22/11/2022

EFLM WEBINAR
RCT Evidence of Clinical Benefit of LDL-Lowering
“The lower the better”
Atherogenic lipoproteins: IMPROVE-IT Trial Data and Statin Trials for Change in LDL
Cholesterol vs. Clinical Benefit. Hazard ratio for reduction in major

which, when, and how to vascular events in the simvastatin (40 mg) – ezetimibe (10 mg)
group as compared with the simvastatin-monotherapy (40 mg)
group in IMPROVE-IT (n=18,144) is plotted against data from

quantify
other statin trials that assessed the association between change in
LDL-C and clinical benefit:
a: Gruppo Italiano per lo Studio della Sopravvivenza nell’Infarto
Miocardico (GISSI Prevenzione);
b: Antihypertensive and Lipid-Lowering Treatment to Prevent
Heart Attack Trial–Lipid Lowering Trial (ALLHAT-LLT);
c: Assessment of Lescol in Renal Transplantation (ALERT);
d: Lescol Intervention Prevention Study (LIPS);
e: Air Force/Texas Coronary Atherosclerosis Prevention Study
(AFCAPS/TexCAPS);
f: Cholesterol and Recurrent Events (CARE);
g: Long-term Intervention with Pravastatin in Ischaemic Disease
(LIPID);
h: Prospective Study of Pravastatin in the Elderly at Risk
(PROSPER);
i: Anglo-Scandinavian Cardiac Outcomes Trial–Lipid Lowering Arm
(ASCOT-LLA);
Dr. Langlois Michel j: West of Scotland Coronary Prevention Study (WOSCOPS);
k: Post–Coronary Artery Bypass Graft (Post CABG);
l: Collaborative Atorvastatin Diabetes Study (CARDS);
m: Heart Protection Study (HPS);
n: Scandinavian Simvastatin Survival Study (4S).

22nd November 2022

Cannon CP et al. N Engl J Med 2015;372:2387-97.

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Lifetime reduction of LDL-C vs. 5-year reduction of LDL-C by statin therapy

“the sooner the better”
Familial Hypercholesterolemia
(FH)
(N=194,427)
Median follow-up 52 years

(N=196,552)
Median follow-up 5
years High ASCVD* risk even
without other risk factors!

Ference BA et al. Eur Heart J. 2017;38:2459-72 BG Nordestgaard et al. Eur Heart J 2013;34:3478-90 *Atherosclerotic cardiovascular disease
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Are we measuring the Which Atherogenic lipoproteins should be measured?

appropriate biomarker?
B
 Remnant particles

 LDL-particles

 Lipoprotein(a)
Khera AV & Kathiresan S. Nature Reviews Genetics 2017;18:331–344
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Atherogenic Dyslipidemia

Remnants

LDL-R

HDL-C

CETP: cholesteryl ester transfer protein; LCAT: Lecithine cholesterol acyltransferase;

Chapman MJ et al. Eur Heart J 2011;32:1345-61. LPL: lipoprotein lipase; HL: hepatic lipase.
Khera AV & Kathiresan S. Nature Reviews Genetics 2017;18:331–344.
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Transient vs. End Product Remnants

Atherosclerosis

Atherogenic lipoproteins Nordestgaard BG et al. Nature Rev Cardiol 2018;15:261-72

HN. Ginsberg et al. Eur Heart J 2021;42:4791-806.
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Lp(a) Genetic Architecture of Plasminogen and Apo(a) Gene

OxPL (oxidized
phospholipids)

apolipo-
protein(a) LDL-like particle
[ ← Isoform variable segment
Schmidt ]
JLR 2016

F. Kronenberg, G. Utermann. J Int Med 2013;273:6–30

G. Lippi et al. Clinica Chimica Acta 2011;412:797–801
Nordestgaard BG et al. Eur Heart J 2010;31:2844-53.
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genetically determined

Low molecular
weight (LMW)
isoforms

apo(a) size
polymorphism

High molecular
weight (HMW)
isoforms
Fast Slow to 0
Hepatic synthesis of apo(a)
Tsimikas S. J Am Coll Cardiol 2017;69:692–711 F. Kronenberg, G. Utermann. J Int Med 2013;273:6–30
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Data from UK Biobank (N=460,506 adults) Meta-analysis of

7 statin trials*

*4D, 4S, FLARE, JUPITER, LIPID,

MIRACL, and TNT (N=29,069)

(B) and (D) adjusted for age, sex,

previous CVD, diabetes, smoking,
systolic blood pressure, LDL-C, HDL-C.

Cardiovascular events, defined as:

fatal or non-fatal coronary heart
disease, stroke, or revascularisation
procedures.

HR=1.11 (95%CI 1.10-1.12) per 50 nmol/L (~23 mg/dL) increment

AP Patel et al. Arterioscler Thromb Vasc Biol 2021;41:465-474. P. Willeit et al. Lancet 2018;392:1311–20
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Atherogenic lipoproteins: direct causal risk factors

Nordestgaard BG et al. Nature Reviews Cardiol 2018;15:261-272

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CDC Reference Method for standardization of LDL-C and HDL-C

Beta-quantification (BQ) “LDL” d = 1.006-1.063 g/ml

includes IDL: 1.006-1.019 g/ml and Lp(a): 1.045-1.130 g/ml !!
Nordestgaard BG et al. Nature Rev Cardiol 2018;15:261-72
 22/11/2022

Friedewald formula: still fit for purpose?

LDL-C predictive equations beyond Friedewald formula

LDL-C = TC – HDL-C – VLDL-C

TG/5 (mg/dL) or TG/2.2 (mmol/L)

Overestimates VLDL-C at high TG

Rossouw HM et al. Clin Chem Lab Med 2021;59:1930-43

Friedewald WT et al. Clin Chem 1972;18:499-502.
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Friedewald calculation at low LDL-C (<1.8 mmol/l) and high TG concentration underestimates LDL-C Reclassification to “high-risk” (>1.8 mmol/l) by Ultracentrifugation LDL-C
Martin S. et al. J Am Coll Cardiol 2013;62:732-9. Martin S. et al. J Am Coll Cardiol 2013;62:732-9.
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Martin-Hopkins equation Novel Martin-Hopkins equation improves accuracy of calculated LDL-C

at low LDL-C, hypertriglyceridemia and non-fasting status

LDL-C = TC – HDL-C – VLDL-C

TG/adjustable factor

Concordance of risk classification with calculated LDL-C (Friedewald or novel) vs. Ultracentrifugation LDL-C

Martin S. et al. JAMA 2013;310:2061-2068. Sathiyakumar V et al. Circulation 2018;137:10-19.

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Sampson equation (LDL-S) vs. Reference Method (BQ) Direct LDL-C EQA survey

Hypertriglyceridemic serum TG 193 mg/dL (2.2 mmol/L)

Calculated vs β-Quantification (BQ) LDL-C concentration, color-coded according to triglyceride (TG) level.
R2 indicates correlation coefficient; RMSE, root mean square error. To convert LDL-C to mmol/L, multiply by 0.0259.
Sampson M. et al. JAMA Cardiol 2020;5:540-48. HW. Vesper et al. Clin Chem 2012;58:523-27.
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Direct HDL-C EQA survey Non-HDL cholesterol includes ‘remnant cholesterol’

Hypertriglyceridemic serum TG 193 mg/dL (2.2 mmol/L)

(non-fasting)

HW. Vesper et al. Clin Chem 2012;58:523-27. A.Varbo et al. Copenhagen General Population Study. J Am Coll Cardiol 2013;61:427-36.
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#particle number #LDL particle number

Unequivocally defined measurand

IFCC Reference Material

Packard CJ. Curr Atheroscler Report 2022;24:133–42. Marcovina SM et al. Clin Chem 1994;40:586–92
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Meta-analysis of population-based cohort studies Data from UK Biobank

*Age, sex, ethnicity, systolic blood pressure, diastolic blood pressure, antihypertensive medication, diabetes, and smoking.

Table 4. Adjusted* Associations of Different Lipid and Lipoprotein Measures With Composite Cardiovascular Disease Using a Linear Model in
Those Without Baseline CVD and Not Taking a Statin (n=346 686)

Adjusted HR per 1 SD Increase P Value

ApoB 1.23 (1.20–1.26) <0.001

Direct LDL-C, 1.20 (1.17–1.23) <0.001

Friedewald LDL-C 1.17 (1.14–1.20) <0.001

Martin/Hopkins LDL-C 1.19 (1.16–1.22) <0.001

Non–HDL-C 1.21 (1.18–1.24) <0.001

HDL-C 0.81 (0.79–0.84) <0.001

ApoA1 0.81 (0.78–0.83) <0.001

Relative Risk(95%CI) per 1-SD increase 1.25 (1.18-1.33) 1.34 (1.24-1.44) 1.43 (1.35-1.51)

Claire Welsh. Comparison of Conventional Lipoprotein Tests and Apolipoproteins in the Prediction of
Sniderman A et al. Circ Cardiovasc Qual Outcomes 2011;4:337-345 Cardiovascular Disease. Circulation 2019;140: 542-552.
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Discordance analysis according to median concentrations of LDL-C, non-HDL-C and apoB Coronary heart disease (CHD) risk in women with LDL-C < Median (120 mg/dl)

Remnant-C

#LDL particles

S. Mora et al. Women’s Health Study (n=27,533)

Circulation 2014;129:553-61 S.Mora et al. Women’s Health Study. Circulation 2014;129:553-61
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Discordance between lipoprotein cholesterol content (nonHDLc) and particle numbers (apoB) Achieved apoB with PCSK9 inhibition predicts residual ASCVD risk after
adjusting for achieved LDL-C or non-HDL-C, but not vice versa

Achieved apoB adjusted for LDL-C and non-HDL-C, and incidence of major adverse cardiovascular events after 4 months alirocumab
Kaplan–Meier curves among women with concordant and discordant treatment in 9245 CHD patients on high-intensity statin therapy.
high and low apoB relative to nonHDLc
Hazard ratio set to 1.00 at median concentration of apoB (39 mg/dL) and LDL-C (33.5 mg/dL) achieved within the alirocumab group at month 4.

Lawler PR et al. Women’s Health Study. Clin Chem 2017;63:870-9

E. Hagström et al. The ODYSSEY OUTCOMES Investigators. Circulation 2022;146:657–672.
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Treatment targeting apoB goals may further reduce residual risk

EAS-EFLM 2020 Consensus

Pre-preanalytical phase (test ordering)

Comprehensive testing of atherogenic lipoproteins should include tests to assess the risk
conferred by LDL particles, remnant particles and, in selected cases, Lp(a).

Preanalytical phase (test sampling)

Fasting is not routinely required for assessing the lipid profile.
Consider fasting sample when nonfasting TG are ≥4.5 mmol/L; however, this is not a
requirement.

Incidence rate for major

adverse cardiovascular
events after month 4 on
PCSK9 inhibition E. Hagström et al. Circulation 2022;146:657–672. M. Langlois et al. Clin Chem Lab Med 2020;58:496-517
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EAS-EFLM 2020 Consensus EAS-EFLM 2020 Consensus

Analytical phase (test measurement) Postanalytical phase (test reporting)

Follow-up of measured or calculated LDL-C and non-HDL-C of a patient, from baseline
to on-treatment measurements, should be ideally performed with the same method.
The Martin-Hopkins equation may be preferable for LDL-C calculation in patients with
low LDL-C <1.8 mmol/L or TG 2.0–4.5 mmol/L, and in nonfasting samples.
Non-HDL-C calculation rather than direct LDL-C assay may be considered for treatment
follow-up when TG concentration is ≥4.5 mmol/l (invalid LDL-C calculation).
ApoB assays currently provide the most accurate measurement of overall burden of
atherogenic particles in the fasting and nonfasting state, and they can be standardized.
Laboratories should automatically calculate and report non-HDL-C on all lipid profiles.
Laboratory reports should flag abnormal concentrations based on risk thresholds.
Extremely high LDL-C (>5 mmol/L) or TG (>10 mmol/L) should alert clinicians for FH or
pancreatitis risk, respectively.
Post-postanalytical phase (test interpretation and use)
LDL-C is the primary target of lipid-lowering therapy.
When LDL-C goal is achieved, apoB or non-HDL-C should be used as secondary treatment
target in patients with TG 2-10 mmol/L, diabetes, obesity or metabolic syndrome.

M. Langlois et al. Clin Chem Lab Med 2020;58:496-517 M. Langlois et al. Clin Chem Lab Med 2020;58:496-517
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EAS-EFLM 2020 Consensus

EAS-EFLM Consensus Statement
ASCVD risk Dyslipidemia Treatment Treatment

estimation characterization choice target

Lp(a) should be measured
Total cholesterol YES Optionala Optionala Optionala

HDL cholesterol YES YES - -  at least once in a person’s lifetime, to identify high inherited Lp(a)
LDL cholesterol YES YES YES YES
 especially in subjects with (family history of) premature* CVD and/or
Triglycerides YES YES YES -
elevated Lp(a), FH, and recurrent CVD despite statin treatment
Non-HDL cholesterol YES - - YESb

ApoB Optionalb YESb - Optionalb  or if the patient shows poor LDL-C response to statin therapy
Lp(a) YES YES Not yetc Not yetc
 using an apo(a) size-independent assay
a When LDL-C is not available
b In patients with mild-to-moderate hypertriglyceridemia: 2-10 mmol/L (*men <55 yrs, women <60 yrs)
c Unless approved treatment is available to substantially reduce Lp(a) and L(a)-related risk M. Langlois et al. Clin Chem Lab Med
2020;58:496-517
M. Langlois, BG. Nordestgaard et al. Clin Chem Lab Med 2020;58:496-517