Decreased expression of HADH is related to poor prognosis and immune infiltration in kidney renal clear cell carcinoma.

Kidney renal clear cell carcinoma (KIRC) is the subtype pf kidney cancer having the highest mortality as well as the highest potential of invasion and metastasis. The expression of HADH, encoding a key enzyme in fatty acid ß-oxidation, has rarely been reported to correlate with prognosis and immune infiltration in cancers. This study aimed to explore the prognostic value of HADH in patients with KIRC. Gene expression profiles and clinical data of KIRC patients were acquired from The Cancer Genome Atlas. We compared the expression of HADH between KIRC tissues and normal tissues. Then, the relationship between HADH expression and the clinicopathological characteristics (survival, age, gender, stage, and grade) of KIRC was explored. Data from several online databases and paraffin-embedded specimens from two cohorts were used for external validation (10 cases from Meizhou People's Hospital and another 75 cases from a tissue chip, with both cohorts including KIRC samples and paired normal tissues). We also predicted the fractions of tumor-infiltrating immune cells (TIICs) in various tissues using CIBERSORT. Next, we estimated the prognostic value of differences in TIIC proportions between the high and low HADH expression groups. Finally, gene set enrichment analysis (GSEA) was performed to explore the potential mechanisms by which HADH expression influences patient survival. The expression of HADH was significantly lower in KIRC tissue than in normal tissue. Decreased expression of HADH was significantly correlated with high histologic grade, advanced stage, and poor prognosis. The differential expression of HADH was validated at the protein level by immunohistochemistry. Multivariate Cox regression analysis indicated that HADH was an independent prognostic factor for KIRC. In addition, HADH expression was significantly associated with the accumulation of several TIICs, especially regulatory T cells. Finally, GSEA revealed that the transcriptome of the low HADH expression group was significantly enriched in genes involved in not only epithelial-mesenchymal transition and inflammatory response but also TNF-α, IL-6-JAK-STAT3, and interferon-γ signaling. In conclusion, our study demonstrated that decreased expression of HADH is related to poor prognosis and immune infiltration in KIRC; this finding may provide crucial information for the development of immunotherapies.

Role of WW Domain-containing Oxidoreductase WWOX in Driving T Cell Acute Lymphoblastic Leukemia Maturation.

Whether tumor suppressor WWOX (WW domain-containing oxidoreductase) stimulates immune cell maturation is largely unknown. Here, we determined that Tyr-33-phosphorylated WWOX physically binds non-phosphorylated ERK and IκBα in immature acute lymphoblastic leukemia MOLT-4 T cells and in the naïve mouse spleen. The IκBα·ERK·WWOX complex was shown to localize, in part, in the mitochondria. WWOX prevents IκBα from proteasomal degradation. Upon stimulating MOLT-4 with ionophore A23187/phorbol myristate acetate, endogenous IκBα and ERK undergo rapid phosphorylation in <5 min, and subsequently WWOX is Tyr-33 and Tyr-287 de-phosphorylated and Ser-14 phosphorylated. Three hours later, IκBα starts to degrade, and ERK returns to basal or non-phosphorylation, and this lasts for the next 12 h. Finally, expression of CD3 and CD8 occurs in MOLT-4 along with reappearance of the IκBα·ERK·WWOX complex near 24 h. Inhibition of ERK phosphorylation by U0126 or IκBα degradation by MG132 prevents MOLT-4 maturation. By time-lapse FRET microscopy, IκBα·ERK·WWOX complex exhibits an increased binding strength by 1-2-fold after exposure to ionophore A23187/phorbol myristate acetate for 15-24 h. Meanwhile, a portion of ERK and WWOX relocates to the nucleus, suggesting their role in the induction of CD3 and CD8 expression in MOLT-4.

Research on the Mechanical, Thermal and Induction Healing Properties of Asphalt Wearing Course with Steel Fibers.

Induction healing technology can effectively repair microcracks in asphalt mixtures and is a promising maintenance technology for asphalt pavements. However, it requires the addition of steel wool fibers to asphalt mixtures and cannot be directly used to repair existing pavements. In order to improve the practicality of the induction healing technology, this article designs a wearing course asphalt mixture with induction healing function that is going to be paved above the existing road surface. The AC-10 asphalt wearing course for induction heating was prepared by adding steel fiber (SF). Analysis of the overall temperature of the surface revealed the unevenness of the temperature distribution, and the healing properties were investigated through protective heating that controlled the maximum temperature of the upper surface. The results show that the addition of SF can improve the high-temperature stability, low-temperature and intermediate-temperature crack resistance, and moisture stability of asphalt wearing courses; however, it has adverse effects on volumetric performance and skid resistance. The heating temperature increases with the increase in SF content, but higher maximum temperature heating rate causes worse heating uniformity and lower healing effect. The maximum heating rate of the sample with 10% SF reaches 3.92 °C/s, while its heating rate at minimum temperature is similar to that of the sample with 6% SF, which is only 0.7 °C/s, indicating the worst heating uniformity. The best healing effect occurs when the maximum temperature of the upper surface reaches 160 °C. The recommended optimal SF content is 6% of the asphalt volume. The asphalt mixture with 6% SF has an appropriate volume performance, moisture stability, and skid resistance; additionally, it has the best high-temperature stability, as well as low-temperature and intermediate-temperature crack resistance. Meanwhile, it also has uniform temperature distribution and efficient healing efficiency.

Identification and multicentric validation of soluble CDCP1 as a robust serological biomarker for risk stratification of NASH in obese Chinese.

The definitive diagnosis of non-alcoholic steatohepatitis (NASH) currently relies on invasive and labor-intensive liver biopsy. Here, we identified soluble CUB domain-containing protein 1 (sCDCP1) as a top-ranked non-invasive biomarker for NASH using Olink-based proteomics in 238 obese individuals with liver biopsies. Both the circulating concentration and hepatic mRNA abundance of sCDCP1 were significantly elevated in patients with NASH and correlated closely with each histological feature of NASH. In the pooled multicenter validation cohort, sCDCP1 as a standalone biomarker achieved an area under the receiver operating characteristic (AUROC) of 0.838 (95% confidence interval [CI] 0.789-0.887) for diagnosing NASH, which is better than those achieved with cytokeratin-18 and other non-invasive tests. Furthermore, the C-DAG model established by the combination of sCDCP1 with diabetes, aspartate aminotransferase (AST), and gender accurately rules in and rules out both NASH and fibrotic NASH (gray zones <20%). Thus, sCDCP1-based non-invasive tests can be potentially implemented for screening and early diagnosis of NASH and for ruling out low-risk individuals to avoid unnecessary liver biopsies.

Self-Healing Properties of Asphalt Concrete with Calcium Alginate Capsules Containing Different Healing Agents.

Calcium alginate capsules encapsulating rejuvenator are a promising self-healing technology for asphalt pavement, but the effects of different healing agents on the self-healing performance of asphalt concrete has not been considered. In view of this, this paper aimed at exploring the effects of calcium alginate capsules containing different healing agents on the self-healing properties of asphalt concrete. Three types of capsules with sunflower oil, waste cooking oil and commercial rejuvenator were fabricated via the orifice-coagulation bath method and the interior structure, mechanical strength, thermal stability and oil content of the prepared capsules were characterized. The healing levels of asphalt mixtures with different capsules under different loading cycles and stress levels were evaluated. Furthermore, the saturates, aromatics, resins and asphaltenes (SARA) fractions and rheological property of extracted asphalt binder within test beams with different capsules after different loading conditions were assessed. The results indicated that all the three types of capsules meet the mechanical and thermal requirement of mixing and compaction of asphalt mixtures. The healing levels of test beams containing vegetable oil capsules were higher than that of waste cooking oil capsules and industrial rejuvenator capsules. The strength recovery ratio and fracture energy recovery ratio of test beams with vegetable oil capsules reached 82.8% and 96.6%, respectively, after 20,000 cycles of compressive loading at 1.4 MPa. The fracture energy recovery ratio of the waste cooking oil capsules also reached as high as 90%, indicating that waste cooking oil can be used as the healing agent of calcium alginate capsules to improve the self-healing property of asphalt mixture. This work provides a significant guide for the selection of healing agent for self-healing capsules in the future.

Self-Healing Performance of Asphalt Concrete with Ca-Alginate Capsules under Low Service Temperature Conditions.

Calcium alginate capsules containing rejuvenators represent a promising method for asphalt concrete premaintenance, but their healing capacities under lower temperature conditions are still unknown. This paper investigated the healing performance of asphalt concrete containing calcium alginate capsules at low service temperatures. The Ca-alginate capsules were synthesized, and their morphology, compressive strength, thermal resistance, and relative oil content were evaluated. Besides, evaluations for the healing of asphalt concrete and the rejuvenator-release ratio of the capsules were determined via fracture-healing-refracture testing and Fourier-transform infrared spectrum experiments. Meanwhile, the glass transition temperature and rheological property of asphalt binder after compressive loading under different temperatures were explored via a differential scanning calorimeter and dynamic shear rheometer. The results showed that the capsules had good thermal resistance and mechanical strength. The capsules released less oil under -15, -10, and -5 °C than at 20 °C, and the healing ratios of the asphalt concrete with the capsules at -15, -10, and -5 °C were obviously lower than that at 20 °C. The released rejuvenator from the capsules could decrease the complex modulus and glass transition temperature of the asphalt binder. When compared with low service temperatures, the asphalt binder containing the capsules and serving at a high temperature has a better softening effect and low-temperature performance due to more oil being released.

Effect of Ageing on Self-Healing Properties of Asphalt Concrete Containing Calcium Alginate/Attapulgite Composite Capsules.

Calcium alginate capsules within asphalt concrete can gradually release interior asphalt rejuvenator under cyclic loading to repair micro cracks and rejuvenate aged asphalt in-situ. However, asphalt pavement will become aged due to environmental and traffic factors during the service period. In view of this, this paper investigated the effect of ageing on the healing properties of asphalt concrete containing calcium alginate/attapulgite composite capsules under cyclic loading. The capsules were fabricated using the orifice-bath method and the morphological structure, mechanical strength, thermal stability, oil release ratios and healing levels of capsules in fresh, short-term ageing and long-term ageing asphalt concrete were explored. The results indicated that the different ageing treatments would not damage the multi-chamber structure nor decrease the mechanical strength of capsules but would induce the capsules release oil prematurely. The premature oil released from capsules in turn can offset the ageing effect owing to ageing treatment. The short-term ageing and long-term ageing plain asphalt mixtures gained strength recovery ratios of 39.3% and 34.2% after 64,000 cycles of compression loading, while the strength recovery ratios of short-term ageing and long-term ageing asphalt mixtures containing capsules were 63.5% and 54.8%, respectively.

Trends in the surgical management of rectal prolapse: An Asian experience.

BACKGROUND: Posterior compartment pelvic floor prolapse (PCPFP) leads to anatomical distortion and functional impairment. Definitive management involves surgery. Ventral mesh rectopexy (VMR) has gained increasing popularity in the West as it emerges as a durable approach. Existing literature and evidence on safety and efficacy of PCPFP surgery in the Asian population remains sparse. Our study aims to review our institution's experience in surgery for PCPFP. METHODS: All cases of PCPFP surgery in Singapore General Hospital between 2014 to 2019 were studied. RESULTS: Eighty-three patients had surgery performed for PCPFP, with the majority (83%) in the last 3 years. Median age was 63 years and 92% were female. Most patients (64%) had obstructive defecation symptoms, while the remaining had fecal incontinence, rectal bleeding, or anal discomfort. Main anatomical indication for surgery was external rectal prolapse (48%). Other indications were rectocele and/or rectal intussusception. The majority (66%) had abdominal rectopexy, while 28 underwent Delorme's procedure. Forty-five of the 50 VMRs were minimally invasive. Patients undergoing rectopexy were observed to be younger. Median length of stay was 3 days. Nine patients had early operative complications of which ileus was most common. Median length of follow-up was 12 months. The majority (93%) had initial symptom satisfaction. Eleven patients had anatomical recurrence with a median length of 9 months to development. There was no significant difference in outcomes between abdominal vs perineal approach, or laparoscopic vs robotic VMR. CONCLUSION: Surgery for PCPFP has gained acceptance in our Asian institution with good symptom improvement, alongside low morbidity and recurrence.

Cloning and characterisation of a novel 2,4-dichlorophenol hydroxylase from a metagenomic library derived from polychlorinated biphenyl-contaminated soil.

A novel 2,4-dichlorophenol hydroxylase (TfdB, EC 1.14.13.20) gene, designated as tfdB-JLU, was identified from a metagenome constructed from polychlorinated biphenyl-contaminated soil by functional screening and heterologously expressed in Escherichia coli. The deduced amino acid sequence of tfdB-JLU exhibited less than 48% homology with other known TfdBs. The enzyme exhibited a wider substrate spectrum than the previously reported TfdBs and higher relative activity towards ortho-substituted dichlorophenols, 2-chlorophenol, and 3-chlorophenol than towards 2,4-dichlorophenol, the preferred substrate of other known TfdBs. The enzyme had a K ( m ) of 5 µM for 2,4-dichlorophenol and 6 µM for NADPH. The optimal temperature and pH of the enzyme were 25°C and 7.5, respectively. Activity of the purified TfdB-JLU was slightly enhanced by Ca(2+), Mn(2+), Co(2+), and Fe(2+), and completely inhibited by Cu(2+), Hg(2+), and Zn(2+). This study is the first report to identify a novel TfdB from a metagenome.

Upregulation of TMEM45A Promoted the Progression of Clear Cell Renal Cell Carcinoma in vitro.

BACKGROUND: Clear cell renal cell carcinoma (ccRCC) is the most common and aggressive type of primary kidney cancer worldwide. Transmembrane protein 45A (TMEM45A) has been reported to be closely associated with the progression of several cancers. However, the role of TMEM45A in ccRCC remains unclear. Our study intended to explore the potential role of TMEM45A in ccRCC. METHODS: Data on the expression of TMEM45A were obtained from multiple databases, including UCSC, GEPIA2, Oncomine and TIMER. Real-world samples of ccRCC and paired normal renal tissues were used to confirm the information obtained from the databases. In addition, the prognostic value of TMEM45A was evaluated. Loss-of-function assays were performed using TMEM45A-targeting lentivirus to evaluate the biological role of TMEM45A in renal cancer cells. Gene set enrichment analysis (GSEA) was performed to investigate the potential molecular mechanisms. RESULTS: TMEM45A was significantly overexpressed in patients with ccRCC and correlated with poor overall survival and disease-free survival. In addition, the expression of TMEM45A was closely associated with various clinicopathological parameters such as histological grade and TNM stage. Knockdown of TMEM45A inhibited the proliferation and migration and promoted the apoptosis of ccRCC cells in vitro. The results of the GSEA suggested that TMEM45A was potentially involved in the promotion of epithelial-mesenchymal transition (EMT) and inflammatory response in ccRCC. CONCLUSION: TMEM45A was overexpressed and associated with poor survival and acted as a tumour promoter in ccRCC; therefore, might be a potential prognostic marker and therapeutic target.

Downregulation of PPA2 expression correlates with poor prognosis of kidney renal clear cell carcinoma.

BACKGROUND: Kidney renal clear cell carcinoma (KIRC) is the most common subtype of kidney cancer. Inorganic pyrophosphatase (PPA2) is an enzyme that catalyzes the hydrolysis of pyrophosphate to inorganic phosphate; few studies have reported its significance in cancers. Therefore, we aimed to explore the prognostic value of PPA2 in KIRC. METHODS: PPA2 expression was detected via immunohistochemistry in a tissue chip containing specimens from 150 patients with KIRC. We evaluated the correlation between PPA2 expression, clinicopathological characteristics, and survival. Data from online databases and another cohort (paraffin-embedded specimens from 10 patients with KIRC) were used for external validation. RESULTS: PPA2 expression was significantly lower in KIRC tissues than in normal renal tissues (p < 0.0001). Low expression of PPA2 was significantly associated with a high histologic grade and poor prognosis. The differential expression of PPA2 was validated at the gene and protein levels. Multivariate Cox regression analysis showed that PPA2 expression was an independent prognostic factor in patients with KIRC. Gene set enrichment analysis suggested that decreased expression of PPA2 might be related to the epithelial-mesenchymal transition in KIRC. CONCLUSIONS: Our study demonstrated that PPA2 is an important energy metabolism-associated biomarker correlated with a favorable prognosis in KIRC.

miR-200a-3p facilitates bladder cancer cell proliferation by targeting the A20 gene.

BACKGROUND: MicroRNAs (miRs) are endogenous, single-stranded, noncoding RNAs that are involved in various physiological processes, and the development and the progression of various types of cancer. Specifically, the role of miR-200a-3p has been implicated in various types of cancer in contributing to a diverse array of cancer types has been previously reported. The present study aimed to investigate the expression levels of miR-200a-3p in human bladder cancer, as well as its potential role in disease pathogenesis. METHODS: Reverse transcription-polymerase chain reaction (RT-PCR) and Western blot were used to detect the expression of has-mir-200a-3p and tumor necrosis factor α (TNF-α) induced protein 3 (A20) in tumor tissues and cell lines. Dual-luciferase reporter assay and combination with the expression intervention of hsa-mir-200a-3p and A20 in bladder cancer cell lines to clarify the binding relationship between hsa-mir-200a-3p and A20.After the expression intervention of hsa-mir-200a-3p and A20 in bladder cancer cells, the changes of cell proliferation, cell apoptosis, cell cycle, wound-healing ability and migration ability were detected by CCK8, flow cytometry, wound-healing and Transwell methods. Xenograft transplantation model was performed subcutaneously in nude mice by implantation of J82 and T24 cells, and then the bladder cancer growth curve was calculated from mice exposed to has-mir-200a-3p minic or minic-NC. RESULTS: Bladder cancer tissues demonstrated significantly upregulated miR-200a-3p expression levels. Moreover, increased miR-200a-3p expression was significantly associated with distant metastasis and advanced stage. In addition, compared with the miR-control (Ctr) group, miR-200a-3p overexpression promoted bladder cancer cell proliferation, migration, invasion, cell cycle, and release of inflammatory cytokines, but inhibited cell apoptosis. Mechanistically, A20 was identified as a target gene of miR-200a-3p in bladder cancer cell lines. Moreover, compared with the miR-Ctr group, the miR-200a-3p overexpression group exhibited significantly promoted tumor growth in vivo, and A20 overexpression blocked the promoting effect of miR-200a-3p on bladder cancer. CONCLUSIONS: The results of the present study indicated that miR-200a-3p might serve act as an oncogene in human bladder cancer by targeting a novel the gene A20 gene; therefore, miR-200a-3p and A20 might serve could serve as novel therapeutic targets for bladder cancer.

A cost-effective smartphone-based device for ankle-brachial index (ABI) detection.

BACKGROUND AND OBJECTIVE: Detectors of ankle-brachial index (ABI) are commonly used in cardiovascular patients who have high-risk levels of arteriosclerosis. Increased evidences suggest that patients with arteriosclerosis possess many risks of geriatric and chronic diseases. Meanwhile, new chronic treatments trend from the hospitals toward family and community health centers, but for arteriosclerosis cases have delivered benefits far below instrument costs. Compared to traditional devices based on cuff pressure, cuffless and non-invasive measures have wider application potential in home health care, especially in the case of physically-restricted or severely symptomatic patients. METHODS: In this study, we developed a simple smartphone-based device for non-invasive ABI monitoring, which consists of four wireless cuffless limbs blood sensors. By identifying and tracking blood flow waveform, a multiparameter fusion (MPF) algorithm is used to estimate blood pressure and generate ABI value. An ARM-based chip STM32 has been adopted as the microcontroller. The ABI calculating program is embedded in C++ and executed by the processor. After generating data, ABI information can be delivered to the smartphone by using Bluetooth. Relying on mobile apps to visualize the data and display on the screen, doctors can monitor cardiovascular patients in real time and analyze the risk levels of arteriosclerosis online. RESULTS: In this paper, the detection conducted by the classical Doppler equipment and prototype were recorded respectively. A statistical evaluation of the verification results obtained from 29 patients and 7 sub-health volunteers is given, which shows that our device can achieve 91.80% and 93.84% accuracy for patients and sub-health volunteers, respectively. In addition, the prototype can be performed stably for a continuous long time monitoring. CONCLUSIONS: According to our studies, the accuracy of our device is sufficient for home medical and chronic disease monitoring within a certain time interval. The smartphone-based ABI device has several apparent advantages over traditional devices, such as portability, cost-effectiveness and energy-efficiency.

Stapled anastomosis for anterior resection is safe even after previous stapled haemorrhoidectomy--a report of five cases.

INTRODUCTION: Stapled haemorrhoidectomy has become popular for the treatment of symptomatic bleeding or prolapsing piles. There are concerns about the safety of another stapled low colorectal anastomosis after rectal resection if a patient who has had previous stapled haemorrhoidectomy subsequently develops colorectal neoplasia requiring an anterior resection. METHODS: A retrospective review of patients who underwent stapled haemorrhoidectomy and subsequently had anterior resection from 1999 to 2008 was performed. RESULTS: Five patients (all male) were found to have distal sigmoid or rectal tumours on surveillance colonoscopy after stapled haemorrhoidectomy. Median age was 65 years (range 58-71). All underwent anterior resection with stapled end-to-end colorectal anastomosis at median of 29 months (range 18-60 months) after the initial stapled haemorrhoidectomy. Median anastomotic height was 12 cm (range 1-12 cm). A defunctioning ileostomy was created for three out of five patients. All the colorectal anastomoses healed uneventfully. CONCLUSIONS: Stapled colorectal anastomosis may be safely performed after previous stapled haemorrhoidectomy.

Downregulation of enhancer RNA EMX2OS is associated with poor prognosis in kidney renal clear cell carcinoma.

Enhancer RNAs are a subclass of long non-coding RNAs transcribed from enhancer regions that play an important role in the transcriptional regulation of genes. However, their role in kidney renal clear cell carcinoma (KIRC) is largely unknown. Herein, we identified the key enhancer RNAs in KIRC via an integrated data analysis method. Gene expression profiles and clinical data of KIRC and 32 other cancer types were acquired using the University of California Santa Cruz Xena platform. Reported enhancer RNAs and genes regulated by them were selected as putative enhancer RNA-target pairs. Kaplan-Meier survival and correlation analyses were performed to identify the key enhancer RNAs. Finally, EMX2OS was identified as the enhancer RNA most associated with survival, with EMX2 as its target. EMX2OS downregulation was significantly associated with higher histological grade, advanced stage, and poorer prognosis. The results were validated in pan-cancer data from The Cancer Genome Atlas and RT-qPCR analysis of 12 pairs of KIRC and normal real-world samples. Functional enrichment analysis indicated that several metabolism-associated signaling pathways were enriched. This study demonstrated that EMX2OS is a key metabolism-associated enhancer RNA in KIRC with a favorable impact on survival and may be a novel therapeutic target in KIRC.

BRDT is a novel regulator of eIF4EBP1 in renal cell carcinoma.

Among all types of kidney diseases, renal cell carcinoma (RCC) has the highest mortality, recurrence and metastasis rates, which results in high numbers of tumor­associated mortalities in China. Identifying a novel therapeutic target has attracted increasing attention. Bromodomain and extraterminal domain (BET) proteins have the ability to read the epigenome, leading to regulation of gene transcription. As an important member of the BET family, bromodomain testis­specific protein (BRDT) has been well studied; however, the mechanism underlying BRDT in the regulation of RCC has not been fully investigated. Eukaryotic translation initiation factor 4E­binding protein 1 (eIF4EBP1) is a binding partner of eIF4E that is involved in affecting the progression of various cancer types via regulating gene transcription. To identify novel regulators of eIF4EBP1, an immunoprecipitation assay and mass spectrometry analysis was performed in RCC cells. It was revealed that eIF4EBP1 interacted with BRDT, a novel interacting protein. In addition, the present study further demonstrated that BRDT inhibitors PLX51107 and INCB054329 blocked the progression of RCC cells, along with suppressing eIF4EBP1 and c­myc expression. Small interfering (si) RNAs were used to knock down BRDT expression, which suppressed RCC cell proliferation and eIF4EBP1 protein expression. In addition, overexpression of eIF4EBP1 partially abolished the inhibited growth function of PLX51107 but knocking down eIF4EBP1 improved the inhibitory effects of PLX51107. Furthermore, treatment with PLX51107 or knockdown of BRDT expression decreased c­myc expression at both the mRNA and protein levels, and attenuated its promoter activity, as determined by luciferase reporter assays. PLX51107 also significantly altered the interaction between the c­myc promoter with eIF4EBP1 and significantly attenuated the increase of RCC tumors, accompanied by decreased c­myc mRNA and protein levels in vivo. Taken together, these data suggested that blocking of BRDT by PLX51107, INCB054329 or BRDT knockdown suppressed the growth of RCC via decreasing eIF4EBP1, thereby leading to decreased c­myc transcription levels. Considering the regulatory function of BET proteins in gene transcription, the present study suggested that there is a novel mechanism underlying eIF4EBP1 regulation by BRDT, and subsequently decreased c­myc in RCC, and further identified a new approach by regulating eIF4EBP1 or c­myc for enhancing BRDT­targeting RCC therapy.

Therapeutic Zfra4-10 or WWOX7-21 Peptide Induces Complex Formation of WWOX with Selective Protein Targets in Organs that Leads to Cancer Suppression and Spleen Cytotoxic Memory Z Cell Activation In Vivo.

Synthetic Zfra4-10 and WWOX7-21 peptides strongly suppress cancer growth in vivo. Hypothetically, Zfra4-10 binds to the membrane Hyal-2 of spleen Z cells and activates the Hyal-2/WWOX/SMAD4 signaling for cytotoxic Z cell activation to kill cancer cells. Stimulation of membrane WWOX in the signaling complex by a WWOX epitope peptide, WWOX7-21, is likely to activate the signaling. Here, mice receiving Zfra4-10 or WWOX7-21 peptide alone exhibited an increased binding of endogenous tumor suppressor WWOX with ERK, C1qBP, NF-κB, Iba1, p21, CD133, JNK1, COX2, Oct4, and GFAP in the spleen, brain, and/or lung which led to cancer suppression. However, when in combination, Zfra4-10 and WWOX7-21 reduced the binding of WWOX with target proteins and allowed tumor growth in vivo. In addition to Zfra4-10 and WWOX7-21 peptides, stimulating the membrane Hyal-2/WWOX complex with Hyal-2 antibody and sonicated hyaluronan (HAson) induced Z cell activation for killing cancer cells in vivo and in vitro. Mechanistically, Zfra4-10 binds to membrane Hyal-2, induces dephosphorylation of WWOX at pY33 and pY61, and drives Z cell activation for the anticancer response. Thus, Zfra4-10 and WWOX7-21 peptides, HAson, and the Hyal-2 antibody are of therapeutic potential for cancer suppression.

Eupatilin Inhibits Renal Cancer Growth by Downregulating MicroRNA-21 through the Activation of YAP1.

Renal cell carcinoma (RCC) is the second most common human urinary tumor. Eupatilin is the main active ingredient of the traditional Chinese medicine Artemisia asiatica. The effect of Eupatilin on RCC and the underlying mechanism remain unknown. Here, we investigated the anticancer effects and mechanisms of Eupatilin in RCC in vivo and in vitro, laying an experimental foundation for the clinical application of Eupatilin in the treatment of RCC. The results showed that Eupatilin significantly inhibited 786-O cell viability and migration and promoted apoptosis. Eupatilin inhibited the expression of miR-21 in 786-O cells, and overexpression of miR-21 suppressed the effect of Eupatilin on viability, apoptosis, and migration in 786-O cells. Eupatilin inhibited the growth of renal tumors in nude mice by downregulating miR-21. YAP1, which was identified as a target of miR-21, showed significantly lower expression in RCC tissues than in healthy tissues. miR-21 significantly inhibited YAP1 protein expression in 786-O cells and tumor tissues isolated from nude mice, and YAP1 attenuated the effect of miR-21 on the viability, apoptosis, and migration of 786-O cells. In conclusion, Eupatilin inhibited the expression of miR-21, which mediated the proapoptotic and antimigratory effects of Eupatilin by suppressing YAP1 in renal cancer cells. These results suggested that Eupatilin could be a potent agent for the treatment of RCC.

WWOX Possesses N-Terminal Cell Surface-Exposed Epitopes WWOX7-21 and WWOX7-11 for Signaling Cancer Growth Suppression and Prevention In Vivo.

Membrane hyaluronidase Hyal-2 supports cancer cell growth. Inhibition of Hyal-2 by specific antibody against Hyal-2 or pY216-Hyal-2 leads to cancer growth suppression and prevention in vivo. By immunoelectron microscopy, tumor suppressor WWOX is shown to be anchored, in part, in the cell membrane by Hyal-2. Alternatively, WWOX undergoes self-polymerization and localizes in the cell membrane. Proapoptotic pY33-WWOX binds Hyal-2, and TGF-ß induces internalization of the pY33-WWOX/Hyal-2 complex to the nucleus for causing cell death. In contrast, when pY33 is downregulated and pS14 upregulated in WWOX, pS14-WWOX supports cancer growth in vivo. Here, we investigated whether membrane WWOX receives extracellular signals via surface-exposed epitopes, especially at the S14 area, that signals for cancer growth suppression and prevention. By using a simulated 3-dimentional structure and generated specific antibodies, WWOX epitopes were determined at amino acid #7 to 21 and #286 to 299. Synthetic WWOX7-21 peptide, or truncation to 5-amino acid WWOX7-11, significantly suppressed and prevented the growth and metastasis of melanoma and skin cancer cells in mice. Time-lapse microscopy revealed that WWOX7-21 peptide potently enhanced the explosion and death of 4T1 breast cancer stem cell spheres by ceritinib. This is due to rapid upregulation of proapoptotic pY33-WWOX, downregulation of prosurvival pERK, prompt increases in Ca2+ influx, and disruption of the IkBα/WWOX/ERK prosurvival signaling. In contrast, pS14-WWOX7-21 peptide dramatically increased cancer growth in vivo and protected cancer cells from ceritinib-mediated apoptosis in vitro, due to a prolonged ERK phosphorylation. Further, specific antibody against pS14-WWOX significantly enhanced the ceritinib-induced apoptosis. Together, the N-terminal epitopes WWOX7-21 and WWOX7-11 are potent in blocking cancer growth in vivo. WWOX7-21 and WWOX7-11 peptides and pS14-WWOX antibody are of therapeutic values in suppressing and preventing cancer growth in vivo.

Strategies by which WWOX-deficient metastatic cancer cells utilize to survive via dodging, compromising, and causing damage to WWOX-positive normal microenvironment.

Proapoptotic tumor suppressor WWOX is upregulated in the early stage of cancer initiation, which probably provides limitation to cancer growth and progression. Later, WWOX protein is reduced to enhance cancer cell growth, migration, invasiveness and metastasis. To understand how WWOX works in controlling cancer progression, here we demonstrate that apoptotic stress mediated by ectopic WWOX stimulated cancer cells to secrete basic fibroblast growth factor (bFGF) in order to support capillary microtubule formation. This event may occur in the cancer initiation stage. Later, when WWOX loss occurs in cancer cells, hyaluronidase production is then increased in the cancer cells to facilitate metastasis. We determined that inhibition of membrane hyaluronidase Tyr216-phosphorylated Hyal-2 by antibody suppresses cancer growth in vivo. WWOX-negative (WWOX-) cells dodged WWOX+cells in the microenvironment by migrating individually backward to avoid physical contacts and yet significantly upregulating the redox activity of WWOX+parental cells or other WWOX+cell types for causing apoptosis. Upon detecting the presence of WWOX+cells from a distance, WWOX- cells exhibit activation of MIF, Hyal-2, Eph, and Wnt pathways, which converges to MEK/ERK signaling and enables WWOX- cells to evade WWOX+cells. Inhibition of each pathway by antibody or specific chemicals enables WWOX- cells to merge with WWOX+cells. In addition, exogenous TGF-ß assists WWOX- cells to migrate collectively forward and merge with WWOX+cells. Metastatic WWOX- cancer cells frequently secrete high levels of TGF-ß, which conceivably assists them to merge with WWOX+cells in target organs and secure a new home base in the WWOX+microenvironment. Together, loss of WWOX allows cancer cells to develop strategies to dodge, compromise and even kill WWOX-positive cells in microenvironment.