Apoptosis In HepG2 Cell Line Research Articles

Design and In Silico and In Vitro Evaluations of a Novel Nicotinamide Derivative as a VEGFR-2 Inhibitor

A new nicotinamide derivative, (E)-N-(4-(1-(2-(4-benzamidobenzoyl)hydrazone)ethyl)phenyl)nicotinamide, was designed as a VEGFR-2 inhibitor. Utilizing the density functional theory (DFT) calculations, the three-dimensional structure of the designed compound was determined, shedding light on its stability and reactivity. Molecular docking revealed its capability to inhibit VEGFR-2, which was further supported by molecular dynamics (MD) simulations confirming its binding to the target protein. In addition, molecular mechanics-generalized born surface area (MM-GBSA), protein-ligand interactions profiler (PLIP), and essential dynamics studies provided further validation of the compound’s precise binding with optimal energy. Then, the “compound 10” was synthesized and subjected to in vitro assays. Compound 10 inhibited VEGFR-2 with an IC50 value of 105.4 ± 0.896 nM, comparing sorafenib’s IC50 value of 61.65 ± 0.934 nM. Besides, it exhibited cytotoxicity against HepG2 and MCF-7 cancer cell lines, with IC50 values of 35.78 ± 0.863 μM and 57.62 μM ± 0.871, comparing sorafenib’s IC50 values of 5.95 ± 0.917 μM and 8.45 ± 0.912 μM. Furthermore, compound 10 demonstrated a lower level of toxicity towards Vero cell lines, with an IC50 value of 127.3 μM. Likewise, compound 10 induced apoptosis in HepG2 cell lines through a flow cytometric analysis in addition to an increase in the levels of caspase-3 and caspase-9. Moreover, compound 10 hindered the migration and healing abilities of HepG2 cells. In conclusion, our study positions compound 10 as a promising candidate for further chemical modifications and biological evaluations.

Exploring the anticancer properties of a new nicotinamide analogue: Investigations into in silico analysis, antiproliferative effects, selectivity, VEGFR-2 inhibition, apoptosis induction, and migration suppression

BackgroundThis study focuses on the development and evaluation of (E)-N-(3-(1-(2-(4-bromobenzoyl)hydrazono)ethyl)phenyl)nicotinamide (BHEPN) as a potential inhibitor of Vascular Endothelial Growth Factor Receptor-2 (VEGFR-2). MethodsComputational investigations as density function theory (DFT), docking, molecular dynamics (MD) simulations, and ADMET) in addition to in vitro (VEGFR-2 inhibition, cytotoxicity against HepG2 and MCF-7 cancer cell lines, selectivity index, cells cycle analysis, apoptosis investigation, and cells migration assay) studies were conducted. ResultsDFT calculations determined the three-dimensional structure and indicated the reactivity of BHEPN. Molecular docking, and MD simulations analysis showed the BHEPN's binding affinity and its potential as a VEGFR-2 inhibitor. ADMET assessments predicted BHEPN's safety and drug-like characteristics. In vitro investigations confirmed the inhibition of VEGFR-2 with an IC50 value of 0.320 ± 0.012 µM. BHEPN also exhibited remarkable cytotoxic effects against HepG2 and MCF-7 cancer cell lines, with IC50 values of 0.19 ± 0.01 µM and 1.18 ± 0.01 µM, respectively, outperforming Sorafenib’s IC50 values (2.24 ± 0.06 µM and 3.17 ± 0.01 µM), respectively. Notably, BHEPN displayed a higher IC50 value of 4.11 ± 0 µM against the non-carcinogenic Vero cell lines, indicating selectivity index values of 21.6 and 3.4 against the tested cancer cell lines, respectively. In a flow cytometry assay, BHEPN induced HepG2 cell cycle arrest at the G1/S phase. Moreover, BHEPN increased the incidence of early and late apoptosis in HepG2 cell lines (from 1.38% and 0.22%) in control cells to (4.11–26.02%) in the treated cells, respectively. Additionally, the percentage of necrosis raised to 13.39%, in contrast to 0.62% in control cells. Finally, BHEPN was able to reduce the migration and wound healing abilities in HepG2 cells to 38.89% compared to 87.92% in untreated cells after 48 h. These in vitro results aligned with the computational predictions, providing strong evidence of BHEPN's efficacy and safety in anticancer applications. ConclusionsBHEPN is a promising candidate for the development of novel anticancer agents through further in vitro and in vivo investigations.

A Theobromine Derivative with Anticancer Properties Targeting VEGFR-2: Semisynthesis, in silico and in vitro Studies.

A computer-assisted drug design (CADD) approach was utilized to design a new acetamido-N-(para-fluorophenyl)benzamide) derivative of the naturally occurring alkaloid, theobromine, (T-1-APFPB), following the pharmacophoric features of VEGFR-2 inhibitors. The stability and reactivity of T-1-AFPB were assessed through density functional theory (DFT) calculations. Molecular docking assessments showed T-1-AFPB's potential to bind with and inhibit VEGFR-2. The precise binding of T-1-AFPB against VEGFR-2 with optimal energy was further confirmed through several molecular dynamics (MD) simulations, PLIP, MM-GBSA, and PCA studies. Then, T-1-AFPB (4-(2-(3,7-Dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-purin-1-yl)acetamido)-N-(4-fluorophenyl)benzamide) was semi-synthesized and the in vitro assays showed its potential to inhibit VEGFR-2 with an IC50 value of 69 nM (sorafenib's IC50 was 56 nM) and to inhibit the growth of HepG2 and MCF-7 cancer cell lines with IC50 values of 2.24±0.02 and 3.26±0.02 μM, respectively. Moreover, T-1-AFPB displayed very high selectivity indices against normal Vero cell lines. Furthermore, T-1-AFPB induced early (from 0.72 to 19.12) and late (from 0.13 to 6.37) apoptosis in HepG2 cell lines. In conclusion, the combined computational and experimental approaches demonstrated the efficacy and safety of T-1-APFPB providing it as a promising lead VEGFR-2 inhibitor for further development aiming at cancer therapy.

Effects of single or combined exposure to bisphenol A and mono(2-ethylhexyl)phthalate on oxidant/antioxidant status, endoplasmic reticulum stress, and apoptosis in HepG2 cell line.

Endocrine disrupting chemicals (EDCs) may affect many biological processes like growth and stress response. Bisphenol A (BPA) is a plasticizer that is used to harden plastics and polycarbonates. Phthalates are used to add flexibility to polyvinyl chloride containing plastics. The main metabolite of di(2-ethylhexyl) phthalate (DEHP) is mono(2-ethylhexyl) phthalate (MEHP) and it is even more toxic than the parent compound. Humans are usually exposed to these chemicals in mixtures by different routes starting from fetal period. However, there are not many studies in literature that investigate the combined effects of these chemicals. The aim of this study is to investigate toxic effects of BPA and/or MEHP on HepG2 cell line. We have evaluated cytotoxicity, cytomorphological, apoptotic changes, oxidative stress, oxidant/antioxidant status alterations, and endoplasmic reticulum (ER) stress. Combined exposure to BPA and MEHP caused alterations in oxidant/antioxidant status and ER stress marker proteins in both cytoplasmic and nuclear cellular fractions. We can suggest that combined exposure to EDCs may cause serious toxicological outcomes and more mechanistic studies are needed to determine the combined toxic effects.

Development of a controlled sustainable anticancer drug delivery nanosystem comprising doxorubicin and functionalized MCM-48

The present work describes the synthesis of functionalized MCM-48 nanoparticles (nMCM-48) with 12-Tungstophosphoric acid (TPA) and its characterization using various techniques like elemental analysis, FT-IR, TGA, HRTEM and N2 adsorption measurements at 77K. It also describes its application as a drug carrier for the controlled release of doxorubicin hydrochloride (DOX). The DOX loaded TPA functionalized nMCM-48 was assessed for its drug delivery potential both in-vitro and in-vivo. In-vitro release study was carried out in simulated gastric fluid (pH-1.2, 37 °C) and simulated body fluid (pH-7.4, 37 °C). The carrier was found to be non-toxic whereas the drug loaded carrier demonstrated apoptosis in HepG2 cell lines. The in-vivo studies performed on the nude mice model showed good tumor regression when treated with DOX loaded TPA/nMCM-48.

Eco-friendly sequential one-pot synthesis, molecular docking, and anticancer evaluation of arylidene-hydrazinyl-thiazole derivatives as CDK2 inhibitors

One current approach in the treatment of cancer is the inhibition of cyclin dependent kinase (CDK) enzymes with small molecules. CDK are a class of enzymes, which catalyze the transfer of the terminal phosphate of a molecule of ATP to a protein that acts as a substrate. Among CDK enzymes, CDK2 has been implicated in a variety of cancers, supporting its potential as a novel target for cancer therapy across many tumor types. Here the discovery and development of arylidene-hydrazinyl-thiazole as a potentially CDK2 inhibitors is described, including details of the design and successful synthesis of the series analogs (27a-r) using one-pot approach under eco-friendly ultrasound and microwave conditions. Most of the newly synthesized compounds showed good growth inhibition when assayed for their in-vitro anti-proliferative activity against three cancer cell lines (HepG2, MCF-7 and HCT-116) compared to the reference drug roscovitine, with little toxicity on the normal fibroblast cell lines (WI-38). Furthermore, the compounds exhibiting the highest anti-proliferative activities were tested against a panel of kinase enzymes. These derivatives displayed an outstanding CDK2 inhibitory potential with varying degree of inhibition in the range of IC50 0.35–1.49 μM when compared with the standard inhibitor roscovitine having an IC50 value 0.71 μM. The most promising CDK2 inhibitor (27f) was selected for further studies to determine its effect on the cell cycle progression and apoptosis in HepG2 cell line. The results indicated that this compound implied inhibition in the G2/M phase of the cell cycle, and it is a good apoptotic agent. Finally, Molecular docking study was performed to identify the structural elements which involved in the inhibitory activity with the prospective target, CDK2, and to rationalize the structure-activity relationship (SAR).

Antiproliferative Activity, Proapoptotic Effect, and Cell Cycle Arrest in Human Cancer Cells of Some Marine Natural Product Extract.

Bioactive constituents of numerous marine organisms have been investigated recently for their preclinical and clinical anticancer activity. Three marine organisms: black-spotted sea cucumber: Pearsonothuria graeffei (Pg), lollyfish: Holothuria atra (Ha), and sea hare: Aplysia dactylomela (Ad), were collected during winter 2019 from Gulf of Aqaba, Red Sea, Egypt, and macerated with ethanol into three different extracts: PgE, HaE, and AdE, where each was in vitro assessed for its antiproliferative and proapoptotic properties on HepG2, HCT-116, and MCF-7 cancer cells. PgE dose-dependently inhibited the growth of HepG2, HCT-116, and MCF-7 cells within IC50 values 16.22, 13.34, and 18.09 μg/mL, respectively, while the IC50 values for the antiproliferative activity of HaE were 12.48, 10.45, and 10.36 μg/mL, respectively, and the IC50 values of AdE were 6.51, 5.33, and 6.87 μg/mL, respectively. All extracts were found to induce G0/G1 cell cycle arrest for HepG2 cells side by side with their inhibition of CDK2 on all three cell lines while all extracts were also showed to induce apoptosis in HepG2 cell line at pre-G1 phase supplemented by their anticancer activity via proapoptotic protein Bax, caspase-3, and cleavage PARP increase, and antiapoptotic protein Bcl-2 downturn. Moreover, necrosis has been relatively noticed in HepG2 cell line as an additional anticancer activity for each extract. Our data introduced three ethanolic marine extracts as natural chemotherapeutic agents to be further developed for cancer control.

Abstract 136: A novel Ruthenium complex induces apoptosis in HepG2 cell line

Abstract Liver cancer is listed among the most common causes o f cancer-related death. Nowadays the clinical diagnosis of early-stage hepatocellular carcinoma (HCC) has improved significantly; nevertheless, HCC prognosis is still highly poor. Therefore, new effective and well-designed therapeutic strategies are urgently recommended. Ruthenium complexes represent a new promising family of metal-based anticancer complexes that exhibit reduced toxicity compared to platinum (II) complexes c urrently used clinically. These complexes show a novel mechanism of action with different spectrum of activity compared to platinum-based drugs. The present study was performed to investigate the effect of new formulated Rhuthenium complex {Bis (quinolin-8-olato) bis (triphenylphosphine) Ruthenium(II)} (Ru(quin)2) with or without sorafenib; the gold standard treatment for advanced HCC, as a combined treatment on different molecular signaling pathway such as ERK1/2, Stat3, AKT/m- TOR and Wnt/β-catenin that are responsible for HCC progression. In addition, the cytotoxic effect of Ru(quin)2 was also examined on phase I and phase II drug metabolizing enzyme in vivo. HepG2 cells were treated with different concentration of Ru(quin)2 to evaluate its IC50 with or without 5µM sorafenib treatment to investigate morphological changes, apoptotic induction, histone release, caspase-3 activity and clonogenic formation. Furthermore, the protein expression levels of P-ERK1/2, Bax, β-catenin, cyclin D1, Stat3, m-TO R and S6- Kinase were also examined by western immunoblotting and gene expression of Aurora kinase B was assayed using qRT-PCR. Flow cytometric analysis was performed to analyze cell cycle machinery and immunocytochemical assay was used to visualize the nuclear and cytoplasmic localization of oncogenic β-catenin protein. In vitro data revealed that Ru(quin)2 exerts potent antitumor effect on HepG2 cells compared to sorafenib. There was a significant decrease in cell viability, proliferation and clonogenic inhibition of HepG2 cells after treatment for 24 h in a dose-dependent manner. In addition, a significant increase in histone release and caspase-3 activity was observed. Moreover, Ru(quin)2 significantly upregulated proapoptotic Bax protein and significantly downregulated the expression of Stat3, m- TOR, S6-Kinase, P-ERK1/2, oncogenic β-catenin and cyclin D1 which in turn was accompanied with G0/G1 cell cycle arrest and upregulation of Aurora kinase B. Immunocytochemical analysis of β-catenin showed a reduction in the nuclear localization and expression compared to control. Moreover, combined treatment of Ru(quin)2 and sorafenib showed more profound effect on proteins expression and apoptotic induction compared to each compound alone. In vivo results showed that Ru(quin)2 treatment was associated with stimulation of phase I and phase II drug metabolizingenzymes. The activities of phase 1 drug- metabolizing enzyme changed significantly in the low and high dose treated groups (2.5 mg/kg and 5 mg/kg every other day for 2 weeks) compared to control group. Meanwhile, Ru(quin)2 treatment of Albino mice upregulated CYP3A4 and NADPH-cytochrome-c-reductase expression compared to the control. All in all, formulated Ru(quin)2 showed a promising inhibitor for HepG2 cell line proliferation, however further investigation for its complete mechanism is warranted. Citation Format: Reem EL Sayed Bakir, Salah Ahmed Sheweita, Mohamed Ali Khalifa, Ahmed Samir Sultan. A novel Ruthenium complex induces apoptosis in HepG2 cell line [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 136.

Design and Synthesis of Molecular Hybrids of Sophora Alkaloids and Cinnamic Acids as Potential Antitumor Agents.

Twenty-five sophora alkaloids-cinnamic acid hybrids (including matrine-cinnamic acid hybrids, sophoridine-cinnamic acid hybrids, and sophocarpine-cinnamic acid hybrids) were designed, synthesized, and evaluated in vitro against three human tumor cell lines (HeLa, HepG2 and A549) with cisplatin as a positive control. Some matrine-cinnamic acid and sophoridine-cinnamic acid compounds exhibited potent effect against all three cancer cell lines, such as compounds 5b, 5e, 5g, and 6d. The structure-activity relationship study of the synthesized compounds was also performed. Preliminary mechanistic studies indicated that compounds 5e and 6d could induce apoptosis in HepG2 cell line. Further, compounds 5e and 6d altered mitochondrial membrane potential and produced ROS leading to cell apoptosis of HepG2 cells. Overall, our findings suggested that these compounds may provide promising lead compounds for further development as antitumor agents by structural modification.

Synthesis of naphthalimide-carborane and metallacarborane conjugates: Anticancer activity, DNA binding ability.

The development of 1,8-naphthalimide derivatives as DNA-targeting anticancer agents is a rapidly growing area and has resulted in several derivatives entering into clinical trials. One of original recent developments is the use of boron clusters: carboranes and metallacarboranes in the design of pharmacologically active molecules. In this direction several naphthalimide-carborane and metallacarborane conjugates were synthesized in the present study. Their effect on a cancer cell line - cytotoxicity, type of cell death, cell cycle, and ROS production were investigated. The tested conjugates revealed different activities than the leading members of the naphthalimides family, namely mitonafide and pinafide. These derivatives could induce G0/G1 arrest and promote mainly apoptosis in HepG2 cell line. Our investigations demonstrated that the most promising molecule is N-{[2-(3,3'-commo-bis(1,2-dicarba-3-cobalta(III)-closo-dodecaborate-1-yl)ethyl]-1'-aminoethyl)}-1,8-naphthalimide] (17). It was shown that 17 exhibited cytotoxicity against HepG2 cells, activated cell apoptosis, and caused cell cycle arrest in HepG2 cells. Further investigations in HepG2 cells revealed that compound 17 can also induce ROS generation, particularly mitochondrial ROS (mtROS), which was also proved by increased 8-oxo-dG level in DNA. Additionally to biological assays the interaction of the new compounds with ct-DNA was studied by CD spectra and melting temperature, thus demonstrating that these compounds were rather weak classical DNA intercalators.

Effect of silver nanoparticles in the induction of apoptosis on human hepatocellular carcinoma (HepG2) cell line

Silver nanoparticles (Ag NPs) serve numerous chief functions in cosmetics, engineering, textile, food technology and medicine. These nanoparticles are also utilized in the pharmaceutical industry particularly in the production of novel antimicrobial agents. However, despite the various studies of Ag NPs induced toxicity, there is a lack of information concerning cellular toxicity mechanisms of these nanoparticles on human cells. In the current project, we investigate the anti-cancer effects of Ag NPs in HepG2 (liver hepatocellular adenocarcinoma) cells. The mean particle size and morphology for the prepared nanoparticles were determined by dynamic light scattering (DLS) and transmission electron microscopy (TEM), respectively. Cell viability, reactive oxygen species (ROS) formation, cytochrome c amount and expression level of BAX/CASP 3/CASP 8/CASP 9 were assayed in HepG2 cells after incubation with Ag NPs. The prepared nanoparticles showed the mean particle size of 30.71 nm with polydispersity index (PDI) of 0.21. Our results revealed decreased cell viability in a concentration-dependent manner and the IC50 of 75 μg/mL for Ag NPs. Ag NPs cytotoxicity was associated with induction of ROS and cell apoptosis in HepG2 cell line. According to our findings, Ag NPs could be considered as potential chemotherapeutic agents in the treatment of liver hepatocellular carcinoma.

The study on effect of the expression of interleukin-35 in HepG2 cell by lipopolysaccharide induced

Objective To investigate the effects of lipopolysaccharide (LPS) on the expression of p35 and Ebi3 subunits in HepG2 cell and the effects of LPS on proliferation and apoptosis of HepG2 cell line in vitro. Methods The effects of LPS on the cell proliferation activity of HepG2 in vitro was measured using cell counting kit-8 (CCK-8) assay and cell apoptosis was detected by Flowcytometry (FCM). FCM were used to test the expression level of p35 and Ebi3 in the LPS treated groups and control group. Enzyme-linked immunosorbent assay (ELISA) were used to test the cultured supernatants expression levelof IL-35. Results The cell proliferative activity were 0.66±0.07, 0.67±0.05, 0.68±0.09 after stimulation with LPS group (10, 100, 1000 μg/L) for 24 hours. Compared with the control group were statistically significant (P values were 0.031, 0.017, 0.007 respectively). The content of IL-35 protein in LPS (100 ng/ml) treated groups [(77.06±35.15) ng/L] had significantly higher compared with control group [(34.13±19.95) ng/L, P=0.035]. The content of p35 and Ebi3 had significantly higher when compared with control group (P values were 0.011 and 0.019 respectively). Conclusion LPS can promote proliferation of HepG2 cells and regulate the expression of interleukin-35 in HepG2 cells. Key words: Lipopolysaccharide; HepG2 cell; Proliferation

ID: 1085 Sodium citrate induces apoptosis in HepG2 cell lines

PURPOSES: Cancer cells were observed to increase glucose uptake and fermentation of glucose to lactate to to synthesis rapidly ATP for cell growth, survival and proliferation. Thus, inhibition of glycolysis might be useful in antitumor treatment. This phenomenon occurred even with fully functioning mitochondria, and known as Warberg effect. Sodium citrate, an inhibitor of Warberg effect, was reported to antiproliferate many cancer cells line. However, sodium citrate has not been studied in Hepatocellular Carcinoma cells line yet. Here we aimed to investigate the effect of sodium citrate in HepG2 cells line.
 
 MATERIAL AND METHODS: HepG2 cell lines was treated with sodium citrate at different concentrations. Viable cells were determined by Alamar Blue. The apoptosis induced-cells was detected by Annexin V with FCM technique. Disintegrated nuclei and DNA fragmentation was analyzed. The activity of caspase-3 was also tested.
 
 RESULTS: We observed that the IC50 value of sodium citrate on HepG2 is at 10mM. FCM analysis showed that sodium citrate induced apoptosis in HepG2 cell line in dose-dependent manner. At 10mM sodium citrate, the caspase-3/7 was observed to be activated in time-dependent manner. Sodium citrate also induced nuclei disintergated in HepG2. DNA fragmentation was observed when HepG2 cells were treated with 10mM sodium citrate.
 
 CONCLUSIONS: We have shown that sodium citrate possesses the antiproliferative ability on HepG2 at IC50 10mM. Sodidum citrate induces apoptosis cells in hepatocellular carcinoma HepG2 by capases-3 activation. More investigation of glycolysis inhibition of sodium citrate on HepG2 should be performed in animals

CS-PEG decorated PLGA nano-prototype for delivery of bioactive compounds: A novel approach for induction of apoptosis in HepG2 cell line

PurposePolymer-based nanoparticles are used as vectors for cancer drug delivery. The bioactive compounds (quercetin, ellagic acid and gallic acid) are well known to be not only antioxidants but also chemopreventive candidates against various types of cancers. To circumvent the low bioavailability and the short half-life time obstacles, we hypothesized a novel PLGA nano-platform functionalized with CS and PEG to encapsulate these phytochemicals. This encapsulation will protect the compounds from the phagocytic uptake and deliver PLGA-CS-PEG nano-prototype with high biodegradability and biosafety. Materials and methodsThree consequent types of PLGA-based nanocomposites were prepared and characterized. Furthermore, we investigated the newly synthesized nano-formulations against human hepatocellular carcinoma (HepG2) and colorectal cancer (HCT 116) cell lines using cell growth inhibition assays, followed by apoptosis and necrosis assays using flow cytometry to detect the underlying mechanism of HepG2 cell death. ResultsThrough Malvern Zeta Sizer, we recorded that the average diameters of the nano-prototypes ranged from 150 to 300nm. The cytotoxic activity of quercetin, ellagic acid, and gallic acid-encapsulated PLGA, PLGA-CS, and PLGA-CS-PEG nano-prototypes it has been found that they reduce the IC50s of the HepG2 cells values by 2.2, 2.9, 2.8-folds, 1, 1.5, 2.7-folds, and 0.9, 0.7, 1.5-folds, respectively. Mechanistically, the nano-platforms of quercetin seem to be dependent on both apoptosis and necrosis, while those of ellagic acid and gallic acid are mainly dependent on apoptosis. ConclusionsCS-PEG-blended PLGA nano-delivery system of quercetin, ellagic acid and gallic acid can potentiate apoptosis-mediated cell death in HepG2 cell line.

Synergistic complex from plants Solanaceae exhibits cytotoxicity for the human hepatocellular carcinoma cell line HepG2.

BackgroundIt had been demonstrated that sugars from various plants can act as potent agents, which induce apoptosis of cancer cells.MethodsUsing HPLC, we fractionated a mixture of two plant extracts from the plant family Solanaceae, namely Capsicum chinense and the plant family Amaryllidaceae namely Allium sativum. We evaluated the effect of different fractions on apoptosis of HepG2 cell line. The most effective fraction was further studied to determine its molecular composition using mass spectrometry (MS) and NMR. We further evaluated the effect of determined molecular composition found in the selected fraction by using a mixture of commercially available substances, which were found in the fraction and tested its pro-apoptotic effect on HepG2 cells. To get some insight into potential apoptotic mechanisms we studied caspase-3 activity and mitochondrial integrity in treated cells.ResultsOut of 93 fractions obtained by HPLC from the plant extract we found HPLC fraction 10 (10 min elution) was the most effective. MS and NMR studies revealed high presence of cellobiose together with vitamin C, sulphur (S) and trace amounts of selenium (Se). HPLC fraction 10 triggered apoptosis of HepG2 within 3 h in the 0.01–1.0 mg/mL concentration range. Furthermore, a mixture of pure cellobiose, vitamin C, S and Se (complex cellobiose/C/S/Se) had a very similar capacity in inducing apoptosis of HepG2 cells compared to HPLC fraction 10. Complex cellobiose/C/S/Se was capable of inducing caspase-3 activity and led to loss of mitochondrial integrity. The capacity of cellobiose alone to induce apoptosis of HepG2 was approximately 1000-fold lower compared to complex cellobiose/C/S/Se.ConclusionIn this study we present the highly synergistic effect of a unique complex consisting of cellobiose, vitamin C, sulphur and selenium on triggering the apoptosis of human hepatocellular carcinoma (HepG2) cell line.

Molecular Pathway of Psoralidin-Induced Apoptosis in HepG2 Cell Line.

To test the role of psoralidin in human liver cancer HepG2 cells in vitro. Cell viability was assessed by methylthiazolyldiphenyl-tetrazolum bromide assay and apoptotic cells were labeled by annexin V then sorted by flow cytometry. Protein expressions of caspase-3, caspase-8, caspase-9, Bax, Bid, Bcl-2, Bcl-xL and p53 were examined by western blot while activity of caspase-3, -8 and -9 were also determined. Psoralidin reduces cell viability greatly in a time dependent manner (64%, 40%, 21%, 12% at 2, 6, 24 and 48 h treatment with 64 μmol/L psoralidin respectively) and up-regulates activities of caspase-3, -8 and -9 in a concentration dependent manner (between 4 to 64 μmol/L). Psoralidin also increases the expression of pro-apoptosis genes Bax, Bid and p53 while decreases the expression of pro-survival genes Bcl-2 and Bcl-xL, both in a concentration dependent manner between 4 and 64 μmol/L (P<0.05 at 16 and 64 μmol/L). Caspase-3 inhibitor (Ac-DEVD-CHO at concentrations between 10 to 20 μmol/L), p53 inhibitor (pifithrin-α at 5 μmol/L) and cyclosporin A can attenuate the apoptotic effect of psoralidin. The cytotoxic role of psoralidin might work through both intrinsic and extrinsic apoptotic pathway.

ML-7 amplifies the quinocetone-induced cell death through akt and MAPK-mediated apoptosis on HepG2 cell line

The study aims at evaluating the combination of the quinocetone and the ML-7 in preclinical hepatocellular carcinoma models. To this end, the effect of quinocetone and ML-7 on apoptosis induction and signaling pathways was analyzed on HepG2 cell lines. Here, we report that ML-7, in a nontoxic concentration, sensitized the HepG2 cells to quinocetone-induced cytotoxicity. Also, ML-7 profoundly enhances quinocetone-induced apoptosis in HepG2 cell line. Mechanistic investigations revealed that ML-7 and quinocetone act in concert to trigger the cleavage of caspase-8 as well as Bax/Bcl-2 ratio up-regulation and subsequent cleavage of Bid, capsases-9 and -3. Importantly, ML-7 weakened the quinocetone-induced Akt pathway activation, but strengthened the phosphorylation of p-38, ERK and JNK. Further treatment of Akt activator and p-38 inhibitor almost completely abolished the ML-7/quinocetone-induced apoptosis. In contrast, the ERK and JNK inhibitor aggravated the ML-7/quinocetone-induced apoptosis, indicating that the synergism critically depended on p-38 phosphorylation and HepG2 cells provoke Akt, ERK and JNK signaling pathways to against apoptosis. In conclusion, the rational combination of quinocetone and ML-7 presents a promising approach to trigger apoptosis in hepatocellular carcinoma, which warrants further investigation.

Synthesis of novel anticancer iridoid derivatives and their cell cycle arrest and caspase dependent apoptosis

Nyctanthes arbortristis Linn (Oleaceae) is widely distributed in sub-Himalayan regions and southwards to Godavari, India commonly known as Harsingar and Night Jasmine. In continuation of our drug discovery programme on Indian medicinal plants, we isolated arbortristoside-A (1) and 7-O-trans-cinnamoyl 6β-hydroxyloganin (2) from the seeds of N. Arbortristis, which exhibited moderate in vitro anticancer activity. Chemical transformation of 2 led to significant improvement in the activity in derivative 8 and 15 against HepG2 (human hepatocellular carcinoma), MCF-7 (breast adenocarcinoma) cell lines. The compounds 8 and 15 were also capable of cell cycle arrest and caspase dependent apoptosis in HepG2 cell lines. These iridoid derivatives hold promise for developing safer alternatives to the marketed drugs.

Preparation of novel antiproliferative emodin derivatives and studies on their cell cycle arrest, caspase dependent apoptosis and DNA binding interaction

Emodin (1) is the major bioactive compound of several herb species, which belongs to anthraquinone class of compound. As a part of our drug discovery program, large quantities of emodin (1) was isolated from the roots of Rheum emodi and a library of novel emodin derivatives 2–15 were prepared to evaluate their antiproliferative activities against HepG2, MDA-MB-231 and NIH/3T3 cells lines. The derivatives 3 and 12 strongly inhibited the proliferation of HepG2 and MDA-MB-231 cancer cell line with an IC50 of 5.6, 13.03 and 10.44, 5.027, respectively, which is comparable to marketed drug epirubicin (III). The compounds 3 and 12 were also capable of inducing cell cycle arrest and caspase dependent apoptosis in HepG2 cell lines and exhibit DNA intercalating activity. These emodin derivatives hold promise for developing safer alternatives to the marketed epirubicin.

Dihydroartemisinin Induces Endoplasmic Reticulum Stress-Mediated Apoptosis in HepG2 Human Hepatoma Cells

Previous studies showed that dihydroartemisinin (DHA) possessed antitumor activity in many human tumor cells through the induction of apoptosis. The aim of this study was to investigate the effects of DHA on apoptosis in the human hepatocellular carcinoma cell line HepG2 and the possible molecular mechanisms involved. The inhibitory effect of DHA on HepG2 cells was measured by MTT assay. The percentage of apoptotic cells was detected by flow cytometry with double staining of fluorescein isothiocyanate-annexin V/propidium iodide. The intracellular production of reactive oxygen species (ROS) and intracellular Ca2+ concentration ([Ca2+]i) were detected by fluorescence spectrophotometry. Protein expression of GADD153, Bcl-2 and Bax in HepG2 cells was examined by Western blot and immunocytochemistry. DHA significantly inhibited proliferation of HepG2 cells in a dose- and time-dependent manner. The apoptosis rates in HepG2 cells treated with 0, 50, 100 and 200 μmol/L DHA for 24 hours were 2.53 ± 0.88%, 24.85 ± 3.63%, 35.27 ± 5.92% and 48.53 ± 7.76%, respectively. Compared with the control group, DHA significantly increased ROS generation and [Ca2+]i level (P <0.05), with the generation of ROS preceding the increase in [Ca2+]i. An increase in GADD153 and Bax expression and a decrease in Bcl-2 were observed in DHA-treated cells. Pretreatment with the antioxidant N-acetylcysteine could attenuate the effects of DHA in the experiments. DHA could inhibit proliferation and induce apoptosis in HepG2 cell lines through increasing the intracellular production of ROS and [Ca2+]i. Endoplasmic reticulum stress-induced apoptosis may contribute to this effect by regulating the expression of GADD153, proapoptotic Bax, and antiapoptotic Bcl-2.