Ascites Tumor Cells Research Articles

Intraperitoneal administration of doxorubicin-encapsulated Brucea javanica oil nanoemulsion against malignant ascites

Malignant ascites is a common complication of advanced cancers, which reduces survival rates and diminishes patients’ quality of life. Intraperitoneal chemotherapy is a conventional method for treating cancer-related ascites, but the poor drug retention of conventional drugs requires frequent administration to maintain sustained anti-tumor effects. In this study, we encapsulated doxorubicin (DOX) into Brucea javanica oil (BJO) to develop a water-in-oil (W/O) nanoemulsion called BJO@DOX for the treatment of malignant ascites through in-situ intraperitoneal administration. BJO significantly induced apoptosis of S180 cells by upregulating the expression of p53 and caspase-3 (cleaved). Additionally, BJO notably downregulated the expression of Bcl-2, further promoting apoptosis of S180 cells. Cell apoptosis significantly inhibited ascites formation and tumor cell proliferation in a mouse model. The combination of DOX and BJO exhibited satisfactory synergistic effects, consequently prolonging the survival period of mice. Histological examination of major organs indicated that the nanoemulsion had excellent biosafety in vivo. The BJO@DOX nanoemulsion represents a promising platform for in-situ chemotherapy of malignant ascites.

Synergic activity of FGFR2 and MEK inhibitors in the treatment of FGFR2-amplified cancers of unknown primary

Patients with cancer of unknown primary (CUP) carry the double burden of an aggressive disease and reduced access to therapies. Experimental models are pivotal for CUP biology investigation and drug testing. We derived two CUP cell lines (CUP#55 and #96), and corresponding patient-derived xenografts (PDXs), from ascites tumor cells. CUP cell lines and PDXs underwent histological, immune-phenotypical, molecular, and genomic characterization confirming the features of the original tumor. The tissue-of-origin prediction was obtained from the tumor microRNA expression profile and confirmed by single-cell transcriptomics. Genomic testing and FISH analysis identified FGFR2 gene amplification in both models, in the form of homogenously staining region (HSR) in CUP#55 and double minutes in CUP#96. FGFR2 was recognized as the main oncogenic driver and therapeutic target. FGFR2-targeting drug BGJ-398 (infigratinib) in combination with the MEK inhibitor trametinib proved to be synergic and exceptionally active, both in vitro and in vivo. The effects of the combined treatment by single-cell gene expression analysis revealed a remarkable plasticity of tumor cells and the greater sensitivity of cells with epithelial phenotype. This study brings personalized therapy closer to CUP patients and provides the rationale for FGFR2 and MEK targeting in metastatic tumors with FGFR2 pathway activation.

Suppression of Ehrlich Ascites Tumor Cell Proliferation via G1 Arrest Induced by Dietary Nucleic Acid-Derived Nucleosides

Suppression of Ehrlich Ascites Tumor Cell Proliferation via G1 Arrest Induced by Dietary Nucleic Acid-Derived Nucleosides

Homologous-magnetic dual-targeted metal-organic framework to improve the Anti-hepatocellular carcinoma efficacy of PD-1 inhibitor

The insufficient abundance and weak activity of tumour-infiltrating lymphocytes (TILs) are two important reasons for the poor efficacy of PD-1 inhibitors in hepatocellular carcinoma (HCC) treatment. The combined administration of tanshinone IIA (TSA) and astragaloside IV (As) can up-regulate the abundance and activity of TILs by normalising tumour blood vessels and reducing the levels of immunosuppressive factors respectively. For enhancing the efficacy of PD-1 antibody, a magnetic metal–organic framework (MOF) with a homologous tumour cell membrane (Hm) coating (Hm@TSA/As-MOF) is established to co-deliver TSA&As into the HCC microenvironment. Hm@TSA/As-MOF is a spherical nanoparticle and has a high total drug-loading capacity of 16.13 wt%. The Hm coating and magnetic responsiveness of Hm@TSA/As-MOF provide a homologous-magnetic dual-targeting, which enable Hm@TSA/As-MOF to counteract the interference posed by ascites tumour cells and enhance the precision of targeting solid tumours. Hm coating also enable Hm@TSA/As-MOF to evade immune clearance by macrophages. The release of TSA&As from Hm@TSA/As-MOF can be accelerated by HCC microenvironment, thereby up-regulating the abundance and activity of TILs to synergistic PD-1 antibody against HCC. This study presents a nanoplatform to improve the efficacy of PD-1 inhibitors in HCC, providing a novel approach for anti-tumour immunotherapy in clinical practice.

Specific lineage transition of tumor-associated macrophages elicits immune evasion of ascitic tumor cells in gastric cancer with peritoneal metastasis.

Gastric cancer with peritoneal metastasis (PM-GC), recognized as one of the deadliest cancers. However, whether and how the tumor cell-extrinsic tumor microenvironment (TME) is involved in the therapeutic failure remains unknown. Thus, this study systematically assessed the immunosuppressive tumor microenvironment in ascites from patients with PM-GC, and its contribution to dissemination and immune evasion of ascites-disseminated tumor cells (aDTCs). Sixty-three ascites and 43 peripheral blood (PB) samples from 51 patients with PM-GC were included in this study. aDTCs in ascites and circulating tumor cells (CTCs) in paired PB were immunophenotypically profiled. Using single-cell RNA transcriptional sequencing (scRNA-seq), crosstalk between aDTCs and the TME features of ascites was inspected. Further studies on the mechanism underlying aDTCs-immune cells crosstalk were performed on in vitro cultured aDTCs. Immune cells in ascites interact with aDTCs, prompting their immune evasion. Specifically, we found that the tumor-associated macrophages (TAMs) in ascites underwent a continuum lineage transition from cathepsinhigh (CTShigh) to complement 1qhigh (C1Qhigh) TAM. CTShigh TAM initially attracted the metastatic tumor cells to ascites, thereafter, transitioning terminally to C1Qhigh TAM to trigger overproliferation and immune escape of aDTCs. Mechanistically, we demonstrated that C1Qhigh TAMs significantly enhanced the expression of PD-L1 and NECTIN2 on aDTCs, which was driven by the activation of the C1q-mediated complement pathway. For the first time, we identified an immunosuppressive macrophage transition from CTShigh to C1Qhigh TAM in ascites from patients with PM-GC. This may contribute to developing potential TAM-targeted immunotherapies for PM-GC.

Abstract B066: Elucidating the impact of ascites on high grade serous ovarian cancer through in vitro multi-model characterization

Abstract High grade serous ovarian cancer (HGSOC) is one of the most lethal gynecological neoplasia worldwide, due to late diagnosis, high relapse rate and drug resistance. At the time of the diagnosis, HGSOC patients frequently present ascites with intraperitoneal tumor dissemination. Metastasis occurs through the shedding of cells from the primary tumor and movement to the peritoneal cavity, with the generation of ascites, where cells can grow in the absence of adhesion and eventually adhere to distant tissues. In this context, malignant ascites acts as a source of soluble factors which provide a pro-inflammatory, immunosuppressive and tumour-promoting microenvironment for tumoral cells in the peritoneum, contributing to tumor progression. Thus, understanding the influence of ascitic factors on tumoral cells would allow to identify pathogenetic mechanisms associated to metastasis, cancer stemness and chemoresistance, and be crucial for target discovery and drugs development. Our approach is based on the use of 2D and 3D patient-derived in vitro culturing models obtained from ascites of HGSOC patients, cultured in parallel in the standard conditions and with the supplementation of ascitic fluid. Specifically, we compared freshly isolated ascites tumor cells (i.e. never put in culture), 2D primary cells and two types of 3D spheroids, as models that enrich for ovarian cancer stem cells (CSCs), namely bulk spheroids and single-cell Metastatic Ovarian Cancer Spheroids (sMOCS) (Velletri, Villa, Cilli, Barzaghi et al. 2021). First, we investigated the role of ascitic fluid in supporting the growth of our models, recreating the peculiar tumor microenvironment of peritoneal ascites: we showed an increase in proliferation rate, propagability, sphere forming efficiency and dimension in the presence of ascitic fluid. Also, we performed bulk RNAseq analysis showing that ascitic fluid supplementation allows to preserve the metabolic status of fresh samples, typically lost in standard culturing conditions. In parallel, 2D cells and 3D spheroids showed a model-specific effect of ascites’ supplementation. In 2D primary cells it caused the downregulation of genes involved in the biosynthesis of cholesterol and in inflammatory response, while in 3D spheroids it induced upregulation of pathways involved in proliferation and glycolysis, suggesting a phenotype specific for CSCs. Moreover, our data allowed the scoring of CSC-related genes, upregulated in our 3D models compared to fresh ascites and 2D cells, and ascites-dependent genes, as they are up/downregulated in 2D cells and/or spheroids in the presence of ascitic fluid. These results shed light on the specific effects of ascites acting as microenvironment on tumor cells, highlighting its relevance for in vitro patient-derived models to better capture tumor features and to improve culturing conditions. Also, they pave the way to the identification of new disease-relevant pathways and targets to be exploited for more refined care of patients. Citation Format: Bianca Barzaghi, Pietro Lo Riso, Marta Rosa Sallese, Carlo Emanuele Villa, Giulia Picco, Alessia Gatto, Ugo Cavallaro, Giuseppe Testa. Elucidating the impact of ascites on high grade serous ovarian cancer through in vitro multi-model characterization [abstract]. In: Proceedings of the AACR Special Conference on Ovarian Cancer; 2023 Oct 5-7; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_2):Abstract nr B066.

Suppression of Ehrlich ascites tumor cell proliferation via G1 arrest induced by dietary nucleic acid-derived nucleosides.

The nucleic acids found in food play a crucial role in maintaining various bodily functions. This study investigated the potential anticancer effects of dietary nucleic acids, an area that is still not fully understood. By utilizing an in vivo mouse model and an in vitro cell model, we discovered an anti-proliferative impact of RNA in both systems. DNA exhibited anti-proliferative effects in the mouse model, while this phenomenon wasn't observed in the in vitro cell model using Ehrlich ascites tumor (EAT) cells. Conversely, DNA hydrolysate demonstrated distinct anti-proliferative effects in EAT cells, suggesting that nucleotides or nucleosides generated during nucleic acid digestion act as active constituents. Furthermore, we examined various nucleosides and two sodium-independent equilibrative nucleoside transporter inhibitors (ENTs), identifying guanosine and 2'-deoxyguanosine as pivotal in the anti-proliferative effect. We also found that the anti-proliferation activity with both nucleosides was suppressed by the treatment of dipyridamole, a non-selective inhibitor for ENT1 and ENT2, but not nitrobenzylthioinosine, a low inhibitor for ENT2. The uptake of these compounds into cells is likely facilitated by ENT2. These nucleotides impeded the progression of cancer cells from the G1 phase to the S phase in the cell cycle. Another significant finding is the increased expression of CCAAT/enhancer-binding protein (C/EBPβ) induced by guanosine and 2'-deoxyguanosine. Furthermore, immunostaining revealed that C/EBPβ diffuses into the nucleus, indicating its presence. This suggests that guanosine or 2-deoxyguanosine induces G1 arrest in cancer cells via the activation of C/EBPβ. Encouraged by these promising results, guanosine and 2'-deoxyguanosine show potential applications in cancer prevention.

Euphorbia grantii Oliv. standardized extract and its fraction ameliorate doxorubicin-induced cardiomyopathy in Ehrlich carcinoma bearing mice

Ethnopharmacological relevanceEuphorbia plants have long been used as traditional medicine in China, Europe, America, Turkey, India, Africa, Iran, and Pakistan because of its high medicinal value and health advantages especially as a remedy for several types of cancer. Aim of the studyDoxorubicin (DOX) is one of the most frequently prescribed drugs in cancer chemotherapy, with dose-limiting cardiotoxicity. The development of medicinal approaches to attenuate drug's toxicity represents an area of great concern in cancer research. Because research on this topic is still disputed and limited, we aim to investigate the potential of supplementation with Euphorbia grantii Oliv. on DOX-induced cardiomyopathy in Ehrlich carcinoma bearing mice. Materials and methodsThe high-performance thin layer chromatography (HPTLC) analysis of total methanolic extract (TE), and its bioactive dichloromethane fraction (DCMF) was applied for the determination of friedelin. Male BALB/c mice were used to keep the Ehrlich ascites tumor cells. The experiment was performed for a 2-weeks period. ResultsA good linearity relationship was found to be with correlation coefficient (r2) value of 0.9924 for the isolated friedelin. Limit of detection (LOD) and limit of quantitation (LOQ) was found to be 0.00179, and 0.000537 ng/band respectively for friedelin. The amount of friedelin in the TE and DCMF were determined by using calibration curve of standard as 106.32 ± 5.69 μg, and 159.2 ± 4.24 μg friedelin/mg extract, respectively. DOX-induced cardiomyopathy by decreasing the ejection fraction (EF) compared to the Ehrlich and negative control groups. It resulted in a decrease in the EF by 30 and 39% compared to the other groups. High and low doses of the TE and DCMF did not result in significantly different ejection fractions compared to the Ehrlich group. Co-administration of DCMF with DOX ameliorated the alteration in the serum CKMB and LDH levels. As revealed from histopathological study, DOX impairs viability of cardiac myocytes and DCMF could effectively and extensively counteract this action of DOX and potentially protect the heart from severe toxicity of DOX. ConclusionsFinally, our results indicated that Euphorbia grantii Oliv. would be the best option to reduce DOX adverse effects.

Synthesis and characterisation of copper acetate mediated oxidative coupled product of naturally occurring hydroxy benzoquinone, embelin and its in vitro and in vivo cytotoxic studies in Dalton Lymphoma ascites tumour cells

Embelin is an active phytoconstituent known to exhibit a variety of biological activities, especially active against various cancer and tumour cell lines. In the present work, embelin was extracted and isolated from Embelia ribes and was structurally modified by incorporating different fluoro substituted aniline in the quinone motif with a view of enhancing the biological activity. The synthesis was carried out in presence of copper acetate catalyst in a protic solvent, glacial acetic acid to obtain EDFA, ETFA and EOCF and were characterised by various spectral techniques. Embelin and its derivatives were then subjected to in vitro studies in DLA cell lines. Antiangeogenic activities were tested using CAM assay. EOCF was identified as the most active derivative and hence subjected to in vivo studies in tumour induced albino mice. The activity was compared with currently used anticancer drug, cyclophosphamide. The study revealed that EOCF was effective in inhibiting tumour growth.

Fermented milk supplemented with water-soluble curcumin ameliorates survival rate, selected biochemical parameters and fecal microbiota of Ehrlich ascites carcinoma-bearing mice

This study assessed the impact of fermented cow and soy milk supplemented with curcumin on enhancement the health status of female Swiss mice under Ehrlich ascites carcinoma (EAC) condition. Both types of milk were fermented by Lactobacillus plantarum EMCC 1027. Ninety-five grams of basal diet mixed with five grams of either fermented cow or soy milk supplemented with curcumin. Induction of Ehrlich tumor-bearing mice synergized by intraperitoneal injection with 1 × 106 Ehrlich ascites tumor cells after one week of the adaptation period. Our results showed that levels of survival rate were increased to 117% in mice that were fed fermented cow and soy milk while supplementation of fermented soy milk with curcumin improved the survival rate to 150%. The concentration of glutathione and superoxide dismutase in Ehrlich tumor-bearing was increased after feeding with both types of fermented milk. Fermented soy milk supplemented with curcumin showed the highest reduction levels of plasma tumor necrosis factor (TNF)-α and interleukin-6. The relative gene expression TNF-α was also significantly (P < 0.05) down-regulated in mice that were fed fermented soy milk with curcumin. The viable count of lactobacilli and bifidobacteria in feces samples was increased when mice fed both types of fermented milk. Dietary feeding with fermented cow or soy milk supplemented with curcumin has an ameliorative impact on the survival rate, antioxidant capacity, inflammation, fecal microbiota in EAC mouse model. Further research is still needed to investigate the molecular mechanisms in this tumor.

Suppression of VEGF and inflammatory cytokines, modulation of Annexin A1 and organ functions by galloylquinic acids in breast cancer model

The ongoing development of novel drugs for breast cancer aims to improve therapeutic outcomes, reduce toxicities, and mitigate resistance to chemotherapeutic agents. Doxorubicin (Dox) is known for its significant side effects caused by non-specific cytotoxicity. In this study, we investigated the antitumor activity of galloylquinic acids (BF) and the beneficial role of their combination with Dox in an Ehrlich ascites carcinoma (EAC)-bearing mouse model, as well as their cytotoxic effect on MCF-7 cells. The EAC-mice were randomized into five experimental groups: normal saline, Dox (2 mg/kg, i.p), BF (150 mg/kg, orally), Dox and BF combined mixture, and a control group. Mice were subjected to a 14-day treatment regimen. Results showed that BF compounds exerted chemopreventive effects in EAC mice group by increasing mean survival time, decreasing tumor volume, inhibiting ascites tumor cell count, modulating body weight changes, and preventing multi-organ histopathological alterations. BF suppressed the increased levels of inflammatory mediators (IL-6 and TNF-α) and the angiogenic marker VEGF in the ascitic fluid. In addition, BF and their combination with Dox exhibited significant cytotoxic activity on MCF-7 cells by inhibiting cell viability and modulating Annexin A1 level. Moreover, BF treatments could revert oxidative stress, restore liver and kidney functions, and normalize blood cell counts.

Kaempferia galanga L. extract and its main component, ethyl p-methoxycinnamate, inhibit the proliferation of Ehrlich ascites tumor cells by suppressing TFAM expression

Kaempferia galanga L. shows anti-cancer effects; however, the underling mechanism remains unclear. In this study, we explored the underlying mechanism of the anti-cancer effects of Kaempferia galanga L. Kaempferia galanga L. rhizome extracts (KGEs) suppressed Ehrlich ascites tumor cell (EATC) proliferation by inhibiting S-phase progression. The main component of KGE is ethyl p-methoxycinnamate (EMC), which exhibits the same anti-proliferative effect as KGE. Furthermore, EMC induced the downregulation of cyclin D1 and upregulation of p21. EMC also decreased the expression of mitochondrial transcription factor A (TFAM) but did not significantly change mitochondrial DNA copy number and membrane potential. Phosphorylation at Ser62 of c-Myc, a transcription factor of TFAM, was decreased by EMC treatment, which might be due to the suppression of H-ras expression. These results indicate that EMC is the active compound responsible for the anti-cancer effect of KGE and suppresses EATC proliferation by regulating the protein expression of cyclin D1 and p21; TFAM may also regulate the expression of these genes. In addition, we investigated the anticancer effects of KGE and EMC in vivo using EATC bearing mice. The volume of ascites fluid was significantly increased by intraperitoneal administration of EATC. However, the increase in the volume of ascites fluid was suppressed by oral administration of EMC and KGE. This study provides novel insights into the association between the anti-cancer effects of natural compounds and TFAM, indicating that TFAM might be a potential therapeutic target.

A Pronectin™ AXL-targeted first-in-class bispecific T cell engager (pAXLxCD3ε) for ovarian cancer

BackgroundPronectins™ are a new class of fibronectin-3-domain 14th-derived (14Fn3) antibody mimics that can be engineered as bispecific T cell engager (BTCE) to redirect immune effector cells against cancer. We describe here the in vitro and in vivo activity of a Pronectin™ AXL-targeted first-in-class bispecific T cell engager (pAXLxCD3ε) against Epithelial Ovarian Cancer (EOC).MethodspAXLxCD3ε T-cell mediated cytotoxicity was evaluated by flow cytometry and bioluminescence. pAXLxCD3ε mediated T-cell infiltration, activation and proliferation were assessed by immunofluorescence microscopy and by flow cytometry. Activity of pAXLxCD3ε was also investigated in combination with poly-ADP ribose polymerase inhibitors (PARPi). In vivo antitumor activity of pAXLxCD3ε was evaluated in immunocompromised (NSG) mice bearing intraperitoneal or subcutaneous EOC xenografts and immunologically reconstituted with human peripheral blood mononuclear cells (PBMC).ResultspAXLxCD3ε induced dose-dependent cytotoxicity by activation of T lymphocytes against EOC cells, regardless of their histologic origin. The addition of PARPi to cell cultures enhanced pAXLxCD3ε cytotoxicity. Importantly, in vivo, pAXLxCD3ε was highly effective against EOC xenografts in two different NSG mouse models, by inhibiting the growth of tumor cells in ascites and subcutaneous xenografts. This effect translated into a significantly prolonged survival of treated animals.ConclusionpAXLxCD3ε is an active therapeutics against EOC cells providing a rational for its development as a novel agent in this still incurable disease. The preclinical validation of a first-in-class agent opens the way to the development of a new 14Fn3-based scaffold platform for the generation of innovative immune therapeutics against cancer.

Visualization of hypoxia in cancer cells from effusions in animals and cancer patients.

Tumor hypoxia is frequently observed in primary solid malignancies, but the hypoxic status of tumor cells floating in body cavity effusions is largely unknown, especially in patients. This study was to observe the hypoxia and proliferation status of cancer cells floating in effusions in mice and patients. The distribution of hypoxia in cancer cells floating in ascites was first studied in nude mice. Hypoxia was detected by immunofluorescent visualization of pimonidazole and GLUT-1. For cancer patients, we retrospectively collected 21 ascites and 7 pleural effusion sample blocks of cancer patients, which were confirmed to contain tumor cells. Immunohistochemistry was performed to detect the expression of endogenous hypoxic markers HIF-1α and GLUT-1, proliferation index Ki-67. 18F-FDG PET/CT was performed to detect the glucose metabolism status of tumor cells in effusions. The tumor cells collected from ascites were positive for pimonidazole and GLUT-1, which suggesting that the cancer cells floating in ascites were hypoxic. Patterns of tumor hypoxia in human patients are similar to those observed in animal. HIF-1α and GLUT-1 were expressed by tumor cells in nearly all 28 cytological cases. For Ki-67 index, ascites tumor cells had a relatively low expression level compared with their corresponding primary or its metastatic lesions. Tumor cells in effusions showed high 18F-FDG uptake indicated the enhanced activity of glucose metabolism. Tumor cells in body cavity effusions, as a unique subgroup of tumor, are in a state of hypoxia and low proliferation, which would be one of the driven causes of chemo-radiotherapy resistance. Novel therapeutic interventions are urgently needed to overcome tumor hypoxia.

Root Bark Extract of Oroxylum indicum Vent. Inhibits Solid and Ascites Tumors and Prevents the Development of DMBA-Induced Skin Papilloma Formation

Oroxylum indicum is a traditionally used plant in Ayurvedic and folk medicines. The plant is useful for the management of gastrointestinal diseases as well as skin diseases. In the present study, we analyzed the antitumor potential of O. indicum in Dalton's lymphoma ascites tumor cells (DLA) and Ehrlich ascites carcinoma (EAC)-induced solid and ascites tumors. Further, the potential of O. indicum extract (OIM) on skin papilloma induction by dimethyl benz(a) anthracene (DMBA) and croton oil was evaluated. The chemical composition of the extract was analyzed using UPLC-Q-TOF-MS. The predominant compounds present in the extract were demethoxycentaureidin 7-O-rutinoside, isorhamnetin-3-O-rutinoside, baicalein-7-O-glucuronide, 5,6,7-trihydroxyflavone, 3-Hydroxy-3',4',5'-trimethoxyflavone, 5,7-dihydroxy-3-(4-methoxyphenyl) chromen-4-one, and 4'-Hydroxy-5,7-dimethoxyflavanone. Treatment with high-dose OIM enhanced the percentage of survival in ascites tumor-bearing mice by 34.97%. Likewise, high and low doses of OIM reduced the tumor volume in mice by 61.84% and 54.21%, respectively. Further, the skin papilloma formation was brought down by the administration of low- and high-dose groups of OIM (by 67.51% and 75.63%). Overall, the study concludes that the Oroxylum indicum root bark extract is a potentially active antitumor and anticancer agent.

Detection of circulating tumor cells and evaluation of epithelial-mesenchymal transition patterns of circulating tumor cells in ovarian cancer.

BackgroundCirculating tumor cells (CTCs) have considered to be promising liquid biopsy in cancer due to the intact information of whole cells and the potential to reflect micrometastasis. However, CTCs research are extremely limited in ovarian cancer, probably due to their rarity. The predictive value of CTCs and circulating tumor microemboli (CTM) in metastasis remains to be elucidated in ovarian cancer. This study tried to identify CTCs/CTM in ovarian cancer with considerably positive rate. To preliminarily identify the invasive capacity of CTCs/CTM, the epithelial-mesenchymal transition (EMT) patterns of CTCs/CTM was evaluated. Moreover, for comprehensive understanding of invasiveness of disseminated cells in ovarian cancer, EMT pattern of exfoliated tumor cells in ascites were also confirmed in this study.MethodsPeripheral blood samples and ascites samples were collected from 22 ovarian cancer patients. The Microfiltration combined with morphological analysis was used to detect CTC single cells or cell clusters. Microfiltration combined with morphological analysis was applied in the CTC isolation and identification. EMT was evaluated by immunofluorescence via markers including vimentin and cytokeratin.ResultsMicrofiltration combined with morphological analysis was introduced to detect CTCs/CTM with a positivity rate of 40.9% in ovarian cancer patients. The number of CTC varied from 1 to 8, with CTM number from 4 to 30. CTCs/CTM of all samples have experienced EMT process. Vimentin was expressed in all CTC samples and all tumor cells in ascites, while cytokeratin was expressed in 44.4% (4/9) of CTC samples. There were no significant differences of the clinical parameters between the CTC-positive and CTC-negative patients.ConclusionsThis study showed that both CTCs/CTM and detached tumor cells in ascites might have undergone complete or partial EMT in ovarian cancer. Moreover, microfiltration combined with cytomorphological analysis showed a considerable CTC detection rate.

Effect of a novel thiazole derivative and its complexes with polymeric carriers on the processes of lipid peroxidation in lymphoma cells

Background. Many types of tumors are sensitive to changes in prooxidant-antioxidant balance. Thus, further studies on reactive oxygen species inducing antitumor drugs that generate oxidative stress-dependent cytotoxic effects are promising. Our previous works showed that thiazole derivatives in combination with polymeric carriers have a pronounced cytotoxic effect on tumor, while not being cytotoxic against pseudo-normal cells in vitro. It was found that thiazole derivatives in complex with PEG-based polymeric carriers affected the antioxidant system of lymphoma cells in vitro. The aim of this work was to study the in vitro effect of the complex of thiazole derivative N-(5-benzyl-1,3-thiazol-2-yl)-3,5-dimethyl-1-benzofuran-2-carboxamide (BF1) in combination with polymeric carriers poly(VEP-co-GMA)-graft-mPEG (Th1), poly(PEGMA) (Th3) and poly(PEGMA-co-DMM) (Th5) on the level of lipid peroxidation products in NK/Ly cells. Materials and Methods. The experiments were conducted on white wild-type male mice with a grafted NK/Ly lymphoma. Ascites tumor cells were inoculated into mice intraperitoneally. Abdominal drainage with ascites of anesthetized mice was performed with a sterile syringe on the 7th–10th days after inoculation. Investigated compounds BF1, polymeric carriers Th1, Th3, Th5 and combination of BF1 + Th1 (Th2), BF1 + Th3 (Th4) and BF1 + Th5 (Th6) at a final concentration of 10 μM were added to the lymphoma samples and incubated for 10 minutes. The level of lipid peroxidation products, such as lipid hydroperoxides and thiobarbituric acid-positive products) were determined according to the techniques described below. Results. All applied complexes based on thiazole derivative BF1 and PEG-based polymeric carriers at a concentration of 10 μM increased the activity of lipid hydroperoxi­des in the lymphoma cells by 29–36% compared to control. Complexes Th2 and Th6 increased the significance of BF1 influence on lymphoma cells from P &lt;0.05 to P &lt;0.01. Among all of the studied complexes, Th4 and Th6 significantly increased the level of TBA-positive products, while Th2 and BF1 did not change the content of the secondary products of lipid peroxidation. None of the unconjugated polymeric carriers affected the level of lipid peroxidation products. Conclusions. Thus, based on the results of this work, thiazole derivative BF1 in complex with polymeric carriers increases the level of primary and secondary products of lipid peroxidation in lymphoma cells. Polymeric carriers enhanced the effect of thiazole derivative on the studied parameters, so complexes of thiazole derivatives and PEG-containing polymeric carriers should be taken into consideration and further investigated as potential antitumor agents.

Spheroid Formation and Peritoneal Metastasis in Ovarian Cancer: The Role of Stromal and Immune Components.

Ovarian cancer (OC) is one of the most common gynecological cancers, with the worst prognosis and the highest mortality rate. Peritoneal dissemination (or carcinomatosis) accompanied by ascites formation is the most unfavorable factor in the progression and recurrence of OC. Tumor cells in ascites are present as either separate cells or, more often, as cell aggregates, i.e., spheroids which promote implantation on the surface of nearby organs and, at later stages, metastases to distant organs. Malignant ascites comprises a unique tumor microenvironment; this fact may be of relevance in the search for new prognostic and predictive factors that would make it possible to personalize the treatment of patients with OC. However, the precise mechanisms of spheroid formation and carcinomatosis are still under investigation. Here, we summarize data on ascites composition as well as the activity of fibroblasts and macrophages, the key stromal and immune components, in OC ascites. We describe current knowledge about the role of fibroblasts and macrophages in tumor spheroid formation, and discuss the specific functions of fibroblasts, macrophages and T cells in tumor peritoneal dissemination and implantation.

DNA Repair Protein HELQ and XAB2 as Chemoresponse and Prognosis Biomarkers in Ascites Tumor Cells of High-Grade Serous Ovarian Cancer.

Nucleotide excision repair (NER) is an important mediator for responsiveness of platinum-based chemotherapy. Our study is aimed at investigating the NER-related genes expression in ascites tumor cells and its application in the prediction of chemoresponse in high-grade serous ovarian cancer (HGSC) patients. The relationship between 16 NER-related genes and the prognosis of ovarian cancer was analyzed in the TCGA database. NER-related genes including HELQ and XAB2 expressions were determined via immunocytochemistry in ascites cell samples from 92 ovarian cancer patients prior to primary cytoreduction surgery. Kaplan-Meier analysis and Cox model were used to investigate the association between NER-related gene expression and prognosis/chemotherapeutic response. Predicting models were constructed using a training cohort of 60 patients and validated in a validation cohort of 32 patients. We found that high expression of HELQ and XAB2 in the training cohort was associated with poor prognosis (for HELQ, P = 0.001, HR = 2.83, 95% CI: 1.46-5.49; for XAB2, P = 0.008, HR = 2.38, 95% CI: 1.23-4.63) and platinum resistance (for HELQ, P < 0.001; for XAB2, P = 0.006). In the validation cohort, the combination of HELQ and XAB2 (AUC = 0.863) showed the highest AUC. The expression levels of HELQ (RR 5.7, 95% CI 1.7-19.2) and XAB2 (RR 3.2, 95% CI 0.9-10.8) in ascites tumor cells were positively correlated to the risk of platinum resistance. In summary, we revealed that the expression levels of HELQ and XAB2 are candidate predictors for primary chemotherapy responsiveness and prognosis in HGSC. Ascites cytology is applicable as a promising method for chemosensitivity prediction in HGSC.

Evaluation of antitumoral effect of mistletoe fruit extract on Ehrlich ascites tumor cells with muse cell analyzer and argyrophilic nucleolar organizer region staining method

Abstract Introduction Mistletoe has been used alone or as a complementary therapy in the treatment of different diseases for years. In this study, the antitumoral effect of mistletoe fruit extract on Ehrlich ascites tumor (EAT) cells was evaluated. Materials and Methods EAT cells from preformed stock mice were transferred to culture dishes containing 5-fluorouracil (5-FU) and mistletoe extracts at different doses (100, 200, 400, and 800 μg/ml). These cells were incubated at 37 °C in an environment with 95% humidity and 5% CO2. At the end of the incubations, the apoptosis status of the cells, cell cycle, mitochondrial membrane potential, and proliferation status with the argyrophilic (Ag) nucleolar organizer region staining (NORs) method were evaluated. Results As a result, it was observed that the mistletoe fruit extract and 5-FU induce apoptosis of EAT cells. It was concluded that the 5-FU substance arrests the cell cycle at the G0/G1 stage, while the mistletoe arrests the cell cycle at the S and G2/M stages. The depolarization rate of the mistletoe treated cells was higher. As a result of the evaluation made with the AgNORs method, it was seen that mistletoe and 5-FU could be effective in reducing the proliferation of EAT cells. Conclusions It was seen that mistletoe fruit extract could be effective in stimulating the apoptosis and depolarization of cancer cells. The results of other studies in the literature and our study support each other. It was concluded that the mistletoe plant may be useful in cancer treatment.