Dalcetrapib and anacetrapib differently impact HDL structure and function in rabbits and monkeys
2017; Elsevier BV; Volume: 58; Issue: 7 Linguagem: Inglês
10.1194/jlr.m068940
ISSN1539-7262
AutoresMathieu R. Brodeur, David Rhainds, Daniel Charpentier, Téodora Mihalache-Avram, Mélanie Mecteau, Geneviève Brand, Evelyne Chaput, Anne Perez, Eric J. Niesor, Éric Rhéaume, Cyrille Maugeais, Jean‐Claude Tardif,
Tópico(s)Drug Transport and Resistance Mechanisms
ResumoInhibition of cholesteryl ester transfer protein (CETP) increases HDL cholesterol (HDL-C) levels. However, the circulating CETP level varies and the impact of its inhibition in species with high CETP levels on HDL structure and function remains poorly characterized. This study investigated the effects of dalcetrapib and anacetrapib, the two CETP inhibitors (CETPis) currently being tested in large clinical outcome trials, on HDL particle subclass distribution and cholesterol efflux capacity of serum in rabbits and monkeys. New Zealand White rabbits and vervet monkeys received dalcetrapib and anacetrapib. In rabbits, CETPis increased HDL-C, raised small and large α-migrating HDL, and increased ABCA1-induced cholesterol efflux. In vervet monkeys, although anacetrapib produced similar results, dalcetrapib caused opposite effects because the LDL-C level was increased by 42% and HDL-C decreased by 48% (P < 0.01). The levels of α- and preβ-HDL were reduced by 16% (P < 0.001) and 69% (P < 0.01), resulting in a decrease of the serum cholesterol efflux capacity. CETPis modulate the plasma levels of mature and small HDL in vivo and consequently the cholesterol efflux capacity. The opposite effects of dalcetrapib in different species indicate that its impact on HDL metabolism could vary greatly according to the metabolic environment. Inhibition of cholesteryl ester transfer protein (CETP) increases HDL cholesterol (HDL-C) levels. However, the circulating CETP level varies and the impact of its inhibition in species with high CETP levels on HDL structure and function remains poorly characterized. This study investigated the effects of dalcetrapib and anacetrapib, the two CETP inhibitors (CETPis) currently being tested in large clinical outcome trials, on HDL particle subclass distribution and cholesterol efflux capacity of serum in rabbits and monkeys. New Zealand White rabbits and vervet monkeys received dalcetrapib and anacetrapib. In rabbits, CETPis increased HDL-C, raised small and large α-migrating HDL, and increased ABCA1-induced cholesterol efflux. In vervet monkeys, although anacetrapib produced similar results, dalcetrapib caused opposite effects because the LDL-C level was increased by 42% and HDL-C decreased by 48% (P < 0.01). The levels of α- and preβ-HDL were reduced by 16% (P < 0.001) and 69% (P < 0.01), resulting in a decrease of the serum cholesterol efflux capacity. CETPis modulate the plasma levels of mature and small HDL in vivo and consequently the cholesterol efflux capacity. The opposite effects of dalcetrapib in different species indicate that its impact on HDL metabolism could vary greatly according to the metabolic environment. HDL cholesterol (HDL-C) levels are inversely correlated with cardiovascular disease (1Gordon T. Castelli W.P. Hjortland M.C. Kannel W.B. Dawber T.R. 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Asselin G. Provost S. Barhdadi A. Rhainds D. L'Allier P.L. Pharmacogenomic determinants of the cardiovascular effects of dalcetrapib.Circ Cardiovasc Genet. 2015; 8: 372-382Crossref PubMed Scopus (135) Google Scholar, 15Lindpaintner K. Chasing-and catching-the wild goose: hypothesis-free post-hoc stratification studies as a new paradigm for drug development.Circ Cardiovasc Genet. 2015; 8: 253-255Crossref PubMed Scopus (6) Google Scholar, 16Tardif J.C. Rhainds D. Brodeur M. Feroz Zada Y. Fouodjio R. Provost S. Boule M. Alem S. Gregoire J.C. L'Allier P.L. Genotype-dependent effects of dalcetrapib on cholesterol efflux and inflammation: concordance with clinical outcomes.Circ Cardiovasc Genet. 2016; 9: 340-348Crossref PubMed Scopus (52) Google Scholar) (NCT02525939) are being actively tested in phase 3 clinical outcome trials. In particular, the effects of dalcetrapib on cardiovascular outcome have been shown to vary greatly in different patient populations (14Tardif J.C. Rheaume E. Lemieux Perreault L.P. Gregoire J.C. Feroz Zada Y. Asselin G. Provost S. Barhdadi A. Rhainds D. L'Allier P.L. Pharmacogenomic determinants of the cardiovascular effects of dalcetrapib.Circ Cardiovasc Genet. 2015; 8: 372-382Crossref PubMed Scopus (135) Google Scholar, 16Tardif J.C. Rhainds D. Brodeur M. Feroz Zada Y. Fouodjio R. Provost S. Boule M. Alem S. Gregoire J.C. L'Allier P.L. Genotype-dependent effects of dalcetrapib on cholesterol efflux and inflammation: concordance with clinical outcomes.Circ Cardiovasc Genet. 2016; 9: 340-348Crossref PubMed Scopus (52) Google Scholar). This strengthens the importance of evaluating the impact of dalcetrapib in species that could show differential responses and be helpful in the comprehension of the effects of dalcetrapib in the various human populations. Thus, a better understanding of the in vivo effects of compounds affecting CETP activity on lipoprotein structure and function is warranted. HDL remodeling by CETP has been implicated in the production of small lipid-poor preβ-HDL (17Kunitake S.T. Mendel C.M. Hennessy L.K. Interconversion between apolipoprotein A-I-containing lipoproteins of pre-beta and alpha electrophoretic mobilities.J. Lipid Res. 1992; 33: 1807-1816Abstract Full Text PDF PubMed Google Scholar, 18Rye K.A. Barter P.J. Formation and metabolism of prebeta-migrating, lipid-poor apolipoprotein A-I.Arterioscler. Thromb. Vasc. Biol. 2004; 24: 421-428Crossref PubMed Scopus (262) Google Scholar). These particles are assumed to be produced by CETP-mediated heterotypic transfer of lipids, but are also produced following transfers among HDL subclasses (homotypic transfer) (19Ha Y.C. Gorjatschko L. Barter P.J. Changes in the density distribution of pig high density lipoproteins during incubation in vitro. Influence of esterified cholesterol transfer activity.Atherosclerosis. 1983; 48: 253-263Abstract Full Text PDF PubMed Scopus (16) Google Scholar). 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The in vitro inhibition of CETP activity by torcetrapib and anacetrapib is associated with an inhibition of preβ-HDL production, while dalcetrapib preserves this formation (23Niesor E.J. Magg C. Ogawa N. Okamoto H. von der Mark E. Matile H. Schmid G. Clerc R.G. Chaput E. Blum-Kaelin D. Modulating cholesteryl ester transfer protein activity maintains efficient pre-beta-HDL formation and increases reverse cholesterol transport.J. Lipid Res. 2010; 51: 3443-3454Abstract Full Text Full Text PDF PubMed Scopus (151) Google Scholar). However, in vivo studies demonstrated that anacetrapib increased preβ-HDL levels in hamsters, whereas dalcetrapib had no impact on preβ-HDL levels in hamsters (24Wang S.P. Daniels E. Chen Y. Castro-Perez J. Zhou H. Akinsanya K.O. Previs S.F. Roddy T.P. Johns D.G. In vivo effects of anacetrapib on prebeta HDL: improvement in HDL remodeling without effects on cholesterol absorption.J. Lipid Res. 2013; 54: 2858-2865Abstract Full Text Full Text PDF PubMed Scopus (17) Google Scholar) and humans (25Ray K.K. Ditmarsch M. Kallend D. Niesor E.J. Suchankova G. Upmanyu R. Anzures-Cabrera J. Lehnert V. Pauly-Evers M. Holme I. dal-ACUTE Investigators The effect of cholesteryl ester transfer protein inhibition on lipids, lipoproteins, and markers of HDL function after an acute coronary syndrome: the dal-ACUTE randomized trial.Eur. Heart J. 2014; 35: 1792-1800Crossref PubMed Scopus (71) Google Scholar). Importantly, it was recently demonstrated that administration of anacetrapib to transgenic mice expressing human CETP increased HDL-C and that, in striking contrast, dalcetrapib reduced HDL-C, indicating the importance of studying these CETPis in a model that naturally expresses the CETP. These differences could be generated by the mechanism of action of each CETPi. While anacetrapib generates a nonproductive complex between CETP and HDL, dalcetrapib possesses the unique characteristic of a thioester bond, which is hydrolyzed in its active thiol form. This reactive dalcetrapib-thiol inhibits CETP activity by covalently binding the cysteine 13 of the human protein (9Okamoto H. Yonemori F. Wakitani K. Minowa T. Maeda K. Shinkai H. A cholesteryl ester transfer protein inhibitor attenuates atherosclerosis in rabbits.Nature. 2000; 406: 203-207Crossref PubMed Scopus (509) Google Scholar, 26Maugeais C. Perez A. von der Mark E. Magg C. Pflieger P. Niesor E.J. Evidence for a role of CETP in HDL remodeling and cholesterol efflux: role of cysteine 13 of CETP.Biochim. Biophys. Acta. 2013; 1831: 1644-1650Crossref PubMed Scopus (22) Google Scholar). Interestingly, Morton and Izem (27Morton R.E. Izem L. Cholesteryl ester transfer proteins from different species do not have equivalent activities.J. Lipid Res. 2014; 55: 258-265Abstract Full Text Full Text PDF PubMed Scopus (14) Google Scholar) recently demonstrated that thimerosal, a thiol-reactive compound that covalently binds cysteines, reduces rabbit CETP activity, but increases hamster, monkey, and human recombinant CETP activity in vitro. They also demonstrated that thimerosal more efficiently inhibits the human and monkey CETP activity transfer in the direction of liposomes containing only triglyceride (TG) compared with liposomes containing CE and TG, while there are no differences between the two types of liposomes when hamster CETP is used. These results suggest that drugs targeting CETP could differentially affect HDL-C and preβ-HDL depending on the mechanism of action and the species involved. The present study was designed to determine the in vivo impact of dalcetrapib and anacetrapib on HDL subclass distribution in New Zealand White rabbits and vervet monkeys, two species with a high CETP activity level (28Tsutsumi K. Hagi A. Inoue Y. The relationship between plasma high density lipoprotein cholesterol levels and cholesteryl ester transfer protein activity in six species of healthy experimental animals.Biol. Pharm. Bull. 2001; 24: 579-581Crossref PubMed Scopus (74) Google Scholar) that react in opposite directions to a thiol-reactive compound in vitro. Rabbits and monkeys were exposed to dalcetrapib and anacetrapib and we measured the cholesterol associated to the different lipoprotein classes biochemically and by fast performance LC (FPLC) profiling, the apoA-I distribution among HDL subparticles by nondenaturing gradient gel electrophoresis (NDGGE) and agarose gels, and the ability of apoB-depleted serum to induce cholesterol efflux. We showed that dalcetrapib and anacetrapib substantially increased HDL-C and apoA-I found in large and small HDL subparticles in rabbits, and that these effects were associated with increased cholesterol efflux capacity. In contrast, while similar results were found with anacetrapib in vervet monkeys, dalcetrapib unexpectedly reduced HDL-C levels, apoA-I associated to preβ-HDL, and cholesterol efflux capacities, suggesting that dalcetrapib's effects on CETP are species dependent. The J774 cell line and HepG2 cells were obtained from the ATCC (Manassas, VA). Anacetrapib was bought from Acorn Pharmatech (Redwood City, CA) and dalcetrapib was produced and given by Hoffmann-LaRoche Ltd., Basel, Switzerland. Male New Zealand White rabbits (3.0 kg, aged 12–13 weeks) were first acclimatized for 2 weeks under moderate caloric restriction (∼80% of ad libitum caloric intake) presented as 125 g (or 37.1 g/kg body weight) per day of cholesterol-free diet (32% energy from protein, 13% from fat, and 55% from carbohydrates). Before starting the experimental diet, rabbits were randomized, according to their baseline HDL-C levels, to receive food as above to achieve doses of dalcetrapib of 300 mg/kg body weight (n = 8), or of anacetrapib of 30 mg/kg body weight (n = 7), or food only (n = 7) (supplemental Fig. S1). Food consumption and body weight were recorded throughout the studies to assure adequate drug administration. Blood samples were obtained from an ear vein 1 day before treatment and on day 14 from animals fasted for ≥5 h. Plasma total cholesterol (TC), HDL-C, LDL cholesterol (LDL-C), and TG levels were measured with an automated chemistry analyzer (Dimension RxL Max, Dade Behring, Deerfield, IL). All rabbit experiments were approved by the Animal Care and Use Committee of the Montreal Heart Institute. Seven female African green monkeys (Chlorocebus pygerythrus), also called vervet monkeys, were obtained from a breeding colony and maintained in the primate facility of the South African Medical Research Council (MRC; Cape Town, South Africa) according to the National Code for the Care and Use of Animals for Scientific Purposes and the MRC Ethical Guidelines. The closed indoor environment was maintained at a temperature of 24–26°C, 45% humidity, 15–20 air changes per hour, and a photoperiod of 12 h. Animals were fed a Western-type diet composed of normal human food items with no extra cholesterol added (14% energy from proteins, 47% from lipids, and 39% from carbohydrates; net mean cholesterol intake 23 mg/kg/day), as described in Fincham et al. (29Fincham J.E. Faber M. Weight M.J. Labadarios D. Taljaard J.J. Steytler J.G. Jacobs P. Kritchevsky D. Diets realistic for westernized people significantly effect lipoproteins, calcium, zinc, vitamins C, E, B6 and haematology in vervet monkeys.Atherosclerosis. 1987; 66: 191-203Abstract Full Text PDF PubMed Scopus (25) Google Scholar), for over 3 years, except for one individual (over 2 months), prior to the initiation of treatments. Under that diet, TC was increased by 2.6-fold (from 147 to 376 mg/dl) and this increase was principally associated with LDL-C. First, anacetrapib was administered for two consecutive periods of 1 week, starting at 3 mg/kg once daily, and followed by 10 mg/kg once daily. Solutions containing either 12 or 40 mg/ml of anacetrapib in vehicle (soybean oil) were injected (using a syringe) into the center of the food portion. After a washout period of 4 weeks to allow plasmatic parameters to return to baseline values, dalcetrapib was administered at escalating doses for two consecutive periods of 2 weeks, starting at 30 mg/kg three times a day (total dose 90 mg/kg/day) and followed by 60 mg/kg three times a day (total dose 180 mg/kg/day). An exact amount of compound powder, based on the animal's body weight, was placed into the center of the food portion. Blood samples were collected by femoral venipuncture (after ketamine anesthesia at 5 mg/kg body weight) 3 days before each CETPi treatment and again at the end of each treatment period, 6 h after the intake of the morning portion of food (supplemental Fig. S1). Food consumption and body weight were recorded throughout the studies. HDL-C and LDL-C were determined using standard clinical chemistry methods (Roche Diagnostics). The plasma concentration of compounds was determined using liquid chromatography-tandem mass spectrometry methods that measured the dal-thiol (the active form of dalcetrapib in plasma) (30Heinig K. Bucheli F. Kuhlmann O. Zell M. Pahler A. Zwanziger E. Gross G. Tardio J. Ishikawa T. Yamashita T. Determination of dalcetrapib by liquid chromatography-tandem mass spectrometry.J. Pharm. Biomed. Anal. 2012; 66: 314-324Crossref PubMed Scopus (12) Google Scholar). Plasma or serum obtained from rabbits or monkeys was stored at −80°C and used within 1 year after freezing. Measurement of CETP mass in plasma was performed by ELISA using either mouse monoclonal antibody clone 68/5 or 6/2 as capture antibody for rabbit and monkey, respectively. The detection antibodies were JRC-2 (rabbit) and clone 6/17 (monkey) conjugated to HRP. All of these antibodies were kindly provided by F. Hoffmann-La Roche Ltd. Rabbit CETP activity was determined by an ex vivo CETP activity assay kit (Roar Biomedical, Inc., New York, NY) (26Maugeais C. Perez A. von der Mark E. Magg C. Pflieger P. Niesor E.J. Evidence for a role of CETP in HDL remodeling and cholesterol efflux: role of cysteine 13 of CETP.Biochim. Biophys. Acta. 2013; 1831: 1644-1650Crossref PubMed Scopus (22) Google Scholar). Briefly, the fluorescent assays were performed by incubating 95% (v/v) plasma with the commercial reagent up to a total volume of 105 μl at 37°C for 90 min. CETP activity was evaluated in rabbits and monkeys using radiolabeled CE transfer assays, as previously described (23Niesor E.J. Magg C. Ogawa N. Okamoto H. von der Mark E. Matile H. Schmid G. Clerc R.G. Chaput E. Blum-Kaelin D. Modulating cholesteryl ester transfer protein activity maintains efficient pre-beta-HDL formation and increases reverse cholesterol transport.J. Lipid Res. 2010; 51: 3443-3454Abstract Full Text Full Text PDF PubMed Scopus (151) Google Scholar). One microgram of HDL3 (1.125 < d < 1.210 g/ml) was labeled with [3H]CE and incubated with 65% (v/v) plasma from treated animals in the presence of 1% BSA, 21 mM Tris-HCl (pH 7.4), 0.5% NaCl, and 0.006% EDTA for 4 h (monkey) or 1 h (rabbit) at 37°C. After incubation, the apoB-containing lipoprotein fraction was separated by ultracentrifugation (d = 1.07 g/ml) for 16.5 h at 5°C. Total radioactivity in the apoB-lipoprotein (upper layer) and HDL (lower layer) subfractions were measured by scintillation counting. CETP activity was expressed as the mass (micrograms) of CE transferred to the apoB-lipoprotein fraction per hour. The integrity of HDL3 after labeling with CE was verified and confirmed by NDGGE and by size-exclusion chromatography (FPLC) (supplemental Fig. S2). Rabbit or vervet plasma was separated by size-exclusion chromatography (FPLC) using a Superose-6 10/300 GL column, as described in (31Innis-Whitehouse W. Li X. Brown W.V. Le N.A. An efficient chromatographic system for lipoprotein fractionation using whole plasma.J. Lipid Res. 1998; 39: 679-690Abstract Full Text Full Text PDF PubMed Google Scholar). The cholesterol content of each fraction was measured with a fluorometric assay. The 1D-NDGGE was performed by a modification of the protocol described by Asztalos et al. (32Asztalos B.F. Sloop C.H. Wong L. Roheim P.S. Two-dimensional electrophoresis of plasma lipoproteins: recognition of new apo A-I-containing subpopulations.Biochim. Biophys. Acta. 1993; 1169: 291-300Crossref PubMed Scopus (216) Google Scholar). Briefly, samples were electrophoresed on a 4–30% polyacrylamide gradient gel and run at 150 V for 24 h at 4°C in a buffer containing 90 mM Tris, 80 mM boric acid, and 2.5 mM EDTA (pH 8.3). After electrophoresis, lipoproteins were transferred to a 0.2 μm pore size nitrocellulose membrane (BioRad, Mississauga, Ontario, Canada) at 12 V for 20 h at 4°C in a Tris-glycine buffer without methanol. After transfer, the membrane was blocked and immunodetection was performed with anti-apoA-I-HRP (monoclonal antibody 1/40, provided by F. Hoffmann-La Roche Ltd.) followed by enhanced chemiluminescence detection on Kodak Biomax film. Agarose gel electrophoresis was used to determine the preβ-HDL level in monkey plasma. Briefly, after electrophoresis, apoA-I was detected by Western blotting using a HRP-labeled monoclonal antibody against human apoA-I (kindly provided by Dr. Hugues Matile, Hoffmann-La Roche). The apoA-I band corresponding to preβ-HDL migration was identified by comparison with purified human apoA-I. Lipoproteins were isolated from rabbit plasma treated or not treated with CETPi. Before isolation, the plasma was adjusted to 0.01% EDTA, 0.02% sodium azide, 10 μM PMSF, and 10 μM butylated hydroxytoluene. HDLs with a density of 1.055–1.21 g/ml were prepared by ultracentrifugation. Baby hamster kidney cells expressing a mifepristone-inducible vector with an ABCA1 gene insert (BHK-ABCA1) were a generous gift of Drs. Jack Oram and Chongren Tang (University of Washington, Seattle, WA) (33Vaughan A.M. Oram J.F. ABCA1 redistributes membrane cholesterol independent of apolipoprotein interactions.J. Lipid Res. 2003; 44: 1373-1380Abstract Full Text Full Text PDF PubMed Scopus (177) Google Scholar). BHK-ABCA1 and J774A.1 mouse macrophages were grown in DMEM with phenol red. HepG2 hepatocellular carcinoma cells were growth in Eagle's minimum essential medium (EMEM). The medium was supplemented with 10% FBS, 100 units/ml penicillin, and 100 μg/ml streptomycin. Cells were cultured in 5% CO2 at 37°C and were harvested once a week with trypsin-EDTA. For experiments, cells were trypsinized, seeded, and cultured for 4 days prior to the assays. At confluence, cells were labeled in DMEM (EMEM for HepG2 cells) containing 2 μCi/ml [1,2-3H]cholesterol plus 1% FBS for 24 h at 37°C. For J774 cells, 50 μg/ml of human acetylated LDL were also added during the labeling period to generate foam cells. Then, cells were equilibrated with DMEM (EMEM for HepG2 cells) containing 0.2% BSA for 18 h at 37°C with or without 0.3 mM cAMP (J774) or 20 nM mifepristone (BHK-ABCA1) to induce ABCA1 expression. An efflux assay was performed in the absence or presence of 3% rabbit apoB-depleted serum for 4 h, 0.7% vervet monkey apoB-depleted plasma for 6 h, or 50 μg/ml of HDL for BHK-ABCA1, J774, or HepG2 cells, respectively. The apoB-depleted serum was obtained by adding 0.4 vol of polyethylene glycol solution (20% PEG 6000 in 200 mM glycine buffer) to 1 vol of plasma. Samples were vortex-mixed, incubated for 20 min at 4°C, and centrifuged at 10,000 g for 30 min at 4°C. At the end of the incubation, the medium was harvested and cells were solubilized. Medium and cells were counted for radioactivity in a β-counter. The percentage of efflux was calculated by subtracting the radioactive counts in the medium in the absence of cholesterol acceptors from the radioactive counts in the presence of acceptor and then dividing by the sum of the radioactive counts in the medium plus the cell fraction. Statistical analysis was performed independently by the Montreal Health Innovations Coordinating Center statistical analysis group. Data are shown as mean ± SEM unless stated otherwise. Repeated measures ANCOVA models were used for rabbit data, while vervet monkey analyses were done with two-way repeated measures ANOVA. To evaluate the level of CETP inhibition, we first measured the activity of CETP in the plasma of control and CETPi-treated rabbits. As shown in Fig. 1A, dalcetrapib and anacetrapib both reduced rabbit CETP activity by 42% (P < 0.05). To exclude an impact of endogenous lipoproteins, CETP activity was also measured with a commercial kit based on the fluorescent method. Dalcetrapib and anacetrapib caused reductions of CETP activity of 63% and 71%, respectively (supplemental Fig. S3), indicating that the radioactive assay was not affected by the endogenous lipoprotein levels of the samples. In vervet monkeys, anacetrapib significantly reduced CETP activity by 51 and 50% (P < 0.001) at 10 and 30 mg/kg/day, respectively, while dalcetrapib did not decrease CETP activity (Fig. 1B). We also determined the impact of CETPi on the levels of circulating CETP. CETP mass was not modified in any group of rabbits (Fig. 1C). In vervet monkeys, CETP mass was significantly increased by dalcetrapib only at 90 mg/kg/day (+23%, P < 0.05), while 3 and 10 mg/kg/day of anacetrapib raised it by +74 and +59%, respectively (P < 0.001, Fig. 1D). To confirm the intestinal absorption and systemic exposure to dalcetrapib, pharmacokinetic experiments were conducted in monkeys. Plasma concentrations of the active form of dalcetrapib (dal-thiol) were measured following a single-meal administration. Dal-thiol reached a maximal plasmatic concentration of 1.4 and 5.1 μM, 7 h after single dose administration of 30 and 100 mg/kg, respectively. In rabbits, dal-thiol concentration in plasma was also measured after 14 days of dalcetrapib administration at 300 mg/kg. The concentration of active dal-thiol was 15 μM in animals fasted for ≥5 h. Anacetrapib concentration was also measured in monkeys and the results demonstrated that anacetrapib plasma concentrations reached 1.7 and 2.6 μM after 1 week under administration of 3 and 10 mg/kg of anacetrapib, respectively (data not shown). Thus, it appears that monkeys were exposed to significant concentrations of both CETPis, with differential effects on CETP activity and mass. Next, we evaluated the impact of CETPis on plasma lipid levels biochemically. The results presented in Table 1 demonstrate that HDL-C levels were increased by 229% (P < 0.01) and 171% (P < 0.05) in dalcetrapib- and anacetrapib-treated rabbits, respectively. These higher HDL-C levels were associated with an increase of TC (+86%, P < 0.01; +72%, P < 0.05), whereas both CETPis had
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