New Trends in Aging Drug Discovery

Aging is considered the main risk factor for many chronic diseases that frequently appear at advanced ages. However, the inevitability of this process is being questioned by recent research that suggests that senescent cells have specific features that differentiate them from younger cells and that removal of these cells ameliorates senescent phenotype and associated diseases. This opens the door to the design of tailored therapeutic interventions aimed at reducing and delaying the impact of senescence in life, that is, extending healthspan and treating aging as another chronic disease. Although these ideas are still far from reaching the bedside, it is conceivable that they will revolutionize the way we understand aging in the next decades. In this review, we analyze the main and well-validated cellular pathways and targets related to senescence as well as their implication in aging-associated diseases. In addition, the most relevant small molecules with senotherapeutic potential, with a special emphasis on their mechanism of action, ongoing clinical trials, and potential limitations, are discussed. Finally, a brief overview of alternative strategies that go beyond the small molecule field, together with our perspectives for the future of the field, is provided.Depto. de Ingeniería Química y de MaterialesDepto. de Química OrgánicaSección Deptal. de Química Orgánica (Óptica y Optometría)Fac. de Ciencias QuímicasFac. de Óptica y OptometríaTRUEMinisterio de Ciencia e Innovación de Españapu

Cannabinoids induce functional Tregs by promoting tolerogenic DCs via autophagy and metabolic reprograming

The generation of functional regulatory T cells (Tregs) is essential to keep tissue homeostasis and restore healthy immune responses in many biological and inflammatory contexts. Cannabinoids have been pointed out as potential therapeutic tools for several diseases. Dendritic cells (DCs) express the endocannabinoid system, including the cannabinoid receptors CB1 and CB2. However, how cannabinoids might regulate functional properties of DCs is not completely understood. We uncover that the triggering of cannabinoid receptors promote human tolerogenic DCs that are able to prime functional FOXP3+ Tregs in the context of different inflammatory diseases. Mechanistically, cannabinoids imprint tolerogenicity in human DCs by inhibiting NF-κB, MAPK and mTOR signalling pathways while inducing AMPK and functional autophagy flux via CB1- and PPARα-mediated activation, which drives metabolic rewiring towards increased mitochondrial activity and oxidative phosphorylation. Cannabinoids exhibit in vivo protective and anti-inflammatory effects in LPS-induced sepsis and also promote the generation of FOXP3+ Tregs. In addition, immediate anaphylactic reactions are decreased in peanut allergic mice and the generation of allergen-specific FOXP3+ Tregs are promoted, demonstrating that these immunomodulatory effects take place in both type 1- and type 2-mediated inflammatory diseases. Our findings might open new avenues for novel cannabinoid-based interventions in different inflammatory and immune-mediated diseases

Synthetic inhibitors of bacterial cell division targeting the GTP-binding site of FtsZ

Cell division protein FtsZ is the organizer of the cytokinetic Z-ring in most bacteria and a target for new antibiotics. FtsZ assembles with GTP into filaments that hydrolyze the nucleotide at the association interface between monomers and then disassemble. We have replaced FtsZ's GTP with non-nucleotide synthetic inhibitors of bacterial division. We searched for these small molecules among compounds from the literature, from virtual screening (VS), and from our in-house synthetic library (UCM), employing a fluorescence anisotropy primary assay. From these screens we have identified the polyhydroxy aromatic compound UCM05 and its simplified analogue UCM44 that specifically bind to Bacillus subtilis FtsZ monomers with micromolar affinities and perturb normal assembly, as examined with light scattering, polymer sedimentation, and negative stain electron microscopy. On the other hand, these ligands induce the cooperative assembly of nucleotide-devoid archaeal FtsZ into distinct well-ordered polymers, different from GTP-induced filaments. These FtsZ inhibitors impair localization of FtsZ into the Z-ring and inhibit bacterial cell division. The chlorinated analogue UCM53 inhibits the growth of clinical isolates of antibiotic-resistant Staphylococcus aureus and Enterococcus faecalis. We suggest that these interfacial inhibitors recapitulate binding and some assembly-inducing effects of GTP but impair the correct structural dynamics of FtsZ filaments and thus inhibit bacterial division, possibly by binding to a small fraction of the FtsZ molecules in a bacterial cell, which opens a new approach to FtsZ-based antibacterial drug discovery.This work was supported by grants from Plan Nacional de Investigación BFU 2011-23416 (J.M.A.), BFU2099-09552 (P.C.), and SAF2010-22198 (M.L.L.-R.), grant CM S2010/BMD-2353 (M.L.L.-R, P.C., J.M.A.), and fellowships FPI (L.B.R.-A.), FPU (M.A.) and CSIC-JAE (E.R.-A.)

Targeting bacterial cell division protein FtsZ with small molecules and fluorescent probes

Trabajo presentado en el 248th National Meeting of the American-Chemical-Society (ACS), celebrado en San Francisco, CA (Estados Unidos), del 10 al 14 de agosto de 201

The cannabinoid WIN55212-2 restores rhinovirus-induced epithelial barrier disruption

Carta al Editor. Received: 19 September 2020 | Revised: 20 November 2020 | Accepted: 5 December 2020Sección Deptal. de Química Orgánica (Óptica y Optometría)Fac. de Óptica y OptometríaTRUEMinisterio de Economía y Competitividad de España (MINECO)Swiss National Science FoundationChristine Kühne‐Center for Allergy Research and Education (CK‐CARE) (Suiza)Universidad Complutense de Madrid (España)inpres

Cannabinoid WIN55212-2 impairs peanut-allergic sensitization and promotes the generation of allergen-specific regulatory T cells

Background: Cannabinoids are lipid-derived mediators with anti-inflammatory prop-erties in different diseases. WIN55212-2, a non-selective synthetic cannabinoid, re-duces immediate anaphylactic reactions in a mouse model of peanut allergy, but its capacity to prevent peanut-allergic sensitization and the underlying mechanisms re-mains largely unknown. Objective: To investigate the capacity of WIN55212-2 to immunomodulate peanut- stimulated human dendritic cells (DCs) and peanut-allergic sensitization in mice. Methods: Surface markers and cytokines were quantified by flow cytometry, ELISA and qPCR in human monocyte-derived DCs (hmoDCs) and T-cell cocultures after stimulation with peanut alone or in the presence of WIN55212-2. Mice were epicuta-neously sensitized with peanut alone or peanut/WIN55212-2. After peanut challenge, drop in body temperature, haematocrit, clinical symptoms, peanut-specific antibodies in serum and FOXP3+ regulatory (Treg) cells in spleen and lymph nodes were quanti-fied. Splenocytes were stimulated in vitro with peanut to analyse allergen-specific T- cell responses. Results: WIN55212-2 reduced peanut-induced hmoDC activation and promoted the generation of CD4+CD127−CD25+FOXP3+ Treg cells, while reducing the induction of IL- 5- producing T cells. In vivo, WIN55212-2 impaired the peanut-induced migration of DCs to lymph nodes and their maturation. WIN55212-2 significantly reduced the induction of peanut-specific IgE and IgG1 antibodies in serum during epicutaneous peanut sensitization, reduced the clinical symptoms score upon peanut challenge and promoted the generation of allergen-specific FOXP3+ Treg cells. Conclusions: The synthetic cannabinoid WIN55212-2 interferes with peanut sensi-tization and promotes tolerogenic responses, which might well pave the way for the development of novel prophylactic and therapeutic strategies for peanut allergy

Cannabinoids induce functional Tregs by promoting tolerogenic DCs via autophagy and metabolic reprograming

The generation of functional regulatory T cells (Tregs) is essential to keep tissue homeostasis and restore healthy immune responses in many biological and inflammatory contexts. Cannabinoids have been pointed out as potential therapeutic tools for several diseases. Dendritic cells (DCs) express the endocannabinoid system, including the cannabinoid receptors CB1 and CB2. However, how cannabinoids might regulate functional properties of DCs is not completely understood. We uncover that the triggering of cannabinoid receptors promote human tolerogenic DCs that are able to prime functional FOXP3+ Tregs in the context of different inflammatory diseases. Mechanistically, cannabinoids imprint tolerogenicity in human DCs by inhibiting NF-κB, MAPK and mTOR signalling pathways while inducing AMPK and functional autophagy flux via CB1- and PPARα-mediated activation, which drives metabolic rewiring towards increased mitochondrial activity and oxidative phosphorylation. Cannabinoids exhibit in vivo protective and anti-inflammatory effects in LPS-induced sepsis and also promote the generation of FOXP3+ Tregs. In addition, immediate anaphylactic reactions are decreased in peanut allergic mice and the generation of allergen-specific FOXP3+ Tregs are promoted, demonstrating that these immunomodulatory effects take place in both type 1- and type 2-mediated inflammatory diseases. Our findings might open new avenues for novel cannabinoid-based interventions in different inflammatory and immune-mediated diseases

Isoprenylcysteine Carboxylmethyltransferase-Based Therapy for Hutchinson-Gilford Progeria Syndrome.

Hutchinson-Gilford progeria syndrome (HGPS, progeria) is a rare genetic disease characterized by premature aging and death in childhood for which there were no approved drugs for its treatment until last November, when lonafarnib obtained long-sought FDA approval. However, the benefits of lonafarnib in patients are limited, highlighting the need for new therapeutic strategies. Here, we validate the enzyme isoprenylcysteine carboxylmethyltransferase (ICMT) as a new therapeutic target for progeria with the development of a new series of potent inhibitors of this enzyme that exhibit an excellent antiprogeroid profile. Among them, compound UCM-13207 significantly improved the main hallmarks of progeria. Specifically, treatment of fibroblasts from progeroid mice with UCM-13207 delocalized progerin from the nuclear membrane, diminished its total protein levels, resulting in decreased DNA damage, and increased cellular viability. Importantly, these effects were also observed in patient-derived cells. Using the Lmna G609G/G609G progeroid mouse model, UCM-13207 showed an excellent in vivo efficacy by increasing body weight, enhancing grip strength, extending lifespan by 20%, and decreasing tissue senescence in multiple organs. Furthermore, UCM-13207 treatment led to an improvement of key cardiovascular hallmarks such as reduced progerin levels in aortic and endocardial tissue and increased number of vascular smooth muscle cells (VSMCs). The beneficial effects go well beyond the effects induced by other therapeutic strategies previously reported in the field, thus supporting the use of UCM-13207 as a new treatment for progeria.This work was supported by grants from The Progeria Research Foundation (PRF 2016-65) and the Spanish MINECO (PID2019-106279RB-I00, PID2019-108489RBI00). The authors thank Fundación La Caixa (A.G.), CEI Moncloa (N.I.M.-R.), MINECO (F.J.O.-N. and M.B.) and Ministerio de Ciencia, Innovación y Universidades (N.K.-F.) for predoctoral fellowships. The authors thank C. López-Otín for kindly donating LmnaG609G/G609G progeroid and their corresponding wild-type fibroblasts and UCM’s CAIs Cytometry and Fluorescence Microscopy, Genomics, NMR, and Mass Spectrometry, for their assistance. The CNIC is supported by the Ministerio de Ciencia e Innovación, the Instituto de Salud Carlos III, and the pro-CNIC Foundation, and is a Severo Ochoa Center of Excellence (grant SEV-2015- 0505). The generation of the antiprogerin antibody was funded by the Wellcome Trust (098291/Z/12/Z to S.N.).S

The structural assembly switch of cell division protein FtsZ probed with fluorescent allosteric inhibitors

FtsZ is a widely conserved tubulin-like GTPase that directs bacterial cell division and a new target for antibiotic discovery. This protein assembly machine cooperatively polymerizes forming single-stranded filaments, by means of self-switching between inactive and actively associating monomer conformations. The structural switch mechanism was proposed to involve a movement of the C-terminal and N-terminal FtsZ domains, opening a cleft between them, allosterically coupled to the formation of a tight association interface between consecutive subunits along the filament. The effective antibacterial benzamide PC190723 binds into the open interdomain cleft and stabilizes FtsZ filaments, thus impairing correct formation of the FtsZ ring for cell division. We have designed fluorescent analogs of PC190723 to probe the FtsZ structural assembly switch. Among them, nitrobenzoxadiazole probes specifically bind to assembled FtsZ rather than to monomers. Probes with several spacer lengths between the fluorophore and benzamide moieties suggest a binding site extension along the interdomain cleft. These probes label FtsZ rings of live Bacillus subtilis and Staphylococcus aureus, without apparently modifying normal cell morphology and growth, but at high concentrations they induce impaired bacterial division phenotypes typical of benzamide antibacterials. During the FtsZ assembly-disassembly process, the fluorescence anisotropy of the probes changes upon binding and dissociating from FtsZ, thus reporting open and closed FtsZ interdomain clefts. Our results demonstrate the structural mechanism of the FtsZ assembly switch, and suggest that the probes bind into the open clefts in cellular FtsZ polymers preferably to unassembled FtsZ in the bacterial cytosol

Targetting assembly of cell division protein FtsZ with small molecules

9 páginas, 4 figuras, 1 tabla -- PAGS nros. 269-277FtsZ is the key protein of bacterial cell division and an emergent target for new antibiotics. It is a filament-forming GTPase and a structural homologue of eukaryotic tubulin. A number of FtsZ-interacting compounds have been reported, some of which have powerful antibacterial activity. Here we review recent advances and new approaches in modulating FtsZ assembly with small molecules. This includes analyzing their chemical features, binding sites, mechanisms of action, the methods employed, and computational insights, aimed at a better understanding of their molecular recognition by FtsZ and at rational antibiotic design. Cell division protein FtsZ is employed by most bacteria to divide. It forms the cytokinetic Z-ring at the division site(1) (Figure 1), is tethered to the inner face of the plasma membrane by FtsA and ZipA, and recruits other accessory proteins of the cell division machinery (divisome), several of which are essential for remodeling the cell wall peptidoglycan at the septum.(2, 3) The assembly of the Z-ring is coordinated with DNA segregation and cell growth through many regulatory proteins in different bacteria.(2) The bacterial division proteins differ from the proteins of eukaryotic cytokinesis. Thus, bacterial cytoskeleton and cell division have been recognized as attractive targets for seeking new antibiotics(4, 5) with which to fight the widespread emergence of pathogens resistant to current antibiotics.(6) FtsZ has been validated as a target for antibacterial intervention with a synthetic compound active on an in vivo model of infection.(7) Very recently, it has been found that antibiotic acyldepsipeptides activate bacterial ClpP peptidase to degrade FtsZ.(8) Here we review small molecule approaches for targeting FtsZ assemblyWork in the authors’ laboratories has been supported by grants MICINN BFU2008-00013, BFU2009-09552 and SAF2010-22198 and grants CAM S-BIO-0214-2006 and S-SAL-249-2006Peer reviewe