Discovery of trans-3,4'-bispyridinylethylenes as potent and novel inhibitors of protein kinase B (PKB/Akt) for the treatment of cancer: Synthesis and biological evaluation

A novel series of Akt/PKB inhibitors derived from a screening lead (1) has been prepared. The novel trans-3,4'-bispyridinylethylenes described herein are potent inhibitors of Akt/PKB with IC(50) values in the low double-digit nanomolar range against Akt1. Compound 2q shows excellent selectivity against distinct families of kinases such as tyrosine kinases and CAMK, and displays poor to modest selectivity against closely related kinases in the AGC and CMGC families. The cellular activities including inhibition of cell growth and phosphorylation of downstream target GSK3 are also described. The X-ray structure of compound 2q complexed with PKA in the ATP binding site was determined.     

Synthesis and SAR of indazole-pyridine based protein kinase B/Akt inhibitors

A series of heteroaryl-pyridine containing inhibitors of Akt are reported. The synthesis and structure-activity relationships are discussed, leading to the discovery of a indazole-pyridine analogue (K(i)=0.16 nM). These compounds bind in the ATP binding site, are potent, ATP competitive, and reversible inhibitors of Akt activity. No selectivity amongst the Akt isoforms is observed for this analogue, but there is good selectivity against an panel of other kinases. It is least selective for other members of the AGC family of kinases but is nonetheless 40-fold selective for Akt over PKA. The compound shows cellular activity and significantly slows tumor growth in vivo.     

DNA damage bypass operates in the S and G2 phases of the cell cycle and exhibits differential mutagenicity

Translesion DNA synthesis (TLS) employs low-fidelity DNA polymerases to bypass replication-blocking lesions, and being associated with chromosomal replication was presumed to occur in the S phase of the cell cycle. Using immunostaining with anti-replication protein A antibodies, we show that in UV-irradiated mammalian cells, chromosomal single-stranded gaps formed in S phase during replication persist into the G2 phase of the cell cycle, where their repair is completed depending on DNA polymerase ζ and Rev1. Analysis of TLS using a high-resolution gapped-plasmid assay system in cell populations enriched by centrifugal elutriation for specific cell cycle phases showed that TLS operates both in S and G2. Moreover, the mutagenic specificity of TLS in G2 was different from S, and in some cases overall mutation frequency was higher. These results suggest that TLS repair of single-stranded gaps caused by DNA lesions can lag behind chromosomal replication, is separable from it, and occurs both in the S and G2 phases of the cell cycle. Such a mechanism may function to maintain efficient replication, which can progress despite the presence of DNA lesions, with TLS lagging behind and patching regions of discontinuity.     

E-cadherin in oral SCC: an analysis of the confusing literature and new insights related to its immunohistochemical expression

E-cadherin plays a crucial structural role in cell-cell contacts in epithelial tissues, and a functional role in signaling pathways that regulate cell proliferation, differentiation, and survival. Reduced immunoexpression of E-cadherin adhesions is largely considered as being equivalent to defective functionality and malignancy, and has been used as a prognostic parameter. A critical analysis of studies on E-cadherin immunoexpression in oral carcinomas revealed a wide range of both technical and interpretational aspects. This paper highlights biological characteristics of E-cadherin with respect to its expression in normal and neoplastic epithelial cells and to its interrelations with the tumor microenvironment that can have an impact on immunohistochemical results and their application in the clinical setting.     

Gibberellic Acid (GA3) Inhibits ROS Increase in Chloroplasts During Dark-Induced Senescence of Pelargonium Cuttings

The temporal and spatial changes in reactive oxygen species (ROS) during dark treatment of Pelargonium cuttings and the effect of gibberellic acid (GA₃) on ROS levels were studied. ROS-related fluorescence was detected in mitochondria and cytoplasm of epidermal cells and in chloroplasts. By monitoring dichlorofluorescein (DCF) fluorescence, an initial decrease in ROS was observed under darkness in the epidermal cell cytoplasm and the chloroplasts, which was followed by an increase on the third day. Following 3 days under darkness, the size and the structure of the chloroplasts also changed, and they became more sensitive to illumination as judged by a higher accumulation of ROS. Pretreatment of leaves with GA₃ did not prevent the structural changes in the chloroplasts, but it inhibited the increase in ROS levels in all cell compartments, including the chloroplasts. It is suggested that the inhibition of ROS increase by GA₃ prevented complete disintegration of chloroplasts during dark-induced senescence and thereby enabled the maintenance of chlorophyll levels in the tissue.     

Dyslexia Impairs Speech Recognition but Can Spare Phonological Competence

Dyslexia is associated with numerous deficits to speech processing. Accordingly, a large literature asserts that dyslexics manifest a phonological deficit. Few studies, however, have assessed the phonological grammar of dyslexics, and none has distinguished a phonological deficit from a phonetic impairment. Here, we show that these two sources can be dissociated. Three experiments demonstrate that a group of adult dyslexics studied here is impaired in phonetic discrimination (e.g., ba vs. pa), and their deficit compromises even the basic ability to identify acoustic stimuli as human speech. Remarkably, the ability of these individuals to generalize grammatical phonological rules is intact. Like typical readers, these Hebrew-speaking dyslexics identified ill-formed AAB stems (e.g., titug) as less wordlike than well-formed ABB controls (e.g., gitut), and both groups automatically extended this rule to nonspeech stimuli, irrespective of reading ability. The contrast between the phonetic and phonological capacities of these individuals demonstrates that the algebraic engine that generates phonological patterns is distinct from the phonetic interface that implements them. While dyslexia compromises the phonetic system, certain core aspects of the phonological grammar can be spared.     

Lead accumulation in the aquatic fern Azolla filiculoides.

In this study, we characterized lead (Pb2+) accumulation and storage by the aquatic fern Azolla filiculoides. Lead precipitates were detected in the vacuoles of mesophyll cells of Azolla plants cultured for 6 d in rich growth medium containing 20 mg l(-1) Pb2+. Energy dispersive spectroscopy (EDS) analysis of the relative element content of leaves collected from these plants revealed a 100% increase in the levels of P, S, Na and Ca and a 40% decrease in Mg and Cl compared to the untreated plants. Both Azolla whole plants and isolated apoplasts were incubated for 6 d in 20 mg l(-1) Pb2+. Lead content in the whole plant composed 0.37%, 2.3% and 1.8% of the dry weight after 2, 4 and 6 d of growth, respectively, while the isolated Azolla apoplast contained 0.125%, 1.22% and 1.4% Pb2+, respectively. Lead content in Azolla whole plant increase by 200%, 100% and 22% after 2, 4 and 6 d of growth, respectively, when compared to Azolla apoplast. Dark, electron dense deposits of lead were observed in light and transmission electron microscope in leaf cells treated with lead. All the observed lead deposits were localized in vacuoles while larger lead deposits were found in mature leaves than in young leaves. No lead deposits were found in cells of the cyanobiont Anabaena when the plants were exposed to similar conditions. Activity and content of V-H+-ATPase were studied in Azolla plants grown in the presence of 20, 40 and 80 mg l(-1) of lead for a period of 4 d. Activity of V-H+-ATPase was increased by 190%, 210% and 220%, respectively, but the content of V-H+-ATPase was reduced by all lead concentrations.     

PABPN1-Dependent mRNA Processing Induces Muscle Wasting

Poly(A) Binding Protein Nuclear 1 (PABPN1) is a multifunctional regulator of mRNA processing, and its expression levels specifically decline in aging muscles. An expansion mutation in PABPN1 is the genetic cause of oculopharyngeal muscle dystrophy (OPMD), a late onset and rare myopathy. Moreover, reduced PABPN1 expression correlates with symptom manifestation in OPMD. PABPN1 regulates alternative polyadenylation site (PAS) utilization. However, the impact of PAS utilization on cell and tissue function is poorly understood. We hypothesized that altered PABPN1 expression levels is an underlying cause of muscle wasting. To test this, we stably down-regulated PABPN1 in mouse tibialis anterior (TA) muscles by localized injection of adeno-associated viruses expressing shRNA to PABPN1 (shPab). We found that a mild reduction in PABPN1 levels causes muscle pathology including myofiber atrophy, thickening of extracellular matrix and myofiber-type transition. Moreover, reduced PABPN1 levels caused a consistent decline in distal PAS utilization in the 3’-UTR of a subset of OPMD-dysregulated genes. This alternative PAS utilization led to up-regulation of Atrogin-1, a key muscle atrophy regulator, but down regulation of proteasomal genes. Additionally reduced PABPN1 levels caused a reduction in proteasomal activity, and transition in MyHC isotope expression pattern in myofibers. We suggest that PABPN1-mediated alternative PAS utilization plays a central role in aging-associated muscle wasting.     

Prolonged stimuli alter the bacterial chemosensory clusters

The clustering of membrane‐bound receptors plays an essential role in various biological systems. A notable model system for studying this phenomenon is the bacterial chemosensory cluster that allows motile bacteria to navigate along chemical gradients in their environment. While the basic structure of these chemosensory clusters is becoming clear, their dynamic nature and operation are not yet understood. By measuring the fluorescence polarization of tagged receptor clusters in live Escherichia coli cells, we provide evidence for stimulus‐induced dynamics in these sensory clusters. We find that when a stimulus is applied, the packing of the receptors slowly decreases and that the process reverses when the stimulus is removed. Consistent with these physical changes we find that the effective cooperativity of the kinase response slowly evolves in the presence of a stimulus. Time‐lapse fluorescence imaging indicates that, despite these changes, the receptor clusters do not generally dissociate upon ligand binding. These data reveal stimulus‐dependent plasticity in chemoreceptor clusters.     

Soil Microbial Diversity in the Vicinity of Desert Shrubs

Water and nutrient availability are the major limiting factors of biological activity in arid and semiarid ecosystems. Therefore, perennial plants have developed different ecophysiological adaptations to cope with harsh conditions. The chemical profile of the root exudates varies among plant species and this can induce variability in associated microbial populations. We examined the influence of two shrubs species, Artemisia sieberi and Noaea mucronata, on soil microbial diversity. Soil samples were collected monthly, from December 2006 to November 2007, near canopies of both shrubs (0–10-cm depth). Samples were used for abiotic tests and determination of soil bacterial diversity. No significant differences were found in the abiotic variables (soil moisture, total organic matter, and total soluble nitrogen (TSN)) between soil samples collected from under the two shrubs during the study period. No obvious differences in the Shannon–Weaver index, evenness values, or total phylogenetic distances were found for the soil microbial communities. However, detailed denaturing gradient gel electrophoresis (DGGE) clustering as well as taxonomic diversity analyses indicated clear shifts in the soil microbial community composition. These shifts were governed by seasonal variability in water availability and, significantly, by plant species type.