Lower LDL is better – can this be achieved with CETP inhibition therapy?

KEYWORDS:

• CETP
• LDL
• cholesterol
• cardiovascular risk
• inhibitor
• statin

This is an exciting time for the development and use of new and emerging lipid-lowering therapies that reduce atherosclerotic cardiovascular disease (ASCVD). In the last few decades, statins had been widely used to successfully lower ASCVD risk. Their success lay in their ability to lower CVD endpoints by 25% for every 40 mg/dL reduction in low-density lipoprotein cholesterol (LDL-C) [1]. Nonetheless, despite the efficacy of statins in reducing CVD events in primary and secondary prevention patients, there is still substantial residual risk and initial and recurring CVD events remaining in these patient groups. Thus, new lipid therapies are needed to better manage CVD.

Because of the wide body of evidence from clinical trials and from epidemiological, genetic, and experimental studies that consistently demonstrate that the lower the level of LDL-C, the lower the CVD risk, regardless of baseline LDL levels, an approach has emerged that lower LDL-C goals should be targeted, particularly for those in high CVD risk groups [2,3]. In the recently released ESC/EAS lipid guidelines, for example, there is a recommendation for a 50% or greater lowering of LDL-C and an LDL-C target of less than 55 mg/dL in very high risk CVD subjects [3]. This includes those with documented ASCVD, either clinically or on imaging (even without an ASCVD event), patients with type 2 diabetes and those with familial hypercholesterolemia (FH), among other categories [3]. Such a target LDL-C may be difficult to achieve solely with maximum tolerable statin therapy, however, and, thus, as second-line therapies, ezetimibe and PCSK9 monoclonal antibody (mAb) inhibitors are now recommended by the ESC/EAS, based on evidence from the IMPROVE-IT, ODYSSEY, and FOURNIER trials [3].

Despite this array of lipid-lowering therapies as first- and second-line treatments for elevated LDL-C, there is still active development of other lipid therapies to lower LDL-C. Reasons include an intolerance to the statin dose required to achieve target LDL-C levels, as well as the inability to achieve LDL targets even with dual statin/ezetimibe therapy in high CVD risk groups. With respect to PCSK9 inhibitors, despite their great success in substantially lowering LDL-C levels, their relatively high cost and lesser level of convenience, being injectables, needs to be considered [4].

This leads to the question of whether cholesteryl-ester transfer protein (CETP) inhibitors, already well studied and tested in clinical trials, and orally administered, should then be considered as an additional lipid-lowering therapy.

Initial evidence of the potential of CETP inhibition as an effective mode to reduce ASCVD came from animal studies and from genetic studies in humans. In rabbits, which possess CETP levels comparable to humans, inhibition of CETP through various means, including via neutralizing antibodies, anti-CETP vaccines, antisense oligonucleotides, and small molecule inhibitors (later tested in humans) all produced anti-atherosclerotic effects when administered after diet-induced atherosclerosis [5]. Further supporting the hypothesis that CETP inhibition is anti-atherogenic came from multiple genetic studies in humans. Large genetic studies consistently showed favorable lipoprotein profiles and lowered ASVD risk in individuals with CETP polymorphisms that reduced CETP activity [6]. Particularly strong evidence was provided in the Copenhagen City Heart Study, in which 10,261 people were followed for up to 34 years [7]. In this cohort, those with two common protein-truncating variants of the CETP gene, known to lower CETP activity, had significant reductions in the risk of ischemic heart disease, myocardial infarction, ischemic cerebrovascular disease, and ischemic stroke. Interestingly, these CETP variants were also significantly associated with longevity [7].

With this strong body of evidence for CETP inhibition being athero-protective, what then is the mode of action of CETP such that CETP inhibition should be beneficial? CETP promotes the exchange of neutral lipids between HDL and non-HDL lipoproteins, consisting of lipoprotein particles containing apolipoprotein B100 (apoB100) [6,8]. The non-HDL lipoproteins include VLDL, their remnants, and LDL particles, all of which are pro-atherogenic. The net effect is a mass transport of cholesteryl esters (CE) from HDL to VLDL and LDL, with a reverse movement of triglycerides (TG) to HDL [6,8]. This neutral lipid transfer is most commonly believed to occur by a conformational change in CETP upon CE binding, leading to the formation of a continuous tunnel across CETP through which CE and TG transfers occur [6,8]. As a result of the above CETP-mediated process, the overall effect of CETP inhibition should be to increase HDL-cholesterol (HDL-C) levels and, conversely, to decrease the cholesterol concentration in the pro-atherogenic non-HDL lipoproteins [6,8].

This is exactly what occurred in most phase 3 outcome trials with various small molecule inhibitors of CETP. Up to a 130% increase in HDL-C and a 40% decrease in LDL-C (calculated using the Friedewald formula) were observed in phase 3 trials with four different CETP inhibitors, the trial populations of which consisted predominantly of individuals with documented atherosclerotic cardiovascular disease (ASCVD). Note, however, that the reductions in LDL-C, calculated in most CETP trials by the Friedewald equation, more than likely overestimated the extent of LDL-C lowering [8,9]. Overestimation of LDL-C reductions holds particularly true at lower LDL-C levels, as in those patients recruited in the phase 3 CETP inhibitor trials, who were on statin therapy during the trials [9]. LDL-C reductions of up to 20%, determined in later trials by beta quantification, more accurately reflect LDL-C lowering achieved with the addition of CETP inhibitors to statin therapy, and are in effect a proxy for the extent of apoB lowering produced, which is a more accurate predictor of CVD outcomes [8,10].

However, despite favorable changes in HDL-C and LDL-C in most trials, the results of most randomized clinical trials of CETP inhibitors were negative – either no change in CVD outcomes or an increase in CVD events (with the earliest inhibitor tested in these trials, torcetrapib) [11]. Of the four CETP inhibitors tested in phase 3 trials, only the administration of anacetrapib produced favorable CVD outcomes. In the REVEAL trial, 34,449 patients with stable ASCVD on statins (and baseline LDL-C levels of 60 mg/dL and HDL-C of 40 mg/dL) were administered either anacetrapib (100 mg) or matching placebo [10]. The primary composite endpoint of nonfatal myocardial infarction, coronary death, or coronary revascularization was significantly reduced by 9% after 4 years in the anacetrapib-treated group [10].

Why the difference in CVD outcomes with the four CETP inhibitors? With torcetrapib, an off-target drug effect in increasing blood pressure and aldosterone levels, as well as impairments in endothelial function, are widely believed to explain the adverse CVD outcomes [12]. The futility of CVD effects with dalcetrapib is generally attributed to the fact that it is a relatively weak inhibitor of CETP and, thus, produced lesser changes in HDL-C and LDL-C than the other CETP inhibitors [13]. However, evacetrapib, despite being a more potent inhibitor of CETP than dalcetrapib, and inducing greater favorable changes in HDL-C and LDL-C, also had a negligible effect on CVD events [14].

Trial design likely played a role in the discrepant results. REVEAL had a much larger patient size than either dal-OUTCOMES (for dalcetrapib) or ACCELERATE (for evacetrapib), and was, thereby, better powered to detect differences in CVD outcomes [10,13,14]. As well, both the dal-OUTCOMES and ACCELERATE trials were terminated early for futility, whereas REVEAL was continued for 4 years. From earlier observations in statin trials, it is clear that clinically significant CVD risk reductions are not seen with lipid lowering until after 2 years. Indeed, this is reflected in the delay in CVD benefits seen in REVEAL [8].

Even with anacetrapib, the reduction in CVD events in REVEAL was fairly modest [10]. This may also be a reflection of trial design. CVD risk reduction is dependent on the absolute reduction in lipid levels. By design, the patients in REVEAL consisted of individuals on statins with very low baseline LDL-C levels averaging 60 mg/dL. Absolute reductions in LDL-C would be expected to be greater had patients with higher LDL-C levels been recruited, as shown in a subset of patients with higher baseline LDL-C treated with anacetrapib in an earlier study [15].

As well, from statin trials, it is known that CVD benefits increase with the number of years of treatment, even beyond the 4 years of trial length of REVEAL. Thus, had the length of treatment with anacetrapib in REVEAL been increased, greater benefit might have been observed [8]. Evidence for this is derived from recent data presented at the American Heart Association in November 2019 by the Oxford investigators of REVEAL on a 2-year follow-up period after anacetrapib cessation (https://www.revealtrial.org/). This follow-up was carried out to test for a lag in the onset of benefit and a prolonged persistence of the effect of anacetrapib treatment. The findings showed a striking doubling of the percentage risk reduction in major coronary events (from 9% to 19%) in the prior anacetrapib-treated group beyond that achieved during the REVEAL trial period.

What were the mechanisms of action through which anacetrapib reduced CVD? Prior studies show that increased catabolism and clearance of LDL particles through the hepatic LDL receptor pathway, causing reductions in plasma LDL-C and apoB levels, plays a major role [16]. This effect may be due to the increased affinity of LDL particles to LDL receptors upon remodeling of their lipid components and size by CETP [12]. It may also be due to an increase in hepatic LDL receptor numbers, although this has not specifically been tested [12]. An additional CETP inhibitor-mediated decrease in LDL formation, either as a result of decreased hepatic production of precursor VLDL particles or from decreased lipolytic conversion of VLDL to LDL, has not been demonstrated but cannot be ruled out.

If the primary mechanism of action of CETP inhibition by anacetrapib is through LDL-receptor-mediated catabolism of LDL particles, as with statins, this can explain why the incremental benefit of CETP inhibition beyond that of statin therapy with anacetrapib did not approach the CVD benefit of statin monotherapy. Both statins and CETP inhibition act to lower LDL particle numbers through the same pathway via increased hepatic LDL-receptor activity [12]. It is then plausible that a certain level of saturation of the pathway occurs when high dose statins are used together with CETP inhibition therapy. The full effect of CETP inhibition is, conversely, unmasked when it is used as a monotherapy and produces the levels of LDL-C (centrifugally separated) and apoB declines observed in complete genetic CETP deficiency (40% and 35%, respectively) and of that seen in statin trials [12].

Consistent with the above concept are the results of elegant Mendelian randomization analyses comparing the LDL-lowering and CVD effects of variants in the CETP gene alone and in combination with variants in the 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR) gene, which encodes the target of statins [17]. Considered alone, variants related to the action of CETP inhibitors were associated with significant reductions in LDL-C, apoB, and CVD that were similar in magnitude to that of variants related to the action of statins. However, when combined, exposure to variants related to CETP inhibition and statins was associated with similar reductions in LDL-C but attenuated reductions in apoB and nonsignificant effects on CVD events. These findings reflect closely what was observed in the CETP inhibitor trials in terms of the CVD effects of CETP inhibitor monotherapy versus that of combination therapy with statins. The discordant relationship of apoB and LDL-C with CVD risk reduction in the above genetics analyses is also in line with that of the CETP inhibitor trials, where apoB reductions induced by CETP inhibition were a much more reliable predictor of CVD outcomes than LDL-C lowering [8,10]. This is made even more apparent in the fact that the results of the REVEAL trial fit well on the slope of the regression line of non-HDL-C levels (a proxy for apoB) reported in trials of statins, ezetimibe, and PCSK9 inhibitors and the corresponding risk of coronary heart disease (CHD) [18]. This effect is attributed to the fact that apoB plasma concentrations are indicative of the absolute number of LDL particles (one apoB molecule is present per LDL particle) that enter the arterial wall and, thereby, induce and promote ASCVD in proportion to their numbers [9,19].

In contrast to the decline in LDL levels, the substantial increases in HDL produced by anacetrapib (130%) are not thought to have contributed to CVD benefits achieved. This in and of itself is not a refute of the HDL hypothesis that HDL elevation confers CVD protection as the quality, components, and subtypes of the HDL produced by CETP inhibition – large, CE-enriched, slow-turning HDL particles – are not those necessarily found to be athero-protective in multitude of past studies [20]. Although some CETP inhibitors markedly increased cellular HDL efflux, a function shown to be significantly associated with decreased CHD risk in prior studies, it is difficult to assess whether this translated to enhanced hepatic HDL reverse cholesterol transport and, thereby, reduced CHD with the CETP inhibitors [21]. The effects of CETP inhibition by anacetrapib on HDL, overall, likely played a negligible role in CVD outcomes, whereas the effects on apoB can explain in large part its CVD mitigating effects.

Other CVD protective effects of anacetrapib – both lipid and non-lipid – may also have contributed to reduced CVD events, including a 30% reduction in Lp(a), an anti-diabetic effect (improvements in HbA1c and insulin sensitivity and a 10% lowering in new-onset type 2 diabetes incidence was observed with anacetrapib in REVEAL), and a proposed anti-thrombotic effect [10,22].

A word should be included here about safety considerations with the CETP inhibitors. While CETP inhibitors, with the exception of torcetrapib, have not shown large increases in blood pressure, slight increases in blood pressure over time have been observed in all phase 3 trials with CETP inhibitors and these should be monitored, particularly in susceptible patients [12]. Furthermore, unlike other CETP inhibitors, anacetrapib has been found to accumulate in adipose tissue chronically, and, conversely, does not clear completely from the circulation, even years after cessation of treatment [12]. Merck had, in fact, halted the development of anacetrapib due to insufficient benefit in October 2017 and had announced that it would not submit applications for regulatory approval of the drug. However, since then, data presented at the American Heart Association in November 2019 on the 2-year follow-up period after anacetrapib cessation showed no safety concerns for non-vascular mortality or morbidity as a result of anacetrapib accumulation (https://www.revealtrial.org/).

The above safety findings, together with the greatly enhanced efficacy of anacetrapib for the duration of the follow-up, may well renew the possibility of anacetrapib being considered for CVD therapy. There are additionally two other CETP inhibitors currently at play. Subjects with certain SNPs in the ADCY9 gene that are putatively protective are being studied for CVD outcomes with dalcetrapib in the dal-GENE trial. This is due to greater CVD benefits seen earlier in subjects with certain polymorphisms in ADCY9 in a post-hoc analysis of dal-OUTCOMES [23]. While results from the dal-GENE trial have yet to be released, analyses of DNA from patients in ACCELERATE and REVEAL failed to confirm a relationship between ADCY9 SNPs and the impact of CETP inhibition on major adverse CVD events [24,25].

More promising developments may be expected for the CETP inhibitor, obicetrapib, previously known as TA-8995, and later as AMG 899, that had been licensed back from Amgen to NewAmsterdam Pharma (personal communication: John J.P. Kastelein) [26]. In the phase 2 TULIP trial over a 12-week dosing period in 337 subjects at high CVD risk (but without documented ASCVD), optimal TA-8895 monotherapy (10 mg) produced LDL-C (assessed by beta quantification) and apoB reductions of 45% and 35%, respectively, and an HDL-C increase of 180% [26]. In combination with statins, TA-8995 conferred an additional decrease in LDL-C of 40–50% and in apoB of 30–35% [26]. The relatively larger reductions in LDL-C and apoB with TA-8995 on top of statin therapy, in comparison to the values reported with other CETP inhibitors in phase 3 trials, may be attributed in part to the higher baseline LDL-C concentrations of the study subjects in TULIP (mean of 140 mg/dL), as well as to properties of the CETP inhibitor itself. In terms of safety considerations, TA-8995 was well tolerated and did not accumulate during the dosing period or 8 weeks after the cessation of the study period [26]. The investigators further noted no adverse effects on aldosterone, endothelin-1, serum electrolyte concentrations, or blood pressure during the study period [26].

Larger phase 3 trials to determine the long-term effects of obicetrapib on cardiovascular safety as well as on apoB and LDL-C lowering efficacy in subjects with complete or partial statin intolerance are now in the planning stage. Considerations in the trial design include the length of the treatment and follow-up periods to assess the efficacy of obicetrapib on lipid and apolipoprotein parameters, CVD outcomes, and safety. They also include considerations of the nature of therapy – that is the use of the CETP inhibitor as a monotherapy or combined with other lipid-lowering therapies, including ezetimibe.

In light of the evidence amassed thus far, what is the role of CETP inhibitors in the management of CVD risk (Table 1)? First, they may be appropriate as third-line lipid therapies in high ASCVD risk patient groups that are unable to achieve target LDL-C levels with statins in combination with ezetimibe or PCSK9 mAb inhibitors (attributed to issues of accessibility, convenience, and cost in the case of PCSK9 mAb inhibitors). Included in this group are patients with Familial Hypercholesterolemia (FH), including both heterozygous and homozygous FH individuals. Alternately, they may be optimally used as a monotherapy in certain individuals. One example are individuals who need to reduce LDL-C levels due to a high CVD risk profile but who are concomitantly intolerant to statin therapy, reported as between 10% and 17% of individuals on statin therapy [27]. CETP inhibitor monotherapy does produce considerably greater reductions in LDL-C when compared to other lipid therapies, including ezetimibe and the newer ATP citrate lyase inhibitor, bempedoic acid [12,28,29]. Another example are patients who are at an elevated risk for both CVD and developing or worsening type 2 diabetes, which can be exaggerated with both statins and PCSK9 mAb inhibitors, but have been shown to be mitigated with CETP inhibition [10]. Furthermore, in individuals for whom Lp(a) is elevated, along with LDL-C, CETP inhibitors may be more suitable than statins and ezetimibe, neither of which improve Lp(a) levels appreciably [30]. While statins have been the mainstay of therapy to lower CVD risk in individuals with elevated Lp(a) concentrations, they have in fact been shown to increase Lp(a) levels by 10% to 20% in several studies, including in a recent subject-level meta-analysis of randomized statin clinical trials [30]. The Lp(a) effect of statins on overall CVD benefit, however, has yet to be determined [30].

Table 1. Putative CETP inhibitor therapy combinations (or CETP monotherapy) and target patient groups at elevated CVD risk that may be considered for treatment with CETP inhibitors for CVD risk reduction.

Therapy	Target patient groups (at elevated CVD risk)
Statin + CETP Inhibitor	Patients unable to achieve target LDL-C with statin monotherapy at any dose (including heterozygous and homozygous FH patients) Patients intolerant to the dose of statin needed to achieve LDL-C targets Patients with ADCY9 SNPs (if additional CVD benefit is demonstrated) Patients with elevated CETP activity levels (if additional CVD benefit is demonstrated)
Statin + Ezetimibe + CETP Inhibitor	Patients unable to achieve target LDL-C with a statin + ezetimibe combination (including: heterozygous and homozygous FH patients; patients intolerant to the statin dose needed to achieve their target LDL-C; and individuals in whom PCSK9 inhibitors are not an option) Patients with ADCY9 SNPs (if additional CVD benefit is demonstrated) Patients with elevated CETP activity levels (if additional CVD benefit is demonstrated)
CETP Inhibitor Monotherapy	Statin-intolerant patients Patients with glucose intolerance or type 2 diabetes Patients with elevated Lp(a) Patients with ADCY9 SNPs (if additional CVD benefit is demonstrated) Patients with elevated CETP activity levels (if additional CVD benefit is demonstrated)

In other groups, the beneficial effects of CETP inhibitors may be magnified and, thus, their therapeutic role on CVD outcomes in these individuals needs to be determined. For instance, subjects with SNPs in the ADCY9 gene that are putatively protective, described above, may derive greater CVD benefit from CETP inhibition with dalcetrapib, if the results of the dal-GENE CVD outcomes trial are positive, at least [23]. The effects of CETP inhibitors may also be heightened in those with high initial CETP levels, as observed in certain ethnicities and in subgroups with elevated triglyceride levels [31]. Overall, there is good potential for the utility of CETP inhibition therapy using a personalized medicine approach to mitigate ASCVD in currently undertreated patient groups (Table 1).

Declaration of interest

The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.

Reviewer disclosures

Peer reviewers on this manuscript have no relevant financial or other relationships to disclose.

Acknowledgments

The author wishes to thank John J.P. Kastelein and Christopher  Cannon for sharing their personal insights for the manuscript.

Additional information

Funding

This paper was not funded.