Identification of novel small molecule inhibitors against the NS3/4A protease of hepatitis C virus genotype 4a.

BACKGROUND: Hepatitis C virus (HCV) infection poses a considerable threat to the public health. The current standard of care treatment with pegylated interferon-alpha in combination with ribavirin (PEG-IFN- α+RBV) is associated with significant side effects, poorly tolerated, and provides limited efficacy. The development of direct-acting antiviral agents (DAAs) targeting key viral enzymes essential for viral replication represents a significant milestone in the treatment of chronic HCV infection. Given its critical role in the viral polyprotein processing and the evasion of the host innate immunity, the NS3/4A protease has emerged as a promising drug target for the development of anti-HCV therapies. Although several potent NS3/4A protease inhibitors (PIs) have been approved or are in clinical development, the majority of currently available PIs have significant limitations related to untoward adverse events and a lack of pan-genotypic activity, indicating a continuing unmet medical need for the development and optimization of novel PIs with improved efficacy and tolerability, convenient dosing schedules, and shorter treatment durations. METHODS: The inhibitory efficacy of four computer-designed chemically-synthesized compounds was evaluated against in vitro-expressed NS3/4A protease from HCV genotype 4a, the most prevalent genotype in Egypt, using a fluorescence-based enzymatic assay. RESULTS: We successfully identified two non-macrocyclic small molecules, BE113 (7a) and BE114 (7b), which exhibited inhibitory activity against HCV NS3/4A protease from HCV genotype 4a. CONCLUSION: The two compounds presented in this study may be promising inhibitors against NS3/4A protease of HCV genotype 4a and could be novel lead compounds for developing new therapeutics for the treatment of chronic HCV infection.

P450 3A-Catalyzed O-Dealkylation of Lapatinib Induces Mitochondrial Stress and Activates Nrf2.

Lapatinib (LAP), an oral tyrosine kinase inhibitor for the treatment of metastatic breast cancer, has been associated with idiosyncractic hepatotoxicity. Recent investigations have implicated the importance of P450 3A4/5 enzymes in the formation of an electrophilic quinone imine (LAPQI) metabolite generated through further oxidation of O-dealkylated lapatinib (OD-LAP). In the current study, hepatic stress was observed via mitochondrial impairment. OD-LAP caused a time- and concentration-dependent decrease in oxygen consumption in HepG2 cells, whereas LAP did not alter the oxygen consumption rate. Interestingly, however, HepG2 cells transfected with human P450 3A4 did exhibit mitochondrial dysfunction via P450 3A4-mediated metabolism of LAP to OD-LAP. OD-LAP-induced mitochondrial toxicity was enhanced upon depletion of intracellular GSH levels, demonstrating that cellular GSH levels are important in the protection of mitochondrial function against LAPQI. Given the nature of LAPQI and the importance of GSH levels in LAP-induced mitochondrial stress, the activation of nuclear factor erythroid 2-related factor 2 (Nrf2) was evaluated, as this transcription factor induces the expression of NAD(P)H quinone oxidoreductase 1, glutathione S-transferase, UDP-glucuronosyltransferases, and glutathione synthetase, all of which might be expected to decrease the toxicity of LAP. Using a FRET-based target gene assay in HepG2 cells, OD-LAP was indeed found to activate Nrf2. Follow-up assays showed increased mRNA levels of Nrf2 target genes after a 4 h treatment with OD-LAP but not with LAP. LAP activation of Nrf2 was observed only when HepG2 cells were transduced with P450 3A4. The significance of Nrf2 protection was established in vivo in Nrf2-KO mice. Increased transaminase levels were found after a single LAP dose in both Nrf2-KO and control mice, indicating elevated hepatic necrosis, although transaminase levels reverted to baseline levels in the control mice upon repeat dosing. They continued to rise in Nrf2-KO mice, however, indicating the likelihood that Nrf-2 plays a significant role in combatting the hepatotoxicity triggered by LAP.

Broad Anti-tumor Activity of a Small Molecule that Selectively Targets the Warburg Effect and Lipogenesis.

Malignant cells exhibit aerobic glycolysis (the Warburg effect) and become dependent on de novo lipogenesis, which sustains rapid proliferation and resistance to cellular stress. The nuclear receptor liver-X-receptor (LXR) directly regulates expression of key glycolytic and lipogenic genes. To disrupt these oncogenic metabolism pathways, we designed an LXR inverse agonist SR9243 that induces LXR-corepressor interaction. In cancer cells, SR9243 significantly inhibited the Warburg effect and lipogenesis by reducing glycolytic and lipogenic gene expression. SR9243 induced apoptosis in tumors without inducing weight loss, hepatotoxicity, or inflammation. Our results suggest that LXR inverse agonists may be an effective cancer treatment approach.

Pyrrolizines: Promising scaffolds for anticancer drugs.

Pyrrolizine derivatives constitute a class of heterocyclic compounds which can serve as promising scaffolds for anticancer drugs. The unique antitumor properties of mitomycin C inspired chemists to develop different pyrrolizine systems and assess their potential antitumor activities against a wide variety of cancer types. Here we review the different classes of pyrrolizines that possess anticancer potency, with an emphasis on their structure activity relationships, in an effort to pave the way for further development in this promising area of research.

A liver-selective LXR inverse agonist that suppresses hepatic steatosis.

Fatty liver, which often accompanies obesity and type 2 diabetes, frequently leads to a much more debilitating hepatic disease including non-alcoholic steatohepatitis, cirrhosis, and hepatocellular carcinoma. Current pharmacological therapies lack conclusive efficacy and thus treatment options are limited. Novel therapeutics that suppress either hepatic lipogenesis and/or hepatic inflammation may be useful. Here, we describe the development of the first selective synthetic LXR inverse agonist (SR9238) and demonstrate that this compound effectively suppresses hepatic lipogenesis, inflammation, and hepatic lipid accumulation in a mouse model of non-alcoholic hepatosteatosis. SR9238 displays high potency for both LXRα and LXRß (40-200 nM IC50) and was designed to display liver specificity so as to avoid potential side effects due to suppression of LXR in the periphery. Unexpectedly, treatment of diet-induced obese mice with SR9238 suppressed plasma cholesterol levels. These data indicate that liver-selective LXR inverse agonists may hold utility in the treatment of liver disease.

α-Substitution effects on the ease of S→N-acyl transfer in aminothioesters.

In S-acylcysteines and homocysteines, the efficacy and rate of S→N-acyl transfer (5 and 6 cyclic TSs) vary with the size of S-acyl group. Conformational and quantum chemical calculations indicate that the spatial distance, b(N-C), between the terminal amine and the thioester carbon is shortened by α-C(O)X (X = OH, OMe, NH2 ) substituents.

Selective synthesis and structural elucidation of S-acyl- and N-acylcysteines.

N-(Acyl)-1H-benzotriazoles 6a-f react with l-cysteine 5 at 20 degrees C to give exclusively (i) N-acyl-l-cysteines 8a-e in the presence of triethylamine in CH(3)CN-H(2)O (3:1), but (ii) S-acyl-l-cysteines 7a-e in CH(3)CN-H(2)O (5:1) in the absence of base. Structures 7b, 7d and 8b, 8d are supported by 2D NMR spectroscopic methods including gDQCOSY, gHMQC, gHMBC, and (1)H-(15)N CIGAR-gHMBC experiments. The structure of compound 8d was also supported by single-crystal X-ray diffraction.