Human MutT Homolog 1 (MTH1) Inhibitor Reduces the Biological Activity of Ovarian Carcinoma Cells

This study intends to discuss the mechanism of MTH1 inhibitor (TH588) in the biological activity of ovarian carcinoma cells. A2780 and SKOV-3 cells were treated with different concentrations of TH588 and assigned into AT group (control), BT group (8 μmol/L TH588), CT group (16 μmol/L), DT group (32 μmol/L), ET group (64 μmol/L) and FT group (128 μmol/L) followed by measuring level of Bcl-2 and Bax by Western blot and PCR, and cell biological activities by MTT, FCM and Transwell chamber assay. The cell proliferative rate was not affected in AT group, but was lower in other groups in a reverse dose-dependent manner. There was significant difference on apoptotic rate and cell invasion among groups with increased apoptosis and reduce invasion after TH588 treatment. FT group showed lowest expression of Bcl-2 and Bax compared to other groups. In conclusion, the biological activity of A2780/SKOV3 cells could be reduced by MTH1 inhibitor which was probably through regulation of Bax and Bcl-2 expression.

Characterization of a novel peptide mined from the Red Sea brine pools and modified to enhance its anticancer activity

Peptide-based drugs have emerged as highly selective and potent cancer therapy. Cancer is one of the leading causes of death worldwide. Multiple approaches have been developed towards cancer treatment, including chemotherapy, radiation, and hormonal therapy; however, such procedures' non-specificity, toxicity, and inefficiency present a hurdle. In this study, we developed a support vector machine (SVM) model to detect the potential anticancer properties of novel peptides through scanning the American University in Cairo Red Sea metagenomics library. Further, we performed in silico studies on a novel 37-mer antimicrobial peptide mined from SVM pipeline analysis. This peptide was further modified to enhance its anticancer activity, analyzed for gene oncology, and subsequently synthesized. The anticancer properties of this 37-mer peptide were evaluated via cellular viability and cell morphology of SNU449, HepG2, SKOV3, and HeLa cells, using MTT assay. Furthermore, we assessed the migration capability of SNU449 and SKOV3 via scratch wound healing assay. Moreover, the targeted selectivity of the peptide for cancerous cells was assessed by testing its hemolytic activity on human erythrocytes. The peptide caused a significant reduction in cellular viability and critically affected the morphology of hepatocellular carcinoma (SNU449 and HepG2), ovarian cancer (SKOV3), and to a limited extent, cervical cancer cell lines (HeLa), in addition to decreasing viability of human fibroblast cell line (1Br-hTERT). Peptide treatment significantly affected the proliferation and migration ability of SNU449 and SKOV3 cells. Annexin V assay was used to evaluate induced cell death upon peptide treatment, attributing programmed cell death (Apoptosis) as the main cause of cell death in SNU449 cells. Finally, we established broad-spectrum antimicrobial properties of the peptide on both gram-positive and gram-negative bacterial strains. Thus, these findings infer the novelty of the peptide as a potential anticancer and antimicrobial agent.

Hedgehog−Gli2 Signaling Promotes Chemoresistance in Ovarian Cancer Cells by Regulating MDR1

BackgroundCisplatin (DDP) resistance remains a key challenge in improving the clinical outcome of patients with ovarian cancer (OC). Gli2 overexpression can lead to DDP resistance in OC cells, but the specific underlying regulatory mechanism remains unclear. The membrane transporter encoding gene MDR1 positively regulates chemotherapy resistance in various cancer types. We evaluated MDR1 as a potential Gli2 downstream target and the contribution of the Gli2/MDR1 axis in promoting DDP resistance in OC cells.MethodsTo generate drug-resistant SKOV3/DDP cells, SKOV3 cells were grown for six months under continuous induction wherein the DDP concentration was steadily increased. Gli2 expression in OC cells with varying DDP sensitivities was detected using western blot. Cell counting kit-8 assays were used to assess the DDP sensitivity of SKOV3, SKOV3/DDP, A2780, and A2780/DDP cells and reversal of DDP resistance in SKOV3/DDP and A2780/DDP cells. Cell proliferation was analyzed using 5-ethynyl-2′-deoxyuridine (EdU) incorporation assays. The transcriptional regulation of MDR1 by Gli2 was determined using luciferase reporter assays. Finally, xenograft OC tumors were generated in nude mice, which were then treated with intraperitoneal DDP or phosphate-buffered saline (PBS) injections to investigate if Gli2 affected DDP resistance in OC in vivo.ResultsDDP-resistant SKOV3/DDP and A2780/DDP cells showed higher expression of Gli2 and MDR1 as compared with that in DDP-sensitive OC cells. Gli2 knockdown in SKOV3/DDP cells significantly reduced MDR1 expression, whereas it increased DNA damage, thereby sensitizing OC cells to DDP. Similar results were obtained after targeting Gli2 expression with the Gli-antagonist 61 inhibitor (GANT61) in SKOV3/DDP and A2780/DDP cells. In cells stably overexpressing Gli2, treatment with gradient concentrations of verapamil, an MDR1 inhibitor, significantly inhibited MDR1 expression. Our findings indicate that downregulation of MDR1 expression may reverse OC cell resistance to DDP. Moreover, dual-luciferase reporter gene assays confirmed that MDR1 is a direct downstream target of Gli2, with Gli2 positively regulating MDR1 expression. Finally, subcutaneous xenotransplantation in nude mice demonstrated that Gli2 plays a key role in regulating OC drug resistance.ConclusionsWe identified a mechanism by which Hedgehog-Gli signaling regulates OC chemoresistance by modulating MDR1 expression. Hence, Gli2 and MDR1 are potential biomarkers and therapeutic targets in patients with chemoresistant OC.

Pardaxin Activates Excessive Mitophagy and Mitochondria-Mediated Apoptosis in Human Ovarian Cancer by Inducing Reactive Oxygen Species

Most ovarian cancer (OC) patients are diagnosed with stage III or higher disease, resulting in a poor prognosis. Currently, paclitaxel combined with carboplatin shows the best treatment outcome for OC. However, no effective drug is available for patients that do not respond to treatment; thus, new drugs for OC are needed. We evaluated the antimicrobial peptide, pardaxin, in PA-1 and SKOV3 cells. Pardaxin induced apoptosis as determined by MTT and TUNEL assays, as well as activation of caspases-9/3, Bid, t-Bid, and Bax, whereas Bcl-2 was downregulated. The IC50 values for pardaxin were 4.6–3.0 μM at 24 and 48 h. Mitochondrial and intracellular reactive oxygen species (ROS) were overproduced and associated with disrupted mitochondrial membrane potential and respiratory capacity. Additionally, the mitochondrial network was fragmented with downregulated fusogenic proteins, MFN1/2 and L-/S-OPA1, and upregulated fission-related proteins, DRP1 and FIS1. Autophagy was also activated as evidenced by increased expression of autophagosome formation-related proteins, Beclin, p62, and LC3. Enhanced mitochondrial fragmentation and autophagy indicate that mitophagy was activated. ROS-induced cytotoxicity was reversed by the addition of N-acetylcysteine, confirming ROS overproduction as a contributor. Taken together, pardaxin demonstrated promising anticancer activity in OC cells, which warrants further preclinical development of this compound.

Epiphycan Predicts Poor Outcomes and Promotes Metastasis in Ovarian Cancer

The small leucine-rich proteoglycan (SLRP) family is widely expressed in extracellular matrix and aggravates tumor progression. However, epiphycan (EPYC), as a member of the SLRPs family, its biological function in cancer has not been confirmed. Thus, we aimed to clarify the role of EPYC in progression of ovarian cancer (OC), and further analyze the molecular mechanisms implicated in tumorigenesis. Here, we analyzed the differential expression genes of GSE38734, including 4 matched primary OC and metastatic tissues. We obtained OC RNAseqs data from the Cancer Genome Atlas (TCGA) and analyzed the correlation between EPYC expression and OC staging, pathological grading, etc. The expression of EPYC in OC and normal ovarian tissues was compared in Oncomine website. We used siRNAs to interfere the expression of EPYC in ovarian cancer cell line SKOV3. Scratch test, transwell-matrigel chamber, CCK8 assay were used to detect the changes of SKOV3 migration, invasion and proliferation ability after EPYC was interfered. We used R software to make GO and KEGG analysis of related genes of EPYC. We used the Hitpredict website to predict interacting proteins. The results showed that the expression of EPYC in metastatic ovarian cancer was higher than primary ovarian cancer, and that in primary cancer was higher than normal ovaries. After siRNA interferes with EPYC expression, the migration, invasion and proliferation of SKOV3 cells were weakened. EPYC mainly played a role in ECM organization, and involved in PI3K/Akt, focal adhesion signaling pathways. EPYC might interact with PLCG2 and CRK, and be involved in signal transduction.

In Vitro Evaluation of DSPE-PEG (5000) Amine SWCNT Toxicity and Efficacy as a Novel Nanovector Candidate in Photothermal Therapy by Response Surface Methodology (RSM)

Nowadays, finding a novel, effective, biocompatible, and minimally invasive cancer treatment is of great importance. One of the most promising research fields is the development of biocompatible photothermal nanocarriers. PTT (photothermal therapy) with an NIR (near-infrared) wavelength range (700–2000 nm) would cause cell death by increasing intercellular and intracellular temperature. PTT could also be helpful to overcome drug resistance during cancer treatments. In this study, an amine derivative of phospholipid poly ethylene glycol (DSPE-PEG (5000) amine) was conjugated with SWCNTs (single-walled carbon nanotubes) to reduce their intrinsic toxicity. Toxicity studies were performed on lung, liver, and ovarian cancer cell lines that were reported to show some degree of drug resistance to cisplatin. Toxicity results suggested that DSPE-PEG (5000) amine SWCNTs might be biocompatible photothermal nanocarriers in PTT. Therefore, our next step was to investigate the effect of DSPE-PEG (5000) amine SWCNT concentration, cell treatment time, and laser fluence on the apoptosis/necrosis of SKOV3 cells post-NIR exposure by RSM and experimental design software. It was concluded that photothermal efficacy and total apoptosis would be dose-dependent in terms of DSPE-PEG (5000) amine SWCNT concentration and fluence. Optimal solutions which showed the highest apoptosis and lowest necrosis were then achieved.

Identification of Novel Drug Candidate for Epithelial Ovarian Cancer via In Silico Investigation and In Vitro Validation

Epithelial ovarian cancer (EOC) has a poor prognosis and high mortality rate; patients are easy to relapse with standard therapies. So, there is an urgent need to develop novel drugs. In this study, differentially expressed genes (DEGs) of EOC were identified in The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. Enrichment and protein–protein interaction (PPI) analyses were performed. The drug candidate which has the possibility to treat EOC was predicted by Connectivity Map (CMAP) databases. Moreover, molecular docking was selected to calculate the binding affinity between drug candidate and hub genes. The cytotoxicity of drug candidates was assessed by MTT and colony formation analysis, the proteins coded by hub genes were detected by Western blots, and apoptosis analysis was evaluated by flow cytometry. Finally, 296 overlapping DEGs (|log 2 fold change|>1; q-value <0.05), which were principally involved in the cell cycle (p < 0.05), and cyclin-dependent kinase 1 (CDK1) were screened as the significant hub gene from the PPI network. Furthermore, the 21 drugs were extracted from CMAPs; among them, piperlongumine (PL) showed a lower CMAP score (-0.80, -62.92) and was regarded as the drug candidate. Furthermore, molecular docking results between PL and CDK1 with a docking score of –8.121 kcal/mol were close to the known CDK1 inhibitor (–8.24 kcal/mol). Additionally, in vitro experiments showed that PL inhibited proliferation and induced apoptosis via targeting CDK1 in EOC SKOV3 cells. Our results reveal that PL may be a novel drug candidate for EOC by inhibiting cell cycle.

Improvement of Cell Growth of Uterosacral Ligament Fibroblast Derived from Pelvic Organ Prolapse Patients by Cold Atmospheric Plasma Treated Liquid

Pelvic organ prolapse (POP) is a chronic disorder that affects quality of life in women. Several POP treatments may be accompanied by abrasion, constant infection, and severe pain. Therefore, new treatment methods and improvements in current treatments for POP are required. Non-thermal atmospheric-pressure plasma is a rising biomedical therapy that generates a mixed cocktail of reactive species by different mechanisms. In this study, we applied a cylinder-type dielectric barrier discharge plasma device to create a plasma-treated liquid (PTL). The PTL was added to primary cultured human uterosacral ligament fibroblast (hUSLF) cells from POP patients at each stage. Surprisingly, treatment with diluted PTL increased hUSLF cell viability but decreased ovarian cancer cell viability. PTL also decreased cell apoptosis in hUSLF cells but increased it in SKOV3 cells. Our results suggest that PTL protects hUSLF cells from cell apoptosis by controlling the p53 pathway and improves cell viability, implying that PTL is a promising application for POP therapy.

Knockdown CD44 Promotes Amyloid-Beta Degradation in Ovarian Cancer Cells By Regulating CD36 Expression

Background: CD44 is highly expressed in many cancers, including ovarian cancer. Its interactions with ligands are involved in tumor progression, prognosis, and metastasis. However, the function of CD44 in the advancement of ovarian cancer remains unclear. Methods and Results: RNA sequencing was used to investigate the possible molecules and pathways regulated by CD44 in ovarian cancer to compare gene expression in CD44-knockdown SKOV3 cells and control cells. Identify the differentially expressed genes and then proceed to functional enrichment analysis. The results showed that genes differentially expressed were enriched in ECM-receptor interaction, Protein digestion and absorption, Focal adhesion, Notch signaling pathway, microRNA in cancer, and TGF-beta signaling pathway. Furthermore, the analysis of the proteins interaction network revealed the interaction between CD44 and CD36 in SKOV3 cells. Further analysis showed that CD36, a molecule that may be involved in ECM-receptor interaction, was low expressed in CD44-knockdown SKOV3 cells. And the results showed that knockdown CD44 induces amyloid-beta degradation in ovarian cancer cells by regulating CD36 expression. The analyses of the public database demonstrated that the CD36 expression was related to the clinical survival of ovarian cancer. Conclusions: Our study showed that CD44 might up-regulate the CD36 expression in ovarian cancer, thereby exerting a cancer-promoting effect.