AURKA promotes renal cell carcinoma progression via regulation of CCNB1 transcription.

AURKA is a member of the serine/threonine kinase family and its kinase activity is crucial for the progression of mitosis. Recent studies have highlighted the therapeutic significance of AURKA inhibition in multiple cancer types. However, the specific mechanisms by which AURKA contributes to the progression of renal cell carcinoma (RCC) have not been fully elucidated. In this study, AURKA expression level was identified in human RCC tissues by immunohistochemical (IHC) staining. The function of AURKA on cell malignant phenotypes was evaluated in vitro after AURKA inhibition. The subcutaneous xenograft was conducted to confirm the in vivo effect of AURKA knockdown on growth of RCC cells. Finally, Co-IP, luciferase assay and ChIP experiments were performed to reveal the regulatory mechanism of AURKA on CCNB1. Our results showed a significant upregulation of AURKA in RCC tissues and cell lines, and a high AURKA expression was associated with poor prognosis. AURKA knockdown inhibited RCC cell proliferation and migration, induced cell apoptosis, and led to G1/G2 phase arrest. This effect was further confirmed by the use of an AURKA inhibitor. Mechanistically, AURKA interacted with E2F1, and subsequently recruited it to the promoter region of CCNB1. CCNB1 expression was essential for AURKA-induced RCC progression. Collectively, our results suggested that AURKA plays an important role in development of RCC via regulating CCNB1 transcription.

Single-cell analysis of human prepuce reveals dynamic changes in gene regulation and cellular communications.

BACKGROUND: The cellular and molecular dynamics of human prepuce are crucial for understanding its biological and physiological functions, as well as the prevention of related genital diseases. However, the cellular compositions and heterogeneity of human prepuce at single-cell resolution are still largely unknown. Here we systematically dissected the prepuce of children and adults based on the single-cell RNA-seq data of 90,770 qualified cells. RESULTS: We identified 15 prepuce cell subtypes, including fibroblast, smooth muscle cells, T/natural killer cells, macrophages, vascular endothelial cells, and dendritic cells. The proportions of these cell types varied among different individuals as well as between children and adults. Moreover, we detected cell-type-specific gene regulatory networks (GRNs), which could contribute to the unique functions of related cell types. The GRNs were also highly dynamic between the prepuce cells of children and adults. Our cell-cell communication network analysis among different cell types revealed a set of child-specific (e.g., CD96, EPO, IFN-1, and WNT signaling pathways) and adult-specific (e.g., BMP10, NEGR, ncWNT, and NPR1 signaling pathways) signaling pathways. The variations of GRNs and cellular communications could be closely associated with prepuce development in children and prepuce maintenance in adults. CONCLUSIONS: Collectively, we systematically analyzed the cellular variations and molecular changes of the human prepuce at single-cell resolution. Our results gained insights into the heterogeneity of prepuce cells and shed light on the underlying molecular mechanisms of prepuce development and maintenance.

Targeting the chromatin effector Pygo2 promotes cytotoxic T cell responses and overcomes immunotherapy resistance in prostate cancer.

The noninflamed microenvironment in prostate cancer represents a barrier to immunotherapy. Genetic alterations underlying cancer cell-intrinsic oncogenic signaling are increasingly appreciated for their role in shaping the immune landscape. Recently, we identified Pygopus 2 (PYGO2) as the driver oncogene for the amplicon at 1q21.3 in prostate cancer. Here, using transgenic mouse models of metastatic prostate adenocarcinoma, we found that Pygo2 deletion decelerated tumor progression, diminished metastases, and extended survival. Pygo2 loss augmented the activation and infiltration of cytotoxic T lymphocytes (CTLs) and sensitized tumor cells to T cell killing. Mechanistically, Pygo2 orchestrated a p53/Sp1/Kit/Ido1 signaling network to foster a microenvironment hostile to CTLs. Genetic or pharmacological inhibition of Pygo2 enhanced the antitumor efficacy of immunotherapies using immune checkpoint blockade (ICB), adoptive cell transfer, or agents inhibiting myeloid-derived suppressor cells. In human prostate cancer samples, Pygo2 expression was inversely correlated with the infiltration of CD8+ T cells. Analysis of the ICB clinical data showed association between high PYGO2 level and worse outcome. Together, our results highlight a potential path to improve immunotherapy using Pygo2-targeted therapy for advanced prostate cancer.

Targeted delivery of nuclear targeting probe for bladder cancer using cyclic pentapeptide c(RGDfK) and acridine orange.

PURPOSE: Both cyclic pentapeptide c(RGDfK) and acridine orange (AO) exhibit antitumor effects and cell permeability. This study aimed to evaluate the nuclear targeting efficiency and safety of the nuclear targeting probe for bladder cancer (BCa) synthesized by c(RGDfK) and AO. METHODS: The nuclear targeting probe AO-(cRGDfK)2 was synthesized from AO hydrochloride, azided c(RGDfK), and a near-infrared skeleton synthesized via click chemistry reactions. The effect of the AO-(cRGDfK)2 probe on cell viability was assessed in BCa 5637 cells. The tumor cell targeting efficacy of the AO-(cRGDfK)2 probe was evaluated in BCa cells in vitro and in tumor-bearing mice in vivo. Nuclear-specific accumulation of fluorescence probe in BCa tumor cells was evaluated using laser scanning confocal microscopy (LSCM). Hematoxylin and eosin staining was performed to detect histopathological changes in the spleen, heart, liver, and kidney. RESULTS: The AO-(cRGDfK)2 probe did not cause a significant reduction in cell viability. LSCM analysis showed that AO-(cRGDfK)2 exhibited nuclear-specific ambulation in BCa cells and was not accumulated in 293T cells. Also, this probe efficiently targeted tumor cells in the serum and urine samples. In vivo imaging system of tumor-bearing mice showed that ~ 80% percent of fluorescence signal was accumulated in the tumor sites. The probe did not change histopathology in the heart, liver, spleen, and kidney in tumor-bearing mice after the 21-day treatment. CONCLUSIONS: The AO-(cRGDfK)2 probe exhibited nuclear-specific accumulation in BCa cells without cytotoxicity, which provides an innovative alternative to improve anticancer therapy for BCa.

A novel cuproptosis-related lncRNA signature predicts prognosis and therapeutic response in bladder cancer.

Bladder cancer (BC) ranks the tenth in the incidence of global tumor epidemiology. LncRNAs and cuproptosis were discovered to regulate the cell death. Herein, we downloaded transcriptome profiling, mutational data, and clinical data on patients from The Cancer Genome Atlas (TCGA). High- and low-risk BC patients were categorized. Three CRLs (AL590428.1, AL138756.1 and GUSBP11) were taken into prognostic signature through least absolute shrinkage and selection operator (LASSO) Cox regression. Worse OS and PFS were shown in high-risk group (p < 0.05). ROC, independent prognostic analyses, nomogram and C-index were predicted via CRLs. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated IncRNAs play a biological role in BC progression. Immune-related functions showed the high-risk group received more benefit from immunotherapy and had stronger immune responses, and the overall survival was better (p < 0.05). Finally, a more effective outcome (p < 0.05) was found from clinical immunotherapy via the TIDE algorithm and many potential anti-tumor drugs were identified. In our study, the cuproptosis-related signature provided a novel tool to predict the prognosis in BC patients accurately and provided a novel strategy for clinical immunotherapy and clinical applications.

CHD1 and SPOP synergistically protect prostate epithelial cells from DNA damage.

BACKGROUND: Recent genomic profiling has identified a subtype of prostate cancer (PCa) characterized by two key genetic alterations: missense mutation of speckle-type POZ protein (SPOP) and homozygous deletion of chromodomain helicase DNA-binding protein 1 (CHD1). Mutually exclusive with E26 transformation-specific (ETS) rearrangements, this subtype displays high genomic instability. Previous studies indicate that deficient SPOP or CHD1 alone leads to feeble prostate abnormalities and each protein is involved in DNA damage response (DDR). It remains to be determined whether CHD1 and SPOP cooperate to suppress prostate tumorigenesis and DDR. METHODS: Prostate-specific single or double knockout of Spop and Chd1 was generated with the Cre/loxP system in mice. Wild-type or mutant SPOP (F102C, F133V) overexpression and CHD1 knockdown with short hairpin RNA were created in human benign prostatic hyperplasia cell line BPH1. The levels of DNA damage and homologous recombination repair were measured by immunofluorescence staining of γH2AX and RAD51, respectively. RESULTS: Spop/Chd1 double-knockout mice displayed prostatic intraepithelial neoplasia at both young (3 months) and old (12 months) ages and failed to generate prostate adenocarcinoma. Compared with wild-type or single-knockout mice, the double-knockout prostate harbored moderately higher proliferating cells and dramatically augmented the level of γH2AX staining, although androgen receptor-positive cells and apoptotic cells remained at a similar level. In BPH1 cell line, SPOP mutant overexpression and CHD1 silencing synergistically sensitized the cells to DNA damage by camptothecin, an inducer of double-strand breaks. CONCLUSIONS: Our results indicate that SPOP and CHD1 can synergistically promote repair of naturally occurring or chemically induced DNA damages in prostate epithelial cells. Regarding the progression of the SPOP/CHD1 subtype of PCa, other functionally complementary drivers warrant further identification. The clinical implication is that this subtype of PCa may be particularly sensitive to poly(ADP-ribose) polymerase inhibitors or DNA-damaging agents.

Polymorphonuclear MDSCs are enriched in the stroma and expanded in metastases of prostate cancer.

Myeloid-derived suppressor cells with polymorphonuclear morphology (PMN-MDSCs) contribute to the progression and immune evasion of prostate cancer. However, the spatial distribution of tumor-infiltrating PMN-MDSCs in primary and metastatic prostate cancer, especially in the context of comparison between the epithelial and stromal compartments of the tumor, has not been characterized. Here, we describe a multicolor immunofluorescence staining study of 90 primary tumors, 37 lymph node metastases (all with matched primary tumors) and 35 bone metastases using archived samples. CD11b+ CD15+ cells were identified as PMN-MDSCs and pan-cytokeratin+ cells were identified as prostate epithelial cells. We found that, in both primary tumor and metastases, PMN-MDSCs infiltrate much more readily in the stromal area compared with the epithelial area of the tumor regions. In comparison to the stromal area of primary tumors, the stromal area of either lymph node metastases or bone metastases was infiltrated with more PMN-MDSCs. In primary tumors, stromal PMN-MDSCs were associated with vascularization, segmented neutrophils, patient age and close juxtaposition to neoplastic epithelial cells. These results reveal the stroma rather than the epithelia of prostate cancer as the major hotbed for PMN-MDSCs and support the role of PMN-MDSCs in the metastatic progression of prostate cancer.

[Cefoxitin plus levofloxacin for prevention of severe infection after transrectal prostate biopsy].

OBJECTIVE: To evaluate the effect of cefoxitin prophylactic in reducing the incidence of severe infection after transrectal prostate biopsy (TRPB). METHODS: This retrospective study included 155 cases of TRPB with a 5-day administration of oral levofloxacin at 200 mg bid (the control group) and another 167 cases with a 3-day administration of oral levofloxacin at the same dose plus intravenous cefoxitin at 2.0 g 2 hours before TRPB (the experimental group) according to the distribution characteristics of drug-resistance bacteria in our department. The patients of the control and experimental groups were aged (68.68 ± 8.12) and (68.72 ± 7.51) years, with PSA levels of (19.78 ± 21.57) and (21.15 ± 42.63) µg/L, involving (11.68 ± 1.44) and (11.77±1.02) biopsy cores, respectively. Comparisons were made between the two groups of patients in the incidence rate of severe infection, which was defined as lower urinary track symptoms plus the systemic inflammatory response syndrome (SIRS) within 7 days after TRPB. RESULTS: The incidence rate of postoperative severe infection was significantly lower in the experimental group than in the control (0.6% ï¼»1/167ï¼½ vs 5.8% ï¼»9/155ï¼½, P < 0.05). Blood cultures revealed positive E-coli strains in 6 cases in the control group, including 5 ESBL-positive and 4 quinolone-resistant and amikacin-sensitive cases, all sensitive to cefoxitin, cefoperazone/sulbactam and imipenem. The only one case of severe infection was shown to be negative in blood culture. CONCLUSIONS: Preoperative intravenous administration of cefoxitin according to the specific distribution characteristics of drug-resistance bacteria can significantly reduce the incidence of severe infection after TRPB.

The PI3K/AKT pathway in the pathogenesis of prostate cancer.

Despite recent advances in our understanding of the biological behavior of prostate cancer (PCa), PCa is becoming the most common malignancy in men worldwide. The phosphatidylinositol 3-kinase (PI3K)/AKT pathway has been implicated in prostate carcinogenesis. Inflammatory cytokines (CCR9, IL-6, and TLR3) regulate PI3K/AKT signaling during apoptosis of PCa cells, and PI3K/AKT signaling participates with androgen-, 1alpha,25(OH)2-vitamin D3-, and prostaglandin-associated mechanisms and is regulated by ErbB, EGFR, and the HER family during cell growth. During metastasis of PCa cells, the PI3K/AKT/NF-kappaB/BMP-2-Smad axis, PTEN/PI3K/AKT pathway, and PI3K/AKT/mTOR signaling regulates tumor cell metastasis and invasion. The present review focuses on the PI3K/AKT signal pathway and discusses the role of the PI3K/AKT signal pathway in PCa tumorigenesis.

Gene expression profiling analysis of castration-resistant prostate cancer.

BACKGROUND: Prostate cancer is a global health issue. Usually, men with metastatic disease will progress to castration-resistant prostate cancer (CRPC). We aimed to identify the differentially expressed genes (DEGs) in tumor samples from non-castrated and castrated men from LNCaP Orthotopic xenograft models of prostate cancer and to study the mechanisms of CRPC. MATERIAL/METHODS: In this work, GSE46218 containing 4 samples from non-castrated men and 4 samples from castrated men was downloaded from Gene Expression Omnibus. We identified DEGs using limma Geoquery in R, the Robust Multi-array Average (RMA) method in Bioconductor, and Bias methods, followed by constructing an integrated regulatory network involving DEGs, miRNAs, and TFs using Cytoscape. Then, we analyzed network motifs of the integrated gene regulatory network using FANMOD. We selected regulatory modules corresponding to network motifs from the integrated regulatory network by Perl script. We preformed gene ontology (GO) and pathway enrichment analysis of DEGs in the regulatory modules using DAVID. RESULTS: We identified total 443 DEGs. We built an integrated regulatory network, found three motifs (motif 1, motif 2 and motif 3), and got two function modules (module 1 corresponded to motif 1, and module 2 corresponded to motif 2). Several GO terms (such as regulation of cell proliferation, positive regulation of macromolecule metabolic process, phosphorylation, and phosphorus metabolic process) and two pathways (pathway in cancer and Melanoma) were enriched. Furthermore, some significant DEGs (such as CAV1, LYN, FGFR3 and FGFR3) were related to CPRC development. CONCLUSIONS: These genes might play important roles in the development and progression of CRPC.

Regulation of growth of human bladder cancer by miR-192.

The regulation of microRNA-192 (miR-192) is impaired in many cancers. Here, we investigated the role of miR-192 in the proliferation, cell cycle progression, and apoptosis of bladder cancer cells. Human bladder cancer cells were transfected with human miR-192 precursor or non-specific control miRNA. The effect of miR-192 on cell proliferation was assessed by a MTT assay. The effects of miR-192 on cell cycle regulation and apoptosis were evaluated by flow cytometry. Western blot was used to analyze the protein levels of cyclin D1, p21, p27, Bcl-2, Bax, and Mcl-1. We found that overexpression of miR-192 significantly decreased the proliferation of bladder cancer cells by 22 and 54 % at 48 and 72 h, respectively. MiR-192-overexpressing cells exhibited a significant increase in G0/G1 phase and a significant decrease in S phase compared to the control miRNA-transfected cells. Moreover, overexpression of miR-192 significantly induced apoptotic death in bladder cancer cells, increased the levels of p21, p27, and Bax, and decreased the levels of cyclin D1, Bcl-2, and Mcl-1. Taken together, these data suggest that miR-192 may be a suppressor for bladder cancer cells by cell cycle regulation.

Dysregulation of cell cycle related genes and microRNAs distinguish the low- from high-risk of prostate cancer.

BACKGROUND: Prostate cancer (PCa) is a biologically heterogeneous disease with considerable variation in clinical aggressiveness. In this study, bioinformatics was used to detect the patterns of gene expression alterations of PCa patients. METHODS: The gene expression profile GSE21034 and GSE21036 were downloaded from Gene Expression Omnibus (GEO) database. Significantly changed mRNA transcripts and microRNAs were identified between subtypes with favorable (cluster 2) and unfavorable (cluster 5) prognosis by two-side unequal variances t test. MicroRNAs and their potential target genes were identified by TargetScan and miRTarBase, respectively. Besides, the overlapped genes between the target genes of microRNAs and mRNA transcripts were assessed by Fisher' exact test (one side). The functional annotation was performed by DAVID, followed by construction of protein-protein interaction (PPI) network. RESULTS: Compared to cluster 2, 1556 up-regulated and 1288 down-regulated transcripts were identified in cluster 5. Total 28 microRNAs were up-regulated and 30 microRNAs were down-regulated in cluster 5. Besides, 12 microRNAs target transcripts were significantly overlapped with down-regulated transcripts in cluster 5 with none of them was found overlapped with up-regulated transcripts. Functional annotation showed that cell cycle was the most significant function. In the PPI network, BRCA1, CDK1, TK1 and TRAF2 were hub protein of signature genes in cluster 5, and TGFBR1, SMAD2 and SMAD4 were hub proteins of signature gnens in cluster 2. CONCLUSIONS: Our findings raise the possibility that genes related with cell cycle and dysregulated miRNA at diagnosis might have clinical utility in distinguishing low- from high-risk PCa patients. VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/13000_2014_156.

Lipoteichoic acid (LTA) and lipopolysaccharides (LPS) from periodontal pathogenic bacteria facilitate oncogenic herpesvirus infection within primary oral cells.

Kaposi's sarcoma (KS) remains the most common tumor arising in patients with HIV/AIDS, and involvement of the oral cavity represents one of the most common clinical manifestations of this tumor. HIV infection incurs an increased risk for periodontal diseases and oral carriage of a variety of bacteria. Whether interactions involving pathogenic bacteria and oncogenic viruses in the local environment facilitate replication or maintenance of these viruses in the oral cavity remains unknown. In the current study, our data indicate that pretreatment of primary human oral fibroblasts with two prototypical pathogen-associated molecular patterns (PAMPs) produced by oral pathogenic bacteria-lipoteichoic acid (LTA) and lipopolysaccharide (LPS), increase KSHV entry and subsequent viral latent gene expression during de novo infection. Further experiments demonstrate that the underlying mechanisms induced by LTA and/or LPS include upregulation of cellular receptor, increasing production of reactive oxygen species (ROS), and activating intracellular signaling pathways such as MAPK and NF-κB, and all of which are closely associated with KSHV entry or gene expression within oral cells. Based on these findings, we hope to provide the framework of developing novel targeted approaches for treatment and prevention of oral KSHV infection and KS development in high-risk HIV-positive patients.

Chrebp regulates the transcriptional activity of androgen receptor in prostate cancer.

Androgen receptor (AR), a member of nuclear hormone receptor, plays an essential role in the initiation and progression of prostate cancer (PCa). In the present study, by way of immunoprecipitation followed by mass spectrometry (IP/MS) system, we found that carbohydrate-responsive element-binding protein (Chrebp), a glucose sensor in normal and cancer cells, interacted with AR in LNCaP cells. The interaction was further confirmed by coimmunoprecipitation analysis. Besides, Chrebp is required for the optimal transcriptional activity of AR in promoting the transcription of the prostate-specific antigen (PSA) promoter and messenger RNA (mRNA) expression. Consistently, knockdown of Chrebp using small interfering RNA (siRNA) in LNCaP cells reduced endogenous PSA levels. Together, our study demonstrates that Chrebp interacts with AR and regulates its transcriptional activity.

Lentivirus-mediated SMO RNA interference inhibits SMO expression and cell proliferation, and affects the cell cycle in LNCaP and PC3 cancer cell lines.

Smoothened (SMO) is an important member of the Hedgehog signaling pathway. We constructed a specific recombinant lentiviral vector for RNA interference, targeting the SMO gene (NM_005631) to observe its effect on SMO expression, cell proliferation and the cell cycle in the human androgen-sensitive prostate cancer cell line, LNCaP, and in the androgen-independent prostate cancer cell line, PC3. Four siRNA sequences were designed and inserted into a lentiviral vector pGCSIL-GFP to construct four recombinant vectors. The vector with the highest interfering efficiency was co-transfected with packaging vectors (pHelper1.0 and pHelper2.0) in 293T cells to assemble lentivirus particles by liposome for infecting LNCaP and PC3 cell lines, respectively. The expression level of SMO mRNA, tumor cell proliferation and cell cycle were measured by quantitative realtime polymerase chain reaction (qRT-PCR), 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide (MTT) assay and flow cytometry, respectively. Sequence results showed that recombinant lentiviral vectors were constructed successfully. pGCSIL-GFP-723 had the highest interfering efficiency, named Lv-SIL-SMO723 after co-transfection, with which LNCaP and PC3 cell lines were infected. Compared with the control groups, results showed significantly decreased (P < 0.05) SMO mRNA expressions of LNCaP and PC3, lower mean percentage of S-phase cells and higher mean percentage of G(2)/M phase cells, as well as obviously slow proliferation (P < 0.01) of LNCaP in the infected group. Yet, the proliferation of PC3 was not altered (P > 0.05). In conclusion, the recombinant lentivirus particles were able to suppress SMO expression, regulate the cell cycle in the LNCaP and PC3 cell lines and markedly inhibit proliferation of LNCaP cells but not PC3 cells.

Clinical value of minimally invasive percutaneous nephrolithotomy in the supine position under the guidance of real-time ultrasound: report of 92 cases.

To determine the clinical value of the real-time-ultrasound-guided minimally invasive percutaneous nephrolithotomy (m-PCNL) technique in the supine position, 92 patients suffering from renal or upper ureteral stones were treated by m-PCNL with a nephroscope/ureteroscope in the supine position. The ipsilateral flanks of the patients with different body sizes were elevated with a 1,000 or 3,000-ml water bag. Under cystoscopy, a ureteral catheter was inserted into the kidney. Normal saline was infused into the kidney via the ureteral catheter to dilate the entire urinary system. Under the guidance of real-time ultrasound, the needle was inserted into the urinary system to dilate the tract and establish the 16F mini-tract for percutaneous nephrolithotomy. All 92 (100%) m-PCNL procedures were successfully performed in the supine position. Primary stone clearance was achieved in 64 cases (69.6%). Residual stones occurred in 28 cases (30.4%). M-PCNL was performed for a second time in 16 cases to clear the residual stones. In 4 cases, stones remained after the second m-PCNL. Two of them were treated further by extracorporeal shockwave lithotripsy (ESWL). The total stone clearance rate of m-PCNL was 82.6%. Only one case required blood transfusion. No other serious complications occurred. The supine position is a favorable position for the patients, the surgeons and the anesthesiologists during the m-PCNL procedure. Real-time ultrasound is a valuable technique for guiding of the m-PCNL.