Browsing by Author "Schroepfer, George J., Jr."

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    Chemical, biochemical, and bioanalytical studies of sterols and isoprenoids: Smith-Lemli-Opitz syndrome, Langer-Giedion syndrome, activation of meiosis, nuclear orphan receptor LXRalpha and cytochrome P450s
    (2000) Ruan, Benfang; Schroepfer, George J., Jr.; Matsuda, Seiichi P. T.
    Sterols and isoprenoids are vital cellular constituents. In studies of their formation, metabolism, and biological effects, the separation and identification of individual isomers present formidable challenges. Approximately 30 unsaturated C27 sterols were prepared and purified as authentic standards to evaluate and develop chromatographic and spectral methods. Novel chromatographic conditions were devised for silver ion HPLC (Ag+-HPLC), a technique that provided unprecedented separations of these closely related sterols. Ag+-HPLC proved very powerful in addressing several important problems in medicine and biology relating to the biochemical effects of sterols and isoprenoids; described herein. Smith-Lemli-Opitz syndrome (SLOS) is a genetic disorder of development associated with the accumulation of unsaturated C27, sterols. My analysis of SLOS and normal blood samples unequivocally demonstrated that only the Delta5,7, Delta5,8, and Delta5,7,9(11) sterols accumulate significantly in SLOS patients. The 19-nor-Delta5,7,9 sterol, which has been reported to accumulate in SLOS blood by several research groups, was not detected in my analyses and was demonstrated to be a GC artifact arising from the thermal decomposition of cholesta-5,8-dien-3beta-ol. Alternative pathways in the late stages of cholesterol biosynthesis relating to the biological origin and metabolism of the beta5,8, beta6,8, and beta6,8(14) sterols were elucidated by incubation of tritium-labeled substrates in rat liver preparations. Also, a simple and rapid colorimetric method was developed for the clinical screening of SLOS. Ag +-HPLC played a critical role in studying a possible biochemical defect in another genetic disorder, Langer-Giedion syndrome, and in isolating isomers of (20R,22R)-dihydroxycholesterol), all-trans geranylgeranoic acid, other isoprenoids, and 4,4-dimethyl-sterols, compounds obtained from novel and efficient chemical syntheses. (20R,22R)-Dihydroxycholesterol proved to be a moderately potent activator of the nuclear orphan receptor LXRalpha, whereas all trans geranylgeranoic acid was found to be an inhibitor. The synthetic 4,4-dimethyl sterols caused a resumption of meiosis in mouse oocytes. Famesoic acid was demonstrated to be an extraordinarily potent substrate inducer of cytochrome P450BM-3.
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    Process for synthesis of 5.alpha.-cholest-8(14)-en-3.beta.-ol-15-one and other 15-oxygenated sterols
    (1990-01-30) Schroepfer, George J., Jr.; Wilson, William K.; Wang, Ker-shi; Kisic, Alemka; Rice University; United States Patent and Trademark Office
    A process for preparing 15-oxygenated sterols, such as 3β-hydroxy-5α-cholest-8(14)-ene-15 one, comprising converting 7-dehydrocholesterol to 3β-benzoyloxycholesta-5,7-diene, converting the 3β-benzoyloxycholesta-5,7-diene to a 3β-benzoyloxy-5-cholesta-7,14-diene, converting the 3β-benzoyloxy-5-cholesta-7,14-diene to a 3β-benzoyloxy-14α, 15α-epoxy-5-cholest-7-ene and converting the 3β-benzoyloxy-14α, 15α-epoxy-5-cholest-7-ene to a 15-oxygenated sterol. Preferably, the 3β-benzoyloxy-cholesta-5,7-diene is converted to a 3β-benzoyloxy-5-cholesta-7,14-diene by (i) contacting 3β-benzoyloxy-cholesta-5,7-diene, in a solvent at a temperature of at most about -55° C., with HCl at a concentration of at least about 2.0 M for a time sufficient to convert the 3β-benzoyloxycholesta-5,7-diene to a 3β-benzoyloxy-5-cholesta-7,14-diene; (ii) neutralizing the resultant reaction mixture with a base to prevent formation of a significant amount of 3β-benzoyloxy-5-cholesta-8,14-diene; and (iii) recovering the 3β-benzoyloxy-5-cholesta-7,14-diene.
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    Side chain derivatized 15-oxygenated sterols- methods of using them and a process for preparing them
    (1994-12-06) Schroepfer, George J., Jr.; Herz, Josef E.; Swaminathan, Shankar; Wilson, William K.; Rice University; United States Patent and Trademark Office
    Pharmaceutical compositions are provided for lowering the activity of HMG-CoA reductase and/or lowering serum cholesterol, comprising an amount effective to lower the activity of HMG-CoA reductase and/or lower serum cholesterol of a side chain derivatized 15-oxygenated sterol having the formula (I): ##STR1## the basic ring structure being saturated or unsaturated, wherein R1 is --OH, O, --OR7, ##STR2## a sulfate group, a sugar moiety, or a Mg, Na, or K salt of a sulfate group; R2 is --H, --OH, O, mono- or di-halogen, or a C1 to C6 alkyl group, which may be unsaturated or substituted with halogen; R3 is --H, --OH, halogen, or a C1 to C6 alkyl group, which may be unsaturated or substituted with halogen; R4 is nonexistent when there is a double bond between the 8 and 14 carbons or αH, βH, or an αC1 to C6 alkyl group; R5 is --OH, O, NOH, or ##STR3## R6 is --CH2 CH(CH3)2 or CH2 N(CH3)2, in which one or more of the hydrogen atoms is replaced by OH or halogen; R7 is a C1 to C6 alkyl group; R8 is a C1 to C20 aliphatic group, which may be substituted or unsubstituted, or a phenyl group; and n is an integer of from 2 to 6; andoptionally a pharmaceutically acceptable carrier or excipient, with the proviso that R6 is not --CH2 CH(CH3)(CH2 OH). Methods of using the pharmaceutical compositions containing the side chain derivatized 15-oxygenated sterols are also provided.A new process is also provided for preparing side chain derivatized 15-oxygenated sterols. This process includes oxidative cleavage of the saturated side chain of the sterol with trifluoroperacetic acid to give a side chain trifluoroacetate and subsequent hydrolysis of this ester. The resultant side chain alcohol is a valuable and advanced intermediate for the preparation of side chain derivatives of 15-oxygenated sterols.
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    Studies of the effect of 3beta-hydroxy-5alpha-cholest-8(14)-en-15-one on microsomal acyl coenzyme A:cholesterol acyltransferase activity
    (1989) Needleman, Dolores Heidi; Schroepfer, George J., Jr.
    3$\beta$-Hydroxy-5$\alpha$-cholest-8(14)-en-15-one (15-ketosterol) is a hypocholesterolemic agent which affects cholesterol metabolism at several levels, including sterol biosynthesis and cholesterol absorption. This dissertation describes studies on the effect of 15-ketosterol on intestinal acyl CoA:cholesterol acyltransferase (ACAT) activity, an enzyme involved in the esterification and absorption of dietary cholesterol. Addition of 15-ketosterol to jejunal microsomes decreased the level of cholesterol esterified by ACAT. Incubation of 15-ketosterol with rat jejunal microsomes reduced the (1 -$\sp{14}$C) oleoyl CoA-dependent esterification of microsomal cholesterol (50% inhibition: 3.0 $\mu$M). This reduction in cholesterol esterification was accompanied by the formation of ($\sp{14}$C) labeled material which, upon analysis by thin layer chromatography, comigrated with 15-ketosteryl oleate. The esterification of 15-ketosterol was confirmed by incubating (2,4- 3H) 15-ketosterol with unlabeled oleoyl CoA and jejunal microsomes. Analyses using either normal phase thin layer chromatography or reverse phase high pressure liquid chromatography detected the formation of (3H) material comigrating with 15-ketosteryl oleate. Oral administration of 15-ketosterol to rats lowered ACAT activity relative to pair-fed controls. ACAT activity in rat jejunal microsomes was lowered 82% (P $<$ 0.02) and 77% (p $<$ 0.001) in animals fed a chow diet containing 0.05% and 0.10% 15-ketosterol, respectively. Analysis of the cholesterol and cholesterol ester content of rat jejunal microsomes showed that oral administration of either 0.10% or 0.125% 15-ketosterol did not affect the concentrations of these compounds. Additional studies showed that reduction of rat jejunal ACAT activity was dependent upon both the duration of oral administration of 15-ketosterol and the concentration of the compound in the diet. Reduction of ACAT activity was observed as early as 3 hours after ingestion of the first meal containing 0.10% 15-ketosterol. Significant reductions in microsomal ACAT activity were observed at concentrations as low as 0.05% 15-ketosterol in the diet. Little or no incorporation of (1-$\sp{14}$C) oleoyl CoA into material comigrating with 15-ketosteryl oleate was observed. In additional studies, incubation of 10 $\mu$M (25R)-3$\beta$,26-dihydroxy-5$\alpha$-cholest-8(14)-en-15-one, a metabolite of 15-ketosterol, with rat jejunal microsomes resulted in a 55% reduction in the oleoyl CoA-dependent esterification of microsomal cholesterol.