Ho-Shen Lin

Non-peptide angiotensin II receptor antagonists discriminate subtypes of 125I-angiotensin II binding sites in the rat brain.

We have utilized quantitative autoradiography to define subtypes of 125I-angiotensin II (AII) binding in rat brain. AII-1 binding (displaced by DuP 753) was found in the nucleus of the solitary tract and the hypothalamus, while AII-2 binding (displaced by WL 19) was found in the thalamus and lateral septum. These results indicate that subtypes of the AII receptor are present in the brain and the AII-1 receptor subtype is present in regions consistant with the known actions of angiotensin.

Dibasic benzo[b]thiophene derivatives as a novel class of active site directed thrombin inhibitors: 2. Sidechain optimization and demonstration of in vivo efficacy.

Potent, subnanomolar thrombin inhibitors 4, 5, and 6 are developed through side chain optimization of novel, benzo[b]thiophene-based small organic entities 2 and 3 and through SAR additivity studies of the new structural elements identified. X-ray crystallographic studies of 4b-thrombin complex revealed a hydrophobic and an electrostatic interaction of these new elements with thrombin at the S2 and S3 binding sites. In vitro and in vivo pharmacological studies showed that 4, 5, and 6 are potent anticoagulants in human plasma with demonstrated antithrombotic efficacy in a rat model of thrombosis.

Synthesis and in vitro biological activity of 4α-(2-propenyl)-5α-cholest-24-en-3α,12α-diol, a 12α-hydroxyl analog of 4α-(2-propenyl)-5α-cholest-24-en-3α-ol: The latter is a potent activator of the low-density lipoprotein receptor promoter

Abstract 4α-(2-Propenyl)-5α-cholest-24-en-3α-ol (3) was shown recently in a Chinese hamster ovary (CHO) cell-based low-density lipoprotein receptor/luciferase (LDLR/Luc) assay to be a potent transcriptional activator of the LDL receptor promoter in the presence of 25-hydroxycholesterol. Because of the involvement of 12α-hydroxylation in the metabolism of cholesterol, we are interested in investigating the effect of introducing a 12α-hydroxyl group to 3 on the transcriptional activity of the LDL receptor promoter. Thus 4α-(2-propenyl)-5α-cholest-24-en-3α,12α-diol (14), a 12α-hydroxyl analog of 3, was synthesized from deoxycholic acid via the formation of 12α-[[(tert-butyl)dimethylsilyl]oxy]-4α-(2-propenyl)-5α-cholest-24-en-3-one (11). Test results show that 14 is inactive at concentrations of up to 20 μg/ml, compared to 3 with an EC30 value of 2.6 μM, in the CHO cell-based LDLR/Luc assay. Apparently introduction of a 12α-hydroxyl group abolishes the capability of 3α-sterol 14 to activate the transcription of the LDL receptor promoter. However, in the [1-14C-acetate]cholesterol biosynthesis inhibition assay in CHO cells, 14 at 10 μg/ml (23 μM) is shown to inhibit the cholesterol biosynthesis by 51% relative to the control cells. Our previous studies indicated that 3 showed a 38% inhibition, but 4α-(2-propenyl)-5α-cholestan-3α-ol (1) exhibited no inhibition in the same assay at 10 μg/ml. In summary the results indicate that, in addition to the 24,25-unsaturation, the 12α-hydroxyl group in 14 has also conferred an inhibitory effect on cholesterol biosynthesis in CHO cells; however, the inhibition of cholesterol biosynthesis by 14 does not lead to the transcriptional activation of the LDL receptor promoter.

Synthesis and in vitro biological activity of 4α-(2-propenyl)-5α-cholest-24-en-3α-ol: A 24,25-dehydro analog of the hypocholesterolemic agent 4α-(2-propenyl)-5α-cholestan-3α-ol

4Alpha-(2-propenyl)-5alpha-cholestan-3alpha-ol (LY295427) was previously identified from a Chinese hamster ovary (CHO) cell-based low density lipoprotein receptor/luciferase (LDLR/Luc) assay to be a potent transcriptional activator of the LDL receptor promoter in the presence of 25-hydroxycholesterol. To investigate the effect of the 24,25-unsaturation in the D-ring side chain (desmosterol D-ring side chain) on antagonizing the repressing effect of 25-hydroxycholesterol, 4alpha-(2-propenyl)-5alpha-cholest-24-en-3alpha-ol (17), a 24,25-dehydro analog of LY295427, was thus synthesized from lithocholic acid via the formation of 3alpha-[[(1,1-dimethylethyl)dimethylsilyl]oxy]-4alpha- (2-propenyl)-5alpha-cholan-24-al (15). Test results showed that 17 had an EC30 value of 2.6 microM, comparable to 2.9 microM of LY295427, in the CHO cell-based LDLR/Luc assay in the presence of 25-hydroxycholesterol. Apparently, the built-in 24,25-unsaturation in the D-ring side chain of 17 had added little effect to antagonizing the repressing effect of 25-hydroxycholesterol. In the [1-14C-acetate]cholesterol biosynthesis inhibition assay, 17 at 10 microg/ml (23 microM) has been shown to inhibit the cholesterol biosynthesis in CHO cells by 38% relative to the vehicle control; whereas LY295427 showed no inhibition in the same assay in our previous studies. In contrast to LY295427, the built-in 24,25-unsaturation in the D-ring side chain of 17 has conferred an inhibitory effect on cholesterol biosynthesis in CHO cells. In summary, the observed LDL receptor promoter activity of 17 is related to its ability to prevent 25-hydroxycholesterol from exerting the repressing effect via an undetermined mechanism and, in part, to inhibit the cholesterol biosynthesis.

Synthesis of 4α-(2-propenyl)-5,6-secocholestan-3α-ol, a Novel B-ring Seco Analog of the Hypocholesterolemic Agent 4α-(2-propenyl)-5α-cholestan-3α-ol

Abstract 4α-(2-Propenyl)-5α-cholestan-3α-ol (LY295427) was previously identified from a CHO cell-based assay to be a potent LDL receptor up-regulator and had demonstrated to be an effective agent in lowering plasma cholesterol levels in hypercholesterolemic hamsters. In order to investigate the effect of flexibility of the 3α-hydroxy-bearing A-ring on the activity, 4α-(2-propenyl)-5,6-secocholestan-3α-ol ( 11 ), a B-ring seco analog of LY295427, is thus synthesized from cholest-4-en-3-one. Test results indicate that 11 is not active in the CHO cell-based LDL receptor/luciferase assay at concentrations up to 20 μg/mL. The result underlines the importance of maintaining the A-B-C-D ring rigidity of the 3α-sterols in terms of binding to the putative oxysterol receptor.