Probiotic-derived silver nanoparticles target mTOR/MMP-9/BCL-2/dependent AMPK activation for hepatic cancer treatment.

Recent advances in nanotechnology have offered novel ways to combat cancer. By utilizing the reducing capabilities of Lactobacillus acidophilus, silver nanoparticles (AgNPs) are synthesized. The anti-cancer properties of AgNPs have been demonstrated in previous studies against several cancer cell lines; it has been hypothesized that these compounds might inhibit AMPK/mTOR signalling and BCL-2 expression. Consequently, the current research used both in vitro and in silico approaches to study whether Lactobacillus acidophilus AgNPs could inhibit cell proliferation autophagy and promote apoptosis in HepG2 cells. The isolated strain was identified as Lactobacillus acidophilus strain RBIM based on 16 s rRNA gene analysis. Based on our research findings, it has been observed that this particular strain can generate increased quantities of AgNPs when subjected to optimal growing conditions. The presence of silanols, carboxylates, phosphonates, and siloxanes on the surface of AgNPs was confirmed using FTIR analysis. AgNPs were configured using UV-visible spectroscopy at 425 nm. In contrast, it was observed that apoptotic cells exhibited orange-coloured bodies due to cellular shrinkage and blebbing initiated by AgNP treatment, compared to non-apoptotic cells. It is worth mentioning that AgNPs exhibited remarkable selectivity in inducing cell death, specifically in HepG2 cells, unlike normal WI-38 cells. The half-maximum inhibitory concentration (IC50) values for HepG2 and WI-38 cells were 4.217 µg/ml and 154.1 µg/ml, respectively. AgNPs induce an upregulation in the synthesis of inflammation-associated cytokines, including (TNF-α and IL-33), within HepG2 cells. AgNPs co-treatment led to higher glutathione levels and activating pro-autophagic genes such as AMPK.Additionally, it resulted in the suppression of mTOR, MMP-9, BCL-2, and α-SMA gene expression. The docking experiments suggest that the binding of AgNPs to the active site of the AMPK enzyme leads to inhibiting its activity. The inhibition of AMPK ultimately results in the suppression of the mechanistic mTOR and triggers apoptosis in HepG2 cells. In conclusion, the results of our study indicate that the utilization of AgNPs may represent a viable strategy for the eradication of liver cancerous cells through the activation of apoptosis and the enhancement of immune system reactions.

Chitosan-loaded piperlongumine nanoparticles and kaempferol enhance the anti-cancer action of doxorubicin in targeting of Ehrlich solid adenocarcinoma: in vivo and in silico modeling study.

Doxorubicin is a chemotherapeutic drug that generates free radical-induced toxicities. Natural agents are used to potentiate or ameliorate the toxicity of chemotherapy. None of the studies investigating whether antioxidants or prooxidants should be used with chemotherapy have addressed their efficacy in the same study. Therefore, the aim of this study was to investigate the potential synergy between doxorubicin and two natural rarely in vivo studied anticancer agents; the antioxidant "Kaempferol" and prooxidant "Piperlongumine" in Ehrlich tumor mice model. 77 albino mice were divided into 11 groups; Ehrlich ascites carcinoma cells were injected intramuscularly to develop solid tumors. After 14 days, intratumoral injections of single or combinations of free or Chitosan nanoparticles loaded with doxorubicin, Piperlongumine, and Kaempferol were performed. Tumor Characterization of nanoparticles was measured, tumors were histopathologically examined and evaluation of expression for cancer-related genes by real-time PCR. In silico molecular docking was performed to uncover potential novel targets for Piperlongumine and Kaempferol. Despite receiving half of the overall dose compared to the free drugs, the combined doxorubicin/ piperlongumine-chitosan nanoparticles treatment was the most efficient in reducing tumor volume; down-regulating Cyclin D1, and BCL2; as well as the Beclin-1, and Cyclophilin A genes modulating growth, apoptosis, autophagy, and metastasis, respectively; up-regulating the Glutathione peroxidase expression as a defense mechanism protecting from oxidative damage. When combined with doxorubicin, Kaempferol and Piperlongumine were effective against Ehrlich solid tumors. However, the combination with the Piperlongumine-loaded chitosan nanoparticles significantly enhanced its anticancer effect compared to the Kaempferol or the same free compounds.

Novel phloretin-based combinations targeting glucose metabolism in hepatocellular carcinoma through GLUT2/PEPCK axis of action: in silico molecular modelling and in vivo studies.

Hepatocellular carcinoma (HCC) is commonly associated with disturbances in glucose metabolism and enhanced glycolysis. However, a controversial role for gluconeogenesis was reported to be tumor-promoting and tumor-suppressive. We investigated novel anti-HCC treatments through either the simultaneous inhibition of glycolysis and gluconeogenesis by "phloretin" and "sodium meta-arsenite", respectively (Combination 1); or the concurrent inhibition of glycolysis and induction of gluconeogenesis by phloretin and dexamethasone, respectively, (combination 2). A total of 110 Swiss albino mice were divided into eleven groups, HCC was induced by N, N-dimethyl-4-aminoazobenzene. We have measured the expression of the glucose transporter 2 (GLUT2), Phosphoenolpyruvate carboxykinases (PEPCK), Caspase-3, Beclin 1, Cyclin D1, and cytokeratin 18 genes; blood glucose and ATP levels; alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities. Furthermore, in silico molecular docking was performed to investigate the potential drug-receptor interactions. Histologically, the phloretin-based combinations resulted in a significant regression of malignant tissue compared to various treatments. GLUT2 and PEPCK mRNA analysis indicated successful off/on modulation of glycolysis and gluconeogenesis. Docking confirmed the potent binding between phloretin, sodium meta-arsenite, and dexamethasone with GLUT2, PEPCK, and Retinoid X Receptor Alpha, respectively. Molecularly, Combination 2 resulted in the highest reduction in cyclin D1, cytokeratin 18, and Beclin 1 expression contemporaneously with the upregulation in Caspase-3 levels. Biochemically, both combinations caused a significant reduction in ATP levels, ALT, and AST activity compared to the other groups. In conclusion, we propose two novel phloretin-based combinations that can be used in treating HCC through the regulation of glucose metabolism and ATP production.

Ammonia scavenger and glutamine synthetase inhibitors cocktail in targeting mTOR/ß-catenin and MMP-14 for nitrogen homeostasis and liver cancer.

The glutamine synthetase (GS) facilitates cancer cell growth by catalyzing de novo glutamine synthesis. This enzyme removes ammonia waste from the liver following the urea cycle. Since cancer development is associated with dysregulated urea cycles, there has been no investigation of GS's role in ammonia clearance. Here, we demonstrate that, although GS expression is increased in the setting of ß-catenin oncogenic activation, it is insufficient to clear the ammonia waste burden due to the dysregulated urea cycle and may thus be unable to prevent cancer formation. In vivo study, a total of 165 male Swiss albino mice allocated in 11 groups were used, and liver cancer was induced by p-DAB. The activity of GS was evaluated along with the relative expression of mTOR, ß-catenin, MMP-14, and GS genes in liver samples and HepG2 cells using qRT-PCR. Moreover, the cytotoxicity of the NH3 scavenger phenyl acetate (PA) and/or GS-inhibitor L-methionine sulfoximine (MSO) and the migratory potential of cells was assessed by MTT and wound healing assays, respectively. The Swiss target prediction algorithm was used to screen the mentioned compounds for probable targets. The treatment of the HepG2 cell line with PA plus MSO demonstrated strong cytotoxicity. The post-scratch remaining wound area (%) in the untreated HepG2 cells was 2.0%. In contrast, the remaining wound area (%) in the cells treated with PA, MSO, and PA + MSO for 48 h was 61.1, 55.8, and 78.5%, respectively. The combination of the two drugs had the greatest effect, resulting in the greatest decrease in the GS activity, ß-catenin, and mTOR expression. MSO and PA are both capable of suppressing mTOR, a key player in the development of HCC, and MMP-14, a key player in the development of HCC. PA inhibited the MMP-14 enzyme more effectively than MSO, implying that PA might be a better way to target HCC as it inhibited MMP-14 more effectively than MSO. A large number of abnormal hepatocytes (5%) were found to be present in the HCC mice compared to mice in the control group as determined by the histopathological lesions scores. In contrast, PA, MSO, and PA + MSO showed a significant reduction in the hepatic lesions score either when protecting the liver or when treating the liver. The molecular docking study indicated that PA and MSO form a three-dimensional structure with NF-κB and COX-II, blocking their ability to promote cancer and cause gene mutations. PA and MSO could be used to manipulate GS activities to modulate ammonia levels, thus providing a potential treatment for ammonia homeostasis.

Hepatocellular Carcinoma cells: activity of Amygdalin and Sorafenib in Targeting AMPK /mTOR and BCL-2 for anti-angiogenesis and apoptosis cell death.

BACKGROUND: Sorafenib (Sor) is the only approved multikinase inhibitor indicated for the treatment of HCC. Previous studies have shown that amygdalin (Amy) possesses anticancer activities against several cancer cell lines; we suggested that these compounds might disrupt AMPK/mTOR and BCL-2. Therefore, the current study used integrated in vitro and in silico approaches to figure out Amy and Sor's possible synergistic activity in targeting AMPK/mTOR and BCL-2 for anti-angiogenesis and apoptosis cell death in HepG2 cells. RESULTS: Notably, Amy demonstrated exceptional cytotoxic selectivity against HepG2 cells in comparison to normal WI-38 cells (IC50 = 5.21 mg/ml; 141.25 mg/ml), respectively. In contrast, WI-38 cells were far more sensitive to the toxicity of Sor. A substantial synergistic interaction between Amy and Sor was observed (CI50 = 0.56), which was connected to cell cycle arrest at the S and G2/M stages and increased apoptosis and potential necroptosis. Amy and Sor cotreatment resulted in the highest glutathione levels and induction of pro-autophagic genes AMPK, HGMB1, ATG5, Beclin 1, and LC3, suppressed the mTOR and BCL2 anti-apoptotic gene. Finally, the docking studies proposed that Amy binds to the active site of the AMPK enzyme, thus inhibiting its activity. This inhibition of AMPK ultimately leads to inhibition of mTOR and thus induces apoptosis in the HepG2 cells. CONCLUSION: Although more in vivo research using animal models is needed to confirm the findings, our findings contribute to the evidence supporting Amy's potential anticancer effectiveness as an alternative therapeutic option for HCC.

Thymoquinone, piperine, and sorafenib combinations attenuate liver and breast cancers progression: epigenetic and molecular docking approaches.

BACKGROUND: Traditional herbal medicine has been used for centuries to cure many pathological disorders, including cancer. Thymoquinone (TQ) and piperine (PIP) are major bioactive constituents of the black seed (Nigella sativa) and black pepper (Piper nigrum), respectively. The current study aimed to explore the potential chemo-modulatory effects, mechanisms of action, molecular targets, and binding interactions after TQ and PIP treatments and their combination with sorafenib (SOR) against human triple-negative breast cancer (MDA-MB-231) and liver cancer (HepG2) cells. METHODS: We determined drug cytotoxicity by MTT assay, cell cycle, and death mechanism by flow cytometry. Besides, the potential effect of TQ, PIP, and SOR treatment on genome methylation and acetylation by determination of DNA methyltransferase (DNMT3B), histone deacetylase (HDAC3) and miRNA-29c expression levels. Finally, a molecular docking study was performed to propose potential mechanisms of action and binding affinity of TQ, PIP, and SOR with DNMT3B and HDAC3. RESULTS: Collectively, our data show that combinations of TQ and/or PIP with SOR have significantly enhanced the SOR anti-proliferative and cytotoxic effects depending on the dose and cell line by enhancing G2/M phase arrest, inducing apoptosis, downregulation of DNMT3B and HDAC3 expression and upregulation of the tumor suppressor, miRNA-29c. Finally, the molecular docking study has identified strong interactions between SOR, PIP, and TQ with DNMT3B and HDAC3, inhibiting their normal oncogenic activities and leading to growth arrest and cell death. CONCLUSION: This study reported TQ and PIP as enhancers of the antiproliferative and cytotoxic effects of SOR and addressed the mechanisms, and identified molecular targets involved in their action.

A bicistronic CYCLIN D1-TROP2 mRNA chimera demonstrates a novel oncogenic mechanism in human cancer.

A chimeric CYCLIN D1-TROP2 mRNA was isolated from human ovarian and mammary cancer cells. The CYCLIN D1-TROP2 mRNA was shown to be a potent oncogene as it transforms naïve, primary cells in vitro and induces aggressive tumor growth in vivo in cooperation with activated RAS. Silencing of the chimeric mRNA inhibits the growth of breast cancer cells. The CYCLIN D1-TROP2 mRNA was expressed by a large fraction of the human gastrointestinal, ovarian, and endometrial tumors analyzed. It is most frequently detected in intestinal cell aneuploid cancers and it is coexpressed with activated RAS oncogenes, consistent with a cooperative transforming activity in human cancers. The chimeric mRNA is a bicistronic transcript of post transcriptional origin that independently translates the Cyclin D1 and Trop-2 proteins. This is a novel mechanism of CYCLIN D1 activation that achieves the truncation of the CYCLIN D1 mRNA in the absence of chromosomal rearrangements. This leads to a higher CYCLIN D1 mRNA stability, with inappropriate expression during the cell cycle. The stabilized CYCLIN D1 mRNA cooperates with TROP2 in stimulating the growth of the expressing cells. These findings show a novel epigenetic, oncogenic mechanism, which seems to be widespread in human cancers.