From their discovery in the first half of the 20th century, lysosomal cysteine proteases have come a long way: from being the enzymes non‐selectively degrading proteins in lysosomes to being those ...responsible for a number of important cellular processes. Some of the features and roles of their structures, specificity, regulation and physiology are discussed.
Cathepsin B (CtsB) is a lysosomal cysteine proteinase that is specifically translocated to the extracellular milieu during cancer progression. The development of a lipidated CtsB inhibitor ...incorporated into the envelope of a liposomal nanocarrier (LNC‐NS‐629) is described. Ex vivo and in vivo studies confirmed selective targeting and internalization of LNC‐NS‐629 by tumor and stromal cells, thus validating CtsB targeting as a highly promising approach to cancer diagnosis and treatment.
On target: A lipidated cathepsin B (CtsB) inhibitor was incorporated into the envelope of a liposomal nanocarrier. The resulting CtsB‐targeted drug delivery system, which can be loaded with diagnostic or therapeutic agents, was selectively internalized by tumor and stromal cells, thus validating CtsB targeting as a promising approach to cancer diagnosis and treatment.
Summary
The potential link between the microbial dynamics and the environmental parameters was investigated in a semi‐enclosed and highly dynamic coastal system (Gulf of Trieste, northern Adriatic ...Sea, NE Mediterranean Sea). Our comprehensive 2‐year time‐series study showed that despite the shallowness of this area, there was a significant difference between the surface and the bottom bacterial community structure. The bottom bacterial community was more diverse than the surface one and influenced by sediment re‐suspension. The surface seawater temperature had a profound effect on bacterial productivity, while the bacterial community structure was more affected by freshwater‐borne nutrients and phytoplankton blooms. Phytoplankton blooms caused an increase of Gammaproteobacteria (Alteromonadaceae, SAR86 and Vibrionaceae) and shift in dominance from SAR11 to Rhodobacteraceae taxon at the surface. Our results propose the importance of the water mass movements as drivers of freshwater‐borne nutrients and of allochthonous microbial taxa. This study emphasizes the prediction power based on association networks analyses that are fed with long‐term measurements of microbial and environmental parameters. These interaction maps offer valuable insights into the response of marine ecosystem to climate‐ and anthropogenic‐driven stressors.
Ballast water discharges may cause negative impacts to aquatic ecosystems, human health and economic activities by the introduction of potentially harmful species. Fifty untreated ballast water ...tanks, ten in each port, were sampled in four Adriatic Italian ports and one Slovenian port. Salinity, temperature and fluorescence were measured on board. Faecal indicator bacteria (FIB), phyto- and zooplankton were qualitatively and quantitatively determined to identify the species assemblage arriving in ballast water. FIB exceeded the convention standard limits in 12% of the sampled tanks. Vibrio cholerae was not detected. The number of viable organisms in the size groups (minimum dimension) <50 and ≥10 μm and ≥50 μm resulted above the abundances required from the Ballast Water Management Convention in 55 and 86% of the samples, respectively. This is not surprising as unmanaged ballast waters were sampled. Some potentially toxic and non-indigenous species were observed in both phyto- and zooplankton assemblages.
•Ballast water sampling was carried out on 50 ships in five Adriatic ports.•Vibrio cholerae was not detected.•Viable organisms were detected in >90% of ballast tanks.•Six potentially harmful and one non-indigenous phytoplankton species were identified.•Six non-indigenous zooplankton species were observed.
Lysosomal cysteine cathepsins contribute to proteolytic events promoting tumor growth and metastasis. Their enzymatic activity, however, is tightly regulated by endogenous inhibitors. To investigate ...the role of cathepsin inhibitor stefin B (Stfb) in mammary cancer, Stfb null mice were crossed with transgenic polyoma virus middle T oncogene (PyMT) breast cancer mice. We show that ablation of Stfb resulted in reduced size of mammary tumors but did not affect their rate of metastasis. Importantly, decrease in tumor growth was correlated with an increased incidence of dead cell islands detected in tumors of Stfb-deficient mice. Ex vivo analysis of primary PyMT tumor cells revealed no significant effects of ablation of Stfb expression on proliferation, angiogenesis, migration and spontaneous cell death as compared with control cells. However, upon treatment with the lysosomotropic agent Leu-Leu-OMe, cancer cells lacking Stfb exhibited a significantly higher sensitivity to apoptosis. Moreover, Stfb-ablated tumor cells were significantly more prone to cell death under increased oxidative stress. These results indicate an in vivo role for Stfb in protecting cancer cells by promoting their resistance to oxidative stress and to apoptosis induced through the lysosomal pathway.
A sigma-2 receptor agonist siramesine has been shown to trigger cell death of cancer cells and to exhibit a potent anticancer activity in vivo. However, its mechanism of action is still poorly ...understood. We show that siramesine can induce rapid cell death in a number of cell lines at concentrations above 20 μM. In HaCaT cells, cell death was accompanied by caspase activation, rapid loss of mitochondrial membrane potential (MMP), cytochrome c release, cardiolipin peroxidation and typical apoptotic morphology, whereas in U-87MG cells most apoptotic hallmarks were not notable, although MMP was rapidly lost. In contrast to the rapid loss of MMP above 20 μM siramesine, a rapid increase in lysosomal pH was observed at all concentrations tested (5-40 μM); however, it was not accompanied by lysosomal membrane permeabilisation (LMP) and the release of lysosomal enzymes into the cytosol. Increased lysosomal pH reduced the lysosomal degradation potential as indicated by the accumulation of immature forms of cysteine cathepsins. The lipophilic antioxidant α-tocopherol, but not the hydrophilic antioxidant N-acetyl-cysteine, considerably reduced cell death and destabilisation of mitochondrial membranes, but did not prevent the increase in lysosomal pH. At concentrations below 15 μM, siramesine triggered cell death after 2 days or later, which seems to be associated with a general metabolic and energy imbalance due to defects in the endocytic pathway, intracellular trafficking and energy production, and not by a specific molecular event. Overall, we show that cell death in siramesine-treated cells is induced by destabilisation of mitochondria and is independent of LMP and the release of cathepsins into the cytosol. Moreover, it is unlikely that siramesine acts exclusively through sigma-2 receptors, but rather through multiple molecular targets inside the cell. Our findings are therefore of significant importance in designing the next generation of siramesine analogues with high anticancer potential.
We report here that a number of commonly used small peptide caspase inhibitors consisting of a caspase recognition sequence linked to chloromethylketone, fluoromethylketone or aldehyde reactive group ...efficiently inhibit other cysteine proteases than caspases. The in vitro studies included cathepsins B, H, L, S, K, F, V, X and C, papain and legumain. Z-DEVD-cmk was shown to be the preferred irreversible inhibitor of most of the cathepsins in vitro, followed by Z-DEVD-fmk, Ac-YVAD-cmk, Z-YVAD-fmk and Z-VAD-fmk. Inactivation of legumain by all the inhibitors investigated was moderate, whereas cathepsins H and C were poorly inhibited or not inhibited at all. Inhibition by aldehydes was not very potent. All the three fluoromethylketones efficiently inhibited cathepsins in Jurkat and human embryonic kidney 293 cells at concentrations of 100 microM. Furthermore, they completely inhibited cathepsins B and X activity in tissue extracts at concentrations as low as 1 microM. These results suggest that data based on the use of these inhibitors should be taken with caution and that other proteases may be implicated in the processes previously ascribed solely to caspases.
It is more than 50years since the lysosome was discovered. Since then its hydrolytic machinery, including proteases and other hydrolases, has been fairly well identified and characterized. Among ...these are the cysteine cathepsins, members of the family of papain-like cysteine proteases. They have unique reactive-site properties and an uneven tissue-specific expression pattern. In living organisms their activity is a delicate balance of expression, targeting, zymogen activation, inhibition by protein inhibitors and degradation. The specificity of their substrate binding sites, small-molecule inhibitor repertoire and crystal structures are providing new tools for research and development. Their unique reactive-site properties have made it possible to confine the targets simply by the use of appropriate reactive groups. The epoxysuccinyls still dominate the field, but now nitriles seem to be the most appropriate “warhead”. The view of cysteine cathepsins as lysosomal proteases is changing as there is now clear evidence of their localization in other cellular compartments. Besides being involved in protein turnover, they build an important part of the endosomal antigen presentation. Together with the growing number of non-endosomal roles of cysteine cathepsins is growing also the knowledge of their involvement in diseases such as cancer and rheumatoid arthritis, among others. Finally, cysteine cathepsins are important regulators and signaling molecules of an unimaginable number of biological processes. The current challenge is to identify their endogenous substrates, in order to gain an insight into the mechanisms of substrate degradation and processing. In this review, some of the remarkable advances that have taken place in the past decade are presented. This article is part of a Special Issue entitled: Proteolysis 50years after the discovery of lysosome.
► Current advances in the field of cysteine cathepsins and their regulation. ► Cysteine cathepsin activity as a delicate balance of various factors. ► Structure of cysteine cathepsins and their mechanism of interaction with inhibitors. ► Inhibition of cysteine cathepsins by protein and small-molecule inhibitors. ► The increased expression of cysteine cathepsins implicated in various diseases.
Results In pregnancies with available outcomes, the most frequently FDA-X prescribed drugs were oral contraceptives (53%), followed by paroxetin (21%), medroxiprogesterone (8%), retinoids (7%), ...warfarin (4%), methotrexate, statins, ribavirin, ulipristal-acetate, and radioactive iodine.
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