RESUMO
In this report, a thorough evaluation of the use of aerobically initiated, metal-free hydroacylation of various C=C and N=N acceptor molecules with a wide range of aldehydes is presented. The aerobic-activation conditions that have been developed are in sharp contrast to previous conditions for hydroacylation, which tend to use transition metals, peroxides that require thermal or photochemical degradation, or N-heterocyclic carbenes. The mildness of the conditions enables a number of reactions involving sensitive reaction partners and, perhaps most significantly, allows for α-functionalised chiral aldehydes to undergo radical-based hydroacylation with complete retention of optical purity. We also demonstrate how the resulting hydroacylation products can be transformed into other useful intermediates, such as γ-keto-sulfonamides, sultams, sultones, cyclic N-sulfonyl imines and amides.
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Reversible protein biotinylation is readily affected via conjugation with a bromomaleimide-based reagent followed by reductive cleavage. The intermediate biotinylated protein constructs are stable at physiological temperature and pH 8.0. Quantitative reversibility is elegantly delivered under mild conditions of using a stoichiometric amount of a bis-thiol, thus providing an approach that will be of general interest in chemical biology and proteomics.
Assuntos
Marcadores de Afinidade/química , Biotina/química , Maleimidas/química , Estreptavidina/química , Concentração de Íons de Hidrogênio , Hidrólise , Modelos Moleculares , Estrutura Terciária de Proteína , TemperaturaRESUMO
Bromomaleimides are useful building blocks in synthesis and powerful reagents for the selective chemical modification of proteins. A mild new synthesis of these reagents is described, along with the convenient transferability of the approach to dithiomaleimides and bromopyridazinediones.
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Smooth converter: Bioconjugation of superfolder GFP involving the formation of an unusually stable, and unprecedented, cyclic sulfonium species is described. This sulfonium can undergo smooth reaction with a range of nucleophiles to give sulfur-, selenium- and azide-modified GFP derivatives in high conversions.
Assuntos
Proteínas de Fluorescência Verde/química , Compostos de Sulfônio/química , Cisteína , Proteínas de Fluorescência Verde/genética , Modelos Moleculares , Mutação , Conformação ProteicaRESUMO
The potential of the approach combining nuclear magnetic resonance (NMR) spectroscopy, relaxed grid search (RGS), molecular dynamics (MD) simulations, and quantum mechanical (QM) calculations for the determination of diastereomer configurations is demonstrated using four diastereomers of a trisubstituted epoxide. Since the change in configuration of the chiral center is expected to change the distribution of conformer populations (including those of side-chain rotamers), changes in NMR parameters [chemical shifts, J couplings, and nuclear Overhauser effects (NOEs)] are expected. The method therefore relies on (1) identification of possible conformations in each diastereomer using relaxed grid search analysis and MD simulations; (2) geometry optimizations of conformers selected from step (1), followed by calculations of their relative energies (populations) using QM methods; (3) calculations of averaged NMR parameters using QM methods; (4) matching calculated and experimental values of NMR parameters of diastereomers. The diastereomer configurations are considered resolved, if three NMR parameters different in nature, chemical shifts, J couplings, and NOEs, are in agreement. A further advantage of this method is that full structural and dynamics characterization of each of the diastereomers is achieved based on the joint analysis of experimental and computational data.
RESUMO
Isolated model anion chromophores of the green and cyan fluorescent proteins were generated in an electrospray ion source, and their photodetachment spectra were recorded using photoelectron imaging. Vertical photodetachment energies of 2.85(10) and 4.08(10) eV have been measured for the model green fluorescent protein chromophore anion, corresponding to photodetachment from the ground electronic state of the anion to the ground and first excited electronic states of the radical, respectively. For the model cyan fluorescent protein chromophore anion, vertical photodetachment energies of 2.88(10) and 3.96(10) eV have been measured, corresponding to detachment from the ground electronic state of the anion to the ground and first excited electronic states of the neutral radical, respectively. We also find evidence suggesting that autoionization of electronically excited states of the chromophore anions competes with direct photodetachment. For comparison and to benchmark our measurements, the vertical photodetachment energies of deprotonated phenol and indole anions have also been recorded and presented. Quantum chemistry calculations support our assignments. We discuss our results in the context of the isolated protein chromophore anions acting as electron donors, one of their potential biological functions.
Assuntos
Proteínas de Fluorescência Verde/química , Ânions/química , Elétrons , Modelos Moleculares , Estrutura Molecular , Espectroscopia Fotoeletrônica , Prótons , Teoria QuânticaRESUMO
The regioselective 1,3-dipolar cycloaddition of alpha-bromo-pentafluorophenyl vinylsulfonate with nitrile oxides has been used to rapidly access a range of 3,5-isoxazoles which could be converted directly to their corresponding sulfonamides.
Assuntos
Isoxazóis/química , Sulfonamidas/síntese química , Ácidos Sulfônicos/química , Ácidos Sulfônicos/síntese química , Compostos de Vinila/química , Compostos de Vinila/síntese química , Micro-Ondas , Nitrilas/química , Estereoisomerismo , Especificidade por Substrato , Sulfonamidas/químicaRESUMO
A new and simple protocol for the direct functionalisation of aldehydes with concomitant conversion into ketones via C-C bond formation has been developed. The reaction effectively enables the direct C-H activation of an aldehyde by mixing of an aldehyde and a vinyl sulfonate under aerobic conditions or using hydrogen peroxide as a sub-stoichiometric reagent.
Assuntos
Aldeídos/química , Alcanossulfonatos/química , Carbono/química , Cetonas/síntese química , Acilação , Catálise , Hidrogênio/química , Peróxido de Hidrogênio/químicaRESUMO
Microwave enhanced diversity-oriented synthesis (MEDOS) using palladium catalysed protocols is introduced as a powerful new strategy for the synthesis of systematically modified small molecules and is highlighted by application to functionalised flavones.
RESUMO
PURPOSE: The level of hypoxia in primary tumors has been linked both clinically and experimentally to the incidence of metastases. This study was designed to address the effect of selectively targeting hypoxic cells in primary tumors on subsequent presentation of metastasis. EXPERIMENTAL DESIGN: The murine KHT model was used as a reproducible temporal and spatial onset of metastases is revealed following treatment of primary ( approximately 400 mm(3)) s.c. tumors with a 25 Gy radiation dose. The bioreductive drugs tirapazamine and RB6145 were administered in multiple doses before radiotherapy. RESULTS: Fractionated treatment with both tirapazamine and RB6145 significantly reduced the hypoxic fraction of the primary tumor, as assessed by pimonidazole binding, and had no effect on the overall growth rate of the primary tumor. Excision assays showed an increased level of cell kill in tirapazamine-treated versus RB6145-treated tumors consistent with tirapazamine targeting hypoxic cells at a broader range of oxygen tensions than RB6145. Tirapazamine treatment significantly reduced the presentation of metastases following radiotherapy (P = 0.003 versus saline controls) whereas RB6145 had no effect. Local control rates increased from 20% to 32% and 50% when radiation was combined with RB6145 and tirapazamine, respectively. CONCLUSIONS: These data provide direct evidence that selective targeting of hypoxic cells in primary tumors is a viable approach in the control of metastatic disease. The enhanced efficacy of tirapazamine versus RB6145 suggests that the radioresistant cells at intermediate oxygen tensions, conducive to targeting with tirapazamine but not with the more stringent bioreductive RB6145, predominate in terms of linking primary tumor hypoxia and metastases.
Assuntos
Metástase Neoplásica/prevenção & controle , Sarcoma Experimental/tratamento farmacológico , Sarcoma Experimental/radioterapia , Triazinas/administração & dosagem , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Terapia Combinada , Feminino , Hipóxia , Camundongos , Camundongos Endogâmicos C3H , Terapia Neoadjuvante , Metástase Neoplásica/patologia , Transplante de Neoplasias , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/patologia , Nitroimidazóis/administração & dosagem , Nitroimidazóis/uso terapêutico , Radiossensibilizantes/administração & dosagem , Radiossensibilizantes/uso terapêutico , Sarcoma Experimental/patologia , Tirapazamina , Resultado do Tratamento , Triazinas/uso terapêuticoRESUMO
Anaplastic (ATC) and certain follicular thyroid-carcinomas (FTCs) are radioresistant. The Phosphatidylinositide 3-kinase (PI3K) pathway is commonly hyperactivated in thyroid-carcinomas. PI3K can modify the PI3K-related kinases (PIKKs) in response to radiation: How PIKKs interact with PI3K and contribute to radioresistance in thyroid-carcinomas is unknown. Further uncertainties exist in how these interactions function under the radioresistant hypoxic microenvironment. Under normoxia/anoxia, ATC (8505c) and FTC (FTC-133) cells were irradiated, with PI3K-inhibition (via GDC-0941 and PTEN-reconstitution into PTEN-null FTC-133s) and effects on PIKK-activation, DNA-damage, clonogenic-survival and cell cycle, assessed. FTC-xenografts were treated with 5 × 2 Gy, ± 50 mg/kg GDC-0941 (twice-daily; orally) for 14 days and PIKK-activation and tumour-growth assessed. PIKK-expression was additionally assessed in 12 human papillary thyroid-carcinomas, 13 FTCs and 12 ATCs. GDC-0941 inhibited radiation-induced activation of Ataxia-telangiectasia mutated (ATM), ATM-and Rad3-related (ATR) and DNA-dependent protein kinase catalytic subunit (DNA-PKcs). Inhibition of ATM and DNA-PKcs was PI3K-dependent, since activation was reduced in PTEN-reconstituted FTC-133s. Inhibition of PIKK-activation was greater under anoxia: Consequently, whilst DNA-damage was increased and prolonged under both normoxia and anoxia, PI3K-inhibition only reduced clonogenic-survival under anoxia. GDC-0941 abrogated radiation-induced cell cycle arrest, an effect most likely linked to the marked inhibition of ATR-activation. Importantly, GDC-0941 inhibited radiation-induced PIKK-activation in FTC-xenografts leading to a significant increase in time taken for tumours to triple in size: 26.5 ± 5 days (radiation-alone) versus 31.5 ± 5 days (dual-treatment). PIKKs were highly expressed across human thyroid-carcinoma classifications, with ATM scoring consistently lower. Interestingly, some loss of ATM and DNA-PKcs was observed. These data provide new insight into the mechanisms of hypoxia-associated radioresistance in thyroid-carcinoma.
Assuntos
Carcinoma/radioterapia , Fosfatidilinositol 3-Quinases/metabolismo , Neoplasias da Glândula Tireoide/radioterapia , Animais , Carcinoma/metabolismo , Carcinoma Papilar/metabolismo , Carcinoma Papilar/radioterapia , Ciclo Celular , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Dano ao DNA , Proteínas de Ligação a DNA/metabolismo , Relação Dose-Resposta à Radiação , Ativação Enzimática , Feminino , Histonas/metabolismo , Humanos , Hipóxia , Indazóis/farmacologia , Camundongos , Camundongos Nus , Transplante de Neoplasias , Oxigênio/química , Tolerância a Radiação , Transdução de Sinais/fisiologia , Sulfonamidas/farmacologia , Câncer Papilífero da Tireoide , Neoplasias da Glândula Tireoide/metabolismoRESUMO
Complications associated with the development of lung metastases have a detrimental effect on the overall survival rate of many cancer patients. Preclinical models that mimic the clinical aspects of lung metastases are an important tool in developing new therapy options for these patients. The commonly used intravenous models only recapitulate dissemination of cancer cells to the lungs via the haematological route. Here we compared spontaneous and intravenous lung metastases of the highly metastatic KHT mouse fibrosarcoma cells after injecting KHT cells into the subcutaneous layer of the skin or directly into the tail vein. In contrast to the intravenous model, metastases spontaneously arising from the subcutaneous tumours disseminated most consistent with the lymph nodes/lymphatics route and were more hypoxic than the metastases observed following tail-vein administration and haematological spread. To ascertain whether this impacted on drug response, we tested the effectiveness of the hypoxia-sensitive cytotoxin AQ4N (Banoxantrone) in both models. AQ4N was more effective as an anti-metastatic drug in mice with subcutaneous KHT tumours, significantly reducing the metastatic score. Complementing the KHT studies, pathology studies in additional models of spontaneous lung metastases showed haematological (HCT116 intrasplenic implant) or mixed haematological/lymphatic (B16 intradermal implant) spread. These data suggest that preclinical models can demonstrate differing, clinically relevant dissemination patterns, and that careful selection of preclinical models is required when evaluating new strategies for targeting metastatic disease.
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Antraquinonas/farmacologia , Neoplasias Colorretais/patologia , Fibrossarcoma/patologia , Hipóxia/fisiopatologia , Neoplasias Pulmonares/secundário , Linfonodos/patologia , Vasos Linfáticos/patologia , Sarcoma Experimental/patologia , Animais , Neoplasias Colorretais/tratamento farmacológico , Feminino , Fibrossarcoma/tratamento farmacológico , Humanos , Técnicas Imunoenzimáticas , Neoplasias Pulmonares/tratamento farmacológico , Linfonodos/efeitos dos fármacos , Metástase Linfática , Vasos Linfáticos/efeitos dos fármacos , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C3H , Sarcoma Experimental/tratamento farmacológico , Células Tumorais CultivadasRESUMO
In this communication we describe a novel strategy for the formation of valuable diaryl and aryl alkyl ketones from acyl hydrazides. A wide variety of ketones are prepared and the mild reaction conditions allow for the use of a range of functionalities, especially in the synthesis of diaryl ketones.
Assuntos
Hidrazinas/química , Cetonas/síntese química , Acilação , Amidas/química , Cetonas/química , Estrutura MolecularRESUMO
Local protein microenvironment is used to control the outcome of reaction between cysteine residues and 2,5-dibromohexanediamide. The differential reactivity is exploited to introduce two orthogonal reactive handles onto the surface of a double cysteine mutant of superfolder green fluorescent protein in a regioselective manner. Subsequent elaboration with commonly used thiol and alkyne containing reagents affects site-selective protein dual labelling.
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Herein we report the functionalisation of aldehydes via hydroacylation of azodicarboxylates. A range of functionalised aldehydes are employed as the limiting reagent including chiral non-racemic aldehydes bearing α-stereocentres which are functionalised giving access to enantiomerically pure products. The resultant hydrazides can be employed as acyl donors in the synthesis of amides.
Assuntos
Aldeídos/química , Ácidos Carboxílicos/química , Amidas/síntese química , Amidas/química , Catálise , Oxirredução , Estereoisomerismo , Água/químicaRESUMO
The development of methods for carbon-carbon bond formation under benign conditions is an ongoing challenge for the synthetic chemist. In recent years there has been considerable interest in using selective C-H activation as a direct route for generating reactive intermediates. In this article, we describe the use of aldehyde auto-oxidation as a simple, clean and effective method for C-H activation, resulting in the generation of an acyl radical. This acyl radical can be used for carbon-carbon bond formation and herein we describe the application of this method for the hydroacylation of alpha,beta-unsaturated esters without the requirement of additional catalysts or reagents. This methodology generates unsymmetrical ketones, which have been shown to have broad use in organic synthesis.
Assuntos
Carbono/química , Acilação , Aldeídos/química , Alcenos/química , Catálise , Ésteres , Cetonas/química , OxirreduçãoRESUMO
Herein we report a mild, facile method for the preparation of 1,4-keto-sulfonates and sulfones on water. Further synthetic manipulations can result in products that are not readily accessed by hydroacylation of electron rich alkenes.
RESUMO
A facile synthesis of dibenzopyrroloazepinones via an electrophilic cyclisation of a biphenyl-acyliminium ion is described; an unusual 1,2-phenyl shift occurs when the C-1' carbon is the more nucleophilic than the C-2' carbon.
Assuntos
Azepinas/química , Compostos de Bifenilo/química , Carbazóis/química , Colchicina/química , Azepinas/síntese química , Carbazóis/síntese química , Cristalografia por Raios X , Ciclização , Conformação MolecularRESUMO
Targeting host factors is a complementary strategy for the development of new antiviral drugs. We screened a library of isoxazolidine and isoxazole sulfonamides and found four compounds that inhibited HIV-1 infection in human CD4+ lymphocytic T cells with no toxicity at IC(90) concentrations. Structure-activity relationship showed that benzyl sulfonamides and a halo-substituted aromatic ring on the heterocycle scaffold were critical for antiretroviral activity. The size and position of the incorporated halogen had a marked effect on the antiretroviral activity. The sulfonamide derivatives had no significant effect on HIV-1 entry, reverse transcription and integration but impaired a step necessary for activation of viral gene expression. This step was Tat-independent, strongly suggesting that the target is a cell factor. A virus partially resistant to the least potent compounds could be selected but could not be propagated in the long term, consistent with the possibility that HIV-1 may be less likely to develop resistance against drugs targeting some host factors. Here, we provide evidence that novel synthetic methods can be applied to develop small molecules with antiretroviral activity that target host factors important for HIV-1 replication.
Assuntos
Fármacos Anti-HIV/química , HIV-1/efeitos dos fármacos , Isoxazóis/química , Sulfonamidas/química , Fármacos Anti-HIV/síntese química , Fármacos Anti-HIV/toxicidade , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Linhagem Celular , Farmacorresistência Viral/efeitos dos fármacos , HIV-1/fisiologia , Células HeLa , Humanos , Isoxazóis/síntese química , Isoxazóis/toxicidade , Relação Estrutura-Atividade , Sulfonamidas/síntese química , Sulfonamidas/toxicidade , Replicação Viral/efeitos dos fármacosRESUMO
A highly efficient, practical and flexible two-step asymmetric synthesis of the beta(2)-selective beta-blocker ICI 118,551 is reported, allowing an unambiguous determination of the dependency of biological activity with optical activity, revealing the S,S-enantiomer to be the most potent.