Abstract-The inhibition of 22-oxidosqualene-lanosterol cyclase (EC 5.4.99.7) (OSC) by new azasqualene derivatives, mimicking the proC-8 and proC-20 carbocationic high-energy intermediates of the cyclization of 2.3-oxidosqualene to lanosterol, was studied using pig liver microsomes, partially purified preparations of OSC, and yeast microsomes. The azasqualene derivatives tested were: 6E- and 6Z-10aza-10.11-dihydrosqualene-2,3- epoxide 17 and 18, 19-aza-l8,19,22,23-tetrahydrosqualene-2,3-epoxi1d9e and its corresponding N-oxide 20, and 19-aza-18,19.22,23-tetrahydrosqualene 21. The compounds 17 and 19 (i.e. the derivatives bearing the 2.3-epoxide ring and the same geometrical configuration as the OSC substrate) were effective inhibitors, as shown by the K, obtained using partially purified OSC: 2.67 pM and 2.14 pM, respectively. Compound 18, having an incorrect configuration and the 19-aza derivative 21, lacking the 2,3-epoxide ring, were poor inhibitors, with IC,, of 44 pM and 70 pM, respectively. Compound 21 was a competitive inhibitor of OSC. whereas 17 and 19 were noncompetitive inhibitors, and showed a biphasic time-dependent inactivation of OSC, their apparent binding constants being 250 pM and 213 pM, respectively. The inhibition of sterol biosynthesis was studied using human hepatoma HepG2 cells. The incorporation of [I4C] acetate in the C,, sterols was reduced by 50% by 0.55 pM 17, 0.22 pM 19, and 0.45 pM 2 1 , whereas 2 pM 18 did not affect sterol biosynthesis. In the presence of 17, 19 and 21, only the intermediate metabolites 2,3-oxidosqualene and 2,3,22,23-dioxidosqualene accumulated, demonstrating a very specific inhibition of OSC.
Inhibition of 2,3-oxidosqualene cyclase and sterol biosynthesis by 10- and 19-azasqualene derivatives.
VIOLA, Franca Cecilia;BRUSA, Paola;BALLIANO, Gianni;CERUTI, Maurizio;CATTEL, Luigi
1995-01-01
Abstract
Abstract-The inhibition of 22-oxidosqualene-lanosterol cyclase (EC 5.4.99.7) (OSC) by new azasqualene derivatives, mimicking the proC-8 and proC-20 carbocationic high-energy intermediates of the cyclization of 2.3-oxidosqualene to lanosterol, was studied using pig liver microsomes, partially purified preparations of OSC, and yeast microsomes. The azasqualene derivatives tested were: 6E- and 6Z-10aza-10.11-dihydrosqualene-2,3- epoxide 17 and 18, 19-aza-l8,19,22,23-tetrahydrosqualene-2,3-epoxi1d9e and its corresponding N-oxide 20, and 19-aza-18,19.22,23-tetrahydrosqualene 21. The compounds 17 and 19 (i.e. the derivatives bearing the 2.3-epoxide ring and the same geometrical configuration as the OSC substrate) were effective inhibitors, as shown by the K, obtained using partially purified OSC: 2.67 pM and 2.14 pM, respectively. Compound 18, having an incorrect configuration and the 19-aza derivative 21, lacking the 2,3-epoxide ring, were poor inhibitors, with IC,, of 44 pM and 70 pM, respectively. Compound 21 was a competitive inhibitor of OSC. whereas 17 and 19 were noncompetitive inhibitors, and showed a biphasic time-dependent inactivation of OSC, their apparent binding constants being 250 pM and 213 pM, respectively. The inhibition of sterol biosynthesis was studied using human hepatoma HepG2 cells. The incorporation of [I4C] acetate in the C,, sterols was reduced by 50% by 0.55 pM 17, 0.22 pM 19, and 0.45 pM 2 1 , whereas 2 pM 18 did not affect sterol biosynthesis. In the presence of 17, 19 and 21, only the intermediate metabolites 2,3-oxidosqualene and 2,3,22,23-dioxidosqualene accumulated, demonstrating a very specific inhibition of OSC.
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