The Potential for CETP Inhibition to Reduce Cardiovascular Disease Risk

By Ansell, Benjamin; Hobbs, F D Richard

Key words: CETi-I – Investigational drugs – JTT-705 – Torcetrapib

ABSTRACT

Background: Although reductions In cardiovascular risk can be achieved by lowering low-density lipoprotein cholesterol, treated patients remain at substantial risk. Epidemiological studies have established that higher levels of high-density lipoprotein cholesterol (HDL-C) are strongly associated with reduced cardiovascular risk, and therefore raising levels of HDL-C may be beneficial. The activity of cholesteryl ester transfer protein (CETP) appears to be Inversely correlated with HDL-C levels and thus CETP is an attractive target for intervention to raise levels of HDL- C and potentially reduce residual cardiovascular risk.

Objectives: This paper reviews the evidence for an atheroprotective role of higher levels of HDL-C, the function of CETP in cholesterol metabolism, and the concept of CETP inhibition as a potential new strategy for decreasing cardiovascular risk. An analysis of clinical studies of CETP inhibition was also performed.

Methods: MEDLINE (1966 to June 2006), EMBASE (1974 to June 2006), and cardiology conference proceedings were searched for clinical trials of CETP Inhibition.

Results: Thirteen reports Involving vaccinebased and pharmacological Inhibition of CETP were found. Modest and inconsistent elevation of HDL-C was observed with vaccine-based therapy, whereas HDL-C elevation with pharmacological Inhibitors was greater and more consistent.

Conclusions: Elevation of HDL-C via CETP inhibition appears to be a potentially promising approach to reduce cardiovascular disease. Preliminary studies suggest benefits of CETP inhibition on serum llpid levels, and ongoing studies should establish the effects on atherosclerosis and cardiovascular events.

Introduction

Cardiovascular disease (CVD) remains the leading cause of mortality in Western populations1’2. A large body of evidence has established that lowering circulating levels of low-density lipoprotein cholesterol (LDL-C) reduces risk for CVD. Thus, all national and international guidelines regarding cholesterol levels for the prevention and treatment of CVD currently focus on lowering levels of LDL-C2,3. Statins are the therapy of choice for lowering LDL-C and have been shown to reduce cardiovascular event rates by up to approximately 40% in patients with, or at risk for, coronary heart disease (CHD)3-16. However, despite this huge treatment effect, the majority of events are not prevented with statin therapy, and there remains the potential to achieve further risk reductions. Even with the most intensive statin regimens, cardiovascular events are not entirely eliminated; in the Pravastatin or Atorvastatin Evaluation and Infection Therapy (PROVE IT) study, 22.4% of patients in the high-dose atorvastatin 80mg/day group experienced a major cardiovascular event or died during 2 years of followup4, and in the Treating to New Targets (TNT) study, nearly 9% of patients receiving high-dose atorvastatin SOmg/day experienced a major cardiovascular event during 5 years of follow- up13. Hence, there is a need for additional strategies that may complement statin therapy and reduce the residual risk for CVD.

Epidemiological studies have indicated that elevated levels of high-density lipoprotein cholesterol (HDL-C) are strongly and independently associated with a decreased risk for CHD17-20. Raising levels of HDL-C may therefore be an additional therapeutic option for managing CVD. However, aside from data from a limited number of studies (including the Veterans Affairs HDL Intervention Trial [VA- HIT]21, the Helsinki Heart Study22, and the Coronary Drug Project23), evidence from clinical trials that therapeutic elevation of HDL-C reduces cardiovascular events is limited. This may be because current therapies are only moderately effective, as with fibrates (HDL-C increases [asymptotically =] 10-20%), or are more effective but poorly tolerated, as with niacin (HDL-C increases [asymptotically =] 15-35%)24. Thus, novel therapies that will effectively elevate HDL-C levels may help to strengthen the rationale for targeting HDL-C in the management of CVD.

One mechanism for raising HDL-C currently under investigation is the inhibition of cholesteryl ester transfer protein (CETP). CETP plays a crucial role in cholesterol metabolism, and congenital CETP deficiency has been shown to be associated with large increases in HDL-C but varying risks of atherosclerosis25.

In this review, we reassess the relationship of HDL-C with cardiovascular risk and examine the role of CETP in cholesterol metabolism, atherosclerosis, and clinical outcomes. We also report the findings from searches of MEDLINE (1966 to present), EMBASE (1974 to present), and proceedings of cardiology conferences with search terms ‘cholesteryl ester transfer protein’ and ‘inhibition1 or ‘inhibitor’ for clinical trials of CETP inhibition and appraise the data to investigate the effectiveness of CETP inhibition as a means of raising HDL-C levels and as a potential new strategy for the prevention and treatment of CVD.

Figure 1. Formation of high-density lipoprotein (HDL) particles. Lipid-poor apolipoprotein A-I (A-I) acquires phospholipids and free cholesterol (FC) from peripheral tissues to form nascent, discoidal HDL (also known as pre β-HDL). Lecithin-cholesterol acyltransferase (LCAT) converts FC in nascent, discoidal HDL to cholesteryl ester (CE), resulting in the formation of the mature, spherical HDL particle (also known as a-HDL). In addition to large quantities of CE, the core of mature HDL particles also contains small amounts of triglyceride (TC)

Relationship between HDL cholesterol and CHD risk

Of the various risk factors examined in the long-running Framingham study, HDL-C was found to be the strongest predictor of CHD risk17. In a 12-year follow-up of the Framingham study population, participants with high HDL-C (80th percentile) had half the risk of developing CHD compared with those with low HDL-C (20th percentile)18. Similarly, in the Prospective Cardiovascular Munster (PROCAM) study, CHD risk was four times lower in patients whose HDL- C levels were ≥ 35 mg/dL (0.9 mmol/L) compared with patients whose HDL-C levels were < 35 mg/dL (0.9mmol/L)19. A combined analysis of four large studies found that each 1 mg/dL (0.026 mmol/ L) increase in HDL-C was associated with a 2-3% decrease in CHD risk26.

Understanding the atheroprotective nature of HDL cholesterol

In common with all lipoproteins, HDL particles contain a core of triglycerides and cholesteryl esters (CEs) and an outer layer of phospholipids and apolipoproteins (Figure 1)27,28. The major protein of HDL is apolipoprotein (apo) A-I, although other apolipoproteins may be present, and additional enzymes, such as lecithincholesterol acyltransferase (LCAT) and paraoxonase are also associated with HDL. The remaining mass of HDL is composed of phospholipids, free and esterified cholesterol, triglycerides, and other minor components such as liposoluble vitamins and antioxidants.

A broad spectrum of effects is now known to contribute to the atheroprotective nature of HDL-C (Figure 2). First, HDL particles appear to alleviate endothelial dysfunction29-31, an early sign of atherosclerosis that is characterized by reduced nitric oxide (NO)- dependent vasodilation in response to physiological stimuli32. Second, HDL particles and their components normally possess potent antioxidant activity33’34, which may protect against oxidative damage that contributes to initiation and progression of atherosclerosis35. Third, HDL particles have antiinflammatory properties, including inhibition of adhesion/transmigration of infiltrating leukocytes36-39 and down-regulation of proinflammatory factors40 that characterize the atherosclerotic plaque41. However, the role of HDL in inflammation is likely to be complex, as the acute phase reaction and/or some chronic diseases may result in conversion of HDL to a proinflammatory state42. Fourth, HDL exerts antithrombotic effects, including inhibition of erythrocyte coagulation43, promotion of prostacyclin synthesis (a platelet aggregation inhibitor)44, and inhibition of thrombin-induced expression of endothelial tissue factor45. Fifth, HDL plays a role in the innate immune system and can aid in combating infection by lysing parasites46 and inhibiting the pyrogenic effects of bacterial lipopolysaccharide47. Finally, HDL’s most pivotal function may be to facilitate reverse cholesterol transport, the process by which cholesterol is transported from peripheral cells to the liver for excretion. This is discussed in greater detail below.

Figure 2. The properties of atheroprotective high-density lipoprotein (HDL) particles include stimulation of reverse cholesterol transport and improvement of endothelial function, as well as inhibition of inflammation, oxidation, thrombosis, and infection. Under inflammatory conditions, HDL can reverse its usually protective functions and paradoxically display pro- inflammatory features itself40.

Cholesterol metabolism: the roles of LDL, HDL, and CETP

The key pathways of cholesterol metabolism and the key molecules involved are summarized in Figure 3.

Role of VLDL/LDL

Very low-density lipoproteins (VLDL) are triglyceriderich particles secreted by the liver whose major apolipoprotein is apo B- 100. Triglycerides in VLDL may be tra\nsferred to muscle and adipose tissue, exchanged for CE from HDL particles via CETP activity, or hydrolyzed by lipoprotein lipase and hepatic lipase, resulting in formation of LDL particles containing apo B-100 (Figure 3). Apo B- 100 facilitates hepatic uptake of LDL (indirect reverse cholesterol transport; Figure 3, pathway 1), although oxidative modification of LDL promotes the preferential uptake by macrophages leading to cholesterol accumulation in the arterial wall.

Role of HDL

As shown in Figure 1 and Figure 3, nascent, discoidal HDL (pre β-HDL) is formed when lipid-poor apo A-I acquires free cholesterol from peripheral tissues via the ABCA1 receptor. This is the first step in reverse cholesterol transport. LCAT esterifies the free cholesterol in nascent, discoidal HDL, resulting in the formation of CE and maturation of the HDL particle (α-HDL). Mature HDL particles transfer cholesterol to the liver via the scavenger receptor Bl (SR-B1), completing reverse cholesterol transport by the direct route (Figure 3, pathway 2).

More recently, an alternative cholesterol efflux pathway via the ATP binding cassette transporters Gl and G4 (ABCG1/4) has been identified. ABCG1/4 transporters expressed on the surface of macrophages in the arterial wall mediate the efflux of free cholesterol directly to mature HDL, thereby providing an alternate pathway for reverse cholesterol transport48,49. In addition, SR-Bl receptors on macrophages in the arterial wall can facilitate efflux of free cholesterol to mature HDL50,51.

Figure 3. Cholesterol metabolism, reverse cholesterol transport, and the role of cholesteryl ester transfer protein (CETP). A-I = apolipoprotein A-I; ABCA1/G1/G4 = ATP-binding cassette A1/G1/G4; B = apolipoprotein B-IOO; CE = cholesteryl ester; FC = free cholesterol; HDL = high-density lipoprotein; LCAT = lecithin-cholesterol acyltransferase; LDL = lowdensity lipoprotein; LDL-R = LDL receptor; LPL = lipoprotein lipase; SR-B1 = scavenger receptor B1 ; TG = triglyceride; VLDL = very low-density lipoprotein

Role of CETP

CETP, secreted by the liver, is mostly bound to HDL and promotes exchange of CE and triglycerides between plasma lipoproteins according to their concentration gradients52. Since VLDL particles are relatively rich in triglycerides and HDL particles are rich in CE, the net effect of CETP is the transfer of CE from HDL to VLDL and LDL particles, and of triglycerides from VLDL to HDL and LDL particles (Figure 4).

In cholesterol metabolism, CETP serves to link the direct and indirect reverse cholesterol transport pathways (Figure 3) by mediating exchange of CE contained within HDL particles for triglycerides from LDL and VLDL particles. As such, high CETP activity may be associated with both anti- and pro-atherogenic effects. Direct reverse cholesterol transport via HDL particles is decreased, the pool of HDL is reduced, and the pool of LDL-C increased (pro-atherogenic). On the other hand, indirect reverse cholesterol transport via LDL particles is increased (anti- atherogenic). Furthermore, by exchanging triglycerides in VLDL for CE in LDL, CETP promotes the formation of triglycerideenriched LDL. This may make LDL more susceptible to endothelium-bound hepatic lipase, resulting in the formation of small dense LDL53 (Figure 4). Small dense LDL is more atherogenic than large LDL54, perhaps because it is more readily oxidized and taken up by macrophages in the arterial wall. In a similar manner, CETP promotes the formation of small dense HDL (Figure 4)55. While small dense HDL may serve as a source of lipid-poor apo A-I for further cholesterol efflux, it is more susceptible to excretion via the kidneys, which may reduce plasma concentrations of HDL-C.

The relationship between CETP and CVD: evidence from studies

As described above, greater understanding of the role that CETP plays in cholesterol metabolism has highlighted several possible mechanisms through which CETP activity may contribute to the development of atherosclerosis and CVD. However, while there are much data from scientific studies to support the concept that high CETP activity is associated with elevated CVD risk52,56, the body of published literature also provides contradictory findings, and the precise relationship between CETP activity and CVD risk remains to be elucidated.

Figure 4. The role of cholesteryl ester transfer protein (CETP) and change in lipoprotein composition and size with CETP activity. CE = cholesteryl ester; HDL = high-density lipoprotein cholesterol; LDL = low-density lipoprotein; TG = triglyceride; VLDL = very low- density lipoprotein

The notion that CETP may be a potential target for reducing CVD risk originated from studies of apparently healthy individuals in Japanese populations that lacked a functional copy of the CETP gene25,57. Compared with unaffected individuals, those who were CETP deficient and who had no measurable CETP activity exhibited substantially elevated HDL-C (+209%) and decreased LDL-C (-44%). These individuals were apparently healthy and, not surprisingly, given the lipid profile, did not show signs of premature atherosclerosis25. These observational studies of CETP-deficient subjects helped establish a link between reduced CETP activity and elevated HDL-C levels, yet were unable to demonstrate a clear link between reduced CETP activity and decreased CVD risk. Indeed, analysis of data from Japanese-American men followed in the Honolulu Heart Program initially showed a 50% increase in CHD among participants with CETP deficiency and HDL-C levels of 41-60mg/dL (1.1-1.6mmol/L)58. Nevertheless, a prospective 7-year follow-up of the Honolulu Heart Program did not confirm the initial observations, instead noting that individuals with a CETP mutation showed a statistically insignificant trend toward fewer CHD events than those without a mutation59.

Apparent contradictions have also been reported regarding the cardiovascular risk or longevity associated with single-nucleotide polymorphisms (SNPs) of the CETP gene, reflecting such limitations as the subtlety of differences in CETP activity conferred by specific polymorphisms, the relatively small numbers of evaluated individuals, and the presence of confounding factors52,56. Data based on analysis of the best characterized fSNP, Taq1B (in the first intron of the CETP gene), have tended to support the association of high circulating levels of CETP with lower levels of HDL-C and increased cardiovascular risk. Subjects homozygous for the B1 allele (B1/B1) have elevated levels of CETP and lower levels of HDL-C compared with heterozygotes (B1/B2) or homozygous B2/B2 subjects60-63. For example, in a case-control sub-study of the West of Scotland Coronary Prevention Study (WOSCOPS) involving 498 men who experienced cardiovascular events (cases) and 1108 men who did not (controls) over 5 years, subjects with homozygous B1 alleles had lower levels of HDL-C compared with B2 homozygous subjects (p < 0.001), and heterozygous subjects had intermediate levels63. Homozygosity for the B2 allele was associated with a 30% risk reduction for cardiovascular events (p = 0.03). In the Framingham Offspring study, a similar gradient of HDL levels among Bl homozygous, B1/B2 heterozygous, and B2 homozygous subjects was observed among 2916 men and women60. Presence of the B2 allele was significantly associated with decreased CETP activity (p < 0.05), and in men (but not women) it was similarly associated with a lower risk of CHD (p = 0.035). Evidence from the Regression Growth Evaluation Statin Study (REGRESS) also supports the association of the Taq 1B1 allele and elevated CETP concentrations with increased atherosclerotic progression64. In a sub-analysis of 807 men with coronary atherosclerosis followed for 2 years, the presence of the Bl allele was associated once again with increased CETP concentrations and decreased HDL-C levels (p < 0.001 for each). In patients randomized to placebo, a significant association was observed between the Bl allele and decreases in the luminal diameter assessed with angiography (p 0.05 for all measures).

A recent meta-analysis of common CETP polymorphisms draws the general conclusion that, when reduced CETP activity is associated with elevated HDL-C levels, there is more likely to be an associated protective effect for CVD65. One example of the beneficial effects that may be conferred by a CETP gene polymorphism is the study published by Barzilai et /.66 of exceptionally long-lived individuals from a genetically homogeneous population of Ashkenazi Jews. Healthy individuals aged 95-107 years were observed to have a unique lipoprotein profile consisting of large HDL and LDL particles. Their offspring also had lipoprotein particles of larger size when compared with an age-matched control group of Ashkenazi Jews and with individuals from the Framingham Offspring Study. In both the individuals with enhanced longevity and their offspring, the prevalence of homozygosity for a CETP polymorphism (I405V) was far greater than in either of the control groups. The 1405 V polymorphism correlated with reduced CETP levels, CETP activity, and larger lipoprotein particle size, and this was believed to be the reason for the survival advantage observed in the long-lived individuals.

Figure 5. Carotid intima-media thickness after 2 years according to the cholesteryl ester transfer protein (CETP) concentration at baseline in the Atorvastatin versus Simvastatin on Atherosclerotic Progression (ASAP) study. Reprinted from de Crooth et al.67 – copyright 2004 with permission from Elsevier

The relationship of CETP with cardiovascular risk factors and responses to statin treatment were examined in a sub-analysis of the Atorvastatin versus Simvastatin on Atherosclerotic Progression (ASAP) study in patients with familial hypercholesterolemia67. In 281 analyzed patients, increasing levels of baseline CETP concentrations were a\ssociated with increasing levels of LDL-C and triglycerides, as well as decreasing levels of HDL-C (p < 0.0001 for each). Higher CETP levels were also associated with reduced particle size of both HDL (p < 0.0001) and LDL (p = 0.002). Statin treatment over 2 years significantly reduced overall levels of CETP (p < 0.0001). Interestingly, ultrasound measurement of the thickness of the wall of the carotid artery (carotid intima-media thickness) revealed that patients with higher baseline CETP concentrations subsequently had increased progression of atherosclerosis (p = 0.014 for trend; Figure 5). Similar relationships between CETP, serum lipids, and atherosclerotic progression were observed in REGRESS68. Among 674 men in that study, those with CETP concentrations in the highest quartile also had the highest concentrations of LDL-C and triglycerides (p < 0.001) and increased plaque progression over 2 years as assessed by quantitative coronary angiography (p < 0.001).

Some of the strongest evidence Unking CETP activity with CVD risk comes from a nested case-control analysis from the European Prospective Investigation into Cancer and Nutrition (EPIC) – Norfolk Population study. When 735 apparently healthy subjects who subsequently developed CHD were compared with 1400 matched controls, it was observed that CETP levels were directly correlated with levels of total cholesterol and LDL-C, and inversely correlated with levels of HDL-C (p < 0.0001 for all)69. Furthermore, the risk for CHD was strongly correlated with levels of CETP such that the adjusted risk of patients in the highest quintile was 1.43 compared with those in the lowest quintile (p = 0.02 for linearity), which is consistent with a pro-atherogenic role for CETP activity. The association of CHD risk with elevated CETP levels in the EPIC- Norfolk Population study was especially apparent in patients with triglyceride levels above the median, as the adjusted risks associated with CETP levels progressively increased to reach 1.87 for patients in the highest quintile compared with those in the lowest quintile (p = 0.02 for linearity).

Accepting the limitations of extrapolating from observational data on CETP activity that have been published to date, the weight of available evidence would appear to favor the hypothesis that elevated levels of CETP activity contribute to a more atherogenic lipid profile and may increase cardiovascular risk.

CETP inhibition as a therapeutic strategy

Given this epidemiological evidence suggesting that CETP activity can contribute to progression of atherosclerosis, it is not surprising that inhibition of CETP is being evaluated as a potential new strategy for elevating HDL-C levels and treating CVD. Systematic searches of the literature (MEDLINE, 1966 to present; EMBASE, 1974 to present; proceedings of cardiology conferences) revealed 13 reports involving two approaches that are currently being tested in clinical trials: the first is a vaccine-based strategy, designed to elicit an antibody response to reduce CETP activity, while the second involves pharmacological inhibition of CETP activity.

Vaccine-based strategy

The anti-CETP vaccine, CETi-1, was developed by linking a B-cell epitope of human CETP to a T-cell epitope of the tetanus toxin in order to generate an antibody response against CETP70. In a Phase 1 trial, 36 subjects were randomized to receive doses of 10, 25, 100, or 250 ug of CETi-1 or placebo, and analyses were performed over 25 weeks. Subjects that completed the initial phase received a second dose of vaccine in a 25-week extension phase. In the initial phase, only one treated subject developed anti-CETP antibodies. In the second phase, eight of 15 treated subjects showed antibody responses. However, there was no significant reduction of CETP activity or increase in HDL-C levels observed in these subjects. In a subsequent Phase 2 trial, for which specific details have not been published, CETi-1 (administered at three different doses; details not available) was compared with placebo in 203 patients. After a 1- year regimen involving multiple vaccinations, about 90% of treated patients developed anti-CETP antibodies. Levels of HDL-C in the highest dose group were raised by 8.4%, which was significant compared with baseline but not compared with the placebo group71.

Although this approach has theoretical merit, it is not difficult to envision problems that might arise from eliciting an autoimmune response as a clinical treatment, including the potential for unwanted side effects and the lack of control over the extent of inhibition. More studies will need to be performed to optimize the efficacy and establish the safety of this technique.

Chemical inhibitor-based strategy

The first reported chemical inhibitor of CETP was JTT-705, which was shown to increase serum HDL-C levels and decrease aortic lesions in a rabbit model of atherosclerosis72. Three studies have since evaluated the efficacy and safety of this compound in humans. In 198 subjects with mild hyperlipidemia, JTT-705 dose dependently inhibited CETP activity and raised HDL-C levels by 37% at the highest dose (900mg/day) over 4 weeks73. Levels of LDL-C were also modestly reduced (7% at 900mg/day). In the second study, JTT-705 at doses of 300 and 600mg/day or placebo were tested for 4 weeks in 155 subjects with type II dyslipidemia (LDL-C > 160mg/dL [4.1 mmol/L], HDL-C < 60 mg/dL [1.6mmol/L], and triglycerides < 400mg/dL [4.5mmol/ L]) already receiving therapy with pravastatin 40mg/day74. At 600mg/ day, JTT-705 reduced CETP activity by 30%, raised HDL-C levels by 28%, and lowered LDL-C by 5%. Although this drug was well tolerated, the frequency of gastrointestinal effects such as diarrhea, nausea, and flatulence increased with increasing doses of the drug (diarrhea occurred in 5, 4, 3, and 2 individuals in 900mg, 600 mg, 300 mg, and placebo groups, respectively; nausea occurred in 3, 2, 2, 0 individuals; flatulence occurred in 2, 2, 3, and 1 individuals, respectively)73.

The third study involving JTT-705 was a double-blind, randomized sequence, crossover trial of 600mg/day for 4 weeks in 19 patients with familial low levels of HDL-C (mean baseline HDL-C level, 30.0mg/ dL)75. Treatment-associated inhibition of CETP activity by almost 25% from baseline resulted in a 19% increase in HDL-C (p = 0.01), which was mirrored by a 14% increase in apo A-I (p = 0.02). The sub- fraction of large HDL particles was increased by 42% (p = 0.01) and the numbers of small LDL particles were reduced by 25% (p = 0.05), suggesting that relatively modest levels of CETP inhibition may improve the atherogenic profile of serum lipids. Furthermore, treatment with JTT-705 increased serum paraoxonase-1 activity (p = 0.04) and decreased levels of oxidized LDL autoantibodies, suggesting that the antioxidant activities of HDL may have been improved by treatment. Unfortunately, no safety data were provided in this report. Although these results are promising, larger scale trials will be required to confirm these observations and to determine the effects on atherosclerosis and clinical outcomes.

The second CETP inhibitor to be reported was torcetrapib, which was shown to substantially raise levels of HDL-C and modestly lower levels of LDL-C in early phase 1 clinical trials76,77. Torcetrapib inhibits CETP activity by 80-90% at doses of 2 ≥ 30 mg76, compared with 37% inhibition by JTT-705 at the highest tested dose (900mg)73. Further analyses established that torcetrapib increased apo A-I by slowing the rate of catabolism, and torcetrapib treatment normalized the distribution of apo A-I within HDL fractions of patients with low baseline levels of HDL-C78. In addition, excretion of products of cholesterol metabolism was unaffected suggesting that reverse cholesterol transport was unlikely to have been adversely affected. Torcetrapib has also been shown to inhibit the activity of at least nine genetic variants of CETP to a similar extent as the wild type protein79.

Subsequently, two larger 8-week phase 2 studies demonstrated the efficacy and safety of doses of 10-90mg/day torcetrapib either alone or on a background of atorvastatin 20mg/day in subjects with low levels of HDL-C (men, <44mg/dL [1.1 mmol/L]; women, < 54mg/dL [1.4mmol/L])80. During the 8-week atorvastatin run-in period, levels of HDL-C remained relatively constant at approximately 40mg/dL in all groups, whereas levels of LDL-C were reduced from approximately 125-140 mg/dL to approximately 80-90mg/dL (specific data not provided). In patients treated with torcetrapib alone (n = 162), the mean increase in HDL-C levels from baseline relative to placebo ranged from 9.0% to 54.5% (p ≤ 0.0001 for 30mg, 60mg, and 90mg doses). In addition, mean LDL-C levels were decreased by up to 16.5% (p < 0.01 for the 90mg dose). In patients treated with torcetrapib and atorvastatin (n = 174), mean HDL-C levels increased by 8.3- 40.2% (p ≤ 0.00001 for 30 mg, 60 mg, and 90 mg doses) and mean LDL-C levels decreased by up to 18.9% (p < 0.01 for 60mg and 90mg doses), relative to values recorded during an atorvastatin run-in phase (i.e., reductions in LDL-C levels in patients treated with torcetrapib and atorvastatin were incremental to decreases with atorvastatin alone). Furthermore, data from these studies showed that torcetrapib treatment produced increases in HDL and LDL particle size, regardless of whether atorvastatin was administered. A larger phase 2 trial of patients with LDL-C levels ≤ 130mg/ day (n = 493) confirmed the effect of torcetrapib on HDL and LDL particle size and showed that treatment with torcetrapib 60 mg/ atorvastatin 10-80mg (the doses chosen for further clinical development of torcetrapib/atorvastatin) increases HDL-C levels by 44-66% (p < 0.0001) and decreases LDL-C levels by -41% to -60% (p < 0.0001)81,82. Torcetrapib is well tolerated, although a small percentage of patients have s\hown increases in systolic blood pressure. In the torcetrapib 0 mg, 30 mg, 60 mg, and 90 mg treatment groups, elevations in systolic blood pressure ≥ 15 mm Hg for ≥ 3 consecutive study visits or an adverse event recorded as hypertension were reported in 3.4%, 0%, 5.4%, and 8.8% of patients, respectively. In all cases, blood pressure returned to normal levels within 1 week of discontinuation. The basis for this effect is unknown, and it is not clear whether this effect may interfere with potential benefits from raising HDL-C in those patients.

Phase 2 studies had also suggested that the LDL-C lowering effect of torcetrapib may be reduced by high baseline triglyceride levels81, and so this possibility was explored further in a post- hoc analysis83. When subjects receiving torcetrapib (0 mg, 10 mg, 30 mg, 60mg, or 90 mg/day) with or without atorvastatin were stratified by high levels of triglycerides (> 150mg/dL [3.9mmol/L]) or low (≤ 150mg/dL [3.9mmol/L]), the LDL-C-lowering effects of torcetrapib were blunted in subjects with high triglyceride levels compared with those having low triglyceride levels. However, when added to a background of atorvastatin, torcetrapib lowered levels of LDL-C to a similar extent in both groups of subjects, suggesting that the potential benefits of torcetrapib treatment may be optimized by concurrent statin therapy.

Further analyses of the phase 2 studies have also been used to determine whether torcetrapib effects differ according to gender or baseline HDL-C levels84,85. When groups of approximately 40 male and female subjects, each with either below or above average HDL-C levels (defined as follows: men, ≥ 44mg/dL [n = 43], > 44mg/ dL [n = 41]; women, ≥ 54mg/dL [n = 39], > 54 mg/dL [n = 40]) were randomized to torcetrapib 120 mg/day for 8 weeks, torcetrapib treatment significantly raised HDL-C levels and lowered LDL-C levels compared with placebo (p < 0.05 for both), but no significant differences in the proportionate changes were observed between any treatment groups84. Furthermore, HDL isolated from equivalent volumes of serum from those individuals showed enhanced ability to promote cholesterol efflux via the SR-B1 pathway in vitro, and the increase in SR-B1-mediated efflux correlated with increases in HDL- C and apo A-I85. Given the preferential increases in large HDL particles by torcetrapib, it was not surprising that torcetrapib treatment had no net effect on ABCA1-mediated efflux, which occurs via small, pre -HDL particles. Ongoing studies will determine whether ABCGlmediated efflux is affected by torcetrapib treatment. Although the effects on net cholesterol movement has not been determined, the reduced atherosclerosis observed in preclinical studies of rabbits treated with either JTT-705(72) or torcetrapib86 strongly suggests that CETP inhibition induces a net efflux from the plaque.

Pharmacokinetic and pharmacodynamic analyses of phase 1 study data have also recently been reported87. These analyses showed that morning dosing with torcetrapib, with or without atorvastatin, resulted in greater exposure and enhanced HDL-C elevation compared with evening dosing. It is notable that LDL-C lowering by statins with short half-lives may be more effective when they are taken in the evening87-90. Because LDL-C lowering with statins that have longer half-lives, such as atorvastatin91 and rosuvastatin92 may be less affected by the time of administration, combination of torcetrapib with a longer-lived statin may help to reduce the complexity of treatment and thereby improve compliance93.

Torcetrapib and atorvastatin are currently being evaluated in several phase 3 clinical trials to determine whether the beneficial effects on lipids will translate into reductions in atherosclerotic progression and cardiovascular events.

Implications of HDL-C elevation with CETP inhibitors

Ongoing trials to compare torcetrapib/atorvastatin with atorvastatin alone may help to establish a causal role for HDL in the prevention of CVD and to determine more precisely the magnitude of HDL-related benefits. In addition, they should help to ascertain whether HDL-related benefits are additive or synergistic to those conferred by lowering LDL-C. If epidemiological data suggesting a 2- 3% decrease in the risk of CHD for each 1 mg/dL increment in HDL-C, independent of LDL-C levels94, are reproduced in these intervention trials, even modest elevations in HDL-C would have substantial effects on CHD risk beyond those evident with statin treatment. Thus, the development of CETP inhibitors may allow for a more comprehensive approach to lipid management than has been possible with statins and other available agents.

If CETP inhibitors should be shown to provide clinical benefits, still other questions will need to be addressed. For example, it is not clear whether there exists an upper limit of benefit for raising HDL-C levels. Indeed, recent statin trials have continued to support further lowering of LDL-C targets4,13, and it is conceivable that the reverse will hold true for HDL-C levels. In addition, it is not clear whether the effects and potential benefits of CETP inhibition may vary according to the patients’ lipid phenotype.

Conclusion

Pharmacological elevation of HDL-C levels holds much promise as a strategy to achieve reductions in cardiovascular risk. CETP inhibition is a novel therapeutic strategy that has been shown to produce substantial elevations in HDL-C and modest decreases in LDL- C levels. Ongoing studies are underway to determine whether the significant lipid-modifying effects of CETP inhibitors can provide benefits in terms of reductions in atherosclerotic progression and coronary events in at-risk patients. The results of these studies may herald a treatment paradigm shift in the approach to managing atherosclerosis and CVD.

Acknowledgment

Declaration of interest: Funding for the development of this manuscript was provided by Pfizer Inc. The authors thank Edward Parr, PhD, for editorial input in developing the manuscript.

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CrossRef links are available in the online published version of this paper: http://www.cmrojournal.com

Paper CMRO-3618_4, Accepted for publication: 26 September 2006

Published Online: 02 November 2006

doi: 10.1185/030079906X148634

Benjamin Ansell(a) and F. D. Richard Hobbs(b)

a Divisions of General Internal Medicine/Health Services Research and Cardiology, University of California Los Angeles School of Medicine, Los Angeles, CA 90095, USA

b Primary Care Clinical Sciences Building, University of Birmingham, Birmingham B15 2TT, UK

Address for correspondence: Professor Richard Hobbs, Primary Care Clinical Sciences Building, University of Birmingham, Birmingham B15 2TT, UK. Tel.: +44-121-414-6764; Fax: +44-121-414-3050; email: f.d.r.hobbs@bham.ac.uk

Copyright Librapharm Dec 2006

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