Liver Cancer Cell Lines Research Articles

Abstract 577: Deciphering the mechanism of the menin-MLL complex dependency in HCC

Abstract Liver cancer is the third leading cause of cancer-related death worldwide, with hepatocellular carcinoma (HCC) being the most common primary liver cancer (~90%). The 5-year survival rate of patients with advanced HCC is ~18% due to the late prognosis and the moderate clinical benefit of available systemic therapy options. Mutations in chromatin regulators were detected in ~50% of liver cancer. Since many chromatin regulators have been shown a dependency in various cancer types and can modulate therapy response, we focused on identifying new therapeutic targets of HCC among epigenetic modifiers and evaluating them as potential targets for future drug development. We constructed a CRISPR library of 6000 gRNAs targeting 737 genes involved in chromatin-mediated gene regulation. Using the epigenome-focused CRISPR/Cas9 screening in two-dimensional and three-dimensional settings in multiple liver cancer cell lines (HepG2, HLF, PLC/PRF5), we identified a list of epigenetic regulators as potential new targets in HCC. Two of them, MEN1 and ASH2L, are core subunits of the menin-MLL complex mediating H3K4 trimethylation (me3). We validated dependencies of HCC on MEN1 and ASH2L by performing a negative selection CRISPR/Cas9 competitive growth assay in HLF and PLC/PRF5 cell lines. Moreover, treatment of HCC cell lines (HLF, PLC/PRF5, HepG2) with recently developed menin inhibitor SNDX-5613 (revumenib) revealed a dose-dependent reduction in cell proliferation. To determine transcriptional changes associated with disruption of the menin-MLL complex, we performed RNA sequencing of HLF and PLC/PRF5 cells following either inhibition with 5µM SNDX-5613 for 4 days or knockout of the MEN1 or ASH2L gene. We observed the upregulation of KRAS and the oncogenic signature of TGFβ signaling pathways in all menin-MLL complex disrupting conditions. We did not detect global changes in H3K4me3 levels upon the menin inhibition or MEN1 or ASH2L knockouts assessed by immunoblotting. To explore the mechanism of the antiproliferative effect of the menin inhibition on HCC further, we evaluated changes in the binding of menin to chromatin and H3K4me3 levels in HCC cells by performing CUT&RUN following treatment of HLF cells with 5µM SNDX-5613 for 4 days. Menin inhibition led to a drastic removal of menin from the chromatin, while H3K4me3 levels were only mildly decreased. We identified 8765 genes near promoter regions with menin binding and H3K4me3 mark, of which 1952 genes were near regions with simultaneous loss of menin and decreased H3K4me3 levels upon menin inhibition. Integration of our genomics data revealed a list of 30 genes, which are downregulated upon menin inhibition in both HLF and PLC/PRF5 cells (FC≥1.5) and the direct targets of the menin-MLL complex in HLF cells, suggesting their potential role in HCC cell survival. Altogether, we anticipate that menin and ASH2L serve as promising targets and represent an appealing therapeutic strategy for HCC treatment. Citation Format: Margarita M. Dzama, Peyton C. Kuhlers, Jesse R. Raab. Deciphering the mechanism of the menin-MLL complex dependency in HCC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 577.

Abstract 2662: Deciphering the cellular and molecular determinants of immunotherapy resistance in NASH-associated hepatocellular carcinoma by single-cell analysis

Abstract Introduction: Non-alcoholic steatohepatitis (NASH) has emerged as a prominent risk and the most rapidly increasing aetiology of hepatocellular carcinoma (HCC). Despite the effectiveness of immune checkpoint blockade (ICB), like anti-programmed cell death-ligand 1 (anti-PD-L1) antibodies, in a subset of HCC patients, NASH has been shown to impair the anti-tumor immunity in ICB-treated HCC. We aim to elucidate the mechanisms responsible for ICB resistance in NASH-HCC at single-cell resolution. Methods: Orthotopic NASH-HCC mouse models were generated by orthotopically injecting liver cancer cell line into NASH liver induced by methionine-and choline-deficient diet (MCD) or choline-deficient, L-amino acid-defined, high-fat diet (CDAHFD). Multi-color flow cytometry and single-cell RNA-sequencing (scRNA-seq) were utilized to dissect the liver and tumor immune microenvironment upon anti-PD-L1 treatment. Results: Anti-PD-L1 treatment failed to alleviate tumor burden in NASH mice, indicating that orthotopic NASH-HCC mouse models were resistant to ICB therapy. Compared to the control liver, multi-color flow cytometry demonstrated that CD11b+F4/80+CD206+M2 macrophages accumulated in NASH liver and further increased in liver and tumor upon anti-PD-L1 treatment. Tumor-infiltrating M2 macrophages exhibited the most significant and robust correlation with tumor weight among all immune cell types. Moreover, PD-1 expression on tumor-infiltrating CD8+ T cells positively correlated with tumor weight and intratumoral M2 macrophages. Furthermore, scRNA-seq analysis identified a NASH-specific tumor-infiltrating monocyte subcluster, which may differentiate into an ‘M2-like’ macrophage subtype upon anti-PD-L1 treatment as demonstrated by cell trajectory analysis. Conclusion: Our study provides insights into the significance of monocyte/macrophage reprogramming in ICB resistance in NASH-HCC. Further investigation is conducted to identify the molecular mechanisms during monocyte-macrophage differentiation and explore potential targeted strategies to overcome ICB resistance in NASH-HCC. Acknowledgment: This study is supported by the General Research Fund (14120621 and 14119023), the Collaborative Research Fund (C4045-18W), the Li Ka Shing Foundation and CUHK Strategic Seed Funding for Collaborative Research Scheme. Keywords: hepatocellular carcinoma, single-cell RNA-sequencing, anti-PD-L1, macrophages. Citation Format: Lingyun Zhang, Yiling Zhang, Wenshu Tang, Zhewen Xiong, Xiaoyu Liu, Zhixian Liang, Yuk Wah Tsang, Weiqin Yang, Xiaohang Long, Alfred Sze Lok Cheng. Deciphering the cellular and molecular determinants of immunotherapy resistance in NASH-associated hepatocellular carcinoma by single-cell analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2662.

Development of a Novel Peptide with RGD Tumor Homing Motif: Evaluation of its Anticancer Potential in Hepatocellular Carcinoma and Colon Cancer Cells

Background: Peptide-based therapy has emerged as a promising avenue for treating various disorders, and recent research has highlighted the potential of anti-cancer peptides (ACPs) in cancer treatment. In this context, this study aimed to design a novel peptide incorporating a tumor-homing peptide (RGD) and C-amidation to enhance its anticancer activity, particularly against liver (HepG2) and colon (HCT-116) cancer cell lines. Objectives: The primary objective was to design a peptide with improved anticancer properties by leveraging the tumor-homing capabilities of RGD and enhancing its activity through C-amidation. The study sought to evaluate the cytotoxicity of the designed peptide against red blood cells (RBCs) and normal Vero cells. Furthermore, the anticancer efficacy of the peptide was assessed in hepatocellular carcinoma (HepG2) and colon cancer (HCT-116) cell lines. The specific objectives included examining the apoptotic induction and morphological changes in treated cells compared to untreated cells. Methods: The peptide was designed using the ACPred-FL bioinformatics tool, and its cytotoxicity was assessed through hemolysis assays against RBCs and normal Vero cells. Anticancer activity was evaluated against HepG2 and HCT-116 cell lines. The analysis of apoptotic induction involved measuring the relative gene expression of oncogenic marker BCL2 and apoptotic markers (BAX, BID, CAS-8). Additionally, Cytopathological examination and Western Blot analysis were employed to study morphological changes and confirm the quantification of relevant markers. Results: The designed peptide, consisting of twelve amino acids with a molecular mass of 1230.6233 Da and an isoelectric point of 9.81, exhibited low erythrocyte lysis and minimal toxicity to normal cells. The IC50 values demonstrated significant anticancer activity against both HepG2 (36.49±2.6 μg/mL) and HCT-116 (11.03±2.5 μg/mL) cell lines. Treated cells exhibited a significant decrease in the oncogenic marker BCL2 and an upregulation of apoptotic markers (BAX, BID, CAS-8). Western Blot analysis confirmed these findings, and Cytopathological examination revealed scattered apoptotic and degenerative changes. Conclusion: The designed peptide displayed remarkable anticancer activity against hepatocellular carcinoma and colon cancer cell lines, effectively modulating apoptotic and oncogenic markers. These findings highlight the potential of the peptide as a therapeutic agent for cancer treatment, emphasizing its clinical significance in combating liver and colon cancers. Nonetheless, further research and development are warranted to explore the translational potential of this peptide in clinical settings.

Benzohydrazide derivative metal complex's antimicrobial and inhibitory effects on liver cancer cell lines and quinone oxidoreductase 2: Experimental, molecular docking, and DFT investigations

The activity of benzohydrazide derivatives has good biological activity against many bacteria species and different cancer cell lines, and the significance of metal complexes in boosting bioavailability and inhibitory action have both been highlighted in recent research. Here we synthesized (E)-N'-(4-(dimethylamino)benzylidene)benzohydrazide (HDmby) and its seven metal (II) complexes, including [MCl2(HDmby)] {where MII = Pd (1), Pt(2), Zn (3), Cd(4), and Hg(5)} and [M(Dmby)2] {where MII = Pd (6), and Hg(7)}, with good yields (71–93 %) and characterized by molar conductivities, elemental analysis, FTIR, and NMR (1H and 13C) spectroscopic techniques. This study focuses on the analysis of the HDmby ligand and its metal complexes using theoretical calculations. The results reveal excellent agreement between the calculated and experimental data, highlighting the stability of both the ligand and its metal complexes. The HOMO and LUMO energies indicate their stability, with negative energy values. The prepared complexes displayed respectable activity against S. aureus, B. subtilis, and E. coli. [PtCl2(HDmby)] complex showed the highest inhibition value (DIZ is 20, 23, and 25 mm) against all types of bacteria. In other combinations, the free ligand doesn't show outstanding results. Also screened the cytotoxic activity against the human liver cancer cells, and the results indicated that complexes (2) and (3) showed higher cytotoxic activity with IC50 = 8.08 ± 0.736 and 4.03 ± 0.271 µM, respectively. As the high activity of quinone oxidoreductase 2 (NQO2) was correlated to liver cancer incidence, the inhibitory activity of HDmby and its complexes on NQO2 was investigated using a molecular docking approach. HDmby has demonstrated a remarkable capacity to form hydrogen bonds, whereas its zinc complex has demonstrated an enhanced energy binding score that is on par with doxorubicin, which is being studied as a reference inhibitor. The outcomes demonstrated a strong degree of concordance with the experimental data and emphasized the significance of organometallic compounds in cancer chemotherapy.

Bee chitosan nanoparticles loaded with apitoxin as a novel approach to eradication of common human bacterial, fungal pathogens and treating cancer.

Antimicrobial resistance is one of the largest medical challenges because of the rising frequency of opportunistic human microbial infections across the globe. This study aimed to extract chitosan from the exoskeletons of dead bees and load it with bee venom (commercially available as Apitoxin [Api]). Then, the ionotropic gelation method would be used to form nanoparticles that could be a novel drug-delivery system that might eradicate eight common human pathogens (i.e., two fungal and six bacteria strains). It might also be used to treat the human colon cancer cell line (Caco2 ATCC ATP-37) and human liver cancer cell line (HepG2ATCC HB-8065) cancer cell lines. The x-ray diffraction (XRD), Fourier transform infrared (FTIR), and dynamic light scattering (DLS) properties, ζ-potentials, and surface appearances of the nanoparticles were evaluated by transmission electron microscopy (TEM). FTIR and XRD validated that the Api was successfully encapsulated in the chitosan nanoparticles (ChB NPs). According to the TEM, the ChB NPs and the ChB NPs loaded with Apitoxin (Api@ChB NPs) had a spherical shape and uniform size distribution, with non-aggregation, for an average size of approximately 182 and 274 ± 3.8 nm, respectively, and their Zeta potential values were 37.8 ± 1.2 mV and - 10.9 mV, respectively. The Api@ChB NPs had the greatest inhibitory effect against all tested strains compared with the ChB NPs and Api alone. The minimum inhibitory concentrations (MICs) of the Api, ChB NPs, and Api@ChB NPs were evaluated against the offer mentioned colony forming units (CFU/mL), and their lowest MIC values were 30, 25, and 12.5 μg mL-1, respectively, against Enterococcus faecalis. Identifiable morphological features of apoptosis were observed by 3 T3 Phototox software after Api@ChB NPs had been used to treat the normal Vero ATCC CCL-81, Caco2 ATCC ATP-37, and HepG2 ATCC HB-8065 cancer cell lines for 24 h. The morphological changes were clear in a concentration-dependent manner, and the ability of the cells was 250 to 500 μg mL-1. These results revealed that Api@ChB NPs may be a promising natural nanotreatment for common human pathogens.

Two new cytotoxic diterpenoid alkaloids from Delphinium grandiflorum

Delphinium grandiflorum, one of the well-known Delphinium species, contains diverse bioactive diterpenoid alkaloids (DAs), which promoted a systematic phytochemical study on this plant. As a result, two new DAs, namely 16-demethyl tuguaconitine (1) and 6-O-acetyl-16-demethyldelsoline (2), together with one known analogue pseudophnine A (3) were obtained from the whole herbs of D. grandiflorum. Their structures were elucidated by use of extensive spectroscopic analysis (HR-ESIMS, IR, 1D and 2D NMR). Additionally, the inhibitory effects of these obtained individual compounds were also evaluated in vitro for lung cancer cell line (A549), liver cancer cell line (SMMC-7721), and breast cancer cell line (MCF-7). The results of cytotoxic activities showed that only compound 2 exhibit weak inhibitory effect on SMMC-7721 and MCF-7, and their IC50 values were 37.4 and 33.1 μM, respectively. However, compound 1 and 3 had no obvious cytotoxic activities (IC50>80 μM).

Synthesis, spectroscopic and biological characterizations of platinum(IV), ruthenium(III) and iridium(III) theophylline complexes

This study used micro-analyses, (FTIR, UV-Vis) spectra, magnetic, thermogravimetric, X-ray powder diffraction (XRD) patterns, and transmittance electron microscopy (TEM) techniques to characterize three synthesized theophylline (TPH) complexes with ruthenium(III), platinum(IV), and iridium(III) metal ions. The metal ions indicated above were found to align with the TPH drug chelate as a mono-dentate ligand via the deprotonated NH group at the nitrogen atom position N7, as verified by FTIR measurements. Additionally, the complexes conductivity and magnetic susceptibility were examined. The octahedral geometry for the synthesized complexes was proposed by the current data. Except for the iridium(III) complex, which has a non-electrolytic nature due to the presence of a chlorine atom inside the chelation sphere, the molar conductivity of the complexes in DMSO solution was electrolyte in nature. Theophylline complexes with a (metal: ligand) stoichiometry of 1:2 were produced. The TPH complexes have also been tested in vitro against G(+) bacteria (Bacillus subtilis and Staphylococcus Aureus) and G(-) bacteria (Pseudomonas aeruginosa and Escherichia coli) to evaluate their antibacterial efficacy. Human breast and liver cancer cell lines were used as a test for the TPH complexes in vitro anti-cancer properties.
 KEY WORDS: Theophylline, Metal ions, Chelation, Octahedral, Spectral analysis, Nano-particles, Biological evaluation
 Bull. Chem. Soc. Ethiop. 2024, 38(3), 725-738. 
 DOI: https://dx.doi.org/10.4314/bcse.v38i3.14

Enoxacin fluoroquinolone antibacterial agent complexes with Pt(IV), Ru(III) and Ir(III) ions: Synthesis, spectroscopic, physicochemical characterizations and biological studies

The synthesis, physicochemical characterizations, and biological investigations on the chelation of the antibiotic medication enoxacin (EXN) with certain vital metal ions, such as platinum(IV), ruthenium(III), and iridium(III) are discussed in this study. These compounds structures were interpreted based on elemental analysis and spectral measurements (FTIR, 1H-NMR, UV-Vis electronic, and XRD). The findings show that there is same coordination pathway for the chelation behavior of enoxacin complexes: a monodentate manner. The Pt(IV), Ru(III), and Ir(III) complexes are coordinated through the N atom of the piperazine ring, which is an unusual mode of coordination for this class of compounds. The dissolved complexes in DMSO were found to have non-electrolyte nature based on their molar conductance measurements. The formula for these complexes is [Pt(EXN)3(H2O)2Cl] (1), [Ru(EXN)3(H2O)3] (2), and [Ir(EXN)3(H2O)3] (3). Using scanning electron microscope (SEM) analysis and X-ray powder diffraction (XRD), the nano-scale range of the Pt(IV), Ru(III), and Ir(III) complexes has been determined. Results of in vitro cytotoxicity were examined using MCF-7 cells (human breast cancer cell line) and HepG-2 cells (human hepatocellular carcinoma) that had been grown. The cytotoxic activity of the metal complexes demonstrated a cytotoxic effect against the growth of human breast and liver cancer cell lines.
 KEY WORDS: Enoxacin, Platinum(IV), Complexity, Spectral analyses, Biological assessments
 Bull. Chem. Soc. Ethiop. 2024, 38(3), 685-699. 
 DOI: https://dx.doi.org/10.4314/bcse.v38i3.11

Application of thiourea polyurethane foam/zinc oxide nanocomposite for anticancer effects and antimicrobial potential

There is a continuous need to fabricate new antimicrobial agents due to the continuous development of microbial drug resistance. Recently, zinc oxide nanoparticles (ZnONPs) have emerged as a promising choice for developing new drugs owing to their exceptional broad-spectrum antimicrobial and anticancer action. In the current work, thiourea polyurethane foam/zinc oxide nanocomposite was synthesized by coupling ZnONPs and thiourea polyurethane foam (TPUF) to test its antimicrobial and anticancer activities. Ultraviolet–visible spectra, Fourier transform infrared, X-ray diffraction (XRD), zeta potential and dynamic light scattering analyses were investigated to study the characterization of thiourea polyurethane foam/zinc oxide nanocomposite (TPUF/ZnO). The synthesized TPUF/ZnO was examined as an antimicrobial agent against gram-positive bacterium (Bacillus cereus), gram-negative bacterium (Escherichia coli), and fungus (Aspergillus niger). The results of the agar well-diffusion test showed good inhibition zones against E. coli (16 mm), B. cereus (13 mm), and A. niger (26 mm). The minimum inhibition concentration (MIC) values of TPUF/ZnO against B. cereus and E. coli were 350 μg/mL and 400 μg/mL. TEM of TPUF/ZnO-treated B. cereus and E. coli bacteria displayed different ultrastructural modifications such as separation of the plasma membrane from the cell wall and vacuole formation that led to a complete lysis and bacterial death. The anticancer activity of TPUF/ZnO was also evaluated against liver and breast cancer cell lines (HepG-2 and MCF-7). TPUF/ZnO showed good cytotoxicity activities against HepG-2, and MCF-7, with IC50 values of 77.08 ± 1.36 and 109.01 ± 3.98 µg/ml, respectively. Thus, TPUF/ZnO could be a promising raw material in the field of food packaging and provide a possible anticancer compound for hepatocellular carcinoma and breast carcinoma.Graphic abstract

Long non-coding RNA LINC01554 overexpression suppresses viability, migration, and invasion of liver cancer cells through regulating miR-148b-3p/EIF4E3

BackgroundLong non-coding RNAs (lncRNAs) can be severed as competing endogenous RNAs (ceRNAs) to regulate target genes or mRNAs via sponging microRNAs (miRNAs). This study explored the effect of LINC01554 on liver cancer cells through the ceRNA mechanism. MethodsFive significantly down-regulated lncRNAs were selected for further verification, and then through bioinformatics, interactive miRNAs and mRNAs of lncRNAs were identified. The relationship between LINC01554, miR-148b-3p and EIF4E3 was detected by the dual luciferase reporter gene assay. Afterwards, HCCLM3 cells were transfected with pCDH-LINC01554, miR-148b-3p inhibitor and miR-148b-3p mimics. Cell viability, apoptosis, migration and invasion were measured by Cell Counting Kit-8, flow cytometer, and Transwell assays. Real-time quantitative PCR (RT-qPCR) and Western blot were used to measure the expressions of related genes and proteins. ResultsLINC01554 was significantly down-regulated in the liver cancer cell lines, and was expressed in the cytoplasm of HCCLM3 cells. LINC01554 overexpression inhibited proliferation, migration, and invasion of HCCLM3 cells, and promote their apoptosis (P < 0.05). Besides, LINC01554 overexpression also significantly increased the levels of BAX, BCL2/BAX, P53, cleaved-Caspase3, TIMP3, E-cadherin and EIF4E3 (P < 0.05). Through bioinformatics and dual-luciferase reporter gene assay, LINC01554, miR-148b-3p and EIF4E3 were proved to interact with each other. Furthermore, the effects of miR-148b-3p knockdown on HCCLM3 cells were similar with those of LINC01554 overexpression, and miR-148b-3p mimics could reverse the changes of cell viability, apoptosis, migration, and invasion induced by LINC01554 overexpression. ConclusionsLINC01554 overexpression could suppress the growth and metastasis of HCCLM3 cells via miR-148b-3p/EIF4E3.

Drug-Loaded Tumor-Derived Microparticles Elicit CD8+ T Cell-Mediated Anti-Tumor Response in Hepatocellular Carcinoma.

Hepatocellular Carcinoma (HCC) poses significant challenges due to limited effective treatments and high recurrence rates. Immunotherapy, a promising approach, faces obstacles in HCC patients due to T-cell exhaustion and immunosuppression within the tumor microenvironment. Using doxorubicin-loaded tumor-derived microparticles (Dox-TMPs), the mice with H22 ascites model and subcutaneous tumors model were treated. Following the treatment, mice were re-challenged with H22 cells to compare the therapeutic effects and recurrence among different groups of mice, alongside examining the changes in the proportions of immune cells within the tumor microenvironment. Furthermore, Dox-TMPs were combined with anti-PD-1 to further validate their anti-tumor efficacy. In vitro studies using various liver cancer cell lines were conducted to verify the tumor-killing effects of Dox-TMPs. Additionally, CD8+ T cells from the abdominal cavity of tumor-free mice were co-cultured with H22 cells to confirm their specific tumor-killing abilities. Dox-TMPs demonstrate effective anti-tumor effects both in vitro and in vivo. In vivo, their effectiveness primarily involves enhancing CD8+ T cell infiltration, alleviating T cell immunosuppression, and improving the immune microenvironment to combat tumors. When used in combination with anti-PD-1, their anti-tumor effects are further enhanced. Moreover, some mice treated with Dox-TMPs developed anti-tumor immunity, displaying a self-specific T-cell immune response upon re-challenged with tumor cells. This suggests that Dox-TMPs also have the potential to act as a long-term immune response against tumor recurrence, indicating their capability as a tumor vaccine. Dox-TMPs exhibit a dual role in liver cancer by regulating T cells within the tumor microenvironment, functioning both as an anti-tumor agent and a potential tumor vaccine.

Investigating the effect of polymerase inhibitors on cellular proliferation: Computational studies, cytotoxicity, CDK1 inhibitory potential, and LC-MS/MS cancer cell entrapment assays.

Directly acting antivirals (DAAs) are a breakthrough in the treatment of HCV. There are controversial reports on their tendency to induce hepatocellular carcinoma (HCC) in HCV patients. Numerous reports have concluded that the HCC is attributed to patient-related factors while others are inclined to attribute this as a DAA side-effect. This study aims to investigate the effect of polymerase inhibitor DAAs, especially daclatasivir (DLT) on cellular proliferation as compared to ribavirin (RBV). The interaction of DAAs with variable cell-cycle proteins was studied in silico. The binding affinities to multiple cellular targets were investigated and the molecular dynamics were assessed. The invitro effect of the selected candidate DLT on cancer cell proliferation was determined and the CDK1 inhibitory potential in was evaluated. Finally, the cellular entrapment of the selected candidates was assessed by an in-house developed and validated LC-MS/MS method. The results indicated that polymerase inhibitor antiviral agents, especially DLT, may exert an anti-proliferative potential against variable cancer cell lines. The results showed that the effect may be achieved via potential interaction with the multiple cellular targets, including the CDK1, resulting in halting of the cellular proliferation. DLT exhibited a remarkable cell permeability in the liver cancer cell line which permits adequate interaction with the cellular targets. In conclusion, the results reveal that the polymerase inhibitor (DLT) may have an anti-proliferative potential against liver cancer cells. These results may pose DLT as a therapeutic choice for patients suffering from HCV and are liable to HCC development.

Abstract 2308 In-vitro Combo Treatment of Human Liver Cancer Cell (Hep-G2) Line with a Cocktail that had Inhibited Human Melanoma Cell Growth

Abstract 2308 In-vitro Combo Treatment of Human Liver Cancer Cell (Hep-G2) Line with a Cocktail that had Inhibited Human Melanoma Cell Growth

LASS2 suppresses metastasis in multiple cancers by regulating the ferroptosis signalling pathway through interaction with TFRC

BackgroundAs a key enzyme in ceramide synthesis, longevity assurance homologue 2 (LASS2) has been indicated to act as a tumour suppressor in a variety of cancers. Ferroptosis is involved in a variety of tumour processes; however, the role of LASS2 in regulating ferroptosis has yet to be explored. This article explores the potential underlying mechanisms involved.MethodsBioinformatics tools and immunohistochemical staining were used to evaluate LASS2 expression, and the results were analysed in relation to overall survival and clinical association in multiple cancers. Coimmunoprecipitation-coupled liquid chromatography-mass spectrometry (co-IP LC-MS) was performed to identify potential LASS2-interacting proteins in thyroid, breast, and liver cancer cell lines. Transcriptomics, proteomics and metabolomics analyses of multiple cancer cell types were performed using MS or LC–MS to further explore the underlying mechanisms involved. Among these tumour cells, the common LASS2 interaction partner transferrin receptor (TFRC) was analysed by protein–protein docking and validated by coimmunoprecipitation western blot, immunofluorescence, and proximity ligation assays. Then, we performed experiments in which tumour cells were treated with Fer-1 or erastin or left untreated, with or without inducing LASS2 overexpression, and assessed the molecular biological and cellular functions by corresponding analyses.ResultsLow LASS2 expression is correlated with adverse clinical characteristic and poor prognosis in patients with thyroid cancer, breast cancer or HCC. Multiomics analyses revealed significant changes in the ferroptosis signalling pathway, iron ion transport and iron homeostasis. Our in vitro experiments revealed that LASS2 overexpression regulated ferroptosis status in these tumour cells by affecting iron homeostasis, which in turn inhibited tumour migration, invasion and EMT. In addition, LASS2 overexpression reversed the changes in tumour cell metastasis induced by either Fer-1 or erastin. Mechanistically, LASS2 interacts directly with TFRC to regulate iron homeostasis in these tumour cells.ConclusionsIn summary, our study reveals for the first time that LASS2 can inhibit tumour cell metastasis by interacting with TFRC to regulate iron metabolism and influence ferroptosis status in thyroid, breast, and liver cancer cells, these results suggest potential universal therapeutic targets for the treatment of these cancers.

MicroRNA-302a enhances 5-fluorouracil sensitivity in HepG2 cells by increasing AKT/ULK1-dependent autophagy-mediated apoptosis

Abstract Objectives MicroRNA-302a (miR-302a) has been implicated in the oncogenic processes, but its role in hepatocellular carcinoma (HCC) chemoresistance and related mechanisms are still unclear. Therefore, the aim of this study was to investigate the role of miR-302a in HCC chemoresistance and elucidate its underlying mechanism. Methods In this study, we detected the level of miR-302a in HCC tissues (including chemoresistant and chemosensitive tissues), non-tumor tissues, liver cancer cell lines, and 5-fluorouracil (5-FU)-resistant cells (HepG2/R). Additionally, we conducted cell viability, apoptosis, and autophagy analyses as well as assessed the levels of cleaved caspase-3, cleaved caspase-9, microtubule-associated protein 1 light chain 3 beta II (LC3B-II), Akt, and UNC-51 like kinase 1 (ULK1) in HepG2 cells transfected with miR-302a mimic or inhibitor prior to 5-FU treatment. Lastly, we predicted the target of miR-302a and verified the relationship between miR-302a and Akt by luciferase reporter and functional repair assays. Results Our results revealed that miR-302a was down-regulated in HCC tissues (p&lt;0.01), especially in chemoresistant tissues (p&lt;0.01). Consistently, the miR-302a level exhibited a lower expression in HepG2/R cells compared to their parental cells (p&lt;0.01). Furthermore, the 5-FU-induced apoptosis and autophagy of HepG2 cells were promoted by miR-302a over-expression and diminished by miR-302a inhibition (p&lt;0.01). Target analysis revealed that miR-302a could directly target Akt. Moreover, miR-302a inhibited Akt expression and subsequently elevated ULK1 expression (p&lt;0.01). Inhibition of ULK1 could abrogate the sensitization of overexpressed miR-302a to 5-FU in HepG2 cells (p&lt;0.01). Conclusions Altogether, our results demonstrate that the down-regulation of miR-302a promotes 5-FU resistance in HCC by attenuating the Akt/ULK1 axis-dependent autophagy and apoptosis.

Enzymatic synthesis of biologically active fructose-based saccharides by Aspergillus niger MK788296 levansucrase

Fructose-based saccharides (FBS) and levans received much attention in recent years because of their diverse biological activities and safety which help in their usability in several medical and pharmaceutical applications. In the present study, FBS and levan were synthesized by Aspergillus niger MK788296 levansucrase. Fifteen fructose-based saccharides were synthesized through the interaction between levansucrase concentration (units), duration of synthesis and substrate (sucrose) concentration through a central composite design (CCD). The synthesized FBS and levans exhibited several biological activities including prebiotic, anti-microbial, fibrinolytic, antioxidant and antitumor activities. The prebiotic effect was emphasized by its promotion effect on the growth of Bifidobacterium animalis subsp. lactis BB-12 and Lactobacillus plantarum DMS 20174. They also exerted antimicrobial activity against Escherichia coli ATCC 8739 and Staphylococcus aureus ATCC 6538. In addition, they exerted prominent antitumor activity against colorectal, liver and breast cancer cell lines.

Iron oxide nanoparticles coated with Glucose and conjugated with Safranal (Fe3O4@Glu-Safranal NPs) inducing apoptosis in liver cancer cell line (HepG2)

Magnetic nanoparticles can be considered a reliable tool for targeted drug delivery to cancer tissues. Based on this, in this study, the anticancer effect of iron oxide nanoparticles coated with glucose and conjugated with Safranal (Fe3O4@Glu-Safranal NPs) on a liver cancer cell line (HepG2) was investigated. Physicochemical properties of nanoparticles were characterized using FT-IR, XRD, VSM, EDS-mapping, SEM and TEM imaging, zeta potential, and DLS analyses. MTT test was used to investigate the inhibitory effect of nanoparticles on cancer and normal cell lines. Also, the reactive oxygen species (ROS) level, the population of apoptotic cells, and cell cycle analysis were evaluated in control and nanoparticle-treated cells. The synthesized particles were spherical, in a size range of 17–49 nm, without impurities, with a surface charge of − 13 mV and hydrodynamic size of 129 nm, and with magnetic saturation of 22.5 emu/g. The 50% inhibitory concentration (IC50) of Safranal, Fe3O4, Fe3O4@Glu-Safranal and Cisplatin drug on liver cancer cells were 474, 1546, 305 and 135 µg/mL, respectively. While, the IC50 of Fe3O4@Glu-Safranal for normal cell line was 680 µg/mL. Treating liver cancer cells with nanoparticles significantly increased the population of apoptotic cells from 2.5% to 34.7%. Furthermore, the population of the cells arrested at the G2/M phase increased in nanoparticle-treated cells. Due to the biocompatibility of the constituent compounds of these nanoparticles, their magnetic properties, and their inhibitory effects on cancer cells, Fe3O4@Glu-Safranal NPs can be further considered as a promising anticancer compound.

Cobalt oxide nanoparticles functionalized with glucose and conjugated to Ellagic acid (Co3O4@Glu-Ellagic acid NPs) induce apoptotic genes in HepG2, liver cancer cell line

Surface modification of metal nanoparticles can be a solution to obtain more efficient and biocompatible drugs. In this study, the anticancer mechanism of cobalt oxide nanoparticles (NPs) functionalized with glucose and conjugated with Ellagic acid (Co3O4@Glu-Ellagic acid NPs) in a liver cancer cell line was investigated. Physicochemical characteristics of NPs were investigated by Fourier-transform infrared spectroscopy (FT-IR), Transmission electron microscopy (TEM), (Scanning Electron Microscopy) SEM, X-Ray diffraction analysis (XRD), Energy Dispersive Spectroscopy mapping (EDS-mapping), Dynamic light scattering (DLS), and zeta potential. Cytotoxicity of Co3O4@Glu-Ellagic acid NPs in normal (THLE2) and cancer (HepG2) hepatocyte cell lines was characterized by MTT and flow cytometry assays. Relative expression of the CASP8, CASP9, and p53 genes was assessed by Real-Time PCR. The synthesized nanoparticles were spherical, without considerable agglomeration, and their size was in the range of 20 to 60 nm. Surface charge and DLS size were − 30.1 mV and 280 nm, respectively. Treatment of the studied cells with the nanoparticles displayed their antiproliferative potential; however, liver cancer cells were considerably more sensitive to the nanoparticles than normal hepatocyte cells. The 50 % inhibitory concentration of Co3O4@Glu-Ellagic acid NPs for normal and cancer hepatocyte cells were 201 and 97 μg/mL, respectively. Treating HepG2 cells with Co3O4@Glu-Ellagic acid NPs led to a significant increase in the percentage of apoptotic cells (from 0.66 % to 50.8 %) and expression of the CASP8, CASP9, and p53 genes by 2.88, 2.11, and 2.0 folds that suggests apoptogenic property of the nanoparticles through activation of the intrinsic and extrinsic apoptosis pathways. Measuring Reactive Oxygen Species (ROS) level showed that in treated cancer cells with Co3O4@Glu-Ellagic acid nanoparticles the ROS level increased. Moreover, Hoechst staining indicated the anti-tumor effect of Co3O4@Glu-Ellagic acid NP by appearance of apoptotic bodies in HepG2 cells. This work revealed that Co3O4@Glu-Ellagic acid NPs are potent apoptogenic agent against liver cancer cells.

Synthesis, characterization, molecular modeling studies, and biological evaluation of metal piroxicam complexes (M = Ni(II), Pt(IV), Pd(II), Ag(I)) as antibacterial and anticancer agents.

Four piroxicam metal complexes; NiL2 , PtL2 , PdL2 , and AgL were synthesized and characterized by different techniques with enhanced antibacterial and anticancer activity. Regarding in vitro antimicrobial activity, complex NiL2 displayed potent antibacterial effect against Escherichia coli and Pseudomonas aeruginosa that was 1.9-folds higher than piroxicam (minimum inhibitory concentration [MIC] = 31.85, 65.32 µM), respectively. In case of G+ve bacteria, complex PtL2 had potent activity on Staphylococcus aureus which was 2.1-folds higher than piroxicam (MIC = 43.12 µM), while activity of complex AgL against Enterococcus faecalis was threefolds higher than piroxicam (MIC = 74.57 µM. Complexes PtL2 and PdL2 exhibited higher inhibition of DNA gyrase than piroxicam (IC50 = 6.21 µM) in the range of 1.9-1.7-folds. The in vitro antiproliferative activity depicted that all investigated complexes showed better cytotoxic effect than piroxicam, specifically Pt and Pd complexes which had lower IC50 values than piroxicam on human liver cancer cell line HepG2 by 1.8 and 1.7-folds, respectively. While Pd and Ag complexes showed 2 and 1.6-folds better effect on human colon cancer cell line HT-29 compared with piroxicam. Molecular modeling studies including docking on Stranded DNA Duplex (1juu) and DNA gyrase enzyme (1kzn) that gave good insight about interaction of complexes with target molecules, calculation of electrostatic potential map and global reactivity descriptors were performed.

Coculture of bacterial levans and evaluation of its anti-cancer activity against hepatocellular carcinoma cell lines

This research represents a novel study to assess how coculture affects levan yield, structure, bioactivities, and molecular weight. Among the 16 honey isolates, four bacterial strains recorded the highest levan yield. The Plackett–Burman design showed that the coculture (M) of isolates G2 and K2 had the maximum levan yield (52 g/L) and the effective factors were sucrose, incubation time, and sugarcane bagasse. The CCD showed that the most proper concentrations for maximum levan yield (81 g/L): were 130 g/L of sucrose and 6 g/f of sugarcane bagasse. Levan’s backbone was characterized, and the molecular weight was determined. G2 and K2 isolates were identified based on 16 sRNA as Bacillus megaterium strain YM1C10 and Rhizobium sp. G6-1. M levan had promising antioxidant activity (99.66%), slowed the migration activity to a great extent, and recorded 70.70% inhibition against the hepatoblastoma cell line (HepG2) at 1000 µg/mL. Gene expression analysis in liver cancer cell lines (HePG2) revealed that M levan decreased the expression of CCL20), 2GRB2, and CCR6) genes and was superior to Doxo. While increasing the expression of the IL4R and IL-10 genes. The DNA damage values were significantly increased (P < 0.01) in treated liver cancer cell lines with levan M and Doxo. The results referred to the importance of each of the hydroxyl and carboxyl groups and the molecular weight in levans bioactivities.