HepG2 IC50 Research Articles

Citrinin Provoke DNA Damage and Cell-Cycle Arrest Related to Chk2 and FANCD2 Checkpoint Proteins in Hepatocellular and Adenocarcinoma Cell Lines.

Citrinin (CIT), a polyketide mycotoxin produced by Penicillium, Aspergillus, and Monascus species, is a contaminant that has been found in various food commodities and was also detected in house dust. Several studies showed that CIT can impair the kidney, liver, heart, immune, and reproductive systems in animals by mechanisms so far not completely elucidated. In this study, we investigated the CIT mode of action on two human tumor cell lines, HepG2 (hepatocellular carcinoma) and A549 (lung adenocarcinoma). Cytotoxic concentrations were determined using an MTT proliferation assay. The genotoxic effect of sub-IC50 concentrations was investigated using the alkaline comet assay and the impact on the cell cycle using flow cytometry. Additionally, the CIT effect on the total amount and phosphorylation of two cell-cycle-checkpoint proteins, the serine/threonine kinase Chk2 and Fanconi anemia (FA) group D2 (FANCD2), was determined by the cell-based ELISA. The data were analyzed using GraphPad Prism statistical software. The CIT IC50 for HepG2 was 107.3 µM, and for A549, it was >250 µM. The results showed that sensitivity to CIT is cell-type dependent and that CIT in sub-IC50 and near IC50 induces significant DNA damage and cell-cycle arrest in the G2/M phase, which is related to the increase in total and phosphorylated Chk2 and FANCD2 checkpoint proteins in HepG2 and A549 cells.

Biochemical and pathophysiological improvements in rats with thioacetamide induced-hepatocellular carcinoma using aspirin plus vitamin C

BackgroundHepatocellular carcinoma (HCC) is one of the leading causes of cancer-related death, so we should be concerned and look for effective/less-harmful treatments than chemotherapeutics already clinically in application. Aspirin works well ''in conjunction'' with other therapies for HCC since aspirin can boost the sensitivity of anti-cancer activity. Vitamin C also was shown to have antitumor effects. In this study, we examined the anti-HCC activities of synergistic combination (aspirin and vitamin C) vs. doxorubicin on HCC-bearing rats and hepatocellular carcinoma (HepG-2) cells.MethodsIn vitro, we evaluated IC50 and selectivity index (SI) using HepG-2 and human lung fibroblast (WI-38) cell lines. In vivo, four rat groups were used: Normal, HCC (intraperitoneally (i.p.) administered 200 mg thioacetamide/kg/twice a week), HCC + DOXO (HCC-bearing rats i.p. administered 0.72 mg doxorubicin (DOXO)/rat/once a week), and HCC + Aspirin + Vit. C (i.p. administered vitamin C (Vit. C) 4 g/kg/day after day concomitant with aspirin 60 mg/kg/orally day after day). We evaluated biochemical factors [aminotransferases (ALT and AST), albumin, and bilirubin (TBIL) spectrophotometrically, caspase 8 (CASP8), p53, Bcl2 associated X protein (BAX), caspase 3 (CASP3), alpha-fetoprotein (AFP), cancer antigen 19.9 (CA19.9), tumor necrosis factor-alpha (TNF-α), and interleukin-6 (IL-6) using ELISA], and liver histopathologically.ResultsHCC induction was accompanied by significant time-dependent elevations in all measured biochemical parameters except the p53 level significantly declined. Liver tissue architecture organization appeared disturbed with cellular infiltration, trabeculae, fibrosis, and neovascularization. Following drug medication, all biochemical levels significantly reversed toward normal, with fewer signs of carcinogenicity in liver tissues. Compared to doxorubicin, aspirin & vitamin C therapy ameliorations were more appreciated. In vitro, combination therapy (aspirin & vitamin C) exhibited potent cytotoxicity (HepG-2 IC50 of 17.41 ± 1.4 µg/mL) and more excellent safety with a SI of 3.663.ConclusionsBased on our results, aspirin plus vitamin C can be considered reliable, accessible, and efficient synergistic anti-HCC medication.

Computational insights into novel benzenesulfonamide-1,3,4-thiadiazole hybrids as a possible VEGFR-2 inhibitor: design, synthesis and anticancer evaluation with molecular dynamics studies

Thiadiazole benzenesulfonamide derivatives 8c and 8e showed the best HepG-2 IC50 of 11.80 and 4.08 μM, respectively with corresponding VEGFR-2 ΔGbinding, −9.1 and −9.8 kcal mol−1, respectively.

Extraction, Purification, and Elucidation of Six Ginkgol Homologs from Ginkgo biloba Sarcotesta and Evaluation of Their Anticancer Activities.

Ginkgols are active constituents from Ginkgo biloba L. (GB) and have pharmacological activities, such as antibacterial and antioxidant activities. In our previous report, only five ginkgols were separated. However, ginkgol C17:1 had two isomers, for which their separation, identification, and bioactivities have not yet been investigated. Hence, this research reports the successful isolation of six ginkgol homologs with alkyl substituents-C17:1-Δ12, C15:1-Δ8, C13:0, C17:2, C17:1-Δ10, and C15:0-for the first time using HPLC. This was followed by the identification of their chemical structures using Fourier transform infrared (FTIR), ultraviolet (UV), gas chromatography and mass spectrometry (GC-MS), carbon-13 nuclear magnetic resonance (13C-NMR), and proton nuclear magnetic resonance (1H-NMR) analysis. The results showed that two ginkgol isomers, C17:1-Δ12 and C17:1-Δ10, were obtained simultaneously from the ginkgol C17:1 mixture and identified entirely for the first time. That aside, the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay showed that the six ginkgol homologs possessed significant antiproliferation effects against HGC and HepG2 cells. Furthermore, the ginkgols with unsaturated side chains (C17:2, C15:1-Δ8, C17:1-Δ12, and C17:1-Δ10) exhibited more potent inhibitory effects than ginkgols with saturated side chains (C13:0, C15:0). In addition, unsaturated ginkgol C15:1-Δ8 showed the most potent cytotoxicity on HepG2 and HGC cells, of which the half-maximal inhibition concentrations (IC50) were 18.84 ± 2.58 and 13.15 ± 2.91 μM, respectively. The IC50 for HepG2 and HGC cells for the three unsaturated ginkgols (C17:1-Δ10, C17:2 and C17:1-Δ12) were ~59.97, ~60.82, and ~68.97 μM for HepG2 and ~30.97, ~33.81, and ~34.55 μM for HGC cells, respectively. Comparing the ginkgols' structure-activity relations, the findings revealed that the position and number of the double bonds of the ginkgols with 17 side chain carbons in length had no significant difference in anticancer activity.

Abstract 1040: Imipridones exhibit synergy with sorafenib, HDAC inhibition, PARP inhibition, and proteasome inhibition in liver cancer cell lines

Abstract Liver cancer is a significant cause of cancer-related death worldwide. We previously showed that the imipridone class of small molecule therapeutics is active against hepatocellular malignancies. ONC201, the first imipridone to reach clinical trials, antagonizes the G protein-coupled receptor DRD2 and is an agonist of mitochondrial caseinolytic protease ClpP. ONC206 also antagonizes DRD2 and is a ClpP agonist. ONC212 is an agonist of both the orphan G protein-coupled receptor GPR132 and ClpP. We investigated the efficacy of ONC201, ONC206, and ONC212 in combination with histone deacetylase inhibition (HDACi), poly ADP ribose polymerase inhibition (PARPi), proteasome inhibition, and current standard of care multi-kinase inhibitor sorafenib. Liver cancer cell lines Hep3B and HepG2 were treated with ONC201, ONC206, and ONC212 to determine dose-response effect. Both cell lines were also treated with each imipridone in combination with sorafenib. Combination treatment effect was also tested between ONC201 and the HDACi vorinostat, PARPi nariparib, and the proteasome inhibitor marizomib. Cell viability was measured with CellTiter-Glo. Synergy was quantified using the combination index (CI). Protein expression was assessed using Western blot. As has been previously reported, both HepG2 and Hep3B cell lines were sensitive to ONC201 (Hep3B IC50 = 2.29μM; HepG2 IC50 = 2.69μM), ONC206 (Hep3B IC50 = 0.11μM; HepG2 IC50 = 0.15μM), and ONC212 (Hep3B IC50 = 0.07μM; HepG2 IC50 = 0.05μM) administered as single-agent therapy with ONC212 demonstrating the greatest potency. In combination with sorafenib, all three imipridones demonstrated synergy in both cell lines. ONC201 and ONC206 exhibited stronger synergy with CI < 0.3 at multiple dose combinations. ONC201 was also found to exert a synergistic reduction in cell viability when Hep3B and HepG2 cells were treated with vorinostat, nariparib, and marizomib. These results suggest that the imipridone family of small molecules is a novel therapy for liver cancer with efficacy as a single agent and in combination with existing targeted agents. Future studies will clarify the mechanisms of synergy and expand the findings into small animal models. Citation Format: Joshua N. Honeyman, Wen-I Chang, Varun V. Prabhu, Joshua E. Allen, Lanlan Zhou, Wafik S. El-Deiry. Imipridones exhibit synergy with sorafenib, HDAC inhibition, PARP inhibition, and proteasome inhibition in liver cancer cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1040.

Design, synthesis and SAR of antitubercular benzylpiperazine ureas.

N-furfuryl piperazine ureas disclosed by scientists at GSK Tres Cantos were chosen as antimycobacterial hits from a phenotypic whole-cell screen. Bioisosteric replacement of the furan ring in the GSK Tres Cantos molecules with a phenyl ring led to molecule (I) with an MIC of 1μM against Mtb H37Rv, low cellular toxicity (HepG2 IC50 ~ 80μM), good DMPK properties and specificity for Mtb. With the aim of delineating the SAR associated with (I), fifty-five analogs were synthesized and screened against Mtb. The SAR suggests that the piperazine ring, benzyl urea and piperonyl moieties are essential signatures of this series. Active compounds in this series are metabolically stable, have low cellular toxicity and are valuable leads for optimization. Molecular docking suggests these molecules occupy the Q0 site of QcrB like Q203. Bioisosteric replacement of N-furfuryl piperazine-1-carboxamides yielded molecule (I) a novel lead with satisfactory PD, metabolism, and toxicity profiles.

Alkaloids from the nearly ripe fruits of Evodia rutaecarpa and their bioactivities

Two novel quinolone alkaloids (1 and 2) and two novel indole alkaloids (5 and 8), together with eleven known analogues, were isolated from the nearly ripe fruits of Evodia rutaecarpa. Their structures were determined by extensive spectroscopic data, including NMR, HRESIMS, and ECD. Additionally, the anti-tumor, hypoglycemic, and anti-bacterial activities of the isolated alkaloids were evaluated in vitro. Compound 5 as a new alkaloid displayed moderate inhibitory effect against four human cancer cell lines (MCF-7 IC50 = 30.7 μM, Hepg-2 IC50 = 65.2 μM, A549 IC50 = 39.1 μM, and SHSY-5Y IC50 = 24.7 μM), α-glucosidase (IC50 = 23.9 μM) and PTP1B (IC50 = 75.8 μM). Compound 11 showed better inhibitory effect against PTP1B (IC50 = 16.2 μM) compared with that of the positive control. Compounds 5, 13, and 14 showed moderate inhibitory effects against Bacillus cereus with MIC values of 50, 25, and 10 μM, respectively.

Synthesis and Anticancer Activity of Benzimidazole/Benzoxazole Substituted Triazolotriazines in Hepatocellular Carcinoma

Receptor Tyrosine Kinases (RTK) are the main family of cell surface receptors for growth factors, hormones and cytokines which are responsible for cell growth and differentiation and are considered as an important therapeutic target in cancer. The aim of this study was to design, synthesise and conduct the biological evaluation of benzimidazole/ benzoxazole substituted triazolotriazines as new anticancer agents. A series of benzimidazolyl and benzoxazolyl-linked triazolotriazines 8a-e and 9a-e were synthesized as receptor tyrosine kinase inhibitors. Target compounds were evaluated in HGF-induced cell proliferation assay in A549, MCF-7, HepG2 and MDA-MB-231 cancer cells. Hepatocellular carcinoma was the most sensitive cell line towards the tested compounds and 8e was the most potent one on HepG2 cells with an IC50 value of 5.13µM which was close to crizotinib (HepG2 IC50 = 4.35µM) as a standard c-Met kinase inhibitor. c-Met kinase assay of 8e showed that this compound is not capable of inhibiting this enzyme and subsequently molecular docking confirmed the low affinity of 8e towards c- Met active site and its possible anticancer mechanism through VEGFR-2 inhibition. Further in silico predictions revealed that 8e can be a drug candidate with favorable pharmacokinetic properties.

In Vitro Cytotoxicity of Secondary Metabolites Extracted from Pseudomonas aeruginosa BS25 Strain

Pseudomonas includes ubiquitous, opportunistic, pathogenic bacteria known to produce a variety of bioactive compounds. Objective of present study was to evaluate bioactive secondary metabolites produced by novel Pseudomonas aeruginosa BS25 strain. The bacterium isolated from patient’s wound was identified via 16S rRNA sequencing. To prepare extracts, nonpolar to polar solvents, i.e., hexane [H], ethyl acetate [E], acetone–ethyl acetate [AE] and water [W], were used. Cytotoxicity screening was performed against three cancer cell lines: HeLa, HepG2 and SHSY5Y. The effect on cell proliferation and mitochondrial activity was measured in HepG2 and Leishmania major. Furthermore, induction of apoptosis, necrosis, singlet oxygen release, lipid peroxidation and DNA binding were evaluated. For chemical composition colorimetric methods, TLC, HPLC and GC–MS analysis were performed. Extract [H] was effective against all cancer cell lines (relative viability: HeLa = 42.97 ± 3.46%, HepG2 = 41.54 ± 4.26%, SHSY5Y = 49.18 ± 2.98%). It strongly inhibited cell proliferation and mitochondrial activity (HepG2 IC50: 168.93 µg/ml), inducing apoptosis (apoptotic cells: 150 µg/ml = 24.89 ± 1.7%, 200 µg/ml = 43.83 ± 2.92%). Furthermore, [H] induced oxidative stress (ΦΔ = 0.5 ± 0.17) and generated TBARs (150 µg/ml = 1.81 ± 0.02 and 200 µg/ml = 2.1 ± 0.2) and DNA degradation. On the other hand, IC50 for L. major was 94.2 µg/ml. Chemical composition analysis indicated the presence of flavonoids, whereas TLC and HPLC revealed two major fractions, and GC–MS showed similarity with l-(+)-ascorbic acid 2,6-dihexadecanoate and 7,9-di-tertbutyl-1-oxaspiro (4,5) deca-6,9-diene-8-one. [H] of BS25 strain exhibits strong biological activity including ROS production, DNA damage and induction of apoptosis. Further investigations on normal cells and in vivo effects are required to fully understand its therapeutic potential.

Design, synthesis, and anticancer properties of isocorydine derivatives

Isocorydine (ICD), an aporphine alkaloid, is widely distributed in nature. Its ability to target side population (SP) cells found in human hepatocellular carcinoma (HCC) makes it and its derivative 8-amino-isocorydine (NICD) promising chemotherapeutic agents for the treatment of HCC. To improve the anticancer activity of isocorydine derivatives, twenty derivatives of NICD were designed and synthesized through chemical structure modifications of the aromatic amino group at C-8. The anti-proliferative activities of all synthesized compounds against human hepatocellular (HepG2), cervical (HeLa), and gastric (MGC-803) cancer cell lines were evaluated using an MTT assay. The results showed that all the synthetic compounds had some tumor cell growth inhibitory activity. The compound COM33 (24) was the most active with IC50 values under 10 μM (IC50 for HepG2 = 7.51 µM; IC50 for HeLa = 6.32 μM). FICD (12) and COM33 (24) were selected for further investigation of their in vitro and in vivo activities due to their relatively good antiproliferative properties. These two compounds significantly downregulated the expression of four key proteins (C-Myc, β-Catenin, CylinD1, and Ki67) in HepG2 cells. The tumor inhibition rate of COM33 (24) in vivo was 73.8% after a dose 100 mg/kg via intraperitoneal injection and the combined inhibition rate of COM33 (24) (50 mg/kg) with sorafenib (50 mg/kg) was 66.5%. The results indicated that these isocorydine derivatives could potentially be used as targeted chemotherapy agents or could be further developed in combination with conventional chemotherapy drugs to target cancer stem cells (CSCs) and epithelial-to-mesenchymal transition (EMT), the main therapeutic targets in HCC.

Phenolic composition, antioxidant and anti-proliferative activities of edible and medicinal plants from the Peruvian Amazon

Among 23 extracts of medicinal and edible plants tested, Mauritia flexuosa L.f., Arecaceae, showed significant antioxidant ability (DPPH and ORAC=1062.9 and 645.9±51.4μg TE/mg extract, respectively), while Annona montana Macfad., Annonaceae, demonstrated the most promising anti-proliferative effect (IC50 for Hep-G2 and HT-29=2.7 and 9.0μg/ml, respectively). However, combinatory antioxidant/anti-proliferative effect was only detected in Oenocarpus bataua Mart., Arecaceae (DPPH=903.8 and ORAC=1024μg TE/mg extract; IC50 for Hep-G2 and HT-29 at 102.6 and 38.8μg/ml, respectively) and Inga edulis Mart., Fabaceae (DPPH=337.0 and ORAC=795.7μg TE/mg extract; IC50 for Hep-G2 and HT-29 at 36.3 and 57.9μg/ml, respectively). Phenolic content was positively correlated with antioxidant potential, however not with anti-proliferative effect. None of these extracts possessed toxicity towards normal foetal lung cells, suggesting their possible use in development of novel plant-based agents with preventive and/or therapeutic action against oxidative stress-related diseases.

The cytotoxicity of benzaldehyde nitrogen mustard-2-pyridine carboxylic acid hydrazone being involved in topoisomerase IIα inhibition.

The antitumor property of iron chelators and aromatic nitrogen mustard derivatives has been well documented. Combination of the two pharmacophores in one molecule in drug designation is worth to be explored. We reported previously the syntheses and preliminary cytotoxicity evaluation of benzaldehyde nitrogen mustard pyridine carboxyl acid hydrazones (BNMPH) as extended study, more tumor cell lines (IC50 for HepG2: 26.1 ± 3.5 μM , HCT-116: 57.5 ± 5.3 μM, K562: 48.2 ± 4.0 μM, and PC-12: 19.4 ± 2.2 μM) were used to investigate its cytotoxicity and potential mechanism. In vitro experimental data showed that the BNMPH chelating Fe2+ caused a large number of ROS formations which led to DNA cleavage, and this was further supported by comet assay, implying that ROS might be involved in the cytotoxicity of BNMPH. The ROS induced changes of apoptosis related genes, but the TFR1 and NDRG1 metastatic genes were not obviously regulated, prompting that BNMPH might not be able to deprive Fe2+ of ribonucleotide reductase. The BNMPH induced S phase arrest was different from that of iron chelators (G1) and alkylating agents (G2). BNMPH also exhibited its inhibition of human topoisomerase IIα. Those revealed that the cytotoxic mechanism of the BNMPH could stem from both the topoisomerase II inhibition, ROS generation and DNA alkylation.

Cytotoxicity Screening for the Cyanobacterial Toxin Cylindrospermopsin

The cell lines C3A, HepG2, NCI-87, HCT-8, HuTu-80, Caco-2, and Vero were screened for sensitivity to the cyanobacterial toxin cylindrospermopsin (CYN), with the aim of determining the most sensitive cells to be used in cytotoxicity tests. Cell lines were chosen to be representative of the organs targeted by the toxin; liver, kidney, intestine, and were expected to have different metabolic activities and uptake capabilities. Over the range of cell lines tested, IC50 determinations at 24 h (MTT assay) ranged fourfold, from 1.5 μM for hepatocyte-derived cell lines (C3A IC50 = 1.5 ± 0.54; HepG2 IC50 = 1.5 ± 0.87) to 6.5 ± 3.3μM for the colon-derived Caco-2 cell line. The cell-line sensitivity seemed to decrease in cell lines derived from progressively more distal regions of the gastrointestinal tract: gastric > duodenal > ileal > colonic. The greater sensitivity of the hepatic cell lines to CYN was also apparent in 7-d exposure studies, with low toxin concentrations exerting cytotoxic effects that were not seen in other cell lines. Short-term exposure of C3A cells to CYN (1–6 h) was shown to induce cytotoxicity at 24 h despite a washout and recovery incubation, demonstrating the protracted and apparently irreversible nature of CYN's toxic effects.