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In vitro testing of Aspergillus fumigatus susceptibility to echinocandins has always been a challenge. Using a simple and quick colorimetric method to analyze the activity of inhibitors of β-1,3-glucan synthesis, we found that the composition of the culture medium significantly influences glucan synthesis and consequently the antifungal properties of inhibitors of β-1,3-glucan synthesis when they are tested alone or in combination with chitin synthase inhibitors.

International audience; In this study, we report a novel mechanism of action for a cytotoxic derivative of betulinic acid (BA). B10 is a semi-synthetic glycosylated derivative of BA selected for its enhanced cytotoxic activity. Interestingly, although B10 induces apoptosis, caspase-3 downregulation incompletely prevents B10-induced cell death, Bcl-2 overexpression fails to protect cells and DNA fragmentation rates do not reflect cell death rates in contrast to cytoplasmic membrane permeabilization. These results implicate that apoptotic and non-apoptotic cell death coexist upon B10 treatment. Unexpectedly, we found that B10 induces autophagy and also abrogates the autophagic flux. B10 destabilizes lysosomes as shown by Lysotracker Red staining and by cathepsin Z and B release from lysosomes into the cytoplasm. Consistently, the cathepsin inhibitor Ca074Me significantly decreases B10-induced cell death, further supporting the fact that the release of lysosomal enzymes contributes to B10-triggered cell death. Downregulation of ATG7, ATG5 or BECN1 by RNAi significantly decreases caspase-3 activation, lysosomal permeabilization and cell death. Thus, by concomitant induction of autophagy and inhibition of the autophagic flux, B10 turns autophagy into a cell death mechanism. These findings have important implications for the therapeutic exploitation of BA derivatives, particularly in apoptosis-resistant cancers.

Aminopeptidase N (APN)/CD13 (EC 3.4.11.2) is a transmembrane protease present in a wide variety of human tissues and cell types (endothelial, epithelial, fibroblast, leukocyte). APN/CD13 expression is dysregulated in inflammatory diseases and in cancers (solid and hematologic tumors). APN/CD13 serves as a receptor for coronaviruses. Natural and synthetic inhibitors of APN activity have been characterized. These inhibitors have revealed that APN is able to modulate bioactive peptide responses (pain management, vasopressin release) and to influence immune functions and major biological events (cell proliferation, secretion, invasion, angiogenesis). Therefore, inhibition of APN/CD13 may lead to the development of anti‐cancer and anti‐inflammatory drugs. This review provides an update on the biological and pharmacological profiles of known natural and synthetic APN inhibitors. Current status on their potential use as therapeutic agents is discussed with regard to toxicity and specificity. © 2005 Wiley Periodicals, Inc. Med Res Rev

International audience; Bevirimat (2), the first-in-class HIV-1 maturation inhibitor, shows a low efficacy due essentially to the natural polymorphism of its target, the CA-SP1 junction. Moreover, its low hydrosolubility makes it difficult to study its interaction with the CA-SP1 junction. We have synthesized new derivatives of bevirimat by adding different hydrophilic substituents at the C-28 position to improve their hydrosolubility and perform the structural study of a complex by NMR. Synthesis of the new derivatives, the effect of substituents at the C-28 position and their hydrosolubility are discussed. The ability of these molecules to inhibit viral infection and their cytotoxicity is assessed. Compared to the well-known bevirimat (2), one of our compounds (16) shows a higher hydrosolubility associated with a 2.5 fold increase in activity, a higher selectivity index and a better antiviral profile. Moreover, for the first time a direct interaction between a derivative of bevirimat (16) and the domain CA-SP1eNC is shown by NMR. Information from this study should allow us to decipher the mechanism by which bevirimat inhibits HIV-1 maturation and how the natural polymorphism of the spacer peptide SP1 triggers resistance to inhibitors.

Background Because some of heat shock protein 90's (HSP90) clients are key cell cycle regulators, HSP90 inhibition can affect the cell cycle. Recently, celastrol is identified both as a novel inhibitor of HSP90 and as a potential anti-tumor agent. However, this agent's effects on the cell cycle are rarely investigated. In this study, we observed the effects of celastrol on the human monocytic leukemia cell line U937 cell cycle. Results Celastrol affected the proliferation of U937 in a dose-dependent way, arresting the cell cycle at G0/G1 with 400 nM doses and triggering cell death with doses above 1000 nM. Cell cycle arrest was accompanied by inhibition of HSP90 ATPase activity and elevation in HSP70 levels (a biochemical hallmark of HSP90 inhibition), a reduction in Cyclin D1, Cdk4 and Cdk6 levels, and a disruption of the HSP90/Cdc37/Cdk4 complex. The observed effects of celastrol on the U937 cell cycle were thiol-related, firstly because the effects could be countered by pre-loading thiol-containing agents and secondly because celastrol and thiol-containing agents could react with each other to form new compounds. Conclusions Our results disclose a novel action of celastrol-- causing cell cycle arrest at G0/G1 phase based upon thiol-related HSP90 inhibition. Our work suggests celastrol's potential in tumor and monocyte-related disease management.

Cell adhesion molecules (CAM) expressed by vascular endothelium in response to cytokine stimulation play a key role in leukocyte adhesion to endothelium during the inflammatory response. Tripterine, a chemical compound of the Chinese plant Tripterygium wilfordii Hook f, displays anti-inflammatory properties in several animal models. However, mechanisms of its action are poorly understood. In the present study, we show that in inflammatory conditions, mimicked by tumor necrosis factor α (TNF-α) stimulation, pretreatment for 6 h with tripterine at nontoxic concentrations of 20–200 nM inhibits the expression of E-selectin, vascular cell adhesion molecule (CAM)-1 (VCAM-1), and intercellular adhesion molecule-1 (ICAM-1) in human umbilical vein endothelial cells (HUVEC) in a dose-dependent manner. Tripterine (200 nM) almost completely inhibits expression of VCAM-1 [50% inhibitory concentration (IC50)=52 nM] and ICAM-1 (IC50=51 nM) and 73% of E-selectin (IC50=94 nM). This inhibition effect is prominent, compared with that of dexamethasone, ibuprofen, methotrexate, or probucol, which revealed a much weaker inhibition at doses as high as 1 mM. Effects on endothelial CAM of other proinflammatory cytokines, such as interleukin-1β and interferon-γ, were also inhibited significantly by tripterine. Moreover, significant inhibition was equally observable in postincubation experiments. In addition, tripterine inhibited adhesion of human monocytes and T lymphocytes to TNF-α-stimulated HUVEC. Finally, tripterine inhibited TNF-α-driven CAM mRNA transcription and nuclear factor-κB nuclear (NF-κB) translocation. Hence, we describe a new mechanism of tripterine’s anti-inflammatory action obtained at nanomolar concentrations, owing to the negative regulation of cytokine-induced adhesion molecule expression and adhesiveness in human endothelium.