RESUMO
Alterations in epigenetic silencing have been associated with ageing and tumour formation. Although substantial efforts have been made towards understanding the mechanisms of gene silencing, novel regulators in this process remain to be identified. To systematically search for components governing epigenetic silencing, we developed a genome-wide silencing screen for yeast (Saccharomyces cerevisiae) silent mating type locus HMR. Unexpectedly, the screen identified the mismatch repair (MMR) components Pms1, Mlh1, and Msh2 as being required for silencing at this locus. We further found that the identified genes were also required for proper silencing in telomeres. More intriguingly, the MMR mutants caused a redistribution of Sir2 deacetylase, from silent mating type loci and telomeres to rDNA regions. As a consequence, acetylation levels at histone positions H3K14, H3K56, and H4K16 were increased at silent mating type loci and telomeres but were decreased in rDNA regions. Moreover, knockdown of MMR components in human HEK293T cells increased subtelomeric DUX4 gene expression. Our work reveals that MMR components are required for stable inheritance of gene silencing patterns and establishes a link between the MMR machinery and the control of epigenetic silencing.
Assuntos
Proteína 1 Homóloga a MutL/genética , Proteínas MutL/genética , Proteína 2 Homóloga a MutS/genética , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Acetilação , Reparo de Erro de Pareamento de DNA , Epigênese Genética , Inativação Gênica , Genes Fúngicos Tipo Acasalamento , Hereditariedade , Histonas/metabolismo , Saccharomyces cerevisiae/genética , Proteínas Reguladoras de Informação Silenciosa de Saccharomyces cerevisiae/metabolismo , Sirtuína 2/metabolismo , Telômero/genéticaRESUMO
Cells of hematopoietic origin express high levels of the immunoproteasome, a cytokine-inducible proteasome variant comprising the proteolytic subunits LMP2 (ß1i), MECL-1 (ß2i), and LMP7 (ß5i). Targeting the immunoproteasome in pre-clinical models of autoimmune diseases with the epoxyketone inhibitor ONX 0914 has proven to be effective. ONX 0914 was previously described as a selective LMP7 inhibitor. Here, we show that PRN1126, developed as an exclusively LMP7-specific inhibitor, has limited effects on IL-6 secretion, experimental colitis, and experimental autoimmune encephalomyelitis (EAE). We demonstrate that prolonged exposure of cells with ONX 0914 leads to inhibition of both LMP7 and LMP2. Co-inhibition of LMP7 and LMP2 with PRN1126 and LMP2 inhibitors LU-001i or ML604440 impairs MHC class I cell surface expression, IL-6 secretion, and differentiation of naïve T helper cells to T helper 17 cells, and strongly ameliorates disease in experimental colitis and EAE. Hence, co-inhibition of LMP2 and LMP7 appears to be synergistic and advantageous for the treatment of autoimmune diseases.
Assuntos
Autoimunidade , Complexo de Endopeptidases do Proteassoma/imunologia , Inibidores de Proteassoma/farmacologia , Subunidades Proteicas/antagonistas & inibidores , Animais , Diferenciação Celular , Permeabilidade da Membrana Celular , Colite/imunologia , Colite/patologia , Citocinas/metabolismo , Sulfato de Dextrana , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/patologia , Epitopos/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Camundongos Endogâmicos C57BL , Complexo de Endopeptidases do Proteassoma/metabolismo , Subunidades Proteicas/imunologia , Baço/citologia , Células Th17/citologia , Células Th17/imunologiaRESUMO
Vimentin has been shown to be involved in wound healing, but its functional contribution to this process is poorly understood. Here we describe a previously unrecognized function of vimentin in coordinating fibroblast proliferation and keratinocyte differentiation during wound healing. Loss of vimentin led to a severe deficiency in fibroblast growth, which in turn inhibited the activation of two major initiators of epithelial-mesenchymal transition (EMT), TGF-ß1 signaling and the Zinc finger transcriptional repressor protein Slug, in vimentin-deficient (VIM(-/-)) wounds. Correspondingly, VIM(-/-) wounds exhibited loss of EMT-like keratinocyte activation, limited keratinization, and slow reepithelialization. Furthermore, the fibroblast deficiency abolished collagen accumulation in the VIM(-/-) wounds. Vimentin reconstitution in VIM(-/-) fibroblasts restored both their proliferation and TGF-ß1 production. Similarly, restoring paracrine TGF-ß-Slug-EMT signaling reactivated the transdifferentiation of keratinocytes, reviving their migratory properties, a critical feature for efficient healing. Our results demonstrate that vimentin orchestrates the healing by controlling fibroblast proliferation, TGF-ß1-Slug signaling, collagen accumulation, and EMT processing, all of which in turn govern the required keratinocyte activation.
Assuntos
Diferenciação Celular/genética , Proliferação de Células/genética , Fatores de Transcrição da Família Snail/genética , Fator de Crescimento Transformador beta/genética , Vimentina/genética , Cicatrização/genética , Animais , Animais Recém-Nascidos , Células Cultivadas , Transição Epitelial-Mesenquimal/genética , Feminino , Fibroblastos/citologia , Fibroblastos/metabolismo , Perfilação da Expressão Gênica/métodos , Queratinócitos/citologia , Queratinócitos/metabolismo , Masculino , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Interferência de RNA , Transdução de Sinais/genética , Fatores de Transcrição da Família Snail/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Vimentina/deficiênciaRESUMO
Fatty acid amide hydrolase (FAAH), an amidase-signature family member, is an integral membrane enzyme that degrades lipid amides including the endogenous cannabinoid anandamide and the sleep-inducing molecule oleamide. Both genetic knock out and pharmacological administration of FAAH inhibitors in rodent models result in analgesic, anxiolytic, and antiinflammatory phenotypes. Targeting FAAH activity, therefore, presents a promising new therapeutic strategy for the treatment of pain and other neurological-related or inflammatory disorders. Nearly all FAAH inhibitors known to date attain their binding potency through a reversible or irreversible covalent modification of the nucleophile Ser241 in the unusual Ser-Ser-Lys catalytic triad. Here, we report the discovery and mechanism of action of a series of ketobenzimidazoles as unique and potent noncovalent FAAH inhibitors. Compound 2, a representative of these ketobenzimidazoles, was designed from a series of ureas that were identified from high-throughput screening. While urea compound 1 is characterized as an irreversible covalent inhibitor, the cocrystal structure of FAAH complexed with compound 2 reveals that these ketobenzimidazoles, though containing a carbonyl moiety, do not covalently modify Ser241. These inhibitors achieve potent inhibition of FAAH activity primarily from shape complementarity to the active site and through numerous hydrophobic interactions. These noncovalent compounds exhibit excellent selectivity and good pharmacokinetic properties. The discovery of this distinctive class of inhibitors opens a new avenue for modulating FAAH activity through nonmechanism-based inhibition.
Assuntos
Amidoidrolases/antagonistas & inibidores , Benzimidazóis/isolamento & purificação , Benzimidazóis/metabolismo , Descoberta de Drogas/métodos , Inibidores Enzimáticos/isolamento & purificação , Inibidores Enzimáticos/metabolismo , Modelos Moleculares , Animais , Benzimidazóis/farmacocinética , Cumarínicos , Cristalização , Inibidores Enzimáticos/farmacocinética , Escherichia coli , Humanos , Estrutura Molecular , Ratos , Espectrofotometria Ultravioleta , Espectrometria de Massas em Tandem , Ureia/metabolismoRESUMO
PPARγ is a member of the nuclear hormone receptor family and plays a key role in the regulation of glucose homeostasis. This Letter describes the discovery of a novel chemical class of diarylsulfonamide partial agonists that act as selective PPARγ modulators (SPPARγMs) and display a unique pharmacological profile compared to the thiazolidinedione (TZD) class of PPARγ full agonists. Herein we report the initial discovery of partial agonist 4 and the structure-activity relationship studies that led to the selection of clinical compound INT131 (3), a potent PPARγ partial agonist that displays robust glucose-lowering activity in rodent models of diabetes while exhibiting a reduced side-effects profile compared to marketed TZDs.
Assuntos
PPAR gama/agonistas , Quinolinas/química , Sulfonamidas/química , Administração Oral , Animais , Sítios de Ligação , Cristalografia por Raios X , Citocromo P-450 CYP3A , Inibidores das Enzimas do Citocromo P-450 , Sistema Enzimático do Citocromo P-450/metabolismo , Diabetes Mellitus Experimental/tratamento farmacológico , Meia-Vida , Resistência à Insulina , Masculino , Camundongos , PPAR gama/metabolismo , Estrutura Terciária de Proteína , Quinolinas/farmacocinética , Quinolinas/uso terapêutico , Ratos , Ratos Sprague-Dawley , Ratos Zucker , Relação Estrutura-Atividade , Sulfonamidas/síntese química , Sulfonamidas/farmacocinética , Sulfonamidas/uso terapêuticoRESUMO
We report our successful effort to increase the PDE3 selectivity of PDE10A inhibitor pyridyl cinnoline 1 using a combination of computational modeling and structural-activity relationship investigations. An analysis of the PDE3 catalytic domain compared to the co-crystal structure of cinnoline analog 1 in PDE10A revealed two areas of structural differences in the active sites and suggested areas on the scaffold that could be modified to exploit those unique structural features. Once SAR established the cinnoline as the optimal scaffold, modifications on the methoxy groups of the cinnoline and the methyl group on the pyridine led to the discovery of compounds 33 and 36. Both compounds achieved significant improvement in selectivity against PDE3 while maintaining their PDE10A inhibitory activity and in vivo metabolic stability comparable to 1.
Assuntos
Nucleotídeo Cíclico Fosfodiesterase do Tipo 3/química , Compostos Heterocíclicos com 2 Anéis/química , Inibidores de Fosfodiesterase/química , Diester Fosfórico Hidrolases/química , Piridinas/química , Animais , Sítios de Ligação , Domínio Catalítico , Cristalografia por Raios X , Nucleotídeo Cíclico Fosfodiesterase do Tipo 3/metabolismo , Desenho de Fármacos , Meia-Vida , Compostos Heterocíclicos com 2 Anéis/síntese química , Compostos Heterocíclicos com 2 Anéis/farmacocinética , Inibidores de Fosfodiesterase/síntese química , Inibidores de Fosfodiesterase/farmacocinética , Diester Fosfórico Hidrolases/metabolismo , Piridinas/síntese química , Piridinas/farmacocinética , Ratos , Relação Estrutura-AtividadeRESUMO
An initial SAR study resulted in the identification of the novel, potent MCHR1 antagonist 2. After further profiling, compound 2 was discovered to be a potent inhibitor of the hERG potassium channel, which prevented its further development. Additional optimization of this structure resulted in the discovery of the potent MCHR1 antagonist 11 with a dramatically reduced hERG liability. The decrease in hERG activity was confirmed by several in vivo preclinical cardiovascular studies examining QT prolongation. This compound demonstrated good selectivity for MCHR1 and possessed good pharmacokinetic properties across preclinical species. Compound 11 was also efficacious in reducing body weight in two in vivo mouse models. This compound was selected for clinical evaluation and was given the code AMG 076.
Assuntos
Carbazóis/química , Ácidos Cicloexanocarboxílicos/química , Canais de Potássio Éter-A-Go-Go/antagonistas & inibidores , Receptores do Hormônio Hipofisário/antagonistas & inibidores , Animais , Peso Corporal/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Carbazóis/síntese química , Carbazóis/farmacocinética , Ácidos Cicloexanocarboxílicos/síntese química , Ácidos Cicloexanocarboxílicos/farmacocinética , Dieta Hiperlipídica , Cães , Avaliação Pré-Clínica de Medicamentos , Canal de Potássio ERG1 , Canais de Potássio Éter-A-Go-Go/metabolismo , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ratos , Receptores do Hormônio Hipofisário/genética , Receptores do Hormônio Hipofisário/metabolismo , Relação Estrutura-AtividadeRESUMO
A series of spiropiperidine carbazoles were synthesized and evaluated as MCHR2 antagonists using a FLIPR assay. The pharmacokinetic properties of selected compounds have also been studied. This effort led to the discovery of potent and specific MCHR2 antagonists. Compound 38 demonstrated good pharmacokinetic properties across rat, beagle dog and rhesus monkey and had a favorable selectivity profile against a number of other receptors. These MCHR2 antagonists are considered appropriate tool compounds for study of the function of MCHR2 in vivo.
Assuntos
Carbazóis/química , Química Farmacêutica/métodos , Piperidinas/química , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Receptores do Hormônio Hipofisário/antagonistas & inibidores , Animais , Células CHO , Linhagem Celular , Cricetinae , Cães , Desenho de Fármacos , Humanos , Concentração Inibidora 50 , Macaca mulatta , Modelos Químicos , Ratos , Relação Estrutura-AtividadeRESUMO
A series of benzodiazepine antagonists of the human ghrelin receptor GHSR1a were synthesized and their antagonism and metabolic stability were evaluated. The potency of these analogs was determined using a functional aequorin (Euroscreen) luminescent assay measuring the intracellular Ca(2+) concentration, and their metabolic stability was measured using an in vitro rat and human S9 hepatocyte assay. These efforts led to the discovery of a potent ghrelin antagonist with good rat pharmacokinetic properties.
Assuntos
Benzodiazepinas/química , Benzodiazepinas/farmacologia , Receptores de Grelina/antagonistas & inibidores , Receptores de Grelina/metabolismo , Animais , Benzodiazepinas/metabolismo , Benzodiazepinas/farmacocinética , Cálcio/metabolismo , Linhagem Celular , Hepatócitos/metabolismo , Humanos , Medições Luminescentes , Obesidade/tratamento farmacológico , Ratos , Ratos Sprague-Dawley , Relação Estrutura-AtividadeRESUMO
We report the discovery of 6,7-dimethoxy-4-(pyridin-3-yl)cinnolines as novel inhibitors of phosphodiesterase 10A (PDE10A). Systematic examination and analyses of structure-activity-relationships resulted in single digit nM potency against PDE10A. X-ray co-crystal structure revealed the mode of binding in the enzyme's catalytic domain and the source of selectivity against other PDEs. High in vivo clearance in rats was addressed with the help of metabolite identification (ID) studies. These findings combined resulted in compound 39, a promising potent inhibitor of PDE10A with good in vivo metabolic stability in rats and efficacy in a rodent behavioral model.
Assuntos
Cumarínicos/síntese química , Inibidores de Fosfodiesterase/síntese química , Diester Fosfórico Hidrolases/metabolismo , Psicotrópicos/síntese química , Animais , Aprendizagem da Esquiva/efeitos dos fármacos , Comportamento Animal/efeitos dos fármacos , Sítios de Ligação , Cumarínicos/farmacologia , Descoberta de Drogas , Humanos , Modelos Moleculares , Inibidores de Fosfodiesterase/administração & dosagem , Ligação Proteica , Psicotrópicos/administração & dosagem , Ratos , Ratos Sprague-Dawley , Relação Estrutura-AtividadeRESUMO
Vimentin has been implicated in wound healing, inflammation, and cancer, but its functional contribution to intestinal diseases is poorly understood. To study how vimentin is involved during tissue injury and repair of simple epithelium, we induced colonic epithelial cell damage in the vimentin null (Vim-/-) mouse model. Vim-/- mice challenged with dextran sodium sulfate (DSS) had worse colitis manifestations than wild-type (WT) mice. Vim-/- colons also produced more reactive oxygen and nitrogen species, possibly contributing to the pathogenesis of gut inflammation and tumorigenesis than in WT mice. We subsequently describe that CD11b+ macrophages served as the mainly cellular source of reactive oxygen species (ROS) production via vimentin-ROS-pSTAT3-interleukin-6 inflammatory pathways. Further, we demonstrated that Vim-/- mice did not develop colitis-associated cancer model upon DSS treatment spontaneously but increased tumor numbers and size in the distal colon in the azoxymethane/DSS model comparing with WT mice. Thus, vimentin has a crucial role in protection from colitis induction and tumorigenesis of the colon.
RESUMO
Stress granules (SGs) are non-membranous organelles facilitating stress responses and linking the pathology of age-related diseases. In a genome-wide imaging-based phenomic screen, we identify Pab1 co-localizing proteins under 2-deoxy-D-glucose (2-DG) induced stress in Saccharomyces cerevisiae. We find that deletion of one of the Pab1 co-localizing proteins, Lsm7, leads to a significant decrease in SG formation. Under 2-DG stress, Lsm7 rapidly forms foci that assist in SG formation. The Lsm7 foci form via liquid-liquid phase separation, and the intrinsically disordered region and the hydrophobic clusters within the Lsm7 sequence are the internal driving forces in promoting Lsm7 phase separation. The dynamic Lsm7 phase-separated condensates appear to work as seeding scaffolds, promoting Pab1 demixing and subsequent SG initiation, seemingly mediated by RNA interactions. The SG initiation mechanism, via Lsm7 phase separation, identified in this work provides valuable clues for understanding the mechanisms underlying SG formation and SG-associated human diseases.
Assuntos
Fenômenos Bioquímicos , Proteínas de Saccharomyces cerevisiae , Grânulos Citoplasmáticos/metabolismo , Humanos , Proteínas de Ligação a Poli(A)/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Grânulos de EstresseRESUMO
Starting from a series of ureas that were determined to be mechanism-based inhibitors of FAAH, several spirocyclic ureas and lactams were designed and synthesized. These efforts identified a series of novel, noncovalent FAAH inhibitors with in vitro potency comparable to known covalent FAAH inhibitors. The mechanism of action for these compounds was determined through a combination of SAR and co-crystallography with rat FAAH.
Assuntos
Amidoidrolases/antagonistas & inibidores , Inibidores Enzimáticos/química , Amidoidrolases/metabolismo , Animais , Sítios de Ligação , Simulação por Computador , Cristalografia por Raios X , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacocinética , Humanos , Lactamas/síntese química , Lactamas/química , Lactamas/farmacocinética , Ratos , Compostos de Espiro/química , Relação Estrutura-Atividade , Ureia/síntese química , Ureia/química , Ureia/farmacocinéticaRESUMO
Piperazine-bisamide analogs were discovered as partial agonists of human growth hormone secretagogue receptor (GHSR) in a high throughput screen. The partial agonists were optimized for potency and converted into antagonists through structure-activity relationship (SAR) studies. The efforts also led to the identification of potent antagonist with favorable PK profile suitable as a tool compound for in vivo studies.
Assuntos
Amidas/química , Fármacos Antiobesidade/química , Indóis/química , Piperazinas/química , Receptores de Grelina/antagonistas & inibidores , Amidas/síntese química , Amidas/uso terapêutico , Animais , Fármacos Antiobesidade/síntese química , Fármacos Antiobesidade/uso terapêutico , Ensaios de Triagem em Larga Escala , Humanos , Indóis/síntese química , Indóis/uso terapêutico , Obesidade/tratamento farmacológico , Piperazinas/síntese química , Piperazinas/uso terapêutico , Ratos , Receptores de Grelina/metabolismo , Relação Estrutura-AtividadeRESUMO
Fused in sarcoma (FUS) is a multifunctional DNA/RNA-binding protein predominantly localized in the cell nucleus. However, FUS has been shown to accumulate and form aggregates in the cytoplasm when mislocalized there due to mutations. These FUS protein aggregates are known as pathological hallmarks in a subset of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD) cases. In this review, we discussed recent research developments on elucidating the molecular mechanisms behind FUS protein aggregation and toxicity. We mainly focus on studies using the budding yeast (Saccharomyces cerevisiae) as a model system, especially on results acquired from yeast genome-wide screens addressing FUS aggregation and toxicity. Human homologs of the FUS toxicity suppressors, identified from these studies, indicate a strong relevance and correlation to a human disease model. By using yeast as a FUS cytotoxicity model these studies provided valuable clues on potential novel targets for therapeutic intervention in ALS.
RESUMO
The nuclear hormone receptor peroxisome proliferator-activated receptor gamma (PPAR gamma; NR1C3) plays a central role in adipogenesis and is the molecular target of the thiazolidinedione class of antidiabetic drugs. To overcome the well-known shortcomings of thiazolidinediones, we have identified INT131 (formerly T131 and AMG131) as a potent selective ligand for PPAR gamma that is structurally and pharmacologically distinct from glitazone agonists. In vitro biochemical and cell-based functional assays showed that INT131 mediates a distinct pattern of coregulator recruitment to PPAR gamma. In adipocytes, INT131 showed minimal stimulation of adipocyte differentiation and partially activated PPAR gamma target genes involved in adipogenesis and, at the same time, showed more agonistic activity on another set of target genes that may influence insulin sensitivity directly. These unique properties of INT131 may provide a mechanistic basis for its distinct pharmacological profile. In vivo, increases in glucose tolerance were observed in Zucker (fa/fa) rats following a 14-day oral treatment with INT131. Although the maximal efficacies of INT131 and rosiglitazone were similar with respect to improvements in glucose tolerance, INT131 had less effect on heart and lung weights, weight gain, hemodilution, and plasma volume. Thus, INT131 appears to selectively modulate PPAR gamma responses in an in vivo preclinical model, showing antidiabetic efficacy while exhibiting an improved hemodynamic and cardiovascular adverse effect profile compared to the full agonist rosiglitazone. X-ray crystallography revealed that INT131 interacts with PPAR gamma through a distinct binding mode, forming primarily hydrophobic contacts with the ligand-binding pocket without direct hydrogen-bonding interactions to key residues in helix 12 that are characteristic of full agonists. Mutagenesis studies on Tyr473 in helix 12 demonstrated this residue as essential for rosiglitazone-induced receptor activation, but nonessential for INT131 function in vitro, providing one possible molecular determinant for INT131's distinct pharmacology. INT131 is currently being evaluated in a clinical setting as a therapeutic agent for the treatment of type 2 diabetes.
Assuntos
Interações Hidrofóbicas e Hidrofílicas , Hipoglicemiantes/farmacologia , PPAR gama/agonistas , Quinolinas/farmacologia , Sulfonamidas/farmacologia , Adipócitos/citologia , Adipócitos/efeitos dos fármacos , Adipócitos/fisiologia , Sequência de Aminoácidos , Animais , Glicemia/análise , Diferenciação Celular/fisiologia , Células Cultivadas , Cristalografia por Raios X , Agonismo Parcial de Drogas , Hipoglicemiantes/química , Insulina/sangue , Camundongos , Dados de Sequência Molecular , PPAR gama/química , Ligação Proteica , Quinolinas/química , Ratos , Ratos Zucker , Rosiglitazona , Sulfonamidas/química , Tiazolidinedionas/química , Tiazolidinedionas/farmacologiaRESUMO
The nuclear hormone receptor peroxisome proliferator-activated receptor gamma (PPARgamma) plays central roles in adipogenesis and glucose homeostasis and is the molecular target for the thiazolidinedione (TZD) class of antidiabetic drugs. Activation of PPARgamma by TZDs improves insulin sensitivity; however, this is accompanied by the induction of several undesirable side effects. We have identified a novel synthetic PPARgamma ligand, T2384, to explore the biological activities associated with occupying different regions of the receptor ligand-binding pocket. X-ray crystallography studies revealed that T2384 can adopt two distinct binding modes, which we have termed "U" and "S", interacting with the ligand-binding pocket of PPARgamma primarily via hydrophobic contacts that are distinct from full agonists. The different binding modes occupied by T2384 induced distinct patterns of coregulatory protein interaction with PPARgamma in vitro and displayed unique receptor function in cell-based activity assays. We speculate that these unique biochemical and cellular activities may be responsible for the novel in vivo profile observed in animals treated systemically with T2384. When administered to diabetic KKAy mice, T2384 rapidly improved insulin sensitivity in the absence of weight gain, hemodilution, and anemia characteristics of treatment with rosiglitazone (a TZD). Moreover, upon coadministration with rosiglitazone, T2384 was able to antagonize the side effects induced by rosiglitazone treatment alone while retaining robust effects on glucose disposal. These results are consistent with the hypothesis that interactions between ligands and specific regions of the receptor ligand-binding pocket might selectively trigger a subset of receptor-mediated biological responses leading to the improvement of insulin sensitivity, without eliciting less desirable responses associated with full activation of the receptor. We suggest that T2384 may represent a prototype for a novel class of PPARgamma ligand and, furthermore, that molecules sharing some of these properties would be useful for treatment of type 2 diabetes.
Assuntos
Adipócitos/metabolismo , Adipogenia/efeitos dos fármacos , Benzotiazóis/farmacologia , Diabetes Mellitus Tipo 2/tratamento farmacológico , Hipoglicemiantes/farmacologia , PPAR gama/agonistas , Sulfonamidas/farmacologia , Adipócitos/patologia , Animais , Benzotiazóis/química , Benzotiazóis/uso terapêutico , Sítios de Ligação , Células Cultivadas , Cristalografia por Raios X , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/patologia , Glucose/metabolismo , Homeostase/efeitos dos fármacos , Humanos , Hipoglicemiantes/química , Hipoglicemiantes/uso terapêutico , Ligantes , Camundongos , Modelos Moleculares , PPAR gama/química , PPAR gama/metabolismo , Rosiglitazona , Sulfonamidas/química , Sulfonamidas/uso terapêutico , Tiazolidinedionas/química , Tiazolidinedionas/farmacologia , Tiazolidinedionas/uso terapêutico , Aumento de Peso/efeitos dos fármacosRESUMO
PURPOSE: To compare the visual outcomes and incidence of complications of lifting with recutting the lamellar flap in laser in situ keratomileusis (LASIK) enhancement surgery. DESIGN: Retrospective case-control study. PARTICIPANTS: Two hundred twelve consecutive eyes undergoing a LASIK enhancement procedure at a single surgery location during a 5-year period. METHODS: Charts of participants were obtained and outcome measures obtained. MAIN OUTCOME MEASURES: Uncorrected visual acuity, best-corrected visual acuity, refractive error, complications. RESULTS: Relifting of flaps was performed in 164 of 212 eyes (77.4%), and recutting of flaps was performed in 48 of 212 eyes (22.6%). There were no significant differences in early visual outcomes between the two groups. At 1 year patients had significantly better uncorrected vision if the flap was lifted rather than recut (20/24.7 vs. 20/31.3, P < 0.008). In addition, the flap lift group had a significantly more stable refraction at 1 year than did the recut group (change in spherical equivalent: +0.05 diopters (D) vs. -0.57 D). The incidence of complications did not significantly differ between the two groups. CONCLUSIONS: LASIK enhancement surgery can be performed safely and effectively by either lifting or recutting a flap. Lifting the flap may show better long-term stability of refractive error and uncorrected acuity.
Assuntos
Substância Própria/cirurgia , Ceratomileuse Assistida por Excimer Laser In Situ/métodos , Miopia/cirurgia , Retalhos Cirúrgicos , Adulto , Estudos de Casos e Controles , Feminino , Humanos , Incidência , Complicações Intraoperatórias , Masculino , Pessoa de Meia-Idade , Complicações Pós-Operatórias , Reoperação , Estudos Retrospectivos , Resultado do Tratamento , Acuidade VisualRESUMO
The nuclear hormone receptor peroxisome proliferator-activated receptor gamma (PPARgamma (NR1C3)) plays a central role in adipogenesis and is the molecular target for the thiazolidinedione (TZD) class of antidiabetic drugs. In a search for novel non-TZD ligands for PPARgamma, T0070907 was identified as a potent and selective PPARgamma antagonist. With an apparent binding affinity (concentration at 50% inhibition of [(3)H]rosiglitazone binding or IC(50)) of 1 nm, T0070907 covalently modifies PPARgamma on cysteine 313 in helix 3 of human PPARgamma2. T0070907 blocked PPARgamma function in both cell-based reporter gene and adipocyte differentiation assays. Consistent with its role as an antagonist of PPARgamma, T0070907 blocked agonist-induced recruitment of coactivator-derived peptides to PPARgamma in a homogeneous time-resolved fluorescence-based assay and promoted recruitment of the transcriptional corepressor NCoR to PPARgamma in both glutathione S-transferase pull-down assays and a PPARgamma/retinoid X receptor (RXR) alpha-dependent gel shift assay. Studies with mutant receptors suggest that T0070907 modulates the interaction of PPARgamma with these cofactor proteins by affecting the conformation of helix 12 of the PPARgamma ligand-binding domain. Interestingly, whereas the T0070907-induced NCoR recruitment to PPARgamma/RXRalpha heterodimer can be almost completely reversed by the simultaneous treatment with RXRalpha agonist LGD1069, T0070907 treatment has only modest effects on LGD1069-induced coactivator recruitment to the PPARgamma/RXRalpha heterodimer. These results suggest that the activity of PPARgamma antagonists can be modulated by the availability and concentration of RXR agonists. T0070907 is a novel tool for the study of PPARgamma/RXRalpha heterodimer function.