ABSTRACT
BACKGROUND: Reactive microglia and recruited peripheral macrophages contribute to the pathogenesis of Alzheimer's dementia (AD). Monocytes, macrophages and microglia all express the marker colony-stimulating factor 1 receptor (CSF1R). 4-Cyano-N-(4-(4-methylpiperazin-1-yl)-2-(4-methylpiperidin-1-yl)phenyl)-1H-pyrrole-2-carboxamide (1) is a high-affinity antagonist for CSF1R. We report the radiosynthesis of both [3H]1 and [11C]1. The PET imaging properties of [11C]1 in mice and baboon were investigated. [3H]1 was studied in Bmax measurement in post-mortem autoradiography in the frontal cortex, inferior parietal cortex and hippocampus from donors diagnosed with AD and age-matched controls. In vitro binding affinity of 1 was measured commercially. Nor-methyl-1 precursor was radiolabeled with [11C]iodomethane or [3H]iodomethane to produce [11C]1 and [3H]1, respectively. Ex vivo brain biodistribution of [11C]1 was compared in normal mice versus lipopolysaccharide-administered (LPS) murine model of neuroinflammation. Dynamic PET imaging was performed in a healthy male Papio anubis baboon. Post-mortem autoradiography with [3H]1 was performed in frozen sections using a standard saturation binding technique. RESULTS: Compound 1 exhibits a high in vitro CSF1R binding affinity (0.59 nM). [11C]1 was synthesized with high yield. [3H]1 was synthesized similarly (commercially). Biodistribution of [11C]1 in healthy mice demonstrated moderate brain uptake. In LPS-treated mice the brain uptake of [11C]1 was ~ 50% specific for CSF1R. PET/CT [11C]1 study in baboon revealed low brain uptake (0.36 SUV) of [11C]1. Autoradiography with [3H]1 gave significantly elevated Bmax values in AD frontal cortex versus control (47.78 ± 26.80 fmol/mg vs. 12.80 ± 5.30 fmol/mg, respectively, P = 0.023) and elevated, but not significantly different binding in AD hippocampus grey matter and inferior parietal cortex (IPC) white matter. CONCLUSIONS: Compound 1 exhibits a high in vitro CSF1R binding affinity. [11C]1 specifically labels CSF1R in the mouse neuroinflammation, but lacks the ability to efficiently cross the blood-brain barrier in baboon PET. [3H]1 specifically labels CSF1R in post-mortem human brain. The binding of [3H]1 is significantly higher in the post-mortem frontal cortex of AD versus control subjects.
ABSTRACT
PURPOSE: Macrophages represent an essential means of sequestration and immune evasion for Mycobacterium tuberculosis. Pulmonary tuberculosis (TB) is characterized by dense collections of tissue-specific and recruited macrophages, both of which abundantly express CSF1R on their outer surface. 4-Cyano-N-(5-(1-(dimethylglycyl)piperidin-4-yl)-2',3',4',5'-tetrahydro-[1,1'-biphenyl]-2-yl)-1H-imidazole-2-carboxamide (JNJ-28312141) is a reported high affinity, CSF1R-selective antagonist. We report the radiosynthesis of 4-cyano-N-(5-(1-(N-methyl-N-([11C]methyl)glycyl)piperidin-4-yl)-2',3',4',5'-tetrahydro-[1,1'-biphenyl]-2-yl)-1H-imidazole-2-carboxamide ([11C]JNJ-28312141) and non-invasive detection of granulomatous and diffuse lesions in a mouse model of TB using positron emission tomography (PET). METHODS: Nor-methyl-JNJ-28312141 precursor was radiolabeled with [11C]iodomethane to produce [11C]JNJ-28312141. PET/CT imaging was performed in the C3HeB/FeJ murine model of chronic pulmonary TB to co-localize radiotracer uptake with granulomatous lesions observed on CT. Additionally, CSF1R, Iba1 fluorescence immunohistochemistry was performed to co-localize CSF1R target with reactive macrophages in infected and healthy mice. RESULTS: Radiosynthesis of [11C]JNJ-28312141 averaged a non-decay-corrected yield of 18.7 ± 2.1%, radiochemical purity of 99%, and specific activity averaging 658 ± 141 GBq/µmol at the end-of-synthesis. PET/CT imaging in healthy mice showed hepatobiliary [13.39-25.34% ID/g, percentage of injected dose per gram of tissue (ID/g)] and kidney uptake (12.35% ID/g) at 40-50 min post-injection. Infected mice showed focal pulmonary lesion uptake (5.58-12.49% ID/g), hepatobiliary uptake (15.30-40.50% ID/g), cervical node uptake, and renal uptake (11.66-29.33% ID/g). The ratio of infected lesioned lung/healthy lung uptake is 5.91:1, while the ratio of lesion uptake to adjacent infected radiolucent lung is 2.8:1. Pre-administration of 1 mg/kg of unlabeled JNJ-28312141 with [11C]JNJ-28312141 in infected animals resulted in substantial blockade. Fluorescence microscopy of infected and uninfected whole lung sections exclusively co-localized CSF1R staining with abundant Iba1 + macrophages. Healthy lung exhibited no CSF1R staining and very few Iba1 + macrophages. CONCLUSION: [11C]JNJ-28312141 binds specifically to CSF1R + macrophages and delineates granulomatous foci of disease in a murine model of pulmonary TB.
Subject(s)
Positron Emission Tomography Computed Tomography , Tuberculosis , Animals , Biphenyl Compounds , Disease Models, Animal , Imidazoles , Mice , Positron Emission Tomography Computed Tomography/methods , Positron-Emission Tomography/methods , Receptors, Granulocyte-Macrophage Colony-Stimulating Factor , Tomography, X-Ray Computed , Tuberculosis/diagnostic imagingABSTRACT
Fibroblast activation protein (FAP) has become a favored target for imaging and therapy of malignancy. We have synthesized and characterized two new (4-quinolinoyl)-glycyl-2-cyanopyrrolidine-based small molecules for imaging of FAP, QCP01 and [111In]QCP02, using optical and single-photon computed tomography/CT, respectively. Binding of imaging agents to FAP was assessed in six human cancer cell lines of different cancer types: glioblastoma (U87), melanoma (SKMEL24), prostate (PC3), NSCLC (NCIH2228), colorectal carcinoma (HCT116), and lung squamous cell carcinoma (NCIH226). Mouse xenograft models were developed with FAP-positive U87 and FAP-negative PC3 cells to test pharmacokinetics and binding specificity in vivo. QCP01 and [111In]QCP02 demonstrated nanomolar inhibition of FAP at Ki values of 1.26 and 16.20 nM, respectively. Both were selective for FAP over DPP-IV, a related serine protease. Both enabled imaging of FAP-expressing tumors specifically in vivo. [111In]QCP02 showed high uptake at 18.2 percent injected dose per gram in the U87 tumor at 30 min post-administration.
Subject(s)
Fibroblasts/metabolism , Fluorescent Dyes/chemistry , Gelatinases/metabolism , Membrane Proteins/metabolism , Neoplasms/metabolism , Serine Endopeptidases/metabolism , Animals , Cell Line, Tumor , Endopeptidases , Fluorescent Dyes/chemical synthesis , Fluorometry , Heterografts/metabolism , Humans , Male , Mice, Inbred NOD , Mice, SCID , Pyrrolidines/chemical synthesis , Pyrrolidines/chemistry , Quinolines/chemical synthesis , Quinolines/chemistryABSTRACT
Chiral bisoxazoline ligands containing naphthyridine, pyridazine, pyrazole, and phenol bridging units were prepared and shown to form bimetallic complexes with various metal salts. X-ray crystal structures of bis-nickel naphthyridine-bridged, bis-zinc pyridazine-bridged, and bis-nickel as well as bis-palladium pyrazole-bridged complexes were obtained.
ABSTRACT
CBR hydroxamidines are small-molecule inhibitors of bacterial RNA polymerase (RNAP) discovered through high-throughput screening of synthetic-compound libraries. CBR pyrazoles are structurally related RNAP inhibitors discovered through scaffold hopping from CBR hydroxamidines. CBR hydroxamidines and pyrazoles selectively inhibit Gram-negative bacterial RNAP and exhibit selective antibacterial activity against Gram-negative bacteria. Here, we report crystal structures of the prototype CBR hydroxamidine, CBR703, and a CBR pyrazole in complex with E. coli RNAP holoenzyme. In addition, we define the full resistance determinant for CBR703, show that the binding site and resistance determinant for CBR703 do not overlap the binding sites and resistance determinants of other characterized RNAP inhibitors, show that CBR703 exhibits no or minimal cross-resistance with other characterized RNAP inhibitors, and show that co-administration of CBR703 with other RNAP inhibitors results in additive antibacterial activities. The results set the stage for structure-based optimization of CBR inhibitors as antibacterial drugs.
Subject(s)
Amidines/pharmacology , Anti-Bacterial Agents/pharmacology , DNA-Directed RNA Polymerases/antagonists & inhibitors , Escherichia coli Proteins/antagonists & inhibitors , Escherichia coli/drug effects , Hydroxylamines/pharmacology , Pyrazoles/pharmacology , Transcription, Genetic/drug effects , Amidines/chemistry , Amino Acid Sequence , Animals , Anti-Bacterial Agents/chemistry , Binding Sites , Chlorocebus aethiops , Crystallography, X-Ray , DNA-Directed RNA Polymerases/chemistry , DNA-Directed RNA Polymerases/genetics , Drug Synergism , Escherichia coli/enzymology , Escherichia coli/genetics , Escherichia coli Proteins/chemistry , Escherichia coli Proteins/genetics , HeLa Cells , Humans , Hydroxylamines/chemistry , Ligands , Models, Molecular , Molecular Sequence Data , Mycobacterium tuberculosis/drug effects , Mycobacterium tuberculosis/enzymology , Mycobacterium tuberculosis/genetics , Protein Binding , Protein Structure, Secondary , Protein Structure, Tertiary , Pyrazoles/chemistry , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Sequence Alignment , Vero CellsABSTRACT
Redox-neutral formation of C-P bonds in the α-position of amines was achieved via a process that features a combination of an oxidative α-C-H bond functionalization and a reductive N-alkylation. Benzoic acid functions as an efficient catalyst in this three-component reaction of cyclic secondary amines, aldehydes and phosphine oxides to provide rapid access to α-amino phosphine oxides not easily accessible by classic Kabachnik-Fields reactions.
Subject(s)
Aldehydes/chemistry , Amines/chemistry , Benzoic Acid/chemistry , Alkylation , Catalysis , Molecular Structure , Oxidation-Reduction , Phosphines/chemistrySubject(s)
Amines/chemistry , Carboxylic Acids/chemistry , Copper/chemistry , Catalysis , Isomerism , Microwaves , Oxidation-ReductionABSTRACT
α-Amino acids react with aldehydes in the presence of a cyanide source to form α-amino nitriles in what can be considered a decarboxylative variant of the classical Strecker reaction. This unprecedented transformation does not require the use of a metal catalyst and provides facile access to valuable α-amino nitriles that are inaccessible by traditional Strecker chemistry.
Subject(s)
Amino Acids/chemistry , Nitriles/chemical synthesis , Aldehydes/chemistry , Catalysis , Molecular Structure , Nitriles/chemistry , StereoisomerismABSTRACT
Indolines react with aromatic and heteroaromatic aldehydes to yield N-alkyl indoles in a benzoic acid catalyzed redox isomerization reaction. Azomethine ylides are intermediates in this process which was established by intramolecular [3 + 2] trapping experiments.