Cadherins (calcium-dependent adhesion proteins) are important in cellular adhesion and may play a role in the development and progression of renal cell carcinoma (RCC). This study investigated changes in cadherin 3 (CDH3; P-cadherin) mRNA expression, DNA methylation, and protein expression in RCC and compared the results with the histopathological and clinical characteristics of patients. The possible contribution of CDH3 to tumor cell invasiveness was tested in a functional assay using siRNA-based suppression of CDH3 expression and subsequent real-time impedance analysis using a Matrigel invasion model. Our analyses revealed a tumor-specific loss of CDH3 mRNA expression, CDH3 DNA hypermethylation, and loss of distal tubular and collecting duct CDH3 protein expression in RCC. A relatively higher methylation level in tumors was associated with a loss of cell differentiation and higher clinical stage. siRNA-induced suppression of CDH3 expression modulated the invasion characteristics of tumor cells in the impedance-based real-time cellular analysis. Our results indicate that loss of CDH3 expression is common in RCC and may contribute to the pathogenesis of a subset of RCC. Further studies to reveal the mechanisms of loss of expression and its effects on the invasive behavior of renal tumor cells are required.

The detection of DNA methylation in primary tumor tissues could be relevant for early stratification of aggressive renal cell carcinomas (RCCs) as a basis for future personalized adjuvant therapy. Methylated TCGA KIRC based candidate CpG loci in INA, NHLH2, and THBS4 that are possibly associated with RCC metastasis were evaluated by pyrosequencing in 154 paired normal adjacent and primary tumor tissues, as well as in 202 metastatic tissues. Statistical analysis was carried out by bivariate logistic regression for group comparisons, log rank survival analysis, and unsupervised and supervised analysis for the classification of tumors. Increased methylation of INA, NHLH2, and THBS4 loci were significantly associated with distant metastasis in primary tumors (p < 0.05), tissue-specific hypermethylation in metastatic (p = 7.88 × 10⁻⁸, 5.57 × 10⁻¹⁰, 2.06 × 10⁻⁷) and tumor tissues (p = 3.72 × 10⁻²⁴, 3.17 × 10⁻¹³, 1.58 × 10⁻¹⁹), and shortened progression free survival in patients (p = 0.03). Combined use of CpG site-specific methylation permits the discrimination of tissues with metastatic disease and reveals a significant contribution of CpG sites in all genes to the statistical classification model. Thus, metastasis in RCC is significantly associated with methylation alterations in INA, NHLH2, and THBS4 loci, providing independent information for the potential early detection of aggressive renal cancers as a rationale for stratifying patients to adjuvant therapies.

Background/Objectives: Alterations in long non-protein-coding RNAs (lncRNAs) are known to influence cellular proliferation, apoptosis, and metastasis in human cancers, including renal cell carcinoma (RCC). Methods: Using pyrosequencing, we analyzed DNA methylation (DNAm) at 23 loci within the LINC00404 CpG island across 28 human cancer cell line models, 181 RCC tumor tissues, 154 paired tumor-adjacent normal tissues (adNs), and 194 metastatic tissue samples. Results: Our analysis revealed that all CpG sites exhibited tumor-specific hypermethylation (all p ≤ 1.4 × 10⁻⁵). Moreover, primary RCC tissues with distant metastases (M1) and metastatic tissue samples (Mtx) showed significant hypermethylation compared to RCC without distant metastases (M0). Notably, DNAm in Mtx displayed a significant increase in 22 CpG sites, compared to 12 CpG sites in the M1/M0 comparison, suggesting that DNAm in Mtx differs both qualitatively and quantitatively. Conclusions: Given that elevated levels of DNAm were also observed in the majority of cell line models, our findings suggest that LINC00404 may play a pivotal role in the malignant development and progression of RCC metastasis, as well as in other human cancers.

Background: Long non-coding RNAs (lncRNAs) regulate multiple cellular processes and are involved in the development and progression of human cancers, including renal cell carcinoma (RCC). Methods: Genome-wide DNA methylation profiling using Illumina EPIC arrays was performed to identify differentially methylated regions (DMR) associated with metastatic RCC. Candidate loci within LINC01354 and LINC01510 were validated by pyrosequencing in 145 primary RCC tissues, 124 paired tumor-adjacent normal tissues (adNs), and 194 metastatic tissue samples obtained from 101 patients with metastatic RCC. In addition, data from The Cancer Genome Atlas Kidney Renal Clear Cell Carcinoma Collection (TCGA-KIRC) were analyzed in silico to assess the clinical relevance of DNA methylation (DNAm) and its association with gene expression. Results: Hypermethylation across all analyzed CpG sites at both loci was associated with high-stage (≥ T3; p = 0.001) and high-grade (≥ G3; p = 0.003) RCC and was markedly increased in metastatic tissues compared with localized primary tumors ( p < 0.001). In addition, primary tumors from patients with distant metastases (M+) showed significantly higher methylation at PS_307-CG1 within LINC01510 than tumors from patients without distant metastases (M0; p = 0.031). Analyses of TCGA-KIRC data confirmed associations of increased methylation with adverse clinicopathological features and reduced gene expression, supporting a potential role for epigenetic silencing. Conclusion: Our data provide further insight into the epigenetic alterations associated with RCC progression by demonstrating tumor- and metastasis-associated hypermethylation at the LINC01354 and LINC01510 loci. The association of hypermethylation with advanced disease and metastatic tissues, together with the observed inverse relationship between methylation and expression, suggests that epigenetic inactivation of these lncRNAs may contribute to RCC progression. DNAm at these loci may therefore represent a potential biomarker of aggressive disease.

Background/Objectives: Antibody–drug conjugates (ADCs) targeting Nectin-4 (enfortumab vedotin) and TROP-2 (sacituzumab govitecan) have transformed the management of advanced urothelial carcinoma (UC), but evidence on their molecular targets in rare non-urothelial bladder carcinomas (N-UC) is scarce. We evaluated the immunohistochemical expression of TROP-2 and Nectin-4 in muscle-invasive UC and N-UC and assessed their association with tumour stage, nodal status and overall survival. Methods: In this retrospective single-centre study, 111 consecutive patients with primary muscle-invasive bladder carcinoma (73 UC, 38 N-UC including squamous, adenocarcinoma, neuroendocrine and sarcomatoid variants) were analysed. TROP-2 and Nectin-4 expression was quantified using the H-score; positivity was defined as ≥15. Marker expression was correlated with clinicopathological characteristics and overall survival (OS) using chi-squared, linear-by-linear, log-rank, and Cox regression analyses. Results: TROP-2 positivity was observed in 46.6% of UC and 36.8% of N-UC (p = 0.925); Nectin-4 positivity in 17.8% and 28.9%, respectively (p = 0.275). Among positive cases, TROP-2 H-scores were higher in N-UC than in UC (mean 109 vs. 70; Mann–Whitney p = 0.045; Cliff’s delta 0.37, 95% CI 0.05–0.66); as no adjustment for multiple testing was applied, this difference is regarded as nominally significant and exploratory. Nectin-4 H-scores were numerically higher in UC (mean 82 vs. 53) but did not reach statistical significance (p = 0.84). Across the cohort, TROP-2 expression increased with advancing T stage (linear-by-linear p = 0.026) and was associated with nodal involvement (p = 0.043). Nectin-4 showed no significant stage association. Sarcomatoid carcinomas were negative for both markers. Median OS was 41 months in UC versus 19 months in N-UC (p = 0.668). Neither marker independently predicted OS, whereas advanced T stage (p = 0.003) and nodal involvement (p = 0.021) were significantly associated with poorer survival; T stage remained independently prognostic in multivariable analysis. Conclusions: TROP-2 and Nectin-4 are expressed at comparable rates in muscle-invasive UC and rare N-UC, except in sarcomatoid variants. TROP-2 expression increased with advancing tumour stage, suggesting stage-dependent regulation in muscle-invasive disease. Therefore, both markers should be interpreted as potential therapeutic targets whose expression can be demonstrated in these tumours, rather than as prognostic biomarkers or as validated predictive biomarkers of ADC response; because no patient received an ADC, the present study cannot determine whether expression predicts clinical benefit, and this distinction requires prospective, treatment-linked evaluation that includes patients with N-UC.