Retromer stabilization results in neuroprotection in a model of Amyotrophic Lateral Sclerosis

Amyotrophic Lateral Sclerosis (ALS) is a fatal disease characterized by the degeneration of upper and lower motor neurons (MNs). We find a significant reduction of the retromer complex subunit VPS35 in iPSCs-derived MNs from ALS patients, in MNs from ALS post mortem explants and in MNs from SOD1G93A mice. Being the retromer involved in trafficking of hydrolases, a pathological hallmark in ALS, we design, synthesize and characterize an array of retromer stabilizers based on bis-guanylhydrazones connected by a 1,3-phenyl ring linker. We select compound 2a as a potent and bioavailable interactor of VPS35-VPS29. Indeed, while increasing retromer stability in ALS mice, compound 2a attenuates locomotion impairment and increases MNs survival. Moreover, compound 2a increases VPS35 in iPSCs-derived MNs and shows brain bioavailability. Our results clearly suggest the retromer as a valuable druggable target in ALS

RATIONAL DESIGN AND SYNTHESIS OF SMALL MOLECULES TARGETED AGAINST NEURODEGENERATIVE PROCESSES AND DISEASES

Improved sanitary conditions, disease prevention (e.g., vaccinations) and treatment (e.g., antibiotics) have increased life expectancy, up to the 67.2 years value in 2010. Unfortunately, living longer a poor life does not represent anyone\u2019s dream, mostly due to neurodegenerative diseases (NDDs). Alzheimer\u2019s Disease International (ADI) estimates in its 2013 report that there are more than 35 million people with dementia worldwide as of 2010, and that this number will double by 2030 and triplicate by 2050. In industrialized countries the prevalence of Parkinson\u2019s disease (PD) is about 1% for people over 60, with estimates of up to 4% for people in the highest age groups. In 2016 the Centres for Disease Control and Prevention estimated that between 14,000 - 15,000 Americans suffered from Amyotrophic Lateral Sclerosis (ALS). These and other NDDs are the result of neurodegenerative processes, that entail the progressive loss of structure or function of neurons, eventually causing their death. Symptomatic treatment strategies available on the market for NDDs are inadequate, as they offer only temporary relief without changing the ultimate NDD outcome. Effective treatment of NDDs should be based on small molecules able to modulate disease-modifying pathways involved in the development and/or the progression of NDDs, to cause their remission. In my Ph.D. work I focused on four different pathways (each described in a dedicated Chapter) involved in the development of multiple NDDs. With the aim to therapeutically modulate them by promoting and/or inhibiting those targets, I\u2019ve rationally designed and synthetized several classes of compounds. My research group established a multi-disciplinary approach in collaboration with bioinformatics, biologists, pharmacologists and clinicians; thus, my putative NDD treatments were tested by our collaborators in different Universities and Institutes (CIBIO, University of Trento; San Raffaele Research Institute, Milan; Department of Neuroscience, Federico II University, Naples)

Synthesis of dual action Smac/Zinc-Chelator conjugates as putative proapoptotic agents

Apoptosis, or programmed cell death, is a critical cell process in normal development and homeostasis of multicellular organisms. It is now recognized that dysfunction of the apoptosis machinery is a hallmark of cancer. Accordingly, targeting critical apoptosis regulators is an attractive approach for the development of new classes of therapies for the treatment of cancer and other human diseases. The X-linked inhibitor of apoptosis protein (XIAP) is a member of IAP proteins that potently inhibit apoptosis[1]. XIAP contains three baculovirus IAP repeat (BIR) domains. The mechanism of action of XIAP entails its binding with initiator and effector caspases through its BIR domains. In cells, the anti-apoptotic function of XIAP is antagonized by Smac/DIABLO (second mitochondria-derived activator of caspases or direct IAP binding protein with low pI). Despite the rather complex structure of Smac, its short N-terminal AVPI sequence is sufficient to trigger the inactivation of anti-apoptotic XIAP[2]. Our research group has shown how small monomeric, AVPI-inspired Smac mimics can bind XIAP on its BIR3 domain with sub-micromolar potency[3,4]. Executioner caspase-3, -6 and -7 exist within the cytosol as inactive zymogens (procaspases) activated by limited proteolysis within their inter-domain linker, carried out by an initiator caspaseThe essential executioner caspase-3 is proteolytically activated by either caspase-8 or -9. Zinc ions co-localize with procaspase-3/caspase-3 and inhibit its enzymatic activity in the cell by direct interaction with an Asp-Asp-Asp (DDD) \u201csafety catch\u201d region[5]. Thus, in this work we coupled a zinc chelator moiety based on di(picolylamide)amine (DPA) and its N,N-bis(pyridin-2-ylmethyl)ethane-1,2-diamine (BPEN) derivative to pro-apoptotic Smac mimetics, synthesized starting from known intermediates 1 and 2. Dual action Smac mimetic-zinc chelators 3 and 4 were prepared from compounds 1 and 2 as shown in Scheme 1 and characterized in vitro, using cell-free and cellular assays[6,7]. Their ability to bind XIAP BIR3 domain, to process pro-caspase-3 to caspase-3 and their cytotoxicity have been experimentally determined, and favorably compared with those of a potent Smac mimic compound, especially for the most potent, tribasic dual action compound 4. Furthermore, the Zinc affinity for both compounds was confirmed by fluorescence measurements. References 1. Q. L. Deveraux, J. C. Reed. Genes & Dev. 1999, 13, 239-252 2. J. Chai, C. Du, J. W. Wu, S. Kyin, X. Wang, Y.Shi. Nature 2000, 406, 855-862 3. P. Seneci, A. Bianchi, C. Battaglia, L. Belvisi, M. Bolognesi, A. Caprini, F. Cossu, E. de Franco, M. de, D. Delia, C. Drago, A. Khaled, D. Lecis, L. Manzoni, M. Marizzoni, E. Mastrangelo, M. Milani,I. Motto, E. Moroni, D. Potenza, V. Rizzo, F. Servida, E. Turlizzi, M. Varrone, F. Vasile, C. Scolastico. Bioorg. Med. Chem. 2009, 17, 5834\u20135856. 4. L. Manzoni, D. Arosio, L. Belvisi, A. Bracci, M. Colombo, D. Invernizzi, C. Scolastico. J. Org. Chem. 2005, 70, 4124-4132. 5. K. S. Putt, G. W. Chen, J. M. Pearson, J. S Sandhorst, M. S. Hoagland, J. Kwon, S. Hwang, H. Jin, M. I. Churchwell, M. Cho, D. R. Doerge, W. G. Helferich, P. J. Hergenrother. Nat. Chem. Biol. 2006, 2, 543, 550 6. Z. Nikolovska-Coleska, R. Wang, X. Fang, H. Pan, Y. Tomita, P. Li, P.P. Roller, K. Krajewski, N.G. Saito, J.A. Stuckey, S. Wang, Anal. Biochem. 2004, 332, 261-273. 7. Z. Nikolovska-Coleska, J.L. Meagher, S. Jiang, S.A. Kawamoto, W. Gao, H. Yi, D. Qin, P.P. Roller, J.A. Stuckey, S. Wang, Anal. Biochem. 2008, 374, 87-98

New experimental VLE data for the binary mixture of carbon dioxide + perfluorohexane (CO2 + C6F14) from 273 K to 333 K

Mixtures of carbon dioxide and perfluorocarbons are more frequently used in different fields: in energy conversion systems, in the electronic industry, in chemical plants and for medical applications. Among all perfluorocarbons, perfluorohexane has interesting properties (e.g. high thermal stability and high carbon dioxide solubility) to be used in mixture, as a working fluid in power and refrigeration cycles. The study of these systems relies on the availability of accurate thermodynamic models of the fluid mixture, which usually require to be calibrated on experimental data. Experimental data on the carbon dioxide and perfluorohexane mixture are scarcely available in literature and limited to a very narrow temperature range. This work aims to enrich the available literature data on VLE measurements of carbon dioxide and perfluorohexane mixture. On the basis of the implemented static-analytical method, high-pressure phase equilibrium data have been collected. Isothermal measurements have been performed within the temperature range of 273 K-333 K and at pressures up to 8.5 MPa. Particular attention to calibration procedures of the measuring devices (temperature and pressure probes and gas chromatograph) and to uncertainty calculation has been devoted. Finally, measurements show a good agreement with the data available in literature. (C) 2019 Elsevier B.V. All rights reserved