Rosa Lemmens-Gruber

Cyclodepsipeptides from Marine Sponges: Natural Agents for Drug Research

A number of natural products from marine sponges, such as cyclodepsipeptides, have been identified. The structural characteristics of this family of cyclic peptides include various unusual amino acid residues and unique N-terminal polyketide-derived moieties. Papuamides are representatives of a class of marine sponge derived cyclic depsipeptides, including callipeltin A, celebesides A and B, homophymine A, mirabamides, microspinosamide, neamphamide A and theopapuamides. They are thought to have cytoprotective activity against HIV-1 in vitro by inhibiting viral entry. Jasplakinolide, a representative member of marine sponge-derived cyclodepsipeptides that include arenastatin A, geodiamolides, homophymines, spongidepsin and theopapuamides, is a potent inducer of actin polymerization in vitro. Although actin dynamics is essential for tumor metasasis, no actin targeting drugs have been used in clinical trials due to their severe cytotoxicity. Nonetheless, the actin cytoskeleton remains a potential target for anti-cancer drug development. These features imply the use of cyclodepsipeptides as molecular models in drug research.

TLR 2 and CD14 Mediate Innate Immunity and Lung Inflammation to Staphylococcal Panton–Valentine Leukocidin In Vivo

The pore-forming toxin Panton–Valentine leukocidin (PVL) is carried by community-acquired methicillin-resistant Staphylococcus aureus and associated with necrotizing pneumonia together with poor prognosis of infected patients. Although the cell-death–inducing properties of PVL have previously been examined, the pulmonary immune response to PVL is largely unknown. Using an unbiased transcriptional profiling approach, we show that PVL induces only 29 genes in mouse alveolar macrophages, which are associated with TLR signaling. Further studies indicate that PVL directly binds to TLR2 and induces immune responses via NF-κB in a TLR2, CD14, MyD88, IL-1R–associated kinase 1, and TNFR-associated factor 6-dependent manner. PVL-mediated inflammation is independent of pore formation but strongly depends on the LukS subunit and is suppressed in CD14/TLR2−/− cells. In vivo PVL or LukS induced a robust inflammatory response in lungs, which was diminished in CD14/TLR2−/− mice. These results highlight the proinflammatory properties of PVL and identify CD14/TLR2 as an essential receptor complex for PVL-induced lung inflammation.

Cyclodepsipeptides - Potential Drugs and Lead Compounds in the Drug Development Process

Cyclodepsipeptides show an interesting spectrum of biological activity. Members of this new class of potential drugs may also serve as lead compounds for more pharmacologically potent and toxicologically safe derivatives. Some of these natural products and (semi-)synthetic derivatives have already been evaluated in clinical trials. A common feature of cyclodepsipeptides is their ionophoric properties. However, their pharmacologically relevant action does not seem to correlate with this feature; rather it is based on interactions with distinct cellular compartments and signal transduction pathways. Cyclodepsipeptides, which are currently being evaluated in clinical trials, are used in refractory cancer therapy, usually in combination with other cytotoxic drugs. A series of cyclooctadepsipeptides, however, shows a completely different spectrum of biological activity, namely, potent anthelmintic properties. A number of cyclodepsipeptides have been well characterized in vitro and in vivo, and interesting modes of action, such as antiplasmodial, antiviral, insecticidal, cytotoxic, and antiproliferative properties have been observed. Whether these natural products will be of benefit for patients must be evaluated in clinical trials. Recently, a number of cyclodepsipeptides from marine sponges, bacteria and fungi have been identified. Subsequent structural determination revealed unique structural features within some of these compounds. It was suggested that the cyclic depsipeptide structure is important for the biological activity because the linear homologues were inactive. The scope of activity of these newly isolated natural products spans a range from cytoprotective activity against HIV-1 infection, growth inhibitory effects toward cancer cells, and antimycobacterial, and antimalarial activity.

Sex-Specific differences in metabolic control, cardiovascular risk, and interventions in patients with type 2 diabetes mellitus

BACKGROUND Sex-specific differences appear particularly relevant in the management of type 2 diabetes mellitus (T2DM), with women experiencing greater increases in cardiovascular morbidity and mortality than do men. OBJECTIVE The aim of this article was to investigate the influence of biological sex on clinical care and microvascular and macrovascular complications in patients with T2DM in a Central European university diabetes clinic. METHODS In a cross-sectional study, sex-specific disparities in metabolic control, cardiovascular risk factors, and diabetic complications, as well as concomitant medication use and adherence to treatment recommendations, were evaluated in 350 consecutive patients who were comparable for age, diabetes duration, and body mass index. Study inclusion criteria included age ≤75 years, T2DM, a documented history of presence or absence of coronary heart disease (CHD), and informed consent. Patients were followed in the diabetes outpatient clinic between November 2007 and March 2008. RESULTS Two hundred and one patients with T2DM met inclusion criteria (93 [46.3%] women, 108 [53.7%] men). Women with T2DM had higher mean (SE) systolic blood pressure (155.4 [22.5] vs 141.0 [19.8] mm Hg for men; P < 0.001) and total cholesterol (TC) (5.28 [1.34] vs 4.86 [1.29] mmol/L for men; P < 0.05), but a lower TC:HDL-C ratio (4.1 [1.19] vs 4.5 [1.2] for men; P < 0.05). Slightly more men (32.4%) than women (26.9%) reached the therapeutic goal of <7.0% for glycosylated hemoglobin. Women with shorter diabetes duration (<10 years) received oral antihyperglycemic therapy less frequently (P < 0.05). Women with longer disease duration had hypertension more frequently than did their male counterparts (100% vs 86.0%, respectively; P < 0.01). Despite a similar rate of CHD, men were twice as likely as women to have had coronary interventions (percutaneous transluminal coronary angioplasty/coronary artery bypass graft, 25.0% vs 12.9%, respectively; P < 0.05). Women with CHD also had a higher rate of cerebral ischemia than did men (27.6% vs 5.4%, respectively; P < 0.05) and received aspirin less frequently for secondary prevention (P < 0.001). Men had greater overall adherence to diabetes and cardiovascular risk guidelines than did women (66.4% vs 58.9%, respectively; P < 0.01). CONCLUSIONS In this study of diabetes clinic outpatients, women with T2DM had a worse cardiovascular risk profile and achieved therapeutic goals less frequently than did men. Treatment strategies should be improved in both sexes, but women with diabetes may be in need of more aggressive treatment, especially when cardiovascular disease is present.

Clinical pharmacy activities in chronic kidney disease and end-stage renal disease patients: a systematic literature review

BackgroundChronic kidney disease (CKD) and end-stage renal disease (ESRD) represent worldwide health problems with an epidemic extent. Therefore, attention must be given to the optimisation of patient care, as gaps in the care of CKD and ESRD patients are well documented. As part of a multidisciplinary patient care strategy, clinical pharmacy services have led to improvements in patient care. The purpose of this study was to summarise the available evidence regarding the role and impact of clinical pharmacy services for these patient populations.MethodsA literature search was conducted using the Medline, Embase and International Pharmaceutical Abstracts databases to identify relevant studies on the impact of clinical pharmacists on CKD and ESRD patients, regarding disease-oriented and patient-oriented outcomes, and clinical pharmacist interventions on drug-related problems.ResultsAmong a total of 21 studies, only four (19%) were controlled trials. The majority of studies were descriptive (67%) and before-after studies (14%). Interventions comprised general clinical pharmacy services with a focus on detecting, resolving and preventing drug-related problems, clinical pharmacy services with a focus on disease management, or clinical pharmacy services with a focus on patient education in order to increase medication knowledge. Anaemia was the most common comorbidity managed by clinical pharmacists, and their involvement led to significant improvement in investigated disease-oriented outcomes, for example, haemoglobin levels. Only four of the studies (including three controlled trials) presented data on patient-oriented outcomes, for example, quality of life and length of hospitalisation. Studies investigating the number and type of clinical pharmacist interventions and physician acceptance rates reported a mean acceptance rate of 79%. The most common reported drug-related problems were incorrect dosing, the need for additional pharmacotherapy, and medical record discrepancies.ConclusionsFew high-quality trials addressing the benefit and impact of clinical pharmacy services in CKD and ESRD patients have been published. However, all available studies reported some positive impact resulting from clinical pharmacist involvement, including various investigated outcome measures that could be improved. Additional randomised controlled trials investigating patient-oriented outcomes are needed to further determine the role of clinical pharmacists and the benefits of clinical pharmacy services to CKD and ESRD patients.

Interactions between ABC‐transport proteins and the secondary Fusarium metabolites enniatin and beauvericin

Enniatins (ENN) and beauvericin (BEA) exert cytotoxic properties. Here, we observed that their impact on Ca(2+)-homeostasis can be reversed by exogenous ATP. Thus, we investigated whether membrane-located ATP-binding cassette (ABC) transporters influence ENNs- and BEA-induced cytotoxicity. In short-term exposure assays breast cancer resistance protein (ABCG2)-overexpression weakly but significantly reduced the cytotoxic activity of BEA but not ENNs. In contrast, multidrug resistance-associated protein-1 (ABCC1)- and P-glycoprotein (ABCB1)-overexpression was not protective under identical conditions. ABCG2-mediated resistance against BEA was reversible by ABCG2 modulators. In long-term exposure assays, ABCG2 and ABCB1 significantly protected against ENNs- and to a lesser extent BEA-induced cytotoxicity. Moreover, both fusariotoxins potently inhibited the ABCG2- and ABCB1-mediated efflux of specific fluorescent substrates, with BEA being more effective. Additionally, ATPase and photoaffinity-labelling assays proofed interaction of both substances with ABCG2 and ABCB1. Remarkably, 2 years selection of KB-3-1 cells against both fusariotoxins resulted only in two-fold ENNs but negligible BEA resistance. Interestingly, the selected sublines displayed upregulation of multidrug resistance proteins and crossresistance to other chemotherapeutics. Summarizing, ABCG2 and ABCB1 slightly but significantly protect human cells against ENNs- and BEA-induced cytotoxicity. However, both mycotoxins potently interact with ABCB1 and ABCG2 transport functions suggesting influences on bioavailability of xenobiotics and pharmaceuticals.

Voltage-Gated Sodium Channels: Mutations, Channelopathies and Targets

Voltage-gated sodium channels produce fast depolarization, which is responsible for the rising phase of the action potential in neurons, muscles and heart. These channels are very large membrane proteins and are encoded by ten genes in mammals. Sodium channels are a crucial component of excitable tissues; hence, they are a target for various neurotoxins that are produced by plants and animals for defence and protection, such as tetrodotoxin, scorpion toxins and batrachotoxin. Several mutations in various sodium channel subtypes cause multiple inherited diseases known as channelopathies. When these mutated sodium channel subtypes are expressed in various tissues, channelopathies in brain, skeletal muscle and cardiac muscle develop as well as neuropathic pain. In this review, we discuss aspects of voltage-gated sodium channel genes with an emphasis on cardiac muscle sodium channels. In addition, we report novel mutations that underlie a spectrum of diseases, such as Brugada, long QT syndrome and inherited conduction disorders. Furthermore, this review explains commonalities and differences among the channel subtypes, the channelopathies caused by the sodium channel gene mutation and the specificity of toxins and blockers of the channel subtypes.

Oxidative stress and DNA interactions are not involved in Enniatin- and Beauvericin-mediated apoptosis induction.

The fusariotoxins beauvericin (BEA) and the structurally related enniatins (ENN) are frequent contaminants of grain-based food and feed. They exert potent cytotoxic activities based on apoptosis induction. Since it is known, that reactive oxygen species (ROS) and DNA damage lead to apoptotic cell death, this study aimed to clarify whether oxidative stress and DNA interactions are involved in ENN- and BEA-induced cytotoxicity. Diverse cellular and molecular assays indicated that oxidative stress does not contribute to ENN- and BEA-induced cytotoxicity. In contrast, both fusariotoxins were shown to exert moderate antioxidative activities. Moreover, only at high concentrations (>100 microM) both mycotoxins were found to intercalate substantially into dsDNA and to inhibit the catalytic activity of topoisomerase I and II. Furthermore, the potent cytotoxic activity of ENN and BEA was shown to be widely independent of cellular mismatch- and nucleotide excision repair pathways. Also the ataxia-telangiectasia mutated (ATM) protein kinase, a well known DNA damage sensor, did not affect BEAs cytotoxic potential while in ENN-induced cytotoxicity ATM had a detectable but not a major modulating influence. Together, our data suggest that ROS and DNA damage are not key factors in ENN- and BEA-mediated cytotoxicity.

IFN-β Increases Listeriolysin O-Induced Membrane Permeabilization and Death of Macrophages

Type I IFN (IFN-I) signaling is detrimental to cells and mice infected with Listeria monocytogenes. In this study, we investigate the impact of IFN-I on the activity of listeriolysin O (LLO), a pore-forming toxin and virulence protein released by L. monocytogenes. Treatment of macrophages with IFN-β increased the ability of sublytic LLO concentrations to cause transient permeability of the plasma membrane. At higher LLO concentrations, IFN-β enhanced the complete breakdown of membrane integrity and cell death. This activity of IFN-β required Stat1. Perturbation of the plasma membrane by LLO resulted in activation of the p38MAPK pathway. IFN-β pretreatment enhanced LLO-mediated signaling through this pathway, consistent with its ability to increase membrane damage. p38MAPK activation in response to LLO was independent of TLR4, a putative LLO receptor, and inhibition of p38MAPK neither enhanced nor prevented LLO-induced death. IFN-β caused cells to express increased amounts of caspase 1 and to produce a detectable caspase 1 cleavage product after LLO treatment. Contrasting recent reports with another pore-forming toxin, this pathway did not aid cell survival as caspase1-deficient cells were equally sensitive to lysis by LLO. Key lipogenesis enzymes were suppressed in IFN-β-treated cells, which may exacerbate the membrane damage caused by LLO.

A novel tumor necrosis factor–mediated mechanism of direct epithelial sodium channel activation

RATIONALE Alveolar liquid clearance is regulated by Na(+) uptake through the apically expressed epithelial sodium channel (ENaC) and basolaterally localized Na(+)-K(+)-ATPase in type II alveolar epithelial cells. Dysfunction of these Na(+) transporters during pulmonary inflammation can contribute to pulmonary edema. OBJECTIVES In this study, we sought to determine the precise mechanism by which the TIP peptide, mimicking the lectin-like domain of tumor necrosis factor (TNF), stimulates Na(+) uptake in a homologous cell system in the presence or absence of the bacterial toxin pneumolysin (PLY). METHODS We used a combined biochemical, electrophysiological, and molecular biological in vitro approach and assessed the physiological relevance of the lectin-like domain of TNF in alveolar liquid clearance in vivo by generating triple-mutant TNF knock-in mice that express a mutant TNF with deficient Na(+) uptake stimulatory activity. MEASUREMENTS AND MAIN RESULTS TIP peptide directly activates ENaC, but not the Na(+)-K(+)-ATPase, upon binding to the carboxy-terminal domain of the α subunit of the channel. In the presence of PLY, a mediator of pneumococcal-induced pulmonary edema, this binding stabilizes the ENaC-PIP2-MARCKS complex, which is necessary for the open probability conformation of the channel and preserves ENaC-α protein expression, by means of blunting the protein kinase C-α pathway. Triple-mutant TNF knock-in mice are more prone than wild-type mice to develop edema with low-dose intratracheal PLY, correlating with reduced pulmonary ENaC-α subunit expression. CONCLUSIONS These results demonstrate a novel TNF-mediated mechanism of direct ENaC activation and indicate a physiological role for the lectin-like domain of TNF in the resolution of alveolar edema during inflammation.