Gregory D Cuny

University of California, Santa Barbara, Santa Barbara, California, United States

Are you Gregory D Cuny?

Claim your profile

Publications (101)405.6 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Inosine 5'-monophosphate dehydrogenase (IMPDH) catalyzes the pivotal step in guanine nucleotide biosynthesis. IMPDH is a target for immunosuppressive, antiviral, and anticancer drugs, but, as of yet, has not been exploited for antimicrobial therapy. We have previously reported potent inhibitors of IMPDH from the protozoan parasite Cryptosporidium parvum (CpIMPDH). Many pathogenic bacteria, including Bacillus anthracis, Staphylococcus aureus, and Listeria monocytogenes, contain IMPDHs that should also be inhibited by these compounds. Herein, we present the structure-activity relationships for the inhibition of B. anthracis IMPDH (BaIMPDH) and antibacterial activity of 140 compounds from five structurally distinct compound series. Many potent inhibitors of BaIMPDH were identified (78% with IC50 ≤ 1 μM). Four compounds had minimum inhibitory concentrations (MIC) of less than 2 μM against B. anthracis Sterne 770. These compounds also displayed antibacterial activity against S. aureus and L. monocytogenes.
    ACS Medicinal Chemistry Letters 08/2014; 5(8):846-50. · 3.31 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: There are currently no effective therapies for fibrodysplasia ossificans progressiva (FOP), a debilitating and progressive heterotopic ossification disease caused by activating mutations of ACVR1 encoding the BMP type I receptor kinase ALK2. Recently a subset of these same mutations of ACVR1 have been identified in diffuse intrinsic pontine glioma (DIPG) tumors. Here we describe the structure activity relationship for a series of novel ALK2 inhibitors based on the 2-aminopyridine compound K02288. Several modifications increased potency in kinase, thermal shift, or cell-based assays of BMP signaling and transcription, as well as selectivity for ALK2 versus closely related BMP and TGF-β type I receptor kinases. Compounds in this series exhibited a wide range of in vitro cytotoxicity that was not correlated with potency or selectivity, suggesting mechanisms independent of BMP or TGF-β inhibition. The study also highlighted a potent 2-methylpyridine derivative 10 (LDN-214117) with high degree of selectivity for ALK2 and low cytotoxicity that could provide a template for pre-clinical development. Contrary to the notion that activating mutations of ALK2 might alter inhibitor efficacy due to potential conformational changes in the ATP-binding site, the compounds demonstrated consistent binding to a panel of mutant and wild-type ALK2 proteins. Thus, BMP inhibitors identified via activity against wild-type ALK2 signaling are likely to be of clinical relevance for the diverse ALK2 mutant proteins associated with FOP and DIPG.
    Journal of medicinal chemistry. 08/2014;
  • Source
    08/2014;
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: STEP (STriatal-Enriched protein tyrosine Phosphatase) is a neuron-specific phosphatase that regulates N-methyl-D-aspartate receptor (NMDAR) and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) trafficking, as well as ERK1/2, p38, Fyn, and Pyk2 activity. STEP is overactive in several neuropsychiatric and neurodegenerative disorders, including Alzheimer's disease (AD). The increase in STEP activity likely disrupts synaptic function and contributes to the cognitive deficits in AD. AD mice lacking STEP have restored levels of glutamate receptors on synaptosomal membranes and improved cognitive function, results that suggest STEP as a novel therapeutic target for AD. Here we describe the first large-scale effort to identify and characterize small-molecule STEP inhibitors. We identified the benzopentathiepin 8-(trifluoromethyl)-1,2,3,4,5-benzopentathiepin-6-amine hydrochloride (known as TC-2153) as an inhibitor of STEP with an IC50 of 24.6 nM. TC-2153 represents a novel class of PTP inhibitors based upon a cyclic polysulfide pharmacophore that forms a reversible covalent bond with the catalytic cysteine in STEP. In cell-based secondary assays, TC-2153 increased tyrosine phosphorylation of STEP substrates ERK1/2, Pyk2, and GluN2B, and exhibited no toxicity in cortical cultures. Validation and specificity experiments performed in wild-type (WT) and STEP knockout (KO) cortical cells and in vivo in WT and STEP KO mice suggest specificity of inhibitors towards STEP compared to highly homologous tyrosine phosphatases. Furthermore, TC-2153 improved cognitive function in several cognitive tasks in 6- and 12-mo-old triple transgenic AD (3xTg-AD) mice, with no change in beta amyloid and phospho-tau levels.
    PLoS biology. 08/2014; 12(8):e1001923.
  • [Show abstract] [Hide abstract]
    ABSTRACT: PERK is serine/threonine kinase localized to the endoplasmic reticulum (ER) membrane. PERK is activated and contributes to cell survival in response to a variety of physiological stresses that affect protein quality control in the ER, such as hypoxia, glucose depravation, increased lipid biosynthesis, and increased protein translation. Pro-survival functions of PERK are triggered by such stresses, suggesting that development of small-molecule inhibitors of PERK may be efficacious in a variety of disease scenarios. Hence, we have conducted a detailed enzymatic characterization of the PERK kinase to develop a high-throughput-screening assay (HTS) that will permit the identification of small-molecule PERK inhibitors. In addition to establishing the Km of PERK for both its primary substrate, eIF2α, and for adenosine triphosphate, further mechanistic studies revealed that PERK targets its substrate via either a random/steady-state ordered mechanism. For HTS, we developed a time-resolved fluorescence resonance energy transfer-based assay that yielded a robust Z' factor and percent coefficient of variation value, enabling the successful screening of 79,552 compounds. This approach yielded one compound that exhibited good in vitro and cellular activity. These results demonstrate the validity of this screen and represent starting points for drug discovery efforts.
    Journal of Biomolecular Screening 03/2014; · 2.21 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Glial glutamate transporter EAAT2 plays a major role in glutamate clearance in synaptic clefts. Several lines of evidence indicate that strategies designed to increase EAAT2 expression have potential for preventing excitotoxicity, which contributes to neuronal injury and death in neurodegenerative diseases. We previously discovered several classes of compounds that can increase EAAT2 expression through translational activation. Here, we present efficacy studies of the compound LDN/OSU-0212320, which is a pyridazine derivative from one of our lead series. In a murine model, LDN/OSU-0212320 had good potency, adequate pharmacokinetic properties, no observed toxicity at the doses examined, and low side effect/toxicity potential. Additionally, LDN/OSU-0212320 protected cultured neurons from glutamate-mediated excitotoxic injury and death via EAAT2 activation. Importantly, LDN/OSU-0212320 markedly delayed motor function decline and extended lifespan in an animal model of amyotrophic lateral sclerosis (ALS). We also found that LDN/OSU-0212320 substantially reduced mortality, neuronal death, and spontaneous recurrent seizures in a pilocarpine-induced temporal lobe epilepsy model. Moreover, our study demonstrated that LDN/OSU-0212320 treatment results in activation of PKC and subsequent Y-box-binding protein 1 (YB-1) activation, which regulates activation of EAAT2 translation. Our data indicate that the use of small molecules to enhance EAAT2 translation may be a therapeutic strategy for the treatment of neurodegenerative diseases.
    The Journal of clinical investigation 02/2014; · 15.39 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Cryptosporidium parasites are a major cause of diarrhea and malnutrition in the developing world, a frequent cause of waterborne disease in the developed world and a potential bioterrorism agent. Currently available treatment is limited and Cryptosporidium drug discovery remains largely unsuccessful. As a result, the pharmacokinetic properties required for in vivo efficacy have not been established. We have been engaged in a Cryptosporidium drug discovery program targeting inosine 5' -monophosphate dehydrogenase (CpIMPDH). Here we report the activity of eight potent and selective inhibitors of CpIMPDH in the IL-12 knockout mouse model, which mimics acute human cryptosporidiosis. Two compounds displayed significant antiparasitic activity, validating CpIMPDH as a drug target. The best compound, P131 (250 mg/kg-day), performed equivalently to paromomycin (2000 mg/kg-day) when administered in a single dose, and better than paromomycin when administered in three daily doses. One compound, A110, appeared to promote Cryptosporidium infection. The pharmacokinetic, uptake and permeability properties of the eight compounds were measured. P131 had the lowest systemic distribution, but accumulated to high concentrations within intestinal cells. A110 had the highest systemic distribution. These observations suggest that systemic distribution is not required, and may be a liability, for in vivo antiparasitic activity. Intriguingly, A110 caused specific alterations in fecal microbiota that were not observed with P131 or vehicle alone. Such changes may explain how A110 promotes parasitemia. Collectively, these observations suggest a blueprint for the development of anticryptosporidial therapy.
    Antimicrobial Agents and Chemotherapy 12/2013; · 4.57 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The photo-stimulated cyclization of 2-(2-halophenylamino)pyridines in liquid ammonia afforded pyrido[1,2-a]benzimidazoles via SRN1 mediated C–N bond forming reactions in moderate to excellent yields (58–94%). This general synthetic strategy was also extended to a 2-(2-bromophenylamino)pyrazine to give pyrazino[1,2-a]benzimidazole. Attempts to employ this reaction using N-(2-chlorophenyl)-3-isoquinolinamine, however, resulted in C–C bond formation generating 7H-indolo[2,3-c]isoquinoline.
    ChemInform 07/2013; 69(26):5487–5494.
  • Xiao Wang, Gregory D Cuny, Timothy Noël
    [Show abstract] [Hide abstract]
    ABSTRACT: Visible advance: A mild, one-pot Stadler-Ziegler process for CS bond formation has been developed. The method employs the photoredox catalyst [Ru(bpy)3 Cl2 ]⋅6 H2 O irradiated with visible light. A variety of aryl-alkyl and diaryl sulfides were prepared from readily available arylamines and aryl/alkylthiols in good yields. The use of a photo microreactor led to a significant improvement with respect to safety and efficiency.
    Angewandte Chemie International Edition 06/2013; · 11.34 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Spinal muscular atrophy (SMA) is a neurodegenerative disease that causes progressive muscle weakness, which primarily targets proximal muscles. About 95% of SMA cases are caused by the loss of both copies of the SMN1 gene. SMN2 is a nearly identical copy of SMN1, which expresses much less functional SMN protein. SMN2 is unable to fully compensate for the loss of SMN1 in motor neurons but does provide an excellent target for therapeutic intervention. Increased expression of functional full-length SMN protein from the endogenous SMN2 gene should lessen disease severity. We have developed and implemented a new high-throughput screening assay to identify small molecules that increase the expression of full-length SMN from a SMN2 reporter gene. Here, we characterize two novel compounds that increased SMN protein levels in both reporter cells and SMA fibroblasts and show that one increases lifespan, motor function, and SMN protein levels in a severe mouse model of SMA.
    EMBO Molecular Medicine 06/2013; · 7.80 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Although tau accumulation is a feature of several neurodegenerative conditions treatment options for these conditions are non-existent. Targeting tau kinases represents a potential therapeutic approach. Small molecules in the diaminothiazole class are potent tau kinase inhibitors that target CDK5 and GSK3β. Lead compounds from the series have IC50 values toward CDK5/p25 and GSK3β in the low nanomolar range and no observed toxicity in the therapeutic dose range. Neuronal protective effects and decreased PHF-1 immunoreactivity were observed in two animal models, 3xTg-AD and CK-p25. Treatment nearly eliminated sarkosyl-insoluble tau with the most prominent effect on the phosphorylation at Ser404. Treatment also induced the recovery of memory in a fear conditioning assay. Given the contribution of both CDK5/p25 and GSK3β to tau phosphorylation, effective treatment of tauopathies may require dual kinase targeting.
    Journal of Biological Chemistry 06/2013; · 4.65 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Cryptosporidium parvum is an enteric protozoan parasite that has emerged as a major cause of diarrhea, malnutrition, and gastroenteritis and poses a potential bioterrorism threat. C. parvum synthesizes guanine nucleotides from host adenosine in a streamlined pathway that relies on inosine 5'-monophosphate dehydrogenase (IMPDH). We have previously identified several parasite-selective C. parvum IMPDH (CpIMPDH) inhibitors by high-throughput screening. In this paper, we report the structure-activity relationship (SAR) for a series of benzoxazole derivatives with many compounds demonstrating CpIMPDH IC50 values in the nanomolar range and >500-fold selectivity over human IMPDH (hIMPDH). Unlike previously reported CpIMPDH inhibitors, these compounds are competitive inhibitors versus NAD(+). The SAR study reveals that pyridine and other small heteroaromatic substituents are required at the 2-position of the benzoxazole for potent inhibitory activity. In addition, several other SAR conclusions are highlighted with regard to the benzoxazole and the amide portion of the inhibitor, including preferred stereochemistry. An X-ray crystal structure of a representative E·IMP·inhibitor complex is also presented. Overall, the secondary amine derivative 15a demonstrated excellent CpIMPDH inhibitory activity (IC50 = 0.5 ± 0.1 nM) and moderate stability (t1/2 = 44 min) in mouse liver microsomes. Compound 73, the racemic version of 15a, also displayed superb antiparasitic activity in a Toxoplasma gondii strain that relies on CpIMPDH (EC50 = 20 ± 20 nM), and selectivity versus a wild-type T. gondii strain (200-fold). No toxicity was observed (LD50 > 50 μM) against a panel of four mammalian cells lines.
    Journal of Medicinal Chemistry 05/2013; · 5.61 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The bone morphogenetic protein (BMP) signaling pathway has essential functions in development, homeostasis, and in the normal and pathophysiologic remodeling of tissues. Small molecule inhibitors of the BMP receptor kinase family have been useful for probing physiologic functions of BMP signaling in vitro and in vivo, and may have roles in the treatment of BMP-mediated diseases. Here we describe the development of a selective and potent inhibitor of the BMP type I receptor kinases, LDN-212854, which in contrast to previously described BMP receptor kinase inhibitors exhibits nearly 4 orders of selectivity for BMP versus the closely related TGF-β and Activin type I receptors. In vitro, LDN-212854 exhibits some selectivity for ALK2 in preference to other BMP type I receptors, ALK1 and ALK3, which may permit the interrogation of ALK2-mediated signaling, transcriptional activity and function. LDN-212854 potently inhibits heterotopic ossification in an inducible transgenic mutant ALK2 mouse model of fibrodysplasia ossificans progressiva. These findings represent a significant step towards developing selective inhibitors targeting individual members of the highly homologous BMP type I receptor family. Such inhibitors would provide greater resolution as probes of physiologic function, and improved selectivity against therapeutic targets.
    ACS Chemical Biology 04/2013; · 5.44 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Receptor Interacting Protein 1 (RIP1) kinase is one of the key mediators of tumor necrosis factor alpha (TNF-α) signaling and is critical for activation of necroptotic cell death. We developed a method for expression of recombinant kinase, utilizing baculovirus co-infection of Cdc37, an Hsp90 co-chaperone, and RIP1-His, followed by a two-step purification scheme. After optimization, 1-3 mg of highly purified RIP1 kinase was typically obtained from a 1 L of Sf9 cells. The recombinant protein displayed kinase activity that was blocked by RIP1 inhibitors, necrostatins. The purified protein was used to develop a simple and robust thermal shift assay for further assessment of RIP1 inhibitors.
    Protein Expression and Purification 03/2013; · 1.43 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: A number of well-known type II inhibitors (ATP non-competitive) that bind kinases in their DFG-out conformation were tested against wild-type LRRK2 and the most common Parkinson's disease-linked mutation G2019S. We found that traditional type II inhibitors exhibit surprising variability in their inhibition mechanism between wild type (WT) and the G2019S mutant of LRRK2. The type II kinase inhibitors were found to work by an ATP-competitive fashion against the G2019S mutant, whereas they appear to work by the expected non-competitive mechanism against WT. Since the G2019S mutation lies in the DXG-motif (DYG in LRRK2 but DFG in most other kinases) of the activation loop, we explored the structural consequence of the mutation on loop dynamics using an enhanced sampling method called metadynamics. The simulations suggest that the G2019S mutation stabilizes the DYG-in state of LRRK2 through a series of hydrogen bonds, leading to an increase in the conformational barrier between the active and inactive forms of the enzyme and a relative stabilization of the active form. The conformational bias toward the active form of LRRK2 mutants has two primary consequences: 1) the mutant enzyme becomes hyperactive, a known contributor to the Parkinsonian phenotype, as a consequence of being "locked" into the activated state and 2) the mutation creates an unusual allosteric pocket that can bind type II inhibitors but in an ATP competitive fashion. Our results suggest that developing type II inhibitors, which are generally considered superior to type I inhibitors due to desirable selectivity profiles, might be especially challenging for the G2019S LRRK2 mutant.
    Biochemistry 02/2013; · 3.38 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Necroptosis is a regulated form of necrotic cell death that has been implicated in the pathogenesis of various diseases including intestinal inflammation and systemic inflammatory response syndrome (SIRS). In this work, we investigated the signaling mechanisms controlled by the necroptosis mediator receptor interacting protein-1 (RIP1) kinase. We show that Akt kinase activity is critical for necroptosis in L929 cells and plays a key role in TNFα production. During necroptosis, Akt is activated in a RIP1 dependent fashion through its phosphorylation on Thr308. In L929 cells, this activation requires independent signaling inputs from both growth factors and RIP1. Akt controls necroptosis through downstream targeting of mammalian Target of Rapamycin complex 1 (mTORC1). Akt activity, mediated in part through mTORC1, links RIP1 to JNK activation and autocrine production of TNFα. In other cell types, such as mouse lung fibroblasts and macrophages, Akt exhibited control over necroptosis-associated TNFα production without contributing to cell death. Overall, our results provide new insights into the mechanism of necroptosis and the role of Akt kinase in both cell death and inflammatory regulation.
    PLoS ONE 01/2013; 8(3):e56576. · 3.73 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Growth factor signaling pathways are tightly regulated by phosphorylation and include many important kinase targets of interest for drug discovery. Small molecule inhibitors of the bone morphogenetic protein (BMP) receptor kinase ALK2 (ACVR1) are needed urgently to treat the progressively debilitating musculoskeletal disease fibrodysplasia ossificans progressiva (FOP). Dorsomorphin analogues, first identified in zebrafish, remain the only BMP inhibitor chemotype reported to date. By screening an assay panel of 250 recombinant human kinases we identified a highly selective 2-aminopyridine-based inhibitor K02288 with in vitro activity against ALK2 at low nanomolar concentrations similar to the current lead compound LDN-193189. K02288 specifically inhibited the BMP-induced Smad pathway without affecting TGF-β signaling and induced dorsalization of zebrafish embryos. Comparison of the crystal structures of ALK2 with K02288 and LDN-193189 revealed additional contacts in the K02288 complex affording improved shape complementarity and identified the exposed phenol group for further optimization of pharmacokinetics. The discovery of a new chemical series provides an independent pharmacological tool to investigate BMP signaling and offers multiple opportunities for pre-clinical development.
    PLoS ONE 01/2013; 8(4):e62721. · 3.73 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Cryptosporidium parvum (Cp) is a potential biowarfare agent and major cause of diarrhea and malnutrition. This protozoan parasite relies on inosine 5'-monophosphate dehydrogenase (IMPDH) for the production of guanine nucleotides. A CpIMPDH-selective N-aryl-3,4-dihydro-3-methyl-4-oxo-1-phthalazineacetamide inhibitor was previously identified in a high throughput screening campaign. Herein we report a structure-activity relationship study for the phthalazinone-based series that resulted in the discovery of benzofuranamide analogs that exhibit low nanomolar inhibition of CpIMPDH. In addition, the antiparasitic activity of select analogs in a Toxoplasma gondii model of C. parvum infection is also presented.
    Bioorganic & medicinal chemistry letters 12/2012; · 2.65 Impact Factor
  • Source
    Dataset: Nature Med
  • [Show abstract] [Hide abstract]
    ABSTRACT: Cryptosporidium parvum and related species are zoonotic intracellular parasites of the intestine. Cryptosporidium is a leading cause of diarrhea in small children around the world. Infection can cause severe pathology in children and immunocompromised patients. This waterborne parasite is resistant to common methods of water treatment and therefore a prominent threat to drinking and recreation water even in countries with strong water safety systems. The drugs currently used to combat these organisms are ineffective. Genomic analysis revealed that the parasite relies solely on inosine-5'-monophosphate dehydrogenase (IMPDH) for the biosynthesis of guanine nucleotides. Herein, we report a selective urea-based inhibitor of C. parvum IMPDH (CpIMPDH) identified by high-throughput screening. We performed a SAR study of these inhibitors with some analogues exhibiting high potency (IC(50) < 2 nM) against CpIMPDH, excellent selectivity >1000-fold versus human IMPDH type 2 and good stability in mouse liver microsomes. A subset of inhibitors also displayed potent antiparasitic activity in a Toxoplasma gondii model.
    Journal of Medicinal Chemistry 09/2012; 55(17):7759-71. · 5.61 Impact Factor

Publication Stats

2k Citations
405.60 Total Impact Points

Institutions

  • 2013
    • University of California, Santa Barbara
      Santa Barbara, California, United States
  • 2012–2013
    • University of Houston
      • Department of Pharmacological and Pharmaceutical Sciences
      Houston, Texas, United States
    • The Ohio State University
      • Department of Neuroscience
      Columbus, OH, United States
  • 2009–2013
    • Brandeis University
      • Department of Biology
      Waltham, MA, United States
  • 2006–2013
    • Tufts University
      • • Department of Medicine
      • • Department of Chemical and Biological Engineering
      Georgia, United States
  • 2003–2013
    • Harvard Medical School
      • • Department of Anesthesia
      • • Department of Cell Biology
      Boston, Massachusetts, United States
  • 2005–2012
    • Brigham and Women's Hospital
      • Department of Medicine
      Boston, MA, United States