RESUMEN
Leishmaniasis is a group of neglected tropical diseases caused by at least 20 species of Leishmania protozoa, which are spread by the bite of infected sandflies. There are three main forms of the disease: cutaneous leishmaniasis (CL, the most common), visceral leishmaniasis (VL, also known as kala-azar, the most serious), and mucocutaneous leishmaniasis. One billion people live in areas endemic to leishmaniasis, with an annual estimation of 30,000 new cases of VL and more than 1 million of CL. New treatments for leishmaniasis are an urgent need, as the existing ones are inefficient, toxic, and/or expensive. We have revised the experimental structure-based drug design (SBDD) efforts applied to the discovery of new drugs against leishmaniasis. We have grouped the explored targets according to the metabolic pathways they belong to, and the key achieved advances are highlighted and evaluated. In most cases, SBDD studies follow high-throughput screening campaigns and are secondary to pharmacokinetic optimization, due to the majoritarian belief that there are few validated targets for SBDD in leishmaniasis. However, some SBDD strategies have significantly contributed to new drug candidates against leishmaniasis and a bigger number holds promise for future development.
Asunto(s)
Leishmania , Leishmaniasis Cutánea , Leishmaniasis Visceral , Humanos , Leishmaniasis Visceral/tratamiento farmacológico , Leishmaniasis Visceral/complicaciones , Leishmaniasis Visceral/epidemiología , Leishmaniasis Cutánea/tratamiento farmacológico , Leishmaniasis Cutánea/complicaciones , Leishmaniasis Cutánea/epidemiología , Ensayos Analíticos de Alto RendimientoRESUMEN
PURPOSE: The function of FAM177A1 and its relationship to human disease is largely unknown. Recent studies have demonstrated FAM177A1 to be a critical immune-associated gene. One previous case study has linked FAM177A1 to a neurodevelopmental disorder in 4 siblings. METHODS: We identified 5 individuals from 3 unrelated families with biallelic variants in FAM177A1. The physiological function of FAM177A1 was studied in a zebrafish model organism and human cell lines with loss-of-function variants similar to the affected cohort. RESULTS: These individuals share a characteristic phenotype defined by macrocephaly, global developmental delay, intellectual disability, seizures, behavioral abnormalities, hypotonia, and gait disturbance. We show that FAM177A1 localizes to the Golgi complex in mammalian and zebrafish cells. Intersection of the RNA sequencing and metabolomic data sets from FAM177A1-deficient human fibroblasts and whole zebrafish larvae demonstrated dysregulation of pathways associated with apoptosis, inflammation, and negative regulation of cell proliferation. CONCLUSION: Our data shed light on the emerging function of FAM177A1 and defines FAM177A1-related neurodevelopmental disorder as a new clinical entity.
RESUMEN
Bacteria of the genus Brucella are facultative intracellular parasites that cause brucellosis, a severe animal and human disease. Recently, a group of taxonomists merged the brucellae with the primarily free-living, phylogenetically related Ochrobactrum spp. in the genus Brucella. This change, founded only on global genomic analysis and the fortuitous isolation of some opportunistic Ochrobactrum spp. from medically compromised patients, has been automatically included in culture collections and databases. We argue that clinical and environmental microbiologists should not accept this nomenclature, and we advise against its use because (i) it was presented without in-depth phylogenetic analyses and did not consider alternative taxonomic solutions; (ii) it was launched without the input of experts in brucellosis or Ochrobactrum; (iii) it applies a non-consensus genus concept that disregards taxonomically relevant differences in structure, physiology, population structure, core-pangenome assemblies, genome structure, genomic traits, clinical features, treatment, prevention, diagnosis, genus description rules, and, above all, pathogenicity; and (iv) placing these two bacterial groups in the same genus creates risks for veterinarians, medical doctors, clinical laboratories, health authorities, and legislators who deal with brucellosis, a disease that is particularly relevant in low- and middle-income countries. Based on all this information, we urge microbiologists, bacterial collections, genomic databases, journals, and public health boards to keep the Brucella and Ochrobactrum genera separate to avoid further bewilderment and harm.
Asunto(s)
Brucella , Ochrobactrum , Ochrobactrum/clasificación , Ochrobactrum/genética , Ochrobactrum/patogenicidad , Ochrobactrum/fisiología , Brucella/clasificación , Brucella/genética , Brucella/patogenicidad , Brucella/fisiología , Terminología como Asunto , Filogenia , Brucelosis/tratamiento farmacológico , Brucelosis/microbiología , Humanos , Infecciones Oportunistas/microbiologíaRESUMEN
Brucellosis is a zoonotic disease caused by Gram-negative bacteria of the genus Brucella. These pathogens cause long-lasting infections, a process in which Brucella modifications in the lipopolysaccharide (LPS) and envelope lipids reduce pathogen-associated molecular pattern (PAMP) recognition, thus hampering innate immunity activation. In vivo models are essential to investigate bacterial virulence, mice being the most used model. However, ethical and practical considerations impede their use in high-throughput screening studies. Although lacking the complexity of the mammalian immune system, insects share key-aspects of innate immunity with mammals, and Galleria mellonella has been used increasingly as a model. G. mellonella larvae have been shown useful in virulence analyses, including Gram-negative pathogens like Klebsiella pneumoniae and Legionella pneumophila. To assess its potential to study Brucella virulence, we first evaluated larva survival upon infection with representative Brucella species (i.e.B. abortus 2308W, B. microti CCM4915 and B. suis biovar 2) and mutants in the VirB type-IV secretion system (T4SS) or in the LPS-O-polysaccharide (O-PS). As compared to K.pneumoniae, the Brucella spp. tested induced a delayed and less severe mortality profile consistent with an escape of innate immunity detection. Brucella replication within larvae was affected by the lack of O-PS, which is reminiscent of their attenuation in natural hosts. On the contrary, replication was not affected by T4SS dysfunction and the mutant induced only slightly less mortality (not statistically significant) than its parental strain. We also evaluated G. mellonella to efficiently recognise Brucella and their LPS by quantification of the pro-phenoloxidase system and melanisation activation, using Pseudomonas LPS as a positive control. Among the brucellae, only B. microti LPS triggered an early-melanisation response consistent with the slightly increased endotoxicity of this species in mice. Therefore, G. mellonella represents a tool to screen for potential Brucella factors modulating innate immunity, but its usefulness to investigate other mechanisms relevant in Brucella intracellular life is limited.
Asunto(s)
Brucella , Mariposas Nocturnas , Animales , Ratones , Mariposas Nocturnas/microbiología , Lipopolisacáridos , Larva/microbiología , Interacciones Huésped-Patógeno , MamíferosRESUMEN
The low aqueous solubility of colchicine site antimitotic agents, of which the trimethoxyphenyl (A ring) is a heavy contributor, is a serious drawback in their clinical development. We have designed new A ring analogs with chameleonic masked polar amino groups able to increase aqueous solubility and also behave as non-polar through intramolecular hydrogen bonds when bound to tubulin. We have incorporated these new A rings in several scaffolds (sulfonamides, combretastatins, phenstatins, isocombretastatins), synthesized, and assayed 43 representatives. The amino analogs show improved aqueous solubility and some of them (8, 60Z, and 67) nanomolar anti-proliferative potencies against human cancer cell lines, with the most favorable substituent being a 3-methylamino group. The antiproliferative effect relates to tubulin inhibition as shown by in vitro tubulin polymerization inhibition, immunofluorescence microscopy, and cell cycle and apoptosis analysis by flow cytometry. The compounds arrest the cell cycle of treated cells in G2/M and later develop an apoptotic response. Docking studies suggested binding at the colchicine site of tubulin with good agreement with the DFT models of the new structural variations made. The 3-methylamino-4,5dimethoxyphenyl moiety is an example of the masked polar group incorporation (MPGI) strategy for soluble ligands binding to hydrophobic sites and a good trimethoxyphenyl ring replacement for the development of new colchicine site ligands.
Asunto(s)
Antineoplásicos , Colchicina , Humanos , Colchicina/química , Línea Celular Tumoral , Sitios de Unión , Tubulina (Proteína)/metabolismo , Solubilidad , Relación Estructura-Actividad , Moduladores de Tubulina/farmacología , Moduladores de Tubulina/química , Antineoplásicos/farmacología , Antineoplásicos/química , Proliferación CelularRESUMEN
Antimitotic agents are one of the more successful types of anticancer drugs, but they suffer from toxicity and resistance. The application of approved drugs to new indications (i.e., drug repurposing) is a promising strategy for the development of new drugs. It relies on finding pattern similarities: drug effects to other drugs or conditions, similar toxicities, or structural similarity. Here, we recursively searched a database of approved drugs for structural similarity to several antimitotic agents binding to a specific site of tubulin, with the expectation of finding structures that could fit in it. These searches repeatedly retrieved frentizole, an approved nontoxic anti-inflammatory drug, thus indicating that it might behave as an antimitotic drug devoid of the undesired toxic effects. We also show that the usual repurposing approach to searching for targets of frentizole failed in most cases to find such a relationship. We synthesized frentizole and a series of analogs to assay them as antimitotic agents and found antiproliferative activity against HeLa tumor cells, inhibition of microtubule formation within cells, and arrest at the G2/M phases of the cell cycle, phenotypes that agree with binding to tubulin as the mechanism of action. The docking studies suggest binding at the colchicine site in different modes. These results support the repurposing of frentizole for cancer treatment, especially for glioblastoma.
Asunto(s)
Antimitóticos , Antineoplásicos , Antimitóticos/farmacología , Tubulina (Proteína)/metabolismo , Línea Celular Tumoral , Relación Estructura-Actividad , Colchicina/química , Antineoplásicos/farmacología , Antineoplásicos/química , Moduladores de Tubulina/química , Ensayos de Selección de Medicamentos Antitumorales , Proliferación Celular , Sitios de UniónRESUMEN
Three Gram-negative, rod-shaped, oxidase-positive, non-spore-forming, non-motile strains (C130915_07T, C150915_16 and C150915_17) were isolated from lymph nodes of Algerian cows. On the basis of 16S rRNA gene and whole genome similarities, the isolates were almost identical and clearly grouped in the genus Pseudochrobactrum. This allocation was confirmed by the analysis of fatty acids (C19:cyclo, C18â:â1, C18â:â0, C16â:â1 and C16â:â0) and of polar lipids (major components: phosphatidylethanolamine, ornithine-lipids, phosphatidylglycerol, cardiolipin and phosphatidylcholine, plus moderate amounts of phosphatidylmonomethylethanolamine, phosphatidyldimethylethanolamine and other aminolipids). Genomic, physiological and biochemical data differentiated these isolates from previously described Pseudochrobactrum species in DNA relatedness, carbon assimilation pattern and growth temperature range. Thus, these organisms represent a novel species of the genus Pseudochrobactrum, for which the name Pseudochrobactrum algeriensis sp. nov. is proposed (type strain C130915_07T=CECT30232T=LMG 32378T).
Asunto(s)
Brucellaceae/clasificación , Bovinos/microbiología , Ganglios Linfáticos , Filogenia , Animales , Técnicas de Tipificación Bacteriana , Composición de Base , Brucellaceae/aislamiento & purificación , ADN Bacteriano/genética , Ácidos Grasos/química , Femenino , Ganglios Linfáticos/microbiología , Fosfolípidos/química , ARN Ribosómico 16S/genética , Análisis de Secuencia de ADNRESUMEN
Brucella melitensis and Brucella ovis are gram-negative pathogens of sheep that cause severe economic losses and, although B. ovis is non-zoonotic, B. melitensis is the main cause of human brucellosis. B. melitensis carries a smooth (S) lipopolysaccharide (LPS) with an N-formyl-perosamine O-polysaccharide (O-PS) that is absent in the rough LPS of B. ovis. Their control and eradication require vaccination, but B. melitensis Rev 1, the only vaccine available, triggers anti-O-PS antibodies that interfere in the S-brucellae serodiagnosis. Since eradication and serological surveillance of the zoonotic species are priorities, Rev 1 is banned once B. melitensis is eradicated or where it never existed, hampering B. ovis control and eradication. To develop a B. ovis specific vaccine, we investigated three Brucella live vaccine candidates lacking N-formyl-perosamine O-PS: Bov::CAΔwadB (CO2-independent B. ovis with truncated LPS core oligosaccharide); Rev1::wbdRΔwbkC (carrying N-acetylated O-PS); and H38ΔwbkF (B. melitensis rough mutant with intact LPS core). After confirming their attenuation and protection against B. ovis in mice, were tested in rams for efficacy. H38ΔwbkF yielded similar protection to Rev 1 against B. ovis but Bov::CAΔwadB and Rev1::wbdRΔwbkC conferred no or poor protection, respectively. All H38ΔwbkF vaccinated rams developed a protracted antibody response in ELISA and immunoprecipitation B. ovis diagnostic tests. In contrast, all remained negative in Rose Bengal and complement fixation tests used routinely for B. melitensis diagnosis, though some became positive in S-LPS ELISA owing to LPS core epitope reactivity. Thus, H38ΔwbkF is an interesting candidate for the immunoprophylaxis of B. ovis in B. melitensis-free areas.
Asunto(s)
Vacuna contra la Brucelosis , Brucella melitensis , Brucella ovis , Brucelosis , Enfermedades de los Roedores , Enfermedades de las Ovejas , Animales , Anticuerpos Antibacterianos , Brucella melitensis/genética , Brucella ovis/genética , Brucelosis/prevención & control , Brucelosis/veterinaria , Masculino , Ratones , Ovinos , Enfermedades de las Ovejas/prevención & controlRESUMEN
OBJECTIVES: To assess the effectiveness and toxicity of regorafenib in patients with metastatic colorectal cancer (mCRC) in routine clinical practice, as well as predictive factors of effectiveness. METHODS: This was a retrospective multicenter study in patients with mCRC who received regorafenib from November 2013 to May 2020. Effectiveness was evaluated by overall survival (OS) and progression-free survival (PFS) using the Kaplan-Meier method. Cox regression was performed to determine survival predictors. RESULTS: Ninety patients were enrolled (median age, 64.3 years). Fifty-two patients (57.8%) were male, and 57 (63.3%) had an ECOG performance status (PS) of 0 to 1. Median follow-up was 2.80 months. Median OS was 8.03 months (95% CI, 5.90-10.17), and median PFS was 2.90 months (95% CI, 2.59-3.21). Eighty-eight patients (97.8%) experienced drug-related adverse events. The most frequent were fatigue in 66 patients (73.3%), followed by palmar-plantar erythrodysesthesia in 40 (44.4%). Low liver tumor burden score (LTBS) and good ECOG PS were independent OS predictive factors. CONCLUSIONS: Patients taking regorafenib had OS and PFS rates similar to those reported in previous randomized trials; the agent had a poor toxicity profile. We identified low LTBS and good ECOG PS as possible predictive factors of better OS, useful in selecting patients with mCRC who might benefit from regorafenib.
Asunto(s)
Neoplasias del Colon , Neoplasias Colorrectales , Neoplasias Hepáticas , Neoplasias del Recto , Humanos , Masculino , Persona de Mediana Edad , Femenino , Neoplasias Colorrectales/patología , Datos de Salud Recolectados Rutinariamente , Compuestos de Fenilurea/efectos adversos , Neoplasias Hepáticas/secundarioRESUMEN
Pan-Gyn cancers entail 1 in 5 cancer cases worldwide, breast cancer being the most commonly diagnosed and responsible for most cancer deaths in women. The high incidence and mortality of these malignancies, together with the handicaps of taxanes-first-line treatments-turn the development of alternative therapeutics into an urgency. Taxanes exhibit low water solubility that require formulations that involve side effects. These drugs are often associated with dose-limiting toxicities and with the appearance of multi-drug resistance (MDR). Here, we propose targeting tubulin with compounds directed to the colchicine site, as their smaller size offer pharmacokinetic advantages and make them less prone to MDR efflux. We have prepared 52 new Microtubule Destabilizing Sulfonamides (MDS) that mostly avoid MDR-mediated resistance and with improved aqueous solubility. The most potent compounds, N-methyl-N-(3,4,5-trimethoxyphenyl-4-methylaminobenzenesulfonamide 38, N-methyl-N-(3,4,5-trimethoxyphenyl-4-methoxy-3-aminobenzenesulfonamide 42, and N-benzyl-N-(3,4,5-trimethoxyphenyl-4-methoxy-3-aminobenzenesulfonamide 45 show nanomolar antiproliferative potencies against ovarian, breast, and cervix carcinoma cells, similar or even better than paclitaxel. Compounds behave as tubulin-binding agents, causing an evident disruption of the microtubule network, in vitro Tubulin Polymerization Inhibition (TPI), and mitotic catastrophe followed by apoptosis. Our results suggest that these novel MDS may be promising alternatives to taxane-based chemotherapy in chemoresistant Pan-Gyn cancers.
Asunto(s)
Antineoplásicos/farmacología , Sulfonamidas/farmacología , Taxoides/farmacología , Moduladores de Tubulina/farmacología , Tubulina (Proteína)/metabolismo , Antineoplásicos/uso terapéutico , Apoptosis/efectos de los fármacos , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Femenino , Células HT29 , Células HeLa , Humanos , Células MCF-7 , Neoplasias Ováricas/metabolismo , Neoplasias Ováricas/patología , Polimerizacion/efectos de los fármacos , Sulfonamidas/química , Sulfonamidas/uso terapéutico , Taxoides/uso terapéutico , Moduladores de Tubulina/química , Moduladores de Tubulina/uso terapéuticoRESUMEN
Brucella ovis is a non-zoonotic rough Brucella that causes genital lesions, abortions and increased perinatal mortality in sheep and is responsible for important economic losses worldwide. Research on virulence factors of B. ovis is necessary for deciphering the mechanisms that enable this facultative intracellular pathogen to establish persistent infections and for developing a species-specific vaccine, a need in areas where the cross-protecting ovine smooth B. melitensis Rev1 vaccine is banned. Although several B. ovis virulence factors have been identified, there is little information on its metabolic abilities and their role in virulence. Here, we report that deletion of pyruvate phosphate dikinase (PpdK, catalyzing the bidirectional conversion pyruvate â phosphoenolpyruvate) in B. ovis PA (virulent and CO2-dependent) impaired growth in vitro. In cell infection experiments, although showing an initial survival higher than that of the parental strain, this ppdK mutant was unable to multiply. Moreover, when inoculated at high doses in mice, it displayed an initial spleen colonization higher than that of the parental strain followed by a marked comparative decrease, an unusual pattern of attenuation in mice. A homologous mutant was also obtained in a B. ovis PA CO2-independent construct previously proposed for developing B. ovis vaccines to solve the problem that CO2-dependence represents for large scale production. This CO2-independent ppdK mutant reproduced the growth defect in vitro and the multiplication/clearance pattern in mouse spleens, and is thus an interesting vaccine candidate for the immunoprophylaxis of B. ovis ovine brucellosis.
Asunto(s)
Proteínas Bacterianas/genética , Brucella ovis/genética , Brucelosis/microbiología , Dióxido de Carbono/metabolismo , Eliminación de Gen , Piruvato Ortofosfato Diquinasa/genética , Animales , Proteínas Bacterianas/metabolismo , Brucella ovis/enzimología , Femenino , Genes Bacterianos , Ratones , Ratones Endogámicos BALB C , Piruvato Ortofosfato Diquinasa/metabolismoRESUMEN
Brucella is a genus of gram-negative bacteria that cause brucellosis. B. abortus and B. melitensis infect domestic ruminants while B. suis (biovars 1-3) infect swine, and all these bacteria but B. suis biovar 2 are zoonotic. Live attenuated B. abortus S19 and B. melitensis Rev1 are effective vaccines in domestic ruminants, though both can infect humans. However, there is no swine brucellosis vaccine. Here, we investigated the potential use as vaccines of B. suis biovar 2 rough (R) lipopolysaccharide (LPS) mutants totally lacking O-chain (Bs2ΔwbkF) or only producing internal O-chain precursors (Bs2Δwzm) and mutants with a smooth (S) LPS defective in the core lateral branch (Bs2ΔwadB and Bs2ΔwadD). We also investigated mutants in the pyruvate phosphate dikinase (Bs2ΔppdK) and phosphoenolpyruvate carboxykinase (Bs2ΔpckA) genes encoding enzymes bridging phosphoenolpyruvate and the tricarboxylic acid cycle. When tested in the OIE mouse model at the recommended R or S vaccine doses (108 and 105 CFU, respectively), CFU/spleen of all LPS mutants were reduced with respect to the wild type and decreased faster for the R than for the S mutants. At those doses, protection against B. suis was similar for Bs2ΔwbkF, Bs2Δwzm, Bs2ΔwadB and the Rev1 control (105 CFU). As described before for B. abortus, B. suis biovar 2 carried a disabled pckA so that a double mutant Bs2ΔppdKΔpckA had the same metabolic phenotype as Bs2ΔppdK and ppdK mutation was enough to generate attenuation. At 105 CFU, Bs2ΔppdK also conferred the same protection as Rev1. As compared to other B. suis vaccine candidates described before, the mutants described here simultaneously carry irreversible deletions easy to identify as vaccine markers, lack antibiotic-resistance markers and were obtained in a non-zoonotic background. Since R vaccines should not elicit antibodies to the S-LPS and wzm mutants carry immunogenic O-chain precursors and did not improve Bs2ΔwbkF, the latter seems a better R vaccine candidate than Bs2Δwzm. However, taking into account that all R vaccines interfere in ELISA and other widely used assays, whether Bs2ΔwbkF is advantageous over Bs2ΔwadB or Bs2ΔppdK requires experiments in the natural host.
Asunto(s)
Vacuna contra la Brucelosis/inmunología , Brucella suis/inmunología , Brucelosis/veterinaria , Enfermedades de los Porcinos/prevención & control , Animales , Brucelosis/prevención & control , Brucelosis/virología , Sus scrofa , Porcinos , Enfermedades de los Porcinos/virología , Vacunas Atenuadas/inmunologíaRESUMEN
In the original publication of this article [1], the corresponding author points out Pilar M. Muñoz and Raquel CondeAlvarez contributed equally to this work.
RESUMEN
Colchicine site antimitotic agents typically suffer from low aqueous solubilities and are formulated as phosphate prodrugs of phenolic groups. These hydroxyl groups are the aim of metabolic transformations leading to resistance. There is an urgent need for more intrinsically soluble analogues lacking these hydroxyl groups. The 3,4,5-trimethoxyphenyl ring of combretastatin A-4 is a liability in terms of solubility but it is considered essential for high cytotoxic and tubulin polymerization inhibitory (TPI) activity. We have synthesized 36 new analogues of combretastatin A-4 replacing the trimethoxyphenyl moiety with more polar pyridine based moieties, measured their aqueous solubility, and studied their anti-proliferative effects against 3 human cancer cell lines. We show here that pyridine rings can be successful replacements for the trimethoxyphenyl ring, resulting in potent and more soluble analogues. The more straightforward replacement, a 2,6-dimethoxypyridine ring led to inactive analogues, but a 2-methoxy-6-methylsulfanylpyridine moiety led to active analogues when combined with different B rings. This replacement led to potent cytotoxic activity against sensitive human cancer cell lines due to tubulin inhibition, as shown by cell cycle analysis, confocal microscopy, and tubulin polymerization inhibitory activity studies. Cell cycle analysis also showed apoptotic responses following treatment. Docking studies suggested binding at the colchicine site of tubulin and provided a good agreement with the observed SAR. A 2-methoxy-6-methylsulfanylpyridine moiety is a good trimethoxyphenyl ring replacement for the development of new colchicine site ligands.
Asunto(s)
Antineoplásicos/farmacología , Colchicina/farmacología , Piridinas/farmacología , Antineoplásicos/síntesis química , Antineoplásicos/química , Ciclo Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Colchicina/síntesis química , Colchicina/química , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Ligandos , Estructura Molecular , Polimerizacion/efectos de los fármacos , Piridinas/química , Solubilidad , Relación Estructura-Actividad , Tubulina (Proteína)/metabolismo , Células Tumorales CultivadasRESUMEN
Tubulin, the microtubules and their dynamic behavior are amongst the most successful antitumor, antifungal, antiparasitic, and herbicidal drug targets. Sulfonamides are exemplary drugs with applications in the clinic, in veterinary and in the agrochemical industry. This review summarizes the actual state and recent progress of both fields looking from the double point of view of the target and its drugs, with special focus onto the structural aspects. The article starts with a brief description of tubulin structure and its dynamic assembly and disassembly into microtubules and other polymers. Posttranslational modifications and the many cellular means of regulating and modulating tubulin's biology are briefly presented in the tubulin code. Next, the structurally characterized drug binding sites, their occupying drugs and the effects they induce are described, emphasizing on the structural requirements for high potency, selectivity, and low toxicity. The second part starts with a summary of the favorable and highly tunable combination of physical-chemical and biological properties that render sulfonamides a prototypical example of privileged scaffolds with representatives in many therapeutic areas. A complete description of tubulin-binding sulfonamides is provided, covering the different species and drug sites. Some of the antimitotic sulfonamides have met with very successful applications and others less so, thus illustrating the advances, limitations, and future perspectives of the field. All of them combine in a mechanism of action and a clinical outcome that conform efficient drugs.
Asunto(s)
Sulfonamidas/farmacología , Moduladores de Tubulina/farmacología , Secuencia de Aminoácidos , Animales , Ensayos Clínicos como Asunto , Humanos , Sulfonamidas/química , Andamios del Tejido/química , Tubulina (Proteína)/metabolismo , Moduladores de Tubulina/químicaRESUMEN
Acinetobacter baumannii causes a wide range of nosocomial infections. This pathogen is considered a threat to human health due to the increasingly frequent isolation of multidrug-resistant strains. There is a major gap in knowledge on the infection biology of A. baumannii, and only a few virulence factors have been characterized, including lipopolysaccharide. The lipid A expressed by A. baumannii is hepta-acylated and contains 2-hydroxylaurate. The late acyltransferases controlling the acylation of lipid A have been already characterized. Here, we report the characterization of A. baumannii LpxO, which encodes the enzyme responsible for the 2-hydroxylation of lipid A. By genetic methods and mass spectrometry, we demonstrate that LpxO catalyzes the 2-hydroxylation of the laurate transferred by A. baumannii LpxL. LpxO-dependent lipid A 2-hydroxylation protects A. baumannii from polymyxin B, colistin, and human ß-defensin 3. LpxO contributes to the survival of A. baumannii in human whole blood and is required for pathogen survival in the waxmoth Galleria mellonella LpxO also protects Acinetobacter from G. mellonella antimicrobial peptides and limits their expression. Further demonstrating the importance of LpxO-dependent modification in immune evasion, 2-hydroxylation of lipid A limits the activation of the mitogen-activated protein kinase Jun N-terminal protein kinase to attenuate inflammatory responses. In addition, LpxO-controlled lipid A modification mediates the production of the anti-inflammatory cytokine interleukin-10 (IL-10) via the activation of the transcriptional factor CREB. IL-10 in turn limits the production of inflammatory cytokines following A. baumannii infection. Altogether, our studies suggest that LpxO is a candidate for the development of anti-A. baumannii drugs.
Asunto(s)
Infecciones por Acinetobacter/microbiología , Acinetobacter baumannii/metabolismo , Acinetobacter baumannii/patogenicidad , Lípido A/metabolismo , Acinetobacter baumannii/genética , Acinetobacter baumannii/crecimiento & desarrollo , Aciltransferasas/genética , Aciltransferasas/metabolismo , Animales , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Humanos , Hidroxilación , Larva/microbiología , Lípido A/química , Masculino , Ratones , Ratones Endogámicos C57BL , Mariposas Nocturnas/microbiología , VirulenciaRESUMEN
Sheep brucellosis is a worldwide extended disease caused by B. melitensis and B. ovis, two species respectively carrying smooth or rough lipopolysaccharide. Vaccine B. melitensis Rev1 is used against B. melitensis and B. ovis but induces an anti-smooth-lipopolysaccharide response interfering with B. melitensis serodiagnosis, which precludes its use against B. ovis where B. melitensis is absent. In mice, Rev1 deleted in wbkC (Brucella lipopolysaccharide formyl-transferase) and carrying wbdR (E. coli acetyl-transferase) triggered antibodies that could be differentiated from those evoked by wild-type strains, was comparatively attenuated and protected against B. ovis, suggesting its potential as a B. ovis vaccine.
Asunto(s)
Amino Azúcares/farmacología , Vacuna contra la Brucelosis/farmacología , Brucella ovis/inmunología , Brucelosis/veterinaria , Polisacáridos/farmacología , Vacunas Atenuadas/farmacología , Animales , Brucelosis/prevención & control , Femenino , Ratones , Ratones Endogámicos BALB CRESUMEN
Colchicine site ligands suffer from low aqueous solubility due to the highly hydrophobic nature of the binding site. A new strategy for increasing molecular polarity without exposing polar groups-termed masked polar group incorporation (MPGI)-was devised and applied to nitrogenated combretastatin analogues. Bulky ortho substituents to the pyridine nitrogen hinder it from the hydrophobic pocket while increasing molecular polarity. The resulting analogues show improved aqueous solubilities and highly potent antiproliferative activity against several cancer cell lines of different origin. The more potent compounds showed moderate tubulin polymerization inhibitory activity, arrested the cell cycle of treated cells at the G2/M phase, and subsequently caused apoptotic cell death represented by the cells gathered at the subG0/G1 population after 48 h of treatment. Annexin V/Propidium Iodide (PI) double-positive cells observed after 72 h confirmed the induction of apoptosis. Docking studies suggest binding at the colchicine site of tubulin in a similar way as combretastatin A4, with the polar groups masked by the vicinal substituents. These results validate the proposed strategy for the design of colchicine site ligands and open a new road to increasing the aqueous solubility of ligands binding in apolar environments.
Asunto(s)
Bibencilos/química , Nitrógeno/química , Moduladores de Tubulina/química , Tubulina (Proteína)/metabolismo , Apoptosis/efectos de los fármacos , Sitios de Unión , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular , Colchicina/química , Diseño de Fármacos , Células HT29 , Células HeLa , Humanos , Ligandos , Células MCF-7 , Simulación del Acoplamiento Molecular , Piridinas/química , Solubilidad/efectos de los fármacos , Relación Estructura-ActividadRESUMEN
Brucella bacteria cause brucellosis, a major zoonosis whose control requires efficient diagnosis and vaccines. Identification of classical Brucella spp. has traditionally relied on phenotypic characterization, including surface antigens and 5-10% CO2 necessity for growth (CO2-dependence), a trait of Brucella ovis and most Brucella abortus biovars 1-4 strains. Although molecular tests are replacing phenotypic methods, CO2-dependence remains of interest as it conditions isolation and propagation and reflects Brucella metabolism, an area of active research. Here, we investigated the connection of CO2-dependence and carbonic anhydrases (CA), the enzymes catalyzing the hydration of CO2 to the bicarbonate used by anaplerotic and biosynthetic carboxylases. Based on the previous demonstration that B. suis carries two functional CAs (CAI and CAII), we analyzed the CA sequences of CO2-dependent and -independent brucellae and spontaneous mutants. The comparisons strongly suggested that CAII is not functional in CO2-dependent B. abortus and B. ovis, and that a modified CAII sequence explains the CO2-independent phenotype of spontaneous mutants. Then, by mutagenesis and heterologous plasmid complementation and chromosomal insertion we proved that CAI alone is enough to support CO2-independent growth of B. suis in rich media but not of B. abortus in rich media or B. suis in minimal media. Finally, we also found that insertion of a heterologous active CAII into B. ovis reverted the CO2-dependence but did not alter its virulence in the mouse model. These results allow a better understanding of central aspects of Brucella metabolism and, in the case of B. ovis, provide tools for large-scale production of diagnostic antigens and vaccines.
Asunto(s)
Proteínas Bacterianas/genética , Brucella abortus/genética , Brucella abortus/patogenicidad , Brucella ovis/genética , Brucella ovis/patogenicidad , Dióxido de Carbono/metabolismo , Anhidrasas Carbónicas/genética , Animales , Proteínas Bacterianas/metabolismo , Brucella abortus/metabolismo , Brucella ovis/metabolismo , Anhidrasas Carbónicas/metabolismo , Femenino , Ratones , Ratones Endogámicos BALB C , VirulenciaRESUMEN
The history of group I introns is characterized by repeated horizontal transfers, even among phylogenetically distant species. The symbiogenetic thalli of lichens are good candidates for the horizontal transfer of genetic material among distantly related organisms, such as fungi and green algae. The main goal of this study was to determine whether there were different trends in intron distribution and properties among Chlorophyte algae based on their phylogenetic relationships and living conditions. Therefore, we investigated the occurrence, distribution and properties of group I introns within the chloroplast LSU rDNA in 87 Chlorophyte algae including lichen and free-living Trebouxiophyceae compared to free-living non-Trebouxiophyceae species. Overall, our findings showed that there was high diversity of group I introns and homing endonucleases (HEs) between Trebouxiophyceae and non-Trebouxiophyceae Chlorophyte algae, with divergence in their distribution patterns, frequencies and properties. However, the differences between lichen Trebouxiophyceae and free-living Trebouxiophyceae were smaller. An exception was the cL2449 intron, which was closely related to ω elements in yeasts. Such introns seem to occur more frequently in lichen Trebouxiophyceae compared to free-living Trebouxiophyceae. Our data suggest that lichenization and maintenance of lichen symbiosis for millions of years of evolution may have facilitated horizontal transfers of specific introns/HEs between symbionts. The data also suggest that sequencing of more chloroplast genes harboring group I introns in diverse algal groups may help us to understand the group I intron/HE transmission process within these organisms.