Effect of Pyridoxine Derivative B6NO on Transcription Factor Nrf2 Activity and Cytotoxic Properties of Doxorubicin In Vitro.

The effect of a new pyridoxine derivative B6NO on doxorubicin cytotoxicity and Nrf2-dependent cellular processes in vitro was studied. Antioxidant B6NO enhances the cytotoxic effect of doxorubicin on tumor cells, which is associated with G2/M cell division arrest and an increase in activity of proapoptotic enzyme caspase-3. The antioxidant promotes intracellular accumulation and nuclear translocation of Nrf2 transcription factor in non-tumor and tumor cells. In non-tumor cells, B6NO increases the expression of antioxidant system proteins and reduces ROS generation in the presence of doxorubicin. In tumor cells, no activation of Nrf2-dependent processes occurs under the action of the antioxidant. Our findings demonstrate the prospect of further studies of pyridoxine derivatives as antioxidants to reduce adverse reactions during chemotherapy.

Investigation of prunetrin induced G2/M cell cycle arrest and apoptosis via Akt/mTOR/MAPK pathways in hepatocellular carcinoma cells.

Hepatocellular carcinoma (HCC) stands as a leading cause of mortality, and despite recent advancements in the overall survival rates, the prognosis remains dismal. Prunetin 4-O-glucoside (Prunetrin or PUR), an active compound derived from Prunus sp., was explored for its impact on HepG2 and Huh7 cells. The cytotoxicity assessment revealed a notable reduction in cell viability in both cell lines, while exhibiting non-toxicity towards HaCaT cells. Colony formation studies underscored PUR's inhibitory effect on cell proliferation, dose-dependently. Mechanistically, PUR downregulated cell cycle proteins (CDC25c, Cdk1/CDC2, and Cyclin B1), inducing G2/M phase arrest, corroborated by flow cytometry. Western blot analyses exhibited dose-dependent cleavages of PARP and caspase 3, indicative of apoptosis. Treatment with the apoptotic inhibitor z-vmd-fmk provided evidence of PUR-induced apoptosis. Annexin V and PI flow cytometry further affirmed apoptotic induction. Enhanced expression of cleaved-caspase 9 and the pro-apoptotic protein Bak, coupled with reduced anti-apoptotic Bcl-xL, and affirmed PUR's induction of intrinsic apoptosis. Additionally, PUR activated the MAPK pathway, evidenced by elevated phospho p38 and phospho ERK expressions in both cell lines. Notably, a concentration-dependent decrease in mTOR and Akt expressions indicated PUR's inhibition of the Akt/mTOR pathway in HepG2 and Huh7 cells. These findings illuminate PUR's multifaceted impact, revealing its potential as a promising therapeutic agent against HepG2 and Huh7 cells through modulation of cell cycle, apoptosis, and key signaling pathways.

Triazole-based estradiol dimers prepared via CuAAC from 17α-ethinyl estradiol with five-atom linkers causing G2/M arrest and tubulin inhibition.

Microtubule dynamic is exceptionally sensitive to modulation by small-molecule ligands. Our previous work presented the preparation of microtubule-targeting estradiol dimer (ED) with anticancer activity. In the present study, we explore the effect of selected linkers on the biological activity of the dimer. The linkers were designed as five-atom chains with carbon, nitrogen or oxygen in their centre. In addition, the central nitrogen was modified by a benzyl group with hydroxy or methoxy substituents and one derivative possessed an extended linker length. Thirteen new dimers were subjected to cytotoxicity assay and cell cycle profiling. Dimers containing linker with benzyl moiety substituted with one or more methoxy groups and longer branched ones were found inactive, whereas other structures had comparable efficacy as the original ED (e.g. D1 with IC50 = 1.53 µM). Cell cycle analysis and immunofluorescence proved the interference of dimers with microtubule assembly and mitosis. The proposed in silico model and calculated binding free energy by the MM-PBSA method were closely correlated with in vitro tubulin assembly assay.

SP/NK1R system regulates carcinogenesis in prostate cancer: Shedding light on the antitumoral function of aprepitant.

AIMS: Prostate cancer continues to be one of the main global health issues in men. Neuropeptide substance P (SP) acting via neurokinin-1receptor (NK1R) promotes tumorigenicity in many human malignant tumors. However, its pro-tumorigenic functions and the therapeutic effects of its inhibition in prostate cancer remain unclear. METHODS: MTT assay was employed for measuring cellular proliferation and cytotoxicity. mRNAs and proteins expression levels were evaluated by qRT-PCR and western blot assay, respectively. Gelatinase activity was assessed by zymography. The migration ability was defined using wound-healing assay. Flow cytometry was employed to evaluate the cell cycle distribution. We also performed an in vivo experiment in a mouse model of prostate cancer to confirm the in vitro therapeutic effect of targeting the SP/NK1R system. RESULTS: We found a noticeable increase in the expression of the truncated isoform of NK1R as an oncogenic NK1R splice variant in tumor cells. We also demonstrated that SP promotes both proliferative and migrative phenotypes of prostate cancer through modifying cell cycle-related proteins (c-Myc, cyclin D1, cyclin B1, p21), and apoptosis-related genes (Bcl-2 and Bax), promoting cell migration and increasing MMP-2 and MMP-9 expression and activity, while aprepitant administration could remarkably reverse these effects. SP also stimulated tumor growth in vivo, which was correlated with shorter survival times, while aprepitant reversed this effect and led to significantly longer survival time. SIGNIFICANCE: Our findings suggest that SP/NK1R system may serve as a novel therapeutic target in prostate cancer and support the possible candidacy of aprepitant in future prostate cancer therapy.

Intracellular Trafficking of Cationic Carbon Dots in Cancer Cell Lines MCF-7 and HeLa-Time Lapse Microscopy, Concentration-Dependent Uptake, Viability, DNA Damage, and Cell Cycle Profile.

Fluorescent carbon dots (CDs) are potential tools for the labeling of cells with many advantages such as photostability, multicolor emission, small size, rapid uptake, biocompatibility, and easy preparation. Affinity towards organelles can be influenced by the surface properties of CDs which affect the interaction with the cell and cytoplasmic distribution. Organelle targeting by carbon dots is promising for anticancer treatment; thus, intracellular trafficking and cytotoxicity of cationic CDs was investigated. Based on our previous study, we used quaternized carbon dots (QCDs) for treatment and monitoring the behavior of two human cancer cell MCF-7 and HeLa lines. We found similarities between human cancer cells and mouse fibroblasts in the case of QCDs uptake. Time lapse microscopy of QCDs-labeled MCF-7 cells showed that cells are dying during the first two hours, faster at lower doses than at higher ones. QCDs at a concentration of 100 µg/mL entered into the nucleus before cellular death; however, at a dose of 200 µg/mL, blebbing of the cellular membrane occurred, with a subsequent penetration of QCDs into the nuclear area. In the case of HeLa cells, the dose-depended effect did not happen; however, the labeled cells were also dying in mitosis and genotoxicity occurred nearly at all doses. Moreover, contrasted intracellular compartments, probably mitochondria, were obvious after 24 h incubation with 100 µg/mL of QCDs. The levels of reactive oxygen species (ROS) slightly increased after 24 h, depending on the concentration, thus the genotoxicity was likely evoked by the nanomaterial. A decrease in viability did not reach IC 50 as the DNA damage was probably partly repaired in the prolonged G0/G1 phase of the cell cycle. Thus, the defects in the G2/M phase may have allowed a damaged cell to enter mitosis and undergo apoptosis. The anticancer effect in both cell lines was manifested mainly through genotoxicity.

Arachidin-1, a Prenylated Stilbenoid from Peanut, Induces Apoptosis in Triple-Negative Breast Cancer Cells.

Triple-negative breast cancer (TNBC) is unresponsive to typical hormonal treatments, causing it to be one of the deadliest forms of breast cancer. Investigating alternative therapies to increase survival rates for this disease is essential. The goal of this study was to assess cytotoxicity and apoptosis mechanisms of prenylated stilbenoids in TNBC cells. The prenylated stilbenoids arachidin-1 (A-1) and arachidin-3 (A-3) are analogs of resveratrol (RES) produced in peanut upon biotic stress. The anticancer activity of A-1 and A-3 isolated from peanut hairy root cultures was determined in TNBC cell lines MDA-MB-231 and MDA-MB-436. After 24 h of treatment, A-1 exhibited higher cytotoxicity than A-3 and RES with approximately 11-fold and six-fold lower IC50, respectively, in MDA-MB-231 cells, and nine-fold and eight-fold lower IC50, respectively, in MDA-MB-436 cells. A-1 did not show significant cytotoxicity in the non-cancerous cell line MCF-10A. While A-1 blocked cell division in G2-M phases in the TNBC cells, it did not affect cell division in MCF-10A cells. Furthermore, A-1 induced caspase-dependent apoptosis through the intrinsic pathway by activating caspase-9 and PARP cleavage, and inhibiting survivin. In conclusion, A-1 merits further research as a potential lead molecule for the treatment of TNBC.

Characterization of Cisplatin Effects in Lenvatinib-resistant Hepatocellular Carcinoma Cells.

BACKGROUND/AIM: Drug resistance to molecular targeted agents, such as lenvatinib, is an important issue. The aim of this study was to explore the mechanism of lenvatinib resistance and to investigate potential drugs that may improve the treatment of lenvatinib-resistant (LR) hepatocellular carcinoma (HCC). MATERIALS AND METHODS: LR cells were developed by long-term culture under lenvatinib exposure. We analyzed the biological characteristics of LR cells in vitro, and investigated the antitumor effects and endogenous mechanisms of cisplatin in LR cells. RESULTS: The proliferative potential of LR cells was enhanced by activation of ERK signaling and changes in several miRNAs. Cisplatin inhibited cell proliferation of LR cells and induced G2/M cell cycle arrest. Furthermore, cisplatin triggered the DNA damage response, via the ATM/ATR-Chk1/Chk2 signaling pathway. CONCLUSION: Proliferation of LR cells was induced upon ERK signaling activation. Cisplatin exerted antitumor effects in LR cells and was involved in the regulation of miRNAs associated with drug resistance.

Mineral Nanomedicine to Enhance the Efficacy of Adjuvant Radiotherapy for Treating Osteosarcoma.

It is vital to remove residual tumor cells after resection to avoid the recurrence and metastasis of osteosarcoma. In this study, a mineral nanomedicine, europium-doped calcium fluoride (CaF2:Eu) nanoparticles (NPs), is developed to enhance the efficacy of adjuvant radiotherapy (i.e., surgical resection followed by radiotherapy) for tumor cell growth and metastasis of osteosarcoma. In vitro studies show that CaF2:Eu NPs (200 µg/mL) exert osteosarcoma cell (143B)-selective toxicity and migration-inhibiting effects at a Eu dopant amount of 2.95 atomic weight percentage. These effects are further enhanced under X-ray irradiation (6 MeV, 4 Gy). Furthermore, in vivo tests show that intraosseous injection of CaF2:Eu NPs and X-ray irradiation have satisfactory therapeutic efficacy in controlling primary tumor size and inhibiting primary tumor metastasis. Overall, our results suggest that CaF2:Eu NPs with their osteosarcoma cell (143B)-selective toxicity and migration-inhibiting effects combined with radiotherapy might be nanomedicines for treating osteosarcoma after tumor resection.

Discovery of Novel Histone Deacetylase 6 (HDAC6) Inhibitors with Enhanced Antitumor Immunity of Anti-PD-L1 Immunotherapy in Melanoma.

A series of 2-phenylthiazole analogues were designed and synthesized as potential histone deacetylase 6 (HDAC6) inhibitors based on compound 12c (an HDAC6/tubulin dual inhibitor discovered by us recently) and CAY10603 (a known HDAC6 inhibitor). Among them, compound XP5 was the most potent HDAC6 inhibitor with an IC50 of 31 nM and excellent HDAC6 selectivity (SI = 338 for HDAC6 over HDAC3). XP5 also displayed high antiproliferative activity against various cancer cell lines including the HDACi-resistant YCC3/7 gastric cancer cells (IC50 = 0.16-2.31 µM), better than CAY10603. Further, XP5 (50 mg/kg) exhibited significant antitumor efficacy in a melanoma tumor model with a tumor growth inhibition (TGI) of 63% without apparent toxicity. Moreover, XP5 efficiently enhanced the in vivo antitumor immune response when combined with a small-molecule PD-L1 inhibitor, as demonstrated by the increased tumor-infiltrating lymphocytes and reduced PD-L1 expression levels. Taken together, the above results suggest that XP5 is a promising HDAC6 inhibitor deserving further investigation.

Structure-Based Design and Synthesis of N-Substituted 3-Amino-ß-Carboline Derivatives as Potent αß-Tubulin Degradation Agents.

So far, relatively few small molecules have been reported to promote tubulin degradation. Our previous studies have found that compound 2, a noncovalent colchicine-site ligand, was capable of promoting αß-tubulin degradation. To further improve its antiproliferative activity, 66 derivatives or analogues of 2 were designed and synthesized based on 2-tubulin cocrystal structure. Among them, 12b displayed nanomolar potency against a variety of tumor cells, including paclitaxel- and adriamycin-resistant cell lines. 12b binds to the colchicine site and promotes αß-tubulin degradation in a concentration-dependent manner via the ubiquitin-proteasome pathway. The X-ray crystal structure revealed that 12b binds in a similar manner as 2, but there is a slight conformation change of the B ring, which resulted in better interaction of 12b with surrounding residues. 12b effectively suppressed tumor growth at an i.v. dose of 40 mg/kg (3 times a week) on both A2780S (paclitaxel-sensitive) and A2780T (paclitaxel-resistant) ovarian xenograft models, with respective TGIs of 92.42 and 79.75% without obvious side effects, supporting its potential utility as a tumor-therapeutic compound.

GPR17 signaling activation by CHBC agonist induced cell death via modulation of MAPK pathway in glioblastoma.

AIM: Glioblastoma multiforme (GBM) is the most common and aggressive primary adult brain tumor. GBM is characterized by a heterogeneous population of cells that are resistant to chemotherapy. Recently, we have synthesized CHBC, a novel indole derivative targeted to GBM biomarker G-protein-coupled receptor 17 and inhibitor of GBM cells. In this study, CHBC was further investigated to characterize the efficiency of this agonist at the molecular level and its underlying mechanism in GBM cell death induction. MATERIALS AND METHODS: The effect of CHBC and TMZ was determined using time dependent inhibitor assay in glioblastoma cells, LN229 and SNB19. Drug induced cell cycle arrest was measured using PI staining followed by image analysis. The induction of apoptosis and mechanism of action of CHBC was studied using apoptosis, caspase 3/7 and mitochondrial membrane permeability assays. Modulation of the key genes involved in MAPK signaling pathway was also measured using immunoblotting array. KEY FINDINGS: The inhibitory kinetic study has revealed that CHBC inhibited SNB19 and LN229 cell growth in a time-dependent manner. Furthermore, CHBC with the IC50 of 85 µM, mediated cell death through an apoptosis mechanism in both studied cell lines. The study also has revealed that CHBC targets GPR17 leading to the induction of apoptosis via the activation of Caspase 3/7 and dysfunction of mitochondrial membrane potential. In addition, CHBC treatment led to marked G2/M cell cycle arrest. The protein array has confirmed the anticancer effect of CHBC by the disruption of the mitogen-activated protein kinase pathway (MAPK). SIGNIFICANCE: Taken together, these results demonstrated that CHBC induced G2/M cell cycle arrest and apoptosis by disrupting MAPK signaling in human glioblastoma cells. This study concludes that CHBC represent a class of compounds for treating glioblastoma.

A Second Near-Infrared Ru(II) Polypyridyl Complex for Synergistic Chemo-Photothermal Therapy.

The clinical success of cisplatin ushered in a new era of the application of metallodrugs. When it comes to practice, however, drug resistance, tumor recurrence, and drug systemic toxicity make it implausible to completely heal the patients. Herein, we successfully transform an electron acceptor [1, 2, 5]thiadiazolo[3,4-g]quinoxaline into a novel second near-infrared (NIR-II) fluorophore H7. After PEGylation and chelation, HL-PEG2k exhibits a wavelength bathochromic shift, enhanced photothermal conversion efficiency (41.77%), and an antineoplastic effect against glioma. Its potential for in vivo tumor tracking and image-guided chemo-photothermal therapy is explored. High levels of uptake and high-resolution NIR-II imaging results are thereafter obtained. The hyperthermia effect could disrupt the lysosomal membranes, which in turn aggravate the mitochondria dysfunction, arrest the cell cycle in the G2 phase, and finally lead to cancer cell apoptosis. HL-PEG2k displays a superior biocompatibility and thus can be a potential theranostic platform to combat the growth and recurrence of tumors.

Design, synthesis and structure-activity relationship studies of novel spirochromanone hydrochloride analogs as anticancer agents.

Aim: Literature reports suggest spirochromanone derivatives exhibit anticancer activity. Methodology: The authors designed and synthesized 18 spirochromanone derivatives (Csp 1-18). The compounds were characterized and evaluated for anticancer activity against human breast cancer (MCF-7) and murine melanoma (B16F10) cell lines. Results: The anticancer activity ranged from 4.34 to 29.31 µm. The most potent compounds, Csp 12 and Csp 18, were less toxic against the human embryonic kidney (HEK-293) cell line and ∼ two/∼fourfold selective toward MCF-7 than B16F10 in comparison to the reference, BG-45. Csp 12 caused 28.6% total apoptosis, leading to significant cytotoxicity, and arrested the G2 phase of the cell cycle in B16F10 cells. A molecular docking study of Csp 12 exhibited effective binding at the active site of the epidermal growth factor receptor kinase domain. Conclusion: This study highlights the importance of spirochromanones as anticancer agents.

Investigation of the antileishmanial activity and mechanisms of action of acetyl-thiohydantoins.

The currently available treatment options for leishmaniasis are associated with high costs, severe side effects, and high toxicity. In previous studies, thiohydantoins demonstrated some pharmacological activities and were shown to be potential hit compounds with antileishmanial properties. The present study further explored the antileishmanial effect of acetyl-thiohydantoins against Leishmania amazonensis and determined the main processes involved in parasite death. We observed that compared to thiohydantoin nuclei, acetyl-thiohydantoin treatment inhibited the proliferation of promastigotes. This treatment caused alterations in cell cycle progression and parasite size and caused morphological and ultrastructural changes. We then investigated the mechanisms involved in the death of the protozoan; there was an increase in ROS production, phosphatidylserine exposure, and plasma membrane permeabilization and a loss of mitochondrial membrane potential, resulting in an accumulation of lipid bodies and the formation of autophagic vacuoles on these parasites and confirming an apoptosis-like process. In intracellular amastigotes, selected acetyl-thiohydantoins reduced the percentage of infected macrophages and the number of amastigotes/macrophages by increasing ROS production and reducing TNF-α levels. Moreover, thiohydantoins did not induce cytotoxicity in murine macrophages (J774A.1), human monocytes (THP-1), or sheep erythrocytes. In silico and in vitro analyses showed that acetyl-thiohydantoins exerted in vitro antileishmanial effects on L. amazonensis promastigotes in apoptosis-like and amastigote forms by inducing ROS production and reducing TNF-α levels, indicating that they are good candidates for drug discovery studies in leishmaniasis treatment. Additionally, we carried out molecular docking analyses of acetyl-thiohydantoins on two important targets of Leishmania amazonensis: arginase and TNF-alpha converting enzyme. The results suggested that the acetyl groups in the N1-position of the thiohydantoin ring and the ring itself could be pharmacophoric groups due to their affinity for binding amino acid residues at the active site of both enzymes via hydrogen bond interactions. These results demonstrate that thiohydantoins are promising hit compounds that could be used as antileishmanial agents.

Branched alkyl of phenyl 4-(2-oxo-3-alkylimidazolidin-1-yl)benzenesulfonates as unique cytochrome P450 1A1-activated antimitotic prodrugs: Biological evaluation and mechanism of bioactivation.

We recently discovered a new family of prodrugs deriving from phenyl 4-(2-oxo-3-imidazolidin-1-yl)benzenesulfonates (PIB-SOs) bioactivatable by cytochrome P450 1A1 (CYP1A1) into potent antimitotics referred to as phenyl 4-(2-oxo-3-alkylimidazolidin-1-yl)benzenesulfonates (PAIB-SOs). PAIB-SOs display significant selectivity toward human breast cancer cells based on the N-dealkylation of PAIB-SOs into their corresponding PIB-SOs by CYP1A1. In this study, we have evaluated the molecular mechanism of the bioactivation of PAIB-SOs into PIB-SOs by branching the linear alkyl chain on the imidazolidin-2-one (IMZ) moiety of PAIB-SOs by branched alkyl groups such as isopropyl, isobutyl and sec-butyl. Our results show that PAIB-SOs bearing an isobutyl group on the IMZ moiety and either a methoxy, a chloro or a bromo group at positions 3, 3,5 or 3,4,5 on the aromatic ring B exhibit antiproliferative activity ranging from 0.13 to 6.9 µM and selectivity toward MCF7 and MDA-MB-468 mammary cancer cells comparatively to other cell lines tested. Moreover, the most potent and selective PAIB-SOs bearing an isobutyl group and either a 3,5-Cl (44), 3,5-Br (45) or a 3,4,5-OMe (46) on the IMZ moiety exhibit antiproliferative activity in the sub-micromolar range and high selectivity ratios toward mammary cancer cells. They stop the cell cycle of MCF7 cells in the G2/M phase and disrupt their cytoskeleton. Furthermore, our studies evidenced that PAIB-SOs bearing either an isopropyl, a sec-butyl or an isobutyl group are hydroxylated on the carbon atom adjacent to the IMZ (Cα-OH) but only PAIB-SOs bearing an isobutyl group are bioactivated into PIB-SOs. Finally, PAIB-SOs 45 and 46 exhibit low toxicity toward normal cells and chick embryos and are thus promising antimitotic prodrugs highly selective toward CYP1A1-expressing breast cancer cells.

Protective role of m6A binding protein YTHDC2 on CCNB2 in manganese-induced spermatogenesis dysfunction.

Human infertility has become the third largest serious disease in the world, seriously affecting the quality of human fertility. Studies have shown that manganese (Mn) can accumulate in the testis through the blood-testicular barrier and damage the male reproductive system. However, the mechanism has not been explored clearly. Recent studies have reported that YTH domain-containing 2 (YTHDC2) can regulate reproductive function. However, none has explored the role of YTHDC2 in Mn-induced reproductive toxicity. The present study investigated whether YTHDC2/CyclinB2 (CCNB2) pathway participates in Mn-induced reproductive toxicity using Kunming mice, spermatogonia, and the seminal plasma of male workers. The mice were received intraperitoneal (i.p.) injections of 0, 12.5, 25, and 50 mg/kg MnCl2 once daily for 2 weeks. The cells were treated with 0, 100, 200 and 400 µM MnCl2 for 24 h. Here, we found that occupational Mn exposure significantly increased Mn levels in the seminal plasma of male workers, while decreased sperm density, semen quality, and the levels of YTHDC2, CCNB1, and CCNB2. We found that Mn can inhibit the YTHDC2/CCNB2 signaling pathway and block the G2/M phase of the cell cycle. Moreover, the morphology of cells and the histomorphology of mice testis were injured. Notably, over-expression (OE) of YTHDC2 increased CCNB2 levels, reduced cell cycle arrest, and improved reproductive toxicity after Mn exposure. These findings suggest that the YTHDC2/CCNB2 signaling pathway participates in Mn-induced reproductive toxicity, and OE of YTHDC2 can mitigate the toxicity of Mn.

Shikonin N-benzyl matrinic acid ester derivatives as novel telomerase inhibitors with potent activity against lung cancer cell lines.

In this study, a series of novel shikonin N-benzyl matrinic acid ester derivatives (PMMB-299-PMMB-310) were synthesized and tested for their ability to inhibit the proliferation of cancer cells. Compared with shikonin and matrine, some of the ester derivatives were found to exhibit better anti-proliferative activity against seven different cancer cell lines, with less cytotoxicity toward non-cancerous cells. The strongest anti-proliferative activity was exhibited by PMMB-302, which had an IC50 value of 2.71 µM against A549 cells. The compound caused cell cycle arrest in the G2/M phase and induced apoptosis. Effects on the expression of apoptosis-related molecules such as Bcl2, Bcl-XL, caspase-3, caspase-9 and FADD suggested that PMMB-302 has tumor suppressive roles in lung cancer cells. In addition, PMMB-302 inhibited expression of telomerase core proteins, dyskerin and NHP2, and telomerase reverse transcriptase RNA. Moreover, molecular docking of PMMB-302 was subsequently conducted to determine the probable binding mode with telomerase. Taken together, the results indicate that PMMB-302 acts as a tumor suppressor in lung cancer cells by negatively regulating telomerase expression.

Theaflavin-3-gallate, a natural antagonist for Hsp90: In-silico and in-vitro approach.

Poor prognosis and metastasis have been recognized as the major cause of breast cancer related deaths worldwide. Recent experimental evidence has shown that Hsp90, the prime chaperone, is overexpressed in many cancers and is responsible if reducing the 5-year survival rate of cancer patients. Therefore, targeted inhibition of Hsp90 may be a new and effective way to target cancer as well as enhancing therapeutic outcomes. In the present study, screening and simulation of potential natural compounds result in the identification of theaflavin-3-gallate as a promising inhibitory compound of Hsp90. Further in-vitro validation of the cytotoxic effect of theaflavin-3-gallate in human breast carcinoma cell line MCF7 and normal cell line MCF10A revealed that theaflavin-3-gallate significantly inhibited the cell proliferation of MCF7 cells whereas no cytotoxic effect was observed on MCF10A cells. We also found that theaflavin-3-gallate significantly induced programmed cell death by arresting cells in the G2/M phase of the cell cycle. A significant decrease in cell migration and colony formation by theaflavin-3-gallate treatment was also observed in MCF7 cells. Furthermore, theaflavin-3-gallate significantly downregulated the mRNA expression patterns of the HSP90, MMP9, VEGFA, and SPP1 genes. Collectively, our results demonstrated theaflavin-3-gallate as a potential natural Hsp90 inhibitor that can be used to enhance the therapeutic efficacy of existing breast cancer therapies and improve overall survival of breast cancer patients.

Control of ER-positive breast cancer by ERα expression inhibition, apoptosis induction, cell cycle arrest using semisynthetic isoeugenol derivatives.

New semi-synthetic effective and safe anticancer agents isoeugenol derivatives were synthesized, characterized, and screened for their cytotoxic activity against MCF-7. Moreover, their selective cytotoxicity was assessed against MCF-10A. Three derivatives, 2, 8 and 10 were significantly more active than the reference drug 5-FU with IC50 values of 6.59, 8.07 and 9.63 and 30.93 µM, respectively. Also interestingly, these derivatives demonstrated some degree of selectivity to cancer cells over normal cells. Furthermore, derivative 2 was subjected to other in vitro experiments against MCF-7 where it inhibited colony formation by 87.5% and lowered ERα concentration to 395.7 pg/mL compared to 1129 pg/mL in untreated control cells. In continuation of the investigation, the apoptotic activity of compound 2, was assessed where it significantly enhanced total apoptotic cell death by 9.16-fold (18.70% compared to 1.64% for the untreated MCF-7 control cells) and arrested the cell cycle at the G2/M phase. Furthermore, the molecular mechanism of apoptotic activity was investigated at both the gene (RT-PCR) and protein (western plotting) levels where upregulation of pro-apoptotic and down regulation of anti-apoptotic genes was detected. Additionally, compound 2 treatment enhanced the antioxidant (GSH, CAT, SOD) activities. Finally, in vivo experiments verified the effective anticancer activity of compound 2 through inhibition of tumor proliferation by 47.6% compared to 22.9% for 5-FU and amelioration of the hematological, biochemical, and histopathological examinations near normal. In effect, compound 2 can be viewed as a promising semi-synthetic derivative of isoeugenol with some degree of selectivity for management of breast cancer through apoptotic induction and ERα downregulation.

The effect of paclitaxel on apoptosis, autophagy and mitotic catastrophe in AGS cells.

Paclitaxel is an anti-microtubule agent that has been shown to induce cell death in gastric cancer. However, the detailed mechanism of action is unclear. In this study, we reveal that the paclitaxel-induced cell death mechanism involves mitotic catastrophe, autophagy and apoptosis in AGS cells. Paclitaxel induced intrinsic apoptosis by activating caspase-3, caspase-9 and PARP. In addition, the significant increase in autophagy marker LC3B-II, together with Atg5, class III PI3K and Beclin-1, and the down-regulation of p62 following paclitaxel treatment verified that paclitaxel induced autophagy. Further experiments showed that paclitaxel caused mitotic catastrophe, cell cycle arrest of the accumulated multinucleated giant cells at the G2/M phase and induction of cell death in 24 h. Within 48 h, the arrested multinucleated cells escaped mitosis by decreasing cell division regulatory proteins and triggered cell death. Cells treated with paclitaxel for 48 h were grown in fresh medium for 24 h and checked for CDC2, CDC25C and lamin B1 protein expressions. These proteins had decreased significantly, indicating that the remaining cells became senescent. In conclusion, it is suggested that paclitaxel-induced mitotic catastrophe is an integral part of the cell death mechanism, in addition to apoptosis and autophagy, in AGS cells.