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1.
Bioconjug Chem ; 34(6): 988-993, 2023 06 21.
Artículo en Inglés | MEDLINE | ID: mdl-37216465

RESUMEN

Macrocycles occupy chemical space "beyond the rule of five". They bridge traditional bioactive small molecule drugs and macromolecules and have the potential to modulate challenging targets such as PPI or proteases. Here we report an on-DNA macrocyclization reaction utilizing intramolecular benzimidazole formation. A 129-million-member macrocyclic library composed of a privileged benzimidazole core, a dipeptide sequence (natural or non-natural), and linkers of varying length and flexibility was designed and synthesized.


Asunto(s)
Compuestos Macrocíclicos , Compuestos Macrocíclicos/química , Biblioteca de Genes , Ciclización , Bencimidazoles , ADN/química
2.
Bioconjug Chem ; 32(9): 1973-1978, 2021 09 15.
Artículo en Inglés | MEDLINE | ID: mdl-34424686

RESUMEN

Enzymatic catalysis is a highly attractive approach to the DNA encoded library technology (DEL) that has not been widely explored. In this paper, we report an l-threonine aldolase (l-TA)-catalyzed on-DNA aldol reaction to form ß-hydroxy-α-amino acids, and its diastereoselectivity determination. l-TAs from three species show good on-DNA aldehyde scope and complementary stereoselectivity. The formed aldol product can be further diversified via various reactions, which demonstrates the utility of this reaction in DEL.


Asunto(s)
Glicina Hidroximetiltransferasa , Aldehídos , Catálisis , Especificidad por Sustrato
3.
Anal Chem ; 88(10): 5498-506, 2016 05 17.
Artículo en Inglés | MEDLINE | ID: mdl-27119259

RESUMEN

Selection of target-binding ligands from DNA-encoded libraries of small molecules (DELSMs) is a rapidly developing approach in drug-lead discovery. Methods of kinetic capillary electrophoresis (KCE) may facilitate highly efficient homogeneous selection of ligands from DELSMs. However, KCE methods require accurate prediction of electrophoretic mobilities of protein-ligand complexes. Such prediction, in turn, requires a theory that would be applicable to DNA tags of different structures used in different DELSMs. Here we present such a theory. It utilizes a model of a globular protein connected, through a single point (small molecule), to a linear DNA tag containing a combination of alternating double-stranded and single-stranded DNA (dsDNA and ssDNA) regions of varying lengths. The theory links the unknown electrophoretic mobility of protein-DNA complex with experimentally determined electrophoretic mobilities of the protein and DNA. Mobility prediction was initially tested by using a protein interacting with 18 ligands of various combinations of dsDNA and ssDNA regions, which mimicked different DELSMs. For all studied ligands, deviation of the predicted mobility from the experimentally determined value was within 11%. Finally, the prediction was tested for two proteins and two ligands with a DNA tag identical to those of DELSM manufactured by GlaxoSmithKline. Deviation between the predicted and experimentally determined mobilities did not exceed 5%. These results confirm the accuracy and robustness of our model, which makes KCE methods one step closer to their practical use in selection of drug leads, and diagnostic probes from DELSMs.


Asunto(s)
ADN/química , Electroforesis Capilar , Proteínas/química , Bibliotecas de Moléculas Pequeñas/química , Biotina/química , Biotina/metabolismo , Anhidrasa Carbónica II/química , Anhidrasa Carbónica II/metabolismo , ADN de Cadena Simple/química , Humanos , Ligandos , Modelos Teóricos , Proteínas/metabolismo , Bibliotecas de Moléculas Pequeñas/metabolismo
4.
Anal Chem ; 87(4): 2474-9, 2015 Feb 17.
Artículo en Inglés | MEDLINE | ID: mdl-25582319

RESUMEN

Selection of protein binders from highly diverse combinatorial libraries of DNA-encoded small molecules is a highly promising approach for discovery of small-molecule drug leads. Methods of kinetic capillary electrophoresis provide the high efficiency of partitioning required for such selection but require the knowledge of electrophoretic mobility of the protein-ligand complex. Here we present a theoretical approach for an accurate estimate of the electrophoretic mobility of such complexes. The model is based on a theory of the thin double layer and corresponding expressions used for the mobilities of a rod-like short oligonucleotide and a sphere-like globular protein. The model uses empirical values of mobilities of free protein, free ligand, and electroosmotic flow. The model was tested with a streptavidin-dsDNA complex linked through biotin (small molecule). The deviation of the prediction from the experimental mobility did not exceed 4%, thus confirming that not only is the model adequate but it is also accurate. This model will facilitate reliable use of KCE methods for selection of drug leads from libraries of DNA-encoded small molecules.


Asunto(s)
ADN/análisis , ADN/química , Electroforesis Capilar/métodos , Estreptavidina/análisis , Estreptavidina/química , Biotina/química
5.
Drug Metab Dispos ; 41(4): 814-26, 2013 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-23355637

RESUMEN

The natural product fumagillin exhibits potent antiproliferative and antiangiogenic properties. The semisynthetic analog PPI-2458, [(3R,4S,5S,6R)-5-methoxy-4-[(2R,3R)-2-methyl-3-(3-methylbut-2-enyl)oxiran-2-yl]-1-oxaspiro[2.5]octan-6-yl] N-[(2R)-1-amino-3-methyl-1-oxobutan-2-yl]carbamate, demonstrates rapid inactivation of its molecular target, methionine aminopeptidase-2 (MetAP2), and good efficacy in several rodent models of cancer and inflammation with oral dosing despite low apparent oral bioavailability. To probe the basis of its in vivo efficacy, the metabolism of PPI-2458 was studied in detail. Reaction phenotyping identified CYP3A4/5 as the major source of metabolism in humans. Six metabolites were isolated from liver microsomes and characterized by mass spectrometry and nuclear resonance spectroscopy, and their structures were confirmed by chemical synthesis. The synthetic metabolites showed correlated inhibition of MetAP2 enzymatic activity and vascular endothelial cell growth. In an ex vivo experiment, MetAP2 inhibition in white blood cells, thymus, and lymph nodes in rats after single dosing with PPI-2458 and the isolated metabolites was found to correlate with the in vitro activity of the individual species. In a phase 1 clinical study, PPI-2458 was administered to patients with non-Hodgkin lymphoma. At 15 mg administered orally every other day, MetAP2 in whole blood was 80% inactivated for up to 48 hours, although the exposure of the parent compound was only ∼10% that of the summed cytochrome P450 metabolites. Taken together, the data confirm the participation of active metabolites in the in vivo efficacy of PPI-2458. The structures define a metabolic pathway for PPI-2458 that is distinct from that of TNP-470 ([(3R,4S,5S,6R)-5-methoxy-4-[(2R,3R)-2-methyl-3-(3-methylbut-2-enyl)oxiran-2-yl]-1-oxaspiro[2.5]octan-6-yl] N-(2-chloroacetyl)carbamate). The high level of MetAP2 inhibition achieved in vivo supports the value of fumagillin-derived therapeutics for angiogenic diseases.


Asunto(s)
Aminopeptidasas/antagonistas & inhibidores , Inhibidores de la Angiogénesis/farmacología , Inhibidores de la Angiogénesis/farmacocinética , Compuestos Epoxi/farmacocinética , Metaloendopeptidasas/antagonistas & inhibidores , Valina/análogos & derivados , Aminopeptidasas/sangre , Animales , Esquema de Medicación , Compuestos Epoxi/administración & dosificación , Compuestos Epoxi/análisis , Compuestos Epoxi/farmacología , Humanos , Leucocitos/efectos de los fármacos , Leucocitos/metabolismo , Ganglios Linfáticos/efectos de los fármacos , Ganglios Linfáticos/metabolismo , Linfoma no Hodgkin/sangre , Linfoma no Hodgkin/tratamiento farmacológico , Metaloendopeptidasas/sangre , Microsomas Hepáticos/química , Microsomas Hepáticos/metabolismo , Ratas , Relación Estructura-Actividad , Timo/efectos de los fármacos , Timo/metabolismo , Valina/administración & dosificación , Valina/análisis , Valina/farmacocinética , Valina/farmacología
6.
Nat Chem Biol ; 5(9): 647-54, 2009 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-19648931

RESUMEN

Biochemical combinatorial techniques such as phage display, RNA display and oligonucleotide aptamers have proven to be reliable methods for generation of ligands to protein targets. Adapting these techniques to small synthetic molecules has been a long-sought goal. We report the synthesis and interrogation of an 800-million-member DNA-encoded library in which small molecules are covalently attached to an encoding oligonucleotide. The library was assembled by a combination of chemical and enzymatic synthesis, and interrogated by affinity selection. We describe methods for the selection and deconvolution of the chemical display library, and the discovery of inhibitors for two enzymes: Aurora A kinase and p38 MAP kinase.


Asunto(s)
ADN/química , Diseño de Fármacos , Inhibidores de Proteínas Quinasas/síntesis química , Bibliotecas de Moléculas Pequeñas/síntesis química , Animales , Aurora Quinasas , Técnicas Químicas Combinatorias , ADN/genética , Modelos Moleculares , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/farmacología , Proteínas Quinasas p38 Activadas por Mitógenos/antagonistas & inhibidores
7.
ACS Med Chem Lett ; 7(4): 379-84, 2016 Apr 14.
Artículo en Inglés | MEDLINE | ID: mdl-27096045

RESUMEN

To identify BCATm inhibitors suitable for in vivo study, Encoded Library Technology (ELT) was used to affinity screen a 117 million member benzimidazole based DNA encoded library, which identified an inhibitor series with both biochemical and cellular activities. Subsequent SAR studies led to the discovery of a highly potent and selective compound, 1-(3-(5-bromothiophene-2-carboxamido)cyclohexyl)-N-methyl-2-(pyridin-2-yl)-1H-benzo[d]imidazole-5-carboxamide (8b) with much improved PK properties. X-ray structure revealed that 8b binds to the active site of BACTm in a unique mode via multiple H-bond and van der Waals interactions. After oral administration, 8b raised mouse blood levels of all three branched chain amino acids as a consequence of BCATm inhibition.

8.
ACS Med Chem Lett ; 6(8): 919-24, 2015 Aug 13.
Artículo en Inglés | MEDLINE | ID: mdl-26288694

RESUMEN

As a potential target for obesity, human BCATm was screened against more than 14 billion DNA encoded compounds of distinct scaffolds followed by off-DNA synthesis and activity confirmation. As a consequence, several series of BCATm inhibitors were discovered. One representative compound (R)-3-((1-(5-bromothiophene-2-carbonyl)pyrrolidin-3-yl)oxy)-N-methyl-2'-(methylsulfonamido)-[1,1'-biphenyl]-4-carboxamide (15e) from a novel compound library synthesized via on-DNA Suzuki-Miyaura cross-coupling showed BCATm inhibitory activity with IC50 = 2.0 µM. A protein crystal structure of 15e revealed that it binds to BCATm within the catalytic site adjacent to the PLP cofactor. The identification of this novel inhibitor series plus the establishment of a BCATm protein structure provided a good starting point for future structure-based discovery of BCATm inhibitors.

9.
J Med Chem ; 57(4): 1276-88, 2014 Feb 27.
Artículo en Inglés | MEDLINE | ID: mdl-24450589

RESUMEN

Tuberculosis (TB) is one of the world's oldest and deadliest diseases, killing a person every 20 s. InhA, the enoyl-ACP reductase from Mycobacterium tuberculosis, is the target of the frontline antitubercular drug isoniazid (INH). Compounds that directly target InhA and do not require activation by mycobacterial catalase peroxidase KatG are promising candidates for treating infections caused by INH resistant strains. The application of the encoded library technology (ELT) to the discovery of direct InhA inhibitors yielded compound 7 endowed with good enzymatic potency but with low antitubercular potency. This work reports the hit identification, the selected strategy for potency optimization, the structure-activity relationships of a hundred analogues synthesized, and the results of the in vivo efficacy studies performed with the lead compound 65.


Asunto(s)
Antituberculosos/farmacología , Proteínas Bacterianas/antagonistas & inhibidores , Descubrimiento de Drogas , Mycobacterium tuberculosis/efectos de los fármacos , Oxidorreductasas/antagonistas & inhibidores , Espectroscopía de Resonancia Magnética , Pruebas de Sensibilidad Microbiana , Mycobacterium tuberculosis/metabolismo , Espectrometría de Masa por Ionización de Electrospray
10.
ACS Med Chem Lett ; 4(4): 381-6, 2013 Apr 11.
Artículo en Inglés | MEDLINE | ID: mdl-24900682

RESUMEN

Semisynthetic analogues of fumagillin, 1, inhibit methionine aminopeptidase-2 (MetAP2) and have entered the clinic for the treatment of cancer. An optimized fumagillin analogue, 3 (PPI-2458), was found to be orally active, despite containing a spiroepoxide function that formed a covalent linkage to the target protein. In aqueous acid, 3 underwent ring-opening addition of water and HCl, leading to four products, 4-7, which were characterized in detail. The chlorohydrin, but not the diol, products inhibited MetAP2 under weakly basic conditions, suggesting reversion to epoxide as a step in the mechanism. In agreement, chlorohydrin 6 was shown to revert rapidly to 3 in rat plasma. In an ex vivo assay, rats treated with purified acid degradants demonstrated inhibition of MetAP2 that correlated with the biochemical activity of the compounds. Taken together, the results indicate that degradation of the parent compound was compensated by the formation of active equivalents leading to a pharmacologically useful level of MetAP2 inhibition.

11.
J Med Chem ; 55(16): 7061-79, 2012 Aug 23.
Artículo en Inglés | MEDLINE | ID: mdl-22891645

RESUMEN

The metalloprotease ADAMTS-5 is considered a potential target for the treatment of osteoarthritis. To identify selective inhibitors of ADAMTS-5, we employed encoded library technology (ELT), which enables affinity selection of small molecule binders from complex mixtures by DNA tagging. Selection of ADAMTS-5 against a four-billion member ELT library led to a novel inhibitor scaffold not containing a classical zinc-binding functionality. One exemplar, (R)-N-((1-(4-(but-3-en-1-ylamino)-6-(((2-(thiophen-2-yl)thiazol-4-yl)methyl)amino)-1,3,5-triazin-2-yl)pyrrolidin-2-yl)methyl)-4-propylbenzenesulfonamide (8), inhibited ADAMTS-5 with IC(50) = 30 nM, showing >50-fold selectivity against ADAMTS-4 and >1000-fold selectivity against ADAMTS-1, ADAMTS-13, MMP-13, and TACE. Extensive SAR studies showed that potency and physicochemical properties of the scaffold could be further improved. Furthermore, in a human osteoarthritis cartilage explant study, compounds 8 and 15f inhibited aggrecanase-mediated (374)ARGS neoepitope release from aggrecan and glycosaminoglycan in response to IL-1ß/OSM stimulation. This study provides the first small molecule evidence for the critical role of ADAMTS-5 in human cartilage degradation.


Asunto(s)
Proteínas ADAM/antagonistas & inhibidores , Cartílago Articular/efectos de los fármacos , Bases de Datos de Compuestos Químicos , Osteoartritis/patología , Sulfonamidas/síntesis química , Triazinas/síntesis química , Proteína ADAMTS5 , Agrecanos/metabolismo , Animales , Cartílago Articular/metabolismo , Cartílago Articular/patología , Endopeptidasas/metabolismo , Epítopos , Glicosaminoglicanos/metabolismo , Humanos , Técnicas In Vitro , Masculino , Persona de Mediana Edad , Osteoartritis/tratamiento farmacológico , Ratas , Ratas Sprague-Dawley , Bibliotecas de Moléculas Pequeñas , Estereoisomerismo , Relación Estructura-Actividad , Sulfonamidas/farmacocinética , Sulfonamidas/farmacología , Triazinas/farmacocinética , Triazinas/farmacología
12.
J Med Chem ; 52(24): 8047-56, 2009 Dec 24.
Artículo en Inglés | MEDLINE | ID: mdl-19929003

RESUMEN

Inhibition of methionine aminopeptidase-2 (MetAP2) represents a novel approach to antiangiogenic therapy. We describe the synthesis and activity of fumagillin analogues that address the pharmacokinetic and safety liabilities of earlier candidates in this compound class. Two-step elaboration of fumagillol with amines yielded a diverse series of carbamates at C6 of the cyclohexane spiroepoxide. The most potent of these compounds exhibited subnanomolar inhibition of cell proliferation in HUVEC and BAEC assays. Although a range of functionalities were tolerated at this position, alpha-trisubstituted amines possessed markedly decreased inhibitory activity, and this could be rationalized by modeling based on the known fumagillin-MetAP2 crystal structure. The lead compound resulting from these studies, (3R,4S,5S,6R)-5-methoxy-4-((2R,3R)-2-methyl-3-(3-methylbut-2-enyl)oxiran-2-yl)-1-oxaspiro[2.5]octan-6-yl (R)-1-amino-3-methyl-1-oxobutan-2-ylcarbamate, (PPI-2458), demonstrated an improved pharmacokinetic profile relative to the earlier clinical candidate TNP-470, and has advanced into phase I clinical studies in non-Hodgkin's lymphoma and solid cancers.


Asunto(s)
Aminopeptidasas/antagonistas & inhibidores , Carbamatos/química , Carbamatos/farmacología , Ciclohexanos/química , Ciclohexanos/farmacología , Ácidos Grasos Insaturados/química , Ácidos Grasos Insaturados/farmacología , Metaloendopeptidasas/antagonistas & inhibidores , Aminoácidos/química , Animales , Bovinos , Procesos de Crecimiento Celular/efectos de los fármacos , Células Endoteliales/citología , Células Endoteliales/efectos de los fármacos , Humanos , Modelos Moleculares , Sesquiterpenos/química , Sesquiterpenos/farmacología , Relación Estructura-Actividad
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