and stage, which are both important prognostic indicators and therefore an expression of these markers might have a potential prognostic value.

Expression of CD10 in urothelial carcinoma of the bladder and its correlation with histopathological grade, pathological stage, and survival of patients.

CD10 plays a role in signal transduction pathway and regulation of cell growth apoptosis, and therefore, it has been evaluated in different malignancies. The present study was conducted to study the immunoexpression of CD10 in urothelial carcinoma and to correlate it with histological grade, pathological stage, and survival of patients. The study included 51 cases of urothelial carcinoma diagnosed on histopathology along with 50 controls having nonneoplastic urothelium. All the cases and controls were subjected to CD10 immunostaining. The CD10 expression was compared between the cases and controls and was also correlated with histological grade, pathological stage, histomorphological features, and 1-year survival. The study included 78.4% of high-grade urothelial carcinoma (HGUC) and 21.6% of low-grade urothelial carcinoma. The positive score of CD10 expression was observed in 68.6% of cases, while 96% of controls observed negative immunostaining. About 90.9% of low-grade carcinoma observed score 0, while 83.7% of high grade observed positive score of 1 and 2. Although there was statistical significant difference between CD 10 score and stage of tumor, its expression did not correlate with 1-year survival of cases. CD10 expression increases with the grade of tumor and thus may be helpful in differentiating low grade from HGUC. Its expression also increases with stage and poor prognostic factors suggesting its possible role in pathogenesis and progression of urothelial carcinoma. CD 10 may be further analyzed for molecular targeted therapy against urothelial carcinoma.

Mutations in Non-Tumoral Human Urothelium: Disease Prelude or Epilogue?

Bladder cancer is characterized by high rates of recurrence and multifocality, features which have commonly been associated with the colonization of widespread areas of non-neoplastic urothelium by mutant cells, a phenomenon known as field change. Whether mutant fields in the bladder arise from tumor cells or develop from the accumulation of somatic mutations followed by clonal expansions of non-transformed progenitor cells during lifetime remains unanswered. In this issue, Strandgaard et al. perform a deep-sequencing analysis of paired samples of tumor and histologically normal-appearing urothelium from four patients with advanced bladder cancer. By using a careful validation process, they report several mutations exclusive of normal, non-neoplastic tissue, suggesting that multiple fields precede (or develop independently from) the disease. Here, we discuss the main results from this work and elaborate on the biological implications and open questions in the context of normal somatic clonal evolution and cancer risk. We finish providing some general guidelines for future experiments to resolve the role of field changes in bladder carcinogenesis and its possible clinical relevance.

Does the addition of AMACR to CK20 help to diagnose challenging cases of urothelial carcinoma in situ?

BackgroundUrothelial carcinoma in situ (CIS) in the bladder can be difficult to diagnose due to factors including procedural artifact, minimal tissue sampled, therapy-related changes, and various CIS growth patterns. Prior data has demonstrated an increase in alpha-methylacyl-CoA-racemase (AMACR) in urothelial CIS, but there is no information on its utility for diagnosing difficult cases. The aim of this investigation was to assess the expression of AMACR that was ordered on equivocal bladder cases during clinical practice.MethodsTransurethral resections of the bladder in which AMACR and CK20 were performed during diagnostic workup were identified and cases with a final diagnosis of CIS (n = 22) or non-neoplastic urothelium (n = 30) were selected. Additionally, cases in which a diagnosis of CIS was rendered without IHC (n = 20) were selected and tested for AMACR expression.ResultsSensitivity of AMACR for CIS diagnosed with IHC during clinical practice was 73% and specificity was 97%, while CK20 was 95% sensitive and 80% specific. Sensitivity of AMACR in CIS diagnosed without IHC was 100%. In all groups, AMACR had inconsistent intensity, compared to CK20 which had consistent, strong intensity.ConclusionsAMACR was usually positive in urothelial CIS and negative in non-neoplastic urothelium. However, it is important to note that AMACR was less sensitive in difficult cases, while CK20 was more sensitive with more consistent, strong staining compared to AMACR.

Diagnostic utility of Ki-67 immunohistochemistry in small endoscopic biopsies of the ureter and renal pelvis

Diagnosis of upper urinary tract urothelial carcinoma in ureteroscopic biopsies is challenging. Therefore, an immunohistochemical marker that can differentiate between malignant and benign urothelium and predict final pathological features is necessary. In this study, we investigated Ki-67 expression in 26 ureteroscopic biopsies of the ureter and renal pelvis diagnosed with urothelial carcinoma (UC) and in 13 biopsies with non-neoplastic urothelium, using digital image analysis. The median Ki-67 labeling index was 1.5% (range: 0.2–13.9%) in non-neoplastic urothelial specimens and 15.0% (range: 0.2–61.3%) in UC specimens (p=0.0001). In 12 of 26 (46%) UC specimens, the Ki-67 labeling index was more than 20%. By contrast, the Ki-67 labeling index was less than 5% in 11 of 13 (85%) non-neoplastic urothelial specimens. Ki-67 expression in ureteroscopic biopsies was significantly correlated with high tumor grade (p=0.013), concomitant carcinoma in situ (p=0.011), and stromal invasion (p=0.048) in surgical resection specimens. Our data suggested that Ki-67 may provide supplemental, objective evidence that can aid diagnosis of upper urinary tract UC in ureteroscopic biopsy specimens. Determination of Ki-67 expression in ureteroscopic biopsy specimens is potentially helpful in clinical decision making for patients with suspected upper urinary tract UC.

GATA3 immunohistochemistry in urothelial carcinoma of the upper urinary tract as a urothelial marker and a prognosticator

Immunohistochemistry of a transcription factor, GATA3, has been widely used as a promising urothelial marker in diagnostic surgical pathology practice. However, the expression status of GATA3 in upper urinary tract urothelial carcinomas (UUTUCs) and its prognostic significance have not been fully investigated. We immunohistochemically stained for GATA3 in 99 UUTUC samples and paired nonneoplastic urothelial tissues. GATA3 was positive in 51 (51.5%; 32 [32.3%] weak, 11 [11.1%] moderate, 8 [8.1%] strong) of 99 UUTUCs, which was significantly lower than in benign urothelium (79 [96.3%] of 82; 33 [40.2%] weak, 35 [42.7%] moderate, 11 [13.4%] strong; P<.001). However, there were no statistically significant associations between GATA3 expression and tumor grade, pT stage, lymph node involvement, or distant metastasis. Meanwhile, the rate of GATA3 positivity was significantly higher (P=.004) in ureteral tumors (66.0%) than in renal pelvic tumors (35.6%). Kaplan-Meier and log-rank tests revealed that GATA3 negativity was significantly associated with lower recurrence-free survival (P=.037 for all cases, P=.026 for muscle-invasive tumors) and cancer-specific survival (P=.007 for all cases, P=.012 for muscle-invasive tumors, P=.035 for cases with adjuvant chemotherapy) rates. Multivariate analysis further identified strong correlations of GATA3 expression with tumor progression (all cases: hazard ratio [HR], 0.479 [95% confidence interval {CI},0.229-1.003; P=.051]; muscle-invasive tumors: HR, 0.387 [95% CI, 0.166-0.903; P=.028) or cancer-specific mortality (all cases: HR, 0.354 [95% CI, 0.135-0.925; P=.034]; muscle-invasive tumors: HR, 0.402 [95% CI, 0.149-1.086; P=.072]). Thus, compared with nonneoplastic urothelium, a significant decrease in the expression of GATA3 in UUTUC was seen. Moreover, loss of GATA3 expression was found to be an independent predictor of poor patient outcomes. Of note was that only roughly half of high-grade and/or muscle-invasive UUTUCs were immunoreactive for GATA3.

MP71-04 GATA3 IMMUNOHISTOCHEMISTRY IN UROTHELIAL CARCINOMA OF THE UPPER URINARY TRACT AS A UROTHELIAL MARKER AS WELL AS A PROGNOSTICATOR

You have accessJournal of UrologyBladder Cancer: Upper Tract Transitional Cell Carcinoma I1 Apr 2017MP71-04 GATA3 IMMUNOHISTOCHEMISTRY IN UROTHELIAL CARCINOMA OF THE UPPER URINARY TRACT AS A UROTHELIAL MARKER AS WELL AS A PROGNOSTICATOR Satoshi Inoue, Kazutoshi Fujita, Hiroki Ide, Seiji Yamaguchi, Hiroaki Fushimi, George Netto, Norio Nonomura, and Hiroshi Miyamoto Satoshi InoueSatoshi Inoue More articles by this author , Kazutoshi FujitaKazutoshi Fujita More articles by this author , Hiroki IdeHiroki Ide More articles by this author , Seiji YamaguchiSeiji Yamaguchi More articles by this author , Hiroaki FushimiHiroaki Fushimi More articles by this author , George NettoGeorge Netto More articles by this author , Norio NonomuraNorio Nonomura More articles by this author , and Hiroshi MiyamotoHiroshi Miyamoto More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2017.02.2260AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES A transcription factor, GATA3, has been one of the most useful urothelial markers in diagnostic surgical pathology practice. However, previous studies have assessed the utility of GATA3 immunohistochemistry often in bladder specimens. Moreover, the prognostic significance of GATA3 expression in upper urinary tract urothelial carcinomas (UUTUCs) has not been fully investigated. Meanwhile, using cell line and animal models, we have demonstrated that GATA3 functions as a tumor suppressor for urothelial cancer. The current study aims to determine the expression status of GATA3 in UUTUC and its prognostic significance. METHODS We immunohistochemically stained for GATA3 in the tissue microarrays consisting of 99 UUTUC samples and paired non-neoplastic urothelium from each case. We then evaluated the associations between GATA3 expression and clinicopathologic features available for our patient cohort. RESULTS GATA3 was positive in 51 [52%; 32 (32%) weak, 11 (11%) moderate, 8 (8%) strong] of 99 UUTUCs, which was significantly lower than in benign urothelium [79 (96%) of 82; 33 (40%) weak, 35 (43%) moderate, 11 (13%) strong] (P < 0.001). Ten (67%) of 15 low-grade versus 41 (48%) of 84 high-grade UUTUCs (P = 0.266) and 20 (54%) of 37 non-muscle-invasive versus 31 (50%) of 62 muscle-invasive UUTUCs (P = 0.836) were immunoreactive for GATA3. There were also no statistically significant associations between GATA3 expression and pN status, distant metastasis, gender of the patients, or the side of UUTUC. However, the rate of GATA3 positivity was significantly higher (P = 0.004) in ureteral tumors (66%) than in renal pelvic tumors (36%). Kaplan-Meier and log-rank tests revealed that GATA3 negativity significantly correlated with lower recurrence-free survival (P = 0.040 for all cases; P = 0.030 for muscle-invasive tumors) and cancer-specific survival (P = 0.007 for all cases; P = 0.012 for muscle-invasive tumors) rates. Multivariate analysis further identified strong correlations of GATA3 expression with cancer-specific mortality of all cases [hazard ratio (HR) = 0.399, 95% confidence interval (CI) = 0.179-0.890, P = 0.025] or muscle-invasive tumors (HR = 0.374, 95% CI = 0.169-0.831, P = 0.016). CONCLUSIONS Compared with non-neoplastic urothelium, a significant decrease in the expression of GATA3 in UUTUC was seen. Of note was that GATA3 was immunohistochemically detected only in roughly half of high-grade and/or muscle-invasive UUTUCs. In addition, loss of GATA3 expression was found to be an independent predictor of poor patient outcomes. © 2017FiguresReferencesRelatedDetails Volume 197Issue 4SApril 2017Page: e945-e946 Advertisement Copyright & Permissions© 2017MetricsAuthor Information Satoshi Inoue More articles by this author Kazutoshi Fujita More articles by this author Hiroki Ide More articles by this author Seiji Yamaguchi More articles by this author Hiroaki Fushimi More articles by this author George Netto More articles by this author Norio Nonomura More articles by this author Hiroshi Miyamoto More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

MP88-08 ESTROGEN RECEPTOR (ER)-β SIGNALS INDUCE UROTHELIAL TUMORIGENESIS VIA DOWN-REGULATION OF A POTENTIAL TUMOR SUPPRESSOR FORKHEAD BOX PROTEIN O1 (FOXO1)

You have accessJournal of UrologyBladder Cancer: Basic Research & Pathophysiology IV1 Apr 2017MP88-08 ESTROGEN RECEPTOR (ER)-β SIGNALS INDUCE UROTHELIAL TUMORIGENESIS VIA DOWN-REGULATION OF A POTENTIAL TUMOR SUPPRESSOR FORKHEAD BOX PROTEIN O1 (FOXO1) Hiroki Ide, Satoshi Inoue, Kazutoshi Fujita, Yi Li, Takashi Kawahara, Eiji Kashiwagi, Taichi Mizushima, Seiji Yamaguchi, Hiroaki Fushimi, Mototsugu Oya, Norio Nonomura, and Hiroshi Miyamoto Hiroki IdeHiroki Ide More articles by this author , Satoshi InoueSatoshi Inoue More articles by this author , Kazutoshi FujitaKazutoshi Fujita More articles by this author , Yi LiYi Li More articles by this author , Takashi KawaharaTakashi Kawahara More articles by this author , Eiji KashiwagiEiji Kashiwagi More articles by this author , Taichi MizushimaTaichi Mizushima More articles by this author , Seiji YamaguchiSeiji Yamaguchi More articles by this author , Hiroaki FushimiHiroaki Fushimi More articles by this author , Mototsugu OyaMototsugu Oya More articles by this author , Norio NonomuraNorio Nonomura More articles by this author , and Hiroshi MiyamotoHiroshi Miyamoto More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2017.02.2733AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Recent preclinical evidence suggests the involvement of ER signaling in urothelial tumorigenesis, although its underlying mechanisms remain unclear. Meanwhile, cross-talk between FOXO1, a transcriptional factor known to induce apoptosis through the PI3K-Akt pathway, and ERβ has been demonstrated in prostate cancer cells. We therefore investigated the role of FOXO1 in neoplastic transformation of urothelial cells in relation to ERβ signaling. METHODS We immunohistochemically stained for ERβ and phospho-FOXO1 (p-FOXO1), an inactive form of FOXO1, in the tissue microarrays consisting of 99 cases of upper urinary tract urothelial carcinoma and paired non-neoplastic urothelium. Then, in ERα(-)/ERβ(+) human normal urothelial SVHUC sublines stably expressing control- or FOXO1-shRNA with or without exposure to a chemical carcinogen 3-methylcholanthrene (MCA), we assessed the expression of FOXO1 and other known tumor suppressors (via RT-PCR and western blot) as well as neoplastic transformation (via MTT assay and mouse xenograft model). RESULTS ERβ and p-FOXO1 were positive in 63% and 100% of urothelial tumors, which were significantly lower and higher than in non-neoplastic urothelial tissues [85% (P=0.001) and 94% (P=0.018)], respectively. In addition, there was a significant correlation between strong (3+) p-FOXO1 expression and ERβ positivity (P=0.002). In SVHUC cells, estradiol reduced FOXO1 expression, which was abolished by an anti-estrogen tamoxifen. FOXO1 knockdown considerably accelerated neoplastic transformation of MCA-SVHUC cells. Moreover, the expression levels of several genes known to inhibit urothelial carcinogenesis (e.g. p21, p27, UGT1A) were significantly lower in MCA-SVHUC-FOXO1-shRNA than in control cells. Notably, tamoxifen treatment resulted in inhibition of neoplastic transformation of control cells, which was abolished by a FOXO1 inhibitor AS1842856. However, no significant effects of tamoxifen on neoplastic transformation were seen in MCA-SVHUC-FOXO1-shRNA cells. CONCLUSIONS FOXO1 appeared to function as a tumor suppressor and could strongly prevent urothelial tumorigenesis. In contrast, ERβ signals were found to promote it presumably via down-regulation of FOXO1 expression. Accordingly, FOXO1 stimulation via an activator, together with ERβ inactivation via an anti-estrogen, has the potential of being an effective chemopreventive approach for urothelial carcinoma. © 2017FiguresReferencesRelatedDetails Volume 197Issue 4SApril 2017Page: e1177 Advertisement Copyright & Permissions© 2017MetricsAuthor Information Hiroki Ide More articles by this author Satoshi Inoue More articles by this author Kazutoshi Fujita More articles by this author Yi Li More articles by this author Takashi Kawahara More articles by this author Eiji Kashiwagi More articles by this author Taichi Mizushima More articles by this author Seiji Yamaguchi More articles by this author Hiroaki Fushimi More articles by this author Mototsugu Oya More articles by this author Norio Nonomura More articles by this author Hiroshi Miyamoto More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

NFATc1 Expression as a Prognosticator in Urothelial Carcinoma of the Upper Urinary Tract

We recently found that NFATc1, a member of the NFAT family and a key regulator of the immune response, could induce bladder carcinogenesis and cancer progression. In this study, we immunohistochemically stained for NFATc1 in upper urinary tract urothelial carcinoma (UUTUC) specimens and paired nonneoplastic urothelial tissues. NFATc1 was positive in 51 [52%; 40 (40%) weak (1+), 9 (9%) moderate (2+), and 2 (2%) strong (3+)] of 99 UUTUCs, which was significantly higher than in benign urothelium [30 (36%) of 83; 28 (34%) weak and 2 (2%) moderate] (0 vs 1+/2+/3+, P=.038; 0/1+ vs 2+/3+, P=.023). There were no significant associations between NFATc1 expression pattern and tumor grade or pT stage. However, the positive rates of NFATc1 expression tended to be higher in renal pelvic tumors (60%) than in ureteral tumors (42%; P=.080) as well as in pN+ tumors (75%) than in pN0 tumors (49%; P=.089). Kaplan-Meier and log-rank tests revealed that moderate (2+) to strong (3+) NFATc1 expression correlated with lower progression-free survival (P=.032) and cancer-specific survival (P=.005) rates in the 99 cases. Patients with high (2+/3+) NFATc1 muscle-invasive tumor (n=9) also had a significantly higher risk of cancer-specific mortality (P=.021) compared to those with low (0/1+) NFATc1 muscle-invasive tumor (n=53). Thus, compared with nonneoplastic urothelium, a significant increase in the expression of NFATc1 in UUTUC was seen, implying the involvement of NFATc1 signals in the development of UUTUC. The current results further suggest that NFATc1 overexpression serves as a predictor of poor prognosis in patients with UUTUC.

Expression of steroid hormone receptors and its prognostic significance in urothelial carcinoma of the upper urinary tract

ABSTRACTTo assess the expression status of steroid hormone receptors in upper urinary tract urothelial carcinoma (UUTUC), we immunohistochemically stained for androgen receptor (AR), estrogen receptor-α (ERα), ERβ, glucocorticoid receptor (GR), and progesterone receptor (PR) in 99 UUTUC specimens and paired non-neoplastic urothelial tissues. AR/ERα/ERβ/GR/PR was positive in 20%/18%/62%/63%/16% of tumors, which was significantly lower (except PR) than in benign urothelial tissues [57% (P < 0.001)/40% (P = 0.001)/85% (P = 0.001)/84% (P = 0.002)/13% (P = 0.489)]. There were no significant associations between each receptor expression pattern and histopathological characteristic of the tumors including tumor grade/stage. Kaplan-Meier and log-rank tests revealed no significant prognostic value of each receptor expression in these 99 patients. However, patients with UUTUC positive for either ERα or PR had a significantly higher risk of disease-specific mortality (P = 0.025), compared with those with UUTUC negative for both. PR positivity alone in pT3 or pT4 tumors was also strongly associated with the risk of disease-specific mortality (P = 0.040). Multivariate analysis further identified the expression of ERα and/or PR as a strong predictor for disease-specific mortality in the entire cohort of the patients (hazard ratio, 2.434; P = 0.037). Thus, in accordance with previous observations in bladder specimens, significant decreases in the expression of AR/ERα/ERβ/GR in UUTUC, compared with that in non-neoplastic urothelium, were observed. Meanwhile, the negativity of both ERα and PR in UUTUC as well as the negativity of PR alone in deeply invasive tumor was suggested to serve as a prognosticator.

ELK1 is up-regulated by androgen in bladder cancer cells and promotes tumor progression.

Little is known about biological significance of ELK1, a transcriptional factor that activates downstream targets including c-fos proto-oncogene, in bladder cancer. Recent preclinical evidence also suggests the involvement of androgen receptor (AR) signaling in bladder cancer progression. In this study, we aim to investigate the functions of ELK1 in bladder cancer growth and their regulation by AR signals. Immunohistochemistry in bladder tumor specimens showed that the levels of phospho-ELK1 (p-ELK1) expression were significantly elevated in urothelial neoplasms, compared with non-neoplastic urothelium tissues, and were also correlated with AR positivity. Patients with p-ELK1-positive non-muscle-invasive and muscle-invasive tumors had significantly higher risks for tumor recurrence and progression, respectively. In AR-positive bladder cancer cell lines, dihydrotestosterone treatment increased ELK1 expression (mRNA, protein) and its nuclear translocation, ELK1 transcriptional activity, and c-fos expression, which was restored by an anti-androgen hydroxyflutamide. ELK1 silencing via short hairpin RNA (shRNA) resulted in decreases in cell viability/colony formation, and cell migration/invasion as well as an increase in apoptosis. Importantly, ELK1 appears to require activated AR to regulate bladder cancer cell proliferation, but not cell migration. Androgen also failed to significantly induce AR transactivation in ELK1-knockdown cells. In accordance with our in vitro findings, ELK1-shRNA expression considerably retarded tumor formation as well as its growth in xenograft-bearing male mice. Our results suggest that ELK1 plays an important role in bladder tumorigenesis and cancer progression, which is further induced by AR activation. Accordingly, ELK1 inhibition, together with AR inactivation, has the potential of being a therapeutic approach for bladder cancer.

Expression of Glypican 3 in low and high grade urothelial carcinomas.

BackgroundGlypican-3 (GPC3) is an oncofetal protein which is encoded by GPC3 gene and takes role in the regulation of cell division and apoptosis. Overexpression of GPC3 has been reported in some types of cancer such as hepatocellular carcinoma (HCC), melanoma, squamous cell carcinoma of the lungs and testicular germ cell tumors. The aim of this study was to investigate the immunohistochemical expression of GPC3 in the non-neoplastic urothelium and in urothelial carcinoma (UC). We also aimed to explore the alterations in the GPC3 expression according to the grade and the invasiveness of UC.MethodsGPC3 expression was studied in 108 UC cases by using immunohistochemistry. Each section was evaluated in terms of the extensiveness and intensity of GPC3 staining. Scores of immunostaining were correlated with tumor grade and stage.ResultsGPC3 expression was observed in 38 cases (35.2%). GPC3 expression was positive in 43.6% of high and in 13.3% of low grade UC (p: 0.003). In 19 UC cases biopsy also harbored non-neoplastic urothelium which showed no staining for GPC3. The difference in staining percentages between low and high grade UCs, suggests that GPC3 staining could be used as an adjunctive marker in cases where the distinction between the low and high grade tumors is difficult. In addition, lack of staining in the non-neoplastic urothelial areas in 19 cases raises the possibility of the use of GPC3 staining for the distinction between neoplastic and non-neoplastic urothelium, especially in punch biopsy samples.ConclusionsBased on our results potential role of GPC3 in urothelial carcinogenesis warrants further investigation, especially the potential use of GPC3 for therapeutic and diagnostic purposes.Virtual SlidesThe virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/2260833001522844

MP7-14 EXPRESSION OF STEROID HORMONE RECEPTORS AND ITS PROGNOSTIC SIGNIFICANCE IN UROTHELIAL CARCINOMA OF THE UPPER URINARY TRACT (UCUUT)

You have accessJournal of UrologyBladder Cancer: Upper Tract TCC II1 Apr 2015MP7-14 EXPRESSION OF STEROID HORMONE RECEPTORS AND ITS PROGNOSTIC SIGNIFICANCE IN UROTHELIAL CARCINOMA OF THE UPPER URINARY TRACT (UCUUT) Eiji Kashiwagi, Kazutoshi Fujita, Seiji Yamaguchi, Hiroaki Fushimi, Leonardo Reis, George Netto, Norio Nonomura, and Hiroshi Miyamoto Eiji KashiwagiEiji Kashiwagi More articles by this author , Kazutoshi FujitaKazutoshi Fujita More articles by this author , Seiji YamaguchiSeiji Yamaguchi More articles by this author , Hiroaki FushimiHiroaki Fushimi More articles by this author , Leonardo ReisLeonardo Reis More articles by this author , George NettoGeorge Netto More articles by this author , Norio NonomuraNorio Nonomura More articles by this author , and Hiroshi MiyamotoHiroshi Miyamoto More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2015.02.221AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Emerging evidence suggests that the steroid hormone receptor signaling pathways play an important role in the outgrowth of urothelial carcinoma. Although recent studies have assessed the expression of androgen receptor (AR), estrogen receptors (ERs), glucocorticoid receptor (GR), and/or progesterone receptor (PR) in bladder tumor specimens, their status in UCUUT remains largely unknown. METHODS We immunohistochemically stained for AR, ERα, ERβ, GR, and PR in the tissue microarrays consisting of 99 UCUUT samples and paired non-neoplastic urothelium from each case. We then analyzed the associations between their expression and clinicopathologic features available for our patient cohort. RESULTS AR/ERα/ERβ/GR/PR was positive in 20%/18%/63%/63%/16% of tumors, which was significantly lower (except PR) than in benign urothelial tissues [58% (P<0.001)/41% (P<0.001)/85% (P<0.001)/84% (P=0.001)/13% (P=0.529)]. AR expression (11% vs. 28%; P=0.045) and ERβ expression (51% vs. 68%; P=0.056) were significantly and marginally, respectively, down-regulated in renal pelvic tumors, compared with ureteral tumors, whereas ERα expression (22% vs. 12%; P=0.2723), GR expression (58% vs. 68%; P=0.395), and PR expression (9% vs. 20%; P=0.155) were similar between the two groups. AR/PR expression also tended to be higher (P=0.072)/lower (P=0.052) in male tumors (27%/10%) than in female tumors (10%/26%), while no significant difference in the positivity of ERα (17% vs. 21%; P=0.790), ERβ (58% vs. 69%; P=0.296), or GR (67% vs. 56%; P=0.395) between genders was seen. In addition, there were no significant associations between expression pattern of each receptor and tumor grade or stage (pT/pN). Kaplan-Meier and log-rank tests further revealed no significant prognostic value of AR, ERα, ERβ, GR, or PR expression in these 99 patients. However, ERα (P=0.040) or PR (P=0.040) positivity in 54 patients with pT3 or pT4 tumor was strongly associated with disease-specific mortality. CONCLUSIONS In accordance with previous observations in bladder specimens, significant decreases in the expression of AR, ERα, ERβ, or GR in UCUUT were observed, compared with non-neoplastic urothelium. In contrast, no strong correlations of each receptor expression with tumor grade or stage were found. Nonetheless, our results further support the involvement of AR/ERα/ERβ/GR/PR signals in the development and progression of UCUUT. © 2015 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 193Issue 4SApril 2015Page: e69 Advertisement Copyright & Permissions© 2015 by American Urological Association Education and Research, Inc.MetricsAuthor Information Eiji Kashiwagi More articles by this author Kazutoshi Fujita More articles by this author Seiji Yamaguchi More articles by this author Hiroaki Fushimi More articles by this author Leonardo Reis More articles by this author George Netto More articles by this author Norio Nonomura More articles by this author Hiroshi Miyamoto More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Endoglin expression in upper urinary tract urothelial carcinoma is associated with intravesical recurrence after radical nephroureterectomy

To evaluate the expression and prognostic significance of endoglin in patients with upper urinary tract urothelial carcinoma. zArchival formalin-fixed and paraffin-embedded tissues from 99 cases of primary upper urinary tract urothelial carcinomas treated with nephroureterectomy were retrieved. Tissue microarrays were constructed with triplicate tumor samples and paired non-neoplastic urothelium. Tissue microarrays were analyzed using immunohistochemistry for endoglin, and the associations between clinicopathological parameters and outcome were studied. Endoglin expression was significantly higher in the endothelium of upper urinary tract urothelial carcinomas than in paired benign urothelium (P < 0.001). Endoglin expression was not associated with pathological T stage or tumor grade, and it was not associated with increased hazard ratios for cancer-specific mortality, tumor recurrence in the lymph node or distant metastasis. However, expression of endoglin was significantly associated with intravesical recurrence, when adjusting for other relevant clinicopathological variables (P = 0.015). Endoglin is overexpressed in the endothelium of upper urinary tract urothelial carcinomas when compared with normal urothelium, and this overexpression seems to be associated with a higher risk of intravesical recurrence. Therefore, endoglin could be a biomarker for the prediction of intravesical recurrence, as well as a potential therapeutic target.

Reduced glucocorticoid receptor expression predicts bladder tumor recurrence and progression.

To assess the levels of glucocorticoid receptor (GR) expression in bladder tumors because the status and its prognostic value remain largely unknown. We immunohistochemically stained for GR in bladder tumor and matched non-neoplastic bladder tissue specimens. Overall, GR was positive in 129 (87%) of 149 urothelial tumors, which was significantly (P=.026) lower than in non-neoplastic urothelium (90 [96%] of 94). Forty-two (79%) of 53 low-grade tumors vs 45 (47%) of 96 high-grade carcinomas (P<.001) and 61 (73%) of 84 non-muscle-invasive (NMI) tumors vs 26 (40%) of 65 muscle-invasive (MI) carcinomas (P<.001) were moderately to strongly immunoreactive for GR. Kaplan-Meier and log-rank tests revealed that loss or weak positivity of GR significantly or marginally correlated with recurrence of NMI tumors (P=.025), progression of MI tumors (P=.082), and cancer-specific survival of MI tumors (P=.067). Multivariate analysis identified low GR expression as a strong predictor for recurrence of NMI tumors (P=.034). GR expression was downregulated in bladder tumors compared with nonneoplastic bladder tumors and in high-grade/MI tumors compared with low-grade/NMI tumors. Decreased expression of GR, as an independent prognosticator, predicted recurrence of NMI tumors. These results support experimental evidence suggesting an inhibitory role of GR signals in bladder cancer outgrowth.