ZINC is a free public resource for ligand discovery. The database contains over twenty million commercially available molecules in biologically relevant representations that may be downloaded in ...popular ready-to-dock formats and subsets. The Web site also enables searches by structure, biological activity, physical property, vendor, catalog number, name, and CAS number. Small custom subsets may be created, edited, shared, docked, downloaded, and conveyed to a vendor for purchase. The database is maintained and curated for a high purchasing success rate and is freely available at zinc.docking.org.
Molecular docking remains an important tool for structure-based screening to find new ligands and chemical probes. As docking ambitions grow to include new scoring function terms, and to address ever ...more targets, the reliability and extendability of the orientation sampling, and the throughput of the method, become pressing. Here we explore sampling techniques that eliminate stochastic behavior in DOCK3.6, allowing us to optimize the method for regularly variable sampling of orientations. This also enabled a focused effort to optimize the code for efficiency, with a three-fold increase in the speed of the program. This, in turn, facilitated extensive testing of the method on the 102 targets, 22,805 ligands and 1,411,214 decoys of the Directory of Useful Decoys-Enhanced (DUD-E) benchmarking set, at multiple levels of sampling. Encouragingly, we observe that as sampling increases from 50 to 500 to 2000 to 5000 to 20,000 molecular orientations in the binding site (and so from about 1×10(10) to 4×10(10) to 1×10(11) to 2×10(11) to 5×10(11) mean atoms scored per target, since multiple conformations are sampled per orientation), the enrichment of ligands over decoys monotonically increases for most DUD-E targets. Meanwhile, including internal electrostatics in the evaluation ligand conformational energies, and restricting aromatic hydroxyls to low energy rotamers, further improved enrichment values. Several of the strategies used here to improve the efficiency of the code are broadly applicable in the field.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Proteins fluctuate between alternative conformations, which presents a challenge for ligand discovery because such flexibility is difficult to treat computationally owing to problems with ...conformational sampling and energy weighting. Here we describe a flexible docking method that samples and weights protein conformations using experimentally derived conformations as a guide. The crystallographically refined occupancies of these conformations, which are observable in an apo receptor structure, define energy penalties for docking. In a large prospective library screen, we identified new ligands that target specific receptor conformations of a cavity in cytochrome c peroxidase, and we confirm both ligand pose and associated receptor conformation predictions by crystallography. The inclusion of receptor flexibility led to ligands with new chemotypes and physical properties. By exploiting experimental measures of loop and side-chain flexibility, this method can be extended to the discovery of new ligands for hundreds of targets in the Protein Data Bank for which similar experimental information is available.
G protein-coupled receptors (GPCRs) are intensely studied as drug targets and for their role in signaling. With the determination of the first crystal structures, interest in structure-based ligand ...discovery increased. Unfortunately, for most GPCRs no experimental structures are available. The determination of the D(3) receptor structure and the challenge to the community to predict it enabled a fully prospective comparison of ligand discovery from a modeled structure versus that of the subsequently released crystal structure. Over 3.3 million molecules were docked against a homology model, and 26 of the highest ranking were tested for binding. Six had affinities ranging from 0.2 to 3.1 μM. Subsequently, the crystal structure was released and the docking screen repeated. Of the 25 compounds selected, five had affinities ranging from 0.3 to 3.0 μM. One of the new ligands from the homology model screen was optimized for affinity to 81 nM. The feasibility of docking screens against modeled GPCRs more generally is considered.
Pathological remodeling of the heart is a hallmark of chronic heart failure (HF) and these structural changes further perpetuate the disease. Cardiac fibroblasts are the critical cell type that is ...responsible for maintaining the structural integrity of the heart. Stress conditions, such as a myocardial infarction (MI), can activate quiescent fibroblasts into synthetic and contractile myofibroblasts. G protein-coupled receptor kinase 5 (GRK5) is an important mediator of cardiovascular homeostasis through dampening of GPCR signaling, and is expressed in the heart and up-regulated in human HF. Of note, GRK5 has been demonstrated to translocate to the nucleus in cardiomyocytes in a calcium-calmodulin (Ca
-CAM)-dependent manner, promoting hypertrophic gene transcription through activation of nuclear factor of activated T cells (NFAT). Interestingly, NFAT is also involved in fibroblast activation. GRK5 is highly expressed and active in cardiac fibroblasts; however, its pathophysiological role in these crucial cardiac cells is unknown. We demonstrate using adult cardiac fibroblasts that genetic deletion of GRK5 inhibits angiotensin II (AngII)-mediated fibroblast activation. Fibroblast-specific deletion of GRK5 in mice led to decreased fibrosis and cardiac hypertrophy after chronic AngII infusion or after ischemic injury compared to nontransgenic littermate controls (NLCs). Mechanistically, we show that nuclear translocation of GRK5 is involved in fibroblast activation. These data demonstrate that GRK5 is a regulator of fibroblast activation in vitro and cardiac fibrosis in vivo. This adds to previously published data which demonstrate the potential beneficial effects of GRK5 inhibition in the context of cardiac disease.
Lead generation is a major hurdle in small-molecule drug discovery, with an estimated 60% of projects failing from lack of lead matter or difficulty in optimizing leads for drug-like properties. It ...would be valuable to identify these less-druggable targets before incurring substantial expenditure and effort. Here we show that a model-based approach using basic biophysical principles yields good prediction of druggability based solely on the crystal structure of the target binding site. We quantitatively estimate the maximal affinity achievable by a drug-like molecule, and we show that these calculated values correlate with drug discovery outcomes. We experimentally test two predictions using high-throughput screening of a diverse compound collection. The collective results highlight the utility of our approach as well as strategies for tackling difficult targets.
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Heart failure (HF) is a global epidemic with the highest degree of mortality and morbidity of any disease presently studied. G protein-coupled receptors (GPCRs) are prominent ...regulators of cardiovascular function. Activated GPCRs are “turned off” by GPCR kinases (GRKs) in a process known as “desensitization”. GRKs 2 and 5 are highly expressed in the heart, and known to be upregulated in HF.
Over the last 20 years, both GRK2 and GRK5 have been demonstrated to be critical mediators of the molecular alterations that occur in the failing heart. In the present review, we will highlight recent findings that further characterize “non-canonical” GRK signaling observed in HF. Further, we will also present potential therapeutic strategies (i.e. small molecule inhibition, microRNAs, gene therapy) that may have potential in combating the deleterious effects of GRKs in HF.
In their study, Morell et al provide a thoughtful and systematic analysis of risk of mortality in 1,709 neonates, children, and young adults (2,602 medical admissions) with PH who received care in 38 ...North American pediatric cardiac ICUs (PCICU) between 2014 and 2019. Their findings lend granularity, a new venue, and eye-opening follow up to previous cautionary notes in the literature regarding the fragility of children with pulmonary vascular diseases of various etiologies. However, details of the specific vasoactive agents that were used in the cases under study are not listed in detail. In summary, Morell et al, in what is a first look at patients with PH admitted to PCICUs, bring to light the challenges and risks these patients face as a result of their PH. This work shows the formidable effect pulmonary vascular disease can have on outcomes and the role various therapeutic interventions may have on mortality risk.
Pulmonary hypertension (PH), an often unrelenting disease that carries with it significant morbidity and mortality, affects not only the pulmonary vasculature but, in turn, the right ventricle as ...well. The survival of patients with PH is closely related to the right ventricular function. Therefore, having an understanding of how to manage right ventricular failure (RVF) and acute pulmonary hypertensive crises is imperative for clinicians who encounter these patients. This review addresses the management of these patients in detail, addressing: (a) the pathophysiology of RVF, (b) intensive care monitoring of these patients in the intensive care unit, (c) imaging of the right ventricle, (d) intubation and mechanical ventilation, (e) inotrope and vasopressor selection, (f) pulmonary vasodilator use, (g) interventional and surgical procedures for the acutely failing right ventricle, and (h) mechanical support for RVF.
BACKGROUND:Cardiac hypertrophic growth is mediated by robust changes in gene expression and changes that underlie the increase in cardiomyocyte size. The former is regulated by RNA polymerase II (pol ...II) de novo recruitment or loss; the latter involves incremental increases in the transcriptional elongation activity of pol II that is preassembled at the transcription start site. The differential regulation of these distinct processes by transcription factors remains unknown. Forkhead box protein O1 (FoxO1) is an insulin-sensitive transcription factor that is also regulated by hypertrophic stimuli in the heart. However, the scope of its gene regulation remains unexplored.
METHODS:To address this, we performed FoxO1 chromatin immunoprecipitation–deep sequencing in mouse hearts after 7 days of isoproterenol injections (3 mg·kg·mg), transverse aortic constriction, or vehicle injection/sham surgery.
RESULTS:Our data demonstrate increases in FoxO1 chromatin binding during cardiac hypertrophic growth, which positively correlate with extent of hypertrophy. To assess the role of FoxO1 on pol II dynamics and gene expression, the FoxO1 chromatin immunoprecipitation–deep sequencing results were aligned with those of pol II chromatin immunoprecipitation–deep sequencing across the chromosomal coordinates of sham- or transverse aortic constriction–operated mouse hearts. This uncovered that FoxO1 binds to the promoters of 60% of cardiac-expressed genes at baseline and 91% after transverse aortic constriction. FoxO1 binding is increased in genes regulated by pol II de novo recruitment, loss, or pause-release. In vitro, endothelin-1– and, in vivo, pressure overload–induced cardiomyocyte hypertrophic growth is prevented with FoxO1 knockdown or deletion, which was accompanied by reductions in inducible genes, including Comtd1 in vitro and Fstl1 and Uck2 in vivo.
CONCLUSIONS:Together, our data suggest that FoxO1 may mediate cardiac hypertrophic growth via regulation of pol II de novo recruitment and pause-release; the latter represents the majority (59%) of FoxO1-bound, pol II–regulated genes after pressure overload. These findings demonstrate the breadth of transcriptional regulation by FoxO1 during cardiac hypertrophy, information that is essential for its therapeutic targeting.
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