RESUMEN
BACKGROUND: Amyloid-ß (Aß) 42 has been implicated as the initiating molecule in the pathogenesis of Alzheimer's disease (AD); thus, therapeutic strategies that target Aß42 are of great interest. γ-Secretase modulators (GSMs) are small molecules that selectively decrease Aß42. We have previously reported that many acidic steroids are GSMs with potencies ranging in the low to mid micromolar concentration with 5ß-cholanic acid being the most potent steroid identified GSM with half maximal effective concentration (EC50) of 5.7 µM. RESULTS: We find that the endogenous cholesterol metabolite, 3ß-hydroxy-5-cholestenoic acid (CA), is a steroid GSM with enhanced potency (EC50 of 250 nM) relative to 5ß-cholanic acid. CA i) is found in human plasma at ~100-300 nM concentrations ii) has the typical acidic GSM signature of decreasing Aß42 and increasing Aß38 levels iii) is active in in vitro γ-secretase assay iv) is made in the brain. To test if CA acts as an endogenous GSM, we used Cyp27a1 knockout (Cyp27a1-/-) and Cyp7b1 knockout (Cyp7b1-/-) mice to investigate if manipulation of cholesterol metabolism pathways relevant to CA formation would affect brain Aß42 levels. Our data show that Cyp27a1-/- had increased brain Aß42, whereas Cyp7b1-/- mice had decreased brain Aß42 levels; however, peripheral dosing of up to 100 mg/kg CA did not affect brain Aß levels. Structure-activity relationship (SAR) studies with multiple known and novel CA analogs studies failed to reveal CA analogs with increased potency. CONCLUSION: These data suggest that CA may act as an endogenous GSM within the brain. Although it is conceptually attractive to try and increase the levels of CA in the brain for prevention of AD, our data suggest that this will not be easily accomplished.
Asunto(s)
Secretasas de la Proteína Precursora del Amiloide/metabolismo , Péptidos beta-Amiloides/metabolismo , Encéfalo/metabolismo , Colesterol/análogos & derivados , Fragmentos de Péptidos/metabolismo , Animales , Barrera Hematoencefálica , Células CHO , Células Cultivadas , Colestanotriol 26-Monooxigenasa/deficiencia , Colestanotriol 26-Monooxigenasa/genética , Colesterol/química , Colesterol/metabolismo , Colesterol/farmacología , Ácidos Cólicos/farmacología , Técnicas de Cocultivo , Cricetinae , Cricetulus , Familia 7 del Citocromo P450 , Relación Dosis-Respuesta a Droga , Evaluación Preclínica de Medicamentos , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Estructura Molecular , Neuroglía/metabolismo , Neuronas/metabolismo , Esteroide Hidroxilasas/deficiencia , Esteroide Hidroxilasas/genética , Relación Estructura-ActividadRESUMEN
Using a cell-based assay, we have identified optimal residues and key recognition elements necessary for inhibition of gamma-secretase. An (S)-hydroxy group or 3,5-difluorophenylacetyl group at the amino terminus and N-methyltertiary amide moiety at the carboxy terminus provided potent gamma-secretase inhibitors with an IC(50) <10 nM.