An efficient synthesis of a 4'-phosphonodifluoromethyl-3'-formyl-phenylalanine containing SRC SH2 ligand.
A CuBr-mediated, regioselective cross-coupling between methyl 2,5-diiodobenzoate (4) and [(diethoxyphosphinyl)difluoromethyl]zinc bromide is reported. Palladium-catalyzed incorporation of an amino acid side chain, followed by subsequent modifications resulted in the rapid construction of 2. Compound 2 was designed to engage Cys188 of the Src SH2 domain, however, this was not observed spectroscopically.
Structure-based design of an osteoclast-selective, nonpeptide src homology 2 inhibitor with in vivo antiresorptive activity.
Targeted disruption of the pp60(src) (Src) gene has implicated this tyrosine kinase in osteoclast-mediated bone resorption and as a therapeutic target for the treatment of osteoporosis and other bone-related diseases. Herein we describe the discovery of a nonpeptide inhibitor (AP22408) of Src that demonstrates in vivo antiresorptive activity. Based on a cocrystal structure of the noncatalytic Src homology 2 (SH2) domain of Src complexed with citrate [in the phosphotyrosine (pTyr) binding pocket], we designed 3',4'-diphosphonophenylalanine (Dpp) as a pTyr mimic. In addition to its design to bind Src SH2, the Dpp moiety exhibits bone-targeting properties that confer osteoclast selectivity, hence minimizing possible undesired effects on other cells that have Src-dependent activities. The chemical structure AP22408 also illustrates a bicyclic template to replace the post-pTyr sequence of cognate Src SH2 phosphopeptides such as Ac-pTyr-Glu-Glu-Ile (1). An x-ray structure of AP22408 complexed with Lck (S164C) SH2 confirmed molecular interactions of both the Dpp and bicyclic template of AP22408 as predicted from molecular modeling. Relative to the cognate phosphopeptide, AP22408 exhibits significantly increased Src SH2 binding affinity (IC(50) = 0.30 microM for AP22408 and 5.5 microM for 1). Furthermore, AP22408 inhibits rabbit osteoclast-mediated resorption of dentine in a cellular assay, exhibits bone-targeting properties based on a hydroxyapatite adsorption assay, and demonstrates in vivo antiresorptive activity in a parathyroid hormone-induced rat model.
Mechanism of the depressor response to dopamine in the rat treated with phenoxybenzamine.
Alterations of hypotensive responses to dopamine by antagonists were characterized in alpha-blocked, anaesthetized rats. Responses were not affected by d-propranolol (0.1 mg/kg) whereas d,1-propranolol (0.1 mg/kg) or haloperidol (1.0 mg/kg) attenuated them; higher doses of inhibitors (1.0 mg/kg; 5.0 mg/kg, respectively) failed to produce a higher inhibition, but combinations of low doses abolished the depressor responses. In adrenalectomized rats, hypotensive responses decreased; haloperidol always attenuated the responses while d,1-propranolol became ineffective. Dopamine produced an enhancement of plasma adrenaline and noradrenaline levels, which was decreased by d,1-propranolol and increased by haloperidol. The data suggest that in rats the depressor component of dopamine is due to activation of both dopaminergic and beta-adrenoceptors. The beta component appears to be due to the release of adrenaline. The results also support the concept of the existence in sympathetic nerve endings and adrenal glands of stimulatory beta-adrenergic and inhibitory dopaminergic prejunctional receptors.