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1.
J Wrist Surg ; 12(1): 2-8, 2023 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-36644726

RESUMO

Background In recent years, new arthroscopic techniques have been introduced to address the irreparable tears of the triangular fibrocartilage complex (TFCC) (Palmer type 1B, Atzei class 4) by replicating the standard Adams-Berger procedure. These techniques, however, show the same limitations of the open procedure in relation to the anatomically defective location of the radial origins of the radioulnar ligaments (RUL) and the risk of neurovascular and/or tendon injury. Aiming to improve the quality of reconstruction and reduce surgical morbidity, a novel arthroscopic technique was developed, with the advantages of reproducing the anatomical origins of the RUL ligaments and providing all-inside tendon graft (TG) deployment and fixation. Description of Technique The Allinside anatomic arthroscopic (3A) technique is indicated for TG reconstruction of irreparable TFCC tears in the absence of distal radioulnar joint (DRUJ) arthritis. Standard wrist arthroscopy portals are used. A small incision in the radial metaphyseal area and arthroscopic control are required to set a Wrist Drill Guide and create two converging tunnels, whose openings are at the radial anatomical origins of the RUL. An ulnar tunnel is drilled at the fovea from inside-out via the 6U portal. A 3-mm tendon strip, from the palmaris longus or extensor carpi radialis brevis, is woven through the tunnels and then secured into the ulnar tunnel with an interference screw. Postoperative immobilization with restricted forearm rotation is discontinued at 5 weeks, and then postoperative rehabilitation is started. Patients and Methods The 3A technique was applied on 5 patients (2 females and 3 males), with an average age 42 years. DRUJ stability, range of motion (ROM), pain (0-10 visual analogue scale [VAS]), grip strength, modified Mayo wrist score (MMWS), and patient satisfaction were used for evaluation before surgery and at follow-up. Results No intraoperative or early complications were registered. At a mean follow-up of 26 months, DRUJ was stable in all patients, which recovered 99% ROM. Pain VAS decreased from 7 to 0.6. Grip strength increased from 38 to 48.8 Kgs. There were 4 excellent results and 1 good result on MMWS. All patient showed high satisfaction. Conclusions Although the 3A technique requires dedicated instrumentation and arthroscopic expertise, it takes advantage of improved intra-articular vision and minimized surgical trauma to reduce the risk of complications and obtain promising functional results.

3.
Nat Commun ; 11(1): 3387, 2020 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-32636417

RESUMO

Biosynthesis of glycosylphosphatidylinositol (GPI) is required for anchoring proteins to the plasma membrane, and is essential for the integrity of the fungal cell wall. Here, we use a reporter gene-based screen in Saccharomyces cerevisiae for the discovery of antifungal inhibitors of GPI-anchoring of proteins, and identify the oligocyclopropyl-containing natural product jawsamycin (FR-900848) as a potent hit. The compound targets the catalytic subunit Spt14 (also referred to as Gpi3) of the fungal UDP-glycosyltransferase, the first step in GPI biosynthesis, with good selectivity over the human functional homolog PIG-A. Jawsamycin displays antifungal activity in vitro against several pathogenic fungi including Mucorales, and in vivo in a mouse model of invasive pulmonary mucormycosis due to Rhyzopus delemar infection. Our results provide a starting point for the development of Spt14 inhibitors for treatment of invasive fungal infections.


Assuntos
Antifúngicos/farmacologia , Glicosiltransferases/antagonistas & inibidores , Policetídeos/farmacologia , Proteínas de Saccharomyces cerevisiae/antagonistas & inibidores , Animais , Proliferação de Células , Modelos Animais de Doenças , Fermentação , Genes Reporter , Glicosilfosfatidilinositóis/biossíntese , Células HCT116 , Células Hep G2 , Humanos , Concentração de Íons de Hidrogênio , Concentração Inibidora 50 , Células K562 , Pulmão/microbiologia , Masculino , Camundongos , Camundongos Endogâmicos ICR , Mucorales , Família Multigênica , Rhizopus , Saccharomyces cerevisiae
4.
Chembiochem ; 20(5): 644-649, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30462880

RESUMO

Selective and specific inhibitors of Plasmodium falciparum lysyl-tRNA synthetase represent promising therapeutic antimalarial avenues. Cladosporin was identified as a potent P. falciparum lysyl-tRNA synthetase inhibitor, with an activity against parasite lysyl-tRNA synthetase >100-fold more potent than that of the activity registered against the human enzyme. Despite its compelling activity, cladosporin exhibits poor oral bioavailability; a critical requirement for antimalarial drugs. Thus, the quest to develop metabolically stable cladosporin-derived analogues, while retaining similar selectivity and potency to that of the natural compound, has begun. Chemogenomic profiling of a designed library allowed an entirely innovative structure-activity relationship study to be initiated; this shed light on structural evidence of a privileged scaffold with a unique activity against tRNA synthetases.


Assuntos
Antimaláricos/síntese química , Descoberta de Drogas , Inibidores Enzimáticos/síntese química , Isocumarinas/síntese química , Lisina-tRNA Ligase/antagonistas & inibidores , Malária Falciparum/tratamento farmacológico , Humanos , Plasmodium falciparum/enzimologia , Relação Estrutura-Atividade
5.
Cell Chem Biol ; 25(3): 279-290.e7, 2018 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-29307839

RESUMO

Invasive fungal infections are accompanied by high mortality rates that range up to 90%. At present, only three different compound classes are available for use in the clinic, and these often suffer from low bioavailability, toxicity, and drug resistance. These issues emphasize an urgent need for novel antifungal agents. Herein, we report the identification of chemically versatile benzamide and picolinamide scaffolds with antifungal properties. Chemogenomic profiling and biochemical assays with purified protein identified Sec14p, the major phosphatidylinositol/phosphatidylcholine transfer protein in Saccharomyces cerevisiae, as the sole essential target for these compounds. A functional variomics screen identified resistance-conferring residues that localized to the lipid-binding pocket of Sec14p. Determination of the X-ray co-crystal structure of a Sec14p-compound complex confirmed binding in this cavity and rationalized both the resistance-conferring residues and the observed structure-activity relationships. Taken together, these findings open new avenues for rational compound optimization and development of novel antifungal agents.


Assuntos
Antifúngicos/metabolismo , Benzamidas/química , Ácidos Picolínicos/química , Amidas/química , Amidas/metabolismo , Amidas/farmacologia , Sequência de Aminoácidos , Antifúngicos/química , Antifúngicos/farmacologia , Aspergillus/efeitos dos fármacos , Benzamidas/metabolismo , Benzamidas/farmacologia , Sítios de Ligação , Candida albicans/efeitos dos fármacos , Cristalografia por Raios X , Farmacorresistência Fúngica/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Simulação de Dinâmica Molecular , Mutagênese Sítio-Dirigida , Proteínas de Transferência de Fosfolipídeos/química , Proteínas de Transferência de Fosfolipídeos/genética , Proteínas de Transferência de Fosfolipídeos/metabolismo , Ácidos Picolínicos/metabolismo , Ácidos Picolínicos/farmacologia , Estrutura Terciária de Proteína , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Alinhamento de Sequência , Relação Estrutura-Atividade
7.
Cell Rep ; 19(3): 451-460, 2017 04 18.
Artigo em Inglês | MEDLINE | ID: mdl-28423309

RESUMO

Flavivirus infections by Zika and dengue virus impose a significant global healthcare threat with no US Food and Drug Administration (FDA)-approved vaccination or specific antiviral treatment available. Here, we present the discovery of an anti-flaviviral natural product named cavinafungin. Cavinafungin is a potent and selectively active compound against Zika and all four dengue virus serotypes. Unbiased, genome-wide genomic profiling in human cells using a novel CRISPR/Cas9 protocol identified the endoplasmic-reticulum-localized signal peptidase as the efficacy target of cavinafungin. Orthogonal profiling in S. cerevisiae followed by the selection of resistant mutants pinpointed the catalytic subunit of the signal peptidase SEC11 as the evolutionary conserved target. Biochemical analysis confirmed a rapid block of signal sequence cleavage of both host and viral proteins by cavinafungin. This study provides an effective compound against the eukaryotic signal peptidase and independent confirmation of the recently identified critical role of the signal peptidase in the replicative cycle of flaviviruses.


Assuntos
Produtos Biológicos/farmacologia , Vírus da Dengue/fisiologia , Lipopeptídeos/farmacologia , Replicação Viral/efeitos dos fármacos , Zika virus/fisiologia , Produtos Biológicos/química , Sistemas CRISPR-Cas/genética , Vírus da Dengue/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Genoma Humano , Genômica , Células HCT116 , Humanos , Lipopeptídeos/química , Proteínas de Membrana , Subunidades Proteicas/metabolismo , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/genética , Serina Endopeptidases , Proteínas Virais/metabolismo , Zika virus/efeitos dos fármacos
8.
J Biomol Screen ; 21(3): 306-15, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26459507

RESUMO

Phenotypic screens are effective starting points to identify compounds with desirable activities. To find novel antifungals, we conducted a phenotypic screen in Saccharomyces cerevisiae and identified two discrete scaffolds with good growth inhibitory characteristics. Lack of broad-spectrum activity against pathogenic fungi called for directed chemical compound optimization requiring knowledge of the molecular target. Chemogenomic profiling identified effects on geranylgeranyltransferase I (GGTase I), an essential enzyme that prenylates proteins involved in cell signaling, such as Cdc42p and Rho1p. Selection of resistant mutants against both compounds confirmed the target hypothesis and enabled mapping of the compound binding site to the substrate binding pocket. Differential resistance-conferring mutations and selective substrate competition demonstrate distinct binding modes for the two chemotypes. Exchange of the S. cerevisiae GGTase I subunits with those of Candida albicans resulted in an absence of growth inhibition for both compounds, thus confirming the identified target as well as the narrow antifungal spectrum of activity. This prenylation pathway is reported to be nonessential in pathogenic species and challenges the therapeutic value of these leads while demonstrating the importance of an integrated target identification platform following a phenotypic screen.


Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Antifúngicos/farmacologia , Descoberta de Drogas/métodos , Inibidores Enzimáticos/farmacologia , Testes de Sensibilidade Microbiana , Alquil e Aril Transferases/química , Alquil e Aril Transferases/genética , Alquil e Aril Transferases/metabolismo , Antifúngicos/química , Farmacorresistência Fúngica , Inibidores Enzimáticos/química , Perfilação da Expressão Gênica , Metabolômica/métodos , Modelos Moleculares , Conformação Molecular , Mutação , Ligação Proteica , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/enzimologia , Saccharomyces cerevisiae/genética
9.
Nat Commun ; 6: 8613, 2015 Oct 12.
Artigo em Inglês | MEDLINE | ID: mdl-26456460

RESUMO

FR171456 is a natural product with cholesterol-lowering properties in animal models, but its molecular target is unknown, which hinders further drug development. Here we show that FR171456 specifically targets the sterol-4-alpha-carboxylate-3-dehydrogenase (Saccharomyces cerevisiae--Erg26p, Homo sapiens--NSDHL (NAD(P) dependent steroid dehydrogenase-like)), an essential enzyme in the ergosterol/cholesterol biosynthesis pathway. FR171456 significantly alters the levels of cholesterol pathway intermediates in human and yeast cells. Genome-wide yeast haploinsufficiency profiling experiments highlight the erg26/ERG26 strain, and multiple mutations in ERG26 confer resistance to FR171456 in growth and enzyme assays. Some of these ERG26 mutations likely alter Erg26 binding to FR171456, based on a model of Erg26. Finally, we show that FR171456 inhibits an artificial Hepatitis C viral replicon, and has broad antifungal activity, suggesting potential additional utility as an anti-infective. The discovery of the target and binding site of FR171456 within the target will aid further development of this compound.


Assuntos
3-Hidroxiesteroide Desidrogenases/antagonistas & inibidores , Antifúngicos/química , Colesterol/análogos & derivados , Proteínas de Saccharomyces cerevisiae/antagonistas & inibidores , Saccharomyces cerevisiae/genética , 3-Hidroxiesteroide Desidrogenases/genética , Candida albicans , Colesterol/química , Farmacorresistência Fúngica/genética , Ergosterol/biossíntese , Mutação , Proteínas de Saccharomyces cerevisiae/genética
10.
Angew Chem Int Ed Engl ; 54(35): 10149-54, 2015 Aug 24.
Artigo em Inglês | MEDLINE | ID: mdl-26179970

RESUMO

Cultivation of myxobacteria of the Nannocystis genus led to the isolation and structure elucidation of a class of novel cyclic lactone inhibitors of elongation factor 1. Whole genome sequence analysis and annotation enabled identification of the putative biosynthetic cluster and synthesis process. In biological assays the compounds displayed anti-fungal and cytotoxic activity. Combined genetic and proteomic approaches identified the eukaryotic translation elongation factor 1α (EF-1α) as the primary target for this compound class. Nannocystin A (1) displayed differential activity across various cancer cell lines and EEF1A1 expression levels appear to be the main differentiating factor. Biochemical and genetic evidence support an overlapping binding site of 1 with the anti-cancer compound didemnin B on EF-1α. This myxobacterial chemotype thus offers an interesting starting point for further investigations of the potential of therapeutics targeting elongation factor 1.


Assuntos
Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Compostos Macrocíclicos/farmacologia , Myxococcales/fisiologia , Neoplasias/patologia , Fator 1 de Elongação de Peptídeos/antagonistas & inibidores , Antifúngicos/química , Antifúngicos/farmacologia , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Candida albicans/efeitos dos fármacos , Genômica/métodos , Humanos , Compostos Macrocíclicos/química , Estrutura Molecular , Neoplasias/tratamento farmacológico , Fator 1 de Elongação de Peptídeos/genética , Fator 1 de Elongação de Peptídeos/metabolismo , Proteômica/métodos , Relação Estrutura-Atividade , Células Tumorais Cultivadas
11.
J Cell Sci ; 128(6): 1217-29, 2015 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-25616894

RESUMO

A new cyclic decadepsipeptide was isolated from Chaetosphaeria tulasneorum with potent bioactivity on mammalian and yeast cells. Chemogenomic profiling in S. cerevisiae indicated that the Sec61 translocon complex, the machinery for protein translocation and membrane insertion at the endoplasmic reticulum, is the target. The profiles were similar to those of cyclic heptadepsipeptides of a distinct chemotype (including HUN-7293 and cotransin) that had previously been shown to inhibit cotranslational translocation at the mammalian Sec61 translocon. Unbiased, genome-wide mutagenesis followed by full-genome sequencing in both fungal and mammalian cells identified dominant mutations in Sec61p (yeast) or Sec61α1 (mammals) that conferred resistance. Most, but not all, of these mutations affected inhibition by both chemotypes, despite an absence of structural similarity. Biochemical analysis confirmed inhibition of protein translocation into the endoplasmic reticulum of both co- and post-translationally translocated substrates by both chemotypes, demonstrating a mechanism independent of a translating ribosome. Most interestingly, both chemotypes were found to also inhibit SecYEG, the bacterial Sec61 translocon homolog. We suggest 'decatransin' as the name for this new decadepsipeptide translocation inhibitor.


Assuntos
Produtos Biológicos/farmacologia , Retículo Endoplasmático/efeitos dos fármacos , Proteínas de Membrana/metabolismo , Transporte Proteico/efeitos dos fármacos , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Animais , Ascomicetos/metabolismo , Células COS , Células Cultivadas , Chlorocebus aethiops , Células HCT116 , Humanos , Proteínas de Membrana/antagonistas & inibidores , Peptídeos Cíclicos/farmacologia , Polimorfismo de Nucleotídeo Único/genética , Canais de Translocação SEC , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crescimento & desenvolvimento
12.
Microbiol Res ; 169(2-3): 107-20, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24360837

RESUMO

Due to evolutionary conservation of biology, experimental knowledge captured from genetic studies in eukaryotic model organisms provides insight into human cellular pathways and ultimately physiology. Yeast chemogenomic profiling is a powerful approach for annotating cellular responses to small molecules. Using an optimized platform, we provide the relative sensitivities of the heterozygous and homozygous deletion collections for nearly 1800 biologically active compounds. The data quality enables unique insights into pathways that are sensitive and resistant to a given perturbation, as demonstrated with both known and novel compounds. We present examples of novel compounds that inhibit the therapeutically relevant fatty acid synthase and desaturase (Fas1p and Ole1p), and demonstrate how the individual profiles facilitate hypothesis-driven experiments to delineate compound mechanism of action. Importantly, the scale and diversity of tested compounds yields a dataset where the number of modulated pathways approaches saturation. This resource can be used to map novel biological connections, and also identify functions for unannotated genes. We validated hypotheses generated by global two-way hierarchical clustering of profiles for (i) novel compounds with a similar mechanism of action acting upon microtubules or vacuolar ATPases, and (ii) an un-annotated ORF, YIL060w, that plays a role in respiration in the mitochondria. Finally, we identify and characterize background mutations in the widely used yeast deletion collection which should improve the interpretation of past and future screens throughout the community. This comprehensive resource of cellular responses enables the expansion of our understanding of eukaryotic pathway biology.


Assuntos
Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/genética , Antifúngicos/farmacologia , Vias Biossintéticas , Farmacorresistência Fúngica , Regulação Fúngica da Expressão Gênica , Ensaios de Triagem em Larga Escala , Dados de Sequência Molecular , Filogenia , Saccharomyces cerevisiae/classificação , Saccharomyces cerevisiae/efeitos dos fármacos , Proteínas de Saccharomyces cerevisiae/metabolismo
13.
J Minim Invasive Gynecol ; 20(6): 891-3, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24183278

RESUMO

We report the case of a 37-year-old woman who developed idiopathic brachial plexus neuritis, also referred to as Parsonage-Turner syndrome, after laparoscopic excision of endometriosis. The differential diagnosis between this non-position-related neuritis and brachial plexus injury is discussed. The aim of this report was to raise awareness on this distressing postoperative complication.


Assuntos
Neurite do Plexo Braquial/diagnóstico , Plexo Braquial/lesões , Endometriose/cirurgia , Complicações Intraoperatórias/diagnóstico , Laparoscopia/efeitos adversos , Doenças Ovarianas/cirurgia , Adulto , Neurite do Plexo Braquial/etiologia , Diagnóstico Diferencial , Feminino , Humanos , Complicações Intraoperatórias/etiologia
14.
ACS Chem Biol ; 8(7): 1519-27, 2013 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-23614532

RESUMO

Translation initiation is an emerging target in oncology and neurobiology indications. Naturally derived and synthetic rocaglamide scaffolds have been used to interrogate this pathway; however, there is uncertainty regarding their precise mechanism(s) of action. We exploited the genetic tractability of yeast to define the primary effect of both a natural and a synthetic rocaglamide in a cellular context and characterized the molecular target using biochemical studies and in silico modeling. Chemogenomic profiling and mutagenesis in yeast identified the eIF (eukaryotic Initiation Factor) 4A helicase homologue as the primary molecular target of rocaglamides and defined a discrete set of residues near the RNA binding motif that confer resistance to both compounds. Three of the eIF4A mutations were characterized regarding their functional consequences on activity and response to rocaglamide inhibition. These data support a model whereby rocaglamides stabilize an eIF4A-RNA interaction to either alter the level and/or impair the activity of the eIF4F complex. Furthermore, in silico modeling supports the annotation of a binding pocket delineated by the RNA substrate and the residues identified from our mutagenesis screen. As expected from the high degree of conservation of the eukaryotic translation pathway, these observations are consistent with previous observations in mammalian model systems. Importantly, we demonstrate that the chemically distinct silvestrol and synthetic rocaglamides share a common mechanism of action, which will be critical for optimization of physiologically stable derivatives. Finally, these data confirm the value of the rocaglamide scaffold for exploring the impact of translational modulation on disease.


Assuntos
Benzofuranos/metabolismo , Fator de Iniciação 4F em Eucariotos/química , Fator de Iniciação 4F em Eucariotos/metabolismo , Saccharomyces cerevisiae/metabolismo , Benzofuranos/química , Sítios de Ligação , Modelos Biológicos , Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/genética , Triterpenos/química , Triterpenos/metabolismo
15.
Antimicrob Agents Chemother ; 57(5): 2272-80, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23478965

RESUMO

High-throughput phenotypic screening against the yeast Saccharomyces cerevisiae revealed a series of triazolopyrimidine-sulfonamide compounds with broad-spectrum antifungal activity, no significant cytotoxicity, and low protein binding. To elucidate the target of this series, we have applied a chemogenomic profiling approach using the S. cerevisiae deletion collection. All compounds of the series yielded highly similar profiles that suggested acetolactate synthase (Ilv2p, which catalyzes the first common step in branched-chain amino acid biosynthesis) as a possible target. The high correlation with profiles of known Ilv2p inhibitors like chlorimuron-ethyl provided further evidence for a similar mechanism of action. Genome-wide mutagenesis in S. cerevisiae identified 13 resistant clones with 3 different mutations in the catalytic subunit of acetolactate synthase that also conferred cross-resistance to established Ilv2p inhibitors. Mapping of the mutations into the published Ilv2p crystal structure outlined the chlorimuron-ethyl binding cavity, and it was possible to dock the triazolopyrimidine-sulfonamide compound into this pocket in silico. However, fungal growth inhibition could be bypassed through supplementation with exogenous branched-chain amino acids or by the addition of serum to the medium in all of the fungal organisms tested except for Aspergillus fumigatus. Thus, these data support the identification of the triazolopyrimidine-sulfonamide compounds as inhibitors of acetolactate synthase but suggest that targeting may be compromised due to the possibility of nutrient bypass in vivo.


Assuntos
Acetolactato Sintase/antagonistas & inibidores , Antifúngicos/farmacologia , Pirimidinas/farmacologia , Proteínas de Saccharomyces cerevisiae/antagonistas & inibidores , Saccharomyces cerevisiae/efeitos dos fármacos , Sulfonamidas/farmacologia , Compostos de Sulfonilureia/farmacologia , Acetolactato Sintase/química , Acetolactato Sintase/genética , Acetolactato Sintase/metabolismo , Aminoácidos de Cadeia Ramificada/metabolismo , Aminoácidos de Cadeia Ramificada/farmacologia , Antifúngicos/química , Domínio Catalítico/efeitos dos fármacos , Ensaios de Triagem em Larga Escala , Humanos , Testes de Sensibilidade Microbiana , Simulação de Acoplamento Molecular , Mutação , Ligação Proteica , Pirimidinas/química , Saccharomyces cerevisiae/enzimologia , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Soro/química , Soro/metabolismo , Sulfonamidas/química , Compostos de Sulfonilureia/química
16.
PLoS One ; 7(9): e42657, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22970117

RESUMO

Argyrins, produced by myxobacteria and actinomycetes, are cyclic octapeptides with antibacterial and antitumor activity. Here, we identify elongation factor G (EF-G) as the cellular target of argyrin B in bacteria, via resistant mutant selection and whole genome sequencing, biophysical binding studies and crystallography. Argyrin B binds a novel allosteric pocket in EF-G, distinct from the known EF-G inhibitor antibiotic fusidic acid, revealing a new mode of protein synthesis inhibition. In eukaryotic cells, argyrin B was found to target mitochondrial elongation factor G1 (EF-G1), the closest homologue of bacterial EF-G. By blocking mitochondrial translation, argyrin B depletes electron transport components and inhibits the growth of yeast and tumor cells. Further supporting direct inhibition of EF-G1, expression of an argyrin B-binding deficient EF-G1 L693Q variant partially rescued argyrin B-sensitivity in tumor cells. In summary, we show that argyrin B is an antibacterial and cytotoxic agent that inhibits the evolutionarily conserved target EF-G, blocking protein synthesis in bacteria and mitochondrial translation in yeast and mammalian cells.


Assuntos
Oligopeptídeos/metabolismo , Fator G para Elongação de Peptídeos/metabolismo , Sítio Alostérico , Sequência de Aminoácidos , Animais , Burkholderia/efeitos dos fármacos , Linhagem Celular Tumoral , Sequência Conservada , Cristalografia por Raios X , Humanos , Mamíferos , Testes de Sensibilidade Microbiana , Proteínas Mitocondriais/metabolismo , Dados de Sequência Molecular , Proteínas Mutantes/química , Proteínas Mutantes/metabolismo , Oligopeptídeos/química , Oligopeptídeos/farmacologia , Fator G para Elongação de Peptídeos/antagonistas & inibidores , Fator G para Elongação de Peptídeos/química , Ligação Proteica/efeitos dos fármacos , Pseudomonas aeruginosa/efeitos dos fármacos , Saccharomyces cerevisiae/metabolismo , Homologia de Sequência de Aminoácidos
17.
Cell Host Microbe ; 11(6): 654-63, 2012 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-22704625

RESUMO

With renewed calls for malaria eradication, next-generation antimalarials need be active against drug-resistant parasites and efficacious against both liver- and blood-stage infections. We screened a natural product library to identify inhibitors of Plasmodium falciparum blood- and liver-stage proliferation. Cladosporin, a fungal secondary metabolite whose target and mechanism of action are not known for any species, was identified as having potent, nanomolar, antiparasitic activity against both blood and liver stages. Using postgenomic methods, including a yeast deletion strains collection, we show that cladosporin specifically inhibits protein synthesis by directly targeting P. falciparum cytosolic lysyl-tRNA synthetase. Further, cladosporin is >100-fold more potent against parasite lysyl-tRNA synthetase relative to the human enzyme, which is conferred by the identity of two amino acids within the enzyme active site. Our data indicate that lysyl-tRNA synthetase is an attractive, druggable, antimalarial target that can be selectively inhibited.


Assuntos
Antimaláricos/farmacologia , Inibidores Enzimáticos/farmacologia , Fungos/química , Isocumarinas/farmacologia , Lisina-tRNA Ligase/antagonistas & inibidores , Plasmodium falciparum/enzimologia , Antimaláricos/isolamento & purificação , Linhagem Celular , Avaliação Pré-Clínica de Medicamentos/métodos , Inibidores Enzimáticos/isolamento & purificação , Humanos , Concentração Inibidora 50 , Isocumarinas/isolamento & purificação , Testes de Sensibilidade Parasitária , Plasmodium falciparum/efeitos dos fármacos , Biossíntese de Proteínas/efeitos dos fármacos , Proteínas de Protozoários/antagonistas & inibidores
18.
J Minim Invasive Gynecol ; 18(6): 792-5, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-22024265

RESUMO

Herein is described an anterior approach to uterine artery ligation during laparoscopic myomectomy and total laparoscopic hysterectomy. The anterior leaf of the broad ligament is opened and the uterine artery is clipped lateral to its crossing over the ureter. Outcome measures were completion of the procedure laparoscopically and the need for transfusion postoperatively. Thirty-eight myomectomies and 28 difficult total laparoscopic hysterectomies (primarily uteri with large myomas) were performed, with 1 conversion to laparotomy during myomectomy and 1 during hysterectomy, and 1 transfusion after total laparoscopic hysterectomy. The anterior approach to uterine artery ligation is an alternative method for treatment of uterine artery occlusion during laparoscopic myomectomy or hysterectomy performed to treat large myomas.


Assuntos
Laparoscopia/métodos , Ligadura/métodos , Miométrio/cirurgia , Artéria Uterina/cirurgia , Adulto , Feminino , Humanos , Histerectomia/métodos , Leiomioma/cirurgia , Resultado do Tratamento , Neoplasias Uterinas/cirurgia
19.
Fertil Steril ; 96(1): e1-3, 2011 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-21620392

RESUMO

OBJECTIVE: To describe a case in which a midline laparotomy for two presumed malignant masses instead revealed parasitic fibroids. DESIGN: Case report. SETTING: Tertiary-level private hospital. PATIENT(S): Woman with 7- and 13-cm abdominopelvic masses 3 years after a total laparoscopic hysterectomy for fibroids. INTERVENTION(S): Midline laparotomy, bilateral salpingooophorectomy, infracolic omentectomy, appendicectomy, para-aortic lymph node dissection, and high rectosigmoid resection with primary anastomosis. MAIN OUTCOME MEASURE(S): Histology of masses showed adenomyoma. RESULT(S): There was no evidence of malignancy. CONCLUSION(S): Morcellation of the fibroids during a total laparoscopic hysterectomy likely left fragments that formed iatrogenic parasitic fibroids, which led to the subsequent laparotomy and bowel resection for potential malignancy.


Assuntos
Adenomioma/parasitologia , Adenomioma/cirurgia , Colo/cirurgia , Doença Iatrogênica , Enteropatias Parasitárias/cirurgia , Leiomioma/parasitologia , Leiomioma/cirurgia , Cuidados Pré-Operatórios , Adenomioma/diagnóstico , Adulto , Colo/parasitologia , Colo/patologia , Feminino , Humanos , Enteropatias Parasitárias/diagnóstico , Leiomioma/complicações
20.
Fertil Steril ; 95(1): 261-3, 2011 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-20663497

RESUMO

OBJECTIVE: To describe the laparoscopic management of an interstitial gestation of a heterotopic pregnancy. DESIGN: Case report and technique description. SETTING: Tertiary-level private practice. PATIENT(S): Woman with a 6-week gestation spontaneous heterotopic twin pregnancy: one twin intrauterine, one interstitial. INTERVENTION(S): A purse-string suture was applied to the proximal portion of the interstitial heterotopic pregnancy. MAIN OUTCOME MEASURE(S): To enable a cornual resection to be performed with minimal bleeding and without recourse to laparotomy. RESULT(S): At 8 weeks gestation an ultrasound scan confirmed a viable singleton intrauterine pregnancy, but a scan at 12 weeks showed a missed miscarriage. CONCLUSION(S): The embedding of the suture into the uterine serosa prevents slipping of the ligature that could occur with a pretied loop.


Assuntos
Morte Fetal/cirurgia , Laparoscopia/métodos , Gravidez Ectópica/cirurgia , Técnicas de Sutura , Feminino , Humanos , Gravidez , Gêmeos
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