RESUMO
Fluoroalkyl groups profoundly affect the physical properties of pharmaceuticals and influence almost all metrics associated with their pharmacokinetic and pharmacodynamic profile1-4. Drug candidates increasingly contain trifluoromethyl (CF3) and difluoromethyl (CF2H) groups, and the same trend in agrochemical development shows that the effect of fluoroalkylation translates across human, insect and plant life5,6. New fluoroalkylation reactions have undoubtedly stimulated this shift; however, methods that directly convert C-H bonds into C-CF2X groups (where X is F or H) in complex drug-like molecules are rare7-13. Pyridines are the most common aromatic heterocycles in pharmaceuticals14, but only one approach-via fluoroalkyl radicals-is viable for achieving pyridyl C-H fluoroalkylation in the elaborate structures encountered during drug development15-17. Here we develop a set of bench-stable fluoroalkylphosphines that directly convert the C-H bonds in pyridine building blocks, drug-like fragments and pharmaceuticals into fluoroalkyl derivatives. No preinstalled functional groups or directing groups are required. The reaction tolerates a variety of sterically and electronically distinct pyridines, and is exclusively selective for the 4-position in most cases. The reaction proceeds through initial formation of phosphonium salts followed by sp2-sp3 coupling of phosphorus ligands-an underdeveloped manifold for forming C-C bonds.
Assuntos
Carbono/química , Flúor/química , Hidrogênio/química , Fósforo/química , Piridinas/química , Alquilação , Animais , Humanos , Ligantes , Preparações Farmacêuticas/química , Farmacocinética , Fosfinas/químicaRESUMO
Fluorine magnetic resonance imaging (19F-MRI) is particularly promising for biomedical applications owing to the absence of fluorine in most biological systems. However, its use has been limited by the lack of safe and water-soluble imaging agents with high fluorine contents and suitable relaxation properties. We report innovative 19F-MRI agents based on supramolecular dendrimers self-assembled by an amphiphilic dendrimer composed of a hydrophobic alkyl chain and a hydrophilic dendron. Specifically, this amphiphilic dendrimer bears multiple negatively charged terminals with high fluorine content, which effectively prevented intra- and intermolecular aggregation of fluorinated entities via electrostatic repulsion. This permitted high fluorine nuclei mobility alongside good water solubility with favorable relaxation properties for use in 19F-MRI. Importantly, the self-assembling 19F-MRI agent was able to encapsulate the near-infrared fluorescence (NIRF) agent DiR and the anticancer drug paclitaxel for multimodal 19F-MRI and NIRF imaging of and theranostics for pancreatic cancer, a deadly disease for which there remains no adequate early detection method or efficacious treatment. The 19F-MRI and multimodal 19F-MRI and NIRF imaging studies on human pancreatic cancer xenografts in mice confirmed the capability of both imaging modalities to specifically image the tumors and demonstrated the efficacy of the theranostic agent in cancer treatment, largely outperforming the clinical anticancer drug paclitaxel. Consequently, these dendrimer nanosystems constitute promising 19F-MRI agents for effective cancer management. This study offers a broad avenue to the construction of 19F-MRI agents and theranostics, exploiting self-assembling supramolecular dendrimer chemistry.
Assuntos
Dendrímeros , Flúor , Nanomedicina Teranóstica , Dendrímeros/química , Animais , Nanomedicina Teranóstica/métodos , Humanos , Camundongos , Flúor/química , Paclitaxel/química , Paclitaxel/uso terapêutico , Imageamento por Ressonância Magnética/métodos , Linhagem Celular Tumoral , Neoplasias Pancreáticas/diagnóstico por imagem , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/terapia , Imagem por Ressonância Magnética de Flúor-19/métodos , Camundongos Nus , Meios de Contraste/químicaRESUMO
Tertiary stereogenic centres containing one fluorine atom are valuable for medicinal chemistry because they mimic common tertiary stereogenic centres containing one hydrogen atom, but they possess distinct charge distribution, lipophilicity, conformation and metabolic stability1-3. Although tertiary stereogenic centres containing one hydrogen atom are often set by enantioselective desymmetrization reactions at one of the two carbon-hydrogen (C-H) bonds of a methylene group, tertiary stereocentres containing fluorine have not yet been constructed by the analogous desymmetrization reaction at one of the two carbon-fluorine (C-F) bonds of a difluoromethylene group3. Fluorine atoms are similar in size to hydrogen atoms but have distinct electronic properties, causing C-F bonds to be exceptionally strong and geminal C-F bonds to strengthen one another4. Thus, exhaustive defluorination typically dominates over the selective replacement of a single C-F bond, hindering the development of the enantioselective substitution of one fluorine atom to form a stereogenic centre5,6. Here we report the catalytic, enantioselective activation of a single C-F bond in an allylic difluoromethylene group to provide a broad range of products containing a monofluorinated tertiary stereogenic centre. By combining a tailored chiral iridium phosphoramidite catalyst, which controls regioselectivity, chemoselectivity and enantioselectivity, with a fluorophilic activator, which assists the oxidative addition of the C-F bond, these reactions occur in high yield and selectivity. The design principles proposed in this work extend to palladium-catalysed benzylic substitution, demonstrating the generality of the approach.
Assuntos
Carbono/química , Flúor/química , Alcenos/química , Catálise , Cátions , Halogenação , Hidrogênio/química , Irídio/química , Compostos Organofosforados/química , Oxirredução , Paládio/químicaRESUMO
Frequently referred to as the 'magic methyl effect', the installation of methyl groups-especially adjacent (α) to heteroatoms-has been shown to dramatically increase the potency of biologically active molecules1-3. However, existing methylation methods show limited scope and have not been demonstrated in complex settings1. Here we report a regioselective and chemoselective oxidative C(sp3)-H methylation method that is compatible with late-stage functionalization of drug scaffolds and natural products. This combines a highly site-selective and chemoselective C-H hydroxylation with a mild, functional-group-tolerant methylation. Using a small-molecule manganese catalyst, Mn(CF3PDP), at low loading (at a substrate/catalyst ratio of 200) affords targeted C-H hydroxylation on heterocyclic cores, while preserving electron-neutral and electron-rich aryls. Fluorine- or Lewis-acid-assisted formation of reactive iminium or oxonium intermediates enables the use of a mildly nucleophilic organoaluminium methylating reagent that preserves other electrophilic functionalities on the substrate. We show this late-stage C(sp3)-H methylation on 41 substrates housing 16 different medicinally important cores that include electron-rich aryls, heterocycles, carbonyls and amines. Eighteen pharmacologically relevant molecules with competing sites-including drugs (for example, tedizolid) and natural products-are methylated site-selectively at the most electron rich, least sterically hindered position. We demonstrate the syntheses of two magic methyl substrates-an inverse agonist for the nuclear receptor RORc and an antagonist of the sphingosine-1-phosphate receptor-1-via late-stage methylation from the drug or its advanced precursor. We also show a remote methylation of the B-ring carbocycle of an abiraterone analogue. The ability to methylate such complex molecules at late stages will reduce synthetic effort and thereby expedite broader exploration of the magic methyl effect in pursuit of new small-molecule therapeutics and chemical probes.
Assuntos
Produtos Biológicos/química , Produtos Biológicos/síntese química , Carbono/química , Técnicas de Química Sintética , Hidrogênio/química , Preparações Farmacêuticas/química , Preparações Farmacêuticas/síntese química , Androstenos/síntese química , Androstenos/química , Catálise , Agonismo Inverso de Drogas , Elétrons , Flúor/química , Hidroxilação , Ácidos de Lewis/química , Manganês/química , Metilação , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/agonistas , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/antagonistas & inibidores , Oxazolidinonas/síntese química , Oxazolidinonas/química , Oxirredução , Receptores de Esfingosina-1-Fosfato/antagonistas & inibidores , Tetrazóis/síntese química , Tetrazóis/químicaRESUMO
Despite the importance of proline conformational equilibria (trans versus cis amide and exo versus endo ring pucker) on protein structure and function, there is a lack of convenient ways to probe proline conformation. 4,4-Difluoroproline (Dfp) was identified to be a sensitive 19F NMR-based probe of proline conformational biases and cis-trans isomerism. Within model compounds and disordered peptides, the diastereotopic fluorines of Dfp exhibit similar chemical shifts (ΔδFF = 0-3 ppm) when a trans X-Dfp amide bond is present. In contrast, the diastereotopic fluorines exhibit a large (ΔδFF = 5-12 ppm) difference in chemical shift in a cis X-Dfp prolyl amide bond. DFT calculations, X-ray crystallography, and solid-state NMR spectroscopy indicated that ΔδFF directly reports on the relative preference of one proline ring pucker over the other: a fluorine which is pseudo-axial (i.e., the pro-4R-F in an exo ring pucker, or the pro-4S-F in an endo ring pucker) is downfield, while a fluorine which is pseudo-equatorial (i.e., pro-4S-F when exo, or pro-4R-F when endo) is upfield. Thus, when a proline is disordered (a mixture of exo and endo ring puckers, as at trans-Pro in peptides in water), it exhibits a small Δδ. In contrast, when the Pro is ordered (i.e., when one ring pucker is strongly preferred, as in cis-Pro amide bonds, where the endo ring pucker is strongly favored), a large Δδ is observed. Dfp can be used to identify inherent induced order in peptides and to quantify proline cis-trans isomerism. Using Dfp, we discovered that the stable polyproline II helix (PPII) formed in the denatured state (8 M urea) exhibits essentially equal populations of the exo and endo proline ring puckers. In addition, the data with Dfp suggested the specific stabilization of PPII by water over other polar solvents. These data strongly support the importance of carbonyl solvation and n â π* interactions for the stabilization of PPII. Dfp was also employed to quantify proline cis-trans isomerism as a function of phosphorylation and the R406W mutation in peptides derived from the intrinsically disordered protein tau. Dfp is minimally sterically disruptive and can be incorporated in expressed proteins, suggesting its broad application in understanding proline cis-trans isomerization, protein folding, and local order in intrinsically disordered proteins.
Assuntos
Flúor , Prolina , Prolina/química , Prolina/análogos & derivados , Flúor/química , Cristalografia por Raios X/métodos , Conformação Proteica , Espectroscopia de Ressonância Magnética/métodos , Peptídeos/química , Ressonância Magnética Nuclear Biomolecular/métodos , Conformação MolecularRESUMO
Global substitution of leucine for analogues containing CH2F instead of methyl groups delivers proteins with multiple sites for monitoring by 19F nuclear magnetic resonance (NMR) spectroscopy. The 19 kDa Escherichia coli peptidyl-prolyl cis-trans isomerase B (PpiB) was prepared with uniform high-level substitution of leucine by (2S,4S)-5-fluoroleucine, (2S,4R)-5-fluoroleucine, or 5,5'-difluoroleucine. The stability of the samples toward thermal denaturation was little altered compared to the wild-type protein. 19F nuclear magnetic resonance (NMR) spectra showed large chemical shift dispersions between 6 and 17 ppm. The 19F chemical shifts correlate with the three-bond 1H-19F couplings (3JHF), providing the first experimental verification of the γ-gauche effect predicted by [Feeney, J. J. Am. Chem. Soc. 1996, 118, 8700-8706] and establishing the effect as the predominant determinant of the 19F chemical shifts of CH2F groups. Individual CH2F groups can be confined to single rotameric states by the protein environment, but most CH2F groups exchange between different rotamers at a rate that is fast on the NMR chemical shift scale. Interactions between fluorine atoms in 5,5'-difluoroleucine bias the CH2F rotamers in agreement with results obtained previously for 1,3-difluoropropane. The sensitivity of the 19F chemical shift to the rotameric state of the CH2F groups potentially renders them particularly sensitive for detecting allosteric effects.
Assuntos
Proteínas de Escherichia coli , Escherichia coli , Peptidilprolil Isomerase , Peptidilprolil Isomerase/metabolismo , Peptidilprolil Isomerase/química , Escherichia coli/metabolismo , Escherichia coli/genética , Escherichia coli/enzimologia , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Ligantes , Ressonância Magnética Nuclear Biomolecular/métodos , Leucina/química , Leucina/metabolismo , Leucina/análogos & derivados , Flúor/químicaRESUMO
Cytochromes P450 (CYPs) are heme-containing enzymes that are present in all kingdoms of life and share a structurally homologous, globular protein fold. CYPs utilize structures distal to the heme to recognize and coordinate substrates, while the necessary interactions with redox partner proteins are mediated at the opposite, proximal surface. In the current study, we investigated the functional allostery across the heme for the bacterial enzyme CYP121A1, which utilizes a non-polar distal-to-distal dimer interface for specific binding of its dicyclotyrosine substrate. Fluorine-detected Nuclear Magnetic Resonance (19F-NMR) spectroscopy was combined with site-specific labeling of a distal surface residue (S171C of the FG-loop), one residue of the B-helix (N84C), and two proximal surface residues (T103C and T333C) with a thiol-reactive fluorine label. Adrenodoxin was used as a substitute redox protein and was found to promote a closed arrangement of the FG-loop, similar to the addition of substrate alone. Disruption of the protein-protein interface by mutagenesis of two CYP121 basic surface residues removed the allosteric effect. Moreover, 19F-NMR spectra of the proximal surface indicate that ligand-induced allostery modulates the environment at the C-helix but not the meander region of the enzyme. In light of the high degree of structural homology in this family of enzymes, we interpret the findings from this work to represent a conserved allosteric network in CYPs.
Assuntos
Sistema Enzimático do Citocromo P-450 , Heme , Sistema Enzimático do Citocromo P-450/química , Sistema Enzimático do Citocromo P-450/genética , Flúor/química , Heme/química , Mutagênese , Oxirredução , Regulação AlostéricaRESUMO
Fluorine NMR is a highly sensitive technique for delineating the conformational states of biomolecules and has shown great utility in drug screening and in understanding protein function. Current fluorinated protein tags leverage the intrinsic chemical shift sensitivity of the 19F nucleus to detect subtle changes in protein conformation and topology. This chemical shift sensitivity can be amplified by embedding the fluorine or trifluoromethyl reporter within a pyridone. Due to their polarizability and rapid tautomerization, pyridones exhibit a greater range of electron delocalization and correspondingly greater 19F NMR chemical shift dispersion. To assess the chemical shift sensitivity of these tautomeric probes to the local environment, 19F NMR spectra of all possible monofluorinated and trifluoromethyl-tagged versions of 2-pyridone were recorded in methanol/water mixtures ranging from 100% methanol to 100% water. 4-Fluoro-2-pyridone and 6-(trifluoromethyl)-2-pyridone (6-TFP) displayed the greatest sensitivity of the monofluorinated and trifluoromethylated pyridones, exceeding that of known conventional CF3 reporters. To evaluate the utility of tautomeric pyridone tags for 19F NMR of biomolecules, the alpha subunit of the stimulatory G protein (Gsα) and human serum albumin (HSA) were each labeled with a thiol-reactive derivative of 6-TFP and the spectra were recorded as a function of various adjuvants and drugs. The tautomeric tag outperformed the conventional tag, 2-bromo-N-(4-(trifluoromethyl)phenyl)acetamide through the improved resolution of several functional states.
Assuntos
Flúor , Metanol , Humanos , Flúor/química , Espectroscopia de Ressonância Magnética/métodos , Conformação Proteica , Água , PiridonasRESUMO
The substitution of a single hydrogen atom in a protein by fluorine yields a site-specific probe for sensitive detection by 19F nuclear magnetic resonance (NMR) spectroscopy, where the absence of background signal from the protein facilitates the detection of minor conformational species. We developed genetic encoding systems for the site-selective incorporation of 4-fluorotryptophan, 5-fluorotryptophan, 6-fluorotryptophan, and 7-fluorotryptophan in response to an amber stop codon and used them to investigate conformational heterogeneity in a designed amino acid binding protein and in flaviviral NS2B-NS3 proteases. These proteases have been shown to present variable conformations in X-ray crystal structures, including flips of the indole side chains of tryptophan residues. The 19F NMR spectra of different fluorotryptophan isomers installed at the conserved site of Trp83 indicate that the indole ring flip is common in flaviviral NS2B-NS3 proteases in the apo state and suppressed by an active-site inhibitor.
Assuntos
Conformação Proteica , Triptofano , Triptofano/química , Triptofano/análogos & derivados , Modelos Moleculares , Ressonância Magnética Nuclear Biomolecular , Flúor/química , Proteínas/químicaRESUMO
The focus of this project is to take advantage of the large NMR chemical shift anisotropy of 19F to determine the orientation of fluorine labeled biomolecules in situ in oriented biological systems such as muscle. The difficulty with a single fluorine atom is that the orientation determined from a chemical shift is not singlevalued in the case of a fully anisotropic chemical shift tensor. The utility of a labeling approach with two fluorine labels in a fixed molecular framework where one of the labels has an axially symmetric chemical shift anisotropy such as a CF3 group and the other has a fully asymmetric chemical shift anisotropy such as 5-fluorotryptophan is evaluated. The result is that the orientation of the label can be determined straightforwardly from a single one-dimensional 19F NMR spectrum. The potential applications are widespread and not limited to biological applications.
Assuntos
Flúor , Ressonância Magnética Nuclear Biomolecular , Flúor/química , Ressonância Magnética Nuclear Biomolecular/métodos , Anisotropia , Marcação por Isótopo/métodosRESUMO
Metabolite profiling serves as a powerful tool that advances our understanding of biological systems, disease mechanisms, and environmental interactions. In this study, we present an approach employing 19F-nuclear magnetic resonance (19F NMR) spectroscopy for plasma amine profiling. This method utilizes a highly efficient and reliable fluorine-labeling reagent, 3,5-difluorosalicylaldehyde, which effectively emulates pyridoxal phosphate, facilitating the formation of Schiff base compounds with primary amines. The fluorine labeling allows for distinct resolution of 19F NMR signals from amine mixtures, leading to precise identification and quantification of amine metabolites in human plasma. This advancement offers valuable tools for furthering metabolomics research.
Assuntos
Aminas , Flúor , Humanos , Flúor/química , Espectroscopia de Ressonância Magnética/métodos , Metabolômica/métodos , Imageamento por Ressonância MagnéticaRESUMO
Kidney diseases have become an important global health concern due to their high incidence, inefficient diagnosis, and poor prognosis. Devising direct methods, especially imaging means, to assess renal function is the key for better understanding the mechanisms of various kidney diseases and subsequent development of effective treatment. Herein, we developed a fluorinated ferrous chelate-based sensitive probe, 1,7-DO2A-Fe(II)-F18 (Probe 1), for 19F magnetic resonance imaging (MRI). This highly fluorinated probe (containing 18 chemically equivalent 19F atoms with a fluorine content at 35 wt %) achieves a 15-time enhancement in signal intensity compared with the fluorine-containing ligand alone due to the appropriately regulated 19F relaxation times by the ferrous ion, which significantly increases imaging sensitivity and reduces acquisition time. Owing to its high aqueous solubility, biostability, and biocompatibility, this probe could be rapidly cleared by kidneys, which provides a means for monitoring renal dysfunction via 19F MRI. With this probe, we accomplish in vivo imaging of the impaired renal dysfunction caused by various kidney diseases including acute kidney injury, unilateral ureteral obstruction, and renal fibrosis at different stages. Our study illustrates the promising potential of Probe 1 for in vivo real-time visualization of kidney dysfunction, which is beneficial for the study, diagnosis, and even stratification of different kidney diseases. Furthermore, the design strategy of our probe is inspiring for the development of more high-performance 19F MRI probes for monitoring various biological processes.
Assuntos
Halogenação , Animais , Camundongos , Sondas Moleculares/química , Rim/diagnóstico por imagem , Rim/patologia , Complexos de Coordenação/química , Complexos de Coordenação/síntese química , Compostos Ferrosos/química , Imageamento por Ressonância Magnética , Nefropatias/diagnóstico por imagem , Imagem por Ressonância Magnética de Flúor-19/métodos , Flúor/químicaRESUMO
The incorporation of fluorine atoms into a biomacromolecule provides a background-free and environmentally sensitive reporter of structure, conformation and interactions using 19Fâ NMR. There are several methods to introduce the 19F reporter - either by synthetic incorporation via solid phase peptide synthesis; by suppressing the incorporation or biosynthesis of a natural amino acid and supplementing the growth media with a fluorinated counterpart during protein expression; and by genetic code expansion to add new amino acids to the amino acid alphabet. This review aims to discuss progress in the field of introducing fluorinated handles into biomolecules for NMR studies by post-translational bioconjugation or 'fluorine-tagging'. We will discuss the range of chemical tagging 'warheads' that have been used, explore the applications of fluorine tags, discuss ways to enhance reporter sensitivity and how the signal to noise ratios can be boosted. Finally, we consider some key challenges of the field and offer some ideas for future directions.
Assuntos
Flúor , Halogenação , Ressonância Magnética Nuclear Biomolecular , Conformação Proteica , Proteínas , Flúor/química , Proteínas/química , Proteínas/metabolismo , HumanosRESUMO
Fluorinated non-natural amino acids are attracting considerable research interest, especially in the biomedical field and in materials science, thanks to their ability to self-assemble into peculiar supramolecular structures. The conformational changes induced by the presence of fluorine atoms obviously affect their functions, as well as the biological activity of the deriving peptides and proteins. Here, we will briefly describe the main effects of fluorination on the aggregation behavior of such building blocks, focusing in particular on their improved tendency to form fibrils, and gels therefrom. Our aim is to underline the promising potential of fluorination as a tool to affect the self-assembly features of amino acids, both when used alone and when inserted into polypeptide sequences. The ability of fluorine to influence physical, chemical, and structural properties of these substrates offers the possibility to engineer bioinspired materials with specific and tunable functions.
Assuntos
Aminoácidos , Halogenação , Peptídeos , Aminoácidos/química , Peptídeos/química , Flúor/químicaRESUMO
Fluorine, the tiny robust atom, with its unique features has captured the attention of scientists in recent times, especially in drug discovery with its integration in small molecules, peptides, and proteins. However, studies to understand the 'fluorine effects' on the conformation of molecules that follow 'beyond the rule of 5' are in the infancy yet significant in molecular design and function. For the first time, using short hybrid peptide sequence as an appropriate model, we examined the substitution effect (size, stereoelectronic effect, and hydrogen bonding) using X-ray diffraction, 2D-NMR, and CD studies. The comparative study on their folding patterns with hydrogen-substituted analogs can provide valuable insights into fluorinated substrates' design.
Assuntos
Flúor , Dobramento de Proteína , Flúor/química , Peptídeos/química , Conformação Molecular , Sequência de Aminoácidos , Ligação de HidrogênioRESUMO
Nuclear magnetic resonance (NMR) spectroscopy NMR is a well-established technique for probing protein structure, dynamics and conformational changes. Taking advantage of the high signal sensitivity and broad chemical shift range of 19F nuclei, 19F NMR has been applied to investigate protein function at atomic resolution. In this report, we extend the unnatural amino acid site-specific incorporation into V. natriegens, an alternate protein expression system. The unnatural amino acid L-4-trifluoromethylphenylalanine (tfmF) was site-specifically introduced into the mitogen-activated protein kinase MEKK3 in V. natriegens using genetically encoded technology, which will be an extensive method for in-cell protein structure and dynamic investigation.
Assuntos
Ressonância Magnética Nuclear Biomolecular , Fenilalanina/química , Fenilalanina/metabolismo , Fenilalanina/genética , Flúor/química , Aminoácidos/química , Aminoácidos/genética , Aminoácidos/metabolismoRESUMO
The design of imaging agents with a high fluorine content is necessary for overcoming the challenges of low sensitivity in 19F magnetic resonance imaging (MRI)-based molecular imaging. Chemically self-assembled nanorings (CSANs) provide a strategy to increase the fluorine content through multivalent display. We previously reported an 19F NMR-based imaging tracer, in which case a CSAN-compatible epidermal growth factor receptor (EGFR)-targeting protein E1-dimeric dihydrofolate (E1-DD) was bioconjugated to a highly fluorinated peptide. Despite good 19F NMR performance in aqueous solutions, a limited signal was observed in cell-based 19F NMR using this monomeric construct, motivating further design. Here, we design several new E1-DD proteins bioconjugated to peptides of different fluorine contents. Flow cytometry analysis was used to assess the effect of variable fluorinated peptide sequences on the cellular binding characteristics. Structure-optimized protein, RTC-3, displayed an optimal spectral performance with high affinity and specificity for EGFR-overexpressing cells. To further improve the fluorine content, we next engineered monomeric RTC-3 into CSAN, η-RTC-3. With an approximate eightfold increase in the fluorine content, multivalent η-RTC-3 maintained high cellular specificity and optimal 19F NMR spectral behavior. Importantly, the first cell-based 19F NMR spectra of η-RTC-3 were obtained bound to EGFR-expressing A431 cells, showing a significant amplification in the signal. This new design illustrated the potential of multivalent fluorinated CSANs for future 19F MRI molecular imaging applications.
Assuntos
Flúor , Imageamento por Ressonância Magnética , Flúor/química , Espectroscopia de Ressonância Magnética , Proteínas , Peptídeos , Receptores ErbB/metabolismoRESUMO
Controlling the enantiomeric purity of chiral drugs is of paramount importance in pharmaceutical chemistry. Isotropic 1H NMR spectroscopy involving chiral agents is a widely used method for discriminating enantiomers and quantifying their relative proportions. However, the relatively weak spectral separation of enantiomers (1H Δδiso(R, S)) in frequency units at low and moderate magnetic fields, as well as the lack of versatility of a majority of those agents with respect to different chemical functions, may limit the general use of this approach. In this article, we investigate the analytical potential of 19F NMR in anisotropic chiral media for the enantiomeric analysis of fluorinated active pharmaceutical ingredients (API) via two residual anisotropic NMR interactions: the chemical shift anisotropy (19F-RCSA) and dipolar coupling ((19F-19F)-RDC). Lyotropic chiral liquid crystals (CLC) based on poly-γ-benzyl-L-glutamate (PBLG) show an interesting versatility and adaptability to enantiodiscrimination as illustrated for two chiral drugs, Flurbiprofen® (FLU) and Efavirenz® (EFA), which have very different chemical functions. The approach has been tested on a routine 300 MHz NMR spectrometer equipped with a standard probe (5 mm BBFO probe) in a high-throughput context (i.e., ≈10 s of NMR experiments) while the performance for enantiomeric excess (ee) measurement is evaluated in terms of trueness and precision. The limits of detection (LOD) determined were 0.17 and 0.16 µmol ml-1 for FLU and EFA, respectively, allow working in dilute conditions even with such a short experimental duration. The enantiodiscrimination capabilities are also discussed with respect to experimental features such as CLC composition and temperature.
Assuntos
Flúor , Espectroscopia de Ressonância Magnética , Estereoisomerismo , Espectroscopia de Ressonância Magnética/métodos , Anisotropia , Preparações Farmacêuticas/química , Preparações Farmacêuticas/análise , Flúor/química , Halogenação , Flurbiprofeno/química , Flurbiprofeno/análise , Cristais Líquidos/química , Princípios AtivosRESUMO
19F parashift probes with paramagnetically shifted reporter nuclei provide attractive platforms to develop molecular imaging probes. These probes enable ratiometric detection of molecular disease markers using a direct detection technique. Here, we describe a series of trivalent lanthanide (Ln(III)) complexes that are structural analogues of the clinically approved MR contrast agent (CA) ProHance to obtain LnL 19F parashift probes. We evaluated trans-gadolinium paramagnetic lanthanides compared to diamagnetic YL for 19F chemical shift and relaxation rate enhancement. The paramagnetic contribution to chemical shift (δPCS) for paramagnetic LnL exhibited either shifts to lower frequency (δPCS < 0 for TbL, DyL, and HoL) or shifts to higher frequency (δPCS > 0 for ErL, TmL, and YbL) compared to YL 19F spectroscopic signal. Zero-echo time pulse sequences achieved 56-fold sensitivity enhancement for DyL over YL, while developing probe-specific pulse sequences with fast delay times and acquisition times achieved 0.6-fold enhancement in limit of detection for DyL. DyL provides an attractive platform to develop 19F parashift probes for ratiometric detection of enzymatic activity.
Assuntos
Elementos da Série dos Lantanídeos , Elementos da Série dos Lantanídeos/química , Estrutura Molecular , Complexos de Coordenação/química , Complexos de Coordenação/síntese química , Imageamento por Ressonância Magnética , Meios de Contraste/química , Flúor/química , HumanosRESUMO
Substitution of two fluorine atoms of the tetrafluoroterephthalonitrile (TFTN) ring (ortho to each other) by amine nucleophiles through SNAr chemistry is achievable. However, tri- and tetra-substitution towards multi-substituted single benzene fluorophores (SBFs) is harder due to increased electron richness of the TFTN moiety. Tertiary amine donors promote the molecule towards such multi-substitution guided by the steric obstruction to intramolecular charge transfer to the TFTN ring. Contrarily, secondary amine substituents with better lone pair donation to the TFTN ring cannot induce the SNAr pathway and instead promote hydrolysis of the nitrile groups of the TFTN moiety. Theoretical investigations have helped unearth the reasons for this observed difference in chemical reactivities and also explain the differences in the emission spectra. Finally, the success of the synthetic method towards multi-substitution is showcased through creation of a highly lipophilic SBF bearing an octyl unit and demonstrating its utility in in vitro cellular imaging.