Inhibition of 3-Hydroxy-3-Methylglutaryl-Coa
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Proc. Natl. Acad. Sci. USA Vol. 81, pp. 2538-2542, April 1984 Medical Sciences Inhibition of 3-hydroxy-3-methylglutaryl-CoA reductase by mevinolin in familial hypercholesterolemia heterozygotes: Effects on cholesterol balance (cholesterol balance/low-density lipoprotein receptors/fecal neutral and acidic steroids) SCOTT M. GRUNDY AND DAVID W. BILHEIMER Department of Internal Medicine and Center for Human Nutrition, University of Texas Health Science Center at Dallas, 5323 Harry Hines Boulevard, Dallas, TX 75235 Communicated by Michael S. Brown, January 9, 1984 ABSTRACT Patients with heterozygous familial hyper- through endocytosis mediated by a specific LDL receptor cholesterolemia (FH) have a deficiency of receptors for plasma (8). low-density lipoprotein (LDL) that impairs removal of LDL When human fibroblasts or cultured liver cells are incu- from plasma. In these patients, mevinolin, an inhibitor of 3- bated with compactin or mevinolin, HMG-CoA reductase is hydroxy-3-methylglutaryl-CoA reductase [mevalonate:NAD' competitively inhibited, and the de novo synthesis of choles- oxidoreductase (CoA-acylating), EC 1.1.1.88], increases re- terol is blocked (9, 10). This block triggers a dual regulatory ceptors for LDL and decreases LDL concentrations. To deter- response that seems designed to provide more cholesterol mine whether mevinolin also causes severe decreases in total for the cell: there is a simultaneous increase in HMG-CoA body synthesis of cholesterol, fecal excretions of neutral ste- reductase and in the number of LDL receptors (9, 11, 12). roids and acidic steroids were determined in five FH heterozy- In dogs, mevinolin has been shown to decrease plasma gotes before and during treatment with mevinolin. The drug LDL by causing an increase in hepatic receptors for LDL produced an average decrease in plasma total cholesterol of (2); the same response probably occurs in humans (7). Me- 23% and in LDL cholesterol of 24%. Mevinolin caused a sig- vinolin thus seems to have the same effect in vivo as in cul- nificant decrease in the output of neutral and acidic steroids in tured cells; i.e., it inhibits HMG-CoA reductase and triggers three patients, but it caused no alterations in two others. a regulatory response that increases LDL receptors and Changes in fecal output of steroids did not correlate with the probably also increases the amount of HMG-CoA reductase. degree of lowering of the patients' LDL-cholesterol level. In The question that arises is whether HMG-CoA reductase can none of the patients did the output of fecal steroids fall below increase sufficiently to return cholesterol synthesis to nor- the values seen in normal subjects studied under similar condi- mal or whether cholesterol synthesis must remain depressed tions. One patient had a previous ileal exclusion operation and for mevinolin to maintain an increase in LDL receptors. In had a massive output of acidic steroids in the control period; the current study, we have attempted to distinguish between mevinolin therapy caused a slight decrease in excretion of acid- these two possibilities in humans by use of the cholesterol- ic steroids, but the output was still markedly above normal. balance technique. We conclude that the LDL lowering action of mevinolin does Normal humans excrete more steroids in the feces than not appear to require a severe decrease in cholesterol synthesis can be accounted for by the cholesterol that they ingest. that might lead to depletion of vital body stores of cholesterol. These steroids emerge as neutral steroids and bile acids. Since the steroid nucleus is not degraded in the body, and An exciting class of drugs for treatment of hypercholester- since the feces are the predominant route of steroid excre- olemia consists of fungal metabolites that are competitive tion, the net fecal steroid excretion (cholesterol intake minus inhibitors of 3-hydroxy-3-methylglutaryl-CoA reductase steroid output) in the steady state approximates the total [HMG-CoA reductase; mevalonate:NAD' oxidoreductase amount of cholesterol synthesized in the body (13). (CoA-acylating), EC 1.1.1.88], a rate-controlling enzyme in In the current investigation we treated five heterozygotes cholesterol synthesis. The prototype compound, compactin, for familial hypercholesterolemia with mevinolin and mea- and its analogue, mevinolin, decrease the levels of low-den- sured the net excretion of fecal steroids. This cholesterol sity lipoprotein (LDL) in normal animals (1, 2), normal hu- balance study was conducted as part of another study in mans (3), and in heterozygotes with familial hypercholester- which turnover of LDL was measured (7). The results indi- olemia (FH) (4-7). Recently, we demonstrated that mevino- cate that mevinolin can cause a moderate decrease in total- lin achieves this effect by increasing the fractional rate of body synthesis of cholesterol, but a marked overall decrease removal of LDL from the circulation of FH heterozygotes in synthesis seemingly is not necessary for mevinolin to en- (7). The higher clearance rate appeared to be due to en- hance the activity of LDL receptors. hanced receptor-mediated catabolism of LDL. A theoretical basis for the ability of mevinolin to stimulate METHODS receptor-mediated catabolism of LDL comes from studies of Patients. Five patients with heterozygous FH were studied human and animal cells in tissue culture. These cells have a on the General Clinical Research Center at Parkland Memo- dual source of the cholesterol required for synthesis of new rial Hospital or in the Metabolic Unit at the Veterans Admin- membranes. Cells can synthesize cholesterol de novo in a istration Medical Center (Dallas, TX). Two patients (M.B. pathway that requires the action of HMG-CoA reductase; and J.P.) were men of ages 36 and 37 yr, respectively; J.P. or, the cells can obtain cholesterol from plasma LDL Abbreviations: FH, familial hypercholesterolemia; HMG-CoA re- The publication costs of this article were defrayed in part by page charge ductase, 3-hydroxy-3-methylglutaryl-CoA reductase; LDL, low- payment. This article must therefore be hereby marked "advertisement" density lipoprotein; VLDL, very-low-density lipoproteins; HDL, in accordance with 18 U.S.C. §1734 solely to indicate this fact. high-density lipoproteins. 2538 Downloaded by guest on September 23, 2021 Medical Sciences: Grundy and Bilheimer Proc. Natl. Acad. Sci. USA 81 (1984) 2539 Table 1. Plasma lipids and lipoproteins Plasma cholesterol,* mg/dl SD Plasma Patient Period Total LDL HDL triglyceride* M.B. Control 340 ± 32 272 ± 24 24 ± 1 208 ± 53 Mevinolin 285 ± 20 239 ± 16 22 ± 6 77 ± 15 J.P. Control 267 ± 17 207 ± 18 24 ± 3 221 ± 21 Mevinolin 226 ± 20 156 ± 4 26 ± 3 205 ± 39 J.C. Control 320 ± 6 281 ± 9 19 ± 2 103 ± 12 Mevinolin 233 ± 9 201 ± 11 20 ± 6 105 ± 10 M.Ba. Control 306 ± 11 254 ± 6 26 ± 3 118 ± 14 Mevinolin 247 ± 12 190 ± 11 29 ± 2 144 ± 8 C.C. Control 319 ± 21 228 ± 18 18 ± 4 311 ± 42 Mevinolin 211 ± 9 153 ± 13 24 ± 5 178 ± 47 Mean + SEM Control 310 ± 12 248 ± 13 22 ± 2 192 ± 38 Mevinolin 240 ± 13 188 ± 16 24 ± 2 142 ± 23 P valuet < 0.005 < 0.002 NS NS NS, not significant. *Mean ± SD of six or seven observations during the LDL turnover study in control period and with mevinolin treatment. tP values were calculated by paired t test. had severe coronary heart disease, but M.B. had no clinical Medical Center. Informed consent was obtained from each evidence of atherosclerosis. The remaining three patients patient. (C.C., J.C., and M.Ba.) were women of ages 52, 32, and 55 Lipid and Lipoprotein Analysis. In each patient, the kinet- yr, respectively. C.C. had coronary heart disease, but J.C. ics of LDL turnover were measured in each study period, and M.Ba. were clinically free of ischemic heart disease. and the results have been reported separately (7). These J.C. is the daughter of M.Ba.; M.B. and C.C. are first cous- turnover studies were initiated about 3 weeks into each peri- ins. od. During the kinetic study, which lasted 20 days, blood Diets. The patients were fed a diet of solid food and liquid samples were obtained daily, and total cholesterol was mea- formula containing 40% of calories as fat, 40% as carbohy- sured on each sample. In this period, samples were taken 6 drate, and 20% as protein (7). The major fat of the diet was or 7 times for estimation of triglycerides, LDL cholesterol, lard, and the polyunsaturated/saturated fat ratio was 0.33. and high-density lipoprotein (HDL) cholesterol. Each sam- Cholesterol intake varied between 170 and 280 mg per day. ple was subjected to ultracentrifugation at density <1.006 Weights were checked daily and caloric intake was adjusted g/ml for isolation of very-low-density lipoprotein (VLDL). to maintain a constant weight throughout the study. Cholesterol was measured on the infranatant both before and Drug Therapy. Capsules containing mevinolin and placebo after precipitation of LDL with heparin-manganese accord- were supplied by Merck Sharp & Dohme (courtesy of J. To- ing to Lipid Research Clinic procedures (14). Cholesterol bert). Mevinolin was given in doses of 20 mg twice daily. and triglyceride concentrations were determined using a Two patients (J.C. and M.B.) received mevinolin first and Boehringer Mannheim enzyme kit. then placebo; the other three (J.P., C.C., and M.Ba.) were Sterol Balance. Cholesterol balance was performed ac- given placebo before mevinolin. The protocol was approved cording to described methods (15, 18). Measurements were by the U.S. Food and Drug Administration and by the Hu- not made in one patient (T.P.), who had been studied previ- man Research Review Boards of the University of Texas ously (7). Serial stool collections were obtained and com- Health Science Center and the Veterans Administrations bined into 4-day pools during each study period (mevinolin Table 2.