Cecilia Astigiano

Recently, the development of sirtuin small molecule inhibitors (SIRTIs) has been gaining attention for the treatment of different cancer types, but also to contrast neurodegenerative disease, diabetes, and autoimmune syndromes. In the search for SIRT2 modulators, the availability of several X-crystallographic data regarding SIRT2−ligand complexes has allowed for setting up a structure-based study, which is herein presented. A set of 116 SIRT2 inhibitors featuring different chemical structures has been collected from the literature and used for molecular docking studies involving 4RMG and 5MAT PDB codes. The information found highlights key contacts with the SIRT2 binding pocket such as Van der Waals and π–π stacking with Tyr104, Phe119, Phe234, and Phe235 in order to achieve high inhibitory ability values. Following the preliminary virtual screening studies, a small in-house library of compounds (1a–7a), previously investigated as putative HSP70 inhibitors, was described to guide the search for dual-acting HSP70/SIRT2 inhibitors. Biological and enzymatic assays validated the whole procedure. Compounds 2a and 7a were found to be the most promising derivatives herein proposed.

Sirtuin 6 (SIRT6) is a member of the mammalian NAD⁺-dependent deac(et)ylase sirtuin family. SIRT6’s anti-inflammatory roles are emerging increasingly often in different diseases and cell types, including endothelial cells. In this study, the role of SIRT6 in pro-inflammatory conditions was investigated by engineering human umbilical vein endothelial cells to overexpress SIRT6 (SIRT6+ HUVECs). Our results showed that SIRT6 overexpression affected the levels of adhesion molecules and sustained megakaryocyte proliferation and proplatelet formation. Interestingly, the pro-inflammatory activation of the ATP/purinergic axis was reduced in SIRT6+ HUVECs. Specifically, the TNFα-induced release of ATP in the extracellular space and the increase in pannexin-1 hemichannel expression, which mediates ATP efflux, were hampered in SIRT6+ cells. Instead, NAD⁺ release and Connexin43 expression were not modified by SIRT6 levels. Moreover, the Ca²⁺ influx in response to ATP and the expression of the purinergic receptor P2X7 were decreased in SIRT6+ HUVECs. Contrary to extracellular ATP, extracellular NAD⁺ did not evoke pro-inflammatory responses in HUVECs. Instead, NAD⁺ administration reduced endothelial cell proliferation and motility and counteracted the TNFα-induced angiogenesis. Altogether, our data reinforce the view of SIRT6 activation as an anti-inflammatory approach in vascular endothelium.

ADP-ribosyl cyclases (ADPRCs) catalyze the synthesis of the Ca²⁺-active second messengers Cyclic ADP-ribose (cADPR) and ADP-ribose (ADPR) from NAD⁺ as well as nicotinic acid adenine dinucleotide phosphate (NAADP⁺) from NADP⁺. The best characterized ADPRC in mammals is CD38, a single-pass transmembrane protein with two opposite membrane orientations. The first identified form, type II CD38, is a glycosylated ectoenzyme, while type III CD38 has its active site in the cytosol. The ectoenzymatic nature of type II CD38 raised long ago the question of a topological paradox concerning the access of the intracellular NAD⁺ substrate to the extracellular active site and of extracellular cADPR product to its intracellular receptors, ryanodine (RyR) channels. Two different transporters, equilibrative connexin 43 (Cx43) hemichannels for NAD⁺ and concentrative nucleoside transporters (CNTs) for cADPR, proved to mediate cell-autonomous trafficking of both nucleotides. Here, we discussed how type II CD38, Cx43 and CNTs also play a role in mediating several paracrine processes where an ADPRC⁺ cell supplies a neighboring CNT-and RyR-expressing cell with cADPR. Recently, type II CD38 was shown to start an ectoenzymatic sequence of reactions from NAD⁺/ADPR to the strong immunosuppressant adenosine; this paracrine effect represents a major mechanism of acquired resistance of several tumors to immune checkpoint therapy.