Expression In Urothelial Carcinoma Research Articles

Preliminary data on computed tomography-based radiomics for predicting programmed death ligand 1 expression in urothelial carcinoma

Background: Programmed death ligand 1 (PD-L1) expression cannot currently be predicted through radiological findings. This study aimed to develop a prediction model capable of differentiating between positive and negative PD-L1 expression through a radiomics-based investigation of computed tomography (CT) images in patients with urothelial carcinoma.Methods: Sixty-four patients with urothelial carcinoma who underwent immunohistochemical testing for PD-L1 were retrospectively reviewed. The number of patients in the positive and negative PD-L1 groups (PD-L1 expression >5%) was 14 and 50, respectively. CT images obtained 90 seconds after contrast medium administration were selected for radiomic extraction. For all tumors, 1,691 radiomic features were extracted from CT using a manually segmented three-dimensional volume of interest. Univariate and multivariate logistic regression analyses were performed to identify radiomic features that were significant predictors of PD-L1 expression. For the radiomics-based model, a receiver operating characteristic (ROC) analysis was performed. Results: Among 64 patients, 14 were included in the PD-L1 positive group. Logistic regression analysis found that the following radiomic features significantly predicted PD-L1 expression: wavelet-low-pass, low-pass, and high-pass filters (LLH)_gray-level size-zone matrix (GLSZM)_SmallAreaEmphasis, wavelet-LLH_firstorder_Energy, log-sigma-0-5-mm-3D_GLSZM_SmallAreaHighGrayLevelEmphasis, original_shape_Maximum2DDiameterColumn, wavelet-low-pass, low-pass, and low-pass filters (LLL)_gray-level run-length matrix (GLRLM)_ShortRunEmphasis, and exponential_firstorder_Kurtosis. The radiomics signature was –4.0934+21.6224 (wavelet-LLH_GLSZM_SmallAreaEmphasis)+0.0044 (wavelet-LLH_firstorder_Energy)–4.7389 (log-sigma-0-5-mm-3D_GLSZM_SmallAreaHighGrayLevelEmphasis)+0.0573 (original_shape_Maximum2DDiameterColumn)–29.5892 (wavelet-LLL_GLRLM_ShortRunEmphasis)–0.4324 (exponential_firstorder_Kurtosis). The area under the ROC curve model representing the radiomics signature for differentiating cases that were deemed PD-L1 positive based on immunohistochemistry was 0.96. Conclusions: This preliminary radiomics model derived from contrast-enhanced CT predicted PD-L1 positivity in patients with urothelial cancer.

An artificial intelligence-powered PD-L1 combined positive score (CPS) analyser in urothelial carcinoma alleviating interobserver and intersite variability.

Immune checkpoint inhibitors targeting programmed death-ligand 1 (PD-L1) have shown promising clinical outcomes in urothelial carcinoma (UC). The combined positive score (CPS) quantifies PD-L1 22C3 expression in UC, but it can vary between pathologists due to the consideration of both immune and tumour cell positivity. An artificial intelligence (AI)-powered PD-L1 CPS analyser was developed using 1,275,907 cells and 6175.42 mm2 of tissue annotated by pathologists, extracted from 400 PD-L1 22C3-stained whole slide images of UC. We validated the AI model on 543 UC PD-L1 22C3 cases collected from three institutions. There were 446 cases (82.1%) where the CPS results (CPS ≥10 or <10) were in complete agreement between three pathologists, and 486 cases (89.5%) where the AI-powered CPS results matched the consensus of two or more pathologists. In the pathologist's assessment of the CPS, statistically significant differences were noted depending on the source hospital (P = 0.003). Three pathologists reevaluated discrepancy cases with AI-powered CPS results. After using the AI as a guide and revising, the complete agreement increased to 93.9%. The AI model contributed to improving the concordance between pathologists across various factors including hospital, specimen type, pathologic T stage, histologic subtypes, and dominant PD-L1-positive cell type. In the revised results, the evaluation discordance among slides from different hospitals was mitigated. This study suggests that AI models can help pathologists to reduce discrepancies between pathologists in quantifying immunohistochemistry including PD-L1 22C3 CPS, especially when evaluating data from different institutions, such as in a telepathology setting.

Scoring PD-L1 Expression in Urothelial Carcinoma: An International Multi-Institutional Study on Comparison of Manual and Artificial Intelligence Measurement Model (AIM-PD-L1) Pathology Assessments.

Assessing programmed death ligand 1 (PD-L1) expression on tumor cells (TCs) using Food and Drug Administration-approved, validated immunoassays can guide the use of immune checkpoint inhibitor (ICI) therapy in cancer treatment. However, substantial interobserver variability has been reported using these immunoassays. Artificial intelligence (AI) has the potential to accurately measure biomarker expression in tissue samples, but its reliability and comparability to standard manual scoring remain to be evaluated. This multinational study sought to compare the %TC scoring of PD-L1 expression in advanced urothelial carcinoma, assessed by either an AI Measurement Model (AIM-PD-L1) or expert pathologists. The concordance among pathologists and between pathologists and AIM-PD-L1 was determined. The positivity rate of ≥ 1%TC PD-L1 was between 20-30% for 8/10 pathologists, and the degree of agreement and scoring distribution for among pathologists and between pathologists and AIM-PD-L1 was similar both scored as a continuous variable or using the pre-defined cutoff. Numerically higher score variation was observed with the 22C3 assay than with the 28-8 assay. A 2-h training module on the 28-8 assay did not significantly impact manual assessment. Cases exhibiting significantly higher variability in the assessment of PD-L1 expression (mean absolute deviation > 10) were found to have patterns of PD-L1 staining that were more challenging to interpret. An improved understanding of sources of manual scoring variability can be applied to PD-L1 expression analysis in the clinical setting. In the future, the application of AI algorithms could serve as a valuable reference guide for pathologists while scoring PD-L1.

Abstract 5131: TYMS overexpression - but not copy number alteration - is associated with cancer aggressiveness in urothelial bladder cancer

Abstract Background: Thymidylate Synthetase (TYMS) is an import enzyme in the early stage of DNA biosynthesis. It is coded by the TYMS gene at 18p11.32, a region which has been found amplified in urothelial carcinoma. TYMS is a target enzyme for 5-fluorouracil (5-FU) and methotrexate. Elevated TYMS expression has been associated with poor prognosis and unfavorable therapy response in different cancer types. The aim of this study was to clarify the clinical relevance of TMYS expression in urothelial carcinoma and to determine the relationship between TYMS expression and gene amplification. Design: TYMS expression was analyzed by immunohistochemistry and TYMS copy number status was determined by fluorescence in-situ hybridization on a tissue microarray (TMA) containing more than 2,700 urothelial carcinomas of the urinary bladder including &amp;gt;600 patients who had undergone cystectomy for muscle-invasive disease and for which follow-up data were available. Results: TMYS immunostaining was detectable in 83.1% of 1,799 analyzable bladder cancers, including 56.4% with weak, 14.4% with moderate, and 12.3% with strong staining. The rate of strongly TMYS positive cancers increased markedly from pTaG2 low (0.3%) to pTaG2 high (4.1%), and pTaG3 cancers (11.8%; p&amp;lt;0.0001) and was even somewhat higher in pT2-4 cancers (16.7%; p=0.6401; pTaG3 vs pT2-4). In muscle-invasive cancers, the frequency of strong TMYS immunostaining increased with tumor stage (p=0.0011) and grade (p=0.0007) but was lower in lymph node positive cancers (p=0.0018). No association was found between TYMS expression and patient prognosis. TYMS amplification was found in 3.1% and TMYS deletion in 1.2% of 1,775 analyzable bladder cancers. The rate of TYMS amplification increased from pTa (0.6%) to pT2-4 (3.7%; p&amp;lt;0.0001) cancers whereas TYMS deletion was only found in pT2-4 (1.4%) cancers. In muscle invasive-cancers, TYMS copy number alterations were unrelated to tumor phenotype and patient prognosis. Strong TYMS positivity was significantly associated with TYMS amplification (p=0.0096) but only 28.6% of 42 amplified cancers showed a strong TMYS staining and only 7.2% of strongly TYMS positive cancers had a TYMS amplification. Conclusion: The significant association between TYMS overexpression and parameters of aggressive phenotype are consistent with a role of TYMS in bladder cancer progression. The incomplete association between TYMS amplification and overexpression demonstrates that TYMS is not the critical target of 18p11 amplification in bladder cancer and that TYMS upregulation is in most cases due to other mechanisms than amplification. Citation Format: Henning Plage, Sebastian Hofbauer, Kira Furlano, Sarah Weinberger, Antonia Franz, Annika Fendler, Florian Roßner, Simon Schallenberg, Sefer Elezkurtaj, Adrian Frericks, Maximilian Lennartz, Andreas H. Marx, Henrik Samtleben, Margit Fisch, Michael Rink, Marcin Slojewski, Krystian Kaczmarek, Thorsten Ecke, Stefan Koch, Nico Adamini, Ronald Simon, Guido Sauter, Henrik Zecha, Joachim Weischenfeldt, Tobias Klatte, Sarah Minner, David Horst, Thorsten Schlomm, Martina Kluth. TYMS overexpression - but not copy number alteration - is associated with cancer aggressiveness in urothelial bladder cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5131.

Expression of HER2 in high-grade urothelial carcinoma based on Chinese expert consensus and the clinical effects of disitamab vedotin-tislelizumab combination therapy in the treatment of advanced patients.

Background: A vast number of researchers have discovered high levels of human epidermal growth factor receptor-2 (HER2) expression in urothelial carcinoma (UC), but they do not use a uniform scoring system. Based on the 2021 edition of clinical pathological expert consensus on HER-2 testing in UC in China, we investigated the expression level and clinical significance of HER2 in high-grade UC. Furthermore, we looked at the prognosis of patients with locally advanced/metastatic UC after combining HER2 targeting antibody-drug conjugates (ADC) medication disitamab vedotin (DV) with programmed cell death protein 1 (PD-1) inhibitor tislelizumab. Patients and methods: From 2019 to 2022, we collected paraffin specimens of UC from the Department of Urology at the Provincial Hospital Affiliated to Shandong First Medical University. HER2 expression-related factors were investigated. Patients with advanced UC who have failed systemic chemotherapy at least once and had received immune checkpoint inhibitor (ICI) medication during second-line treatment were selected and treated with DV in combination with tislelizumab. We assessed the therapy's efficacy and safety. Results: 185 patients with high-grade UC were included in this investigation. 127 patients (68.7%) were HER2 positive (IHC 2+/3+) according to the 2021 Clinical pathological expert consensus on HER2 testing in UC in China. The clinical stage of UC differed statistically significantly between the HER2-and HER2+ groups (p = 0.019). Sixteen advanced UC patients were treated with DV and tislelizumab for a median of 14months. The disease control rate was 87.5%, while the objective response rate (ORR) was 62.5%. The ORR of HER2+ individuals was higher than that of HER2-individuals (70.0% vs. 50.0%). The median progression-free survival or overall survival was not reached. In this study, the incidence of treatment-related adverse events was 68.8% (11/16), with all of them being grade 1 or 2 adverse reactions. Conclusion: HER2 protein expressed at a high percentage in UC, and 68.7% patients expressed HER2 positive (IHC 2+/3+). HER2+ expression is positively correlated with higher clinical stage of UC. HER2 targeted ADC drug disitamab vedotin combining with PD-1 inhibitor tislelizumab has shown efficacy, safety and controllable adverse reactions in the treatment of advanced UC.

Evaluation of GATA3 and p53 Expression in Urothelial Carcinoma and their Association with Clinicopathological Parameters: A Cross-sectional Study

Introduction: Bladder cancer is the tenth most common cancer worldwide, with 90% of bladder cancers are Urothelial Carcinoma (UC). In India, its incidence is 1.6 per lakh population annually. GATA3 is a recently used immunohistochemical marker which is highly sensitive for UC. Alterations in the p53 pathway play an important role in tumour progression and the assessment of prognosis. Aim: To study the expression of GATA3 and p53 in UC and their association with clinicopathological parameters. Materials and Methods: An ambispective cross-sectional study was conducted in the Department of Pathology, Kalinga Institute of Medical Sciences, Bhubaneswar, Odisha, India, from September 2020 to August 2022. A total of 50 cases of histologically proven UC were subjected to immunohistochemistry with GATA3 and p53. The expression of GATA3 and p53 was evaluated in UC, and their association with clinicopathological parameters like age, gender, personal habits, tumour site, symptoms and signs, histological grade, and invasion into lamina propria and muscle was studied. The Chi-square test was used to determine the relationship between all parameters, with a p-value &lt;0.05 considered statistically significant. Results were analysed by standard software (Stata version 23.1). Results: GATA3 expression was observed in 94% (47/50) of cases of UC. GATA3 expression was associated with histological grade and muscle invasion, with 66.7% (4/6) expression in high-grade muscle-invasive UC, compared to 91.30% (21/23) in non muscle invasive UC, and 100% (21/21) in low-grade UC. Additionally, 66.7% (14/21) of low-grade UC showed p53 expression, whereas 80% (24/30) of high-grade UC demonstrated p53 expression. Conclusion: GATA3 is a sensitive and specific marker for UC. Its expression is highly variable among different histological types of UC and decreases with increasing grade of tumour in the study. p53 expression was associated with the histological grade of the tumour and was statistically significant in this study. GATA3 can be utilised to predict the probable grade of the tumour and its behaviour by correlating it with p53.

Expression and clinicopathologic significance of HER2 and PD-L1 in high grade urothelial carcinoma of the urinary tract.

Urothelial carcinoma (UC) is an aggressive tumor with high recurrence rates and poses a great challenge for clinical management. Programmed death ligand-1 (PD-L1) inhibitors and human epidermal growth factor receptor 2 (HER2) blockers have been approved for the treatment of advanced urothelial carcinoma. PD-L1 and HER2 expression in UC will determine whether patients are likely to respond to these targeted treatments. This study assessed the expressions of HER2 and PD-L1 in UC at our institution and investigated their correlations with gender, tumor location (upper genitourinary (GU) tract vs. lower GU tract), tumor stage, and histologic divergent subtypes. Patients with UC who had PD-L1 or HER2 immunostains performed in the past 3 years at our institution were included in our analysis. A total of 97 cases were identified. PD-L1 and HER2 scores were provided by two experienced GU pathologists. HER2 scores were given according to the criteria used in breast cancer, while PD-L1 scores were reported as the combined positive score. We assessed correlation of the scores with the patients' gender, tumor location, tumor stage, and histologic divergent subtypes. The data for PD-L1 expression were analyzed using the Mann-Whitney U Test for gender and urinary tract location, and one-way analysis of variance (ANOVA) for stage and histology. The data for HER2 expression were analyzed using the chi-square test. For all analyses, significance was set at P<0.05. Of the 97 patients, the average age was 69 years. There were 95 patients who had previously reported HER2 results and 86 patients who had PD-L1 results. PD-L1 expression did not show a significant difference among the histological divergent subtypes (P=0.36). However, HER2 status exhibited a significant difference, with more HER2-positive cases observed in the conventional histology (P=0.008). No correlation between HER2 status and either gender or tumor stage was identified. The median PD-L1 combined positive score was significantly higher in lower urinary tract UC than upper (10 and 2, respectively; P=0.049). No significant differences were observed for gender or pathologic stage. Our data suggest that HER2 is more frequently expressed in conventional UC than in divergent subtypes. Additionally, PD-L1 has a higher expression level in lower urinary tract UC compared to upper. However, PD-L1 and HER2 expression are not related to gender or tumor stage in UC.

An iodine‑labelled Antibody–drug conjugate PET probe for noninvasive monitoring of Nectin-4 expression in urothelial carcinoma

Background and PurposeSome kinds of antibody–drug conjugate (ADC) with high affinity to Nectin-4 have demonstrated breakthrough progress in the third-line setting for bladder cancer. However, many patients are still difficult to benefit from treatment based on the heterogeneity of tumour. As the most advanced auxiliary treatment technology, treatment visualization can most intuitively predict the effectiveness of drug treatment, and timely detect the occurrence of drug resistance. Among them, nuclear medicine molecular probes play an important role in this field. Methods124/125I-EV was prepared by labelling Enfortumad Vedetin (EV), an ADC drugs widely used in clinic targeted Nectin-4, with Na124/125I using N-bromine succinimide as oxidant. The radiochemical purity was analyzed via radio-TLC and bioactivity was measured by enzyme-linked immunosorbent assay. Cell uptake assay and small-animal PET imaging were performed to verified the specificity and targeting. Key Results124/125I-EV was prepared with high labeling yield and radiochemical purity. ELISA assays demonstrated that 124I-EV maintained the same high bioactivity as EV with significantly higher uptake in SW780 cells (Nectin-4 positive, 4.05 ± 0.32 %IA/5 × 105 cells at 8 h) than that in T24 cells (Nectin-4 negative, 1.34 ± 0.18 %IA/5 × 105 cells, p < 0.001). In PET imaging, 124I-EV had a significantly higher accumulation in SW780 tumour than that in T24 tumour and the uptake in SW780 tumour could be specifically blocked when co-injected with cold EV. The signal-to-noise ratio at the tumour site gradually increased with time, and peaked at 72 h. Conclusion and Implications124I-EV was successfully prepared with high specificity and binding affinity of Nectin-4. This radioactive probe completely simulates the internal circulation of ADC drugs and tumour uptake and retention, which will greatly improve the clinical application of ADC therapy.

Expression of Preferentially Expressed Antigen in Melanoma, a Cancer/Testis Antigen, in Carcinoma In Situ of the Urinary Tract.

Carcinoma in situ (CIS) of the urinary tract comprises 1-3% of all urothelial malignancies and is often a precursor to muscle-invasive urothelial carcinoma (UC). This study aimed to examine the expression profiles of preferentially expressed antigen in melanoma (PRAME), a cancer/testis antigen, and assess its diagnostic and therapeutic applications in CIS, given that its expression in UC has been minimally studied and has not yet been analyzed in CIS. We selected consecutive patients with CIS who underwent biopsy and/or transurethral tumor resection at the Osaka Medical and Pharmaceutical University Hospital. Immunohistochemical staining for PRAME and p53 was performed. Overall, 53 patients with CIS (6 females and 47 males) were included. Notably, PRAME expression was observed in 23 of the 53 patients (43.4%), whereas it was absent in the non-neoplastic urothelial epithelium. Furthermore, no correlation was found between PRAME expression and aberrant p53 expression. Therefore, PRAME expression may serve as a useful marker for CIS of the urinary tract. Furthermore, PRAME may be a candidate for the novel therapeutic target for standard treatment-refractory CIS patients.

Downregulation of CRTAC1 in Urothelial Carcinoma Promotes Tumor Aggressiveness and Confers Poor Prognosis.

Cartilage acidic protein 1 (CRTAC1) is a glycosylated calcium-binding extracellular matrix protein. The oncological functions of CRTAC1 in urothelial carcinoma (UC) of the urinary bladder (UB) and upper urinary tract (UT) have not yet been elucidated. Based on the published UBUC transcriptome data, we re-evaluated the differential expression profile of calcium ion binding-related genes (GO:0005509), and we found that CRTAC1 was the most significantly downregulated gene in UBUC progression. Therefore, we analyzed the prognostic value and biological significance of CRTAC1 expression in UC. We used immunohistochemistry to determine the CRTAC1 expression levels in 340 patients with UTUC and 295 patients with UBUC. The CRTAC1 expression was compared with the clinicopathological characteristics, and the prognostic impact of CRTAC1 on metastasis-free survival (MFS) and disease-specific survival (DSS) was evaluated. To study the biological functions of CRTAC1, the proliferation, migration, invasion, and tube formation abilities of UC-derived cells were evaluated. A low CRTAC1 expression significantly correlated with high tumor stage, high histological grade, perineural invasion, vascular invasion, nodal metastasis, and high mitotic rate (all p < 0.01). Moreover, the CRTAC1 immunoexpression status was an independent prognostic factor for MFS and DSS in UBUC and UTUC patients (all p < 0.001) in the multivariate analysis. The exogenous expression of CRTAC1 suppressed the cell proliferation, invasion, and angiogenesis, and downregulated the matrix metallopeptidase 2 (MMP2) level in BFTC909 and T24 cells. CRTAC1 may participate in progression of UC and serve as a prognostic marker for metastasis. Low CRTAC1 expression was significantly associated with aggressive UC characteristics and worse clinical outcomes. The inclusion of CRTAC1 immunoexpression in the standard pathological variables may optimize the risk stratification of patients.

Elucidating the Associated Biological Function and Clinical Significance of RHOJ Expression in Urothelial Carcinoma.

Urothelial cancer, a common urinary system malignancy, often presents treatment challenges due to metastasis and chemotherapy side effects. Angiogenesis, crucial for tumor growth, has become a target for drug development. This study explores the expression, prognostic value, and clinical correlation of RHOJ in the TCGA BLCA, GSE31684, and GSE32894 datasets. We identify common differentially expressed genes across these databases and utilize g:Profiler and Cytoscape ClueGO for functional assessment. Further, we perform a gene set enrichment analysis (GSEA) using Hallmark gene sets and use the imsig package for immune cell infiltration analysis. Our analysis indicates that RHOJ expression levels significantly impact survival rates, tumor progression, and immune response in urothelial tumors. High RHOJ expression correlated with poor prognosis, advanced disease stages, and an increase in monocyte population within the tumor microenvironment. This aligns with current literature indicating a key role of immune infiltration in bladder cancer progression and treatment response. Moreover, the GSEA and imsig results further suggest a potential mechanistic link between RHOJ expression and immune-related pathways. Considering the increasing emphasis on immunotherapeutic strategies in bladder cancer management, our findings on RHOJ's potential as a diagnostic biomarker and its association with immune response open new avenues for therapeutic interventions.

Immunohistochemical Expression of Clustered Differentiation 10 (CD10) Across Various Grade &amp; Stage of Urinary BladderCarcinoma

Objective: To assess the Clustered Differentiation 10 (CD10) expression in Urothelial carcinoma and to ascertain how its expression relates to grade and stage. Method: Descriptive cross sectional research was carried out over a period of six months in Pathology Department on eighty five cases of bladder carcinoma from transurethral resection of bladder (TURBT) specimens diagnosed on Hematoxylin-eosin (H &amp; E) stained sections irrespective of patient's age and gender were included in the study. These were stained for CD10 by Immunohistochemical technique. Results: Sixty seven (78.8%) urothelial carcinomas showed positive CD10 staining while 18 cases (21.17%) demonstrated negative expression. In 67 positive cases, 42 high grade tumors had 2+ expression while 13 grade had 1+ staining .All low-grade tumors(12) displayed 1+ score. Sixteen tumorsin pT1, 21 tumorsin pT2 and 3 tumors in pT3 stage displayed 2+ score. Nineteen tumors in pT1 while 4 tumors in pT2 had 1+ score. Conclusion: Our findings indicate that CD10 expression is greater in high grade and invasive urothelial carcinomas and is associated with progression of bladder carcinomas.

Role of GATA-3 Expression in Urothelial Carcinoma and ITS Correlation with p53 by Immunohistochemistry

Background and Objectives: Urothelial carcinomas are known for divergent differentiation, which often poses diagnostic challenges. The use of immunohistochemistry (IHC) markers helps in differentiating these entities and provides insight into tumor pathogenesis. This study is done with the objective of evaluating significant clinicopathological and histomorphological features of urothelial carcinoma and correlating with GATA-3 and P53 staining. Materials and Methods: A total of 59 cases of urothelial tumors were included in this prospective study. Tissue samples were processed, and H and E-stained slides were evaluated and reported as per the WHO/International Society of Urologic Pathology Classification of bladder tumor 2016. IHC for GATA-3 and p53 was performed using respective monoclonal primary antibodies for GATA-3 and p53. Statistical Analysis: The correlation between GATA-3 and other classical clinicopathological parameters was studied by Chi-square test. P &lt;0.05 was considered statistically significant. Results: In the present study, out of 59 cases, there was a male preponderance with an M: F ratio of 7.4:1 and an age range of 25–85 years. High-grade urothelial carcinoma was the most common histologic subtype. GATA-3 expression was seen in 93.2% of cases. There was a statistically significant correlation of GATA-3 expression with hematuria and dysuria. GATA-3 expression was compared with significant histopathological parameters and a significant correlation of the above parameters with GATA-3 expression was observed. High P53 expression was seen in the majority of high-grade tumors, with only a single case of low-grade UC showing strong P 53 reactivity. Conclusion: GATA-3 is a sensitive IHC marker to diagnose urothelial carcinoma. GATA-3 expression intensity and proportion show significant association with histopathological parameters and may play an important role in stratifying high-risk patients.

P16 expression in urothelial carcinoma: Experience from a tertiary care center in coastal South India.

Urothelial carcinomas (UC) account for 6 and 2% of all cancers in men and women, respectively. Human papillomavirus (HPV) is one of the causative agents in cancers of the uterine cervix and head and neck. The role of HPV is also being studied in cancers of the urinary bladder, penis, and prostate. As p16-INK4a is a surrogate marker for high-risk HPVE7 oncoprotein, this study aims to highlight the utility of p16 immunohistochemistry (IHC) in the evaluation of HPV-associated UC. A retrospective study was conducted on UC of the bladder received in the Pathology department between January 2013 and December 2018. Bladder biopsies from non-neoplastic lesions served as controls. IHC was done for the detection of the p16 antigen. The p16 staining was recorded as positive, when there was strong staining in >50% of tumor nuclei. The p16 positive and negative tumors were compared based on age, gender, tumor size, grade, and muscle invasion. P value <0.05 was considered statistically significant. The expression of p16 was analyzed in 72 UC and compared with 20 non-neoplastic cases, of which 26.4% of the cases showed p16 expression. The p16 expression was absent in the non-neoplastic lesions. While the majority (87.5%) of the low-grade tumors were negative for p16 expression, 43.8% high-grade tumors were positive. Similarly, a larger proportion of invasive carcinomas (38.8%) expressed p16 as compared to non-invasive carcinomas (13.8%). Thus, p16 expression showed a significant association with grade and stage in these malignancies (P < 0.05). The p16 expression was associated with high-grade and muscle-invasive UC. The p16 was absent in all non-neoplastic and precursor lesions. Thus, it can provide essential information not only about HPV association but also on the prognostic implications for the patients.

Abstract 4867: Effect of FGFR3 activity on Nectin-4 expression in urothelial carcinoma

Abstract Bladder cancer is a common malignancy in the U.S. and cause of significant morbidity and mortality. Combination platinum-based chemotherapy has long been the mainstay in the treatment of metastatic urothelial carcinoma (mUC); however, over the last decade there have been significant advances in systemic therapy, which includes the FDA approval of the pan-FGFR inhibitor erdafitinib (approved for tumors with FGFR2/3 alterations) and the antibody drug conjugate (ADC) enfortumab vedotin (which targets the cell surface protein Nectin-4). Despite these advances, most patients with mUC will progress and succumb from their disease, highlighting the need for further therapeutic development. In parallel to therapeutic advances, there have been significant advances in the molecular characterization of UC, including identification of luminal and basal subsets. NECTIN4 and FGFR3 alterations are known to be enriched in luminal subtypes and we have found, via query of the TCGA data, that NECTIN4 expression was significantly higher in FGFR3 altered tumors. Given this association we sought to investigate the effect of FGFR3 activity on Nectin-4 expression. Using erdafitinib, in select luminal UC cell lines (RT112, RT4, SW780) with FGFR3 fusion proteins, we found, much to our surprise, that FGFR inhibition increased Nectin-4 expression. Western blots, probing for Nectin-4, following 24H, 48H, and 72H of treatment with erdafitinib (50 nM) showed a significant increase in protein expression in all three cell lines. Importantly, we consistently saw increased expression of what we believe corresponds to the membrane bound form of Nectin-4 and have now confirm such by flow cytometry and immunofluorescence probing for Nectin-4 in the RT112 cell line. We have also seen statistically significant increases in NECTIN4 on RT-PCR following treatment with erdafitinib; however, we believe that Nectin-4 regulation by FGFR inhibition is not merely transcriptional because the induction of Nectin-4 protein appears significantly greater than the fold increase in mRNA. Notably, in two additional UC cell lines (HT1376 - a luminal cell line with high Nectin-4 expression &amp; UMUC3 - a basal cell line with low Nectin-4 expression), with no alterations in FGFR3, we did not see a significant change in Nectin-4 expression following treatment with erdafitinib. Based on our above results, we hypothesize that FGFR3 inhibition, in tumors harboring FGFR3 alterations, may act as a sensitize agent to Nectin-4 ADC targeted therapy; however, our results are preliminary and further investigation, particularly with in vivo models, is warranted. Citation Format: Sean Clark-Garvey, Mi Zhou, Andrew Truong, Wolfgang Beckabir, Michael Sturdivant, William Kim. Effect of FGFR3 activity on Nectin-4 expression in urothelial carcinoma. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4867.

Assessment of the expression pattern of HER2 and its correlation with HER2-targeting antibody-drug conjugate therapy in urothelial cancer

BackgroundHuman epidermal growth factor receptor 2 (HER2) overexpression is related to anti-HER2 therapy in many tumors. RC48- antibody-drug conjugate (ADC) has shown promising efficacy in patients with HER2-positive locally advanced or metastatic urothelial carcinoma (UC). The characteristic expression and scoring systems of HER2 are nonexistent in UC. We aimed to explore HER2 status and its correlation with the efficacy of HER2-targeting ADC therapy in UC. MethodsA total of 137 and 43 patients were enrolled in cohort 1 and cohort 2, respectively, from March 2009 to December 2018. The patients in cohort 2 were enrolled in a phase II study of RC48-ADC. UC samples were tested for HER2 status using immunohistochemistry (IHC) and/or fluorescence in situ hybridization (FISH). The 2018 ASCO/CAP HER2 scoring system was adopted and modified to score HER2 expression in UC. ResultsThe HER2-positive (IHC 2+ or 3+) rate was 24.1% (33/137). In HER2 IHC 2+ or 3+ patients, the HER2 gene amplification rate was 31% (13/42). The objective response rates (ORRs) in RC48-ADC-treated patients with IHC 3+, IHC 2+ and FISH+, IHC 2+ and FISH- were 58.8%, 66.7% and 40%, respectively. The ORR showed a trend toward a better benefit for RC48-ADC therapy in patients with HER2 amplification than in those without amplification (61.5% vs. 44.8%, P = 0.059). The heterogeneity of HER2 expression in the primary tumor was 55.5% (15/27), and the ORR was not significantly different between patients with tumor heterogeneity and homogeneity. ConclusionsIHC testing should be performed to assess the HER2 status before the initiation of HER2-ADC therapy. There was a trend toward a better benefit for patients with HER2 amplification, and tumor heterogeneity did not influence the drug efficacy.

The genomic landscape of urothelial carcinoma with high and low ERBB2/HER2 expression.

555 Background: Human epidermal growth factor receptor 2 (HER2) expression has been associated with poor prognosis in urothelial carcinoma (UC). Recent data suggests that HER2-targeted antibody-drug conjugate (ADC) treatment is efficacious. Here, we explore the role of ERBB2/ HER2 expression in UC by analyzing genomic and clinical outcomes in a large database of real-world patient samples. Methods: NextGen Sequencing (NGS) of DNA (592 genes or whole exome sequencing; WES)/RNA (whole transcriptome sequencing; WTS) was performed for 4,743 UC tumors submitted to Caris Life Sciences (Phoenix, AZ). PD-L1 (SP142; Positive (+): ≥ 2+, ≥ %5) and HER2 (4B5; +: ≥3+ and &gt;10%) expression was tested by IHC. High tumor mutational burden (TMB-high) was defined as ≥10 mutations/MB. ERBB2-high and -low expression were defined as ≥ top and &lt; bottom quartile of ERBB2 transcripts per million (TPM), respectively. Mann-Whitney U and X2/Fisher-Exact tests were applied where appropriate, with P-values adjusted for multiple comparisons ( p &lt; .05). Real-world overall survival (OS) information was obtained from insurance claims data and Kaplan-Meier estimates were calculated for molecularly defined patients. Results: 2.5% (120/4743) of tumors had HER2 IHC and WTS data available, 97% (61/63) of HER2+ tumors were ERBB2-high, and 98% (51/52) were ERBB2-amplified by DNA NGS (copy number ≥4; 77% with copy number ≥6). Tumors from lower tract UC had higher ERBB2 expression compared to upper tract UC (50 v 40 median TPM (mTPM), p &lt; .001). ERBB2 expression was similar between primary and metastatic tumors (47 v 47 mTPM, p = .95) but significantly higher in lung metastases (59 v 47 mTPM, p &lt; .001). Mutation rates of ERRB2 (12%), pTERT (76%) and ELF3 (14 %) were higher in ERBB2-high compared to ERBB2-low tumors (5%, 60%, 5%, all p &lt; .001). The opposite pattern was observed for CDKN2A (4% ERBB2-high, 9% ERBB2-low), KMT2D (20%, 35%) and PIK3CA (13%, 23%) (all p &lt; .001). There was a higher incidence of PD-L1+ staining in ERBB2-low (40.3%) compared to ERBB2-high tumors (18%, p &lt; .001). 45.4% of ERBB2-high tumors were TMB-high compared to 35% of ERBB2-low ( p &lt; .001). ERBB2-high tumors had higher expression of ADC target genes NECTIN4 (12 v 8 mTPM) and TACSTD2 (366 v 74 mTPM) compared to ERBB2-low (each, p &lt; .001). ERBB2-high tumors were associated with better overall survival from time of tissue sampling than ERBB2-low (HR 1.34, 95% CI 1.14-1.60, p &lt; .001) and from start of anti-PD-L1 therapy (HR 2.26, 95% CI 1.02-5.02, p = .04) but no difference in OS from start of anti-PD1 therapy (HR 1.39, 95% CI 0.75-2.58, p = .30). Conclusions: High concordance between HER2 IHC and ERBB2 expression/amplification was observed. Differences in the genomic landscape and ADC target expression of ERBB2-high vs -low UC may provide rationale for combination treatment strategies with HER2-blocking antibodies. The association between high ERBB2 expression and survival advantage warrants further investigation.

CT-based radiomics for prediction of PD-L1 expression in urothelial carcinoma

CT-based radiomics for prediction of PD-L1 expression in urothelial carcinoma

Urine PD-L1 is a tumor tissue candidate substitute and is associated with poor survival in muscle-invasive bladder cancer patients.

Programmed death molecule ligand 1 (PD-L1) expression in urothelial carcinoma is a predictive marker used to guide immunotherapy. As expression of PD-L1 may be heterogeneous in the tumor tissue space, it cannot be accurately determined by immunohistochemical analysis. In this study, we examined PD-L1 protein levels in preoperative urine samples from bladder cancer patients, evaluated the prevalence of PD-L1 in urine, examined the usefulness of urine as a surrogate for PD-L1 expression in tumors, and compared PD-L1 expression in postoperative pathological sections. We found that PD-L1 in urine and tumor tissue correlated well and that it may be able to some extent serve as a surrogate for tissues in bladder cancer and thus predict risk of recurrence in muscle-invasive bladder cancer (MIBC) patients. Our findings reveal the clinical relevance of urine PD-L1 as a noninvasive prognostic indicator for immunotherapy and offer clinical translational suggestions for eventual development of a prognostic model for immunotherapy for bladder cancer.

CD47 expression in urothelial carcinoma and its correlation with histopathological findings.

The incidence of bladder cancers (BCs) is increasing day by day in both males and females with increased smoking. New treatment protocols are required due to the long follow-up times and high recurrence rates. CD47 (integrin-associated protein) is a membrane receptor that is effective in normal and pathophysiological events such as apoptosis, proliferation, adhesion, and immunity. Phagocytosis of both normal and tumor cells is prevented, by binding to the ligand signal regulatory protein-1 (SIRP-1)α on macrophages. Anti-CD47 antibodies can eliminate the binding and may be effective in treatment. The study included 194 cases with urothelial carcinoma (UC) who underwent transurethral resection (TUR) of the bladder and had been followed up for one year. Cytoplasmic and membranous staining was preferred for CD47-positive staining. Staining results were compared with tumor grades, pathologic tumor stage, recurrence, and metastasis grade. The prevalence and intensity of CD47 staining in BCs were evaluated in this study. High-grade tumors were stained more pronouncedly compared with low-grade tumors and normal epithelium. The prevalence of CD47 staining was significant in pT2 (Pathological tumor stage 2, invades of muscularis propria) tumors, and its correlation with pTa (Pathological tumor stage a, noninvasive papillary carcinoma) tumors was significant ( P < 0.001). In the intensity evaluation, there was a significant difference between pTa and pT2 and between pT1 (Pathological tumor stage 1, invades lamina propria) and pT2 ( P = 0.003). CD47 is expressed strongly in high-grade tumors. The prevalence and intensity of CD47 staining are high in pathologic tumor stage, recurrence, and distant metastases and are considered poor prognostic markers. We believe that anti-CD47 antibodies can be used as an alternative to the current treatment or in combination with other medications, and the systemic side effects that may occur with intravesical treatment can be prevented.