Consequences of Cholesteryl Ester Transfer Protein Inhibition in Patients With Familial Hypoalphalipoproteinemia
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动脉硬化血栓血管生物学 2005年第9期
Department of Vascular Medicine (R.J.B., G.K.H., K.E.H., J.H.M.L., J.A.K., J.J.P.K., E.S.G.S) Academic Medical Center, Amsterdam, The Netherlands Department of Medicine (S.T.) University of California, San Diego Biometrics (K.P.), Akros Pharma Inc Princeton, New Jersey Department of Clinical Epidemiology and Biostatistics (A.E.Z.) Academic Medical Center Amsterdam, The Netherlands
To the Editor:
A large proportion of clinical events cannot be prevented during statin therapy, which calls for novel drug targets to further improve cardiovascular outcome. In particular, HDL-increasing strategies hold great promise. The impact of decreased HDL-C on cardiovascular disease (CVD)-related morbidity and mortality has been sharply delineated in individuals affected by familial hypoalphalipoproteinemia (FHA).1 HDL exerts multiple antiatherogenic actions beyond its role in reverse cholesterol transport, comprising antiinflammatory, antioxidative, and direct vascular effects.2 Whereas current strategies to raise HDL-C are limited, novel CETP-inhibitors are capable of mediating significant HDL-C elevation.3,4 Therefore, we evaluated the effects of CETP inhibition on lipid metabolism and markers of oxidation in subjects with FHA.
Subjects were recruited from a Dutch population-based study to identify genes that control HDL-C levels,1 meeting the following criteria: (1) plasma HDL-C level below 10th percentile for age and sex; (2) absence of secondary lipid disorders; and (3) high likelihood of inherited low HDL (defined as HDL-C below 10th percentile in at least one first-degree family member). Nineteen FHA patients (13 men and 6 women; mean±SD age: 42.9±13.9 years), all free of overt macrovascular disease, were enrolled in the study. In 9 of these subjects the underlying defect was defined: heterozygosity for an apolipoprotein A-I (L178P) mutation,1 whereas in the remainder this genetic defect was excluded. The study protocol was approved by the Institutional Review Board at the Academic Medical Center, University Hospital of Amsterdam. All subjects gave written informed consent. This study was designed as a single-center, randomized-sequence, double-blind, crossover study with a 4-week treatment period of JTT-705 600 mg and placebo, respectively.
Lipid parameters were measured by routine methods. Serial ultracentrifugation was used to determine serum HDL subfractions. Lipoprotein profiling was performed by proton nuclear magnetic resonance spectroscopy.5 Fast protein liquid (FPLC) was used to separate VLDL, HDL, and LDL fractions. In individual fractions, total cholesterol (TC), free cholesterol (FC), triglyceride (TG), and phospholipids (PL) concentrations were measured. CETP activity and concentration,6 circulating oxidized LDL (oxLDL) antibodies,7 and serum paraoxonase (PON) activity8 were determined, as described previously. Data are expressed as mean±SD. A paired t test or Wilcoxon Signed rank test was used, depending on distribution of the tested parameter. A probability value of 0.05 was considered significant.
At baseline, HDL-C and apoA-I levels in FHA patients (Table) were below 10th percentile. The number of HDL particles was also decreased compared with reference values derived from the Framingham Offspring study (FOS) (20.7±7.9 μmol/L versus mean reference values of 33±6 μmol/ L5). Whereas LDL-C levels were within the normal range, the number of LDL particles was increased (1912±691 nmol/L; reference values5: men 1535±406, women 1370±427 nmol/L), particularly because of a higher number of very small LDL particles (978±717 nmol/L; reference values5: men 479±391, women 267±261 nmol/L). Plasma triglyceride levels (Table) were within reference ranges. Treatment with JTT-705 was associated with a 24% decrease in CETP activity, with a concomitant 126% increase in CETP mass (Table). JTT-705 increased the levels of HDL-C and apoA-I by 19% and 14%, respectively (Table). The increase in the HDL2 fraction (42%) clearly exceeded the increase in the HDL3 fraction (Table). Using FPLC, JTT-705 was shown to result in an elevation of HDL TC of 24% (P=0.01). NMR analysis showed a significant increase in the HDL particle number from 20.7±7.9 to 23.6±7.2 μmol/L (P=0.008). Particularly within the large HDL subclass, particle number rose from 2.2±1.4 to 3.5±1.9 μmol/L (P=0.004). Total LDL-C did not change during JTT-705 treatment (Table). Using FPLC, JTT-705 was associated with a 165% decrease in TG content in the LDL fraction (P=0.003). LDL subclass analysis with NMR showed that JTT-705 reduced the total number of LDL particles from 1912±691 to 1610±468 nmol/L (P=0.01) with a concomitant reduction in very small LDL particles (from 978±716 to 733±487, P=0.05).
CETP Mass and Activity, Lipoproteins, and Apolipoproteins A-I and B at Baseline and During Placebo and JTT-705 Treatment Periods
OxLDL IgM antibodies showed a modest decrease of 7% (from 8148±3449 to 7293±3601 RLU/100 ms; P=0.001) during JTT-705, compared with baseline. Together with the HDL-C and apoA-I increase, JTT-705 also induced a slight but significant increase in serum PON-1 activity (95.9±91.1 to 105.4±103.7 U/L; P=0.04).
The precise role of CETP activity in the course of atherogenesis has been a matter of debate.9 In the present study, CETP inhibition in FHA individuals is associated with favorable effects on the HDL and LDL subfractions with concomitantly enhanced plasma antioxidant capacity, involving reduced oxLDL-autoantibodies and enhanced serum PON-1 activity. In detail, JTT-705 preferentially increased the number of large HDL particles to a level comparable to that reported in healthy normolipidemic controls in the Framingham Offspring study.5 The clinical relevance of this finding is underscored by observations that particularly larger CE-rich HDL particles show a strong inverse relationship with CVD risk.10 In line with Brousseau,4 we observe an increase in cholesterol content within the HDL particle on JTT-705 treatment, which is likely to contribute to increased plasma residence time of the HDL particle.
With respect to LDL, we find increased levels of atherogenic, very small LDL particles in FHA subjects, in the absence of elevated TG levels. Apparently, in case of very low HDL-C levels, LDL-C takes over as the principle cholesterol ester donor for triglyceride-rich particles. In line, particle concentration of small LDL diminishes significantly on JTT-705 treatment.
Regarding the oxidative status, JTT-705 resulted in a significant reduction of autoantibody levels against oxLDL. Because circulating oxLDL-autoantibodies are thought to reflect LDL oxidation, this may imply that the decreased number of oxidation-prone, very small LDL particles translates into reduced LDL oxidation rates. In parallel, JTT-705 was also associated with an increase in serum PON-1 activity, most likely reflecting delayed HDL apoA-I catabolism,4 thus further enabling HDL to attenuate oxidative modification of the LDL particle.
A limitation of the current study was the presence of a carry-over phenomenon for several parameters of interest, likely because of the lack of a washout between treatment arms. Therefore, we only compared the JTT-705 period to the baseline observations, rather than with the placebo period.
In summary, we provide evidence in patients with familial hypoalphalipoproteinemia that even modest CETP inhibition confers beneficial effects beyond its well recognized HDL-C increasing action, including a reduced number of small LDL particles as well as augmentation of the plasma antioxidant capacity. Because these effects occurred at a JTT-705 dose associated with a modest 24% CETP inhibition, more profound changes can be expected on further optimization of the regimen.3,4 Taken together, these findings suggest an antiatherogenic effect of CETP-inhibition in FHA patients. The results of ongoing trials evaluating the effect of CETP inhibition on hard cardiovascular end points are to be awaited to corroborate the impact of these beneficial changes on cardiovascular outcome.
References
Hovingh GK, Brownlie A, Bisoendial RJ, Dube MP, Levels JH, Petersen W, Dullaart RP, Stroes ES, Zwinderman AH, de Groot E, Hayden MR, Kuivenhoven JA, Kastelein JJ. A novel apoA-I mutation (L178P) leads to endothelial dysfunction, increased arterial wall thickness, and premature coronary artery disease. J Am Coll Cardiol. 2004; 44: 1429–1435.
Bisoendial RJ, Hovingh GK, Levels JH, Lerch PG, Andresen I, Hayden MR, Kastelein JJ, Stroes ES. Restoration of endothelial function by increasing high-density lipoprotein in subjects with isolated low high-density lipoprotein. Circulation. 2003; 107: 2944–2948.
de Grooth GJ, Kuivenhoven JA, Stalenhoef AF, de Graaf J, Zwinderman AH, Posma JL, van Tol A, Kastelein JJ. Efficacy and safety of a novel cholesteryl ester transfer protein inhibitor, JTT-705, in humans: a randomized phase II dose-response study. Circulation. 2002; 105: 2159–2165.
Brousseau ME, Diffenderfer MR, Millar JS, Nartsupha C, Asztalos BF, Welty FK, Wolfe ML, Rudling M, Bjorkhem I, Angelin B, Mancuso JP, Digenio AG, Rader DJ, Schaefer EJ. Effects of cholesteryl ester transfer protein inhibition on high-density lipoprotein subspecies, apolipoprotein A-I Metabolism, and fecal sterol excretion. Arterioscler Thromb Vasc Biol. 2005; 25: 1057–1064.
Freedman DS, Otvos JD, Jeyarajah EJ, Shalaurova I, Cupples LA, Parise H, D’Agostino RB, Wilson PW, Schaefer EJ. Sex and age differences in lipoprotein subclasses measured by nuclear magnetic resonance spectroscopy: the Framingham Study. Clin Chem. 2004; 50: 1189–1200.
Speijer H, Groener JE, van Ramshorst E, van Tol A. Different locations of cholesteryl ester transfer protein and phospholipid transfer protein activities in plasma. Atherosclerosis. 1991; 90: 159–168.
Tsimikas S, Witztum JL, Miller ER, Sasiela WJ, Szarek M, Olsson AG, Schwartz GG. High-dose atorvastatin reduces total plasma levels of oxidized phospholipids and immune complexes present on apolipoprotein B-100 in patients with acute coronary syndromes in the MIRACL trial. Circulation. 2004; 110: 1406–1412.
van Himbergen TM, Roest M, de Graaf J, Jansen EH, Hattori H, Kastelein JJ, Voorbij HA, Stalenhoef AF, van Tits LJ. Indications that paraoxonase-1 contributes to plasma high density lipoprotein levels in familial hypercholesterolemia. J Lipid Res. 2005; 46: 445–451.
Barter PJ, Brewer HB Jr, Chapman MJ, Hennekens CH, Rader DJ, Tall AR. Cholesteryl ester transfer protein: a novel target for raising HDL and inhibiting atherosclerosis. Arterioscler Thromb Vasc Biol. 2003; 23: 160–167.
Asztalos BF, Roheim PS, Milani RL, Lefevre M, McNamara JR, Horvath KV, Schaefer EJ. Distribution of ApoA-I-containing HDL subpopulations in patients with coronary heart disease. Arterioscler Thromb Vasc Biol. 2000; 20: 2670–2676.(Radjesh J. Bisoendial; G.)
To the Editor:
A large proportion of clinical events cannot be prevented during statin therapy, which calls for novel drug targets to further improve cardiovascular outcome. In particular, HDL-increasing strategies hold great promise. The impact of decreased HDL-C on cardiovascular disease (CVD)-related morbidity and mortality has been sharply delineated in individuals affected by familial hypoalphalipoproteinemia (FHA).1 HDL exerts multiple antiatherogenic actions beyond its role in reverse cholesterol transport, comprising antiinflammatory, antioxidative, and direct vascular effects.2 Whereas current strategies to raise HDL-C are limited, novel CETP-inhibitors are capable of mediating significant HDL-C elevation.3,4 Therefore, we evaluated the effects of CETP inhibition on lipid metabolism and markers of oxidation in subjects with FHA.
Subjects were recruited from a Dutch population-based study to identify genes that control HDL-C levels,1 meeting the following criteria: (1) plasma HDL-C level below 10th percentile for age and sex; (2) absence of secondary lipid disorders; and (3) high likelihood of inherited low HDL (defined as HDL-C below 10th percentile in at least one first-degree family member). Nineteen FHA patients (13 men and 6 women; mean±SD age: 42.9±13.9 years), all free of overt macrovascular disease, were enrolled in the study. In 9 of these subjects the underlying defect was defined: heterozygosity for an apolipoprotein A-I (L178P) mutation,1 whereas in the remainder this genetic defect was excluded. The study protocol was approved by the Institutional Review Board at the Academic Medical Center, University Hospital of Amsterdam. All subjects gave written informed consent. This study was designed as a single-center, randomized-sequence, double-blind, crossover study with a 4-week treatment period of JTT-705 600 mg and placebo, respectively.
Lipid parameters were measured by routine methods. Serial ultracentrifugation was used to determine serum HDL subfractions. Lipoprotein profiling was performed by proton nuclear magnetic resonance spectroscopy.5 Fast protein liquid (FPLC) was used to separate VLDL, HDL, and LDL fractions. In individual fractions, total cholesterol (TC), free cholesterol (FC), triglyceride (TG), and phospholipids (PL) concentrations were measured. CETP activity and concentration,6 circulating oxidized LDL (oxLDL) antibodies,7 and serum paraoxonase (PON) activity8 were determined, as described previously. Data are expressed as mean±SD. A paired t test or Wilcoxon Signed rank test was used, depending on distribution of the tested parameter. A probability value of 0.05 was considered significant.
At baseline, HDL-C and apoA-I levels in FHA patients (Table) were below 10th percentile. The number of HDL particles was also decreased compared with reference values derived from the Framingham Offspring study (FOS) (20.7±7.9 μmol/L versus mean reference values of 33±6 μmol/ L5). Whereas LDL-C levels were within the normal range, the number of LDL particles was increased (1912±691 nmol/L; reference values5: men 1535±406, women 1370±427 nmol/L), particularly because of a higher number of very small LDL particles (978±717 nmol/L; reference values5: men 479±391, women 267±261 nmol/L). Plasma triglyceride levels (Table) were within reference ranges. Treatment with JTT-705 was associated with a 24% decrease in CETP activity, with a concomitant 126% increase in CETP mass (Table). JTT-705 increased the levels of HDL-C and apoA-I by 19% and 14%, respectively (Table). The increase in the HDL2 fraction (42%) clearly exceeded the increase in the HDL3 fraction (Table). Using FPLC, JTT-705 was shown to result in an elevation of HDL TC of 24% (P=0.01). NMR analysis showed a significant increase in the HDL particle number from 20.7±7.9 to 23.6±7.2 μmol/L (P=0.008). Particularly within the large HDL subclass, particle number rose from 2.2±1.4 to 3.5±1.9 μmol/L (P=0.004). Total LDL-C did not change during JTT-705 treatment (Table). Using FPLC, JTT-705 was associated with a 165% decrease in TG content in the LDL fraction (P=0.003). LDL subclass analysis with NMR showed that JTT-705 reduced the total number of LDL particles from 1912±691 to 1610±468 nmol/L (P=0.01) with a concomitant reduction in very small LDL particles (from 978±716 to 733±487, P=0.05).
CETP Mass and Activity, Lipoproteins, and Apolipoproteins A-I and B at Baseline and During Placebo and JTT-705 Treatment Periods
OxLDL IgM antibodies showed a modest decrease of 7% (from 8148±3449 to 7293±3601 RLU/100 ms; P=0.001) during JTT-705, compared with baseline. Together with the HDL-C and apoA-I increase, JTT-705 also induced a slight but significant increase in serum PON-1 activity (95.9±91.1 to 105.4±103.7 U/L; P=0.04).
The precise role of CETP activity in the course of atherogenesis has been a matter of debate.9 In the present study, CETP inhibition in FHA individuals is associated with favorable effects on the HDL and LDL subfractions with concomitantly enhanced plasma antioxidant capacity, involving reduced oxLDL-autoantibodies and enhanced serum PON-1 activity. In detail, JTT-705 preferentially increased the number of large HDL particles to a level comparable to that reported in healthy normolipidemic controls in the Framingham Offspring study.5 The clinical relevance of this finding is underscored by observations that particularly larger CE-rich HDL particles show a strong inverse relationship with CVD risk.10 In line with Brousseau,4 we observe an increase in cholesterol content within the HDL particle on JTT-705 treatment, which is likely to contribute to increased plasma residence time of the HDL particle.
With respect to LDL, we find increased levels of atherogenic, very small LDL particles in FHA subjects, in the absence of elevated TG levels. Apparently, in case of very low HDL-C levels, LDL-C takes over as the principle cholesterol ester donor for triglyceride-rich particles. In line, particle concentration of small LDL diminishes significantly on JTT-705 treatment.
Regarding the oxidative status, JTT-705 resulted in a significant reduction of autoantibody levels against oxLDL. Because circulating oxLDL-autoantibodies are thought to reflect LDL oxidation, this may imply that the decreased number of oxidation-prone, very small LDL particles translates into reduced LDL oxidation rates. In parallel, JTT-705 was also associated with an increase in serum PON-1 activity, most likely reflecting delayed HDL apoA-I catabolism,4 thus further enabling HDL to attenuate oxidative modification of the LDL particle.
A limitation of the current study was the presence of a carry-over phenomenon for several parameters of interest, likely because of the lack of a washout between treatment arms. Therefore, we only compared the JTT-705 period to the baseline observations, rather than with the placebo period.
In summary, we provide evidence in patients with familial hypoalphalipoproteinemia that even modest CETP inhibition confers beneficial effects beyond its well recognized HDL-C increasing action, including a reduced number of small LDL particles as well as augmentation of the plasma antioxidant capacity. Because these effects occurred at a JTT-705 dose associated with a modest 24% CETP inhibition, more profound changes can be expected on further optimization of the regimen.3,4 Taken together, these findings suggest an antiatherogenic effect of CETP-inhibition in FHA patients. The results of ongoing trials evaluating the effect of CETP inhibition on hard cardiovascular end points are to be awaited to corroborate the impact of these beneficial changes on cardiovascular outcome.
References
Hovingh GK, Brownlie A, Bisoendial RJ, Dube MP, Levels JH, Petersen W, Dullaart RP, Stroes ES, Zwinderman AH, de Groot E, Hayden MR, Kuivenhoven JA, Kastelein JJ. A novel apoA-I mutation (L178P) leads to endothelial dysfunction, increased arterial wall thickness, and premature coronary artery disease. J Am Coll Cardiol. 2004; 44: 1429–1435.
Bisoendial RJ, Hovingh GK, Levels JH, Lerch PG, Andresen I, Hayden MR, Kastelein JJ, Stroes ES. Restoration of endothelial function by increasing high-density lipoprotein in subjects with isolated low high-density lipoprotein. Circulation. 2003; 107: 2944–2948.
de Grooth GJ, Kuivenhoven JA, Stalenhoef AF, de Graaf J, Zwinderman AH, Posma JL, van Tol A, Kastelein JJ. Efficacy and safety of a novel cholesteryl ester transfer protein inhibitor, JTT-705, in humans: a randomized phase II dose-response study. Circulation. 2002; 105: 2159–2165.
Brousseau ME, Diffenderfer MR, Millar JS, Nartsupha C, Asztalos BF, Welty FK, Wolfe ML, Rudling M, Bjorkhem I, Angelin B, Mancuso JP, Digenio AG, Rader DJ, Schaefer EJ. Effects of cholesteryl ester transfer protein inhibition on high-density lipoprotein subspecies, apolipoprotein A-I Metabolism, and fecal sterol excretion. Arterioscler Thromb Vasc Biol. 2005; 25: 1057–1064.
Freedman DS, Otvos JD, Jeyarajah EJ, Shalaurova I, Cupples LA, Parise H, D’Agostino RB, Wilson PW, Schaefer EJ. Sex and age differences in lipoprotein subclasses measured by nuclear magnetic resonance spectroscopy: the Framingham Study. Clin Chem. 2004; 50: 1189–1200.
Speijer H, Groener JE, van Ramshorst E, van Tol A. Different locations of cholesteryl ester transfer protein and phospholipid transfer protein activities in plasma. Atherosclerosis. 1991; 90: 159–168.
Tsimikas S, Witztum JL, Miller ER, Sasiela WJ, Szarek M, Olsson AG, Schwartz GG. High-dose atorvastatin reduces total plasma levels of oxidized phospholipids and immune complexes present on apolipoprotein B-100 in patients with acute coronary syndromes in the MIRACL trial. Circulation. 2004; 110: 1406–1412.
van Himbergen TM, Roest M, de Graaf J, Jansen EH, Hattori H, Kastelein JJ, Voorbij HA, Stalenhoef AF, van Tits LJ. Indications that paraoxonase-1 contributes to plasma high density lipoprotein levels in familial hypercholesterolemia. J Lipid Res. 2005; 46: 445–451.
Barter PJ, Brewer HB Jr, Chapman MJ, Hennekens CH, Rader DJ, Tall AR. Cholesteryl ester transfer protein: a novel target for raising HDL and inhibiting atherosclerosis. Arterioscler Thromb Vasc Biol. 2003; 23: 160–167.
Asztalos BF, Roheim PS, Milani RL, Lefevre M, McNamara JR, Horvath KV, Schaefer EJ. Distribution of ApoA-I-containing HDL subpopulations in patients with coronary heart disease. Arterioscler Thromb Vasc Biol. 2000; 20: 2670–2676.(Radjesh J. Bisoendial; G.)