Phosphorylated RB Promotes Cancer Immunity by Inhibiting NF-κB Activation and PD-L1 Expression.

Aberrant expression of programmed death ligand-1 (PD-L1) in tumor cells promotes cancer progression by suppressing cancer immunity. The retinoblastoma protein RB is a tumor suppressor known to regulate the cell cycle, DNA damage response, and differentiation. Here, we demonstrate that RB interacts with nuclear factor κB (NF-κB) protein p65 and that their interaction is primarily dependent on CDK4/6-mediated serine-249/threonine-252 (S249/T252) phosphorylation of RB. RNA-seq analysis shows a subset of NF-κB pathway genes including PD-L1 are selectively upregulated by RB knockdown or CDK4/6 inhibitor. S249/T252-phosphorylated RB inversely correlates with PD-L1 expression in patient samples. Expression of a RB-derived S249/T252 phosphorylation-mimetic peptide suppresses radiotherapy-induced upregulation of PD-L1 and augments therapeutic efficacy of radiation in vivo. Our findings reveal a previously unrecognized tumor suppressor function of hyperphosphorylated RB in suppressing NF-κB activity and PD-L1 expression and suggest that the RB-NF-κB axis can be exploited to overcome cancer immune evasion triggered by conventional or targeted therapies.

EZH2 cooperates with gain-of-function p53 mutants to promote cancer growth and metastasis.

In light of the increasing number of identified cancer-driven gain-of-function (GOF) mutants of p53, it is important to define a common mechanism to systematically target several mutants, rather than developing strategies tailored to inhibit each mutant individually. Here, using RNA immunoprecipitation-sequencing (RIP-seq), we identified the Polycomb-group histone methyltransferase EZH2 as a p53 mRNA-binding protein. EZH2 bound to an internal ribosome entry site (IRES) in the 5'UTR of p53 mRNA and enhanced p53 protein translation in a methyltransferase-independent manner. EZH2 augmented p53 GOF mutant-mediated cancer growth and metastasis by increasing protein levels of mutant p53. EZH2 overexpression was associated with worsened outcome selectively in patients with p53-mutated cancer. Depletion of EZH2 by antisense oligonucleotides inhibited p53 GOF mutant-mediated cancer growth. Our findings reveal a non-methyltransferase function of EZH2 that controls protein translation of p53 GOF mutants, inhibition of which causes synthetic lethality in cancer cells expressing p53 GOF mutants.

Dihydromyricetin attenuates cisplatin-induced acute kidney injury by reducing oxidative stress, inflammation and ferroptosis.

BACKGROUND: Cisplatin is effective against various types of cancers. However, its clinical application is limited owing to its adverse effects, especially acute kidney injury (AKI). Dihydromyricetin (DHM), a flavonoid derived from Ampelopsis grossedentata, has varied pharmacological activities. This research aimed to determine the molecular mechanism for cisplatin-induced AKI. METHODS: A murine model of cisplatin-induced AKI (22 mg/kg, I.P.) and a HK-2 cell model of cisplatin-induced damage (30 µM) were established to evaluate the protective function of DHM. Renal dysfunction markers, renal morphology and potential signaling pathways were investigated. RESULTS: DHM decreased the levels of renal function biomarkers (blood urea nitrogen and serum creatinine), mitigated renal morphological damage, and downregulated the protein levels of kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin. It upregulated the expression levels of antioxidant enzymes (superoxide dismutase and catalase expression), nuclear factor-erythroid-2-related factor 2 (Nrf2) and its downstream proteins, including heme oxygenase-1 (HO-1), glutamate-cysteine ligase catalytic (GCLC) and modulatory (GCLM) subunits, thus eventually reducing cisplatin-induced reactive oxygen species (ROS) production. Moreover, DHM partially inhibited the phosphorylation of the active fragments of caspase-8 and -3 and mitogen-activated protein kinase and restored glutathione peroxidase 4 expression, which attenuated renal apoptosis and ferroptosis in cisplatin-treated animals. DHM also mitigated the activation of NLRP3 inflammasome and nuclear factor (NF)-κB, attenuating the inflammatory response. In addition, it reduced cisplatin-induced HK-2 cell apoptosis and ROS production, both of which were blocked by the Nrf2 inhibitor ML385. CONCLUSIONS: DHM suppressed cisplatin-induced oxidative stress, inflammation and ferroptosis probably through regulating of Nrf2/HO-1, MAPK and NF-κB signaling pathways.

Competitive ELISA based on pH-responsive persistent luminescence nanoparticles for autofluorescence-free biosensor determination of ochratoxin A in cereals.

An accurate and sensitive competitive enzyme-linked immunosorbent assay (ELISA) based on persistent luminescence nanoparticles Zn2GeO4:Mn2+, Eu3+ (ZGME) was developed for detecting ochratoxin A (OTA), a powerfully toxic mycotoxin usually found in grains. As a signal output element of autofluorescence-free biosensors, ZGME can be integrated into ELISA with glucose oxidase (GOx)-binding OTA molecules due to its excellent pH-responsive persistent luminescence. In the absence of OTA, the OTA-GOx conjugate was captured by the anti-OTA monoclonal antibody (anti-OTA mAb) pre-coated on the 96-well plate. The results indicate a decrease in the pH value of the solution, which triggered the quenching of ZGME luminescence due to GOx-dependent gluconic acid production. The presence of OTA inhibited the binding of OTA-GOx on the plate, thus decreasing the production of gluconic acid and increasing the persistent luminous intensity of ZGME. Under the optimized concentrations of anti-OTA mAb and OTA-GOx, quantitative determination of OTA was achieved by plotting the increase or decrease in persistent luminescence intensity of ZGME at 535 nm. In this study, the linear range was from 0.1 µg L-1 to 63 µg L-1, and the limit of detection (LOD) was as low as 0.045 µg L-1. In five food samples (corn grit, brown rice, soybean, rice, and wheat), the results exhibited good stability and repeatability, with a recovery range from 81.3% to 94.4% and a relative standard deviation (RSD) of less than 4.2%. Hence, the established method provides a sensitive, accurate, and autofluorescence-free approach for the determination of OTA in different grain samples.

FUNDC1 regulates mitochondrial dynamics at the ER-mitochondrial contact site under hypoxic conditions.

In hypoxic cells, dysfunctional mitochondria are selectively removed by a specialized autophagic process called mitophagy. The ER-mitochondrial contact site (MAM) is essential for fission of mitochondria prior to engulfment, and the outer mitochondrial membrane protein FUNDC1 interacts with LC3 to recruit autophagosomes, but the mechanisms integrating these processes are poorly understood. Here, we describe a new pathway mediating mitochondrial fission and subsequent mitophagy under hypoxic conditions. FUNDC1 accumulates at the MAM by associating with the ER membrane protein calnexin. As mitophagy proceeds, FUNDC1/calnexin association attenuates and the exposed cytosolic loop of FUNDC1 interacts with DRP1 instead. DRP1 is thereby recruited to the MAM, and mitochondrial fission then occurs. Knockdown of FUNDC1, DRP1, or calnexin prevents fission and mitophagy under hypoxic conditions. Thus, FUNDC1 integrates mitochondrial fission and mitophagy at the interface of the MAM by working in concert with DRP1 and calnexin under hypoxic conditions in mammalian cells.

Androgen receptor splice variants bind to constitutively open chromatin and promote abiraterone-resistant growth of prostate cancer.

Androgen receptor (AR) splice variants (ARVs) are implicated in development of castration-resistant prostate cancer (CRPC). Upregulation of ARVs often correlates with persistent AR activity after androgen deprivation therapy (ADT). However, the genomic and epigenomic characteristics of ARV-dependent cistrome and the disease relevance of ARV-mediated transcriptome remain elusive. Through integrated chromatin immunoprecipitation coupled sequencing (ChIP-seq) and RNA sequencing (RNA-seq) analysis, we identified ARV-preferential-binding sites (ARV-PBS) and a set of genes preferentially transactivated by ARVs in CRPC cells. ARVs preferentially bind to enhancers located in nucleosome-depleted regions harboring the full AR-response element (AREfull), while full-length AR (ARFL)-PBS are enhancers resided in closed chromatin regions containing the composite FOXA1-nnnn-AREhalf motif. ARV-PBS exclusively overlapped with AR binding sites in castration-resistant (CR) tumors in patients and ARV-preferentially activated genes were up-regulated in abiraterone-resistant patient specimens. Expression of ARV-PBS target genes, such as oncogene RAP2A and cell cycle gene E2F7, were significantly associated with castration resistance, poor survival and tumor progression. We uncover distinct genomic and epigenomic features of ARV-PBS, highlighting that ARVs are useful tools to depict AR-regulated oncogenic genome and epigenome landscapes in prostate cancer. Our data also suggest that the ARV-preferentially activated transcriptional program could be targeted for effective treatment of CRPC.

JS-K, a nitric oxide donor, induces autophagy as a complementary mechanism inhibiting ovarian cancer.

BACKGROUND: Ovarian cancer (OC) is the second most frequent gynecological cancer and is associated with a poor prognosis because OC progression is often asymptoma-tic and is detected at a late stage. There remains an urgent need for novel targeted therapies to improve clinical outcomes in ovarian cancer. As a nitric oxide prodrug, JS-K is reported highly cytotoxic to human cancer cells such as acute myeloid leukemia, multiple myeloma and breast cancer. This study is aim to investigate the influence of JS-K on proliferation and apoptosis in ovarian cancer cells and explored possible autophagy-related mechanisms, which will contribute to future ovarian cancer therapy and supply theory support that JS-K holds great promise as a novel therapeutic agent against ovarian cancer. METHODS: The cytotoxicity, extracellular ROS/RNS activity and apoptotic effect of JS-K and indicated inhibitors on ovarian cancer cells in vitro were evaluated by MTT assay, extracellular ROS/RNS assay, caspases activities assay and western blot. Further autophagy effect of JS-K and indicated inhibitors were examined by MTT assay, cell transfection, immunofluorescence analysis, transmission electron microscopy (TEM) analysis and western blot on ovarian cancer cells in vitro. In vivo, the BALB/c-nude female mice with SKOV3 ovarian cancer cells xenograft were used to examine the efficacy of JS-K treatment on tumor growth. PCNA and p62 proteins were analyzed by immunohistochemistry. RESULTS: In vitro, JS-K inhibited the proliferation of ovarian cancer cells, induced apoptosis and cell nucleus shrinkage, enhanced the enzymatic activity of caspase-3/7/8/9, and significantly increased the production of ROS/RNS in ovarian cancer A2780 and SKOV3 cells, these effects were attenuated by inhibition of NAC. In addition, JS-K induced autophagy-related proteins and autophagosomes changes in ovarian cancer A2780 and SKOV3 cells. In vivo, JS-K inhibited tumor growth, decreased p62 protein expression and increased the expression levels of PCNA in xenograft models which were established using SKOV3 ovarian cancer cells. CONCLUSION: Taken together, we demonstrated that ROS/RNS stress-mediated apoptosis and autophagy are mechanisms by which SKOV3 cells undergo cell death after treatment with JS-K in vitro. Moreover, JS-K inhibited SKOV3 tumor growth in vivo. An alternative therapeutic approach for triggering cell death in cancer cells could constitute a useful multimodal therapies for treating ovarian cancer, which is known for its resistance to apoptosis-inducing drugs.

In vivo microdialysis with ultra performance liquid chromatography-mass spectrometry for analysis of tetramethylpyrazine and its interaction with borneol in rat brain and blood.

Tetramethylpyrazine (TMP) has been widely used in the treatment of ischemic cerebrovascular disease. However, the mechanism of TMP and how to increase its bioavailability need to be further explored. In our study, an in vivo microdialysis sampling technique coupled with ultra-performance liquid chromatography-mass spectrometry method was developed to investigate the pharmacokinetic properties of TMP and its interaction with different doses of borneol (BO) in rats. Linearity of TMP in brain and blood dialysates exhibited good linear relationships over the concentration range of 0.991-555.14 ng/mL. The specificity, linearity, accuracy, precision, matrix effect and stability were within acceptable ranges. The results demonstrated that BO had a marked impact on the pharmacokinetic properties of TMP. After co-administration, the areas under the concentration-time curve (AUC) of TMP in brain and blood were significantly increased. Meanwhile, the peak concentration of TMP in brain was also enhanced. The AUCBrain /AUCBlood of TMP, increased from 44% to 56 and 60.8% after co-administration with BO (15 and 30 mg/kg). The pharmacodynamic results showed that TMP co-administration with BO enhanced the cerebral blood flow during the period of ischemia and reduced the infarct volume. Overall, it might be an effective way to treat stroke to use TMP co-administered with BO.

Multiple molecular and cellular mechanisms of the antitumour effect of dihydromyricetin (Review).

Dihydromyricetin (DHM) is a natural flavonoid compound with multiple antitumour effects, including inhibition of proliferation, promotion of apoptosis, inhibition of invasion and migration, clearance of reactive oxygen species (ROS) and induction of autophagy. For example, DHM can effectively block the progression of the tumour cell cycle and inhibit cell proliferation. In different types of cancer cells, DHM can regulate the PI3K/Akt pathway, mTOR, and NF-κB pathway components, such as p53, and endoplasmic reticulum stress can alter the accumulation of ROS or induce autophagy to promote the apoptosis of tumour cells. In addition, when DHM is used in combination with various known chemotherapy drugs, such as paclitaxel, nedaplatin, doxorubicin, oxaliplatin and vinblastine, it can increase the sensitivity of tumour cells to DHM and increase the therapeutic effect of chemotherapy drugs. In the present review, the multiple molecular and cellular mechanisms underlying the antitumour effect of DHM, as well as its ability to increase the effects of various traditional antitumour drugs were summarized. Through the present review, it is expected by the authors to draw attention to the potential of DHM as an antitumour drug and provide valuable references for the clinical translation of DHM research and the development of related treatment strategies.

Dihydromyricetin functions as a tumor suppressor in hepatoblastoma by regulating SOD1/ROS pathway.

Background: Hepatoblastoma has an unsatisfactory prognosis, and traditional chemotherapy has strong side effects. Dihydromyricetin is a flavonoid extracted from a woody vine of the genus Serpentine in the family Vitaceae, with effects such as preventing alcoholic liver and reducing the incidence of liver cancer. However, the effect of DHM on hepatoblastoma and its specific pathway are still unclear. Purpose: The purpose of this study was to investigate the effects of DHM on children's hepatoblastoma and its related mechanisms. Methods: CCK-8 assays were used to measure proliferation. Apoptosis and reactive oxygen species (ROS) were analyzed by flow cytometry. Apoptotic cells were observed using Hoechst 33342 staining and fluorescence microscopy. Protein expression levels in HuH-6 and HepG2 cells were determined by western blotting. Results: We found that DHM was able to inhibit the growth and increase cellular mortality in HuH-6 and HepG2 cells. Furthermore, DHM decreased the intracellular ROS level and increased the expression of SOD1. ROS scavenger NAC promoted apoptosis, while the use of SOD1 inhibitor LCS-1 weakened the ROS scavenging effect of DHM , and to some extent reduced the killing effect of DHM on hepatoblastoma cells. Conclusion: These results suggest that regulating SOD1/ROS pathway to induce apoptosis is one of the potential mechanisms of DHM as a tumor suppressor in hepatoblastoma. Therefore, DHM may be a novel candidate for inhibiting hepatoblastoma growth and deserves further study.

Expressions of glucose transporter genes are diversely attenuated and significantly associated with prostate cancer progression.

Prostate cancer is a health-threaten disease in men worldwide, however, lacking is the reliable biomarkers for patient management. Aberrant metabolic events including glucose metabolism are involved in prostate cancer progression. To examine the involvement of glucose metabolic pathways in prostate cancer, we analyzed the expression profiles of glucose transporter family genes using multiple RNA-seq datasets. Our results showed that three SLC2A family genes (SLC2A4/5/9) were significantly downregulated in primary prostate cancers compared to their benign compartments. These down-regulated expressions were inversely correlated with their gene promoter methylation and genome abnormalities. Among these three SLC2A genes, only SLC2A4 showed a significantly reverse correlation with all clinicopathological parameters, including TNM stage, disease relapse, Gleason score, disease-specific survival, and progression-free interval. In addition, the expression levels of these three genes were strongly correlated with anti-cancer immune cell filtration in primary prostate cancers. In a group of patients with early-onset prostate cancers, SLC2A4 also showed a strong negative correlation with multiple clinicopathological parameters, such as tumor mutation burden, biochemical relapse, pre-surgical PSA levels, and Gleason score but a positive correlation with progression-free interval after surgery. In metastatic castration-resistant prostate cancers (CRPC), SLC2A9 gene expression but not SLC2A4 or SLC2A5 genes showed a significant correlation with androgen receptor (AR) activity score and neuroendocrinal (NE) activity score. Meanwhile, SLC2A2/9/13 expression was significantly elevated in CRPC tumors with neuroendocrinal features compared to those without NE features. On the other hand, SLC2A10 and SlC2A12 gene expression were significantly reduced in NEPC tumors compared to CRPC tumors. Consistently, SLC2A10/12 expression levels were significantly reduced in castrated animals carrying the LuCaP35 xenograft models. Survival outcome analysis revealed that SLC2A4 expression in primary tumors is a favorable prognostic factor and SLC2A6 is a worse prognostic factor for disease-specific survival and progression-free survival in prostate cancer patients. In conclusion, our results suggest that SLC2A4/6 expressions are strong prognostic factors for prostate cancer progression and survival. The significance of SLC2A2/9/13 over-expression during NEPC progression needs more investigation.

Hepcidin Downregulation Correlates With Disease Aggressiveness And Immune Infiltration in Liver Cancers.

BACKGROUND: Hepcidin is a polypeptide hormone mainly produced by hepatocytes to modulate systemic iron balance. A drastic downregulation of the hepcidin gene was found in liver cancers. However, there is a paucity of information about the clinical significance of hepcidin gene downregulation in liver cancers. METHODS: Hepcidin expression profiles were assessed using multiple public datasets via several bioinformatics platforms. Clinical and pathological information was utilized to stratify patients for comparison. Patient survival outcomes were evaluated using the Kaplan-Meier plotter, a meta-analysis tool. Tumor immune infiltration was analyzed using the single sample gene set enrichment analysis (ssGSEA) approach on the Cancer Genome Atlas (TCGA) dataset. Hepcidin antagonist Fursultiamine was used to treat liver cancer HepG2 and Huh7 cells together with Sorafenib. RESULTS: Hepcidin gene was predominantly expressed in benign liver tissues but drastically decreased in liver cancer tissues. Hepcidin reduction in liver cancers correlated with risk factors like non-alcoholic fatty liver disease (NAFLD) and liver fibrosis, as well as cancer grade and tumor stage. Hepcidin downregulation was associated with a rapid cancer progression and worse disease-specific survival, especially in patients of the White race without alcohol consumption history. Hepcidin expression in liver cancer tissues positively correlated with the bone morphogenetic protein-6 (BPM6)/interleukin-6 (IL6) cytokines and cytotoxic immune infiltration. Blocking hepcidin action with its antagonist Fursultiamine moderately reduced Sorafenib-induced apoptotic cell death in HepG2 and Huh7 cells. CONCLUSION: Hepcidin downregulation in liver cancers correlated with liver cancer risk factors, cancer aggressiveness, cytotoxic immune cell infiltration, and patient survival outcomes. BMP6/IL6 pathway insufficiency is a potential cause of hepcidin downregulation in liver cancers.

Molecular mechanisms and therapeutic implications of dihydromyricetin in liver disease.

Recent studies demonstrated that dihydromyricetin (DHM) has prominent therapeutic effects on liver injury and liver cancer. By summarizing the current preclinical in vitro and in vivo studies, the present review examines the preventive and therapeutic effects of DHM on liver disorders as well as its potential mechanisms. Briefly, in both chemical- and alcohol-induced liver injury models, DHM ameliorates hepatocyte necrosis and steatosis while promoting liver regeneration. In addition, DHM can alleviate nonalcoholic fatty liver disease (NAFLD) via regulating lipid/glucose metabolism, probably due to its anti-inflammatory or sirtuins-dependent mechanisms. Furthermore, DHM treatment inhibits cell proliferation, induces apoptosis and autophagy and regulates redox balance in liver cancer cells, thus exhibiting remarkable anti-cancer effects. The pharmacological mechanisms of DHM may be associated with its anti-inflammatory, anti-oxidative and apoptosis-regulatory benefits. With the accumulating interests in utilizing natural products to target common diseases, our work aims to improve the understanding of DHM acting as a novel drug candidate for liver diseases and to accelerate its translation from bench to bedside.

The Orphan Nuclear Receptor Gene NR0B2 Is a Favorite Prognosis Factor Modulated by Multiple Cellular Signal Pathways in Human Liver Cancers.

BACKGROUND: Liver cancer is a leading cause of cancer death worldwide, and novel prognostic factor is needed for early detection and therapeutic responsiveness monitoring. The orphan nuclear receptor NR0B2 was reported to suppress liver cancer development in a mouse model, and its expression levels were reduced in liver cancer tissues and cell lines due to hypermethylation within its promoter region. However, it is not clear if NR0B2 expression is associated with cancer survival or disease progression and how NR0B2 gene expression is regulated at the molecular level. METHODS: Multiple cancer databases were utilized to explore NR0B2 gene expression profiles crossing a variety of human cancers, including liver cancers, on several publicly assessable bioinformatics platforms. NR0B2 gene expression with or without kinase inhibitor treatment was analyzed using the qPCR technique, and NR0B2 protein expression was assessed in western blot assays. Two human hepatocellular carcinoma cell lines HepG2 and Huh7, were used in these experiments. NR0B2 gene activation was evaluated using NR0B2 promoter-driven luciferase reporter assays. RESULTS: NR0B2 gene is predominantly expressed in liver tissue crossing human major organs or tissues, but it is significantly downregulated in liver cancers. NR0B2 expression is mostly downregulated in most common cancers but also upregulated in a few intestinal cancers. NR0B2 gene expression significantly correlated with patient overall survival status in multiple human malignancies, including lung, kidney, breast, urinary bladder, thyroid, colon, and head-neck cancers, as well as liposarcoma and B-cell lymphoma. In liver cancer patients, higher NR0B2 expression is associated with favorite relapse-free and progression-free survival, especially in Asian male patients with viral infection history. In addition, NR0B2 expression negatively correlated with immune infiltration and PIK3CA and PIK3CG gene expression in liver cancer tissues. In HepG2 and Huh7 cells, NR0B2 expression at the transcription level was drastically reduced after MAPK inhibition but was significantly enhanced after PI3K inhibition. CONCLUSION: NR0B2 gene expression is altered mainly in most human malignancies and significantly reduced in liver cancers. NR0B2 is a prognosis factor for patient survival in liver cancers. MAPK and PI3K oppositely modulate NR0B2 expression, and NR0B2 gene upregulation might serve as a therapeutic responsiveness factor in anti-PI3K therapy for liver cancer.

PAQR6 Upregulation Is Associated with AR Signaling and Unfavorite Prognosis in Prostate Cancers.

Progesterone-induced rapid non-genomic signaling events have been confirmed through several membrane progesterone receptors (mPR). Some mPRs were reported to correlate with cancer progression and patient prognosis. In this study, we conducted a comprehensive analysis of all progesterone receptor (PGR)-related genes in prostate cancer tissues and examined the correlations of their expression levels with disease progression and patient survival outcomes. We utilized multiple RNA-seq and cDNA microarray datasets to analyze gene expression profiles and performed logistics aggression and Kaplan-Meier survival analysis after stratifying patients based on tumor stages and Gleason scores. We also used NCBI GEO datasets to examine gene expression patterns in individual cell types of the prostate gland and to determine the androgen-induced alteration of gene expression. Spearman coefficient analysis was conducted to access the correlation of target gene expression with treatment responses and disease progression status. The classic PGR was mainly expressed in stromal cells and progestin and adipoQ receptor (PAQR) genes were the predominant genes in prostate epithelial cells. Progesterone receptor membrane component-1 (PGRMC1) was significantly higher than PGRMC2 in all prostate cell types. In prostate cancer tissues, PAQR6 expression was significantly upregulated, while all other genes were largely downregulated compared to normal prostate tissues. Although both PAQR6 upregulation and PAQR5 downregulation were significantly correlated with tumor pathological stages, only PAQR6 upregulation was associated with Gleason score, free-prostate-specific antigen (fPSA)/total-PSA (tPSA) ratio, and patient overall survival outcomes. In addition, PAQR6 upregulation and PGR/PGRMC1 downregulation were significantly associated with a quick relapse. Conversely, in neuroendocrinal prostate cancer (NEPC) tissues, PAQR6 expression was significantly lower, but PAQR7/8 expression was higher than castration-resistant prostate cancer (CRPC) tissues. PAQR8 expression was positively correlated with androgen receptor (AR) score and AR-V7 expression levels but inversely correlated with NEPC score in metastatic CRPC tumors. This study provides detailed expression profiles of membrane progesterone receptor genes in primary cancer, CRPC, and NEPC tissues. PAQR6 upregulation in primary cancer tissues is a novel prognostic biomarker for disease progression, overall, and progression-free survival in prostate cancers. PAQR8 expression in CRPC tissues is a biomarker for AR activation.

PAQR5 Expression Is Suppressed by TGFß1 and Associated With a Poor Survival Outcome in Renal Clear Cell Carcinoma.

BACKGROUND: Renal cell carcinoma (RCC) was sex-hormone responsive, and clinical trials using progesterone significantly reduced the incidence of distal metastasis after radical nephrectomy. Recently membrane-bound progesterone receptors (mPRs) were discovered to mediate the non-genomic effect of progesterone. Aberrant expressions of these mPRs were reported in human breast, ovarian, urinary bladder, brain, uterine, and prostate cancers. However, their expression profiles in RCC are yet to be assessed. METHODS: Multiple datasets from RNA sequencing (RNA-seq), cDNA microarray, and proteomic analysis were used to compare gene expression between cancerous and normal kidney tissues. Immunohistochemistry was conducted to examine protein expression in kidney tissues. Promoter methylation levels were assessed for correlation analysis with gene expression. RESULTS: Of the seven membrane-bound progesterone receptor genes, the progestin and adipoQ receptor-5 (PAQR5) gene is predominantly expressed in normal kidney tissue but was significantly downregulated in RCC tissues. PAQR5 downregulation correlated with tumor stage, cancer grade, lymph node invasion, and distal metastasis only in clear cell RCC (ccRCC) tissues. PAQR5 downregulation was associated with an increased promoter DNA methylation and a poor survival outcome in ccRCC patients. In addition, PAQR5 expression inversely correlated with transforming growth factor beta-1 (TGFB1) expression, and TGFß1 treatment significantly reduced PAQR5 gene expression. CONCLUSION: PAQR5 is a novel prognostic biomarker in ccRCC and is negatively regulated by the TGFß1 pathway.

rhIL-1Ra reduces hepatocellular apoptosis in mice with acetaminophen-induced acute liver failure.

Acute liver failure (ALF) is a life-threatening disease that has proven difficult to cure. In Western countries, acetaminophen (APAP) poisoning is the most common cause of ALF. However, the mode of cell death in APAP-induced ALF cases is controversial. Previous studies have shown that administration of anti-interleukin-1 (anti-IL-1) antibody attenuated APAP-induced liver injury, and that administration of anti-IL-1 receptor antagonist (anti-IL-1Ra) antibody exacerbated organ injury. These results prompted us to investigate the roles of IL-1Ra in APAP-induced ALF mice. Our results show that administration of recombinant human IL-1Ra (rhIL-1Ra) could significantly improve the survival rate of mice with ALF induced by APAP. Furthermore, we found that rhIL-1Ras could dramatically inhibit the activities of alanine aminotransferase and aspartate aminotransferase in serum, reduce the death of hepatocytes and accelerate the proliferation of hepatocytes. In addition, we show that hepatocellular apoptosis rather than necrosis was the major cause of ALF-induced animal death, and that the anti-apoptosis role of rhIL-1Ra was mediated by reducing the release of cytochrome c from the mitochondria, and the activities of caspase-3, caspase-8 and caspase-9 in the liver tissue. In conclusion, these data indicate that rhIL-1Ra is a promising candidate for the treatment of APAP-induced ALF in mice through the reduction of hepatocellular apoptosis.

[Differential expression of MAPK-pathway genes during liver regeneration of mouse.].

To investigate the different expression profiles of MAPK pathway genes and their corresponding functions during liver regeneration, we used a CCl4 induced mouse liver regeneration model in this study. Mouse was injected with CCl4 in the abdominal cavity to cause damage in the liver and followed by liver histology examination and measurement of serum ALT levels in blood sample collected at 0, 0.5, 1.5, 4.5, and 7 d after CCl4 injection. Differentially expressed genes in the MAPK pathway during liver regeneration were analyzed using mouse cDNA microarray method (Affymetrix). The results obtained were further subjected to hierarchical clustering study and were validated with real-time PCR. Microarray hybridization identified 31 out of the 93 MAPK pathway component genes, which have significantly altered their expression levels during liver regeneration. Among them, both up- and down-regulated genes were classified into various groups according to clustering studies and functional analysis. At the initial stage of liver regeneration, the number of up-regulated genes was greater than the down-regulated genes, while at the late stage the situation was reversed. Our results suggest that MAPK pathway might play different regulatory roles in responding to different stages of liver regeneration.

JS­K induces G2/M phase cell cycle arrest and apoptosis in A549 and H460 cells via the p53/p21WAF1/CIP1 and p27KIP1 pathways.

Lung cancer is one of the most common malignancies worldwide, with high mortality and morbidity rates. O2­â€‹(2,4­â€‹dinitrophenyl)­1­â€‹[(4­ethoxycarbonyl)piperazin­1­yl]diazen­1­ium­1,2­diolate (JS­K) is a potent anticancer agent that acts against a subset of human non­small cell lung cancer (NSCLC) cell lines; however, the underlying mechanisms of JS­K in NSCLC remain unclear. The present study aimed to evaluate the anticancer effect of JS­K and investigate its underlying mechanisms in A549 and H460 cells. In the present study, A549 and H460 cells were treated with JS­K, and then evaluated by cell viability assay, flow cytometry and western blot analysis. JS­K markedly induced cell cycle arrest at the G2/M phase in a concentration and time­dependent manner in both cell lines. This was associated with increased expression levels of p53, and the cell cycle inhibitors p21WAF1/CIP1 and p27KIP1, which, in turn, inhibited the expression of Cdc2, cyclin B1 and cyclin­dependent kinase 2. In addition, JS­K­induced inhibition of proliferation was revealed to be partially modulated by the upregulation of p53 and p21WAF1, the ratio of Bax/Bcl­2, and the activation of both the intrinsic and extrinsic apoptotic pathways in A549 and H460 cells. These results demonstrated that JS­K could trigger cell cycle arrest at the G2/M phase and apoptosis in A549 and H460 cells, which was likely mediated via the p53/p21WAF1/CIP1 and p27KIP1 pathways. Overall, the results indicated that JS­K may be used as an anticancer agent for the treatment of NSCLC.

Xanthine oxidase-mediated oxidative stress promotes cancer cell-specific apoptosis.

The natural compound Alternol was shown to induce profound oxidative stress and apoptotic cell death preferentially in cancer cells. In this study, a comprehensive investigation was conducted to understand the mechanism for Alternol-induced ROS accumulation responsible for apoptotic cell death. Our data revealed that Alternol treatment moderately increased mitochondrial superoxide formation rate, but it was significantly lower than the total ROS positive cell population. Pre-treatment with mitochondria-specific anti-oxidant MitoQ, NOX or NOS specific inhibitors had no protective effect on Alternol-induced ROS accumulation and cell death. However, XDH/XO inhibition by specific small chemical inhibitors or gene silencing reduced total ROS levels and protected cells from apoptosis induced by Alternol. Further analysis revealed that Alternol treatment significantly enhanced XDH oxidative activity and induced a strong protein oxidation-related damage in malignant but not benign cells. Interestingly, benign cells exerted a strong spike in anti-oxidant SOD and catalase activities compared to malignant cells after Alternol treatment. Cell-based protein-ligand engagement and in-silicon docking analysis showed that Alternol interacts with XDH protein on the catalytic domain with two amino acid residues away from its substrate binding sites. Taken together, our data demonstrate that Alternol treatment enhances XDH oxidative activity, leading to ROS-dependent apoptotic cell death.