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1.
J Med Chem ; 64(15): 10951-10966, 2021 08 12.
Artigo em Inglês | MEDLINE | ID: mdl-34260245

RESUMO

Influenza viruses cause approximately half a million deaths every year worldwide. Vaccines are available but partially effective, and the number of antiviral medications is limited. Thus, it is crucial to develop therapeutic strategies to counteract this major pathogen. Influenza viruses enter the host cell via their hemagglutinin (HA) proteins. The HA subtypes of influenza A virus are phylogenetically classified into groups 1 and 2. Here, we identified an inhibitor of the HA protein, a tertiary aryl sulfonamide, that prevents influenza virus entry and replication. This compound shows potent antiviral activity against diverse H1N1, H5N1, and H3N2 influenza viruses encoding HA proteins from both groups 1 and 2. Synthesis of derivatives of this aryl sulfonamide identified moieties important for antiviral activity. This compound may be considered as a lead for drug development with the intent to be used alone or in combination with other influenza A virus antivirals to enhance pan-subtype efficacy.


Assuntos
Antivirais/farmacologia , Glicoproteínas de Hemaglutininação de Vírus da Influenza/metabolismo , Vírus da Influenza A/efeitos dos fármacos , Sulfonamidas/farmacologia , Antivirais/síntese química , Antivirais/química , Relação Dose-Resposta a Droga , Estrutura Molecular , Relação Estrutura-Atividade , Sulfonamidas/síntese química , Sulfonamidas/química , Internalização do Vírus/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos
2.
J Med Chem ; 63(17): 9773-9786, 2020 09 10.
Artigo em Inglês | MEDLINE | ID: mdl-32787093

RESUMO

A series of N-acyl benzothiazoles shows selective and potent cytotoxicity against cancer cell lines expressing cytochrome P450 4F11. A prodrug form is metabolized by cancer cells into an active inhibitor of stearoyl-CoA desaturase (SCD). Substantial variation on the acyl portion of the inhibitors allowed the identification of (R)-27, which balanced potency, solubility, and lipophilicity to allow proof-of-concept studies in mice. The prodrugs were activated inside the tumor, where they can arrest tumor growth. Together, these observations offer promise that a tumor-activated prodrug strategy might exploit the essentiality of SCD for tumor growth, while avoiding toxicity associated with systemic SCD inhibition.


Assuntos
Benzotiazóis/farmacologia , Inibidores Enzimáticos/farmacologia , Estearoil-CoA Dessaturase/antagonistas & inibidores , Animais , Benzotiazóis/farmacocinética , Linhagem Celular Tumoral , Família 4 do Citocromo P450/metabolismo , Feminino , Humanos , Camundongos , Pró-Fármacos/metabolismo , Distribuição Tecidual
3.
Antiviral Res ; 158: 288-302, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-30144461

RESUMO

Specific host pathways that may be targeted therapeutically to inhibit the replication of Ebola virus (EBOV) and other emerging viruses remain incompletely defined. A screen of 200,000 compounds for inhibition of an EBOV minigenome (MG) assay that measures the function of the viral polymerase complex identified as hits several compounds with an amino-tetrahydrocarbazole scaffold. This scaffold was structurally similar to GSK983, a compound previously described as having broad-spectrum antiviral activity due to its impairing de novo pyrimidine biosynthesis through inhibition of dihydroorotate dehydrogenase (DHODH). We generated compound SW835, the racemic version of GSK983 and demonstrated that SW835 and brequinar, another DHODH inhibitor, potently inhibit the MG assay and the replication of EBOV, vesicular stomatitis virus (VSV) and Zika (ZIKV) in vitro. Nucleoside and deoxynucleoside supplementation studies demonstrated that depletion of pyrimidine pools contributes to antiviral activity of these compounds. As reported for other DHODH inhibitors, SW835 and brequinar also induced expression of interferon stimulated genes (ISGs). ISG induction was demonstrated to occur without production of IFNα/ß and independently of the IFNα receptor and was not blocked by EBOV-encoded suppressors of IFN signaling pathways. Furthermore, we demonstrated that transcription factor IRF1 is required for this ISG induction, and that IRF1 induction requires the DNA damage response kinase ATM. Therefore, de novo pyrimidine biosynthesis is critical for the replication of EBOV and other RNA viruses and inhibition of this pathway activates an ATM and IRF1-dependent innate immune response that subverts EBOV immune evasion functions.


Assuntos
Ebolavirus/efeitos dos fármacos , Imunidade Inata/efeitos dos fármacos , Nucleosídeos/farmacologia , Pirimidinas/antagonistas & inibidores , Pirimidinas/biossíntese , Replicação Viral/efeitos dos fármacos , Células A549 , Antivirais/farmacologia , Compostos de Bifenilo/química , Compostos de Bifenilo/farmacologia , Carbazóis/química , Carbazóis/farmacologia , Dano ao DNA , Células HEK293 , Doença pelo Vírus Ebola/tratamento farmacológico , Doença pelo Vírus Ebola/virologia , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Humanos , Evasão da Resposta Imune , Imunidade Inata/genética , Fator Regulador 1 de Interferon/metabolismo , Fator Regulador 1 de Interferon/farmacologia , Interferon-alfa/metabolismo , Interferon beta/metabolismo , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/antagonistas & inibidores , Vírus de RNA/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Vesiculovirus/efeitos dos fármacos , Zika virus/efeitos dos fármacos
4.
Environ Microbiol ; 20(4): 1330-1349, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29215193

RESUMO

Ralstonia solanacearum thrives in plant xylem vessels and causes bacterial wilt disease despite the low nutrient content of xylem sap. We found that R. solanacearum manipulates its host to increase nutrients in tomato xylem sap, enabling it to grow better in sap from infected plants than in sap from healthy plants. Untargeted GC/MS metabolomics identified 22 metabolites enriched in R. solanacearum-infected sap. Eight of these could serve as sole carbon or nitrogen sources for R. solanacearum. Putrescine, a polyamine that is not a sole carbon or nitrogen source for R. solanacearum, was enriched 76-fold to 37 µM in R. solanacearum-infected sap. R. solanacearum synthesized putrescine via a SpeC ornithine decarboxylase. A ΔspeC mutant required ≥ 15 µM exogenous putrescine to grow and could not grow alone in xylem even when plants were treated with putrescine. However, co-inoculation with wildtype rescued ΔspeC growth, indicating R. solanacearum produced and exported putrescine to xylem sap. Intriguingly, treating plants with putrescine before inoculation accelerated wilt symptom development and R. solanacearum growth and systemic spread. Xylem putrescine concentration was unchanged in putrescine-treated plants, so the exogenous putrescine likely accelerated disease indirectly by affecting host physiology. These results indicate that putrescine is a pathogen-produced virulence metabolite.


Assuntos
Lycopersicon esculentum/microbiologia , Doenças das Plantas/microbiologia , Putrescina/metabolismo , Ralstonia solanacearum/metabolismo , Ralstonia solanacearum/patogenicidade , Xilema/metabolismo , Metabolômica , Virulência , Fatores de Virulência/metabolismo , Xilema/microbiologia
5.
J Biol Chem ; 292(29): 12041-12053, 2017 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-28546427

RESUMO

Ubiquitous polyamine spermidine is not required for normal planktonic growth of Bacillus subtilis but is essential for robust biofilm formation. However, the structural features of spermidine required for B. subtilis biofilm formation are unknown and so are the molecular mechanisms of spermidine-stimulated biofilm development. We report here that in a spermidine-deficient B. subtilis mutant, the structural analogue norspermidine, but not homospermidine, restored biofilm formation. Intracellular biosynthesis of another spermidine analogue, aminopropylcadaverine, from exogenously supplied homoagmatine also restored biofilm formation. The differential ability of C-methylated spermidine analogues to functionally replace spermidine in biofilm formation indicated that the aminopropyl moiety of spermidine is more sensitive to C-methylation, which it is essential for biofilm formation, but that the length and symmetry of the molecule is not critical. Transcriptomic analysis of a spermidine-depleted B. subtilis speD mutant uncovered a nitrogen-, methionine-, and S-adenosylmethionine-sufficiency response, resulting in repression of gene expression related to purine catabolism, methionine and S-adenosylmethionine biosynthesis and methionine salvage, and signs of altered membrane status. Consistent with the spermidine requirement in biofilm formation, single-cell analysis of this mutant indicated reduced expression of the operons for production of the exopolysaccharide and TasA protein biofilm matrix components and SinR antagonist slrR Deletion of sinR or ectopic expression of slrR in the spermidine-deficient ΔspeD background restored biofilm formation, indicating that spermidine is required for expression of the biofilm regulator slrR Our results indicate that spermidine functions in biofilm development by activating transcription of the biofilm matrix exopolysaccharide and TasA operons through the regulator slrR.


Assuntos
Bacillus subtilis/fisiologia , Proteínas de Bactérias/agonistas , Biofilmes/crescimento & desenvolvimento , Regulação Bacteriana da Expressão Gênica , Polissacarídeos Bacterianos/biossíntese , Espermidina/metabolismo , Fatores de Transcrição/agonistas , Adenosilmetionina Descarboxilase/genética , Adenosilmetionina Descarboxilase/metabolismo , Bacillus subtilis/citologia , Bacillus subtilis/genética , Bacillus subtilis/crescimento & desenvolvimento , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Cadaverina/análogos & derivados , Cadaverina/metabolismo , Deleção de Genes , Perfilação da Expressão Gênica , Metionina/metabolismo , Metilação , Ciclo do Nitrogênio , Óperon , Purinas/metabolismo , S-Adenosilmetionina/metabolismo , Análise de Célula Única , Espermidina/análogos & derivados , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
6.
J Med Chem ; 60(9): 3979-4001, 2017 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-28398755

RESUMO

The enzyme 15-prostaglandin dehydrogenase (15-PGDH) catalyzes the first step in the degradation of prostaglandins including PGE2. It is a negative regulator of tissue repair and regeneration in multiple organs. Accordingly, inhibitors of 15-PGDH are anticipated to elevate in vivo levels of PGE2 and to promote healing and tissue regeneration. The small molecule SW033291 (1) inhibits 15-PGDH with Ki = 0.1 nM in vitro, doubles PGE2 levels in vivo, and shows efficacy in mouse models of recovery from bone marrow transplantation, ulcerative colitis, and partial hepatectomy. Here we describe optimized variants of 1 with improved solubility, druglike properties, and in vivo activity.


Assuntos
Inibidores Enzimáticos/farmacologia , Hidroxiprostaglandina Desidrogenases/antagonistas & inibidores , Animais , Inibidores Enzimáticos/química , Humanos , Hidroxiprostaglandina Desidrogenases/genética , Camundongos , Camundongos Knockout , Relação Estrutura-Atividade
7.
ACS Chem Biol ; 11(10): 2782-2789, 2016 10 21.
Artigo em Inglês | MEDLINE | ID: mdl-27541336

RESUMO

The small polyamine putrescine (1,4-diaminobutane) is ubiquitously and abundantly found in all three domains of life. It is a precursor, through N-aminopropylation or N-aminobutylation, for biosynthesis of the longer polyamines spermidine, sym-homospermidine, spermine, and thermospermine and longer and branched chain polyamines. Putrescine is also biochemically modified for purposes of metabolic regulation and catabolism, e.g. N-acetylation and N-glutamylation, and for incorporation into specialized metabolites, e.g. N-methylation, N-citrylation, N-palmitoylation, N-hydroxylation, and N-hydroxycinnamoylation. Only one example is known where putrescine is modified on a methylene carbon: the formation of 2-hydroxyputrescine by an unknown C-hydroxylase. Here, we report the functional identification of a previously undescribed putrescine 2-hydroxylase, a Rieske-type nonheme iron sulfur protein from the ß-proteobacteria Bordetella bronchiseptica and Ralstonia solanacearum. Identification of the putrescine 2-hydroxylase will facilitate investigation of the physiological functions of 2-hydroxyputrescine. One known role of 2-hydroxyputrescine has direct biomedical relevance: its role in the biosynthesis of the cyclic hydroxamate siderophore alcaligin, a potential virulence factor of the causative agent of whooping cough, Bordetella pertussis. We also report the functional identification of a putrescine N-hydroxylase from the γ-proteobacterium Shewanella oneidensis, which is homologous to FAD- and NADPH-dependent ornithine and lysine N-monooxygenases involved in siderophore biosynthesis. Heterologous expression of the putrescine N-hydroxylase in E. coli produced free N-hydroxyputrescine, never detected previously in a biological system. Furthermore, the putrescine C- and N-hydroxylases identified here could contribute new functionality to polyamine structural scaffolds, including C-H bond functionalization in synthetic biology strategies.


Assuntos
Oxigenases de Função Mista/metabolismo , Putrescina/metabolismo , Escherichia coli/metabolismo , Isomerismo , Estrutura Molecular , Putrescina/química
8.
Science ; 348(6240): aaa2340, 2015 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-26068857

RESUMO

Agents that promote tissue regeneration could be beneficial in a variety of clinical settings, such as stimulating recovery of the hematopoietic system after bone marrow transplantation. Prostaglandin PGE2, a lipid signaling molecule that supports expansion of several types of tissue stem cells, is a candidate therapeutic target for promoting tissue regeneration in vivo. Here, we show that inhibition of 15-hydroxyprostaglandin dehydrogenase (15-PGDH), a prostaglandin-degrading enzyme, potentiates tissue regeneration in multiple organs in mice. In a chemical screen, we identify a small-molecule inhibitor of 15-PGDH (SW033291) that increases prostaglandin PGE2 levels in bone marrow and other tissues. SW033291 accelerates hematopoietic recovery in mice receiving a bone marrow transplant. The same compound also promotes tissue regeneration in mouse models of colon and liver injury. Tissues from 15-PGDH knockout mice demonstrate similar increased regenerative capacity. Thus, 15-PGDH inhibition may be a valuable therapeutic strategy for tissue regeneration in diverse clinical contexts.


Assuntos
Hidroxiprostaglandina Desidrogenases/fisiologia , Prostaglandinas/metabolismo , Regeneração/fisiologia , Animais , Transplante de Medula Óssea , Colite/enzimologia , Colite/prevenção & controle , Dinoprostona/metabolismo , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Hematopoese/efeitos dos fármacos , Hidroxiprostaglandina Desidrogenases/antagonistas & inibidores , Hidroxiprostaglandina Desidrogenases/genética , Regeneração Hepática/efeitos dos fármacos , Camundongos , Camundongos Knockout , Piridinas/química , Piridinas/farmacologia , Regeneração/efeitos dos fármacos , Regeneração/genética , Tiofenos/química , Tiofenos/farmacologia
9.
Artigo em Inglês | MEDLINE | ID: mdl-27158662

RESUMO

BACKGROUND: There are currently no therapeutic options for patients with Parkinson's disease that prevent or slow the death of dopaminergic neurons. We have recently identified the novel P7C3 class of neuroprotective molecules that blocks neuron cell death. AIMS: The aim of this study was to determine whether treatment with highly active members of the P7C3 series blocks dopaminergic neuron cell death and associated behavioral and neurochemical deficits in the rat 6-hydroxydopamine (6-OHDA) model of Parkinson's disease. METHODS: After unilateral injection of 6-OHDA into the median forebrain bundle, rats were assessed for behavioral function in the open field, cylinder test, and amphetamine-induced circling test. Thereafter, their brains were subjected to neurochemical and immunohistochemical analysis of dopaminergic neuron survival. Analysis was conducted as a function of treatment with P7C3 compounds, with administration initiated either before or after 6-OHDA exposure. RESULTS: Animals administered P7C3-A20 or P7C3-S243, two of the most advanced agents in the P7C3 series of neuroprotective compounds, both before and after 6-OHDA exposure showed evidence of protective efficacy in all measures. When P7C3-S243 administration was initiated after 6-OHDA exposure, rats also showed protective efficacy in all measures, which included blocking dopaminergic neuron cell death in ipsilateral substantia nigra pars compacta, preservation of dopamine and its metabolites in ipsilateral striatum, and preservation of normal motor behavior. CONCLUSIONS: The P7C3 series of compounds may form the basis for developing new therapeutic agents for slowing or preventing progression of Parkinson's disease.

10.
Invest Ophthalmol Vis Sci ; 55(12): 8330-41, 2014 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-25468886

RESUMO

PURPOSE: Traumatic brain injury (TBI) frequently leads to chronic visual dysfunction. The purpose of this study was to investigate the effect of TBI on retinal ganglion cells (RGCs), and to test whether treatment with the novel neuroprotective compound P7C3-S243 could prevent in vivo functional deficits in the visual system. METHODS: Blast-mediated TBI was modeled using an enclosed over-pressure blast chamber. The RGC physiology was evaluated using a multielectrode array and pattern electroretinogram (PERG). Histological analysis of RGC dendritic field and cell number were evaluated at the end of the study. Visual outcome measures also were evaluated based on treatment of mice with P7C3-S243 or vehicle control. RESULTS: We show that deficits in neutral position PERG after blast-mediated TBI occur in a temporally bimodal fashion, with temporary recovery 4 weeks after injury followed by chronically persistent dysfunction 12 weeks later. This later time point is associated with development of dendritic abnormalities and irreversible death of RGCs. We also demonstrate that ongoing pathologic processes during the temporary recovery latent period (including abnormalities of RGC physiology) lead to future dysfunction of the visual system. We report that modification of PERG to provocative postural tilt testing elicits changes in PERG measurements that correlate with a key in vitro measures of damage: the spontaneous and light-evoked activity of RGCs. Treatment with P7C3-S243 immediately after injury and throughout the temporary recovery latent period protects mice from developing chronic visual system dysfunction. CONCLUSIONS: Provocative PERG testing serves as a noninvasive test in the living organism to identify early damage to the visual system, which may reflect corresponding damage in the brain that is not otherwise detectable by noninvasive means. This provides the basis for developing an earlier diagnostic test to identify patients at risk for developing chronic CNS and visual system damage after TBI at an earlier stage when treatments may be more effective in preventing these sequelae. In addition, treatment with the neuroprotective agent P7C3-S243 after TBI protects from visual system dysfunction after TBI.


Assuntos
Traumatismos por Explosões/tratamento farmacológico , Lesões Encefálicas/tratamento farmacológico , Carbazóis/farmacologia , Fármacos Neuroprotetores/farmacologia , Células Ganglionares da Retina/efeitos dos fármacos , Transtornos da Visão/prevenção & controle , Análise de Variância , Animais , Traumatismos por Explosões/complicações , Traumatismos por Explosões/fisiopatologia , Lesões Encefálicas/complicações , Lesões Encefálicas/fisiopatologia , Contagem de Células , Dendritos/patologia , Modelos Animais de Doenças , Eletrorretinografia/efeitos dos fármacos , Injeções Intraperitoneais , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Plasticidade Neuronal/fisiologia , Células Ganglionares da Retina/patologia , Células Ganglionares da Retina/fisiologia , Transtornos da Visão/etiologia , Transtornos da Visão/fisiopatologia
11.
Cell Rep ; 8(6): 1731-1740, 2014 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-25220467

RESUMO

The P7C3 class of neuroprotective aminopropyl carbazoles has been shown to block neuronal cell death in models of neurodegeneration. We now show that P7C3 molecules additionally preserve axonal integrity after injury, before neuronal cell death occurs, in a rodent model of blast-mediated traumatic brain injury (TBI). This protective quality may be linked to the ability of P7C3 molecules to activate nicotinamide phosphoribosyltransferase, the rate-limiting enzyme in nicotinamide adenine dinucleotide salvage. Initiation of daily treatment with our recently reported lead agent, P7C3-S243, 1 day after blast-mediated TBI blocks axonal degeneration and preserves normal synaptic activity, learning and memory, and motor coordination in mice. We additionally report persistent neurologic deficits and acquisition of an anxiety-like phenotype in untreated animals 8 months after blast exposure. Optimized variants of P7C3 thus offer hope for identifying neuroprotective agents for conditions involving axonal damage, neuronal cell death, or both, such as occurs in TBI.


Assuntos
Transporte Axonal/efeitos dos fármacos , Axônios/metabolismo , Carbazóis/farmacologia , Fármacos Neuroprotetores/farmacologia , Animais , Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/metabolismo , Lesões Encefálicas/tratamento farmacológico , Carbazóis/química , Carbazóis/uso terapêutico , Modelos Animais de Doenças , Hipocampo/metabolismo , Memória/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Atividade Motora/efeitos dos fármacos , Fármacos Neuroprotetores/uso terapêutico , Nicotinamida Fosforribosiltransferase/metabolismo , Transmissão Sináptica/efeitos dos fármacos
12.
Cell ; 158(6): 1324-1334, 2014 Sep 11.
Artigo em Inglês | MEDLINE | ID: mdl-25215490

RESUMO

The P7C3 class of aminopropyl carbazole chemicals fosters the survival of neurons in a variety of rodent models of neurodegeneration or nerve cell injury. To uncover its mechanism of action, an active derivative of P7C3 was modified to contain both a benzophenone for photocrosslinking and an alkyne for CLICK chemistry. This derivative was found to bind nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme involved in the conversion of nicotinamide into nicotinamide adenine dinucleotide (NAD). Administration of active P7C3 chemicals to cells treated with doxorubicin, which induces NAD depletion, led to a rebound in intracellular levels of NAD and concomitant protection from doxorubicin-mediated toxicity. Active P7C3 variants likewise enhanced the activity of the purified NAMPT enzyme, providing further evidence that they act by increasing NAD levels through its NAMPT-mediated salvage.


Assuntos
NAD/metabolismo , Fármacos Neuroprotetores/farmacologia , Animais , Carbazóis/farmacologia , Linhagem Celular Tumoral , Células Cultivadas , Citocinas/agonistas , Citocinas/genética , Citocinas/metabolismo , Doxorrubicina/farmacologia , Humanos , Redes e Vias Metabólicas , Camundongos , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Nicotinamida Fosforribosiltransferase/genética , Nicotinamida Fosforribosiltransferase/metabolismo
13.
J Med Chem ; 57(9): 3746-54, 2014 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-24697290

RESUMO

(-)-P7C3-S243 is a neuroprotective aminopropyl carbazole with improved druglike properties compared with previously reported compounds in the P7C3 class. It protects developing neurons in a mouse model of hippocampal neurogenesis and protects mature neurons within the substantia nigra in a mouse model of Parkinson's disease. A short, enantioselective synthesis provides the neuroprotective agent in optically pure form. It is nontoxic, orally bioavailable, metabolically stable, and able to cross the blood-brain barrier. As such, it represents a valuable lead compound for the development of drugs to treat neurodegenerative diseases and traumatic brain injury.


Assuntos
Carbazóis/farmacologia , Fármacos Neuroprotetores/farmacologia , Animais , Área Sob a Curva , Modelos Animais de Doenças , Descoberta de Drogas , Espectroscopia de Ressonância Magnética , Camundongos , Camundongos Endogâmicos C57BL , Fármacos Neuroprotetores/farmacocinética , Doença de Parkinson/patologia , Espectrometria de Massas por Ionização por Electrospray , Substância Negra/patologia
14.
J Neurotrauma ; 31(5): 476-86, 2014 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-24070637

RESUMO

Traumatic brain injury (TBI) is characterized by histopathological damage and long-term sensorimotor and cognitive dysfunction. Recent studies have reported the discovery of the P7C3 class of aminopropyl carbazole agents with potent neuroprotective properties for both newborn neural precursor cells in the adult hippocampus and mature neurons in other regions of the central nervous system. This study tested, for the first time, whether the highly active P7C3-A20 compound would be neuroprotective, promote hippocampal neurogenesis, and improve functional outcomes after experimental TBI. Sprague-Dawley rats subjected to moderate fluid percussion brain injury were evaluated for quantitative immunohistochemical and behavioral changes after trauma. P7C3-A20 (10 mg/kg) or vehicle was initiated intraperitoneally 30 min postsurgery and twice per day every day thereafter for 7 days. Administration of P7C3-A20 significantly reduced overall contusion volume, preserved vulnerable anti-neuronal nuclei (NeuN)-positive pericontusional cortical neurons, and improved sensorimotor function 1 week after trauma. P7C3-A20 treatment also significantly increased both bromodeoxyuridine (BrdU)- and doublecortin (DCX)-positive cells within the subgranular zone of the ipsilateral dentate gyrus 1 week after TBI. Five weeks after TBI, animals treated with P7C3-A20 showed significantly increased BrdU/NeuN double-labeled neurons and improved cognitive function in the Morris water maze, compared to TBI-control animals. These results suggest that P7C3-A20 is neuroprotective and promotes endogenous reparative strategies after TBI. We propose that the chemical scaffold represented by P7C3-A20 provides a basis for optimizing and advancing new pharmacological agents for protecting patients against the early and chronic consequences of TBI.


Assuntos
Lesões Encefálicas/tratamento farmacológico , Encéfalo/patologia , Carbazóis/uso terapêutico , Fármacos Neuroprotetores/uso terapêutico , Recuperação de Função Fisiológica/efeitos dos fármacos , Animais , Encéfalo/efeitos dos fármacos , Lesões Encefálicas/patologia , Carbazóis/farmacologia , Masculino , Aprendizagem em Labirinto/efeitos dos fármacos , Aprendizagem em Labirinto/fisiologia , Atividade Motora/efeitos dos fármacos , Atividade Motora/fisiologia , Neurônios/efeitos dos fármacos , Neurônios/patologia , Fármacos Neuroprotetores/farmacologia , Ratos , Ratos Sprague-Dawley , Recuperação de Função Fisiológica/fisiologia
15.
Nat Chem Biol ; 9(4): 271-6, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23434853

RESUMO

Hypoxia inducible factors (HIFs) are heterodimeric transcription factors induced in many cancers where they frequently promote the expression of protumorigenic pathways. Though transcription factors are typically considered 'undruggable', the PAS-B domain of the HIF-2α subunit contains a large cavity within its hydrophobic core that offers a unique foothold for small-molecule regulation. Here we identify artificial ligands that bind within this pocket and characterize the resulting structural and functional changes caused by binding. Notably, these ligands antagonize HIF-2 heterodimerization and DNA-binding activity in vitro and in cultured cells, reducing HIF-2 target gene expression. Despite the high sequence identity between HIF-2α and HIF-1α, these ligands are highly selective and do not affect HIF-1 function. These chemical tools establish the molecular basis for selective regulation of HIF-2, providing potential therapeutic opportunities to intervene in HIF-2-driven tumors, such as renal cell carcinomas.


Assuntos
Antineoplásicos/farmacologia , Fatores de Transcrição Hélice-Alça-Hélice Básicos/antagonistas & inibidores , Proteínas de Neoplasias/antagonistas & inibidores , Bibliotecas de Moléculas Pequenas/farmacologia , Regulação Alostérica , Antineoplásicos/química , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Sítios de Ligação , Linhagem Celular Tumoral , Cristalografia por Raios X , Ensaios de Triagem em Larga Escala , Humanos , Cinética , Ligantes , Simulação de Acoplamento Molecular , Proteínas de Neoplasias/metabolismo , Ressonância Magnética Nuclear Biomolecular , Ligação Proteica , Estrutura Terciária de Proteína , Bibliotecas de Moléculas Pequenas/química
16.
J Med Chem ; 56(4): 1739-47, 2013 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-23363003

RESUMO

Hypoxia inducible factors (HIFs) are heterodimeric transcription factors induced in a variety of pathophysiological settings, including cancer. We describe the first detailed structure-activity relationship study of small molecules designed to inhibit HIF-2α-ARNT heterodimerization by binding an internal cavity of the HIF-2α PAS-B domain. Through a series of biophysical characterizations of inhibitor-protein interactions (NMR and X-ray crystallography), we have established the structural requirements for artificial inhibitors of the HIF-2α-ARNT PAS-B interaction. These results may serve as a foundation for discovering therapeutic agents that function by a novel mode of action.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/antagonistas & inibidores , Translocador Nuclear Receptor Aril Hidrocarboneto/química , Fatores de Transcrição Hélice-Alça-Hélice Básicos/química , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Cristalografia por Raios X , Ensaios de Triagem em Larga Escala , Humanos , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Estrutura Molecular , Mutação , Oxidiazóis/síntese química , Oxidiazóis/química , Ligação Proteica , Multimerização Proteica , Estrutura Terciária de Proteína , Bibliotecas de Moléculas Pequenas/química , Relação Estrutura-Atividade
17.
Nat Chem Biol ; 9(2): 84-9, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23292651

RESUMO

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) activates apoptosis through the death receptors DR4 and DR5. Because of its superior safety profile and high tumor specificity compared to other TNF family members, recombinant soluble TRAIL and agonistic antibodies against its receptors are actively being developed for clinical cancer therapy. Here, we describe the identification and characterization of the small molecules that directly target DR5 to initiate apoptosis in human cancer cells. The activity was initially discovered through a high-throughput chemical screen for compounds that promote cell death in synergy with a small-molecule mimetic of Smac, the antagonist for inhibitor of apoptosis protein. Structure-activity relationship studies yielded a more potent analog called bioymifi, which can act as a single agent to induce DR5 clustering and aggregation, leading to apoptosis. Thus, this study identified potential lead compounds for the development of small-molecule TRAIL mimics targeting DR5 for cancer therapy.


Assuntos
Neoplasias/tratamento farmacológico , Receptores do Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Apoptose , Caspase 3/metabolismo , Caspase 8/metabolismo , Caspases/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular , Relação Dose-Resposta a Droga , Ativação Enzimática , Humanos , Cinética , Modelos Químicos , Ftalimidas/farmacologia , Ligação Proteica , RNA Interferente Pequeno/metabolismo , Receptores do Ligante Indutor de Apoptose Relacionado a TNF/efeitos dos fármacos , Proteínas Recombinantes/metabolismo , Tiazolidinas/farmacologia
18.
Proc Natl Acad Sci U S A ; 109(42): 17016-21, 2012 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-23027932

RESUMO

We previously reported the discovery of P7C3, an aminopropyl carbazole having proneurogenic and neuroprotective properties in newborn neural precursor cells of the hippocampal dentate gyrus. We have further found that chemicals having efficacy in this in vivo screening assay also protect dopaminergic neurons of the substantia nigra following exposure to the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, a mouse model of Parkinson disease. Here, we provide evidence that an active analog of P7C3, known as P7C3A20, protects ventral horn spinal cord motor neurons from cell death in the G93A-SOD1 mutant mouse model of amyotrophic lateral sclerosis (ALS). P7C3A20 is efficacious in this model when administered at disease onset, and protection from cell death correlates with preservation of motor function in assays of walking gait and in the accelerating rotarod test. The prototypical member of this series, P7C3, delays disease progression in G93A-SOD1 mice when administration is initiated substantially earlier than the expected time of symptom onset. Dimebon, an antihistaminergic drug with significantly weaker proneurogenic and neuroprotective efficacy than P7C3, confers no protection in this ALS model. We propose that the chemical scaffold represented by P7C3 and P7C3A20 may provide a basis for the discovery and optimization of pharmacologic agents for the treatment of ALS.


Assuntos
Esclerose Amiotrófica Lateral/prevenção & controle , Carbazóis/farmacologia , Neurônios Motores/citologia , Fármacos Neuroprotetores/farmacologia , Medula Espinal/citologia , Animais , Carbazóis/síntese química , Carbazóis/química , Carbazóis/farmacocinética , Indóis/farmacocinética , Indóis/farmacologia , Camundongos , Atividade Motora/efeitos dos fármacos , Atividade Motora/fisiologia , Neurônios Motores/efeitos dos fármacos , Reação em Cadeia da Polimerase , Teste de Desempenho do Rota-Rod , Medula Espinal/efeitos dos fármacos
19.
Proc Natl Acad Sci U S A ; 109(42): 17010-5, 2012 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-23027934

RESUMO

We previously reported the discovery of P7C3, an aminopropyl carbazole having proneurogenic and neuroprotective properties in newborn neural precursor cells of the dentate gyrus. Here, we provide evidence that P7C3 also protects mature neurons in brain regions outside of the hippocampus. P7C3 blocks 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-mediated cell death of dopaminergic neurons in the substantia nigra of adult mice, a model of Parkinson disease (PD). Dose-response studies show that the P7C3 analog P7C3A20 blocks cell death with even greater potency and efficacy, which parallels the relative potency and efficacy of these agents in blocking apoptosis of newborn neural precursor cells of the dentate gyrus. P7C3 and P7C3A20 display similar relative effects in blocking 1-methyl-4-phenylpyridinium (MPP(+))-mediated death of dopaminergic neurons in Caenorhabditis elegans, as well as in preserving C. elegans mobility following MPP(+) exposure. Dimebon, an antihistaminergic drug that is weakly proneurogenic and neuroprotective in the dentate gyrus, confers no protection in either the mouse or the worm models of PD. We further demonstrate that the hippocampal proneurogenic efficacy of eight additional analogs of P7C3 correlates with their protective efficacy in MPTP-mediated neurotoxicity. In vivo screening of P7C3 analogs for proneurogenic efficacy in the hippocampus may thus provide a reliable means of predicting neuroprotective efficacy. We propose that the chemical scaffold represented by P7C3 and P7C3A20 provides a basis for optimizing and advancing pharmacologic agents for the treatment of patients with PD.


Assuntos
1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina/antagonistas & inibidores , 1-Metil-4-fenilpiridínio/antagonistas & inibidores , Carbazóis/farmacologia , Neurônios Dopaminérgicos/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Doença de Parkinson/prevenção & controle , Substância Negra/citologia , Animais , Apoptose/efeitos dos fármacos , Caenorhabditis elegans , Carbazóis/síntese química , Carbazóis/química , Carbazóis/farmacocinética , Relação Dose-Resposta a Droga , Hipocampo/citologia , Hipocampo/efeitos dos fármacos , Indóis/farmacocinética , Indóis/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Estrutura Molecular , Substância Negra/efeitos dos fármacos
20.
J Am Chem Soc ; 133(5): 1428-37, 2011 Feb 09.
Artigo em Inglês | MEDLINE | ID: mdl-21210688

RESUMO

Degeneration of the hippocampus is associated with Alzheimer's disease and occurs very early in the progression of the disease. Current options for treating the cognitive symptoms associated with Alzheimer's are inadequate, giving urgency to the search for novel therapeutic strategies. Pharmacologic agents that safely enhance hippocampal neurogenesis may provide new therapeutic approaches. We discovered the first synthetic molecule, named P7C3, which protects newborn neurons from apoptotic cell death, and thus promotes neurogenesis in mice and rats in the subgranular zone of the hippocampal dentate gyrus, the site of normal neurogenesis in adult mammals. We describe the results of a medicinal chemistry campaign to optimize the potency, toxicity profile, and stability of P7C3. Systematic variation of nearly every position of the lead compound revealed elements conducive toward increases in activity and regions subject to modification. We have discovered compounds that are orally available, nontoxic, stable in mice, rats, and cell culture, and capable of penetrating the blood-brain barrier. The most potent compounds are active at nanomolar concentrations. Finally, we have identified derivatives that may facilitate mode-of-action studies through affinity chromatography or photo-cross-linking.


Assuntos
Carbazóis/química , Carbazóis/farmacologia , Descoberta de Drogas/métodos , Neurogênese/efeitos dos fármacos , Fármacos Neuroprotetores/química , Fármacos Neuroprotetores/farmacologia , Animais , Carbazóis/uso terapêutico , Carbazóis/toxicidade , Relação Dose-Resposta a Droga , Estabilidade de Medicamentos , Células HeLa , Humanos , Masculino , Camundongos , Doenças Neurodegenerativas/tratamento farmacológico , Fármacos Neuroprotetores/uso terapêutico , Fármacos Neuroprotetores/toxicidade , Relação Estrutura-Atividade
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