SAR Refinement di derivati benzotiopiranopirimidinici e piridotiopiranopirimidinici quali inibitori di VEGFR-2

L’angiogenesi rappresenta uno dei bersagli principali di nuovi composti antitumorali e molti farmaci anti-angiogenesi sono attualmente somministrati per la cura di molte neoplasie in fase avanzata. Gli studi sull’angiogenesi si basano sul presupposto che, prevenire la crescita di nuovi vasi sanguigni possa compromettere la vita delle cellule tumorali. Quindi lo scopo della terapia anti-angiogenica consiste nel stabilizzare la malattia e controllare la cancerogenesi, piuttosto che sradicare la massa tumorale come fanno chemioterapia e radioterapia. Dallo studio delle basi molecolari del processo angiogenico è emerso che la via del Fattore di Crescita dell’Endotelio Vascolare (VEGF) rappresenta uno dei più importanti modulatori positivi di questo meccanismo fisiologico. I membri della famiglia VEGF hanno capacità sovrapposte di interagire con una serie di recettori tirosin chinasi: VEGFR-1 e VEGFR-2 (KDR), che sono prevalentemente espressi nelle cellule endoteliali e coinvolti principalmente nell’angiogenesi e il VEGFR-3, presente nei vasi linfatici dove, sembra, regolare la linfoangiogenesi. La dimostrazione che l’espressione del VEGF e dei relativi recettori sia correlata al grado di vascolarizzazione di svariati tumori sperimentali e clinici ha portato alla progettazione e allo sviluppo di agenti che bersagliano il pathway del VEGF: a partire dagli anticorpi monoclonali anti-VEGF, come il bevacizumab, alle piccole molecole ATP-competitive VEGF inibitrici, compresi composti eterociclici appartenenti a svariate classi. All’interno della classe delle pirimidine, quelle con una funzione anilinica inserita sul sistema pirimidinico hanno mostrato di avere effetti anti-angiogenici notevoli, e per tale motivo sono stati utilizzati come composti lead per la progettazione di nuovi inibitori competitivi del VEGFR-2. In questo contesto, il mio lavoro di tesi ha riguardato la sintesi di composti polieteroeterociclici, caratterizzati da pirimidine fuse con i sistemi benzotiopirano e piridotiopirano in cui sono stati inseriti i sostituenti in posizione meta del sistema anilinico pendente. La progettazione delle strutture sintetizzate è stata sviluppata in seguito ai risultati biologici e di molecular docking effettuati su composti analoghi di tipo benzotiopirano- e piridotiopirano-pirimidinico recentemente sintetizzati presso il laboratorio dove ho svolto la mia tesi sperimentale. In particolare, sono stati inseriti in posizione meta oltre a Cl, ed OCH3, che erano gia presenti in posizione para in composti precedentemente sintetizzati, anche il Br, dato che numerosi inibitori del recettore VEGFR-2, descritti in letteratura, sono caratterizzati dalla presenza di questo tipo di sostituente. La valutazione dell’attività antiproliferativa dei nuovi composti sintetizzati sarà effettuata in collaborazione con l’Università di Padova, e sarà condotta in vitro su varie linee cellulari: HeLa (adenocarcinoma della cervice uterina), A-431 (carcinoma squamoso), MSTO-211H (mesotelioma bifasico) e cellule endoteliali della vena ombelicale umana (HUVEC). Allo scopo di indagare il meccanismo d’azione responsabile dell’eventuale effetto antiproliferativo, sarà valutata la capacità dei composti di inibire l’attività tirosina chinasica del KDR per mezzo di un saggio effettuato su enzimi ricombinanti uman

Snake and spider toxins induce a rapid recovery of function of botulinum neurotoxin paralysed neuromuscular junction

Botulinum neurotoxins (BoNTs) and some animal neurotoxins (-Bungarotoxin, -Btx, from elapid snakes and -Latrotoxin, -Ltx, from black widow spiders) are pre-synaptic neurotoxins that paralyse motor axon terminals with similar clinical outcomes in patients. However, their mechanism of action is different, leading to a largely-different duration of neuromuscular junction (NMJ) blockade. BoNTs induce a long-lasting paralysis without nerve terminal degeneration acting via proteolytic cleavage of SNARE proteins, whereas animal neurotoxins cause an acute and complete degeneration of motor axon terminals, followed by a rapid recovery. In this study, the injection of animal neurotoxins in mice muscles previously paralyzed by BoNT/A or /B accelerates the recovery of neurotransmission, as assessed by electrophysiology and morphological analysis. This result provides a proof of principle that, by causing the complete degeneration, reabsorption, and regeneration of a paralysed nerve terminal, one could favour the recovery of function of a biochemically- or genetically-altered motor axon terminal. These observations might be relevant to dying-back neuropathies, where pathological changes first occur at the neuromuscular junction and then progress proximally toward the cell body

3d printing technologies: Are their materials safe for conservation treatments?

3D printing technologies have been definitively introduced in conservation treatments. Despite the advantage of not requiring direct contact with the artwork, allowing the preservation of fragile objects, the printed item is located in direct contact with the object and the characterization of the filament used for the printing is not often taken into consideration. The following study was undertaken as an evaluation of filaments possibly employed for conservation treatments. The characterisation of the components was carried out through infrared spectroscopy, thermal and chromatographic analyses. Moreover, it was investigated whether such materials release volatile organic compounds (VOC) during their degradation process. Indeed, all of them released styrenic and alkyl compounds, all solvents for materials that can be found on artworks, employed by both, artist and conservator

Coat flexibility in the secretory pathway: a role in transport of bulky cargoes.

Membrane trafficking in eukaryotic cells is a highly dynamic process, which needs to adapt to a variety of cargo proteins. The COPII coat mediates ER export of thousands of proteins with a wide range of sizes by generating coated membrane vesicles that incapsulate cargo. The process of assembly and disassembly of COPII, regulated by GTP hydrolysis, is a major determinant of the size and shape of transport carriers. Here, we analyse our knowledge of the COPII coat architecture and it assembly/disassembly dynamics, and link coat flexibility to the role of COPII in transport of large cargoes. We propose a common mechanism of action of regulatory factors that modulate COPII GTP hydrolysis cycle to promote budding

Current data processing strategies for cryo-electron tomography and subtomogram averaging

Cryo-electron tomography (cryo-ET) can be used to reconstruct three-dimensional (3D) volumes, or tomograms, from a series of tilted two-dimensional images of biological objects in their near-native states in situ or in vitro. 3D subvolumes, or subtomograms, containing particles of interest can be extracted from tomograms, aligned, and averaged in a process called subtomogram averaging (STA). STA overcomes the low signal to noise ratio within the individual subtomograms to generate structures of the particle(s) of interest. In recent years, cryo-ET with STA has increasingly been capable of reaching subnanometer resolution due to improvements in microscope hardware and data processing strategies. There has also been an increase in the number and quality of software packages available to process cryo-ET data with STA. In this review, we describe and assess the data processing strategies available for cryo-ET data and highlight the recent software developments which have enabled the extraction of high-resolution information from cryo-ET datasets

In situ structure determination by subtomogram averaging

Cryo-tomography and subtomogram averaging are increasingly popular techniques for structural determination of macromolecular complexes in situ. They have the potential to achieve high-resolution views of native complexes, together with the details of their location relative to interacting molecules. The subtomogram averaging (StA) pipelines are well-established, with current developments aiming to optimise each step by reducing manual intervention and user decisions, following similar trends in single-particle approaches that have dramatically increased their popularity. Here, we review the main steps of typical StA workflows. We focus on considerations arising from the fact that the objects of study are embedded within unique crowded environments, and we emphasise those steps where careful decisions need to be made by the user. [Abstract copyright: Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.

In Silico Meta-Analysis of Boundary Conditions for Experimental Tests on the Lumbar Spine

The study of the spine range of motion under given external load has been the object of many studies in literature, finalised to a better understanding of the spine biomechanics, its physiology, eventual pathologic conditions and possible rehabilitation strategies. However, the huge amount of experimental work performed so far cannot be straightforwardly analysed due to significant differences among loading set-ups. This work performs a meta-analysis of various boundary conditions in literature, focusing on the flexion/extension behaviour of the lumbar spine. The comparison among range of motions is performed virtually through a validated multibody model. Results clearly illustrated the effect of various boundary conditions which can be met in literature, so justifying differences of biomechanical behaviours reported by authors implementing different set-up: for example, a higher value of the follower load can indeed result in a stiffer behaviour; the application of force producing spurious moments results in an apparently more deformable behaviour, however the respective effects change at various segments along the spine due to its natural curvature. These outcomes are reported not only in qualitative, but also in quantitative terms. The numerical approach here followed to perform the meta-analysis is original and it proved to be effective thanks to the bypass of the natural variability among specimens which might completely or partially hinder the effect of some boundary conditions. In addition, it can provide very complete information since the behaviour of each functional spinal unit can be recorded. On the whole, the work provided an extensive review of lumbar spine loading in flexion/extension

a multibody simulation of a human fall model creation and validation

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