Development of a PAK4-targeting PROTAC for renal carcinoma therapy: concurrent inhibition of cancer cell proliferation and enhancement of immune cell response.

BACKGROUND: Finding the oncogene, which was able to inhibit tumor cells intrinsically and improve the immune answers, will be the future direction for renal cancer combined treatment. Following patient sample analysis and signaling pathway examination, we propose p21-activated kinase 4 (PAK4) as a potential target drug for kidney cancer. PAK4 exhibits high expression levels in patient samples and plays a regulatory role in the immune microenvironment. METHODS: Utilizing AI software for peptide drug design, we have engineered a specialized peptide proteolysis targeting chimera (PROTAC) drug with selectivity for PAK4. To address challenges related to drug delivery, we developed a nano-selenium delivery system for efficient transport of the peptide PROTAC drug, termed PpD (PAK4 peptide degrader). FINDINGS: We successfully designed a peptide PROTAC drug targeting PAK4. PpD effectively degraded PAK4 with high selectivity, avoiding interference with other homologous proteins. PpD significantly attenuated renal carcinoma proliferation in vitro and in vivo. Notably, PpD demonstrated a significant inhibitory effect on tumor proliferation in a fully immunocompetent mouse model, concomitantly enhancing the immune cell response. Moreover, PpD demonstrated promising tumor growth inhibitory effects in mini-PDX and PDO models, further underscoring its potential for clinical application. INTERPRETATION: This PAK4-targeting peptide PROTAC drug not only curtails renal cancer cell proliferation but also improves the immune microenvironment and enhances immune response. Our study paves the way for innovative targeted therapies in the management of renal cancer. FUNDING: This work is supported by Research grants from non-profit organizations, as stated in the Acknowledgments.

Development of a Double-Stapled Peptide Stabilizing Both α-Helix and ß-Sheet Structures for Degrading Transcription Factor AR-V7.

Peptide drugs offer distinct advantages in therapeutics; however, their limited stability and membrane penetration abilities hinder their widespread application. One strategy to overcome these challenges is the hydrocarbon peptide stapling technique, which addresses issues such as poor conformational stability, weak proteolytic resistance, and limited membrane permeability. Nonetheless, while peptide stapling has successfully stabilized α-helical peptides, it has shown limited applicability for most ß-sheet peptide motifs. In this study, we present the design of a novel double-stapled peptide capable of simultaneously stabilizing both α-helix and ß-sheet structures. Our designed double-stapled peptide, named DSARTC, specifically targets the androgen receptor (AR) DNA binding domain and MDM2 as E3 ligase. Serving as a peptide-based PROTAC (proteolysis-targeting chimera), DSARTC exhibits the ability to degrade both the full-length AR and AR-V7. Molecular dynamics simulations and circular dichroism analysis validate the successful constraint of both secondary structures, demonstrating that DSARTC is a "first-in-class" heterogeneous-conformational double-stapled peptide drug candidate. Compared to its linear counterpart, DSARTC displays enhanced stability and an improved cell penetration ability. In an enzalutamide-resistant prostate cancer animal model, DSARTC effectively inhibits tumor growth and reduces the levels of both AR and AR-V7 proteins. These results highlight the potential of DSARTC as a more potent and specific peptide PROTAC for AR-V7. Furthermore, our findings provide a promising strategy for expanding the design of staple peptide-based PROTAC drugs, targeting a wide range of "undruggable" transcription factors.

MYC induces CDK4/6 inhibitors resistance by promoting pRB1 degradation.

CDK4/6 inhibitors (CDK4/6i) show anticancer activity in certain human malignancies, such as breast cancer. However, their application to other tumor types and intrinsic resistance mechanisms are still unclear. Here, we demonstrate that MYC amplification confers resistance to CDK4/6i in bladder, prostate and breast cancer cells. Mechanistically, MYC binds to the promoter of the E3 ubiquitin ligase KLHL42 and enhances its transcription, leading to RB1 deficiency by inducing both phosphorylated and total pRB1 ubiquitination and degradation. We identify a compound that degrades MYC, A80.2HCl, which induces MYC degradation at nanomolar concentrations, restores pRB1 protein levels and re-establish sensitivity of MYC high-expressing cancer cells to CDK4/6i. The combination of CDK4/6i and A80.2HCl result in marked regression in tumor growth in vivo. Altogether, these results reveal the molecular mechanisms underlying MYC-induced resistance to CDK4/6i and suggest the utilization of the MYC degrading molecule A80.2HCl to potentiate the therapeutic efficacy of CDK4/6i.

PrLZ regulates EMT and invasion in prostate cancer via the TGF-ß1/p-smad2/miR-200 family/ZEB1 axis.

BACKGROUND: Prostate leucine zipper (PrLZ) is a prostate-specific protein, and our previous study demonstrated that PrLZ enhances the malignant progression of prostate cancer (Pca). However, the roles of PrLZ in epithelial to mesenchymal transition (EMT) remain unknown. METHODS: Quantitative real-time PCR (qRT-PCR), immunohistochemical (IHC) staining, hematoxylin-eosin (HE) staining, and western blotting were used to analyze the expression of protein and genes level in human PCa cell lines. Invasion assay was used to examine the effect of PrLZ, miR-200a, miR-200b, miR-200c, miR-141, miR-429, miR-205, and ZEB1 on PCa cell line invasion in vitro. Prostate cancer metastasis animal model was designed to assess the effect of PrLZ on PCa cell line invasion in vivo. RESULTS: We proved that high PrLZ expression initiates EMT, which was shown by the downregulation of E-cadherin and upregulation of vimentin in PC-3/PrLZ and ARCaP-E/PrLZ cells. Mechanistic analysis revealed that PrLZ regulates EMT by activating TGF-ß1/p-smad2 signaling and further inhibiting the expression of miR-200 family members, which negatively regulates ZEB1 expression and causes EMT in Pca. Moreover, using two of orthotopic mouse model and tail vein injection of human prostate cancer cells mouse model, we observed that PC-3/PrLZ cells led to the development of distant organ metastases in vivo. CONCLUSIONS: Our results show the mechanism by which PrLZ regulates EMT and metastasis and suggest that PrLZ may be a potential therapeutic target for Pca metastasis.

Knocking down AR promotes osteoblasts to recruit prostate cancer cells by altering exosomal circ-DHPS/miR-214-3p/CCL5 pathway.

ABSTRACT: Tumor-derived exosomes have been shown to play a key role in organ-specific metastasis, and the androgen receptor regulates prostate cancer (PCa) progression. It is unclear whether the androgen receptor regulates the recruitment of prostate cancer cells to the bone microenvironment, even bone metastases, through exosomes. Here, we found that exosomes isolated from PCa cells after knocking down androgen receptor (AR) or enzalutamide treatment can facilitate the migration of prostate cancer cells to osteoblasts. In addition, AR silencing or treatment with the AR antagonist enzalutamide may increase the expression of circular RNA-deoxyhypusine synthase (circ-DHPS) in PCa cells, which can be transported to osteoblasts by exosomes. Circ-DHPS acts as a competitive endogenous RNA (ceRNA) against endogenous miR-214-3p to promote C-C chemokine ligand 5 (CCL5) levels in osteoblasts. Increasing the level of CCL5 in osteoblasts could recruit more PCa cells into the bone microenvironment. Thus, blocking the circ-DHPS/miR-214-3p/CCL5 signal may decrease exosome-mediated migration of prostate cancer cells to osteoblasts.

EZH2-regulated immune risk score prognostic model predicts outcome of clear cell renal cell carcinoma.

Background: The enhancer of zeste homolog 2 (EZH2) plays an important role in the tumor microenvironment (TME), and EZH2 in shaping the epigenetic landscape of CD8+ T cell fate and function, with a particular emphasis on cancer. Here, high EZH2 expression always leads to less CD8+ T cell infiltration. However, clear cell renal cell carcinoma (ccRCC) is reportedly a "hot" tumor, with contradictory high EZH2 expression. Our goal was to construct a EZH2-regulated immune risk score prognostic model to predict ccRCC outcomes, and provide a prospect of clinical EZH2 inhibitors in fine-tuning T cell responses with immune therapy. Methods: We downloaded and analyzed The Cancer Genome Atlas (TCGA), Cancer Cell Line Encyclopedia (CCLE), TISIDB database, and WebGestalt for ccRCC patients, EZH2-related tumor-infiltrating lymphocytes and immunomodulators. R packages "limma", "BiocManager", and "preprocessCore", etc. were downloaded to prepare CIBERSORT files, immune cells heatmap, multivariable Cox model and survival analysis. The EZH2-regulated immune risk model's prognostic ability was calculated by receiver operating characteristic (ROC) and area under the curve (AUC) analyses in R studio. Results: EZH2 was highly expressed and related to poor outcome in ccRCC. However, high-expression EZH2 was not related to a "cool" tumor. Of the 49 immunomodulators significantly regulated by EZH2, forest plot showed 26 immunomodulators signatures independently associated with overall survival. The EZH2-regulated immune-risk score prognostic model was an independent prognostic factor (AUC =0.816), especially combined with clinicopathologic parameters in ccRCC overall survival prediction. Conclusions: The EZH2-regulated immune-risk score prognostic model was an independent prognostic factor, with good accuracy and predictability, and could provide experimental data to the clinical area.

CHD1 deletion stabilizes HIF1α to promote angiogenesis and glycolysis in prostate cancer.

Chromodomain-helicase-DNA-binding protein 1 (CHD1) deletion is among the most common mutations in prostate cancer (PCa), but its role remains unclear. In this study, RNA sequencing was conducted in PCa cells after clustered regularly interspaced palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9)-based CHD1 knockout. Gene set enrichment analysis (GSEA) indicated upregulation of hypoxia-related pathways. A subsequent study confirmed that CHD1 deletion significantly upregulated hypoxia-inducible factor 1α (HIF1α) expression. Mechanistic investigation revealed that CHD1 deletion upregulated HIF1α by transcriptionally downregulating prolyl hydroxylase domain protein 2 (PHD2), a prolyl hydroxylase catalyzing the hydroxylation of HIF1α and thus promoting its degradation by the E3 ligase von Hippel-Lindau tumor suppressor (VHL). Functional analysis showed that CHD1 deletion promoted angiogenesis and glycolysis, possibly through HIF1α target genes. Taken together, these findings indicate that CHD1 deletion enhances HIF1α expression through PHD2 downregulation and therefore promotes angiogenesis and metabolic reprogramming in PCa.

De Novo Design of an Androgen Receptor DNA Binding Domain-Targeted peptide PROTAC for Prostate Cancer Therapy.

Androgen receptor splice variant-7 (AR-V7), one of the major driving factors, is the most attractive drug target in castration-resistant prostate cancer (CRPC). Currently, no available drugs efficiently target AR-V7 in clinical practice. The DNA binding domain (DBD) is indispensable for the transcriptional activity of AR full length and AR splice variants, including AR-V7. Based on the homodimerization structure of the AR DBD, a novel peptide-based proteolysis-targeting chimera (PROTAC) drug is designed to induce AR and AR-V7 degradation in a DBD and MDM2-dependent manner, without showing any activity on other hormone receptors. To overcome the short half-life and poor cell penetrability of peptide PROTAC drugs, an ultrasmall gold (Au)-peptide complex platform to deliver the AR DBD PROTAC in vivo is developed. The obtained Au-AR pep-PROTAC effectively degrades AR and AR-V7 in prostate cancer cell lines, particularly in CWR22Rv1 cells with DC50 values 48.8 and 79.2 nM, respectively. Au-AR pep-PROTAC results in suppression of AR levels and induces tumor regression in both enzalutamide sensitive and resistant prostate cancer animal models. Further optimization of the Au-AR pep-PROTAC can ultimately lead to a new therapy for AR-V7-positive CRPC.

Serum Metabolomic Patterns in Patients With Aldosterone-Producing Adenoma.

Aldosterone-producing adenoma (APA), the main cause of endocrine hypertension, has recently been reported to be associated with other diseases, such as metabolic syndrome, but the detailed mechanism underlying this association remains unclear. Here, we used untargeted metabolomics and compared the abundance of serum metabolites between essential hypertension (EHT) and APA patients, as well as the serum metabolites of APA patients before and after adrenalectomy. Our results revealed 44 differential metabolites between APA and EHT patients and 39 differential metabolites between pre- and postoperative APA patients. Several metabolites involved in cardiovascular disease, obesity, and diabetes were dysregulated in APA patients compared to EHT patients, including arachidonic acid metabolites [e.g., 5(S)-HpETE and 12-HETE], amino acids (e.g., L-carnitine, taurine, and L-arginine), nucleotide metabolites (e.g., hypoxanthine) and cholesterol 3-sulfate. Importantly, the levels of hypoxanthine and cholesterol 3-sulfate, two metabolites that promote the development of atherosclerotic lesions and obesity, were originally increased in APA patients, but those elevated levels were reversed by adrenalectomy. Conversely, levels of L-carnitine and (3-carboxypropyl) trimethylammonium cation, two metabolites participating in lipid metabolism, were decreased in APA patients but increased postoperatively. We conclude that APA might participate in cardiovascular and metabolic diseases by regulating serum metabolites.

Acetylation-dependent regulation of TPD52 isoform 1 modulates chaperone-mediated autophagy in prostate cancer.

Aberrant chaperone-mediated autophagy (CMA) activation has been suggested as a tumorigenesis-promoting event in various cancers, although its roles in prostate cancer (PCa) remain elusive. Emerging evidence indicates that TPD52 isoform 1, a prostate-specific and androgen-responsive gene, contributes to the malignant progression of PCa. Here, we demonstrate that TPD52 enhances CMA activation by interacting with HSPA8/HSC70 and enhancing substrate degradation in PCa. Elevation of TPD52 is essential for CMA-induced PCa cell proliferation and stress resistance in vitro and in vivo. Furthermore, TPD52 is acetylated by KAT2B at K163, which is a process that can be antagonized by HDAC2. Inactivation of HDAC2 results in elevated TPD52 acetylation, which compromises the interaction between TPD52 and HSPA8, leading to impaired CMA function and tumor growth in vivo. Taken together, our findings reveal that acetylation-dependent regulation of TPD52 modulates CMA oncogenic function in PCa, thereby suggesting the possibility of targeting the TPD52-mediated CMA pathway to control the progression of PCa.Abbreviations: CMA: chaperone-mediated autophagy; HDAC2: histone deacetylase 2; HSPA8/HSC70: heat shock protein family A (Hsp70) member 8; KAT2B: lysine acetyltransferase 2B; LAMP2A: lysosomal associated membrane protein 2A; PCa: prostate cancer; TPD52: tumor protein D52.

Inhibition of Androgen Receptor Signaling Promotes Prostate Cancer Cell Migration via Upregulation of Annexin A1 Expression.

BACKGROUND: Recent studies indicate that androgen deprivation therapy (ADT), the main therapeutic approach for metastatic prostate cancer (PCa), accelerates PCa invasion and metastasis. Annexin A1 (ANXA1) is a Ca2+-regulated phospholipid-binding protein that can promote PCa migration and invasion. AIM OF THE STUDY: The aim of this study is to determine whether ANXA1 is regulated by ADT and participates in PCa progression after ADT, and to explore the possible mechanism of ANXA1-mediated PCa migration. METHODS: Expression of ANXA1 and androgen receptor (AR) in PCa cell lines and tissues was detected, and the association between these two proteins were analyzed. Expression of ANXA1 was evaluated after AR knockdown or AR inhibition in PCa cell lines. Cell migration of PCa cell liness after ANXA1 knockdown or overexpression was determined by in vitro migration assay. Transcriptome analysis was used to explore the possible mechanism of ANXA1-mediated PCa migration. RESULTS: ANXA1 expression in PCa cell lines and tissues was reversely associated with AR. In vitro studies revealed an increase in ANXA1 expression after AR knockdown or treatment with AR antagonist. Moreover, functional assays indicated that ANXA1 knockdown in PCa cells significantly inhibited cell migration, while ANXA1 overexpression in PCa cells significantly accelerated cell migration. Transcriptome analysis showed that ANXA1 regulated multiple genes involved in cell junction organization, such as CADM1, LIMCH1 and PPM1F. CONCLUSIONS: Our results indicate that ADT might accelerate PCa metastasis via ANXA1 expression and PCa cell migration.

CCL5-dependent mast cell infiltration into the tumor microenvironment in clear cell renal cell carcinoma patients.

We investigated the mechanisms affecting tumor progression and survival outcomes in Polybromo-1-mutated (PBRM1MUT) clear cell renal cell carcinoma (ccRCC) patients. PBRM1MUT ccRCC tissues contained higher numbers of mast cells and lower numbers of CD8+ and CD4+ T cells than tissues from PBRM1WT ccRCC patients. Hierarchical clustering, pathway enrichment and GSEA analyses demonstrated that PBRM1 mutations promote tumor progression by activating hypoxia inducible factor (HIF)-related signaling pathways and increasing expression of vascular endothelial growth factor family genes. PBRM1MUT ccRCC tissues also show increased expression of C-C motif chemokine ligand 5 (CCL5). PBRM1-silenced ccRCC cells exhibited greater Matrigel tube formation and cell proliferation than controls. In addition, HMC-1 human mast cells exhibited CCL5-dependent in vitro migration on Transwell plates. High CCL5 expression in PBRM1MUT ccRCC patients correlated with increased expression of genes encoding IFN-γ, IFN-α, IL-6, JAK-STAT3, TNF-α, and NF-ΚB. Moreover, high CCL5 expression was associated with poorer survival outcomes in ccRCC patients. These findings demonstrate that CCL5-dependent mast cell infiltration promotes immunosuppression within the tumor microenvironment, resulting in tumor progression and adverse survival outcomes in PBRM1MUT ccRCC patients.

Global, regional, and national burdens of bladder cancer in 2017: estimates from the 2017 global burden of disease study.

BACKGROUND: The aim of this study is to describe the prevalence and mortality of bladder cancer (BCa) using data obtained in the Global Burden of Disease study performed in 2017 (GBD 2017). METHODS: Data on BCa for 2017, including prevalence, mortality, and disability-adjusted life years (DALYs), were obtained from GBD 2017 at the global, regional, and national levels. We also analyzed the association of BCa burden with the country development level. RESULTS: There were 2.63 million BCa cases estimated from the GBD 2017 data, with 200,000 persons dying of BCa, resulting in 3.60 million DALYs in 2017. The age-standardized prevalence (ASP) of BCa was 32.91/100,000 persons, and age-standardized death rate (ASDR) was 2.57/100,000 persons. The ASP and ASDR of BCa were higher in males than in females, and higher in people older than 60 years. The ASP and ASDR of BCa were higher in Western Europe and Central Europe than in South Asia, Andean Latin America, and Central Latin America, and higher in countries with a higher sociodemographic index (SDI). Correlation analysis identified that the ASP and ASDR of BCa were positively correlated with the country SDI (P < 0.0001 and ρ = 0.68 for ASP, and P = 0.0048 and ρ = 0.20 for ASDR). In addition, 33.72% deaths and 36.80% DALYs caused by BCa could be attributed to smoking globally. CONCLUSION: The prevalence and mortality of BCa were very high in 2017, especially in high-SDI countries. Smoking-cessation strategies should be strengthened to control the burden associated with BCa.

Exosome-mediated miR-9-5p promotes proliferation and migration of renal cancer cells both in vitro and in vivo by targeting SOCS4.

Exosomes secreted by cancer cells play important roles in tumor progression by interacting with cell receptors. Renal cancer derived exosomes contain miRNAs which are associated with cell proliferation and invasion. Micro RNA 9-5 (miR-9-5) is highly expressed in the serum of renal cancer patients with advanced (tumor size - node - metastasis) TNM stage and Fuhrman grade. miR-9-5p is extensively expressed in exosomes derived from renal cancer cells. Overexpression of miR-9-5p promotes proliferation and invasion of A-704 (a cancer cell line of human kidney) cells via targeting and deregulating SOCS4 mRNA. Inhibition of the Janus kinase (JAK)/signaling transducer and activator of transcription (STAT) pathway by SOCS4 will be reduced, which leads to phosphorylation of STAT3 and JAK. Activated cytokine signaling promotes cell proliferation and invasion, and inhibits apoptosis. Moreover, overexpression of SOCS4 reduces miR-9-5p levels and plays an opposite role in cell. To conclude, exosomal miR-9-5p plays important roles in renal cancer both in vivo and in vitro, indicating it may be used as biomarker for diagnosis and for monitoring the efficacy if therapy.

Snail promotes prostate cancer migration by facilitating SPOP ubiquitination and degradation.

Prostate cancer (PCa) is the second leading cause of cancer-associated mortality in men. Speckle-type pox virus and zinc finger protein (SPOP), the most frequently mutated gene in PCa, functions as a tumor suppressor via degradation of cancer-promoting substrates. However, its upstream regulation in PCa metastasis remains poorly determined. Here, in a Snail-induced metastatic PCa model, we observed an accelerated degradation of SPOP protein in cells, which is crucial for the PCa migration and activation of the AKT signaling pathway. Mechanistically, we demonstrated that binding to Snail promoted SPOP ubiquitination and degradation. Moreover, the bric-a-brac/tramtrack/broad complex (BTB) domain of SPOP is turned out to be essential for Snail-mediated SPOP degradation. Thus, our findings reveal a post-translational level regulation of SPOP expression that facilitates the metastasis of PCa cells.

DARS-AS1 promotes clear cell renal cell carcinoma by sequestering miR-194-5p to up-regulate DARS.

Clear cell renal cell carcinoma (ccRCC), the most frequent subtype of renal cell carcinoma (RCC), is characterized by high relapse rate and mortality. Long non-coding RNAs (lncRNAs) are critical participants during cancer development. LncRNA DARS antisense RNA 1 (DARS-AS1), a newly-found lncRNA, is not specifically reported in ccRCC. However, Gene Expression Profiling Interactive Analysis (GEPIA) and starBase databases revealed the up-regulation of DARS-AS1 in ccRCC. Current study investigated the function and mechanism of DARS-AS1 in ccRCC. Functional assays including colony formation assay, EdU assay, caspase-3 activity detection, flow cytometry analysis and JC-1 assay were implemented to identify the role of DARS-AS1 in ccRCC. As a result, silencing of DARS-AS1 retarded proliferation and facilitated apoptosis in ccRCC cells. Moreover, mainly a cytoplasmic localization of lncRNA DARS-AS1 was verified in ccRCC cells. Then, we demonstrated that DARS-AS1 positively regulated its nearby gene, aspartyl-tRNA synthetase (DARS), by sequestering miR-194-5p. Moreover, DARS was testified as the oncogene in ccRCC and DARS-AS1 worked as a tumor-facilitator in ccRCC through miR-194-5p/DARS signaling. In a summary, this study firstly uncovered that DARS-AS1 boosted DARS expression via absorbing miR-194-5p, therefore contributing to malignancy in ccRCC. Our findings may be helpful for opening new strategies for ccRCC treatment.

Decreased glucose bioavailability and elevated aspartate metabolism in prostate cancer cells undergoing epithelial-mesenchymal transition.

Prostate cancer (PCa) is a common malignancy with a high tendency for metastasis. Epithelial-mesenchymal transition (EMT) plays a crucial role in PCa metastasis. Metabolic reprogramming offers metabolic advantages for cancer development and could result in the discovery of novel targets for cancer therapy. However, the metabolic features of PCa cells undergoing EMT remain unclear. We used metabolome and transcriptome analyses and found that PCa cells undergoing EMT showed impaired glucose utilization. In vitro studies demonstrated that PCa cells undergoing EMT were less addicted to glucose than epithelial-like PCa cells. However, cells that underwent EMT had higher levels of aspartate and its downstream metabolites, indicative of upregulated aspartate metabolism. As aspartate is a contributor for EMT and metastasis in human cancer cells, we conclude that this metabolic reprogramming may play a vital role in EMT and PCa progression.

Androgen receptor suppresses prostate cancer metastasis but promotes bladder cancer metastasis via differentially altering miRNA525-5p/SLPI-mediated vasculogenic mimicry formation.

Early studies suggest that the androgen receptor (AR) may play differential roles in influencing prostate cancer (PCa) and bladder cancer (BCa) metastasis, but the underlying mechanisms remain unclear. Here, we found that the AR might function via differentially altering vasculogenic mimicry (VM) formation to either decrease PCa metastasis or increase BCa metastasis. Mechanism dissection showed that the AR could differentially alter the expression of the VM marker SLPI through miR-525-5p to regulate SLPI; moreover, it could either increase miR-525-5p transcription in PCa or decrease it in BCa via binding to different androgen-response-elements (AREs) located at different positions in the miR-525 precursor promoter. Further, results from liquid chromatography-mass spectrometry (LC-MS) showed that the co-factors of AR in PCa and BCa are NFIX and HDAC2, respectively. Together, these results provide the first detailed mechanism of how the AR can differentially alter PCa and BCa metastasis; thus, targeting the newly identified AR-miR-525-5p-SLPI axis may help suppress metastasis.

E-cadherin is downregulated in benign prostatic hyperplasia and required for tight junction formation and permeability barrier in the prostatic epithelial cell monolayer.

BACKGROUND: We previously reported the presence of prostate-specific antigen (PSA) in the stromal compartment of benign prostatic hyperplasia (BPH). Since PSA is expressed exclusively by prostatic luminal epithelial cells, PSA in the BPH stroma suggests increased tissue permeability and the compromise of epithelial barrier integrity. E-cadherin, an important adherens junction component and tight junction regulator, is known to exhibit downregulation in BPH. These observations suggest that the prostate epithelial barrier is disrupted in BPH and E-cadherin downregulation may increase epithelial barrier permeability. METHODS: The ultra-structure of cellular junctions in BPH specimens was observed using transmission electron microscopy (TEM) and E-cadherin immunostaining analysis was performed on BPH and normal adjacent specimens from BPH patients. In vitro cell line studies using benign prostatic epithelial cell lines were performed to determine the impact of small interfering RNA knockdown of E-cadherin on transepithelial electrical resistance and diffusion of fluorescein isothiocyanate (FITC)-dextran in transwell assays. RESULTS: The number of kiss points in tight junctions was reduced in BPH epithelial cells as compared with the normal adjacent prostate. Immunostaining confirmed E-cadherin downregulation and revealed a discontinuous E-cadherin staining pattern in BPH specimens. E-cadherin knockdown increased monolayer permeability and disrupted tight junction formation without affecting cell density. CONCLUSIONS: Our results indicate that tight junctions are compromised in BPH and loss of E-cadherin is potentially an important underlying mechanism, suggesting targeting E-cadherin loss could be a potential approach to prevent or treat BPH.

G Protein γ subunit 7 loss contributes to progression of clear cell renal cell carcinoma.

Clear cell renal cell carcinoma (ccRCC) is a common urinary neoplasm, looking for useful candidates to establish scientific foundation for the therapy of ccRCC is urgent. We downloaded genomic profiles of GSE781, GSE6244, GSE53757, and GSE66271 from the Gene Expression Omnibus (GEO) database. GEO2R was used to analyze the derivative genes, while hub genes were screened by protein-protein interactions and cytoscape. Further, overall survival, gene methylation, gene mutation, and gene expression were all analyzed using bioinformatics tools. Colony formation and cell-cycle assay were used to detect the biological function of GNG7 in vitro. We found that GNG7 was downregulated in ccRCC tissues and negatively associated with overall survival in ccRCC patients. We also found that promoter methylation and frequent gene mutation were responsible for GNG7 gene suppression. GNG7 low expression was related to upregulation of enhancer of zeste homolog 2 and downregulation of disabled homolog 2-interacting protein. Further, Gene Set Enrichment Analysis results showed that mTOR1, E2F, G2M, and MYC pathways were all significantly altered in response to GNG7 low expression. In vitro, A498 and 786-O cells in which GNG7 expression was silenced, exhibited a lower G1 phase when compared to the negative control cells. Taken together, our findings suggest that GNG7 is a tumor suppressor gene in ccRCC progression and represents a novel candidate for ccRCC treatment.