RESUMO
The NADase SARM1 (sterile alpha and TIR motif containing 1) is a key executioner of axon degeneration and a therapeutic target for several neurodegenerative conditions. We show that a potent SARM1 inhibitor undergoes base exchange with the nicotinamide moiety of nicotinamide adenine dinucleotide (NAD+) to produce the bona fide inhibitor 1AD. We report structures of SARM1 in complex with 1AD, NAD+ mimetics and the allosteric activator nicotinamide mononucleotide (NMN). NMN binding triggers reorientation of the armadillo repeat (ARM) domains, which disrupts ARM:TIR interactions and leads to formation of a two-stranded TIR domain assembly. The active site spans two molecules in these assemblies, explaining the requirement of TIR domain self-association for NADase activity and axon degeneration. Our results reveal the mechanisms of SARM1 activation and substrate binding, providing rational avenues for the design of new therapeutics targeting SARM1.
Assuntos
Proteínas do Domínio Armadillo , NAD , Proteínas do Domínio Armadillo/genética , Proteínas do Citoesqueleto/química , Proteínas do Citoesqueleto/genética , NAD/metabolismo , NAD+ Nucleosidase/metabolismo , Domínios ProteicosRESUMO
Axonal degeneration is an early and ongoing event that causes disability and disease progression in many neurodegenerative disorders of the peripheral and central nervous systems. Chemotherapy-induced peripheral neuropathy (CIPN) is a major cause of morbidity and the main cause of dose reductions and discontinuations in cancer treatment. Preclinical evidence indicates that activation of the Wallerian-like degeneration pathway driven by sterile alpha and TIR motif containing 1 (SARM1) is responsible for axonopathy in CIPN. SARM1 is the central driver of an evolutionarily conserved programme of axonal degeneration downstream of chemical, inflammatory, mechanical or metabolic insults to the axon. SARM1 contains an intrinsic NADase enzymatic activity essential for its pro-degenerative functions, making it a compelling therapeutic target to treat neurodegeneration characterized by axonopathies of the peripheral and central nervous systems. Small molecule SARM1 inhibitors have the potential to prevent axonal degeneration in peripheral and central axonopathies and to provide a transformational disease-modifying treatment for these disorders. Using a biochemical assay for SARM1 NADase we identified a novel series of potent and selective irreversible isothiazole inhibitors of SARM1 enzymatic activity that protected rodent and human axons in vitro. In sciatic nerve axotomy, we observed that these irreversible SARM1 inhibitors decreased a rise in nerve cADPR and plasma neurofilament light chain released from injured sciatic nerves in vivo. In a mouse paclitaxel model of CIPN we determined that Sarm1 knockout mice prevented loss of axonal function, assessed by sensory nerve action potential amplitudes of the tail nerve, in a gene-dosage-dependent manner. In that CIPN model, the irreversible SARM1 inhibitors prevented loss of intraepidermal nerve fibres induced by paclitaxel and provided partial protection of axonal function assessed by sensory nerve action potential amplitude and mechanical allodynia.
Assuntos
Proteínas do Domínio Armadillo/antagonistas & inibidores , Axônios/efeitos dos fármacos , Proteínas do Citoesqueleto/antagonistas & inibidores , Paclitaxel/toxicidade , Doenças do Sistema Nervoso Periférico/induzido quimicamente , Doenças do Sistema Nervoso Periférico/tratamento farmacológico , Tiazóis/uso terapêutico , Animais , Antineoplásicos Fitogênicos/toxicidade , Proteínas do Domínio Armadillo/deficiência , Proteínas do Domínio Armadillo/genética , Axônios/metabolismo , Células Cultivadas , Proteínas do Citoesqueleto/deficiência , Proteínas do Citoesqueleto/genética , Relação Dose-Resposta a Droga , Células HEK293 , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/metabolismo , Camundongos , Camundongos Knockout , Doenças do Sistema Nervoso Periférico/genética , Doenças do Sistema Nervoso Periférico/metabolismo , Tiazóis/farmacologiaRESUMO
The optimisation of two series of 4-hydroxybenzothiazolone derived ß2-adrenoceptor agonists, bearing α-substituted cyclopentyl and ß-phenethyl amino-substituents, as inhaled long-acting bronchodilators is described. Analogues were selected for synthesis using a lipophilicity based hypothesis to achieve the targeted rapid onset of action in combination with a long duration of action. The profiling of the two series led to identification of the α-substituted cyclopentyl analogue 2 as the optimal compound with a comparable profile to the inhaled once-daily long-acting ß2-adrenoceptor agonist indacaterol. On the basis of these data 2 was promoted as the backup development candidate to indacaterol from the Novartis LABA project.
Assuntos
Agonistas de Receptores Adrenérgicos beta 2/administração & dosagem , Agonistas de Receptores Adrenérgicos beta 2/farmacologia , Benzotiazóis/administração & dosagem , Benzotiazóis/farmacologia , Receptores Adrenérgicos beta 2/metabolismo , Administração por Inalação , Agonistas de Receptores Adrenérgicos beta 2/química , Animais , Benzotiazóis/química , Relação Dose-Resposta a Droga , Cobaias , Estrutura MolecularRESUMO
The synthesis of two series of 4'-aza-carbocyclic nucleosides are described in which the 4'-substituent is either a reversed amide, relative to the carboxamide of NECA, or an N-bonded heterocycle. Using established purine substitution patterns, potent and selective examples of agonists of the human adenosine A(2A) receptor have been identified from both series. The propionamides 14-18 and the 4-hydroxymethylpyrazole 32 were determined to be the most potent and selective examples from the 4'-reversed amide and 4'-N-bonded heterocyclic series, respectively.
Assuntos
Agonistas do Receptor A2 de Adenosina , Compostos Aza/síntese química , Ácidos Carboxílicos/síntese química , Nucleosídeos/síntese química , Nucleotídeos de Pirimidina/síntese química , Animais , Compostos Aza/metabolismo , Compostos Aza/farmacologia , Células CHO , Ácidos Carboxílicos/metabolismo , Ácidos Carboxílicos/farmacologia , Cricetinae , Cricetulus , Avaliação Pré-Clínica de Medicamentos/métodos , Humanos , Nucleosídeos/metabolismo , Nucleosídeos/farmacologia , Nucleotídeos de Pirimidina/metabolismo , Nucleotídeos de Pirimidina/farmacologia , Ratos , Receptor A2A de Adenosina/metabolismoRESUMO
The chiral synthesis of a 4-hydroxybenzothiazolone based series of beta(2)-adrenoceptor agonists is described. Using this methodology a library of N-substituted analogues were prepared for the rapid identification of leads with the potential to be fast onset and long-acting inhaled bronchodilators with improved therapeutic margins. The design of the library to achieve the targeted profile was based upon lipophilicity and metabolism based hypotheses. This approach identified beta-phenethyl, alpha-substituted cyclopentyl and monoterpene N-substituents to be of particular interest for further evaluation, as exemplified by structures 19, 29 and 33, respectively.
Assuntos
Antagonistas de Receptores Adrenérgicos beta 2/uso terapêutico , Broncodilatadores/uso terapêutico , Tiazóis/uso terapêutico , Administração por Inalação , Antagonistas de Receptores Adrenérgicos beta 2/farmacologia , Broncodilatadores/administração & dosagem , Broncodilatadores/farmacologia , Tiazóis/farmacologiaRESUMO
A kinetic template-guided tethering (KTGT) strategy has been developed for the site-directed discovery of fragments that bind to defined protein surfaces, where acrylamide-modified fragments can be irreversibly captured in a protein-templated conjugate addition reaction. Herein, an efficient and facile method is reported for the preparation of acrylamide libraries from a diverse range of amine fragments using a solid-supported quaternary amine base.