A Peptidomimetic Antibiotic Targets Outer Membrane Proteins and Disrupts Selectively the Outer Membrane in Escherichia coli.

Increasing antibacterial resistance presents a major challenge in antibiotic discovery. One attractive target in Gram-negative bacteria is the unique asymmetric outer membrane (OM), which acts as a permeability barrier that protects the cell from external stresses, such as the presence of antibiotics. We describe a novel ß-hairpin macrocyclic peptide JB-95 with potent antimicrobial activity against Escherichia coli. This peptide exhibits no cellular lytic activity, but electron microscopy and fluorescence studies reveal an ability to selectively disrupt the OM but not the inner membrane of E. coli. The selective targeting of the OM probably occurs through interactions of JB-95 with selected ß-barrel OM proteins, including BamA and LptD as shown by photolabeling experiments. Membrane proteomic studies reveal rapid depletion of many ß-barrel OM proteins from JB-95-treated E. coli, consistent with induction of a membrane stress response and/or direct inhibition of the Bam folding machine. The results suggest that lethal disruption of the OM by JB-95 occurs through a novel mechanism of action at key interaction sites within clusters of ß-barrel proteins in the OM. These findings open new avenues for developing antibiotics that specifically target ß-barrel proteins and the integrity of the Gram-negative OM.

Fluorescent rhodanine-3-acetic acids visualize neurofibrillary tangles in Alzheimer's disease brains.

There is a high demand for the development of an imaging agent for neurofibrillary tangles (NFTs) detection in Alzheimer's diagnosis. In the present study, a series of rhodanine-3-acetic acids was synthesized and evaluated for fluorescence imaging of NFTs in brain tissues of AD patients. Five out of seven probes have shown excellent binding affinity to NFTs over amyloid plaques in the Thiazine red R displacement assay. However, the selectivity in this in vitro assay is not confirmed by the histopathological evaluation, which indicates significant differences in the binding sites in the assays. Probe 6 showed binding affinity (IC50=19nM) to tau aggregates which is the highest among this series. Probes 2, 3, 4 and 5 display IC50 values of lower than 100nM to tau aggregates to displace Thiazine red R. Evaluation of the cytotoxicity of these five probes with human liver carcinoma cells revealed that these compounds excert negligible cytotoxicity. The in vivo studies with zebrafish embryos confirmed negligible cytotoxicity at 24 and 72h post fertilization.

Structure-based optimization of oxadiazole-based GSK-3 inhibitors.

Inhibition of glycogen synthase kinase-3 (GSK-3) induces neuroprotective effects, e.g. decreases ß-amyloid production and reduces tau hyperphosphorylation, which are both associated with Alzheimer's disease (AD). The two isoforms of GSK-3 in mammalians are GSK-3α and ß, which share 98% homology in their catalytic domains. We investigated GSK-3 inhibitors based on 2 different scaffolds in order to elucidate the demands of the ATP-binding pocket [1]. Particularly, the oxadiazole scaffold provided potent and selective GSK-3 inhibitors. For example, the most potent inhibitor of the present series, the acetamide 26d, is characterized by an IC50 of 2 nM for GSK-3α and 17 nM for GSK-3ß. In addition, the benzodioxane 8g showed up to 27-fold selectivity for GSK-3α over GSK-3ß, with an IC50 of 35 nM for GSK-3α. Two GSK-3 inhibitors were further profiled for efficacy and toxicity in the wild-type (wt) zebrafish embryo assay to evaluate simultaneously permeability and safety.

2-Styrylindolium based fluorescent probes visualize neurofibrillary tangles in Alzheimer's disease.

We evaluated 2-styrylindolium derivatives (6-11) as novel and selective probes for neurofibrillary tangles (NFTs) on brain sections of AD patients. The staining experiments indicated that these compounds may bind selectively to NFTs in the presence of ß-amyloid (Aß) plaques. Cell free binding assays confirmed that 2-[2-[4-(1-pyrrolidinyl)phenyl]ethenyl]-1,3,3-trimethyl-3H-indolium iodide (9) and 2-[2-[4-(diethylamino)phenyl]ethenyl]-1-butyl-3,3-dimethyl-3H-indolium iodide (11) display excellent affinities to Tau-aggregates (IC(50) values of 5.1 and 1.4 nM, respectively) in the displacement of Thiazin Red R. These probes have good solubility in distilled water and low or no cytotoxicity in zebrafish embryo and liver hepatocellular carcinoma cell assays.

Modification of a promiscuous inhibitor shifts the inhibition from γ-secretase to FLT-3.

The inhibition of FLT-3 activity is an interesting target for the treatment of acute myeloid leukemia (AML). The serendipitous identification of FLT-3 inhibitors from a CK1/γ-secretase programme provided compounds with dual inhibitory activity. We analyzed the structure-activity relationship of these inhibitors and derivatized them to arrive at compounds with reduced impact on γ-secretase activity and enhanced FLT-3 inhibition.

Small molecule kinase inhibitors for LRRK2 and their application to Parkinson's disease models.

Parkinson's disease (PD) is the second most common neurodegenerative disorder. Several single gene mutations have been linked to this disease. Mutations in the gene encoding leucine-rich repeat kinase 2 (LRRK2) indicate LRRK2 as promising therapeutic target for the treatment of PD. LRRK2 mutations were observed in sporadic as well as familial PD patients and have been investigated intensively. LRRK2 is a large and complex protein, with multiple enzymatic and protein-interaction domains, each of which is effected by mutations. The most common mutation in PD patients is G2019S. Several LRRK2 inhibitors have been reported already, although the crystal structure of LRRK2 has not yet been determined. This review provides a summary of known LRRK2 inhibitors and will discuss recent in vitro and in vivo results of these inhibitors.

Small-Molecule Inhibitors of GSK-3: Structural Insights and Their Application to Alzheimer's Disease Models.

The world health organization (WHO) estimated that 18 million people are struck by Alzheimer's disease (AD). The USA, France, Germany, and other countries launched major programmes targeting the identification of risk factors, the improvement of caretaking, and fundamental research aiming to postpone the onset of AD. The glycogen synthase kinase 3 (GSK-3) is implicated in multiple cellular processes and has been linked to the pathogenesis of several diseases including diabetes mellitus, cancer, and AD. Inhibition of GSK-3 leads to neuroprotective effects, decreased ß-amyloid production, and a reduction in tau hyperphosphorylation, which are all associated with AD. Various classes of small molecule GSK-3 inhibitors have been published in patents and original publications. Herein, we present a comprehensive summary of small molecules reported to interact with GSK-3. We illustrate the interactions of the inhibitors with the active site. Furthermore, we refer to the biological characterisation in terms of activity and selectivity for GSK-3, elucidate in vivo studies and pre-/clinical trials.

Identification of glycogen synthase kinase-3 inhibitors with a selective sting for glycogen synthase kinase-3α.

The glycogen synthase kinase-3 (GSK-3) has been linked to the pathogenesis of colorectal cancer, diabetes, cardiovascular disease, acute myeloid leukemia (AML), and Alzheimer's disease (AD). The debate on the respective contributions of GSK-3α and GSK-3ß to AD pathology and AML is ongoing. Thus, the identification of potent GSK-3α-selective inhibitors, endowed with favorable pharmacokinetic properties, may elucidate the effect of GSK-3α inhibition in AD and AML models. The analysis of all available crystallized GSK-3 structures provided a simplified scheme of the relevant hot spots responsible for ligand binding and potency. This resulted in the identification of novel scorpion shaped GSK-3 inhibitors. It is noteworthy, compounds 14d and 15b showed the highest GSK-3α selectivity reported so far. In addition, compound 14d did not display significant inhibition of 48 out of 50 kinases in the test panel. The GSK-3 inhibitors were further profiled for efficacy and toxicity in the wild-type (wt) zebrafish embryo assay.