Synthesis and biological evaluation of novel acylhydrazone derivatives as potential antitumor agents

We have designed, synthesized, and evaluated as potential antitumor agents a series of 2-hydroxybenzylidene derivatives of the N-(2-trifluoromethylpiridyn-4-yl)anthranilic acid hydrazide, and some analogues bearing a (2-trifluoromethyl)piridyn-4-ylamino group in 3- or 4-position of benzohydrazide or 4-position of phenylacetohydrazide. Compounds 12e, 13e, 15e, and 16e, bearing a 4-(diethylamino)salicylidene group exhibited potent cytotoxicity, with averaged GI₅₀ values in sub-micromolar range, and a variety of cell selectivity at nanomolar concentrations. The determination of acute toxicity in athymic nudes mice proved some compounds to be non-toxic, making them good candidates for further study as antitumor agents.     

Minimized combinatorial CRISPR screens identify genetic interactions in autophagy

Combinatorial CRISPR-Cas screens have advanced the mapping of genetic interactions, but their experimental scale limits the number of targetable gene combinations. Here, we describe 3Cs multiplexing, a rapid and scalable method to generate highly diverse and uniformly distributed combinatorial CRISPR libraries. We demonstrate that the library distribution skew is the critical determinant of its required screening coverage. By circumventing iterative cloning of PCR-amplified oligonucleotides, 3Cs multiplexing facilitates the generation of combinatorial CRISPR libraries with low distribution skews. We show that combinatorial 3Cs libraries can be screened with minimal coverages, reducing associated efforts and costs at least 10-fold. We apply a 3Cs multiplexing library targeting 12,736 autophagy gene combinations with 247,032 paired gRNAs in viability and reporter-based enrichment screens. In the viability screen, we identify, among others, the synthetic lethal WDR45B-PIK3R4 and the proliferation-enhancing ATG7-KEAP1 genetic interactions. In the reporter-based screen, we identify over 1,570 essential genetic interactions for autophagy flux, including interactions among paralogous genes, namely ATG2A-ATG2B, GABARAP-MAP1LC3B and GABARAP-GABARAPL2. However, we only observe few genetic interactions within paralogous gene families of more than two members, indicating functional compensation between them. This work establishes 3Cs multiplexing as a platform for genetic interaction screens at scale.     

De novo synthesis of diacylglycerol from glucose. A new pathway of signal transduction in human neutrophils stimulated during phagocytosis of beta-glucan particles.

The phagocytosis of beta-glucan particles by human neutrophils and the associated activation of NADPH O2- forming oxidase were accompanied by an increased hydrolysis of phosphoinositides by phospholipase C, hydrolysis of phosphatidylcholine by phospholipase D, accumulation of diglyceride (DG) mass, and [Ca2+]i rise. The reaction of phospholipid hydrolysis played a minor role in the formation of DG, which was mainly formed by de novo synthesis from glucose. The activation of this pathway was shown by the stimulation of the incorporation of [U-14C]glucose into DG, which occurred very rapidly after the challenge of neutrophils with beta-glucan particles. This DG derived from glucose was found almost completely as 1-acyl-2-acyl-glycerol (DAG). On the basis of the finding that phosphatidic acid was the precursor of DAG, an increase in the incorporation of [U-14C]acetate into DAG did not occur, and the [14C]radioactivity was in the glycerol backbone, the synthesis of DAG from [U-14C]glucose occurred very likely via dihydroxyacetone phosphate and glycerol 3-phosphate, stepwise acylation to phosphatidic acid, and dephosphorylation by phosphatidate phosphatase.     

Location and functional characterization of the right atrioventricular pacemaker ring in the adult avian heart

Previous histological studies showed that in addition to a sinus node, an atrioventricular (AV) node, an AV bundle, left and right bundle branches, birds also possess a right AV‐Purkinje ring that is located in the atrial sheet of the right muscular AV‐valve along all its base length. The functionality of the AV‐Purkinje ring is unknown. In this work, we studied the topology of pacemaker myocytes in the atrial side of the isolated chicken spontaneously contracting right muscular AV‐valve using the method of microelectrode mapping of action potentials. We show that AV‐cells having the ability to show pacemaking reside in the right muscular AV‐valve. Pacemaker action potentials were exclusively recorded close to the base of the valve along its whole length from dorsal to the ventral attachment to the interventricular septum. These action potentials have much slower rate of depolarization, lower amplitude, and higher diastolic depolarization than action potentials of Purkinje (conducting) cells. We conclude the right AV‐valve has a ring bundle of pacemaker cells (but not Purkinje cells) in the adult chicken heart. J. Morphol. 277:363–369, 2016. © 2015 Wiley Periodicals, Inc.     

An efficient transformation system for Bifidobacterium spp.

This study describes a broad host transformation protocol that enables the uptake of plasmid DNA into 10 different species of Bifidobacterium , some of which have never been transformed before. The vector pNC7 (4·9 kb) was used to optimize the electroporation protocol. Transformation efficiencies ranged from 3·6×10⁻¹ to 1·2×10⁵ transformations per μg DNA. The impact of growth medium composition and electric field strength on transformation efficiency were independently optimized. Electrocompetent cells were grown in Iwata medium broth enriched with Actilight^RP 16%, harvested during the early exponential growth phase, and pulsed at 12·5 kV cm⁻¹, 100 Ω and 25 μF.     

Physique and Performance of Young Wheelchair Basketball Players in Relation with Classification

The relationships among physical characteristics, performance, and functional ability classification of younger wheelchair basketball players have been barely investigated to date. The purpose of this work was to assess anthropometry, body composition, and performance in sport-specific field tests in a national sample of Italian younger wheelchair basketball players as well as to evaluate the association of these variables with the players’ functional ability classification and game-related statistics. Several anthropometric measurements were obtained for 52 out of 91 eligible players nationwide. Performance was assessed in seven sport-specific field tests (5m sprint, 20m sprint with ball, suicide, maximal pass, pass for accuracy, spot shot and lay-ups) and game-related statistics (free-throw points scored per match, two- and three-point field-goals scored per match, and their sum). Association between variables, and predictivity was assessed by correlation and regression analysis, respectively. Players were grouped into four Classes of increasing functional ability (A-D). One-way ANOVA with Bonferroni’s correction for multiple comparisons was used to assess differences between Classes. Sitting height and functional ability Class especially correlated with performance outcomes, but wheelchair basketball experience and skinfolds did not. Game-related statistics and sport-specific field-test scores all showed significant correlation with each other. Upper arm circumference and/or maximal pass and lay-ups test scores were able to explain 42 to 59% of variance in game-related statistics (P<0.001). A clear difference in performance was only found for functional ability Class A and D. Conclusion: In younger wheelchair basketball players, sitting height positively contributes to performance. The maximal pass and lay-ups test should be carefully considered in younger wheelchair basketball training plans. Functional ability Class reflects to a limited extent the actual differences in performance.     

Pollen wall ontogeny in <Emphasis Type="Italic">Polemonium caeruleum</Emphasis> (Polemoniaceae) and suggested underlying mechanisms of development

By a detailed ontogenetic study of Polemonium caeruleum pollen, tracing each stage of development at high TEM resolution, we aim to understand the establishment of the pollen wall and to unravel the mechanisms underlying sporoderm development. The main steps of exine ontogeny in Polemonium caeruleum, observed in the microspore periplasmic space, are spherical units, gradually transforming into columns, then to rod-like units (procolumellae), the appearance of the initial tectum, growth of columellae in height and tectum in thickness and initial sporopollenin accumulation on them, the appearance of the endexine lamellae and of dark-contrasted particles on the tectum, the appearance of a sponge-like layer and of the intine in aperture sites, the appearance of the foot layer on the base of the sponge-like layer and of spinules on the tectum, and massive sporopollenin accumulation. This sequence of developmental events fits well to the sequence of self-assembling micellar mesophases. This gives (together with earlier findings and experimental exine simulations) strong evidence that genome and self-assembly probably share control of exine formation. It is highly probable that self-assembly is an intrinsic instrument of evolution.     

Novel sulfenamides and sulfonamides based on pyridazinone and pyridazine scaffolds as CB1 receptor ligand antagonists

A series of sulfenamide and sulfonamide derivatives was synthesized and evaluated for the affinity at CB₁ and CB₂ receptors. The N-bornyl-S-(5,6-di-p-tolylpyridazin-3-yl)-sulfenamide, compound 11, displayed good affinity and high selectivity for CB₁ receptors (K_i values of 44.6?nM for CB₁ receptors and >40?μM for CB₂ receptors, respectively). The N-isopinocampheyl-sulfenamide 12 and its sulfonamide analogue 22 showed similar selectivity for CB₁ receptors with K_i values of 75.5 and 73.2?nM, respectively. These novel compounds behave as antagonists/inverse agonists at CB₁ receptor in the [³⁵S]-GTPγS binding assays, and none showed adequate predictive blood–brain barrier permeation, exhibiting low estimated LD₅₀. However, testing compound 12 in a supraspinal analgesic test (hot-plate) revealed that it was as effective as the classic CB₁ receptor antagonist rimonabant, in reversing the analgesic effect of a cannabinoid agonist.     

Is volumetric modulated arc therapy with constant dose rate a valid option in radiation therapy for head and neck cancer patients?

Background

Intensity-modulated radiotherapy (IMRT) improves dose distribution in head and neck (HN) radiation therapy. Volumetric-modulated arc therapy (VMAT), a new form of IMRT, delivers radiation in single or multiple arcs, varying dose rates (VDR-VMAT) and gantry speeds, has gained considerable attention. Constant dose rate VMAT (CDR-VMAT) associated with a fixed gantry speed does not require a dedicated linear accelerator like VDR-VMAT. The present study explored the feasibility, efficiency and delivery accuracy of CDR-VMAT, by comparing it with IMRT and VDR-VMAT in treatment planning for HN cancer.