Inhibition of PCSK9: A powerful weapon for achieving ideal LDL cholesterol levels

Plasma LDL cholesterol level is a major determinant of cardiovascular disease (CVD) risk. The striking results of the early large-scale statin trials showed that LDL levels can be reduced by 30–40% with comparable percentage reductions in CVD risk. More recent trials show that there is progressively greater relative risk reduction as the on-treatment level of LDL drops. In the JUPITER trial (1), rosuvastatin treatment of healthy subjects who had elevated CRP levels lowered LDL cholesterol levels ≈50% [from a baseline median of 108 mg/dL to 55 mg/dL (25% had values <44 mg/dL)] and resulted in a ≈50% reduction in all CVD events. We recently suggested that an “ideal” LDL cholesterol level of 50 mg/dL should be the target goal of therapy in all high-risk subjects (2), but the Jupiter trial extends this concept to an even larger population and supports the concept that “the lower the LDL the better” is a reasonable position.

However, achieving LDL levels of 50 mg/dL or even lower in patients with high initial values is often not possible with statins alone, even with maximum dosages. Such patients often require 2 or even 3 different drugs to reach a LDL level <100 mg/dL, much less 50 mg/dL, and such combinations increase the risk of side effects. Consequently, the search for new cholesterol-lowering interventions continues apace. In this issue of PNAS, Chan et al. (3) describe dramatic cholesterol lowering in nonhuman primates with the use of a mAb against proprotein convertase subtilisin/kexin type 9 (PCSK9), a recently described key regulator of hepatocyte LDL receptor (LDL-R) expression.

Role of PCSK9 in Regulating Plasma LDL

The number of LDL-R expressed on the surface of hepatocytes is the primary determinant of plasma LDL levels. It binds plasma LDL on the hepatocyte surface, mediating LDL uptake and delivery to the endosomal system, where the LDL particle is released, and the LDL-R is recycled back to the hapatocyte surface to bind LDL particles anew (4). Wild-type PCSK9 decreases the steady-state level of expression of the LDL-R on the hepatocyte cell membrane. Recent studies have now elucidated many of the steps by which this occurs (5). In the hepatocyte, PCSK9 undergoes an obligatory autocatalytic cleavage in the endoplasmic reticulum before being secreted, but its catalytic activity is not required for its ability to modulate LDL-R surface expression. Rather, PCSK9 is secreted into plasma by hepatocytes and acts by binding to the LDL-R extracellularly, being internalized with it and facilitating its enhanced degradation. PCSK9 does not itself degrade the LDL-R but binds tightly to it and channels it toward the lysosomal compartment for degradation. Thus, it inhibits the recycling of the LDL-R back to the cell surface, which results in decreased LDL-R number and

Blocking PCSK9 binding to the LDL-R can profoundly lower plasma LDL levels.

increased plasma LDL levels. Importantly, stimuli that lead to activation of LDL-R synthesis, such as statins, also coordinately activate expression and secretion of PCSK9, which attenuates the LDL lowering activity of the LDL-R induction. Indeed, overexpression of PCSK9 in mice (6) and gain-of-function mutations in humans (7) enhance hepatocyte degradation of the LDL-R, resulting in dramatic elevations of plasma LDL. In fact, the phenotype in humans is difficult to distinguish from that of familial hypercholesterolemia because of mutations in the LDL receptor gene itself.

However, loss-of-function mutations of PCSK9 have the opposite effect: they increase the density of the LDL-R on the hepatocyte cell membrane and increase the rate of removal of LDL from plasma and lower LDL levels (5, 8). Thus, it was expected that strategies that resulted in the inhibition of PCSK9 synthesis or inhibition of the binding of PCSK9 to the LDL-R should lower plasma cholesterol levels. Indeed, lowering of LDL has been demonstrated with siRNAs (9), antisense oligonucleotides (10), or polyclonal antibodies against PCSK9 (11). Chan et al. (3) now show dramatic effects on plasma LDL cholesterol levels in cynomolgus monkeys after infusion of a humanized murine mAb that binds to PCSK9 and blocks its ability to bind the LDL-R. A single i.v. injection of this mAb reduced LDL levels by 80% at 10 days, and the levels returned to normal after 14 days. HDL cholesterol levels decreased 18%, likely because of clearance of apolipoprotein E-containing HDL. In preliminary studies in mice, Chan et al. show that the reduction of plasma LDL levels absolutely depends on the presence of hepatic LDL-R and that infusion of the mAb results in increased hepatic LDL-R expression. Furthermore, they show a synergistic effect of the mAb in statin-treated cells in increasing LDL-R surface expression, which suggests that blocking PCSK9 in statin-treated humans would yield synergistic hypolipidemic effects.

Large Benefits Conferred by Lifetime Low LDL Levels

One of the most important lessons PCSK9 has taught us comes from the work of Hobbs and colleagues (12) on loss-of-function mutations in humans. They show that the low LDL in these cohorts confers a surprising degree of protection against CVD, disproportionate to what one would have predicted based on data from the 5-year statin trials. Two nonsense mutations, relatively more common among African-Americans, were associated with plasma LDL cholesterol levels ≈28% lower than that in African-American subjects with the wild-type allele. In a 5-year statin trial in adults, that 28% reduction in LDL would be predicted to be accompanied by a 25–35% reduction in CVD risk. However, the observed reduction in CVD risk in the subjects with the PCSK9 mutations was 88%. A missense mutation, relatively more common in Caucasians, was associated with a modest 15% decrease in LDL, yet a 47% reduction in risk. Although other factors may be involved, the simplest assumption is that having the low LDL for a lifetime slowed atherogenesis much more than treatment not begun until late in life and lasting only for 5 years. This is a powerful argument for earlier intervention in treating hypercholesterolemia and reducing LDL to ideal levels.

One reason for excitement about the prospects for PCSK9 inhibition is that it is almost certainly going to be additive to, or even synergistic with, the effects of the statins as noted above. Statin treatment increases hepatocyte LDL-R number, but at the same time it increases synthesis and plasma levels of PCSK9 (5). The latter effect leads to decreased hepatocyte LDL-R number, thus blunting the effectiveness of the statin. Mice lacking PCSK9 have been shown to be hypersensitive to statins (13). The combined use of a statin and a PCSK9 inhibitor could make it much easier to reach ideal LDL cholesterol levels in patients at high risk. Used in combination, the dosages of each needed could be reduced, hopefully reducing side effects.

Clinical Development of PCSK9 Inhibitors

Much remains to be done to bring any of these PCSK9 inhibitors to the bedside. Although parenteral administration is not particularly attractive for lifelong treatment, such therapies are now widely used for many chronic conditions, including diabetes. Such injections would likely be readily accepted by high-risk patients who cannot achieve LDL cholesterol levels of 50 mg/dL on available combination therapies or who have limiting side effects to statins or other hypolipidemic agents. For many patients with familial forms of hypercholesterolemia, who begin with LDL cholesterol levels of 200–400 mg/dL, or higher, even achieving 75% reductions with available combinations of 3 and even 4 medications fail to reduce LDL to ideal levels. Thus, a parenteral therapy administered once a week, if relatively free of side effects, would be of great value and acceptable. Of course, unanticipated and and/or disqualifying toxic side effects (e.g., what happens to the mAb:PCSK9 immune complexes?) can only be ruled out in large-scale, multicenter trials, but from a theoretical perspective, it is probably safe to say that the inhibition of PCSK9 per se will not be a source of toxicity. A young woman has been described who is a compound homozygote lacking any detectable circulating PCSK9 and has an LDL cholesterol level of 14 mg/dL with normal levels of HDL and triglycerides (8). At 32 she is alive and well and teaching aerobics.

Although the use of a parenterally administered mAb to block PCSK9 binding to the LDL-R may limit its use to high-risk individuals, the studies of Chan et al. (3) demonstrate in principle that blocking PCSK9 binding to the LDL-R can profoundly lower plasma LDL levels. In the future it may be possible to develop orally available small molecules that can block PCSK9 binding to the LDL-R, or that inhibit the intracellular autocatalytic cleavage required for PCSK9 secretion (5, 14). Such a combined PCSK9 inhibitor–statin approach may be of great value in the treatment of hypercholesterolemia even in patients with less profound elevations in LDL levels and importantly, in “younger” populations. As pointed out above, statin treatment prevents 25–50% of CVD events in 5-year trials. To which the optimist says, “good show!” but the pessimist says “what about the 50–75% of statin-treated subjects who go on to experience a CVD event?” The pessimists have concluded that we have reached the limit of what we can do by lowering LDL. The optimists say, “not at all. We just have to initiate treatment at an earlier age, and lower cholesterol to targets much closer to the ideal.” Subjects and patients enrolled in the 5-year trials were mostly in their 60s or older. Most physicians, using current guidelines, do not treat hypercholesterolemia until the fifth–sixth decades. Yet, as we discuss elsewhere (2), fatty streak lesions and even raised lesions are prevalent by the third decade. Lowering cholesterol to near-ideal levels earlier in life would likely produce a much greater impact on preventing CVD, as suggested by study of patients with PCSK9 loss-of-function mutations described above. The availability of a combined treatment with few side effects would strongly encourage initiation of therapy at a younger age when it is likely to be much more efficacious and might help to motivate practitioners to initiate effective treatment earlier in life.