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how statins work 4: blocking the mevalonate pathway and the question of side-effects

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I may be going into this topic in too much detail, but I’m enjoying what I’m learning along the way, and our biochemistry is enormously complex and fascinating and worth knowing a lot about for its own sake as well as a guard against false claims and over-simplification. I haven’t yet ordered Ben Goldacre’s new book Do statins work?: the battle for perfect evidence-based medicine, which might render all this (superficial) research superfluous, but I doubt it…. [I’ve since discovered that the book won’t be available for quite a while – I wonder what’s holding it back, as clearly it has already been written…]

So I left off with the production of citrate. Thiol (HS) groups, like alcohol (HO) groups can be linked to carboxylic acid groups. The alcohol group can be taken from the carboxylic acid group and the hydrogen from the thiol group, and these can be combined to create water molecules. The sulphur atom is bound to the carbon of the carboxylic acid group, and this is called a thioester link (similar to an ester link but with a thiol rather than an alcohol group). The resulting molecule is Acetyl-CoA. Acetyl is essentially acetic acid with the alcohol group removed.

Oxaloacetate combines with Acetyl-CoA and water to create citrate. The water molecule is involved in breaking the thioester link, via hydrolysis, between the carbon atom (of the carboxylic acid group of the acetic acid molecule), and the sulphur atom of the coenzyme A. These are connected by a covalent bond, in which two electrons are shared, one from the carbon and one from the sulphur. This hydrolytic process results in an alcohol group combining with the carbon atom, with the other hydrogen atom combining with the sulphur to recreate a coenzyme A (CoA-SH).

A break in the covalent bond between oxygen and carbon in the ketone group of the oxaloacetate molecule, as well as a break in the covalent bond in the methyl group of the acetic acid component of the Acetyl-CoA molecule, leads to a new molecule with an extra deprotonated carboxylic acid group, the conjugate base of citric acid, i.e. citrate. The conversion is catalyzed by the enzyme citrate synthase.

Next is the transportation of citrate into the inter-membrane space of the mitochondrion. A protein (SLC25A1) magically transports the citrate into the inter-membrane space from where it diffuses through the outer membrane into the cell cytoplasm. The citrate then undergoes a reverse process, thanks to an enzyme called ATP citrate lyase, I won’t go into all the details here, but oxaloacetate is recreated, a condensation reaction occurs, and ADP and inorganic phosphate is formed. The oxaloacetate is returned to the mitochondrial matrix in the form of pyruvate, via malate. We now have Acetyl-CoA in the cytoplasm, and the pyruvate is transported by another protein (SLC16A1) back into the matrix, completing the cycle, as the pyruvate is reconverted to oxaloacetate. As I’ve pointed out, there’s a lot more to this cycle, in terms of enzymes, hydrolysis, ATP and ADP, but we’re now at the point where we can look at cholesterol synthesis in the cytoplasm.

The first stage of this is the formation of mevalonate. Three Acetyl-CoA molecules are required for this. So, according to the videos I’m relying on, the first reaction joins together two Acetyl-CoA’s. This involves breaking a bond between the carbon and the sulphur atom in one (which sends one electron back to the carbon, the other back to the sulphur), and in the other, breaking a bond between carbon and hydrogen. Then the free-electron carbon is bound to the other molecule’s free-electron carbon. This results in a molecule with a methyl group (CH3), a carbonyl group (C=O), a methylene group (CH2<) and a carboxyl group (COOH) bound to the coenzyme A molecule by a thioester link. In the same reaction the hydrogen atom is bound to the sulphur atom to recreate an intact molecule of coenzyme A, with its thiol group. All of this is a reversible reaction catalysed by an enzyme in the cytoplasm called Acetyl-CoA Acetyl Transferase, and the resulting molecular product is called Acetoacetyl-CoA. This is the result of binding two Acetyl-CoA’s together, and the next step is to bind a third Acetyl-CoA to this molecule. This involves breaking one of the carbon-oxygen bonds in the Acetoacetyl-CoA molecule, breaking a carbon-hydrogen bond in the new third molecule, and introducing a water molecule to hydrolyse the thioester link. The free-electron carbons from the two molecules are bound together, and the hydrogen from the new molecule is bound to the oxygen. The product is catalysed by the enzyme HMG-CoA synthase, and the name of the product is HMG-CoA, aka beta-hydroxy-beta-methylglutaryl-CoA. The ‘beta’ refers to the middle carbon in the 5-carbon chain, to which the methyl and alcohol groups are attached.

So, we’re now at the stage of having produced HMG-CoA. The next reaction is vitally important for a couple of reasons. It’s the rate-limiting step for the entire biosynthesis of cholesterol, and it’s the site of action of statins, which block the synthesis here by blocking the enzyme that catalyses this reaction. That enzyme is called HMG-CoA reductase, and the various statins that can block this enzyme have names such as simvastatin, atorvastatin and rosuvastatin.

So I’ll try to make sense of this step, then I’ll go back to how statins interfere with it. HMG-CoA reductase catalyses the reaction which transforms HMG-CoA into mevalonate. This reaction requires two molecules of reduced NADP (2NADPH) which carry hydride anions, and also two protons (2H+). This effectively brings in four neutral hydrogen atoms, resulting in the creation of two oxidised NADP molecules (2NADP+). A thioester link in the HMG-CoA is broken, ultimately recreating a coenzyme A with an intact thiol group and and an aldehyde group. Also one of the bonds between the carbon and the oxygen can be broken, allowing for the binding of other hydrogens, in such a way that two hydrogen atoms are bound to the end carbon of the molecule, as well as an alcohol group. This is the mevanolate molecule, to which phosphate groups are added, converting it to ‘activated isoprenes’.

So the question of whether inhibiting of HMG-CoA reductase by statins has effects beyond the inhibition of cholesterol production de novo, is obviously essential, so I won’t be focusing so much on cholesterol biosynthesis from here, I’ll be looking, in my amateur way, at claims that the inhibition of HMG-CoA reductase by statins can sometimes have somatic side-effects, including SAMS (statin-associated muscle symptoms) ‘which are the most well-documented side effect of statins, although there appears to be no unifying mechanism’, according to this research article published in Circulation Research. Note the vagueness of this acronym, which might be seen as a red flag. Circulation Research is a journal of the American Heart Association, and looks pretty kosher to me, so I will rely on it heavily in this and possibly other posts. The article is also pretty up-to-date having been published earlier this year (2019).

The article summarises the mechanism of action of statins in a single paragraph which I’ll reproduce here, because it also reports on possible positive pleiotropic effects:

Statins work by competitively blocking the active site of the first and key rate-limiting enzyme in the mevalonate pathway, HMG-CoA reductase. Inhibition of this site prevents substrate access, thereby blocking the conversion of HMG-CoA to mevalonic acid. Within the liver, this reduces hepatic cholesterol synthesis, leading to increased production of microsomal HMG-CoA reductase and increased cell surface LDL receptor expression. This facilitates increased clearance of LDL-c from the bloodstream and a subsequent reduction in circulating LDL-c levels by 20% to 55%. In addition to reducing LDL-c and cardiovascular morbidity and mortality, statins may have additional non–lipid-related pleiotropic effects. These include improvements in endothelial function, stabilization of atherosclerotic plaques, anti-inflammatory, immunomodulatory and antithrombotic effects, effects on bone metabolism, and reduced risk of dementia. These additional benefits are primarily thought to arise because of inhibition of the synthesis of isoprenoid intermediates of the mevalonate pathway.

Pleiotropy is a term I’m just learning about. A pleiotropic gene is one that ‘exhibits multiple phenotypic expression’. For our purposes, the word ‘multiple’ is key. The point here is that the blocking of HMG-CoA reductase may have multiple benefits, not all related to high LDL-c levels, which raises the obvious question about the positive purposes of HMG-CoA reductase production. As to the harms or side-effects, the article has this to say:

Currently, no universally accepted definition of statin toxicity/intolerance exists, with several groups attempting to define the condition. The prevalence of statin intolerance is also widely debated, in part because of difficulties in identification and diagnosis, particularly with respect to muscle symptoms. Observational studies suggest it occurs in 10% to 15% of patients, with clinic data putting it as high as 30%. In randomized controlled trials, the incidence is thought to be 1.5% to 5% of patients, although this is believed to be an underestimation as most studies exclude patients with a history of statin intolerance either before randomization or during the run-in period.

As can be seen, it’s very difficult to find a reliable rate of statin intolerance or toxicity, or even a reliable definition of same – the article lists four separate definitions from different associations monitoring lipid problems and atherosclerosis. It concludes the section with this statement:

Despite the difficulties in identifying and diagnosing statin toxicity, however, several international organizations have identified statin intolerance to be of major clinical importance that warrants further research and investigation.

Clearly the statin wars – if I can be so controversial as to use such a term – are far from over. I’ll continue to educate myself about the issue and promise to write more in the future.

it’s complicated


Cholesterol biosynthesis part 3, by Ben1994 (video)

Cholesterol biosynthesis part 4, by Ben1994 (video)



Written by stewart henderson

October 19, 2019 at 6:50 pm

How statins work 2: atherosclerosis and LDL cholesterol

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Recent studies have revealed that children 8-10 years old are being diagnosed with Type II diabetes, high cholesterol, and high blood pressure at an alarming rate.

Lee Haney

Picture taken from ‘LDL in atherosclerosis and heart disease’ by Axel Sigurdsson, MD, PHD

As I said in my previous post, biochemistry is almost infinitely complex, so bear with me as I crawl towards an understanding of the role of statins in reducing LDL cholesterol in the blood stream, thus reducing atherosclerosis, a major feature of heart disease.

Remembering, first, that cholesterol is a sterol, which is a modified steroid with a hydroxyl (alcohol) group coming off carbon 3. It’s a mostly hydrophobic lipid with this tiny polar hydroxyl group added. It’s carried around in the bloodstream by lipoproteins.

I’ll turn now to atherosclerosis – though I don’t currently know whether statins can perform roles other than reducing the build-up of plaque in the arteries.

So, generally, our arteries carry oxygenated blood from the heart to other organs and regions of the body. Atherosclerosis is sometimes called ‘hardening of the arteries’, as sklerosis is from Greek, meaning ‘hardening’, but it’s really a narrowing rather than a hardening, or perhaps it’s better described as both, as we’ll see. Arteriosclerosis is a more general name, while atherosclerosis means blockage or narrowing (stenosis) due to an atheroma, an abnormal accumulation of ‘debris’ or plaque consisting of fat (mostly), calcium and sometimes fibrous tissue in the inner arterial wall (endothelium). These atheroma are difficult to detect before they cause heart attacks or disease, because heart arteries are very small and hidden deep within the chest. They’re also quite mobile and elastic with blood flow. Heart attack and stroke sometimes happen when the atheroma ‘bursts’ – the fibrous cap (of smooth muscles cells, cholesterol-rich foam cells, collagen and elastin) which surrounds the atheroma is ruptured, or breaks free from the arterial wall, causing a blood clot (thrombus). These are sudden events, not easily detected beforehand. Alternatively, major problems arise when the atheroma becomes large enough to defeat arterial flexibility.

There can be symptoms, apart from such major dramas as heart attacks and stroke, which may act as warning signs for atherosclerosis. The narrowing of the arteries means that less blood and oxygen is reaching the cardiovascular system (ischemia), and this may result in vomiting, angina (chest pain), and general feelings of faintness and anxiety. Atherosclerosis of the carotid artery, which feeds the brain, may have different symptoms, including headaches, dyspnea and facial numbness. Atherosclerosis can also affect the function of the liver, kidneys and other organs, and the vascular system.

So what causes atheromas? It seems that these accumulations of plaque are the result of monocyte-macrophage activity. Macrophages are types of white blood cells (leucocytes) that perform immune and cleansing functions. However, we don’t really know why the plaque build-up occurs – though it might be initiated by damage to the endothelium. We do know that atherosclerosis can begin early, and that blood LDL cholesterol is a major factor in the activity that leads to this build-up. That’s why researchers have been rather single-minded about ways of reducing LDL cholesterol, and even on increasing HDL cholesterol levels, though there’s little evidence, apparently, that higher HDL levels are beneficial. Nor, interestingly, is there much evidence that lowering triglycerides has a positive effect on heart disease, while study after study has shown that low LDL cholesterol levels are key to avoiding cardiovascular problems.

Okay, now I’m going to take a few steps back to look more deeply at the role of LDL cholesterol in building atheromas and so causing atherosclerosis. Returning to my vague mention of macrophages and monocytes, here’s a clearer picture, drawn mainly from this excellent video.

  1. Structure of arterial wall

First, we need to know that the arterial walls are layered. The first layer surrounding the lumen (the tunnel space where the blood flows and where you find red blood cells or RBCs, leukocytes and lipoproteins, etc) is the epithelium, a thin layer of squamous cells. This layer is surrounded by the tunica intima (sometimes the epithelium is described as part of the T intima), an elastic layer quite rich in collagen. It also contains structural cells called fibroblasts, and smooth muscle cells (SMCs). Surrounding the T intima is the tunica media (particularly rich in SMCs), which in turn is surrounded by the thicker, tougher tunica adventitia. In general, the arterial wall becomes stiffer and more fibrous as you move from inner to outer. Atherosclerosis is apparently more of a problem in large and middle-sized arteries which contain more of the protein elastin.

2. Plaque formation

Plaque formation begins, it’s believed, when there’s damage to the thin endothelial layer (only one cell thick) as well as an abundance of circulating low density lipoproteins (LDLs). LDLs (mostly lipid with a small amount of protein) can then move through the damaged layer into the T intima where they become oxidised by ‘reactive oxygen species’ (free radicals) and other enzymes such as metallo-proteases, released by the endothelial cells. These oxidised LDLs, which are now ‘trapped’ in the T intima, will activate endothelial cells to express receptors for white blood cells (leukocytes), particularly the largest types of leucocyte, known as monocytes. So we have this accumulation of oxidised LDLs activating endothelial cells to express adhesion molecules for leucocytes, which brings monocytes and T helper cells into the T intima layer. This movement into tissue transforms monocytes into macrophages (not sure how that happens), and these macrophages then ‘take up’ or engulf the oxidised LDLs and form foam cells. By this time the lipid material dumped into the T intima has created something like a lake of fat, known as a ‘fatty streak’. Foam cells are central to the process of plaque formation and atherosclerosis, as they induce more SMCs into the T intima from the T media by means of a released growth factor, IGF-1 (insulin-like growth factor), and this leads to increased synthesis of collagen in the region, which hardens the plaque build-up, a build-up further fostered by foam cell death which releases more lipid material. Foam cells also release pro-inflammatory cytokines and reactive oxygen species as well as chemokines which attract more macrophages to the site. Upon death they also release DNA material that attracts neutrophils, a very common type of white blood cell. All of this will increase inflammation or plaque build-up in the region.

3. Effects

As mentioned, SMCs contribute to the containment of this inflamed lipid area by releasing proteins such as collagen and elastin, which is used to build a fibrous cap around it. They also stiffen the formation, the atheroma as it’s called, by adding calcium. All of this has the effect of enlarging the atheroma and so reducing the diameter of the arterial lumen in the area, which raises blood pressure as the blood tries to maintain an adequate flow. The calcification of the area also considerably reduces the flexibility of the arterial wall, again resulting in increased blood pressure. Rupture of the fibrous cap may result, which may lead to thrombosis.

So where do statins come in here? Let me quote from an abstract of one academic paper: Statins in atherosclerosis: lipid-lowering agents with antioxidant capabilities, published in 2004:

Statins (3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors) are the first-line choice for lowering total and LDL cholesterol levels and they have been proven to reduce the risk of CHD [chronic heart disease]. Recent data suggest that these compounds, in addition to their lipid-lowering ability, can also reduce the production of reactive oxygen species and increase the resistance of LDL to oxidation. It may be that the ability of statins to limit the oxidation of LDL contributes to their effectiveness at preventing atherosclerotic disease.

Note that oxidation of LDL has the effect of fixing it in the T intima, as mentioned above, so if it’s true, as I presume it is, that statins inhibit LDL oxidation, as well as having other benefits, then they can’t be a bad thing, as long as there aren’t serious side-effects. I’ll continue to explore this topic, as it’s teaching me a lot about the blood, the liver and the circulatory system, inter alia – and it’s great fun. Dr Ben Goldacre has written a book Do statins work? the battle for perfect evidence-based medicine, which hasn’t been released yet, but I intend to get my hands on it and devour it, along with more videos and articles. In the meantime I hope it’s not too controversial to go on saying that the best way to reduce that nasty (but not too nasty) LDL cholesterol is to eat a healthy diet and engage in effective exercise.

PS: haha I know this’ll be unreadable to most, but if anybody finds any egregious error in this, let me know.


Atherosclerosis video – Nucleus Medical Media (2009)

Atherosclerosis – pathophysiology, video by Armando Hasudungen (2014)

Atherosclerosis – part 1, Khan Academy video



Cholesterol metabolism part 1, video by Ben1994 (2015)

Cholesterol metabolism part 2, video by Ben1994 (2015)

Cholesterol metabolism part 3, video by Ben1994 (2015)

Written by stewart henderson

September 21, 2019 at 5:20 pm