Ovarian Cell Lines Research Articles

Fostamatinib Inhibits the Proliferation of Ovarian Cancer Cells Through Apoptosis Induction.

Ovarian cancer remains a significant challenge due to its high mortality rate and poor prognosis, especially in advanced stages. Despite treatment advancements, issues with resistance and recurrence persist, highlighting the urgent need for new and effective therapies. This study aimed to evaluate fostamatinib, an oral spleen tyrosine kinase inhibitor initially developed for autoimmune diseases, as a potential treatment for ovarian cancer. The effects of fostamatinib on ovarian cancer cell lines were assessed using WST-1 assays for cell proliferation. Apoptosis was evaluated through TUNEL assays, DNA fragmentation analysis, and flow cytometry. Western blot analysis was used to detect cleavage of apoptotic proteins, including caspase-3 and PARP, and flow cytometry analyzed cell cycle changes. Fostamatinib treatment resulted in a dose- and time-dependent reduction in ovarian cancer cell growth and induced apoptosis, as indicated by increased TUNEL-positive cells, DNA fragmentation, and rises in both early and late apoptosis. Western blot analysis showed increased cleavage of apoptotic proteins, including caspase-3 and PARP. Flow cytometry also demonstrated an increase in the sub-G1 phase of the cell cycle, further supporting apoptosis induction. Fostamatinib, by inhibiting cell proliferation and inducing apoptosis, shows promise as a repurposed therapeutic agent for ovarian cancer, potentially offering a new approach to improve patient outcomes.

TNFRSF10D expression as a potential biomarker for cisplatin-induced damage and ovarian tumor relapse prediction

Among gynecological malignancies, ovarian cancer (OC) presents the most challenging diagnostic scenario. Despite exhaustive efforts, up to 90% of patients treated with taxane/platinum-based chemotherapy experience relapse, leading to poor survival rates. Identifying new molecular markers that can characterize disease aggressiveness, chemoresistance, recurrence risk, and metastasis is crucial. This study aimed to assess the susceptibility of three ovarian tumor cell lines (TOV-21G, SKOV-3, and OV-90) to cisplatin and paclitaxel, and to investigate the influence of these treatments on the mRNA expression of TANK, RIPK1, NFKB1, TNFRSF10D, and TRAF2. Among the cell lines, SKOV-3 ovarian adenocarcinoma cells demonstrated the highest resistance to cisplatin treatment (0.125mg/mL), followed by TOV-21G (0.076mg/mL) and OV-90 cells (0.028mg/mL). Regarding paclitaxel treatment, the SKOV-3 cell line exhibited the highest resistance (1.4µg/mL), followed by OV-90 (1.3µg/mL) and TOV-21G cells (0.9µg/mL). Gene expression analysis after paclitaxel treatment remained unchanged; however, after cisplatin treatment, TNFRSF10D was observed to be upregulated nearly 100-fold in SKOV-3 compared to all other cell lines studied. SKOV-3 is described as cisplatin and tumor necrosis factor-resistant. Despite the defective signaling of the TNFRSF10D receptor for apoptosis, it can activate the NFKB transcription factor through non-canonical TRAIL signaling, contributing to a pro-inflammatory immune response. In light of this, damage associated with cisplatin increases TNFRSF10D expression and may promote cell survival through non-canonical NFKB pathway activation. This suggests that resistance to TRAIL-induced apoptosis in these cells could serve as a promising chemoresistance biomarker in OC.

Deciphering the effect of Potentilla fulgens root extract against healthy HUVEC cell line and cancer cell lines (A549 and SKOV-3)

BackgroundPotentilla fulgens, a highly valued indigenous medicinal herb grown in high altitudes of the Himalayan region with anticancer, hypoglycaemic, antibacterial, anti-inflammatory, and antiulcerogenic properties, are used in traditional systems of medicine. This study was aimed to investigate the effect of P. fulgens root extract, as one of the natural alternatives to chemotherapeutic drugs used in cancer treatment, on proliferation, apoptosis, and autophagy of human non-small cell lung cancer cell line (A549), human ovarian cancer cell line (SKOV-3), and healthy human umbilical vein endothelial cell line (HUVEC). MethodsAnti-proliferative effect was assessed by MTT assay. The expression of autophagy and apoptosis-related proteins was evaluated by western blotting. Total oxidant status (TOS) and total antioxidant capacity (TAC) test were determined using standard kit methods. ResultsOur results showed that the extract inhibited proliferation of HUVEC, A549, and SKOV-3 cells in a dose-dependent manner. MTT assay analysis revealed that the extract significantly (P<0.05) induced mortality in HUVEC, A549, and SKOV-3 cells. Western blot results revealed increased expression of NF-κB after the extract treatment but led to the down-regulation in Beclin-1, Bax, extracellular-signal-related kinase 1 and 2, Sequestosome-1, and Cleaved Casp-3 levels. Treatment groups showed an increase in TOS and TAC values in A549 and SKOV-3 cell lines, while HUVEC cell line showed an increase in TAC and a decrease in TOS values, compared to the control group. ConclusionsOur findings indicated that P. fulgens root extract inhibited the proliferation of healthy cells and cancer cells through cell cycle arrest, representing its limited application as therapeutic agent in cancer treatment.

PGRMC1 and PAQR4 are promising molecular targets for a rare subtype of ovarian cancer.

The heterogeneity of ovarian cancer (OC) has made developing effective treatments difficult. Nowadays, hormone therapy plays a growing role in the treatment of OC; however, hormone modulators have had only limited success so far. To provide a more rigorous foundation for hormonal therapy for different OC subtypes, the current study used a series of bioinformatics approaches to analyse the expression profiles of genes encoding membrane progesterone (PGRMC1, progestins and the adipoQ receptor [PAQR] family), and androgen (zinc transporter member 9 [ZIP9], OXER1) receptors. Our work investigated also their prognostic value in the context of OC. We found differences in expression of ZIP9 and OXER1 between different OC subtypes, as well as between patient tumour and normal tissues. Expression of mRNA encoding PAQR7 and PAQR8 in a panel of OC cell lines was below the qPCR detection limit and was downregulated in tumour tissue samples, whereas high expression of PGRMC1 and PAQR4 mRNA was observed in rare subtypes of OC cell lines. In addition, chemical inhibition of PGRMC1 reduced the viability of rare OCs represented by COV434 cells. In conclusion, PGRMC1 and PAQR4 are promising targets for anticancer therapy, particularly for rare subtypes of OC. These findings may reflect differences in the observed responses of various OC subtypes to hormone therapy.

Long Non-coding RNA FOXD2-AS1 Silencing Inhibits Malignant Behaviors of Ovarian Cancer Cells Via miR-324-3p/SOX4 Signaling Axis.

It is urgent to develop new therapeutic strategies for ovarian cancer (OC). Long-noncoding RNAs (lncRNAs) have participated in multiple biological processes including tumor recurrence and progression. This study aimed to determine the effects and potential regulatory mechanism of lncRNA FOXD2-AS1 in OC progression. Levels of lncRNA FOXD2-AS1 and miR-324-3p in OC tissues and cell lines were analyzed using quantitative real-time PCR (qRT-PCR). The direct target between FOXD2-AS1 or miR-324-3p was determined using bioinformatics tools and further verified by dual-luciferase reporter assay. Cell viability, apoptosis, migration, along invasion were assessed by MTT, flow cytometry, as well as Transwell assays, respectively. In addition, the levels of miR-324-3p, PCNA, MMP9, Bax, Bcl-2, and SOX4 in OC cells were evaluated using qRT-PCR and western blot assays. We observed that lncRNA FOXD2-AS1 was up-regulated while miR-324-3p was down-regulated in OC tissues and cell lines, especially in SKOV3 cells. Moreover, miR-324-3p was a direct target of lncRNA FOXD2-AS1. Meanwhile, SOX4 interacted with miR-324-3p and was negatively regulated by miR-324-3p in SKOV3 cells. Function assays confirmed that lncRNA FOXD2-AS1 silenced depressed cell proliferation, migration, and invasion while accelerating apoptosis. These functions of lncRNA FOXD2-AS1 were attenuated by miR-324-3p inhibition. Our research demonstrated that FOXD2-AS1 silencing restrained cell growth and metastasis of OC via regulating miR-324-3p/SOX4 axis, indicating that lncRNA FOXD2-AS1 could be a novel potential therapeutic target for OC.

Development of tripeptide-cyclotriphosphazene derivatives: In vitro cytotoxicity, genotoxicity studies and molecular docking analysis within ovarian and prostate cancer cell line receptors

Peptide-phosphazene compounds are important compounds of growing interest in biomedical research and have potential therapeutic effects. The tripeptide-cyclotriphosphazenes conjugates were synthesized and analyzed for their molecular docking analysis, visualizing their binding profiles within various cancer cell line receptors and tested in vitro cytotoxic and genotoxic properties. Determining in vitro cytotoxic studies of obtained compounds displayed cytotoxic effect against two selected human cancer cell lines, including ovarian (A2780) and prostate (PC-3), cancer cells. The compound DTAP demonstrated significantly higher efficacy at 100 µM in the PC-3 cancer cell line compared to the reference drug docetaxel at 50 µM. Among the tripeptide-phosphazene conjugtates, DTGG demonstrates the most promising anticancer activity with a logIC50 of 1.23 µM, forming five hydrogen bonds and a favorable salt bridge interaction, along with several hydrophobic interactions, thereby stabilizing its binding within the human ovarian tumor domain based on molecular docking analysis. The derivative DTGP emerges as the most potent among the DTG derivatives, achieving a ΔG model value of −108 kcal/mol, primarily due to a π-cationic interaction with the LYS204 amino acid in chain C, which significantly enhances its binding affinity. Additionally, DNA damage studies on human ovarian and prostate cancer cell lines determined that cell death due to DNA damage was the basis of the decrease in cell viability. These results support the evaluation of the compounds as potential drug candidates.

Targeting Mitochondrial Cholesterol Efflux via TCF21/ABCA10 Pathway to Enhance Cisplatin Efficacy in Ovarian Cancer.

Cisplatin (DDP) resistance is one of the causes of treatment failure for ovarian cancer (OV). Mitochondrial cholesterol level was reported to be associated with OV chemoresistance. We found that ABCA10, a potential cholesterol transport protein, was highly expressed in ovarian tissues and downregulated in OV tissues. Our study aimed to explore TCF21/ABCA10 axis resistance to DDP therapy in ovarian cancer based on regulating mitochondrial cholesterol efflux. Thirty epithelial ovarian cancer tumors and thirty ovarian tissues from non-cancer patients were collected. Western blot and RT-qPCR were used to measure ABCA10 and TCF21 expression levels in these tissues, as well as in a human ovarian epithelial cell line (IOSE-80), OV cells (A2780 and SKOV3), and DDP-resistant OV cell lines (A2780/DDP and SKOV3/DDP). IOSE-80 cells were also infected with ABCA10 knockdown lentivirus to identify the most effective ABCA10 knockdown plasmid. Lentiviral infection was used to create ABCA10 knockdown, ABCA10 overexpression, and TCF21 overexpression anti-DDP OV cell lines. Cell proliferation was detected by CCK-8 and EDU staining, flow cytometry for apoptosis, MTT for metabolic activity, calcium-induced Cytochrome C release, and mitochondrial matrix swelling for mitochondrial function and Oil Red O staining for lipid accumulation. Cholesterol metabolism was evaluated by measuring mitochondrial cholesterol and cholesterol efflux. Protein concentration was determined using the BCA method. A dual-luciferase reporter assay confirmed TCF21's interaction with ABCA10. ChIP also verified this interaction. The mRNA level (P < 0.01) and protein level (P < 0.001) of ABCA10 were downregulated in cancer tissues of OV patients relative to normal ovarian tissues. Relative to human ovarian epithelial cells, ABCA10 expression was significantly downregulated in OV cells (P < 0.01) and even more significantly downregulated in DDP-resistant OV cells (P < 0.001). Compared to the group treated solely with DDP, the overexpression of ABCA10 significantly inhibited the proliferation of DDP-resistant OV cells (P < 0.01), markedly reduced the staining intensity of EDU in these cells (P < 0.05), and substantially accelerated apoptosis in DDP-resistant OV cells (P < 0.01).Overexpression of ABCA10 further accelerated Cytochrome C expression and mitochondrial matrix swelling in DDP-resistant OV cells compared to the DDP-alone group (P < 0.01). The addition of cholesterol reversed the decrease in lipid accumulation, the decrease in mitochondrial cholesterol levels (P < 0.05), and the increase in cholesterol efflux (P < 0.01) in DDP-resistant OV cells caused by overexpression of ABCA10. The transcription factor TCF21 was bound to the promoter of ABCA10. Overexpression of TCF21 significantly increased ABCA10 expression in DDP-resistant OV cells (P < 0.01) and increased cytochrome C expression in A2780/DDP (P < 0.05) and SKOV3/DDP (P < 0.01) cells, with accelerated mitochondrial matrix swelling in A2780/DDP (P < 0.01) and SKOV3/DDP (P < 0.001) cells, while knockdown of ABCA10 reversed these effects. Our study found that TCF21 boosts ABCA10 expression, which in turn reduces DDP resistance in OV cells by enhancing mitochondrial cholesterol efflux. This mechanism increases the sensitivity of DDP-resistant OV cells to DDP. Our findings will provide new therapeutic targets for the treatment of ovarian cancer.

Anti-ovarian cancer migration and toxicity characteristics of a platinum(IV) pro-drug with axial HDAC inhibitor ligands in zebrafish models.

Ovarian cancer is the fifth leading cause of cancer related death in the United States. Cisplatin is a platinum-based anti-cancer drug used against ovarian cancer that enters malignant cells and then damages DNA causing cell death. Typically, ovarian cancer cells become resistant to cisplatin making it necessary to increase subsequent dosage, which usually leads to side-effects including irreversible damage to kidney and auditory system tissue. Ovarian cancer resistance is often associated with upregulation of histone deacetylase (HDAC) enzymes that cause DNA to adopt a closed configuration which reduces the ability of cisplatin to target and damage DNA. Compound B, a platinum(IV) complex with two axial phenylbutyrate (PBA) HDAC inhibitor ligands attached to a cisplatin core, can simultaneously inhibit HDAC activity and damage DNA causing decreased cancer cell viability in cisplatin-sensitive (A2780) and -resistant (A2780cis) ovarian cancer cell lines. However, compound B was not previously evaluated in vivo. As simultaneously inhibiting HDAC-mediated resistance with cisplatin treatment could potentiate the platinum drug's effect, we first confirmed the anti-cancer effect of compound B in the A2780 and A2780cis cell lines using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide spectrophotometric assay. Then, we used zebrafish embryo and transgenic animal models to comparatively analyze the effect of cisplatin, compound B, and controls on general organismal, auditory, and renal system toxicity, and cancer metastasis. We found that lower dosages of compound B (0.3 or 0.6 µM) than of cisplatin (2.0 µM) could cause similar or decreased levels of general, auditory, and renal tissue toxicity, and at 0.6 µM, compound B reduces cancer metastasis more than 2.0 µM cisplatin.

Protective Effects of Nettle Tea on SKOV-3 Ovarian Cancer Cells Through ROS Production, Apoptosis Induction, and Motility Inhibition Without Altering Autophagy.

Urtica dioica L. (UD), also known as the stinging nettle, has long been used in traditional medicine for its wide range of health benefits. The current study focuses on the effect of nettle tea on the growth and proliferation of one of the most aggressive ovarian adenocarcinoma cell line, SKOV-3 cells. To examine this, cytotoxicity, cell cycle analysis, and ROS assays were performed, along with Annexin V/PI dual staining, cell death ELISA, Western blot analysis, and motility assays. The results showed that a UD aqueous extract (UDAE) can inhibit the growth and proliferation of SKOV-3 cells in a dose- and time-dependent manner by promoting cellular fragmentation. This was accompanied by an increase in two apoptotic hallmarks, the flipping of phosphatidylserine to the outer membrane leaflet and DNA fragmentation as revealed by cell death ELISA. This aqueous extract showed a pro-oxidant activity while also activating the extrinsic caspase-dependent apoptotic pathway with no alteration in autophagy markers. Furthermore, the extract showed promising inhibitory effect on the migratory capacities of aggressive ovarian cancer cells, in vitro.

Exploring the Antitumor Potential of New Indazole-indolizines Designed by Molecular Hybridization

Background: Cancer represents the major health problem faced by the population of the world, remaining one of the main causes of death. Hence, the development of new targeted antitumor drugs with high efficacy and lower toxicity is still needed. Objective: As a continuation of our work to discover new molecules with cytotoxic properties, two heterocyclic scaffolds, namely indolizine, and indazole, were combined in the same molecule, aiming to improve the bioactivity. This article focused on the synthesis, characterization, and biological evaluation of a series of new indazole-indolizine hybrid compounds. Methods: The biological potential of the synthesized compound was investigated in vitro against the human farnesyltransferase enzyme and NCI 60 tumor cell lines panel. While the farnesyltransferase inhibitory activity was modest, a very good antiproliferative action was observed for compound 4a, which, at a concentration of 10 μM, inhibited the growth of 20 types of cancer cells by more than 50% and showed cytotoxic action against the ovarian cancer cell line OVCAR-4. Results: A series of novel indazole-indolizine hybrids were synthesized via a [3+2] cycloaddition reaction, fully characterized and biologically evaluated for antitumor potential. Conclusion: Compound 4a could be a promising starting point in the development of new antitumoral agents. Further biological investigations will be performed to identify the biological target of the compounds. Moreover, different synthetic strategies to introduce new substituents on the indolizine core will be addressed.

Identification of three subtypes of ovarian cancer and construction of prognostic models based on immune-related genes

BackgroundImmunotherapy has revolutionized the treatment of ovarian cancer (OC), but different immune microenvironments often constrain the efficacy of immunotherapeutic interventions. Therefore, there is an imperative to delineate novel immune subtypes for development of efficacious immunotherapeutic strategies.MethodsThe immune subtypes of OC were identified by consensus cluster analysis. The differences in clinical features, genetic mutations, mRNA stemness (mRNAsi) and immune microenvironments were analyzed among subtypes. Subsequently, prognostic risk models were constructed based on differentially expressed genes (DEGs) of the immune subtypes using weighted correlation network analysis.ResultsOC patients were classified into three immune subtypes with distinct survival rates and clinical features. Different subtypes exhibited varying tumor mutation burdens, homologous recombination deficiencies, and mRNAsi levels. Significant differences were observed among immune subtypes in terms of immune checkpoint expression and immunogenic cell death. Prognostic risk models were validated as independent prognostic factors demonstrated great predictive performance for survival of OC patients.ConclusionIn this study, three distinct immune subtypes were identified based on gene sets related to vaccine response, with the C2 subtype exhibiting significantly worse prognosis. While no statistically significant differences in tumor mutation burden (TMB) were observed across the three subtypes, the homologous recombination deficiency (HRD) score and mRNA stemness index (mRNAsi) were notably elevated in the C2 group compared to the others. Immune infiltration analysis indicated that the C2 subtype may have an increased presence of regulatory T (Treg) cells, potentially contributing to a more favorable response to combination therapies involving PARP inhibitors and immunotherapy. These findings offer a precision medicine approach for tailoring immunotherapy in ovarian cancer patients. Moreover, the C3 subtype demonstrated significantly lower expression levels of immune checkpoint genes, a pattern validated by independent datasets, and associated with a better prognosis. Further investigation revealed that the immune-related gene FCRL5 correlates with ovarian cancer prognosis, with in vitro experiments showing that it influences the proliferation and migration of the ovarian cancer cell line SKOV3.

Non-B DNA-informed mutation burden as a marker of treatment response and outcome in cancer.

Genomic instability is crucial in tumorigenesis, with Tumour Mutation Burden (TMB) being a biomarker to indicate therapeutic effectiveness, particularly in immunotherapy. However, TMB is not always a reliable predictor and displays heterogeneity. Non-B DNA, susceptible to mutations, play a significant role in cancer development, indicating their potential merit when combined with mutation for enhanced markers in cancer. We assessed mutations and non-B DNA interplay as biomarkers. Our methodology quantifies tumour mutations and their co-localization with non-B DNA, using survival and drug sensitivity assessments for clinical relevance. We introduce two novel markers, 'nbTMB' (non-B-informed tumour mutation burden) and 'mlTNB' (mutation-localised tumour non-B burden). In case studies: (1) nbTMB informs on survival heterogeneity among TMB-high patients undergoing immunotherapy whereas TMB is unable to further differentiate; (2) nbTMB informs on altered cisplatin sensitivity among ovarian cancer cell lines whereas TMB is unable to differentiate; and (3) mlTNB informs on survival heterogeneity among early-stage pancreatic cancer progressors in whom other markers of genomic instability fail to differentiate. These novel markers offer a nuanced approach to enhance our understanding of treatment responses and outcomes in cancer, underscoring the need for a comprehensive exploration of the interplay between non-B and B-DNA features.

GC-MS/HPLC Profiling and Sono-maceration Mediated Extraction of Osbeckia parvifolia Polyphenols: In Silico and In Vitro Analysis on Anti-Proliferative activity in Ovarian Cancer Cell Lines.

Osbeckia parvifolia, an endemic edible plant of Western Ghats, was investigated in the present study for its polyphenolic compounds, including content, constituents, extraction through an ultrasonic-assisted maceration technique and therapeutic potential in biomedical applications. The methanolic extract (OPM) exhibited an IC50 value of 1.25 µg/mL against 2,2-Diphenyl-1-picrylhydrazyl (DPPH) radicals. Furthermore, the ethyl acetate and methanolic extracts also strongly inhibited 5-lipoxygenase, especially OPM (84.93%), which was comparable to standard curcumin. OPM also elicited cytotoxicity in SKOV3 ovarian cancer cells (93.80%), surpassing paclitaxel. Bio-accessibility analysis demonstrated that the release of phenolic compounds and antioxidant potential were very high (above 100%), revealing the possibility of synergistic efficacy of polyphenolic complexes in drug development. Gas Chromatography -Mass Spectrometry (GC-MS) analysis revealed 22 bioactive polyphenolic compounds in OPM, such as epicatechin, quercetin, and psoralidin. This was confirmed by High Performance Liquid Chromatography (HPLC) and High-Pressure Thin Layer Chromatography (HPTLC) analyses, which revealed a high quantity of catechin (37.45 mg/g). Molecular docking revealed the significant binding affinity of these proteins for the ovarian oncoproteins PI3K (-8.52 kcal/mol) and Casp-8 (-8.41 kcal/mol). Adsorption, Distribution, Metabolism, Excretion and Toxicity (ADMET) profiling indicated the favorable pharmacokinetic properties of these compounds, supporting their candidacy in drug formulations against ovarian cancer.

Functional Assessments of Gynecologic Cancer Models Highlight Differences Between Single-Node Inhibitors of the PI3K/AKT/mTOR Pathway and a Pan-PI3K/mTOR Inhibitor, Gedatolisib

Background/Objectives: The PI3K/AKT/mTOR (PAM) pathway is frequently activated in gynecological cancers. Many PAM inhibitors selectively target single PAM pathway nodes, which can lead to reduced efficacy and increased drug resistance. To address these limitations, multiple PAM pathway nodes may need to be inhibited. Gedatolisib, a well-tolerated panPI3K/mTOR inhibitor targeting all Class I PI3K isoforms, mTORC1 and mTORC2, could represent an effective treatment option for patients with gynecologic cancers. Methods: Gedatolisib and other PAM inhibitors (e.g., alpelisib, capivasertib, and everolimus) were tested in endometrial, ovarian, and cervical cancer cell lines by using cell viability, cell proliferation, and flow cytometry assays. Xenograft studies evaluated gedatolisib in combination with a CDK4/6 inhibitor (palbociclib) or an anti-estrogen (fulvestrant). A pseudo-temporal transcriptomic trajectory of endometrial cancer clinical progression was computationally modeled employing data from 554 patients to correlate non-clinical studies with a potential patient group. Results: Gedatolisib induced a substantial decrease in PAM pathway activity in association with the inhibition of cell cycle progression and the decreased cell viability in vitro. Compared to single-node PAM inhibitors, gedatolisib exhibited greater growth-inhibitory effects in almost all cell lines, regardless of the PAM pathway mutations. Gedatolisib combined with either fulvestrant or palbociclib inhibited tumor growth in endometrial and ovarian cancer xenograft models. Conclusions: Gedatolisib in combination with other therapies has shown an acceptable safety profile and promising preliminary efficacy in clinical studies with various solid tumor types. The non-clinical data presented here support the development of gedatolisib combined with CDK4/6 inhibitors and/or hormonal therapy for gynecologic cancer treatment.

In Vitro Methylene Blue and Carboplatin Combination Triggers Ovarian Cancer Cells Death.

Ovarian cancer presents a dire prognosis and high mortality rates, necessitating the exploration of alternative therapeutic avenues, particularly in the face of platinum-based chemotherapy resistance. Conventional treatments often overlook the metabolic implications of cancer, but recent research has highlighted the pivotal role of mitochondria in cancer pathogenesis and drug resistance. This study delves into the metabolic landscape of ovarian cancer treatment, focusing on modulating mitochondrial activity using methylene blue (MB). Investigating two epithelial ovarian cancer (EOC) cell lines, OV1369-R2 and OV1946, exhibiting disparate responses to carboplatin, we sought to identify metabolic nodes, especially those linked to mitochondrial dysfunction, contributing to chemo-resistance. Utilizing ARPE-19, a normal retinal epithelial cell line, as a control model, our study reveals MB's distinct cellular uptake, with ARPE-19 absorbing 5 to 7 times more MB than OV1946 and OV1369-R2. Treatment with 50 µM MB (MB-50) effectively curtailed the proliferation of both ovarian cancer cell lines. Furthermore, MB-50 exhibited the ability to quell glutaminolysis and the Warburg effect in cancer cell cultures. Regarding mitochondrial energetics, MB-50 spurred oxygen consumption, disrupted glycolytic pathways, and induced ATP depletion in the chemo-sensitive OV1946 cell line. These findings highlight the potential of long-term MB exposure as a strategy to improve the chemotherapeutic response in ovarian cancer cells. The ability of MB to stimulate oxygen consumption and enhance mitochondrial activity positions it as a promising candidate for ovarian cancer therapy, shedding light on the metabolic pressures exerted on mitochondria and their modulation by MB, thus contributing to a deeper understanding of mitochondrial dysregulation and the metabolic underpinnings of cancer cell proliferation.

Withaferin A ameliorates ovarian cancer-induced renal damage through the regulation of expression of inflammatory cytokines

BackgroundCachexia a multifactorial syndrome is a common sequala in patients with cancer. It varies from 42 to 80% depending upon the oncological stage and is directly responsible for 30% of deaths in these patients. Previous research from our laboratory demonstrated that peritoneal ovarian cancer generated in NSG mice resulted in skeletal and cardiac muscle atrophy - leading to loss of skeletal muscle mass and strength, and cardiac dysfunction (cachexia). Treatment of mice bearing i.p. tumors with withaferin A (WFA) showed reversal of skeletal muscle and cardiac cachexia. The present study is focused on determining effects of peritoneal ovarian tumors on kidney damage and effects of WFA treatment on ameliorating kidney damage.MethodsWe generated intraperitoneal ovarian cancer by injecting female NSG mice with ovarian cancer cell line (A2780). After one week of injecting cancer cells, mice were treated with WFA (4 mg/kg) every third day, for three weeks. After 4 weeks of injection of cancer cells, the mice were sacrificed and various tissues including kidney and blood were collected, snap-frozen in liquid nitrogen, and stored at -800C. The presence of kidney biomarker creatinine, was measured in the plasma by an ELISA. The mRNA was isolated from mouse kidneys and was used to examine the expression levels of signaling proteins, inflammatory cytokines, and genes responsible for inducing cachexia (IL-1β, IL-6, TNF-α, TGF-β, GDF-15, and MYD88).ResultsOur results showed a significant increase in levels of expression of inflammatory cytokine IL-1 β (p < 0.01), IL-6 (p < 0.001), TNF-α (p < 0.001), and other related genes including TRAF6 (p < 0.01), MYD88 (p < 0.01), and GDF-15 (p = 0.005) in tumor-bearing mice compared to controls. Treatment of mice bearing tumors with WFA attenuated the increase in expression of each gene. In addition, our results showed a significant increase in creatinine levels in circulation in tumor-bearing mice compared to control mice. Treatment of tumor-bearing mice with WFA resulted in a significant decrease in plasma creatinine levels compared to tumor-bearing mice.ConclusionsOur results conclude that ovarian tumors in NSG mice caused kidney damage and renal dysfunction, which was effectively ameliorated by WFA treatment, suggesting a protective effect of WFA on kidney injury induced by ovarian cancer.

Multicellular ovarian cancer spheroids: novel 3D model to mimic tumour complexity

In vitro, spheroid models have become well established in cancer research because they can better mimic certain characteristics of in vivo tumours. However, interaction with the tumour microenvironment, such as cancer-associated fibroblasts, plays a key role in tumour progression. We initially focused on the interaction of tumour cells with fibroblasts. To model this interaction, we developed a spheroid model of ovarian cancer and fibroblasts. To this end, ovarian cancer cell lines and ex vivo primary cells were simultaneously and sequentially seeded with fibroblasts in a scaffold-free system at different ratios and subsequently characterized with respect to changes in morphology, proliferation, and viability. We demonstrated that co-cultures are able to form by far more compact spheroids, especially in cells that form aggregates in mono-culture. In addition, the co-cultures were able to increase proliferation and sensitivity to cisplatin. Simultaneous seeding led fibroblasts invade the core in both cell lines and primary cells. These results show differences in formation, firmness, and size between co-culture and mono-culture. Our model is designed to better represent and characterize the mutual influencing factors of fibroblasts and tumour cells. Fibroblast-supplemented multicellular spheroids are a valuable tool for tumour microenvironment interaction and new drug discovery.

Preparation of dual drug-loaded polymer nanoconjugate to enhance treatment efficacy for ovarian cancer cells

Ovarian cancer is the deadliest gynecological malignancy, representing 2.5 % of all female cancers and accounting for 5 % of female cancer-related fatalities. Despite numerous strategies in its treatment, the disease shows a high recurrence rate and a low survival rate. Consequently, there is a growing focus on targeted therapies in ovarian cancer treatment. It is well-known that VEGFR and LPA pathways undergo alterations in ovarian cancer and stimulate survival, adhesion, migration, invasion, tumor growth and angiogenesis. Cabozantinib (CBZ) is a multi-receptor tyrosine kinase inhibitor that effectively targets MET, VEGFR-1, 2, 3, FLT3, c-KIT, and RET. Ki16425 is a selective inhibitor of LPA receptors 1, 2, and 3. Therefore, targeting LPA receptors and combining with VEGFR inhibitor is a strategic approach for ovarian cancer treatment. In this study, it was aimed to prepare polymer-drug nanoconjugate for both VEGFR and LPAR inhibition. For this, O-(2-Carboxyethyl) polyethylene glycol (PEG5000) which advantages are known in cancer studies, was chosen as the carrier system, and a nanoconjugate containing Ki16425 and CBZ (Ki-PEG-CBZ) was synthesized and its potential was evaluated. Initially, CBZ and Ki16425 were conjugated to the PEG5000 through pH-sensitive hydrazone and ester bonds. After nanoconjugate characterization, in vitro release and its ovarian cancer treatment potential were evaluated on A2780, OVCAR3 and SKOV3 ovarian cancer cell lines. A nanoconjugate was obtained with a particle size of 169 ± 15.23 nm, a zeta potential of −13.5 ± 1.21 mV, and a release profile lasting 48 h, containing CBZ and Ki16425 with drug loading efficiencies of 73.71 ± 0.53 % and 77.72 ± 2.51 %, respectively. In vitro studies have demonstrated that Ki-PEG-CBZ is highly effective against ovarian cancer. We suggest that the developed polymer-drug nanoconjugate is an effective and safe nanoconjugate for the treatment of ovarian cancer.

Investigation of the Immunoexpression of SVIP and UPR Pathway Proteins in Ovarian Adenocarcinoma Cell Line OVCAR-3

Ovarian cancer is the deadliest gynecological cancer. The endoplasmic reticulum (ER), a vital cell organelle, is involved in the folding, synthesis, and modification of a wide range of soluble and insoluble proteins. ER stress initiates the unfolded protein response (UPR), an evolutionary conserved cell stress mechanism. The UPR is mediated by three ER transmembrane sensors: IRE1, ATF6, and PERK. An inhibitor of ERAD is a small VCP/p97-interacting protein (SVIP). The study aimed to investigate the relationship between SVIP and the ER stress protein markers in the human ovarian cancer cell line OVCAR-3. The SVIP and GRP78, PERK, ATF4 immunoexpression levels were analyzed. Furthermore, employing immunofluorescence, the colocalization of three ER sensors and SVIP was ascertained. The immunoexpression of SVIP and GRP78, ATF4, and PERK were shown in the OVCAR-3 cell line. Additionally, immunofluorescence results showed the colocalization of SVIP and UPR-related proteins in the cytoplasm of OVCAR-3 cells. In conclusion, we demonstrated the cellular localization of SVIP and the proteins involved in the UPR pathway. However, further studies are needed to determine the relation between SVIP and these proteins in cancer cells.

Data mining of PubChem bioassay records reveals diverse OXPHOS inhibitory chemotypes as potential therapeutic agents against ovarian cancer

Focused screening on target-prioritized compound sets can be an efficient alternative to high throughput screening (HTS). For most biomolecular targets, compound prioritization models depend on prior screening data or a target structure. For phenotypic or multi-protein pathway targets, it may not be clear which public assay records provide relevant data. The question also arises as to whether data collected from disparate assays might be usefully consolidated. Here, we report on the development and application of a data mining pipeline to examine these issues. To illustrate, we focus on identifying inhibitors of oxidative phosphorylation, a druggable metabolic process in epithelial ovarian tumors. The pipeline compiled 8415 available OXPHOS-related bioassays in the PubChem data repository involving 312,093 unique compound records. Application of PubChem assay activity annotations, PAINS (Pan Assay Interference Compounds), and Lipinski-like bioavailability filters yields 1852 putative OXPHOS-active compounds that fall into 464 clusters. These chemotypes are diverse but have relatively high hydrophobicity and molecular weight but lower complexity and drug-likeness. These chemotypes show a high abundance of bicyclic ring systems and oxygen containing functional groups including ketones, allylic oxides (alpha/beta unsaturated carbonyls), hydroxyl groups, and ethers. In contrast, amide and primary amine functional groups have a notably lower than random prevalence. UMAP representation of the chemical space shows strong divergence in the regions occupied by OXPHOS-inactive and -active compounds. Of the six compounds selected for biological testing, 4 showed statistically significant inhibition of electron transport in bioenergetics assays. Two of these four compounds, lacidipine and esbiothrin, increased in intracellular oxygen radicals (a major hallmark of most OXPHOS inhibitors) and decreased the viability of two ovarian cancer cell lines, ID8 and OVCAR5. Finally, data from the pipeline were used to train random forest and support vector classifiers that effectively prioritized OXPHOS inhibitory compounds within a held-out test set (ROCAUC 0.962 and 0.927, respectively) and on another set containing 44 documented OXPHOS inhibitors outside of the training set (ROCAUC 0.900 and 0.823). This prototype pipeline is extensible and could be adapted for focus screening on other phenotypic targets for which sufficient public data are available.Scientific contributionHere, we describe and apply an assay data mining pipeline to compile, process, filter, and mine public bioassay data. We believe the procedure may be more broadly applied to guide compound selection in early-stage hit finding on novel multi-protein mechanistic or phenotypic targets. To demonstrate the utility of our approach, we apply a data mining strategy on a large set of public assay data to find drug-like molecules that inhibit oxidative phosphorylation (OXPHOS) as candidates for ovarian cancer therapies.Graphical