JAM3: A prognostic biomarker for bladder cancer via epithelial­mesenchymal transition regulation

Understanding the intricate relationship between prognosis, immune function, and molecular markers in bladder cancer (BC) demands sophisticated analytical methods. To identify novel biomarkers for predicting prognosis and immune function in BC patients, we combined weighted gene co-expression network analysis (WGCNA) and least absolute shrinkage and selection operator (LASSO) regression analysis. This was conducted using data from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. Ultimately, we screened the junctional adhesion molecule 3 (JAM3) as an independent risk factor in BC. High levels of JAM3 were linked to adverse clinical parameters, such as higher T and N stages. Additionally, a JAM3-based nomogram model accurately predicted 1-, 3- and 5-year survival rates of BC patients, indicating potential clinical utility. Functional enrichment analysis revealed that high JAM3 expression activated the calcium signaling pathway, the extracellular matrix (ECM)-receptor interaction, and the PI3K-Akt signaling pathway, and was positively correlated with genes associated with epithelial­mesenchymal transition (EMT). Subsequently, we found that overexpression of JAM3 promoted the migration and invasion abilities in BC cells, regulating the expression levels of N-Cadherin, matrix metallopeptidase 2 (MMP2), and Claudin-1 thereby promoting EMT levels. Additionally, we showed that JAM3 was negatively correlated with anti-tumor immune cells such as CD8+T cells, while positively correlated with pro-tumor immune cells such as M2 macrophages, suggesting its involvement in immune cell infiltration. The immune checkpoint CD200 also showed a positive correlation with JAM3. Our findings revealed that elevated JAM3 levels are predictive of poor prognosis and immune cell infiltration in BC patients by regulating the EMT process.

A new immune-related gene signature predicts the prognosis and immune escape of bladder cancer.

BACKGROUND: The biological roles of immune-related genes (IRGs) in bladder cancer (BC) need to be further elucidated. OBJECTIVE: To elucidate the predictive value of IRGs for prognosis and immune escape in BC. METHODS: We comprehensively analyzed the transcriptomic and clinical information of 430 cases, including 19 normal and 411 BC patients from the TCGA database, and verified 165 BC cases in the GSE13507 dataset. The risk model was constructed based on IRGs by applying LASSO Cox regression and exploring the relationship between the risk score and prognosis, gene mutations, and immune escape in BC patients. RESULTS: We identified 4 survival-related genes (PSMC1, RAC3, ROBO2 and ITGB3) among 6,196 IRGs in both the TCGA and GES13507 datasets,, which were used to establish a gene risk model by applying LASSO Cox regression. The results showed that the high-risk (HR) group was closely associated with poor survival or advanced pathological stage of BC. Furthermore, the risk score was found to be an independent risk factor for prognosis of BC patients. In addition, high-risk individuals showed a greater prevalence of TP53 mutations lower CD8+ T-cell and NK cell infiltration, higher Treg cell infiltration, higher expression of PD-L1, and higher immune exclusion scores than those in the low-risk (LR) group. Finally, the experimental verification shows that the model construction gene, especially PMSC1, plays an important role in the growth and metastasis of bladder cancer. CONCLUSIONS: These evidences revealed the vital role of IRGs in predicting prognosis, TP53 mutation and immune escape in BC patients.

Comprehensive analysis of integrin αvß3/α6ß1 in prognosis and immune escape of prostate cancer.

Integrin αvß3/α6ß1 are crucial in the transduction of intercellular cancer information, while their roles in prostate cancer (PCa) remain poorly understood. Here, we systematically analyzed the transcriptome, single nucleotide polymorphisms (SNPs) and clinical data of 495 PCa patients from the TCGA database and verified them in 220 GEO patients, and qPCR was used to validate the expression of the model genes in our patients. First, we found that integrin αvß3/α6ß1 was negatively correlated with most immune cell infiltration and immune functions and closely associated with poor survival in TCGA patients. Then, we divided these patients into two groups according to the expression level of αvß3/α6ß1, intersected differentially expressed genes of the two groups with the GEO dataset and identified eight biochemical recurrence-related genes (BRGs), and these genes were verified by qPCR in our patients. Next, these BRGs were used to construct a prognostic risk model by applying LASSO Cox regression. We found that the high-risk (HR) group showed poorer OS, PFS, biochemical recurrence and clinical characteristics than the low-risk (LR) group. In addition, the HR group was mainly enriched in the cell cycle pathway and had a higher TP53 mutation rate than the LR group. More importantly, lower immune cell infiltration and immune function, higher expression of PD-L1, PD-1, and CTLA4, and higher immune exclusion scores were identified in the HR group, suggesting a higher possibility of immune escape. These findings suggested the key role of integrin αvß3/α6ß1 in predicting prognosis, TP53 mutation and immune escape in PCa.

Heat shock protein family A member 8 is a prognostic marker for bladder cancer: Evidences based on experiments and machine learning.

Heat shock protein member 8 (HSPA8) is one of the most abundant chaperones in eukaryotic cells, but its biological roles in bladder cancer (BC) are largely unclear. First, we observed that HSPA8 was abundant in both cell lines and tissues of BC, and the HSPA8-high group had poorer T stages and overall survival (OS) than the HSPA8-low group in the TCGA patients. Next, when we knocked down HSPA8 in BC cells, the growth and migration abilities were significantly decreased, the apoptosis rates were significantly increased, and the Ki67 fluorescence intensity was decreased in BC cells. Moreover, caspase 3 was significantly decreased with overexpression of HSPA8 in BC cells. After that, a machine learning prognostic model was created based on the expression of HSPA8 by applying LASSO Cox regression in TCGA and GEO patients. The model indicated that the low-risk (LR) group with BC had better tumour stages, lymphovascular invasion, and OS than the high-risk (HR) group. Additionally, the risk score was demonstrated to be an independent risk factor for the prognosis of BC by univariate and multivariate Cox analyses. Moreover, the HR group showed a greater rate of TP53 mutations and was mostly enriched in the ECM-receptor interaction pathway than the LR group. Importantly, lower CD8+ T-cell and NK cell infiltration, higher immune exclusion scores, higher expression of PD-L1 and CTLA4 and poorer immune checkpoint therapy effects were found in the HR group. These findings demonstrated how crucial HSPA8 plays a role in determining the prognosis of bladder cancer.

Endostatin 33 Peptide Is a Deintegrin α6ß1 Agent That Exerts Antitumor Activity by Inhibiting the PI3K-Akt Signaling Pathway in Prostate Cancer.

BACKGROUND: Prostate cancer (PCa) is the leading cause of death in men and has poor therapeutic outcomes. METHODS: A novel endostatin 33 peptide was synthesized by adding a specific QRD sequence on the basis of the endostatin 30 peptide (PEP06) with antitumor activity. Then, bioinformatic analysis and subsequent experiments were performed to validate the antitumor function of this endostatin 33 peptide. RESULTS: We found that the 33 polypeptides significantly inhibited growth, invasion and metastasis and promoted the apoptosis of PCa in vivo or vitro, which is more significant than PEP06 under the same conditions. According to 489 cases from the TCGA data portal, the α6ß1 high expression group was closely associated with the poor prognosis (Gleason score, pathological N stage, etc.) of PCa and was mainly enriched in the PI3K-Akt pathway. Subsequently, we demonstrated that endostatin 33 peptide can down-regulate the PI3K-Akt pathway via the targeted inhibition of α6ß1, thereby inhibiting the epithelial-mesenchymal transition and matrix metalloproteinase in C42 cell lines. CONCLUSION: The endostatin 33 peptide can exert antitumor effects by inhibiting the PI3K-Akt pathway, especially in tumors with a high expression of the integrin α6ß1 subtype, such as prostate cancer. Therefore, our study will provide a new method and theoretical basis for the treatment of prostate cancer.

[Research progress on markers related to angiogenesis and stemness characteristics in prostate cancer].

Recurrence, metastasis and drug resistance in patients with prostate cancer (PCa) are still important causes of high mortality in patients. Two important features of solid tumors, including PCa, are tumor angiogenesis and the emergence of cancer stemness. Studies have found that cancer stem cells can promote tumor angiogenesis, and the highly vascularized tumor microenvironment further supports the growth of these stem cells, forming a harmful cycle that leads to tumor recurrence, metastasis, and drug resistance. This article summarizes the regulatory factors of tumor angiogenesis and tumor stemness characteristics in PCa and deeply explores their role in promoting the development of PCa.

The predictive value of pyroptosis for the prognosis and immune escape of bladder cancer.

OBJECTIVE: To evaluate the predictive value of pyroptosis-related genes for the prognosis and immune escape of bladder cancer (BC). METHODS: Transcriptomic and single nucleotide polymorphisms (SNPs) data were downloaded from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) portal. Least absolute shrinkage and selection operator (LASSO) analysis was carried out to construct a prognostic risk model for BC patients. RESULTS: Based on the expression of 50 pyroptosis-related genes, BC patients from TCGA database were divided into two clusters, which showed significant differences in overall survival and disease specific survival. Furthermore, we intersected the differentially expressed genes between these two clusters with those identified from the GSE13507 dataset and finally identified eight survival related genes, which was used to construct a prognostic risk model by LASSO Cox regression. According to the model, the high-risk (HR) group was closely associated with poor survival or the advanced pathological stage of BC. In addition, the HR group was mainly enriched in cell cycle and immune-related pathways and had a higher TP53 mutation rate than the low-risk (LR) group. Furthermore, these two risk groups were significantly related to immune cell composition, immune cell infiltration, and immune response. Importantly, a higher expression of PD-1, PD-L1, and CTLA4 as well as higher immune exclusion scores were found in the HR group, suggesting a higher possibility of immune escape. CONCLUSION: Our studies revealed the key role of pyroptosis in predicting the prognosis, TP53 mutation, and immune escape of patients with BC.

Roles of Loss of Chromosome 14q Allele in the Prognosis of Renal Cell Carcinoma with C-reactive Protein Abnormity.

BACKGROUND:: Renal cell carcinoma (RCC) is frequently associated with paraneoplastic inflammatory syndrome (PIS). This study aimed at exploring the connections between the survival rate and specific gene alterations and the potential mechanism. METHODS:: We retrospectively studied 69 surgical RCC cases from August 2014 to February 2016, including 18 cases of clear cell RCC (ccRCC) demonstrating elevated pretreatment serum C-reactive protein (CRP, Group A). Twelve of the 18 cases were symptomized with febrile episode. We also selected 49 cases of ccRCC with normal pretreatment CRP (Group B). Using 22 microsatellite markers, we compared the incidence of loss of heterozygosity (LOH) between Group A and Group B. All statistical tests are two-sided. RESULTS:: The 3p LOH was common in both Group A (89%) and Group B (92%). The frequency of 14q LOH in Group A (16 of 18) was higher than Group B (4 of 49, χ2 = 40.97 P < 0.0001). The 3p and 14q LOH were the characteristics of ccRCC with elevated acute phase reactants, including PIS, regardless of the presence of metastasis. On the contrary, 14q LOH was a rare genomic alternation in advanced-staged ccRCC without PIS. The overall survival of patients with elevated CRP (33.3%) was lower than its counterparts (6.1%, hazard ratio=1.852, P < 0.0001) in Kaplan-Meier curve. CONCLUSIONS:: The results imply that the disruption of a 14q gene(s) might result in not only the inflammatory manifestations in the tumor host but also the poor survival rate as well. The isolation of the gene(s) on 14q might be a vital goal in the treatment of PIS-associated RCC.

Antitumor effects of mutant endostatin are enhanced by Bcl-2 antisense oligonucleotides in UM-UC-3 bladder cancer cell line.

BACKGROUND: Endostatin is a potent inhibitor of tumor angiogenesis. In the preliminary studies, we developed a mutant endostatin containing Arg-Gly-Asp-Arg-Gly-Asp (RGDRGD) sequences. In this study, we compared the antitumor effects of mutant endostatin and Bcl-2 antisense oligonucleotides both in combination and individually. METHODS: The artificially synthesized Bcl-2 ASODN (antisense oligonucleotides) included a translation-initiation site and was transfected into the bladder cancer cells by Lipofectamine. Cell growth was investigated by the tumor cell growth chart, MTT assay, caspase-3 activity detection assay, AO/EB fluorescein stain, and the annexin V-FITC apoptosis detection assay. In the in vivo study, UM-UC-3 bladder cancer cells were subcutaneously implanted into nude mice and the growth of tumor was examined. The ultrastructure of the tumor tissues in the treated and control groups were observed. RESULTS: The cell growth chart showed that the cell population of the treated combination group decreased by 52.04% compared to the control group. The inhibition rate of the treated combination group was (79.66 ± 6.79)%, whereas those of the individual ASODN and ES groups were (53.39 ± 3.22)% and (50.22 ± 5.46)% respectively. In the caspase-3 activity detection using AO/EB fluorescein stain and annexin V-FITC apoptosis detection assay, the co-inhibitory effect was higher than the individual inhibitory effects (P < 0.05). There were significant differences in the inhibition of the solid tumor growth in the in vivo study. CONCLUSIONS: Our findings indicated that Bcl-2 antisense oligonucleotides enhance the antitumor effects of mutant endostatin both in vitro and in vivo. We noted the synergistic effects of Bcl-2 antisense oligonucleotides combined with mutant endostatin.

Ultrasound-guided minimally invasive percutaneous nephrolithotomy in flank position for management of complex renal calculi.

OBJECTIVES: To evaluate the safety and efficacy of performing ultrasound-guided minimally invasive percutaneous nephrolithotomy (MPCNL) in the flank position for the management of complex renal calculi. Percutaneous nephrolithotomy is usually performed with the patient in the prone position under fluoroscopic guidance; however, this position, and guidance method have some limitations. METHODS: From January 2007 to December 2009, 93 patients (101 kidneys) with complex renal calculi underwent ultrasound-guided MPCNL in the flank position. RESULTS: The mean age of the patients was 45.3 years (range 29-71). The calculi-free rate in the patients who underwent a single procedure was 78.2% (79 of 101 kidneys). The average operative duration was 82.6 minutes (range 45-190). Although the perioperative blood loss was not significantly different between single-tract and double-tract MPCNL (P = .087, F = 2.981), the calculi-free rate was significantly greater in the patients who underwent double-tract MPCNL than in those who underwent single-tract MPCNL (P = .027, chi-square = 4.873). Perioperative blood transfusions were not required in any patient. Similarly, ureteral calculi due to percutaneous nephrolithotomy were not observed. Secondary renal hemorrhage occurred in 3 patients who had undergone single-tract MPCNL and 1 underwent nephrectomy. CONCLUSIONS: The results of our study have shown that ultrasound-guided MPCNL with the patient in the flank position is safe and effective for treating complex renal calculi, without the side effects of radiation to the patient and surgeon. Double-tract MPCNL is suitable for complex renal calculi and, in some cases, is required to increase the calculi-free rate. The insertion of twin ureteral catheters before lithotripsy might be helpful in avoiding residual ureteral calculi after percutaneous nephrolithotomy.

[Effects of irrigating fluid absorption in percutaneous nephrolithotripsy].

OBJECTIVE: To determine the hemodynamic status, fluid-electrolyte changes and complications associated with irrigation time in percutaneous nephrolithotripsy. METHODS: A total of 68 renal calculi patients (31 males and 37 females) were recruited. The lateral recumbent percutaneous nephrolithotripsy was operated with Ho laser under ultrasonic guidance. 0.9% NaCI was used as perfusion fluid. The following items were recorded: mean arterial blood pressure (MAP), heart rate, central venous pressure (CVP), hemoglobin, sodium, potassium and chloride; perfusion time during operation; peri-operative and post-operative complications. RESULTS: (1) Peri-operative and post-operative conditions: the average operative time was 83.1 +/- 22.21 minutes. Two cases stopped because of bleeding after puncture and the tube of stoma was placed for stone clearance of the second time. There was more bleeding in 11 patients, but the operations were continued with blood transfusion and close monitoring. Two operations ceased because of a premunition of congestive heart failure. Nine patients needed post-operative blood transfusion and 18 had a post-operative fever. One patient bled in and around the tube and had a peri-renal infection a week later. (2) Changes of observation parameters: there was no significant difference in CVP, heart rate, hemoglobin, sodium, potassium and chloride (P > 0.05). The post-perfusion value of MAP increased (P < 0.05) especially in the cases of more bleeding and long time of irrigation. Peri-operative and post-operative injection of furosemide could reduce the CVP value. The average irrigation time in the fever group was longer than the non-fever group (P < 0.05) and the CVP value of the fever group was higher than the non-fever group (P < 0.05). CONCLUSION: Low pressure and short time of perfusion are safe in clinical practice. Congestive heart failure after the perfusion and the occurrence of post-operative infections are difficult to avoid when there are a long time of irrigation and more bleeding during operation.

[Treatment of ureteral obstruction by holmium: YAG laser endoureterotomy: a report of 18 cases].

OBJECTIVE: To investigate the clinical value and safety of holmium: YAG laser endoureterotomy in the treatment of ureteral obstruction. METHODS: Holmium: YAG laser endoureterotomy, with the laser optic fiber 550 microm in diameter and the output power of 3.5 Watt, via ureteroscopy, was performed on 18 patients ureteral obstruction, 8 males and 10 females, aged 52.1 (34-67), 11 with the stricture in the upper segment (complete obstruction in 4 cases), 5 in the middle segment, and 2 in lower segment; and 6 cases complicated with ureteral calculus. Postoperatively, an orthopedic ureteral stent (a 6-Fr double-J ureteral stent with a movable 5 cm length 9-Fr orthopedic cannula) was remained indwelling for 3-6 months. Follow-up was conducted for 10.7 (2-14) months. RESULTS: The operative duration was 32 (25-70) minutes. One patient underwent failed endoureterotomy and was turned to percutaneous nephroscopy. Success was achieved in 16 patients. The glomerular filtration rate (GFR) of these affected kidneys increased from 16.4+/-6.9 ml/min ante-operatively to 24.9+/-8.2 ml/min (P<0.01) postoperatively. One kidney was resected because of non-function, with GFR of 2 ml/min and intractable pyelitis. No recurrence of ureteral stricture was observed. CONCLUSION: Holmium: YAG laser endoureterotomy with insertion of orthopedic ureteral stent is an efficient and safe treatment for ureteral strictures with minimal invasion, less complications and easy recovery. This operation should be performed with a thorough preparation and severely restricted indication.

Mutational analysis of proto-oncogene Dbl on Xq27 in testicular germ cell tumors reveals a rare SNP in a patient with bilateral undescended testis.

OBJECTIVES: An abundance of X chromosomes in testicular germ cell tumors (TGCTs), and a candidate TGCTs susceptibility gene (TGCT1) on Xq27 highlight the potential involvement of X chromosomes in TGCT pathogenesis. However, the TGCT1 on Xq27 has so far not been identified. We hypothesized that a somatic mutation of dbl oncogene on Xq27 may play a role for the development of TGCTs. METHODS: We have screened 41 TGCT tissues for dbl mutations using single-strand conformation polymorphism (SSCP) analysis. These tissues are composed of 25 seminomatous TGCTs tissues and 16 non-seminomatous TGCTs tissues, including two cases with a rhabdomyosarcoma component. RESULTS: Somatic mutations were not detected in the 25 exons of dbl in these TGCTs. However, we found a rare single nucleotide polymorphism (SNP) (T to C nucleotide change) within intron 22 in one out of the 41 TGCTs cases (2%). Furthermore, the sample with the rare SNP was identified as the sole TGCTs case associated with bilateral undescended testis in our series. CONCLUSIONS: Our results indicate that proto-oncogene dbl is not a major target for sporadic TGCTs. However, the rare SNP in dbl may affect the susceptibility to undescended testis. Determining the frequency of this SNP in patients with various types of undescended testis in different ethnic groups is a warranted study.

[Optimization of extracting method and inhibitory action of recombinant human endostatin in bladder cancer].

OBJECTIVE: To explore the optimal method for protein expression in rhES (recombinant human endostatin) and study the anti-tumor activities of rhES in solid tumor and established cell line. METHODS: Different IPTG concentrations, the timing of adding IPTG into the culture medium and the different time of expression were employed to explore the optimized conditions of protein expression. Activity examination: (1) animal experiment: nude mice bearing subcutaneous cancer in treated group and controlled group were observed. (2) cellular experiment: the inhibitory effect of rhES in T-24 established cell line were observed by MTT assay and cancer cell growth curve. RESULTS: The expression of rhES protein was 58.65%. Of all the E. coli body proteins, the protein purity after purification was 96.22%. Activity examination indicated that rhES could inhibit the growth of solid tumor and the established cell line. In animal experiment, the tumor inhibition rate was 66.8%. Cell experiment: the 50% inhibitory concentration (IC(50)) was 22 microg/ml. The cell population decreased 58.75% than the control group at Day 7 in the tumor cell growth curve. CONCLUSION: A high expression and activity of rhES protein can be obtained by the optimized expression conditions. rhES can inhibit the cellular growth in both solid tumor and established cell line of bladder cancer.

[Endostatin in the treatment of the transplantable model of human laryngeal squamous carcinoma in nude mice].

OBJECTIVE: To evaluate the inhibitory effect of endostatin on tumor growth of human laryngeal squamous carcinoma in nude mice and to explore the possible mechanism of the inhibition and the possible way of biological therapy. METHODS: Nude mice model bearing laryngocarcinoma was established by using human laryngeal squamous carcinoma cell line ( Hep-II). The animals were given endostatin (20 mg x kg(-1) x d(-1)) or PBS, for 21 consecutive days. The volumes of the subcutaneous tumor were observed. The microstructure in which the general 2-step immuohistochemical examination was adopted and ultra-microstructural changes of carcinoma after administration of endostatin were observed under light and electron microscopes for pathology examination. RESULTS: The differences were statistically significant for the net mice weight, tumor weight, and tumor volume and weight/net mice weight between the treatment group and the control group. The restrained percentage of tumor was 45.9%. The necrosis and apoptosis of the tumor cell and the angiogenesis reduction were found under light and electron microscope in the treatment group. The expression of MVD, PCNA and VEGF of the treatment group is lower than that of the control group, and T test showed that P < 0.01, P < 0.05, P < 0.05 respectively, the differences were statistically significant. CONCLUSIONS: These studies showed that endostatin could significantly restrain the development of laryngocarcinoma. The mechanism may be due to the effect of antiangiogesis.