Why focus on triglyceride-rich lipoproteins and their remnants?

Hello. I’m Henry Ginsberg from the Vagelos College of Physicians and Surgeons of Columbia University and I’ll be speaking today about TG-rich remnant lipoproteins as target for residual risk. These are my disclosures.

We know that dyslipidemia has been a target for reducing cardiovascular disease but the focus, of course, and rightfully so, has been on LDL reduction. These are the latest guidelines from the ESC/EAS showing different levels of LDL goals and targets for very high risk, high risk, moderate risk and low risk. However, even though we have the ability to reach those targets with our potent LDL-lowering drugs, cardiovascular disease is still a major challenge. There are millions and millions of deaths each year from cardiovascular disease and the overall incidents of cardiovascular disease has actually been increasing in the past two decades.

When we look at the causes of cardiovascular death, we see that it’s ischemic heart disease as number one with cerebrovascular disease close behind. So, what are the causes of residual risk for cardiovascular disease? On this slide, we portray all of the different factors that can contribute to or decrease cardiovascular disease. And at the top, we’ve highlighted dyslipidemia and what we’re talking about today is dyslipidemia separate from LDL, the rest of the lipid profile that can cause cardiovascular disease.

To point out the importance of this, these are two studies, IMPROVE-IT where simvastatin plus ezetimibe was compared to simvastatin alone and the FOURIER Trial with evolocumab plus statin as compared to statin alone. In both trials, the green area is the difference that you obtain in reducing risk with either ezetimibe added to statin or evolocumab added to statin but below that in each slide is a large area. Those are the people in the study, regardless of what treatment group they were in, that were having cardiovascular events. This is the residual risk of cardiovascular disease. And we know that triglyceride is very important as one of the residual lipid factors for the risk of cardiovascular disease.

This is a very important study from now 20 plus years ago from Melissa Austin showing that increased triglycerides of about 1 mmol, 89 mg/dL, increased risk for both men and women. And this was at the lower part of the graphic independent of all adjusted factors, including HDL cholesterol.

These are more recent data from NHANES showing increasing triglyceride levels from very levels, 1.69, which would be about 150 mg/dL up to about 500 mg/dL, 5.65 mmol/L and in each case, in the overall group for those on statin, for those not on statin, increasing triglycerides were associated with increased risk for cardiovascular disease.

Here we see similar data from the ODYSSEY Trial with alirocumab, both short-term on the left at 16 weeks and long-term on the right the entire three plus years of follow-up. And we see baseline triglycerides were associated with higher and higher risk in these groups of individuals despite the treatment they were getting to lower their LDL to optimal levels.

So, triglycerides we believe are a very important risk factor for cardiovascular disease and mandelian randomization studies suggests they’re actually causal that carried on lipoproteins, thus the triglyceride-rich lipoprotein as the title of our topic today.

And these are the TG-rich lipoproteins. On the left is the chylomicron, made in your intestine. It’s a big oil droplet because you need grams of fat and it’s wrapped in phospholipid. There’s a little bit of cholesterol ester in the core and proteins on the surface. And on the right is VLDL, made in the liver, regardless of the time of day but insensitive to a number of regulatory factors. Triglyceride droplet in the middle, more cholesterol ester than is seen in the chylomicron remnant and the VLDL is smaller than the chylomicron remnant and has its proteins.

And the protein of interest is apolipoprotein B or ApoB, one of the largest proteins we make and this is the full length ApoB, 4,536 amino acids that comes out on VLDL. It has the domain for binding to the LDL receptor. It’s also necessary for the assembly and secretion of VLDL.

And in the intestine, we make a truncated form of that, about 48-percent of the full length and we call it B48 and it lacks the LDL receptor binding domain but is also necessary for the assembly and secretion of the chylomicron.

So, what are TGRL remnants? And I’ve shown you what the TGRLs are. Well, very simply, when you eat a meal and you digest fat in the intestine and the fatty acids and monoglycerides are taken up into the enterocytes, they’re reassembled to the triglyceride and the intestine secretes a chylomicron into the lymphatic system, makes its way to the circulation. In the circulation, it interacts with lipoprotein lipase and, under the best conditions, removal of about 85-percent of the triglycerides in the chylomicron follows and we’re left with a chylomicron remnant. And that goes into the liver. Chylomicron remnants really are not converted to LDL, they’re an end product going into the liver.

When we have VLDL to look at, we see the liver here, as I’ve said, makes VLDL, has the full length ApoB on it. It enters the circulation immediately, interacts again with lipoprotein lipase. About 85-percentm under normal circumstances, of the triglyceride it’s carrying is removed in adipose tissue and in muscle beds and the remaining VLDL remnant is taken up by the liver or undergoes further lipolysis by hepatic lipase and maybe some other factors are involved with remodeling and you’re making LDL.

So, what are remnants? They are the chylomicrons and the VLDL that have lost most of their TG via lipoprotein lipase mediated lipolysis. But they also have increased their content of cholesterol esters from HDL and LDL via the action of cholesterol ester transfer protein. All of this happens as they circulate in the plasma.

So, how to TGRL remnants differ from LDL and do these differences make them more atherogenic? And on the left, we have an LDL 23 nm in diameter, 2,000 to 2,700 approximately molecules of cholesterol per particle, ApoB is the protein there and it’s interacting with the LDL receptor around the body and at the surface of the liver. And then we have remnant particles, which they’re chylomicron remnants. They have B48. If they’re VLDL remnants, they have B100. They’re larger in diameter and they carry many, many more cholesterol molecules per particle. They also have TG. Obviously, they’ve lost most of their TG from the nascent chylo and VLDL but they have more TG than LDL does and, in the artery wall, that TG can be lipolyzed by lipoprotein lipase locally in the fatty acids that are released, might be atherogenic, might be lipotoxic and the remnant at the liver can interact with several pathways for removal. So, per particle, remnants can deliver two to three times more cholesterol to the artery wall than LDL. They have more TG than LDL and lipolysis of that TG can be proinflammatory and they have several apoproteins in addition to ApoB that may be proatherogenic and there’s been a fair amount of work about apo(c3) in that regard. But LDL particles do outnumber the remnants by five to 10-fold in someone with an average LDL cholesterol. And if you were on a statin and a PCSK9 and ezetimibe, then your LDL particles may be so low they’re almost equal to the remnant particles.

So, how do we know that TGRL remnants are atherogenic? We have a rare disorder, one in 10,000 or fewer called type III hyperlipoproteinemia or dysbetalipoproteinemia where ApoB is lacking or E is defective in its ability to bind to the LDL receptor because it’s the ApoB 2 isoform and so on the background of overproduction of VLDL or chylomicron, insulin resistant people, obese people, diabetics, we have a remnant that cannot be removed by the liver that accumulate and these people get early and severe atherosclerosis.

On the other hand, the vast majority of people who have atherosclerotic cardiovascular disease with increased remnants are someone like this, a 60-year-old man, already has had an MI three years before, hypertensive, obese, diabetic and recently had an ASC and needed a CABG. And the lipids in this gentleman at the time of that acute coronary syndrome and the coronary artery bypass procedure had an LDL of 60 because he was on drugs to lower his LDL but his triglyceride was 250 and that would be the lipoprotein VLDL and some chylomicrons that would be carrying about 50 mg/dL of cholesterol, a remnant cholesterol. His HDL is low. His non-HDL is high. His ApoB is pretty good because of all those LDL-lowering drugs but he still has an event.

Are there good surrogates that we can measure in individuals like this that tell us they have high remnants? Well, here’s a schema of, again, VLDL that enters the circulation of the liver, undergoing lipolysis, getting smaller, becoming a remnant. We also sometimes call it intermediate density lipoprotein and then that going on to LDL or being taken out of the system in the liver. And for the chylomicron, we have those large lipid droplet particles entering the circulation, losing their triglyceride LDL and becoming remnants. And the lower part of the schema shows in a normal individual with low TG, very few of these ApoB48 and ApoB100 particles are actually in the plasma at any point in time. But if your TG is elevated, 3 mmol or approximately 300 mg/dL, you will have a lot of B48 and B100 particles in the plasma, particularly after a meal. So, we can measure TG. That’s what’s a major lipid in those particles. We can measure remnant-like particle cholesterol, which in most cases and most of the papers you’re reading, is actually TG divided by five but there are direct assays and we could also look at non-HDL cholesterol. ApoB is probably the best surrogate but a little bit more difficult to obtain for most people.

Now, this is the distribution of triglyceride in Denmark, courtesy of Borge Nordestgaard and Anne Tybjaerg-Hansen and their colleagues. You can see that if you look at the X axis go up to about 300 mg/dL, between 3 and 4 mmol of triglycerides, you’re dealing with probably 80 to 85-percent of the population. And in that curvilinear portion of this picture are the remnants.

So, we have TG there, we have that cholesterol, we have ApoB. Does TG lowering prevent cardiovascular disease? Well, what about fibrates? Maybe. And if you believe in subgroup analyses where the post hoc or ACCORD prespecified, if you look to the right, the primary endpoint subgroup of TG high and HDL low all had significant benefit from fibrates.

What about omega-3 fatty acids? Well, maybe it’s only EPA, icosapent ethyl in particular, and not EPA/DHA combinations based on REDUCE-IT and the STRENGTH Studies. But when you look at those studies and look at the lipid changes, TG went down about the same in both groups. ApoB went down about the same in both groups. LDL went up about the same in both groups. CRP went down about the same in both groups. When you look at the placebo arms of those studies, they differ dramatically and that’s the question that remains.

So, what do we have coming next? We have pemafibrate in PROMINENT Study, which is a slightly different kind of fibrate that we’re looking at in people with high TG and low HDL. The whole study is the right subgroup of individuals. We have APOC3 inhibition and we have ANGPTL3 inhibition. And with that, I’ll end. Thank you.