Proteasome activation: A novel strategy for targeting undruggable intrinsically disordered proteins.

Intrinsically disordered proteins (IDPs) are characterized by their inability to adopt well-defined tertiary structures under physiological conditions. Nonetheless, they often play pivotal roles in the progression of various diseases, including cancer, neurodegenerative disorders, and cardiovascular ailments. Owing to their inherent dynamism, conventional drug design approaches based on structural considerations encounter substantial challenges when applied to IDPs. Consequently, the pursuit of therapeutic interventions directed towards IDPs presents a complex endeavor. While there are indeed existing methodologies for targeting IDPs, they are encumbered by noteworthy constrains. Hence, there exists an imminent imperative to investigate more efficacious and universally applicable strategies for modulating IDPs. Here, we present an overview of the latest advancements in the research pertaining to IDPs, along with the indirect regulation approach involving the modulation of IDP degradation through proteasome. By comprehending these advancements in research, novel insights can be generated to facilitate the development of new drugs targeted at addressing the accumulation of IDPs in diverse pathological conditions.

Recent advances and future perspectives of noncompetitive proteasome inhibitors.

The proteasome regulates intracellular processes, maintains biological homeostasis, and has shown great significance in the study of various diseases, such as neurodegenerative diseases, immune-related diseases, and cancer, especially in hematologic malignancies such as multiple myeloma (MM) and mantle cell lymphoma (MCL). All clinically used proteasome inhibitors bind to the active site of the proteasome and thus exhibit a competitive mechanism. The development of resistance and intolerance during treatment drives the search for inhibitors with different mechanisms of action. In this review, we provide an overview of noncompetitive proteasome inhibitors, including their mechanisms of action, function, possible applications, and their advantages and disadvantages compared with competitive inhibitors.

pH-Responsive and liver-targeting drug delivery system for combination delivery of artesunate with arsenic trioxide prodrug against hepatocellular carcinoma.

OBJECTIVE: Arsenic trioxide (ATO) exerts therapeutic effects on various solid tumors, and artesunate (ART) synergizes with antitumor drugs. We herein combined ART and an ATO prodrug (ATOP) in pH-responsive and liver-targeting liposomes to improve targeted hepatocellular carcinoma (HCC) treatment. METHODS: 1,2-Distearoyl-sn-glycero-3-phosphoethanolamine (DSPE)-hydrazone (HYD)-polyethylene glycol (PEG)-glycyrrhetinic acid (GA) (DSPE-HYD-PEG-GA) was synthesized and characterized. The optimal ratio of ART and ATOP was selected. Calcium arsenate nanoparticles (CaAs NPs) and DSPE-HYD-PEG-GA@ART/CaAs NPs liposomes were prepared and their physicochemical properties were characterized. Their intracellular uptake, intracellular localization, uptake pathway identification, cytotoxicity, proapoptotic effects, and relevant mechanisms were studied. RESULTS: The DSPE-HYD-PEG-GA was successfully synthesized. The best ratio of ART and ATOP was 7:1. The particle size of CaAs NPs under transmission electron microscopy was 142.39 ± 21.50 nm. Arsenic (As), calcium, and oxygen elements were uniformly distributed in CaAs NPs, and the drug loading and encapsulation efficiency of As are 37.28% and 51.40%, respectively. The liposomes were elliptical, and the particle size was 100.91 ± 39.31 nm. The liposome cell intake was significantly increased in Huh-7 cells. The liposomes entered the cell through macropinocytosis and caveolin-mediated endocytosis and were predominantly distributed in the cytoplasm. They exerted an excellent inhibitory effect on Huh-7 cells and promoted tumor cell apoptosis through lipid peroxidation, mitochondrial membrane potential reduction, and cell-cycle blockage. CONCLUSIONS: The pH-responsive and liver-targeting drug delivery system for the combination delivery of ART with ATOP showed promising effects on hepatocellular carcinoma (HCC).

Optimization of piperidine constructed peptidyl derivatives as proteasome inhibitors.

A series of non-covalent piperidine-containing peptidyl derivatives with various substituents at side chains of different residues were designed, synthesized and evaluated as proteasome inhibitors. After proteasome inhibitory evaluations of all the synthesized target compounds, selected ones were tested for their anti-proliferation activities against three multiple myeloma (MM) cell lines. 8 analogues displayed more potent activities than carfilzomib, and the most promising compound 24 showed IC50 values of 0.8 ± 0.2 nM against 20S proteasome and 8.42 ± 0.74 nM, 7.14 ± 0.52 nM, 14.20 ± 1.08 nM for RPMI 8226, NCI-H929 and MM.1S cell lines, respectively. Additionally, mechanisms of anti-cancer activity of representative compound 24 were further investigated. Apoptosis of RPMI-8226 cells were achieved through accumulating polyubiquitin and inducing the cleavage of caspase and PARP. Besides, half-life in rat plasma of compound 24 was prolonged after optimization, which would be helpful for increasing in vivo activities of this series of derivatives. All the studies confirmed that piperidine-containing non-covalent proteasome inhibitors can be potential leads for anti-MM drug development.

Preparation, evaluation, and in vitro cytotoxicity studies of artesunate-loaded glycyrrhetinic acid decorated PEG-PLGA nanoparticles.

OBJECTIVE: The objective of this study was to prepare the liver targeting drug delivery system (TDDS) of artesunate (ART)-loaded polyethylene glycol (PEG)-poly(d,l-lactic-co-glycolic) acid (PLGA) nanoparticles (NPs) modified by glycyrrhetinic acid (GA), and evaluate its in vitro cytotoxicity. SIGNIFICANCE: The GA-PEG-PLGA-ART NPs enhanced the in vitro cytotoxicity on HCC cell lines. The development of GA-PEG-PLGA NPs will greatly push the clinical applications of ART as a novel anticancer drug. METHODS: The NPs were prepared using solvent evaporation method, and the formulation was optimized through an orthogonal design. In addition, physical properties were determined, including particle size, polydispersity index (PDI), zeta potential (ZP), morphology, drug loading capacity (LC) and encapsulation efficiency (EE), and in vitro drug release. Moreover, the in vitro cytotoxicity of NPs with three human cancer cell lines viz. HepG2, Hep3B, and SMCC-7721 was conducted using the SRB assay. Additionally, lyophilization was conducted to improve the long-term physical stability. RESULTS: The GA-PEG-PLGA-ART NPs have spherical shape, small particle size (around 88 nm) with a narrow size distribution (PDI < 0.3), high drug LC (up to 59.3 ± 1.65%), and high EE (up to 73.13 ± 5.17%). In vitro drug release behavior showed that drugs were released from NPs in a sustained and controlled release pattern. Cytotoxicity study indicated the NPs achieved lower cancer cell survival fraction. The GA-PEG-PLGA NPs freeze-dried with 3% (w/v) of mannitol showed better effect on long-term physical stability. CONCLUSION: The GA-PEG-PLGA-ART NPs appear as a potential liver targeted intracellular delivery platform for ART.

Protective effect of apigenin magnesium complex on H2O2-induced oxidative stress and inflammatory responses in rat hepatic stellate cells.

Context: Apigenin displays antioxidant and anti-inflammatory effects. However, effects of apigenin magnesium (AM) complex on these aspects remain unknown.Objective: This study investigated the effects of AM complex on oxidative stress and inflammatory responses in hydrogen peroxide (H2O2)-induced rat hepatic stellate cells (HSCs).Materials and methods: The antioxidant and anti-inflammatory effects of AM complex at concentrations of 0.625, 1.25, and 2.5 mg/mL were evaluated, comparing to HSCs treated by H2O2 alone. Cell viability, reactive oxygen species (ROS), the activity of superoxide dismutase (SOD), glutathione (GSH), malondialdehyde (MDA), nitric oxide (NO), interleukin 6 (IL-6), and nuclear factor-kappa B (NF-κB) levels were measured. Moreover, cell apoptosis, mRNA expression levels of transforming growth factor-ß (TGF-ß), NF-κB, and inducible nitric oxide synthase (iNOS) were assessed.Results: AM complex significantly inhibited oxidative stress and inflammatory response at concentrations of 0.625, 1.25, and 2.5 mg/mL (IC50 = 1.679 mg/mL). AM complex elevated the survival rate of H2O2-treated HSCs and had no toxic effects on HSCs. AM complex also promoted SOD activity and GSH levels but suppressed ROS, MDA, and NO levels. Additionally, AM complex decreased IL-6 and NF-κB levels, gene expression of TGF-ß, NF-κB, and iNOS, as well as induced apoptosis of HSCs.Discussion and conclusions: Data indicated that AM complex mitigated oxidative stress and inflammatory responses on H2O2-treated HSCs, suggesting that AM complex is a possible candidate for anti-hepatic diseases. Additional efforts, both in vivo and in humans, are required to assess of AM complex as a potential therapeutic drug in liver diseases.

Design, synthesis, and biological evaluation of novel 3-(thiophen-2-ylthio)pyridine derivatives as potential multitarget anticancer agents.

A series of novel 3-(thiophen-2-ylthio)pyridine derivatives as insulin-like growth factor 1 receptor (IGF-1R) inhibitors was designed and synthesized. IGF-1R kinase inhibitory activities and cytotoxicities against HepG2 and WSU-DLCL2 cell lines were tested. For all of these compounds, potent cancer cell proliferation inhibitory activities were observed, but not through the inhibition of IGR-1R. Selected compounds were further screened against various kinases. Typical compound 22 (50% inhibitory concentration [IC50 ] values, HepG2: 2.98 ± 1.11 µM and WSU-DLCL2: 4.34 ± 0.84 µM) exhibited good inhibitory activities against fibroblast growth factor receptor-2 (FGFR2), FGFR3, epidermal growth factor receptor, Janus kinase, and RON (receptor originated from Nantes), with IC50 values ranging from 2.14 to 12.20 µM. Additionally, the cell-cycle analysis showed that compound 22 could arrest HepG2 cells in the G1/G0 phase. Taken together, all the experiments confirmed that the compounds in this series were multitarget anticancer agents worth further optimizing.

Synthesis and evaluation of a series of pyridine and pyrimidine derivatives as type II c-Met inhibitors.

In this study, a series of novel pyridine and pyrimidine-containing derivatives were designed, synthesized and biologically evaluated for their c-Met inhibitory activities. In the biological evaluation, half of the target compounds exhibited moderate to potent c-Met inhibitory activities. Among which, it is noteworthy that compounds 13d not only showed most potent c-Met inhibitory potency but also displayed excellent anti-proliferative activity (IC50=127nM against EBC-1 cell line) as well as an acceptable kinase selectivity profile. Moreover, the western blot assay indicated that 13d inhibited c-Met phosphorylation in EBC-1 cells in a dose-dependent manner, with complete abolishment at 0.1mM. All these experimental results suggested that 13d could be served as a promising lead compound for the development of anticancer agents.

Design, synthesis and biological evaluation of novel non-covalent piperidine-containing peptidyl proteasome inhibitors.

A series of novel non-covalent piperidine-containing dipeptidyl derivatives were designed, synthesized and evaluated as proteasome inhibitors. All target compounds were tested for their proteasome chymotrypsin-like inhibitory activities, and selected derivatives were evaluated for the anti-proliferation activities against two multiple myeloma (MM) cell lines RPMI 8226 and MM-1S. Among all of these compounds, eight exhibited significant proteasome inhibitory activities with IC50 less than 20nM, and four are more potent than the positive control Carfilzomib. Compound 28 displayed the most potent proteasome inhibitory activity (IC50: 1.4±0.1nM) and cytotoxicities with IC50 values at 13.9±1.8nM and 9.5±0.5nM against RPMI 8226 and MM-1S, respectively. Additionally, the ex vivo blood cell proteasome inhibitory activities of compounds 24 and 27-29 demonstrated that the enzymatic metabolism in the whole blood could be well tolerated. All these experiments confirmed that the piperidine-containing non-covalent proteasome inhibitors are potential leads for exploring new anti-cancer drugs.

[Effect of apigenin on protein expressions of PPARs in liver tissues of rats with nonalcoholic steatohepatitis].

OBJECTIVE: To investigate the effect of apigenin on the protein expression levels of peroxisome proliferator-activated receptors (PPARs) in liver tissues of rats with nonalcoholic steatohepatitis (NASH). METHODS: The NASH rat model was established by feeding of a high-fat diet. Unmodeled rats served as the normal controls. The modeled rats were divided into a model control group, Essentiale treatment grouP(300 mg/kg/day),and three apigenin treatment groups for low-dose (15 mg/kg/day), moderate-dose (30 mg/kg/day) and high-dose (60 mg/kg/day). After 13 weeks of treatment,changes in insulin sensitivity from pre-treatment baseline were assessed by measuring the alanine aminotransferase (ALT), aspartate aminotransferase (AST),total cholesterol (TC),triglycerides (TG),low-density and high-density lipoprotein cholesterol (LDL-C and HDL-C),fasting blood glucose (FBG) and fasting insulin (FINS).The liver index and HOMA-IR were also calculated.Protein and gene expression of PPARα and PPARgamma in liver tissue were assessed by immunohistochemistry and RT-PCR.Statistical analysis was performed by the LSD test and Games-Howell test. RESULTS: The apigenin-treated groups showed a significantly greater change in insulin sensitivity than the untreated model group,with the most significant change occurring in the high-dose grouP(P less than 0.05).Compared with the untreated model group,the apigenin-treated groups showed lower levels of ALT (95.4+/-7.3),AST (183.7+/-14.3),TC (1.61+/-0.25),TG (1.23+/-0.21),LDL-C (1.86+/-0.14),FBG (5.29+/-1.45) and FINS (0.76+/-0.86),but a higher level of HDL-C (1.04+/-0.17); again,the high-dose group showed the greatest change (all P less than 0.05).Compared to the untreated model group,the apigenin-treated groups showed significantly lower liver index (3.75+/-0.25 vs.2.90+/-0.17) and HOMA-IR (1.34+/-0.06 vs.0.18+/-0.04),with the high-dose group showing the greatest change (both P less than 0.05). Compared to the untreated model group,the apigenin-treated groups showed higher levels of protein and mRNA of PPARα (18.27+/-4.05 and 0.63+/-0.02,respectively) and PPARgamma(8.48+/-5.05 and 0.39+/-0.02),with the high-dose group showing the greatest change (all P < 0.05). CONCLUSION: Apigenin can improve glucose tolerance,lipid metabolism and insulin resistance while decreasing blood levels of TC,TG,LDL-C,FBG,FINS and HOMA-IR,and increasing HDL-C in NASH,as shown in a high-fat diet induced rat model, and may have therapeutic potential.