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1.
Nat Chem Biol ; 2024 Mar 22.
Artigo em Inglês | MEDLINE | ID: mdl-38519575

RESUMO

Cotransins target the Sec61 translocon and inhibit the biogenesis of an undefined subset of secretory and membrane proteins. Remarkably, cotransin inhibition depends on the unique signal peptide (SP) of each Sec61 client, which is required for cotranslational translocation into the endoplasmic reticulum. It remains unknown how an SP's amino acid sequence and biophysical properties confer sensitivity to structurally distinct cotransins. Here we describe a fluorescence-based, pooled-cell screening platform to interrogate nearly all human SPs in parallel. We profiled two cotransins with distinct effects on cancer cells and discovered a small subset of SPs, including the oncoprotein human epidermal growth factor receptor 3 (HER3), with increased sensitivity to the more selective cotransin, KZR-9873. By comparing divergent mouse and human orthologs, we unveiled a position-dependent effect of arginine on SP sensitivity. Our multiplexed profiling platform reveals how cotransins can exploit subtle sequence differences to achieve SP discrimination.

2.
Nat Chem Biol ; 19(9): 1054-1062, 2023 09.
Artigo em Inglês | MEDLINE | ID: mdl-37169961

RESUMO

Preventing the biogenesis of disease-relevant proteins is an attractive therapeutic strategy, but attempts to target essential protein biogenesis factors have been hampered by excessive toxicity. Here we describe KZR-8445, a cyclic depsipeptide that targets the Sec61 translocon and selectively disrupts secretory and membrane protein biogenesis in a signal peptide-dependent manner. KZR-8445 potently inhibits the secretion of pro-inflammatory cytokines in primary immune cells and is highly efficacious in a mouse model of rheumatoid arthritis. A cryogenic electron microscopy structure reveals that KZR-8445 occupies the fully opened Se61 lateral gate and blocks access to the lumenal plug domain. KZR-8445 binding stabilizes the lateral gate helices in a manner that traps select signal peptides in the Sec61 channel and prevents their movement into the lipid bilayer. Our results establish a framework for the structure-guided discovery of novel therapeutics that selectively modulate Sec61-mediated protein biogenesis.


Assuntos
Proteínas de Membrana , Sinais Direcionadores de Proteínas , Animais , Camundongos , Transporte Proteico , Proteínas de Membrana/metabolismo , Canais de Translocação SEC/química , Canais de Translocação SEC/genética , Canais de Translocação SEC/metabolismo , Biossíntese de Proteínas
3.
Br J Haematol ; 198(1): 137-141, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35434798

RESUMO

Glucocorticoid (GC) resistance is a poor prognostic factor in T-cell acute lymphoblastic leukaemia (T-ALL). Interleukin-7 (IL-7) mediates GC resistance via GC-induced upregulation of IL-7 receptor (IL-7R) expression, leading to increased pro-survival signalling. IL-7R reaches the cell surface via the secretory pathway, so we hypothesized that inhibiting the translocation of IL-7R into the secretory pathway would overcome GC resistance. Sec61 is an endoplasmic reticulum (ER) channel that is required for insertion of polypeptides into the ER. Here, we demonstrate that KZR-445, a novel inhibitor of Sec61, potently attenuates the dexamethasone (DEX)-induced increase in cell surface IL-7R and overcomes IL-7-induced DEX resistance.


Assuntos
Leucemia-Linfoma Linfoblástico de Células T Precursoras , Canais de Translocação SEC , Citocinas/metabolismo , Dexametasona/farmacologia , Glucocorticoides/farmacologia , Humanos , Interleucina-7 , Erros Inatos do Metabolismo , Leucemia-Linfoma Linfoblástico de Células T Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células T Precursoras/metabolismo , Receptores de Glucocorticoides/deficiência , Canais de Translocação SEC/metabolismo , Linfócitos T/metabolismo
4.
Drug Metab Dispos ; 49(9): 810-821, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34234005

RESUMO

KZR-616 is an irreversible tripeptide epoxyketone-based selective inhibitor of the human immunoproteasome. Inhibition of the immunoproteasome results in anti-inflammatory activity in vitro and based on promising therapeutic activity in animal models of rheumatoid arthritis and systemic lupus erythematosus KZR-616 is being developed for potential treatment of multiple autoimmune and inflammatory diseases. The presence of a ketoepoxide pharmacophore presents unique challenges in the study of drug metabolism during lead optimization and clinical candidate profiling. This study presents a thorough and systematic in vitro and cell-based enzymatic metabolism and kinetic investigation to identify the major enzymes involved in the metabolism and elimination of KZR-616. Upon exposure to liver microsomes in the absence of NADPH, KZR-616 and its analogs were converted to their inactive diol derivatives with varying degrees of stability. Diol formation was also shown to be the major metabolite in pharmacokinetic studies in monkeys and correlated with in vitro stability results for individual compounds. Further study in intact hepatocytes revealed that KZR-616 metabolism was sensitive to an inhibitor of microsomal epoxide hydrolase (mEH) but not inhibitors of cytochrome P450 (P450) or soluble epoxide hydrolase (sEH). Primary human hepatocytes were determined to be the most robust source of mEH activity for study in vitro. These findings also suggest that the exposure of KZR-616 in vivo is unlikely to be affected by coadministration of inhibitors or inducers of P450 and sEH. SIGNIFICANCE STATEMENT: This work presents a thorough and systematic investigation of metabolism and kinetics of KZR-616 and related analogs in in vitro and cell-based enzymatic systems. Information gained could be useful in assessing novel covalent proteasome inhibitors during lead compound optimization. These studies also demonstrate a robust source in vitro test system that correlated with in vivo pharmacokinetics for KZR-616 and two additional tripeptide epoxyketones.


Assuntos
Cisteína Endopeptidases/imunologia , Sistema Enzimático do Citocromo P-450/metabolismo , Epóxido Hidrolases/metabolismo , Morfolinas/farmacologia , Complexo de Endopeptidases do Proteassoma/imunologia , Proteínas/imunologia , Animais , Doenças Autoimunes/tratamento farmacológico , Células Cultivadas , Cisteína Endopeptidases/metabolismo , Epóxido Hidrolases/imunologia , Hepatócitos/metabolismo , Humanos , Inativação Metabólica , Inflamação/tratamento farmacológico , Macaca fascicularis , Inibidores de Proteassoma/farmacologia
5.
Front Immunol ; 14: 1043680, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36969170

RESUMO

Zetomipzomib (KZR-616) is a selective inhibitor of the immunoproteasome currently undergoing clinical investigation in autoimmune disorders. Here, we characterized KZR-616 in vitro and in vivo using multiplexed cytokine analysis, lymphocyte activation and differentiation, and differential gene expression analysis. KZR-616 blocked production of >30 pro-inflammatory cytokines in human peripheral blood mononuclear cells (PBMCs), polarization of T helper (Th) cells, and formation of plasmablasts. In the NZB/W F1 mouse model of lupus nephritis (LN), KZR-616 treatment resulted in complete resolution of proteinuria that was maintained at least 8 weeks after the cessation of dosing and was mediated in part by alterations in T and B cell activation, including reduced numbers of short and long-lived plasma cells. Gene expression analysis of human PBMCs and tissues from diseased mice revealed a consistent and broad response focused on inhibition of T, B, and plasma cell function and the Type I interferon pathway and promotion of hematopoietic cell lineages and tissue remodeling. In healthy volunteers, KZR-616 administration resulted in selective inhibition of the immunoproteasome and blockade of cytokine production following ex vivo stimulation. These data support the ongoing development of KZR-616 in autoimmune disorders such as systemic lupus erythematosus (SLE)/LN.


Assuntos
Leucócitos Mononucleares , Nefrite Lúpica , Humanos , Animais , Camundongos , Leucócitos Mononucleares/metabolismo , Citocinas/metabolismo , Imunidade
6.
Bioorg Med Chem Lett ; 22(11): 3786-90, 2012 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-22542009
7.
Bioorg Med Chem Lett ; 21(18): 5206-9, 2011 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-21840217

RESUMO

A bis-amide antagonist of Smoothened, a seven-transmembrane receptor in the Hedgehog signaling pathway, was discovered via high throughput screening. In vitro and in vivo experiments demonstrated that the bis-amide was susceptible to N-acyl transferase mediated amide scission. Several bioisosteric replacements of the labile amide that maintained in vitro potency were identified and shown to be metabolically stable in vitro and in vivo.


Assuntos
Aciltransferases/antagonistas & inibidores , Amidas/farmacologia , Descoberta de Drogas , Inibidores Enzimáticos/farmacologia , Aciltransferases/metabolismo , Amidas/química , Amidas/metabolismo , Inibidores Enzimáticos/química , Inibidores Enzimáticos/metabolismo , Ensaios de Triagem em Larga Escala , Estrutura Molecular , Estereoisomerismo , Relação Estrutura-Atividade
9.
Bioorg Med Chem Lett ; 20(15): 4607-10, 2010 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-20594845

RESUMO

Pyridopyridazine antagonists of the hedgehog signaling pathway are described. Designed to optimize our previously described phthalazine smoothened antagonists, a representative compound eliminates a PXR liability while retaining potency and in vitro metabolic stability. Moreover, the compound has improved efficacy in a hedgehog/smoothened signaling mouse pharmacodynamic model.


Assuntos
Proteínas Hedgehog/antagonistas & inibidores , Ftalazinas/química , Piperazinas/química , Piridazinas/química , Receptores de Esteroides/química , Animais , Proteínas Hedgehog/metabolismo , Humanos , Camundongos , Microssomos Hepáticos/metabolismo , Ftalazinas/síntese química , Ftalazinas/farmacocinética , Piperazinas/síntese química , Piperazinas/farmacocinética , Receptor de Pregnano X , Piridazinas/síntese química , Piridazinas/farmacocinética , Ratos , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Esteroides/metabolismo , Transdução de Sinais , Receptor Smoothened , Relação Estrutura-Atividade , Tilosina/análogos & derivados
10.
Bioorg Med Chem Lett ; 20(12): 3618-22, 2010 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-20493695

RESUMO

The Hedgehog (Hh) signaling pathway regulates cell proliferation and differentiation in developing tissues, and abnormal activation of the Hh pathway has been linked to several tumor subsets. As a transducer of Hh signaling, the GPCR-like protein Smoothened (Smo) is a promising target for disruption of unregulated Hh signaling. A series of 1-amino-4-arylphthalazines was developed as potent and orally bioavailable inhibitors of Smo. A representative compound from this class demonstrated significant tumor volume reduction in a mouse medulloblastoma model.


Assuntos
Ftalazinas/química , Ftalazinas/farmacologia , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Administração Oral , Animais , Disponibilidade Biológica , Linhagem Celular Tumoral , Sistema Enzimático do Citocromo P-450/efeitos dos fármacos , Desenho de Fármacos , Proteínas Hedgehog , Humanos , Meduloblastoma/tratamento farmacológico , Meduloblastoma/patologia , Camundongos , Ftalazinas/síntese química , Transdução de Sinais , Receptor Smoothened
11.
Bioorg Med Chem Lett ; 20(23): 7071-5, 2010 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-20971000

RESUMO

In this communication, human 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) inhibitory activities of a novel series of diarylsulfones are described. Optimization of this series resulted in several highly potent 11ß-HSD1 inhibitors with excellent pharmacokinetic (PK) properties. Compound (S)-25 showed excellent efficacy in a non-human primate ex vivo pharmacodynamic model.


Assuntos
Inibidores Enzimáticos/síntese química , Sulfonas/síntese química , 11-beta-Hidroxiesteroide Desidrogenase Tipo 1/antagonistas & inibidores , Animais , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacocinética , Humanos , Farmacocinética , Relação Estrutura-Atividade , Sulfonas/química , Sulfonas/farmacocinética
12.
Bioorg Med Chem Lett ; 19(6): 1797-801, 2009 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-19217779

RESUMO

Discovery and optimization of a piperidyl benzamide series of 11beta-HSD1 inhibitors is described. This series was derived from a cyclohexyl benzamide lead structures to address PXR selectivity, high non-specific protein binding, poor solubility, limited in vivo exposure, and in vitro cytotoxicity issues observed with the cyclohexyl benzamide structures. These efforts led to the discovery of piperidyl benzamide 15 which features improved properties over the cyclohexyl benzamide derivatives.


Assuntos
11-beta-Hidroxiesteroide Desidrogenase Tipo 1/antagonistas & inibidores , Benzamidas/síntese química , Diabetes Mellitus Tipo 2/tratamento farmacológico , Insulina/metabolismo , Piperidinas/química , 11-beta-Hidroxiesteroide Desidrogenase Tipo 1/química , Benzamidas/farmacologia , Cristalografia por Raios X/métodos , Desenho de Fármacos , Hepatócitos/efeitos dos fármacos , Humanos , Concentração Inibidora 50 , Microssomos/metabolismo , Modelos Químicos , Estrutura Molecular , Solubilidade , Relação Estrutura-Atividade
13.
Bioorg Med Chem Lett ; 19(5): 1446-50, 2009 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-19185488

RESUMO

Novel 4,4-disubstituted cyclohexylbenzamide inhibitors of 11beta-HSD1 were optimized to account for liabilities relating to in vitro pharmacokinetics, cytotoxicity and protein-related shifts in potency. A representative compound showing favorable in vivo pharmacokinetics was found to be an efficacious inhibitor of 11beta-HSD1 in a rat pharmacodynamic model (ED(50)=10mg/kg).


Assuntos
11-beta-Hidroxiesteroide Desidrogenase Tipo 1/antagonistas & inibidores , Benzamidas/química , 11-beta-Hidroxiesteroide Desidrogenase Tipo 1/metabolismo , Animais , Benzamidas/farmacologia , Relação Dose-Resposta a Droga , Células HeLa , Humanos , Macaca fascicularis , Ratos , Ratos Sprague-Dawley , Relação Estrutura-Atividade
14.
J Med Chem ; 61(24): 11127-11143, 2018 12 27.
Artigo em Inglês | MEDLINE | ID: mdl-30380863

RESUMO

Selective immunoproteasome inhibition is a promising approach for treating autoimmune disorders, but optimal proteolytic active site subunit inhibition profiles remain unknown. We reveal here our design of peptide epoxyketone-based selective low molecular mass polypeptide-7 (LMP7) and multicatalytic endopeptidase complex subunit-1 (MECL-1) subunit inhibitors. Utilizing these and our previously disclosed low molecular mass polypeptide-2 (LMP2) inhibitor, we demonstrate a requirement of dual LMP7/LMP2 or LMP7/MECL-1 subunit inhibition profiles for potent cytokine expression inhibition and in vivo efficacy in an inflammatory disease model. These and additional findings toward optimized solubility led the design and selection of KZR-616 disclosed here and presently in clinical trials for treatment of rheumatic disease.


Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Artrite Reumatoide/tratamento farmacológico , Morfolinas/farmacologia , Inibidores de Proteassoma/farmacologia , Administração Intravenosa , Animais , Anti-Inflamatórios não Esteroides/química , Anti-Inflamatórios não Esteroides/farmacocinética , Artrite Experimental/tratamento farmacológico , Linhagem Celular Tumoral , Cisteína Endopeptidases/metabolismo , Citocinas/metabolismo , Desenho de Fármacos , Feminino , Humanos , Camundongos Endogâmicos BALB C , Morfolinas/química , Morfolinas/farmacocinética , Oligopeptídeos/farmacologia , Complexo de Endopeptidases do Proteassoma/imunologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Inibidores de Proteassoma/química , Inibidores de Proteassoma/farmacocinética , Relação Estrutura-Atividade
15.
ACS Med Chem Lett ; 8(4): 413-417, 2017 Apr 13.
Artigo em Inglês | MEDLINE | ID: mdl-28435528

RESUMO

Building upon the success of bortezomib (VELCADE) and carfilzomib (KYPROLIS), the design of a next generation of inhibitors targeting specific subunits within the immunoproteasome is of interest for the treatment of autoimmune disease. There are three catalytic subunits within the immunoproteasome (low molecular mass polypeptide-7, -2, and multicatalytic endopeptidase complex subunit-1; LMP7, LMP2, and MECL-1), and a campaign was undertaken to design a potent and selective LMP2 inhibitor with sufficient properties to allow for sustained inhibition in vivo. Screening a focused library of epoxyketones revealed a series of potent dipeptides that were optimized to provide the highly selective inhibitor KZR-504 (12).

16.
Nucleic Acids Res ; 31(4): 1282-91, 2003 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-12582248

RESUMO

Polyamides are a class of heterocyclic small molecules with the potential of controlling gene expression by binding to the minor groove of DNA in a sequence-specific manner. To evaluate the feasibility of this class of compounds as antiviral therapeutics, molecules were designed to essential sequence elements occurring numerous times in the HPV genome. This sequence element is bound by a virus-encoded transcription and replication factor E2, which binds to a 12 bp recognition site as a homodimeric protein. Here, we take advantage of polyamide:DNA and E2:DNA co-crystal structural information and advances in polyamide synthetic chemistry to design tandem hairpin polyamides that are capable of displacing the major groove-binding E2 homodimer from its DNA binding site. The binding of tandem hairpin polyamides and the E2 DNA binding protein to the DNA site is mutually exclusive even though the two ligands occupy opposite faces of the DNA double helix. We show with circular permutation studies that the tandem hairpin polyamide prevents the intrinsic bending of the E2 DNA site important for binding of the protein. Taken together, these results illustrate the feasibility of inhibiting the binding of homodimeric, major groove-binding transcription factors by altering the local DNA geometry using minor groove-binding tandem hairpin polyamides.


Assuntos
DNA Viral/metabolismo , Proteínas de Ligação a DNA , Nylons/metabolismo , Proteínas Oncogênicas Virais/metabolismo , Sítios de Ligação/genética , DNA Viral/química , Modelos Biológicos , Conformação de Ácido Nucleico , Nylons/química , Oligonucleotídeos/genética , Oligonucleotídeos/metabolismo , Proteínas Oncogênicas Virais/química , Papillomaviridae/genética , Papillomaviridae/metabolismo , Ligação Proteica
17.
ACS Med Chem Lett ; 5(11): 1245-50, 2014 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-25408839

RESUMO

In this letter, we reported the design and synthesis of three potent, selective, and orally bioavailable 11ß-HSD1 inhibitors labeled with (14)C: AMG 456 (1), AM-6949 (2), and AM-7715 (3). We evaluated the covalent protein binding of the labeled inhibitors in human liver microsomes in vitro and assessed their potential bioactivation risk in humans. We then studied the in vitro mechanism of 2 in human hepatocytes and the formation of reactive intermediates. Our study results suggest that 1 and 3 have low potential for metabolic bioactivation in humans, while 2 has relatively high risk.

18.
J Med Chem ; 57(6): 2472-88, 2014 Mar 27.
Artigo em Inglês | MEDLINE | ID: mdl-24548297

RESUMO

We previously reported the discovery of AMG 232, a highly potent and selective piperidinone inhibitor of the MDM2-p53 interaction. Our continued search for potent and diverse analogues led to the discovery of novel morpholinone MDM2 inhibitors. This change to a morpholinone core has a significant impact on both potency and metabolic stability compared to the piperidinone series. Within this morpholinone series, AM-8735 emerged as an inhibitor with remarkable biochemical potency (HTRF IC50 = 0.4 nM) and cellular potency (SJSA-1 EdU IC50 = 25 nM), as well as pharmacokinetic properties. Compound 4 also shows excellent antitumor activity in the SJSA-1 osteosarcoma xenograft model with an ED50 of 41 mg/kg. Lead optimization toward the discovery of this inhibitor as well as key differences between the morpholinone and the piperidinone series will be described herein.


Assuntos
Acetatos/síntese química , Acetatos/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Morfolinas/síntese química , Morfolinas/farmacologia , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-mdm2/química , Proteína Supressora de Tumor p53/antagonistas & inibidores , Proteína Supressora de Tumor p53/química , Animais , Linhagem Celular Tumoral , Cristalografia por Raios X , Descoberta de Drogas , Humanos , Indicadores e Reagentes , Camundongos , Modelos Moleculares , Conformação Molecular , Morfolinas/farmacocinética , Ratos , Relação Estrutura-Atividade , Ensaios Antitumorais Modelo de Xenoenxerto
19.
ACS Med Chem Lett ; 5(8): 894-9, 2014 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-25147610

RESUMO

Continued optimization of the N-substituent in the piperidinone series provided potent piperidinone-pyridine inhibitors 6, 7, 14, and 15 with improved pharmacokinetic properties in rats. Reducing structure complexity of the N-alkyl substituent led to the discovery of 23, a potent and simplified inhibitor of MDM2. Compound 23 exhibits excellent pharmacokinetic properties and substantial in vivo antitumor activity in the SJSA-1 osteosarcoma xenograft mouse model.

20.
J Med Chem ; 57(8): 3464-83, 2014 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-24670009

RESUMO

The discovery and optimization of a series of acyl CoA:diacylglycerol acyltransferase 1 (DGAT1) inhibitors based on a pyrimido[4,5-b][1,4]oxazine scaffold is described. The SAR of a moderately potent HTS hit was investigated resulting in the discovery of phenylcyclohexylacetic acid 1, which displayed good DGAT1 inhibitory activity, selectivity, and PK properties. During preclinical toxicity studies a metabolite of 1 was observed that was responsible for elevating the levels of liver enzymes ALT and AST. Subsequently, analogues were synthesized to preclude the formation of the toxic metabolite. This effort resulted in the discovery of spiroindane 42, which displayed significantly improved DGAT1 inhibition compared to 1. Spiroindane 42 was well tolerated in rodents in vivo, demonstrated efficacy in an oral triglyceride uptake study in mice, and had an acceptable safety profile in preclinical toxicity studies.


Assuntos
Diacilglicerol O-Aciltransferase/antagonistas & inibidores , Inibidores Enzimáticos/síntese química , Oxazinas/síntese química , Alanina Transaminase/sangue , Animais , Aspartato Aminotransferases/sangue , Cães , Descoberta de Drogas , Inibidores Enzimáticos/farmacologia , Ensaios de Triagem em Larga Escala , Macaca mulatta , Camundongos , Camundongos Endogâmicos C57BL , Oxazinas/farmacologia , Ratos , Ratos Sprague-Dawley , Relação Estrutura-Atividade , Triglicerídeos/metabolismo
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