11 results on '"Camilo Rojas"'
Search Results
2. Thieno[2,3-d]pyrimidine-Based Positive Allosteric Modulators of Human Mas-Related G Protein-Coupled Receptor X1 (MRGPRX1)
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Ilyas Berhane, Niyada Hin, Ajit G. Thomas, Qian Huang, Chi Zhang, Vijayabhaskar Veeravalli, Ying Wu, Justin Ng, Jesse Alt, Camilo Rojas, Hiroe Hihara, Mika Aoki, Kyoko Yoshizawa, Tomoki Nishioka, Shuichi Suzuki, Shao-Qiu He, Qi Peng, Yun Guan, Xinzhong Dong, Srinivasa N. Raja, Barbara S. Slusher, Rana Rais, and Takashi Tsukamoto
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Male ,Proton Magnetic Resonance Spectroscopy ,Article ,Mass Spectrometry ,Receptors, G-Protein-Coupled ,Mice ,HEK293 Cells ,Pyrimidines ,Allosteric Regulation ,Drug Discovery ,Animals ,Humans ,Molecular Medicine ,Carbon-13 Magnetic Resonance Spectroscopy ,Chromatography, Liquid - Abstract
Mas-related G protein-coupled receptor X1 (MRGPRX1) is a human sensory neuron-specific receptor and potential target for the treatment of pain. Positive allosteric modulators (PAMs) of MRGPRX1 have the potential to preferentially activate the receptors at the central terminals of primary sensory neurons and minimize itch side effects caused by peripheral activation. Using a high-throughput screening (HTS) hit, a series of thieno[2,3-d]pyrimidine-based molecules were synthesized and evaluated as human MRGPRX1 PAMs in HEK293 cells stably transfected with human MrgprX1 gene. An iterative process to improve potency and metabolic stability led to the discovery of orally available 6-(tert-butyl)-5-(3,4-dichlorophenyl)-4-(2-(trifluoromethoxy)phenoxy)thieno[2,3-d]pyrimidine (1t), which can be distributed to the spinal cord, the presumed site of action, following oral administration. In a neuropathic pain model induced by sciatic nerve chronic constriction injury (CCI), compound 1t (100 mg/kg, po) reduced behavioral heat hypersensitivity in humanized MRGPRX1 mice, demonstrating the therapeutic potential of MRGPRX1 PAMs in treating neuropathic pain.
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- 2022
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3. High Throughput Screening Cascade To Identify Human Aspartate N-Acetyltransferase (ANAT) Inhibitors for Canavan Disease
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Camilo Rojas, Ondřej Nešuta, Anna Neužilová, Niyada Hin, Ajit G. Thomas, Huijun Wei, Takashi Tsukamoto, Barbara S. Slusher, Jesse Alt, and Shunyou Long
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chemistry.chemical_classification ,Mutation ,Physiology ,Cognitive Neuroscience ,High-throughput screening ,Acetyl-CoA ,Aspartate N-acetyltransferase ,Cell Biology ,General Medicine ,medicine.disease ,medicine.disease_cause ,Biochemistry ,Molecular biology ,Canavan disease ,law.invention ,chemistry.chemical_compound ,Enzyme ,chemistry ,Acetylation ,law ,medicine ,Recombinant DNA - Abstract
Canavan disease (CD) is a progressive, fatal neurological disorder that begins in infancy resulting from a mutation in aspartoacyclase (ASPA), an enzyme that catalyzes the deacetylation of N-acetyl aspartate (NAA) into acetate and aspartate. Increased NAA levels in the brains of affected children are one of the hallmarks of CD. Interestingly, genetic deletion of N-acetyltransferase-8-like (NAT8L), which encodes aspartate N-aceyltransferase (ANAT), an enzyme responsible for the synthesis of NAA from l-aspartate and acetyl-CoA, leads to normalization of NAA levels and improvement of symptoms in several genetically engineered mouse models of CD. Therefore, pharmacological inhibition of ANAT presents a promising therapeutic strategy for treating CD. Currently, however, there are no clinically viable ANAT inhibitors. Herein we describe the development of fluorescence-based high throughput screening (HTS) and radioactive-based orthogonal assays using recombinant human ANAT expressed in E. coli. In the fluorescence-based assay, ANAT activity was linear with respect to time of incubation up to 30 min and protein concentration up to 97.5 ng/μL with Km values for l-aspartate and acetyl-CoA of 237 μM and 11 μM, respectively. Using this optimized assay, we conducted a pilot screening of a 10 000-compound library. Hits from the fluorescence-based assay were subjected to an orthogonal radioactive-based assay using L-[U-14C] aspartate as a substrate. Two compounds were confirmed to have dose-dependent inhibition in both assays. Inhibitory kinetics studies of the most potent compound revealed an uncompetitive inhibitory mechanism with respect to l-aspartate and a noncompetitive inhibitory mechanism against acetyl-CoA. The screening cascade developed herein will enable large-scale compound library screening to identify novel ANAT inhibitors as leads for further medicinal chemistry optimization.
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- 2021
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4. Novel Human Neutral Sphingomyelinase 2 Inhibitors as Potential Therapeutics for Alzheimer’s Disease
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Kristen R. Hollinger, Jenny Lam, Camilo Rojas, Radim Nencka, Vijayabhaskar Veeravalli, Hubert Hřebabecký, Ajit G. Thomas, Michal Šála, Barbara S. Slusher, Rana Rais, Eliška Procházková, Ondřej Nešuta, Martin Kögler, Ranjeet Prasad Dash, Lyndah Lovell, Amanda Donoghue, and Carolyn Tallon
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Male ,Ceramide ,Mice, Transgenic ,Pharmacology ,Exosomes ,01 natural sciences ,Exosome ,Article ,Mice ,Structure-Activity Relationship ,03 medical and health sciences ,chemistry.chemical_compound ,Alzheimer Disease ,In vivo ,Drug Discovery ,Animals ,Humans ,Structure–activity relationship ,Enzyme Inhibitors ,030304 developmental biology ,0303 health sciences ,Chemistry ,Phosphorylcholine ,Body Weight ,Brain ,Microvesicles ,0104 chemical sciences ,Bioavailability ,Pyridazines ,Disease Models, Animal ,010404 medicinal & biomolecular chemistry ,Sphingomyelin Phosphodiesterase ,Molecular Medicine ,Female ,Sphingomyelin - Abstract
Neutral sphingomyelinase 2 (nSMase2) catalyzes the cleavage of sphingomyelin to phosphorylcholine and ceramide, an essential step in the formation and release of exosomes from cells that is critical for intracellular communication. Chronic increase of brain nSMase2 activity and related exosome release have been implicated in various pathological processes, including the progression of Alzheimer’s disease (AD), making nSMase2 a viable therapeutic target. Recently, we identified phenyl (R)-(1-(3-(3,4-dimethoxyphenyl)-2,6-dimethylimidazo[1,2-b]pyridazin-8-yl)-pyrrolidin-3-yl)carbamate 1 (PDDC), the first nSMase2 inhibitor that possesses both favorable pharmacodynamics and pharmacokinetic (PK) parameters, including substantial oral bioavailability, brain penetration, and significant inhibition of exosome release from the brain in vivo. Herein we demonstrate the efficacy of 1 (PDDC) in a mouse model of AD and detail extensive structure–activity relationship (SAR) studies with 70 analogues, unveiling several that exert similar or higher activity against nSMase2 with favorable pharmacokinetic properties.
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- 2020
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5. Discovery of 6-Diazo-5-oxo-<scp>l</scp>-norleucine (DON) Prodrugs with Enhanced CSF Delivery in Monkeys: A Potential Treatment for Glioblastoma
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Judson Englert, Kelly A. Metcalf Pate, Rana Rais, Jessica Tan, Lukáš Tenora, Camilo Rojas, Lenka Monincová, Robert J. Adams, Andrej Jančařík, Anne Le, Pavel Majer, Dana Ferraris, Jesse Alt, Michael T. Nedelcovych, Jonathan D. Powell, Amira Elgogary, and Barbara S. Slusher
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0301 basic medicine ,Antimetabolites, Antineoplastic ,Glutamine ,Diazooxonorleucine ,Mice, Nude ,Pharmacology ,Mice ,03 medical and health sciences ,chemistry.chemical_compound ,Therapeutic index ,Drug Discovery ,Animals ,Humans ,Moiety ,Prodrugs ,Brain Neoplasms ,Chemistry ,Antagonist ,Haplorhini ,Metabolism ,Prodrug ,6-Diazo-5-oxo-L-norleucine ,030104 developmental biology ,Toxicity ,Molecular Medicine ,Female ,Glioblastoma - Abstract
The glutamine antagonist 6-diazo-5-oxo-l-norleucine (DON, 1) has shown robust anticancer efficacy in preclinical and clinical studies, but its development was halted due to marked systemic toxicities. Herein we demonstrate that DON inhibits glutamine metabolism and provides antitumor efficacy in a murine model of glioblastoma, although toxicity was observed. To enhance DON's therapeutic index, we utilized a prodrug strategy to increase its brain delivery and limit systemic exposure. Unexpectedly, simple alkyl ester-based prodrugs were ineffective due to chemical instability cyclizing to form a unique diazo-imine. However, masking both DON's amine and carboxylate functionalities imparted sufficient chemical stability for biological testing. While these dual moiety prodrugs exhibited rapid metabolism in mouse plasma, several provided excellent stability in monkey and human plasma. The most stable compound (5c, methyl-POM-DON-isopropyl-ester) was evaluated in monkeys, where it achieved 10-fold enhanced cerebrospinal fluid to plasma ratio versus DON. This strategy may provide a path to DON utilization in glioblastoma multiforme patients.
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- 2016
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6. Unprecedented Binding Mode of Hydroxamate-Based Inhibitors of Glutamate Carboxypeptidase II: Structural Characterization and Biological Activity
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Rana Rais, Krystyna M. Wozniak, Niyada Hin, Zora Novakova, Takashi Tsukamoto, Jan Vávra, Camilo Rojas, Barbara S. Slusher, Andrej Jancarik, Jiri Pavlicek, Ying Wu, Pavel Majer, Dana Ferraris, Cyril Barinka, Barbora Havlinova, and Jakub Ptacek
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Glutamate Carboxypeptidase II ,Models, Molecular ,0301 basic medicine ,Stereochemistry ,Hydroxamic Acids ,Structure-Activity Relationship ,03 medical and health sciences ,chemistry.chemical_compound ,Glutamatergic ,0302 clinical medicine ,Drug Discovery ,Hydrolase ,Glutamate carboxypeptidase II ,Humans ,Moiety ,Structure–activity relationship ,Enzyme Inhibitors ,Hydroxamic acid ,Dose-Response Relationship, Drug ,Molecular Structure ,Chemistry ,Biological activity ,030104 developmental biology ,Biochemistry ,Antigens, Surface ,Molecular Medicine ,030217 neurology & neurosurgery ,In vivo pharmacokinetics - Abstract
Inhibition of glutamate carboxypeptidase II (GCPII) is effective in preclinical models of neurological disorders associated with excessive activation of glutamatergic systems. Here we report synthesis, structural characterization, and biological activity of new hydroxamic acid-based inhibitors with nanomolar affinity for human GCPII. Crystal structures of GCPII/hydroxamate complexes revealed an unprecedented binding mode in which the putative P1' glutarate occupies the spacious entrance funnel rather than the conserved glutamate-binding S1' pocket. This unique binding mode provides a mechanistic explanation for the structure-activity relationship data, most notably the lack of enantiospecificity and the tolerance for bulky/hydrophobic functions as substituents of a canonical glutarate moiety. The in vivo pharmacokinetics profile of one of the inhibitors will be presented along with analgesic efficacy data from the rat chronic constrictive injury model of neuropathic pain.
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- 2016
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7. Design, Synthesis, and Pharmacological Evaluation of Fluorinated Tetrahydrouridine Derivatives as Inhibitors of Cytidine Deaminase
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Edgar Schuck, Christopher Rowbottom, Camilo Rojas, Greg Delahanty, Barbara Slusher, Dana Ferraris, Bipin Mistry, Jesse Alt, Sanjeev Redkar, Bridget Duvall, Kristen Sanders, Takashi Tsukamoto, and Kuan-Chun Huang
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Models, Molecular ,Molecular Conformation ,Biological Availability ,Decitabine ,Pharmacology ,Structure-Activity Relationship ,Drug Stability ,Pharmacokinetics ,Cytidine Deaminase ,Drug Discovery ,Tetrahydrouridine ,medicine ,Animals ,Structure–activity relationship ,Enzyme Inhibitors ,Gastric Juice ,Chemistry ,Excitatory Postsynaptic Potentials ,Fluorine ,Cytidine deaminase ,Metabolism ,Plasma levels ,Macaca mulatta ,Design synthesis ,Biochemistry ,Drug Design ,Azacitidine ,Molecular Medicine ,medicine.drug - Abstract
Several 2'-fluorinated tetrahydrouridine derivatives were synthesized as inhibitors of cytidine deaminase (CDA). (4R)-2'-Deoxy-2',2'-difluoro-3,4,5,6-tetrahydrouridine (7a) showed enhanced acid stability over tetrahydrouridine (THU) 5 at its N-glycosyl bond. As a result, compound 7a showed an improved oral pharmacokinetic profile with a higher and more reproducible plasma exposure in rhesus monkeys compared to 5. Co-administration of 7a with decitabine, a CDA substrate, boosted the plasma levels of decitabine in rhesus monkeys. These results demonstrate that compound 7a can serve as an acid-stable alternative to 5 as a pharmacoenhancer of drugs subject to CDA-mediated metabolism.
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- 2014
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8. Synthesis and SAR of 1-Hydroxy-1H-benzo[d]imidazol-2(3H)-ones as Inhibitors of <scp>d</scp>-Amino Acid Oxidase
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Jesse Alt, Bridget Duvall, Barbara S. Slusher, Dana Ferraris, Camilo Rojas, Niyada Hin, Kenji Hashimoto, Takashi Tsukamoto, Rana Rais, Ajit G. Thomas, and James F. Berry
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Oxidase test ,Stereochemistry ,Organic Chemistry ,Glucuronidation ,D-amino acid oxidase ,Pharmacology ,Biochemistry ,chemistry.chemical_compound ,Inhibitory potency ,Pharmacokinetics ,chemistry ,Drug Discovery ,Ic50 values ,Benzene - Abstract
A series of 1-hydroxy-1H-benzo[d]imidazol-2(3H)-ones were synthesized and evaluated for their ability to inhibit human and porcine forms of d-amino acid oxidase (DAAO). The inhibitory potency is largely dependent on the size and position of substituents on the benzene ring with IC50 values of the compounds ranging from 70 nM to greater than 100 μM. Structure–activity relationships of this new class of DAAO inhibitors will be presented in detail along with comparisons to previously published SAR data from other classes of DAAO inhibitors. Two of these compounds were given to mice orally together with d-serine to assess their effects on plasma d-serine pharmacokinetics.
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- 2012
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9. Synthesis and Biological Evaluation of Thiol-Based Inhibitors of Glutamate Carboxypeptidase II: Discovery of an Orally Active GCP II Inhibitor
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Krystyna M. Wozniak, Jana Polakova, Takashi Tsukamoto, Brian Grella, Tharin Limsakun, Greg Delahanty, Paul F. Jackson, Kathryn Ann Shaffer, Maclin Keith M, Camilo Rojas, Barbara S. Slusher, Eric Wang, Weixing Li, Eric Burak, Pavel Majer, Dilrukshi Vitharana, Qun Liu, Yao Sen Ko, Anthony Zakrzewski, and Doris Stoermer
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Glutamate Carboxypeptidase II ,Male ,Hot Temperature ,Carboxylic acid ,Administration, Oral ,Biological Availability ,Pain ,Carboxypeptidases ,Constriction, Pathologic ,Chemical synthesis ,Glutarates ,Rats, Sprague-Dawley ,Structure-Activity Relationship ,chemistry.chemical_compound ,Drug Discovery ,Glutamate carboxypeptidase II ,Animals ,Sulfhydryl Compounds ,Enzyme Inhibitors ,IC50 ,chemistry.chemical_classification ,Analgesics ,biology ,Peripheral Nervous System Diseases ,Sciatic Nerve ,Phosphonate ,Rats ,Enzyme ,chemistry ,Biochemistry ,Hyperalgesia ,Enzyme inhibitor ,biology.protein ,Thiol ,Molecular Medicine - Abstract
A series of 2-(thioalkyl)pentanedioic acids were synthesized and evaluated as inhibitors of glutamate carboxypeptidase II (GCP II, EC 3.4.17.21). The inhibitory potency of these thiol-based compounds against GCP II was found to be dependent on the number of methylene units between the thiol group and pentanedioic acid. A comparison of the SAR of the thiol-based inhibitors to that of the phosphonate-based inhibitors provides insight into the role of each of the two zinc-binding groups in GCP II inhibition. The most potent thiol-based inhibitor, 2-(3-mercaptopropyl)pentanedioic acid (IC(50) = 90 nM), was found to be orally bioavailable in rats and exhibited efficacy in an animal model of neuropathic pain following oral administration.
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- 2003
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10. Synthesis and Biological Evaluation of <scp>d</scp>-Amino Acid Oxidase Inhibitors
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Takashi Tsukamoto, Yao Sen Ko, Dana Ferraris, Camilo Rojas, Bridget Duvall, Kenji Hashimoto, Pavel Majer, and Ajit G. Thomas
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D-Amino-Acid Oxidase ,Male ,Agonist ,Stereochemistry ,medicine.drug_class ,D-amino acid oxidase ,Administration, Oral ,Rats, Sprague-Dawley ,Structure-Activity Relationship ,Drug Discovery ,Serine ,medicine ,Animals ,chemistry.chemical_classification ,Benzoxazoles ,Oxidase test ,biology ,Brain ,Drug Synergism ,Biological activity ,Isoxazoles ,Rats ,Enzyme ,chemistry ,Biochemistry ,Enzyme inhibitor ,Glycine ,biology.protein ,Molecular Medicine ,NMDA receptor - Abstract
D-amino acid oxidase (DAAO) catalyzes the oxidation of D-amino acids including d-serine, a full agonist at the glycine site of the NMDA receptor. A series of benzo[ d]isoxazol-3-ol derivatives were synthesized and evaluated as DAAO inhibitors. Among them, 5-chloro-benzo[ d]isoxazol-3-ol (CBIO) potently inhibited DAAO with an IC50 in the submicromolar range. Oral administration of CBIO in conjunction with d-serine enhanced the plasma and brain levels of d-serine in rats compared to the oral administration of d-serine alone.
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- 2008
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11. Spectral observation of an acyl-enzyme intermediate of lipoprotein lipase
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Chris R. Lively, James T. McFarland, William G. Gustafson, Leslie O. Schulz, Camilo Rojas, and Huei Hsiang Wang
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Lipoprotein lipase ,Chemistry ,Stereochemistry ,Acylation ,Hydrolysis ,Glyceride ,Leaving group ,Substrate (chemistry) ,Hydrogen-Ion Concentration ,In Vitro Techniques ,Biochemistry ,Rats ,Catalysis ,Serine ,Kinetics ,Lipoprotein Lipase ,Adipose Tissue ,Animals ,Spectrophotometry, Ultraviolet ,lipids (amino acids, peptides, and proteins) ,Histidine - Abstract
There have been several studies indicating that hydrolysis reactions of fatty acid esters catalyzed by lipases proceed through an acyl-enzyme intermediate typical of serine proteases. In particular, one careful kinetic study with the physiologically important enzyme lipoprotein lipase (LPL) is consistent with rate-limiting deacylation of such an intermediate. To observe the spectrum of acyl-enzyme and study the mechanism of LPL-catalyzed hydrolysis of substrate, we have used a variety of furylacryloyl substrates including 1,2-dipalmitoyl-3-[(beta-2-furylacryloyl)triacyl]glyceride (DPFATG) to study the intermediates formed during the hydrolysis reaction catalyzed by the enzyme. After isolation and characterization of the molecular weight of adipose LPL, we determined its extinction coefficient at 280 nm to quantitate the formation of any acyl-enzyme intermediate formed during substrate hydrolysis. We observed an intermediate at low pH during the enzyme-catalyzed hydrolysis of (furylacryloyl)imidazole. This intermediate builds early in the reaction when a substantial amount of substrate has hydrolyzed but no product, furylacrylate, has been formed. The acyl-enzyme has a lambda max = 305 nm and a molar extinction coefficient of 22,600 M-1 cm-1; these parameters are similar to those for furylacryloyl esters including the serine ester. These data provide the first spectral evidence for a serine acyl-enzyme in lipase-catalyzed reactions. The LPL hydrolysis reaction is base catalyzed, exhibiting two pKa values; the more acidic of these is 6.5, consistent with base catalysis by histidine. The biphasic rates for substrate disappearance or product appearance and the absence of leaving group effect indicate that deacylation of intermediate is rate limiting.
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- 1989
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