-
Kyle D Konze,
Anqi Ma,
Fengling Li,
Dalia Barsyte-Lovejoy,
Trevor Parton,
Christopher J Macnevin,
Feng Liu,
Cen Gao, Xi-Ping Huang,
Ekaterina Kuznetsova, [......],
Jacqueline L Norris,
Lindsey I James,
Bryan L Roth,
Peter J Brown,
Stephen V Frye,
Cheryl H Arrowsmith,
Klaus M Hahn,
Gang Greg Wang,
Masoud Vedadi,
Jian Jin
[show abstract]
[hide abstract]
ABSTRACT: EZH2 or EZH1 is the catalytic subunit of the polycomb repressive complex 2 that catalyzes methylation of histone H3 lysine 27 (H3K27). The trimethylation of H3K27 (H3K27me3) is a transcriptionally repressive post-translational modification. Overexpression of EZH2 and hypertrimethylation of H3K27 have been implicated in a number of cancers. Several selective inhibitors of EZH2 have been reported recently. Herein we disclose UNC1999, the first orally bioavailable inhibitor that has high in vitro potency for wild-type and mutant EZH2 as well as EZH1, a closely related H3K27 methyltransferase that shares 96% sequence identity with EZH2 in their respective catalytic domains. UNC1999 was highly selective for EZH2 and EZH1 over a broad range of epigenetic and non-epigenetic targets, competitive with the cofactor SAM and non-competitive with the peptide substrate. This inhibitor potently reduced H3K27me3 levels in cells and selectively killed diffused large B cell lymphoma cell lines harboring the EZH2(Y641N) mutant. Importantly, UNC1999 was orally bioavailable in mice, making this inhibitor a valuable tool for investigating the role of EZH2 and EZH1 in chronic animal studies. We also designed and synthesized UNC2400, a close analogue of UNC1999 with potency >1,000-fold lower than that of UNC1999 as a negative control for cell-based studies. Finally, we created a biotin-tagged UNC1999 (UNC2399), which enriched EZH2 in pull-down studies, and a UNC1999-dye conjugate (UNC2239) for co-localization studies with EZH2 in live cells. Taken together, these compounds represent a set of useful tools for the biomedical community to investigate the role of EZH2 and EZH1 in health and disease.
ACS Chemical Biology 04/2013; · 6.45 Impact Factor
-
Lindsey I James,
Dalia Barsyte-Lovejoy,
Nan Zhong,
Liubov Krichevsky,
Victoria K Korboukh,
J Martin Herold,
Christopher J Macnevin,
Jacqueline L Norris,
Cari A Sagum,
Wolfram Tempel, [......],
Shili Duan,
Andrew Emili,
Jack F Greenblatt,
Dmitri B Kireev,
Jian Jin,
William P Janzen,
Peter J Brown,
Mark T Bedford,
Cheryl H Arrowsmith,
Stephen V Frye
[show abstract]
[hide abstract]
ABSTRACT: We describe the discovery of UNC1215, a potent and selective chemical probe for the methyllysine (Kme) reading function of L3MBTL3, a member of the malignant brain tumor (MBT) family of chromatin-interacting transcriptional repressors. UNC1215 binds L3MBTL3 with a K(d) of 120 nM, competitively displacing mono- or dimethyllysine-containing peptides, and is greater than 50-fold more potent toward L3MBTL3 than other members of the MBT family while also demonstrating selectivity against more than 200 other reader domains examined. X-ray crystallography identified a unique 2:2 polyvalent mode of interaction between UNC1215 and L3MBTL3. In cells, UNC1215 is nontoxic and directly binds L3MBTL3 via the Kme-binding pocket of the MBT domains. UNC1215 increases the cellular mobility of GFP-L3MBTL3 fusion proteins, and point mutants that disrupt the Kme-binding function of GFP-L3MBTL3 phenocopy the effects of UNC1215 on localization. Finally, UNC1215 was used to reveal a new Kme-dependent interaction of L3MBTL3 with BCLAF1, a protein implicated in DNA damage repair and apoptosis.
Nature Chemical Biology 01/2013; · 14.69 Impact Factor
-
Martilias S Farrell,
Ying Pei,
Yehong Wan,
Prem N Yadav,
Tanya L Daigle,
Daniel J Urban,
Hyeong-Min Lee,
Noah Sciaky,
Arkeen Simmons,
Randal J Nonneman, Xi-Ping Huang,
Sandy J Hufeisen,
Jean-Marc Guettier,
Sheryl S Moy,
Jürgen Wess,
Marc G Caron,
Nicole Calakos,
Bryan L Roth
[show abstract]
[hide abstract]
ABSTRACT: Here, we describe a newly generated transgenic mouse in which the Gs DREADD (rM3Ds), an engineered G protein-coupled receptor, is selectively expressed in striatopallidal medium spiny neurons (MSNs). We first show that in vitro, rM3Ds can couple to Gα(olf) and induce cAMP accumulation in cultured neurons and HEK-T cells. The rM3Ds was then selectively and stably expressed in striatopallidal neurons by creating a transgenic mouse in which an adenosine2A (adora2a) receptor-containing bacterial artificial chromosome was employed to drive rM3Ds expression. In the adora2A-rM3Ds mouse, activation of rM3Ds by clozapine-N-oxide (CNO) induces DARPP-32 phosphorylation, consistent with the known consequence of activation of endogenous striatal Gα(s)-coupled GPCRs. We then tested whether CNO administration would produce behavioral responses associated with striatopallidal G(s) signaling and in this regard CNO dose-dependently decreases spontaneous locomotor activity and inhibits novelty induced locomotor activity. Lastly, we show that CNO prevented behavioral sensitization to amphetamine and increased AMPAR/NMDAR ratios in transgene-expressing neurons of the nucleus accumbens shell. These studies demonstrate the utility of adora2a-rM3Ds transgenic mice for the selective and non-invasive modulation of Gα(s) signaling in specific neuronal populations in vivo. This unique tool provides a new resource for elucidating the roles of striatopallidal MSN Gα(s) signaling in other neurobehavioral contexts.Neuropsychopharmacology accepted article preview online, 5 December 2012; doi:10.1038/npp.2012.251.
Neuropsychopharmacology: official publication of the American College of Neuropsychopharmacology 12/2012; · 6.99 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The 5-HT(2C) receptor is an attractive drug target in the quest for new therapeutics to treat a variety of human disorders. We have previously undertaken a structural optimization campaign that has led to some potent and moderately selective 5-HT(2C) receptor agonists. After expanding our structure-function library, we were able to combine our datasets so as to allow the design of compounds of improved selectivity and potency. We disclose herein the structural optimization of our previously reported 5-HT(2B)/5-HT(2C) agonists, which has led to the identification of a highly selective 5-HT(2C) agonist, (+)-trans-[2-(2-cyclopropylmethoxyphenyl)cyclopropyl]methylamine hydrochloride, with an EC(50) of 55 nM and no detectable agonism at the 5-HT(2B) receptor.
ACS Medicinal Chemistry Letters 12/2011; 2(12):929-932. · 3.36 Impact Factor
-
John A Allen,
Julianne M Yost,
Vincent Setola,
Xin Chen,
Maria F Sassano,
Meng Chen,
Sean Peterson,
Prem N Yadav, Xi-ping Huang,
Bo Feng,
Niels H Jensen,
Xin Che,
Xu Bai,
Stephen V Frye,
William C Wetsel,
Marc G Caron,
Jonathan A Javitch,
Bryan L Roth,
Jian Jin
[show abstract]
[hide abstract]
ABSTRACT: Elucidating the key signal transduction pathways essential for both antipsychotic efficacy and side-effect profiles is essential for developing safer and more effective therapies. Recent work has highlighted noncanonical modes of dopamine D(2) receptor (D(2)R) signaling via β-arrestins as being important for the therapeutic actions of both antipsychotic and antimanic agents. We thus sought to create unique D(2)R agonists that display signaling bias via β-arrestin-ergic signaling. Through a robust diversity-oriented modification of the scaffold represented by aripiprazole (1), we discovered UNC9975 (2), UNC0006 (3), and UNC9994 (4) as unprecedented β-arrestin-biased D(2)R ligands. These compounds also represent unprecedented β-arrestin-biased ligands for a G(i)-coupled G protein-coupled receptor (GPCR). Significantly, UNC9975, UNC0006, and UNC9994 are simultaneously antagonists of G(i)-regulated cAMP production and partial agonists for D(2)R/β-arrestin-2 interactions. Importantly, UNC9975 displayed potent antipsychotic-like activity without inducing motoric side effects in inbred C57BL/6 mice in vivo. Genetic deletion of β-arrestin-2 simultaneously attenuated the antipsychotic actions of UNC9975 and transformed it into a typical antipsychotic drug with a high propensity to induce catalepsy. Similarly, the antipsychotic-like activity displayed by UNC9994, an extremely β-arrestin-biased D(2)R agonist, in wild-type mice was completely abolished in β-arrestin-2 knockout mice. Taken together, our results suggest that β-arrestin signaling and recruitment can be simultaneously a significant contributor to antipsychotic efficacy and protective against motoric side effects. These functionally selective, β-arrestin-biased D(2)R ligands represent valuable chemical probes for further investigations of D(2)R signaling in health and disease.
Proceedings of the National Academy of Sciences 11/2011; 108(45):18488-93. · 9.68 Impact Factor
-
Masoud Vedadi,
Dalia Barsyte-Lovejoy,
Feng Liu,
Sylvie Rival-Gervier,
Abdellah Allali-Hassani,
Viviane Labrie,
Tim J Wigle,
Peter A Dimaggio,
Gregory A Wasney,
Alena Siarheyeva, [......],
Aled Edwards,
Bryan L Roth,
William P Janzen,
Benjamin A Garcia,
Arturas Petronis,
James Ellis,
Peter J Brown,
Stephen V Frye,
Cheryl H Arrowsmith,
Jian Jin
Nature Chemical Biology 01/2011; 7(9):648. · 14.69 Impact Factor
-
Masoud Vedadi,
Dalia Barsyte-Lovejoy,
Feng Liu,
Sylvie Rival-Gervier,
Abdellah Allali-Hassani,
Viviane Labrie,
Tim J Wigle,
Peter A Dimaggio,
Gregory A Wasney,
Alena Siarheyeva, [......],
Aled Edwards,
Bryan L Roth,
William P Janzen,
Benjamin A Garcia,
Arturas Petronis,
James Ellis,
Peter J Brown,
Stephen V Frye,
Cheryl H Arrowsmith,
Jian Jin
[show abstract]
[hide abstract]
ABSTRACT: Protein lysine methyltransferases G9a and GLP modulate the transcriptional repression of a variety of genes via dimethylation of Lys9 on histone H3 (H3K9me2) as well as dimethylation of non-histone targets. Here we report the discovery of UNC0638, an inhibitor of G9a and GLP with excellent potency and selectivity over a wide range of epigenetic and non-epigenetic targets. UNC0638 treatment of a variety of cell lines resulted in lower global H3K9me2 levels, equivalent to levels observed for small hairpin RNA knockdown of G9a and GLP with the functional potency of UNC0638 being well separated from its toxicity. UNC0638 markedly reduced the clonogenicity of MCF7 cells, reduced the abundance of H3K9me2 marks at promoters of known G9a-regulated endogenous genes and disproportionately affected several genomic loci encoding microRNAs. In mouse embryonic stem cells, UNC0638 reactivated G9a-silenced genes and a retroviral reporter gene in a concentration-dependent manner without promoting differentiation.
Nature Chemical Biology 01/2011; 7(8):566-74. · 14.69 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The human Ether-à-go-go related gene (hERG) potassium channel is responsible for the rapid delayed rectifier potassium current that plays a critical role in the repolarization of cardiomyocytes during the cardiac action potential. In humans, inhibition of hERG by drugs can prolong the electrocardiographic QT interval, which, in rare instance, leads to ventricular arrhythmia and sudden cardiac death. As such, several medications that block hERG channels in vitro have been withdrawn from the market due to QT prolongation and arrhythmias. The current FDA guidelines recommend that drug candidates destined for human use be evaluated for potential hERG activity ( www.fda.gov/downloads/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/ucm074963.pdf ). Here, we employed automated planar patch clamp (APPC), high-throughput fluorescent Tl(+) flux, and moderate-throughput [³H]dofetilide competition binding assays to characterize a panel of 49 drugs for their activities at the hERG channel. Notably, we used the same HEK293-hERG cell line for all assays, facilitating comparisons of hERG potencies across screening platforms. In general, hERG inhibitors were most potent in APPC assays, intermediate potent in [³H]dofetilide binding assays, and least potent in Tl(+) flux assays. Binding affinity constants (pK(i) values) and Tl(+) flux potencies (pEC₅₀ values) correlated well with APPC pEC₅₀ values. Further, the inhibitory potencies of many known hERG inhibitors in APPC matched literature values from manual and/or automated patch clamp systems. We also developed a novel fluorescent Tl(+) flux assays to measure the effects of drugs that modulate hERG trafficking and surface expression.
Assay and Drug Development Technologies 12/2010; 8(6):727-42. · 1.73 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Cardiac cell therapy for older patients who experience a myocardial infarction may require highly regenerative cells from young, healthy (allogeneic) donors. Bone marrow mesenchymal stem cells (MSCs) are currently under clinical investigation because they can induce cardiac repair and may also be immunoprivileged (suitable for allogeneic applications). However, it is unclear whether allogeneic MSCs retain their immunoprivilege or functional efficacy late after myocardial implantation. We evaluated the effects of MSC differentiation on the immune characteristics of cells in vitro and in vivo and monitored cardiac function for 6 months after post-myocardial infarction MSC therapy.
In the in vitro experiments, inducing MSCs to acquire myogenic, endothelial, or smooth muscle characteristics (via 5-azacytidine or cytokine treatment) increased major histocompatibility complex-Ia and -II (immunogenic) expression and reduced major histocompatibility complex-Ib (immunosuppressive) expression, in association with increased cytotoxicity in coculture with allogeneic leukocytes. In the in vivo experiments, we implanted allogeneic or syngeneic MSCs into infarcted rat myocardia. We measured cell differentiation and survival (immunohistochemistry, real-time polymerase chain reaction) and cardiac function (echocardiography, pressure-volume catheter) for 6 months. MSCs (versus media) significantly improved ventricular function for at least 3 months after implantation. Allogeneic (but not syngeneic) cells were eliminated from the heart by 5 weeks after implantation, and their functional benefits were lost within 5 months.
The long-term ability of allogeneic MSCs to preserve function in the infarcted heart is limited by a biphasic immune response whereby they transition from an immunoprivileged to an immunogenic state after differentiation, which is associated with an alteration in major histocompatibility complex-immune antigen profile.
Circulation 12/2010; 122(23):2419-29. · 14.74 Impact Factor
-
Prem N Yadav,
Atheir I Abbas,
Martilias S Farrell,
Vincent Setola,
Noah Sciaky, Xi-Ping Huang,
Wesley K Kroeze,
LaTasha K Crawford,
David A Piel,
Michael J Keiser,
John J Irwin,
Brian K Shoichet,
Evan S Deneris,
Jay Gingrich,
Sheryl G Beck,
Bryan L Roth
[show abstract]
[hide abstract]
ABSTRACT: Clozapine, by virtue of its absence of extrapyramidal side effects and greater efficacy, revolutionized the treatment of schizophrenia, although the mechanisms underlying this exceptional activity remain controversial. Combining an unbiased cheminformatics and physical screening approach, we evaluated clozapine's activity at >2350 distinct molecular targets. Clozapine, and the closely related atypical antipsychotic drug olanzapine, interacted potently with a unique spectrum of molecular targets. This distinct pattern, which was not shared with the typical antipsychotic drug haloperidol, suggested that the serotonergic neuronal system was a key determinant of clozapine's actions. To test this hypothesis, we used pet1(-/-) mice, which are deficient in serotonergic presynaptic markers. We discovered that the antipsychotic-like properties of the atypical antipsychotic drugs clozapine and olanzapine were abolished in a pharmacological model that mimics NMDA-receptor hypofunction in pet1(-/-) mice, whereas haloperidol's efficacy was unaffected. These results show that clozapine's ability to normalize NMDA-receptor hypofunction, which is characteristic of schizophrenia, depends on an intact presynaptic serotonergic neuronal system.
Neuropsychopharmacology: official publication of the American College of Neuropsychopharmacology 11/2010; 36(3):638-51. · 6.99 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Autologous mesenchymal stem cells (MSCs) have been proven safe in phase I and II clinical trials in patients who have suffered a myocardial infarction. However, their potential for proliferation and differentiation decreases with age, which limits their efficacy in elderly patients. Allogeneic MSCs offer several key advantages over autologous MSCs, including a high regenerative potential and availability for clinical use without the delay required for expansion. It was believed that allogeneic MSCs were immune privileged and thus able to escape the recipient's immune system. In several preclinical studies, allogeneic MSCs were successful in regenerating the myocardium, and the transplanted MSCs improved heart function early after implantation. However, the long-term ability of allogeneic MSCs to preserve heart function is limited because of a transition from an immune privileged to an immunogenic phenotype after the cells differentiate. The initial phase I/II clinical study using allogeneic MSCs in patients with acute myocardial infarction was safe, and no side effects were observed. However, the long-term safety and efficacy of allogeneic MSCs remain to be established. In this review, we discuss the challenges of using allogeneic MSCs for cardiac repair and present strategies to prevent the immune rejection of allogeneic MSCs to increase their potential for use in cardiac patients.
Trends in cardiovascular medicine 11/2010; 20(8):263-8. · 4.37 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Injectable scaffolds made of biodegradable biomaterials can stabilize a myocardial infarct and promote cardiac repair. Here, we describe the synthesis of a new, temperature-sensitive, aliphatic polyester hydrogel (HG) conjugated with vascular endothelial growth factor (VEGF) and evaluate its effects on cardiac recovery after a myocardial infarction (MI). Seven days after coronary ligation in rats, PBS, HG, or HG mixed or conjugated with VEGF (HG + VEGF or HG-VEGF, respectively) was injected around the infarct (n = 8-11/group). Function was evaluated by echocardiography at multiple time points. Pressure-volume measurements were taken and infarct morphometry and blood vessel density were assessed at 35 days after injection. HG-VEGF provided localized, sustained VEGF function. Compared with outcomes in the PBS group, fractional shortening, ventricular volumes, preload recruitable stroke work, and end-systolic elastance were all preserved (p < 0.05) in the HG and HG + VEGF groups, and further preserved in the HG-VEGF group. Conjugated VEGF also produced the highest blood vessel density (p < 0.05). The infarct thinned and dilated after PBS injection, but was smaller and thicker in hearts treated with HG (p < 0.05). Our temperature-sensitive HG attenuated adverse cardiac remodeling and improved ventricular function when injected after an MI. VEGF delivery enhanced these effects when the VEGF was conjugated to the HG.
Biomaterials 10/2010; 32(2):579-86. · 7.40 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Some antipsychotic drugs are known to cause valvular heart disease by activating serotonin 5-HT(2B) receptors. We have developed and validated binary classification QSAR models capable of predicting potential 5-HT(2B) actives. The classification accuracies of the models built to discriminate 5-HT(2B) actives from the inactives were as high as 80% for the external test set. These models were used to screen in silico 59,000 compounds included in the World Drug Index, and 122 compounds were predicted as actives with high confidence. Ten of them were tested in radioligand binding assays and nine were found active, suggesting a success rate of 90%. All validated actives were then tested in functional assays, and one compound was identified as a true 5-HT(2B) agonist. We suggest that the QSAR models developed in this study could be used as reliable predictors to flag drug candidates that are likely to cause valvulopathy.
Journal of Medicinal Chemistry 10/2010; 53(21):7573-86. · 4.80 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Late after a myocardial infarction (MI), surgical ventricular restoration (SVR) can reduce left ventricular volumes, but an enhanced cardiac patch may be required to restore function. We developed a new, biodegradable patch (modified gelfoam, MGF) consisting of a spongy inner core (gelfoam) to encourage cell engraftment and an outer coating (poly epsilon-caprolactone) to provide sufficient strength to permit ventricular repair. Two weeks after coronary ligation in rats, SVR was performed using one of the following: gelfoam, MGF, MGF patches with hydrogel alone, or with hydrogel and cytokines (stem cell factor, stromal cell-derived factor-1alpha), bone marrow mesenchymal stem cells, or both. Cardiac function and morphology were evaluated by echocardiography, conduction catheterization, magnetic resonance imaging, and histology. Animals whose hearts were repaired with untreated gelfoam died of ventricular rupture. The MGF groups had significantly improved myocardial systolic function vs. MI controls. Enhancement with cytokines and/or cells promoted more alpha-smooth muscle actin-positive cells, more capillaries, greater wall thickness, a more ellipsoid shape, greater fractional shortening, and better-preserved systolic elastance than MGF alone. This combination of the new, reinforced, biodegradable biomaterial and cytokine/cell treatment created a viable tissue after SVR and produced better functional outcomes than un-reinforced gelfoam or MGF alone.
Biomaterials 10/2010; 31(30):7684-94. · 7.40 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The serendipitous discovery of N-cyclohexyl-8-fluoro-3,3a,4,9b-tetrahydro-1H-thiochromeno[4,3-c]isoxazole-1-carboxamide as a selective human serotonin 5-HT2B antagonist with Ki of 42+/-5 nM is reported herein. A subsequent functional assay indicated little agonist activity compared to 5-HT itself.
Bioorganic & medicinal chemistry letters 09/2010; 20(18):5488-90. · 2.65 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Iron chelators are effective at removing iron from the body in iron overload, but little is known about the handling of iron chelates by the kidney. We studied the transport of deferoxamine, deferasirox, and three hydroxypyridones, and their iron chelates, in polarized renal epithelial MDCK cells growing on Transwell inserts. Directional iron efflux was also studied in (59)Fe-loaded cells. The chelators were transported at comparable rates in the apical and basolateral directions and moved faster than their corresponding chelates, except for deferoxamine, which did not move from the basolateral to the apical side. In contrast, the chelates were transported faster in the apical-to-basolateral direction. More permeable chelators were more efficient at removing iron from iron-loaded cells compared with deferoxamine. Iron is preferentially removed from the basolateral side, and kinetic modeling suggests facilitated diffusion of chelates in some cases. Basolateral iron efflux is temperature-dependent and partially sensitive to ATP depletion. Polarized transport of chelates suggests the kidney may be involved in reabsorption of iron bound to chelators, with a temperature-sensitive facilitated removal of some iron complexes from the basolateral side. Further studies are warranted to determine if these processes may contribute to the observed nephrotoxicity of some iron chelators.
International journal of hematology 03/2010; 91(3):401-12. · 1.17 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Cardiac cell therapies offer distinct and exciting advantages over current treatments to prevent postinfarction heart failure because they can reverse ventricular remodeling and improve function, but only if the implanted stem cells contribute biological functions and achieve prolonged engraftment within the hostile environment of the damaged heart. Unfortunately, function is diminished in autologous stem cells isolated from older patients and those with comorbidities, and so clinical trials testing the implantation of healthy, allogeneic bone marrow-derived stromal cells (MSCs) isolated from young donors are currently underway. MSCs are unique because, in addition to exerting paracrine effects that restore blood flow and recruit endogenous stem cells to the infarct, they exhibit immune-modulating properties in culture that-if retained after allogeneic implantation-imply the cells may escape immune recognition within the heart. At present, the scope of MSC immune modulation after implantation is unclear.
Journal of Cardiac Surgery 02/2010; 25(3):359-66. · 0.87 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Drug-induced valvular heart disease (VHD) is a serious side effect of a few medications, including some that are on the market. Pharmacological studies of VHD-associated medications (e.g., fenfluramine, pergolide, methysergide, and cabergoline) have revealed that they and/or their metabolites are potent 5-hydroxytryptamine(2B) (5-HT(2B)) receptor agonists. We have shown that activation of 5-HT(2B) receptors on human heart valve interstitial cells in vitro induces a proliferative response reminiscent of the fibrosis that typifies VHD. To identify current or future drugs that might induce VHD, we screened approximately 2200 U.S. Food and Drug Administration (FDA)-approved or investigational medications to identify 5-HT(2B) receptor agonists, using calcium-based high-throughput screening. Of these 2200 compounds, 27 were 5-HT(2B) receptor agonists (hits); 14 of these had previously been identified as 5-HT(2B) receptor agonists, including seven bona fide valvulopathogens. Six of the hits (guanfacine, quinidine, xylometazoline, oxymetazoline, fenoldopam, and ropinirole) are approved medications. Twenty-three of the hits were then "functionally profiled" (i.e., assayed in parallel for 5-HT(2B) receptor agonism using multiple readouts to test for functional selectivity). In these assays, the known valvulopathogens were efficacious at concentrations as low as 30 nM, whereas the other compounds were less so. Hierarchical clustering analysis of the pEC(50) data revealed that ropinirole (which is not associated with valvulopathy) was clearly segregated from known valvulopathogens. Taken together, our data demonstrate that patterns of 5-HT(2B) receptor functional selectivity might be useful for identifying compounds likely to induce valvular heart disease.
Molecular pharmacology 08/2009; 76(4):710-22. · 4.53 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Inhibitors of histone deacetylases (HDACIs) have emerged as a new class of drugs for the treatment of human cancers and other diseases because of their effects on cell growth, differentiation, and apoptosis. In this study we have developed several quantitative structure-activity relationship (QSAR) models for 59 chemically diverse histone deacetylase class 1 (HDAC1) inhibitors. The variable selection k nearest neighbor (kNN) and support vector machines (SVM) QSAR modeling approaches using both MolconnZ and MOE chemical descriptors generated from two-dimensional rendering of compounds as chemical graphs have been employed. We have relied on a rigorous model development workflow including the division of the data set into training, test, and external sets and extensive internal and external validation. Highly predictive QSAR models were generated with leave-one-out cross-validated (LOO-CV) q2 and external R2 values as high as 0.80 and 0.87, respectively, using the kNN/MolconnZ approach and 0.93 and 0.87, respectively, using the SVM/MolconnZ approach. All validated QSAR models were employed concurrently for virtual screening (VS) of an in-house compound collection including 9.5 million molecules compiled from the ZINC7.0 database, the World Drug Index (WDI) database, the ASINEX Synergy libraries, and other commercial databases. VS resulted in 45 structurally unique consensus hits that were considered novel putative HDAC1 inhibitors. These computational hits had several novel structural features that were not present in the original data set. Four computational hits with novel scaffolds were tested experimentally, and three of them were confirmed active against HDAC1, with IC50 values for the most active compound of 1.00 microM. The fourth compound was later identified to be a selective inhibitor of HDAC6, a Class II HDAC. Moreover, two of the confirmed hits are marketed drugs, which could potentially facilitate their further development as anticancer agents. This study illustrates the power of the combined QSAR-VS method as a general approach for the effective identification of structurally novel bioactive compounds.
Journal of Chemical Information and Modeling 02/2009; 49(2):461-76. · 4.68 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Iron that is not specifically chaperoned through its essential functional pathways is damaging to biological systems, in major part by catalyzing the production of reactive oxygen species. Iron serves in several essential roles in the mitochondrion, as an essential cofactor for certain enzymes of electron transport, and through its involvement in the assembly of iron-sulfur clusters and iron-porphyrin (heme) complexes, both processes occurring in the mitochondrion. Therefore, there are mechanisms that deliver iron specifically to mitochondria, although these are not well understood. Under normal circumstances the mitochondrion has levels of stored iron that are higher than other organelles, though lower than in cytosol, while in some disorders of iron metabolism, mitochondrial iron levels exceed those in the cytosol. Under these circumstances of excess iron, protective mechanisms are overwhelmed and mitochondrial damage ensues. This may take the form of acute oxidative stress with structural damage and functional impairment, but also may result in long-term damage to the mitochondrial genome. This review discusses the evidence that mitochondria do indeed accumulate iron in several genetic disorders, and are a direct target for iron toxicity when it is present in excess. We then consider two classes of genetic disorders involving iron and the mitochondrion. The first include defects in genes directly regulating mitochondrial iron metabolism that lead to Friedreich's ataxia and the various sideroblastic anemias, with excessive mitochondrial iron accumulation. Under the second class, we discuss various primary hemochromatoses that lead to direct mitochondrial damage, with reference to mutations in genes encoding HFE, hepcidin, hemojuvelin, transferrin receptor-2, ferroportin, transferrin, and ceruloplasmin.
Chemico-Biological Interactions 11/2006; 163(1-2):68-76. · 2.46 Impact Factor
