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Cell surface hyaluronidase transmembrane protein 2 (TMEM2), which also serves as a reportedly functions in malignancy of several solid tumors. However, TMEM2 involvement in bladder cancer (BCa) is unknown. Therefore, we investigate potential changes in expression of TMEM2 during BCa invasion and over the course of the epithelial mesenchymal transition (EMT). Immunohistochemical analysis of 127 clinical specimens revealed that TMEM2 expression changed with pathological stage (pT) and infiltration pattern (INF) and was highest in pTa-pT1 of INFa tumors and significantly lower at stages from pTa-pT1 to pT2 or 3 in INFb or INFc. E-cadherin expression was highest in INFa and lowest in INFc, a pattern comparable to TMEM2 expression. TMEM2 protein expression analysis of BCa cell lines showed that muscle-invasive T24 and YTS-1 cells with low TMEM2 expression exhibited EMT phenotypes in vitro, in contrast to high TMEM2-expressing non-muscle invasive RT4 cells. EMT-induced non-muscle invasive RT4 cells also showed significantly decreased plasma membrane expression of TMEM2. Our data suggested TMEM2 expression is higher in non-invasive cancers, whereas invasive cancer cells are less likely to express TMEM2 during muscle-invasion and "partial EMT"., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

The evaluation of surgical damage is challenging because of the lack of specific biomarkers. Total cell-free DNA (cfDNA) levels have been reported to increase with external trauma and may be a biomarker for tissue damage. To investigate the utility of perioperative total cfDNA levels in evaluating surgical damage in urological surgeries. This multicenter, prospective, observational study included 196 patients scheduled for urological surgeries between September 2020 and July 2021. The primary outcome was the change in total cfDNA levels before and after urological surgery. The secondary outcome was the effect of surgical type on total cfDNA ratio before and after urological surgery. The postoperative median total cfDNA level of the 196 patients was significantly increased 2.5-fold compared to the preoperative level (185.2 ng/mL vs. 406.7 ng/mL, P < 0.001). The median total cfDNA before/after ratio was greater than four-fold for kidney transplantation, open cystectomy, and open adrenalectomy. The ratio was less than two-fold for laparoscopic adrenalectomy and robot-assisted radical prostatectomy. Major surgery showed a significant postoperative increase in total cfDNA levels, while minor surgery did not. Total cfDNA levels increased 2.5-fold after urological surgery and it can be used as an acute-phase biomarker for surgical damage., (© 2021. The Author(s).)

Purpose: To investigate the prognostic significance of total cell-free DNA (cfDNA) level and androgen receptor amplification (AR-amp) in patients with castration-resistant prostate cancer (CRPC)., Methods: We retrospectively compared the total cfDNA level and AR-amp in 42 individuals without prostate cancer, 57 patients with localized prostate cancer without androgen-deprivation therapy (ADT), 97 patients with castration-sensitive prostate cancer (CSPC) with ADT, and 97 patients with CRPC. The association of these cfDNA biomarkers on disease status and overall survival was evaluated using Kaplan-Meier analysis and multivariable Cox regression analysis. Finally, a simple risk model was developed including total cfDNA and AR-amp to predict poor prognosis., Results: The median total cfDNA level and AR-amp in patients with CRPC was 387 pg/μL and 1.07 copies, respectively. The total cfDNA levels and AR-amp were significantly higher in the patients with CRPC than in individuals without prostate cancer, patients with localized prostate cancer without ADT, and patients with CSPC with ADT. Total cfDNA-high (> 600 pg/μL) and AR-amp-high (> 1.26 copies) were significantly associated with poor overall survival. Multivariable Cox regression analysis showed cfDNA-high and AR-amp-high were significantly associated with poor overall survival in patients with CRPC. We developed a risk model using cfDNA-high (score 1) and AR-amp-high (score 1). The risk score 1-2 was significantly associated with worse overall survival than score 0., Conclusion: Total cfDNA level and AR-amp are potential biomarkers for poor prognosis in patients with CRPC., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH, DE part of Springer Nature.)

Objectives: To investigate the association between Ki67 index and programmed death-ligand 1 (PD-L1) expression in muscle-invasive bladder cancer (MIBC) patients after RC., Materials and Methods: We retrospectively evaluated 262 MIBC patients treated with RC between April 2004 and April 2020. The impact of Ki67 index and PD-L1 expression on prognosis was evaluated by univariate Cox regression analysis. In addition, a pathomolecular risk score, including Ki67 and PD-L1, was developed to predict prognosis and pathological factors. We also evaluated the link between the Ki67 index and PD-L1 under the IL-6 stimulation in the bladder cancer cell lines of T24 and 5637 cells., Results: The median age and follow-up period was 69 years and 52 months, respectively. Ki67 index and PD-L1 expression were significantly associated with tumor recurrence. Univariate Cox regression analysis showed that pT3-4, mixed histology, lymphovascular invasion positive (LVI+), pN+, Ki67-high (>17%), and PD-L1+ were significantly associated with recurrence-free survival (RFS). The pathomolecular risk score was developed using resection margin+ (1 point), mixed histology (1 point), LVI+ (1 point), pN+ (1 point), and Ki67-high (1 point). RFS and overall survival were significantly shorter in patients with higher pathomolecular risk scores (>1) than in those with lower risk scores (≤1). Cell proliferation was significantly increased in the T24 and 5637 cells under the IL-6 stimulation, while PD-L1 expression was not., Conclusions: A significant effect of Ki67-high and PD-L1 expression on poor prognosis was observed in patients with MIBC. Further studies are necessary to elucidate the precise mechanisms of cell proliferation and PD-L1 expression in patients with MIBC., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The LIM and SH3 protein-1 (LASP-1) is a multi-domain protein that is involved in several malignant cancers. The role of LASP-1 in malignant phenotypes including high invasive properties and unrestricted cell proliferation, remain to be elucidated. The present study reported the association of LASP-1 expression with bladder cancer malignancy and its role in cancer cell invasion and proliferation. The immunohistochemical analysis of the expression status of LASP-1 in radical cystectomy specimens from invasive bladder cancer patients revealed that the LASP-1-positive patients demonstrated a decreased survival rate compared with the LASP-1-negative patients. The expression level of LASP-1 was increased in invasive bladder cancer cell lines compared with the non-invasive bladder cancer cell lines. Invasive cancer cells form invadopodia, the filamentous actin-based membrane protrusions that are essential in cancer cell invasion. Knockdown of LASP-1 reduced the ability to form invadopodia, resulting in decreased invasive capacity of the LASP-1 knockdown cells. In addition, knockdown of LASP-1 reduced cell proliferation. These results suggest that LASP-1 is important in invadopodia formation and cell proliferation of bladder cancer cells, promoting the malignant properties and resulting in poor-prognosis.

Muscle invasive bladder cancer is an aggressive type of epithelial tumor with a high rate of metastasis. For bladder cancer cells to reach the muscle layer, cells must invade through an urothelial cell monolayer (transurothelial invasion) and basement membrane. However, the process by which transurothelial invasion occurs has not been fully characterized. In this study we developed a novel method to evaluate the transurothelial invasion capacity and investigated its cellular and molecular processes using primary culture cells from bladder cancer patients. The analysis revealed that compared with the prognosis for patients with non‑muscle invasive bladder cancer that of patients with muscle invasive bladder cancer was particularly poor due to metastatic recurrence. Cancer cells from patients with muscle invasive bladder cancer exhibited a higher invasive capacity through the urothelial cell monolayer compared with those from non‑invasive bladder cancer patients. Furthermore, muscle invasive bladder cancer cells demonstrated a greater ability to form invadopodia, the filamentous actin‑based membrane protrusions required for matrix degradation and invasion compared with non‑invasive cells. Bladder cancer cell lines were established with reduced invadopodia formation by silencing the expression of cortactin, an essential component of invadopodia. The cortactin knockdown bladder cancer cells with reduced invadopodia formation demonstrated a markedly reduced ability to invade through the urothelial cell monolayer, indicating that invadopodia are essential for transurothelial invasion. The results indicate that invadopodia formation is required for muscle invasion of aggressive bladder cancer cells.

Invasive cancer cells form the filamentous actin‑based membrane protrusions known as invadopodia. Invadopodia are thought to play a critical role in cancer cell invasion and metastasis due to their ability to degrade the extracellular matrix. The present study assessed whether invadopodia formation is essential in extravasation of circulating bladder cancer cells and lung metastasis. To analyze the importance of invadopodia, bladder cancer cell lines with reduced invadopodia formation were established by silencing the expression of cortactin, an essential component of invadopodia, using cortactin short hairpin RNA. Bladder cancer cells with cortactin knockdown demonstrated a markedly decreased ability to form invadopodia, secrete matrix metalloproteinases and invade the extracellular matrix. In addition, the knockdown cells exhibited a reduced transendothelial invasion capacity and decreased formation of metastatic foci in the lungs. The present study demonstrated that bladder cancer cells with cortactin knockdown have a reduced capacity to extravasate into the lung from the circulation, due to the decreased invasive character of invadopodia. This suggests that invadopodia formation is a critical process for cancer cell extravasation.

Core2 β-1,6-N-acetylglucosaminyltransferase (C2GnT) forms an N-acetylglucosamine branch in the O-glycans (core2 O-glycans) of cell surface glycoproteins. We previously revealed that the expression of C2GnT is positively correlated with poor prognosis in prostate cancer patients. However, the detailed mechanisms underlying their poor prognosis remain unclear. In the current study, we report that the core2 O-glycans carried by the surface MUC1 glycoproteins of prostate cancer cells play an important role in the evasion of NK cell immunity. In C2GnT‑expressing prostate cancer cells, the MUC1 core2 O-glycans are modified with poly-N-acetyllactosamine. MUC1 glycoproteins carrying poly-N-acetyllactosamine attenuated the interaction of the cancer cells with NK cells, resulting in decreased secretion of granzyme B by the NK cells. Poly‑N‑acetyllactosamine also interfered with the ability of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) to access the cancer cell surface. These effects of poly-N-acetyllactosamine on NK cells render C2GnT-expressing prostate cancer cells resistant to NK cell cytotoxicity. By contrast, C2GnT-deficient prostate cancer cells carrying a lower amount of poly-N-acetyllactosamine than the C2GnT-expressing prostate cancer cells were significantly more susceptible to NK cell cytotoxicity. Our results strongly suggest that C2GnT-expressing prostate cancer cells evade NK cell immunity and survive longer in the host blood circulation, thereby resulting in the promotion of prostate cancer metastasis.

The majority of deaths associated with solid tumors are caused by tumor metastasis. To prevent metastasis, it is vital to understand its detailed process. In hematogenous metastasis of bladder cancer, some cancer cells disseminating into blood circulation extravasate into the lung tissues to form metastases. To study the molecular basis of the lung metastasis of bladder cancer, we employed an in vivo selection system that mimics hematogenous metastasis of bladder cancer on a low-metastatic bladder cancer cell line (KK-47). We have successfully isolated a high-metastatic bladder cancer subline, KK-47HM4, from KK-47 cells. We characterized KK-47HM4 in in vitro experimental systems. No significant difference in growth rate and susceptibility to NK cell attack between KK-47 and KK-47HM4 cells was observed. However, KK-47HM4 exhibited the higher capacities of Matrigel Matrix invasion and transendothelial invasion than KK-47. These results suggest that the extravasation of KK-47HM4 cells was enhanced among the multiple steps of the lung metastasis of bladder cancer. Our cDNA microarray analysis identified 67 genes whose expression was up- or downregulated in KK-47HM4 cells compared with KK-47 cells. This analysis data implied that one possible cause for enhanced extravasation of KK-47HM4 is its higher adhesion to extracellular matrix proteins. KK-47HM4 is the first bladder cancer subline with enhanced extravasation potential using the in vivo selection system. The information provided by our cDNA microarray analysis using KK-47HM4 will be useful for further investigation into the molecular basis of extravasation of cancer cells.