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1.
ACS Chem Biol ; 18(2): 377-384, 2023 02 17.
Artigo em Inglês | MEDLINE | ID: mdl-36745020

RESUMO

Phosphatidylserine (PS) is a key lipid that plays important roles in disease-related biological processes, and therefore, the means to track PS in live cells are invaluable. Herein, we describe the metabolic labeling of PS in Saccharomyces cerevisiae cells using analogues of serine, a PS precursor, derivatized with azide moieties at either the amino (N-l-SerN3) or carbonyl (C-l-SerN3) groups. The conservative click tag modification enabled these compounds to infiltrate normal lipid biosynthetic pathways, thereby producing tagged PS molecules as supported by mass spectrometry studies, thin-layer chromatography (TLC) analysis, and further derivatization with fluorescent reporters via click chemistry to enable imaging in yeast cells. This approach shows strong prospects for elucidating the complex biosynthetic and trafficking pathways involving PS.


Assuntos
Fosfatidilserinas , Saccharomyces cerevisiae , Fosfatidilserinas/metabolismo , Saccharomyces cerevisiae/metabolismo , Química Click
2.
Chembiochem ; 20(2): 172-180, 2019 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-30098105

RESUMO

Phosphatidylinositol (PI) lipids control critical biological processes, so aberrant biosynthesis often leads to disease. As a result, the capability to track the production and localization of these compounds in cells is vital for elucidating their complex roles. Herein, we report the design, synthesis, and application of clickable myo-inositol probe 1 a for bioorthogonal labeling of PI products. To validate this platform, we initially conducted PI synthase assays to show that 1 a inhibits PI production in vitro. Fluorescence microscopy experiments next showed probe-dependent imaging in T-24 human bladder cancer and Candida albicans cells. Growth studies in the latter showed that replacement of myo-inositol with probe 1 a led to an enhancement in cell growth. Finally, fluorescence-based TLC analysis and mass spectrometry experiments support the labeling of PI lipids. This approach provides a promising means for tracking the complex biosynthesis and trafficking of these lipids in cells.


Assuntos
Corantes Fluorescentes/química , Inositol/química , Engenharia Metabólica , Fosfatidilinositóis/química , Candida albicans/citologia , Candida albicans/crescimento & desenvolvimento , Candida albicans/metabolismo , Células Cultivadas , Química Click , Corantes Fluorescentes/síntese química , Humanos , Inositol/síntese química , Imagem Óptica
3.
ACS Chem Neurosci ; 8(9): 1823-1829, 2017 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-28737885

RESUMO

Memantine was the first breakthrough medication for the treatment of moderate to severe Alzheimer's disease (AD) patients and represents a fundamentally new mechanism of action (moderate-affinity, uncompetitive, voltage-dependent, N-methyl-d-aspartate (NMDA) receptor antagonist that exhibits fast on/off kinetics) to modulate glutamatergic dysfunction. Since its approval by the FDA in 2003, memantine, alone and in combination with donepezil, has improved patient outcomes in terms of cognition, behavioral disturbances, daily functioning, and delaying time to institutionalization. In this review, we will highlight the historical significance of memantine to AD (and other neuropsychiatric disorders) as well as provide an overview of the synthesis, pharmacology, and drug metabolism of this unique NMDA uncompetitive antagonist that clearly secures its place among the Classics in Chemical Neuroscience.


Assuntos
Antagonistas de Aminoácidos Excitatórios/farmacologia , Memantina/farmacologia , Nootrópicos/farmacologia , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/metabolismo , Animais , Antagonistas de Aminoácidos Excitatórios/síntese química , Antagonistas de Aminoácidos Excitatórios/farmacocinética , Antagonistas de Aminoácidos Excitatórios/uso terapêutico , Humanos , Memantina/síntese química , Memantina/farmacocinética , Memantina/uso terapêutico , Estrutura Molecular , Nootrópicos/síntese química , Nootrópicos/farmacocinética , Nootrópicos/uso terapêutico , Receptores de N-Metil-D-Aspartato/metabolismo
4.
Bioconjug Chem ; 28(4): 923-932, 2017 04 19.
Artigo em Inglês | MEDLINE | ID: mdl-28248084

RESUMO

Artificial systems for controlled membrane fusion applicable for drug delivery would ideally use triggers that are orthogonal to biology. To apply the strain-promoted alkyne-azide cycloaddition (SPAAC) to drive membrane fusion, oxo-dibenzocyclooctyne (ODIBO)-lipid 1 was designed, synthesized, and studied alongside azadibenzocyclooctyne (ADIBO)-lipids 2-4 to assess fusion with liposomes containing azido-lipid 5. Lipids 1-2 were first shown to be effective for liposome derivatization. Next, fusion was evaluated using liposomes containing 1 and varying ratios of PC and PE via a FRET dilution fusion assay, and a 1:1 PC-to-PE ratio yielded the greatest signal change attributed to fusion. Finally, lipids 1-4 were compared, and 1 yielded the greatest triggering of fusion, while 2-4 yielded varying efficacies depending on the structural features of each lipid. Fusion was further validated through STEM studies showing larger multilamellar assemblies after liposome mixing, and FRET assay results supporting the mixing of liposome aqueous contents. This work provides a platform for triggered fusion toward drug delivery applications and an understanding of the effects of lipid structure and membrane composition on fusion.


Assuntos
Alcinos/química , Azidas/química , Ciclo-Octanos/química , Lipídeos/química , Lipossomos/química , Fusão de Membrana , Compostos Aza/química , Reação de Cicloadição , Lipossomos/ultraestrutura
5.
Bioconjug Chem ; 26(6): 1021-31, 2015 Jun 17.
Artigo em Inglês | MEDLINE | ID: mdl-25927978

RESUMO

For drug delivery purposes, the ability to conveniently attach a targeting moiety that will deliver drugs to cells and then enable controlled release of the active molecule after localization is desirable. Toward this end, we designed and synthesized clickable and photocleavable lipid analogue 1 to maximize the efficiency of bioconjugation and triggered release. This compound contains a dibenzocyclooctyne group for bioorthogonal derivatization linked via a photocleavable 2-nitrobenzyl moiety at the headgroup of a synthetic lipid backbone for targeting to cell membranes. To assess delivery and release using this system, we report fluorescence-based assays for liposomal modification and photocleavage in solution as well as through surface immobilization to demonstrate successful liposome functionalization and photoinduced release. In addition, fluorophore delivery to and release from live cells was confirmed and characterized using fluorescence microscopy and flow cytometry analysis in which 1 was delivered to cells, derivatized, and photocleaved. Finally, drug delivery studies were performed using an azide-tagged analogue of camptothecin, a potent anticancer drug that is challenging to deliver due to poor solubility. In this case, the ester attachment of the azide tag acted as a caging group for release by intracellular esterases rather than through photocleavage. This resulted in a dose-dependent response in the presence of liposomes containing delivery agent 1, confirming the ability of this compound to stimulate delivery to the cytoplasm of cells.


Assuntos
Antineoplásicos Fitogênicos/administração & dosagem , Camptotecina/administração & dosagem , Preparações de Ação Retardada/química , Lipídeos/química , Lipossomos/química , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/farmacologia , Azidas/química , Camptotecina/análogos & derivados , Camptotecina/farmacologia , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , Sistemas de Liberação de Medicamentos , Células HeLa , Humanos , Luz , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Imagem Óptica , Processos Fotoquímicos , Saccharomyces cerevisiae/citologia , Saccharomyces cerevisiae/metabolismo
6.
Chemistry ; 20(12): 3350-7, 2014 Mar 17.
Artigo em Inglês | MEDLINE | ID: mdl-24615893

RESUMO

Liposomes represent promising carriers for drug delivery applications. To maximize this potential, there has been significant interest in developing liposomal systems encapsulating molecular cargo that are highly stable until their contents are released remotely in a controlled manner. Herein, we describe the design, synthesis, and analysis of a photocleavable analogue of the ubiquitous lipid phosphoatidylcholine (PC) for the development of highly stable and controllable photodisruptable membranes. Our strategy was to develop a lipid that closely mimics the structure of PC to optimize favorable properties including biocompatibility and stability of subsequent liposomes when mixed with lipids possessing a broad range of physicochemical properties. Thus, NB-PC was designed, which contains a photocleavable 2-nitrobenzyl group embedded within the acyl chain at the sn-2 position. Following the synthesis of NB-PC, liposome disruption efficacy was evaluated through photolysis studies involving the detection of nile red release. Studies performed using a range of liposomes with different percentages of NB-PC, PC, phosphatidylethanolamine (PE), cholesterol, and polyethylene glycol-PE (PEG-PE) demonstrated minimal background release in controls, release efficacies that correlate directly with the amount of NB-PC incorporation, and that release is only minimally impacted by the inclusion of the lipids PE and cholesterol that possess disparate properties. These results demonstrate that the NB-PC system is a highly stable, flexible, and tunable system for photoinitiated release from liposomal systems.


Assuntos
Colesterol/química , Lipídeos/química , Lipossomos/química , Fosfatidilcolinas/síntese química , Fosfatidiletanolaminas/química , Polietilenoglicóis/química , Fenômenos Químicos , Sistemas de Liberação de Medicamentos , Cinética , Fosfatidilcolinas/química
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