APOC3, Coronary Disease, and Complexities of Mendelian Randomization


Cell Metabolism

Previews
of global metabolic control, affecting adi-

pose tissue physiology and glucose use

by the liver through a novel UPR cell-

nonautonomous mechanism. It remains

to be shown whether long-term manipula-

tion of this pathway influences other func-

tional outputs of POMC neurons, but for

the time being, the specificity and cell-

type-restricted mode of action of XBP1s

in the hypothalamus can be considered

a promising therapeutic candidate to treat

prevalent metabolic diseases.

ACKNOWLEDGMENTS

The authors are supported by grants P09-015-F,
FONDECYT 1140549, CONICYT-USA2013-0003,
and ACT1109 (C.H.); FONDECYT 3140388 (P.M.);
and 1R01AG042679 (A.D.).
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471–482.
APOC3, Coronary Disease,
and Complexities
of Mendelian Randomization
Jonathan C. Cohen,1,* Stefan Stender,2 and Helen H. Hobbs1,2,3,*
1Department of Internal Medicine
2Department of Molecular Genetics
3Howard Hughes Medical Institute
University of Texas Southwestern Medical Center, Dallas, TX 75390, USA
*Correspondence: jonathan.cohen@utsouthwestern.edu (J.C.C.), helen.hobbs@utsouthwestern.edu (H.H.H.)
http://dx.doi.org/10.1016/j.cmet.2014.08.007

Two new studies report that triglyceride (TG)-lowering mutations in APOC3 reduce coronary heart disease
(CHD) (Crosby et al., 2014; Jørgensen et al., 2014). Here, we explore limitations of using Mendelian random-
ization to evaluate CHD risk, including potential confounding by the widespread use of statin therapy.
The status of plasma TG levels as a risk

factor for CHD has been debated for de-

cades. In most studies, plasma TG levels

are associated with CHD, but adjusting

for confounding variables (e.g., smoking,

insulin resistance, and diabetes) sub-

stantially attenuates the association (Di

Angelantonio et al., 2009). Two recent

studies take a genetic approach to untan-

gle the Gordian knot between TG levels

and CHD (Crosby et al., 2014; Jørgensen

et al., 2014).

Instead of stratifying individuals based

on plasma TG levels, the studies divide

participants into two groups according

to their APOC3 genotypes. Plasma

APOC3 and TG levels are highly corre-
lated. Stratifying by APOC3 genotype

rather than plasma level of TG circum-

vents confounding by factors that affect

both plasma TG levels and CHD, an

approach referred to as Mendelian

randomization (Katan, 1986).

One study, led by Sekar Kathiresan,

identified four rare variants in APOC3

that were associated with a 39% reduc-

tion in plasma TG levels (Crosby et al.,

2014). The variants were then tested for

association with CHD in 110,097 individ-

uals from 15 different studies. Mutation

carriers had a 40% reduction in CHD

compared to noncarriers. The other

study, led by Anne Tybjærg-Hansen,

used a similar strategy (Jørgensen et al.,
2014). They found three APOC3 variants

that were associated with a 44% reduc-

tion in plasma TG levels. In a cohort of

75,725 Danes, carriers of these variants

had a 41% reduction in CHD. Taken

together, these findings provide compel-

ling evidence that reducing APOC3

expression will reduce CHD risk. The

question remains as to whether the

reduced CHD risk in APOC3 variant

carriers is due to lower plasma TG levels

or to other associated factors, such as

lower plasma levels of LDL cholesterol

(LDL-C), APOC3, or remnant lipoproteins,

or to increased levels of HDL-C.

Reductions in LDL-C are consistently

associated with reduced CHD. Figure 1A
eptember 2, 2014 ª2014 Elsevier Inc. 387

mailto:jonathan.cohen@utsouthwestern.edu
mailto:helen.hobbs@utsouthwestern.edu
http://dx.doi.org/10.1016/j.cmet.2014.08.007
http://crossmark.crossref.org/dialog/?doi=10.1016/j.cmet.2014.08.007&domain=pdf


Figure 1. Genetic and Pharmacological Reduction in LDL-C and Coronary Heart Disease
(A) Reduction in CHD risk associated with genetic variants (blue circles) and pharmacological agents
(green circles) that lower plasma levels of LDL-C. Genetic variations that reduce plasma LDL-C levels
are associated with a greater reduction in CHD compared to that seen in statin trials. The sources of
the data shown in this figure are as follows: APOB rs754523 (PMID: 18193043), LDLR rs2228671
(PMID: 18714375), ABCG8 rs4245791 (PMID: 24657701), and PCSK9 (PMID: 16554528). The red circle
represents the CHD reduction (�46%) that is predicted for a loss-of-function mutation in APOC3
(R19X) (Crosby et al., 2014; Pollin et al., 2008). WOSCOPS, The West of Scotland Coronary Prevention
Study (PMID: 7566020); CARE, Cholesterol and Recurrent Events Trial (PMID: 8801446); HPS, Heart Pro-
tection Study (PMID: 12114036); 4S, The Scandinavian Simvastatin Survival Study (PMID: 7968073).
(B) Effects of APOC3 loss-of-function variants on circulating lipid and lipoprotein levels and on CHD.
Proven causal links are indicated by blue arrows. Question marks indicate where causality has not
been established. The red X indicates no causal relationship. IDL, intermediate density lipoprotein; CR,
chylomicron remnant.

Cell Metabolism

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plots the reduction in CHD as a function of

the reduction in LDL-C in four studies in

which subjects were treated for 5 years

with a cholesterol-lowering statin (green

line). The blue line shows the reduction

in CHD in subjects with DNA variations

that lower LDL-C levels. For each per-

centage reduction in LDL-C, the LDL-C-

lowering variants produce a much greater

reduction in CHD than seen in the statin

trials. Presumably this reflects the fact
388 Cell Metabolism 20, September 2, 2014 ª
that DNA variants lower LDL-C levels

from birth, whereas statin treatment is

initiated when atherosclerotic plaques

have already developed.

Can the reduction in CHD in the APOC3

mutant carriers be explained by a

reduction in LDL-C levels? The effects

of APOC3 inactivation on LDL-C levels

remain inconclusive (Pollin et al., 2008;

Tachmazidou et al., 2013). Pollin et al.

identified a nonsense mutation (R19X) in
2014 Elsevier Inc.
APOC3 that is common in the Amish

and is associated with a 17% reduction

in plasma LDL-C levels (Pollin et al.,

2008). Based on prior genetic studies,

mutations that lower LDL-C by 17%

should decrease CHD by �46% (Fig-
ure 1A), which is similar to the reduction

observed in the two APOC3 studies

(40% and 41%).

The APOC3 carriers in the discovery

cohort of the Kathiresan study (Crosby

et al., 2014) had a 16% reduction in

LDL-C level, which is similar to that

observed in the Amish, but corresponding

data were provided for only a subset of

the cohorts in the CHD association study.

In the Tybjærg-Hansen study (Jørgensen

et al., 2014), the mean plasma LDL-C level

was only 3% lower in APOC3 carriers than

in noncarriers. This modest reduction in

LDL-C cannot account for the dramatic

reduction in CHD associated with the

APOC3 variants.

A factor that may mask the contribu-

tion of plasma LDL-C levels to the reduc-

tion in CHD in APOC3 carriers is statin

treatment. Statins are more likely to

be prescribed to individuals who have

higher plasma LDL-C levels, multiple

CHD risk factors, or established CHD. If

APOC3 noncarriers have higher plasma

LDL-C levels and/or more CHD, they

would be more likely to be treated with

statins. This would lower their LDL

levels and obscure differences in LDL-C

levels between APOC3 carriers and

noncarriers.

Could an excess of statin use among

noncarriers mask the effect of APOC3

variants on LDL-C levels in these two

studies? Statin use was not described

in the Kathiresan study. In the Tybjærg-

Hansen study, statin use was more prev-

alent among noncarriers than carriers,

although the difference did not reach

statistical significance. Given the large

effects of LDL-C on CHD risk and the as-

sociation between APOC3 mutations

and LDL-C levels, it is premature to

conclude that the reduction in CHD in

APOC3 variant carriers is independent

of plasma LDL-C levels. Resolution of

this issue will require additional studies

in statin-naive individuals or in which

pre-statin LDL-C levels are used in the

analysis.

Alternatively, factors other than LDL-C

levels may contribute to the reduction in

CHD. APOC3 has pleiotropic effects



Cell Metabolism

Previews
(Figure 1B). It is possible that APOC3 itself

promotes atherosclerosis (Ginsberg and

Brown, 2011), or alternatively, that

APOC3 retards clearance of atherogenic

lipoprotein remnants. If the lower levels

of TG in APOC3 carriers are atheroprotec-

tive, studies using other variants that

lower TG levels without affecting other

CHD risk factors should replicate the

association. Carriers of the APOC3 vari-

ants also have higher plasma levels of

HDL-C, which are inversely associated

with CHD. Ironically, Tybjærg-Hansen

and Kathiresan performed the key genetic

studies that showed plasma HDL-C levels

are not causally related to CHD risk

(Frikke-Schmidt et al., 2008; Voight

et al., 2012). Thus, it is unlikely that

HDL-C is conferring the cardioprotective

effect of the APOC3 mutations.

The identification of other sequence

variations that lower plasma TG levels

without altering other risk factors would

bolster the contention that TG lowering

is causally linked to reduction in CHD.

APOC3 may be an excellent therapeutic

target for patients with severe hypertri-

glyceridemia. These patients are at risk

of developing pancreatitis, a potentially

life-threatening disorder, and the arma-

mentarium of drugs to treat severe hy-

pertriglyceridemia is extremely limited.
The high circulating levels of APOC3

(10–20 mg/dl) may limit the efficacy of

targeting APOC3 using antibody-based

therapies, but strategies that target

hepatic APOC3 mRNA may prove

efficacious.

Homozygotes for loss-of-function mu-

tations can provide important clues as

to the safety, as well as the efficacy, of

therapies targeting a specific protein.

Neither study identified any individuals

with total APOC3 deficiency. Identifica-

tion of such individuals would provide

reassurance that extreme pharmacolog-

ical inhibition of APOC3 would not have

unforeseen detrimental effects.

As these two studies attest, human

genetic studies have fueled a resurgence

of interest in lipid-modifying therapies

for CHD prevention. Studies using a

Mendelian randomization approach hold

the promise of identifying new thera-

peutic agents for CHD, but they must

address problems associated with pleiot-

ropy and account for effects of statin

treatment on plasma LDL-C levels.

Short-term reductions in LDL-C levels

due to statin therapy do not reflect lifetime

exposure to this atherogenic lipoprotein.

Mendelian randomization, although a

powerful approach, does not eliminate

all confounding factors.
Cell Metabolism 20, S
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Stem Cell Lineage Specification:
You Become What You Eat
Clifford D.L. Folmes1,* and Andre Terzic1,*
1Center for Regenerative Medicine, Mayo Clinic, Rochester, MN 55905, USA
*Correspondence: folmes.clifford@mayo.edu (C.D.L.F.), terzic.andre@mayo.edu (A.T.)
http://dx.doi.org/10.1016/j.cmet.2014.08.006

Nutrient availability and intermediate metabolism are increasingly recognized to govern stem cell behavior.
Oburoglu et al. (2014) now demonstrate that glutamine- and glucose-dependent nucleotide synthesis segre-
gate erythroid versus myeloid differentiation during hematopoietic stem cell specification, implicating a
metabolism-centric regulation of lineage choices.
Flexibility in energy metabolism enables

cells to prioritize metabolic pathways in or-

der to support stage-specific energetic de-

mands (Folmes et al., 2012). Interrogation

of stem cell metabolism has identified
glycolysis as a key player in the mainte-

nance of stemness through provision of

energy and anabolic precursors (Folmes

et al., 2011), while oxidative metabolism al-

lows for more efficient energy production
to match energy-demanding processes

of differentiating progeny (Chung et al.,

2007). Furthermore, individual metabolic

pathways underlying stem cell renewal

versus lineage specification are starting
eptember 2, 2014 ª2014 Elsevier Inc. 389

mailto:folmes.clifford@mayo.edu
mailto:terzic.andre@mayo.edu
http://dx.doi.org/10.1016/j.cmet.2014.08.006
http://crossmark.crossref.org/dialog/?doi=10.1016/j.cmet.2014.08.006&domain=pdf

	APOC3, Coronary Disease, and Complexities of Mendelian Randomization
	References