LIPID - the British Columbia Society of Laboratory Science

LIPID:
Moving from
Theory to Practice
Dr. Gordon Hoag
Dept of Laboratory Medicine, Pathology & Medical Genetics
Vancouver Island Health Authority
Victoria Lipid Clinic

Disclosure Information
Honoraria for presentations from Merck Frosst, Schering, Pfizer,
AstraZeneca, Fournier and Roche.
A member of scientific advisory boards from AstraZeneca, Schering,
Merck Frosst, Oryx Pharma and Beckman Coulter.
A recipient of unrestricted educational grants from Merck Frosst and
AstraZeneca.
A recipient of research grants from Merck Frosst, Sanofi, Pfizer and
GlaxoSmithKline.

Mean Total Cholesterol mg/dL
200
180
160
140
120
100
80
60
40
20
0
Hazda !Kung San Howler
Monkey
Horse Peccary African
Elephant
Hunter-Gatherer Humans Wild Primates Wild Mammals Modern Human

What is a logical approach?

Extra Hepatic
Organs
Net Cholesterol Balance in Humans
Liver
Intestine
Dietary Cholesterol
(400 mg/day)
LDL
VLDL
Cholesterol
LDLR
Cholesterol
1200 mg/day
(cholesterol+bile acids)
Cholesterol
Acetyl-CoA
ABCA1
HDL
Acetyl-CoA
SRB1
800 mg/day
Synthesized Cholesterol
(400 mg/day)
Fecal Sterols
(800 mg/day)
Courtesy Dr Murray Huff

M24
Ezetimibe : Mechanism of Action
Apo B 48
CE Poor
Chylomicron
ABCG5/G8
ACAT
NPC1L1
Protein
X
Less free
cholesterol
& sterol
absorption
Intestinal Lumen
EZETIMIBE
Enterocyte
Lymphatic Vessel
Ezetimibe has specific, high affinity binding
to a structural protein on the brush border

Slide 6
M24
same
Merck, 07/02/2008

POSCH-Study
Program on the Surgical Control of the Hyperlipidemia
*5 years after randomization
"The POSCH trial provides long-term evidence supporting
effective lipid modification in the management of atherosclerosis"
n=421 Follow up = 14.7 years
Influence on Lipids*:
Total cholesterol:
LDL-C
HDL-C
Reduction of Endpoints:
Total mortality:
CHD-mortality:
PTCA/Bypass
- 23 %, p

hs-CRP Adds to Predictive Value of TC:HDL
Ratio in Determining Risk of First MI
5.0
4.0
Relative Risk
3.0
2.0
1.0
0.0
High Medium Low Low
Total Cholesterol:HDL-C Ratio
Medium
High
Ridker et al, Circulation 1998;97:2007–2011.

JUPITER: Evidence supporting pharmacological
intervention in medium risk patients
Primary Endpoint: Time to first occurrence of a CV death, non-fatal stroke, non-fatal MI,
unstable angina or arterial revascularization
Cumulative incidence
0.08
0.06
0.04
0.02
0.00
N at risk
Rosuvastatin
8,901
Placebo
8,901
HR 0.56 (95% CI 0.46 – 0.69)
P < 0.00001
0 1 2 3 4
Follow-up (years)
8,631
8,621
8,412
8,353
6,540
6,508
3,893
3,872
1,958
1,963
1,353
1,333
Placebo
251 / 8,901
Ridker P et al. Rosuvastatin to prevent vascular events in men and women with elevated C-
reactive protein.N Engl J Med 2008; 359: 2195-207
983
955
544
534
Rosuvastatin 20
mg
142 / 8,901
157
174
RRR
-44%
LDL reduction 50%

Evidence favouring LDL reduction for the prevention and
treatment of atherosclerosis is strong and compelling
A prospective meta-analysis of data from 90,056 individuals from 14 trials of statins 1
A 1 mmol/L (39 mg/dL) reduction in LDL-C was associated with a…
50
23% reduction in
major coronary events
50
21% reduction in
major vascular events
40
40
Proportional reduction in
event rate (%±SE)
30
20
10
0
-10
0.5
(19)
1.0
(38)
Reduction in
LDL-C mmol/L (mg/dL)
1.5
(58)
2.0
(77)
Proportional reduction in
event rate (%±SE)
30
20
10
0
-10
0.5
(19)
1.0
(38)
1.5
(58)
Reduction in
LDL-C mmol/L (mg/dL)
2.0
(77)
Adapted from Baigent C, et al, Cholesterol Treatment Trialists’ (CTT) Collaborators. Lancet
2005;366:1267–1278.

CD1
Nonfatal MI and CHD death RR reduction (%)
Multiple studies showed a relationship between
LDL-C Reduction & CHD relative risk
100
80
60
40
20
0
-20
London
Oslo
MRC
Los Angeles
Upjohn
LRC
NHLBI
POSCH
4S
WOSCOPS
CARE
LIPID
AF/TexCAPS
HPS
ALERT
PROSPER
ASCOT-LLA
CARDS
15 20 25 30 35 40
LDL-C reduction (%)
Adapted with permission from Robinson et al. JACC 2005; 46:1855-62

Slide 16
CD1 Even when looking at the original slide Dr Ftichett sent, I still can't find where London is ?????
Charles Dupont, 05/03/2008

Relationship Between Mean Low-Density Lipoprotein
Cholesterol Levels and Median Change in Percent Atheroma
Volume for Several Intravascular Ultrasound Trials
Nissen, S. E. et al. JAMA 2006;295:1556-1565.
Copyright restrictions may apply.

Wide Variation of Response to Statins
% LDL-C reduction to
simvastatin or pravastatin up
to 60 mg/d for 3 months
Mean reduction 20%
12% had LDL-C ↓

M19
Individual LDL-C % Response
to Atorvastatin 10mg
% LDL-C change
0
-10
-20
-30
-40
-50
0
-10
-20
-30
-40
-50
-60
-70
Men
-60
-70
Women
Pedro-Botet J et al. Atherosclerosis 2001; 158:183-93

Slide 22
M19
is author's name Pedro-Bolet or Botet.. need to check
Merck, 07/02/2008

Limitations of Statin Mono-therapy in
Achieving LDL-Cholesterol Targets
Attenuated dose response for LDL-C lowering
» Largest effect at lowest dose
» Doubling statin dose reduces LDL-C by ~5-6%
Wide inter-individual variation of statin response
Higher doses associated with increased toxicity
» Hepatic
» Musculoskeletal

The Majority of Statin LDL-C Efficacy
is with the Starting Dose
0
Atorvastatin Rosuvastatin Simvastatin
Mean % Change in LDL-C From
Untreated Baseline
-10
-20
-30
-40
-50
-37%
-6%
-5%
-3%
-28%
-46% *† -7%
-4%
-7%
-6%
-3%
10mg
20mg
40mg
80mg
-60
10 mg 20 mg 40 mg 80 mg
Jones PH et al. Am J Cardiol 2003;92:152-60

Genetic Variants of NPC1L1 and Cholesterol Absorption
Numerous rare protein
variants of NPC1L1 exist in
the patient population
NPC1L1 Protein Variants
The relationship between
these and both ezetimibe
response and altered sterol
absorption remains to be
determined
Huff et al ATVB 2006;26:2437

The Genetics of the Cholesterol Transport
Protein, Neimann Pick C1 like 1 Protein, in the
Intestine
Eight genetic variants described
Variants have a pharmacological effect on
response
Normal variant (60% of the population) has
a 15-25% response
Non-responders and hyper-responders are
expected
Two week course of Ezetimibe is
informative to the genetics of the patient

Case 1
M.M. 43 y.o., British Origin, Photographer, Married, 1 son
Identified high cholesterol 15 years ago, Father age 65
died.
AMI recently (2004)
On Lipitor severe jaundice.
Several episodes of chest pain. No L arm radiation
No: DM, HBP, TIA’s, PVD.
Exercise level very high
Alcohol: wine/beer. 2-6 oz/wk

Case 1 continued
On examination: Arcus Bilaterally, Tendons
thickened, abdominal girth 89.5 cm, height
5’6”, and weight 175 lbs
Lab Value Initial 6 wk Ezetrol
– TG(mmol/L) 3.92 1.52
– Chol(mmol/L) 8.27 6.47
– LDL(mmol/L) 5.6 4.1
– HDL(mmol/L) 0.93 0.94
Dx F.H.

Benefit from LDL-C reduction
independent of mechanism
31

Pharmacotherapy (LDL Cholesterol):
Immediate treatment for high-risk patients
Concomitant diet and lifestyle changes
Statin monotherapy
Moderate risk patients:
- Initiate health behavior interventions
- Pharmacological treatment is indicated if:
LDL-C >3.5 mmol/L
TC/HDL-C ratio >5.0
Hs-CRP > 2 mg/L in men older than 50 years and in
women older than 60 years of age, irrespective of
LDL-C
Genest J et al: 2009 Canadian Cardiovascular Society/Canadian guidelines for the diagnosis and
treatment of dyslipidemia and prevention of cardiovascular disease in the adult- 2009
recommendations. Can J Cardiol 2009; 25(10): 567-579..

Pharmacotherapy (LDL Cholesterol):
Significant minority of patients will require combination therapy
• Ezetimibe (inhibits cholesterol absorption)
Additional LDL-C reductions of up to 20%
• Bile acid sequestrants (inhibits bile acid reabsorption)
Additional LDL-C reductions of 10-15%
• Niacin
Additional LDL-C reductions of up to 20%
Combinations are generally safe
Clinical outcome trials underway

Optional Secondary Targets when LDL-C at Target
Test Cut-point Intervention
TC/HDL-C >4.0 Fibrate
Niacin
Non HDL-C >3.5 mmol/L Fibrate
Niacin
Apo B/AI >0.8 Ezetimibe
Niacin
Triglycerides >1.7 mmol/L Fibrate
Niacin
hsCRP >2.0 mg/L Ezetimibe
Statin
Adjusting lipid-lowering therapy to optimize one or more of these
secondary targets may be considered in the high risk patient, after
achieving a target LDL-C or ApoB, but the clinical advantages of this
approach, with respect to patient outcomes, remain to be proven.
Genest J et al: Can J Cardiol 2009; 25(10): 567-579.

Important Take Home Points
Evidence favouring LDL reduction for the
prevention and treatment of atherosclerosis
is strong and compelling
New guidelines recommend a more
aggressive approach to management of both
high and medium risk patients
– Target LDL-C

CASE STUDY
A 63yo male diabetic patient was referred to the Victoria
Lipid Clinic in October 2008. An appointment was made
for May 2009.

Test Range Results:
Sep-08
TRIG

What would you do?
- Further analysis
- Intervention
- What are the targets?
- Other clinical findings
- Other diagnosis
- Would you have considered this an “Urgent” referral?

Test Range Results:
Apr-09
TRIG

What would you do?
- Further analysis
- What are the targets?

Test Range Results:
Jul-09
TRIG

What would you do?
- Further analysis

Test Range Results:
Aug-09
TRIG

What is the appropriate target?

Risk Categories and Treatment
Recommendations

What is Atherogenic Dyslipidemia?
Normal ratio of VLDL
to LDL particles in
plasma
Atherogenic
dyslipidemia –
increased VLDL and
sd-LDL
VL
DL
apo
B
sd-
LDL
norm
al
LDL
adapted from:
Sniderman et al
CMAJ Jan 9 2001; 164 (1)

Mortality in Statin Secondary Prevention Trials
Mortality (%)
10
9
8
7
6
5
4
3
2
1
0
4S
S40
4S
PL
CARE
P40
CARE
PL
LIPID
P40
LIPID
PL
HPS
S40
HPS
PL
non-CV
CV
TNT
A80
TNT
A10

Individual LDL-C Response to Ezetimibe 10mg
% change in LDL-C mmol/L
20
10
0
-10
-20
-30
-40
-50
-60
-70
-80
haplotype 2
mean = -23.6±1.6%
P=0.0054
non haplotype 2
mean = -35.9±4.0%
Subjects with NPC1L1 haplotype 2
(1735C-25342A-27677T)
Subjects not carrying NPC1L1 haplotype 2
Hegele RA et al. Lipids in Health and Disease 2005; 4:16

Effects of Ezetimibe as Mono or Combination Therapy over Any Time Frame on
Total Cholesterol in 141 Patients at the Victoria Lipid Clinic
100%
Fig 1: Total Cholesterol (A) and LDL-c (B) response and therapeutic catagories for individual patients
80%
Percent Change in Total Cholesterol
60%
40%
20%
0%
-20%
-40%
Individual Patients
Non-effective
Non-Responsive
Minimally Responsive
Moderately Responsive
Hyper-Responsive

Does one size fit all?

Unimodel for Cholesterol Metabolism
Synthesizers Mixed Absorbers
Hoenig MR, et al. Atherosclerosis 184 (2006) 247-254
Miettinen TA, et al. Atherosclerosis 164 (2002) 147-152

Reciprocal Model for Cholesterol Metabolism
Absorbers
S
y
n
t
h
e
s
i
z
e
r
s
Low Moderate High
High Moderate Low
Santosa S, et al. Life Sciences xx (2006) xxx-xxx
Harats D. XIV Int’l Symposium on Atherosclerosis 2006 Rome, Italy

Matrix Model for Cholesterol Metabolism
Absorbers
S
y
n
t
h
e
s
i
z
e
r
s
Low
Low
Low
Moderate
Low
High
Moderate
Low
Moderate
Moderate
Moderate
High
High
Low
High
Moderate
High
High

Percent Reduction in LDL by Statin negatively
correlates to percent reduction by Ezetimibe
80
% reduction in LDL-C by ezetimibe
60
40
20
0
-20
r = -0.29, p = 0.002
-20 0 20 40 60 80
% reduction in LDL-C by statin
Senaratne et al. Am Coll Cardiol, New Orleans March 2007

Effects of Ezetimibe Co-Administered with Statin on
Cholesterol Production, LDL-R Expression and LDL-C
Levels
Theoretical response
Relative change
Cholesterol Production
Inhibitor (Statin)
Cholesterol Absorption
Inhibitor (Ezetimibe)
Cholesterol Absorption
Inhibitor (Ezetimibe)
+
Cholesterol Production
Inhibitor (Statin)
Content Synthesis
LDL-R
LDL-C
Content Synthesis
LDL-R
LDL-C
Content Synthesis
LDL-R
LDL-C
Turley S.D. and Dietschy J.M., Preventive Cardiology 2003; Vol VI

Baseline Cholesterol Synthesis as a Predictor of
Recurrent Coronary Events and Response to Statins
Risk reduction with simvastatin not associated
with baseline cholesterol
? Is reduction of risk related to
– Baseline intestinal absorption
– Baseline endogenous cholesterol synthesis
4S Sub-study in 866 subjects
Measured Cholestanol / Cholesterol
– Index of absorption / synthesis
Miettinen et al. BMJ 1998; 316:1127

Simvastatin Did Not Reduce Major Coronary Events in the
Highest Quartiles of Cholesterol Absorption - 4S Sub-Study
40
38%
34% 25% 17%
Relative Risk
% Major coronary events
35
30
25
20
15
10
5
Placebo (n=434)
Simvastatin
20-40 mg (n=434)
0
1 2 3 4
Low Cholesterol
Absorption
4S = Scandinavian Simvastatin Survival Study.
High Cholesterol
Absorption
Miettinen et al. BMJ 1998; 316:1127

Achievement of target LDL-C concentrations:
In high risk individuals, treatment should be
started immediately, concomitant with diet and
therapeutic lifestyle changes. The primary target
of therapy is to achieve an LDL-C level of less
than 2.0 mmol/L. For patients with established
CAD, a reduction of LDL-C of at least 50% is
generally required to prevent progression or elicit
regression of atherosclerosis.

Target lipid levels:
The majority of patients, including those with
the metabolic syndrome, diabetes mellitus and
combined dyslipidemia, are able to achieve
target levels of LDL-C on statin monotherapy.
However, a significant minority of patients may
require combination therapy with an agent that
inhibits cholesterol absorption (ezetimibe) or
bile acid reabsorption (cholestyramine).

Ezetimibe and Cholesterol Balance
Conclusions
LDL-C response to both statins and ezetimibe is variable
A possible contributor might be the wide range of intestinal cholesterol absorption
Genetic variation appears to underlie the intestinal cholesterol absorption, ranging
from high to usual to low
Ezetimibe inhibits 17% (average) of intestinal cholesterol uptake (50% in animal
studies)
Variation in response to ezetimibe might reflect genetic difference in intestinal
absorption and /or expression of NPC1L1
High intestinal absorption might be associated with reduced synthesis – and
reduced response to statin
Poor response to statin may be associated with enhanced response to ezetimibe,
while enhanced response to statin might be associated with poor response to
ezetimibe.

Risk Level
Initiate Treatment if:
High
patients
CAD, PVD
Atherosclerosis
2009 Target Lipid Levels
Consider treatment in all
Most Pts with Diabetes
FRS>20%
RRS>20%
Moderate
FRS 10-19% LDL-C>3.5 mmol/L
TC/HDL > 5.0
hsCRP > 2
– men 50+, women 60+
Family history and hsCRP modulate risk
Low
FRS 5.0 mmol/L
Primary LDL-C
< 2 mmol/L
Or ≥ 50 % ↓ LDL-C
< 2 mmol/L*
Or ≥ 50% ↓ LDL-C
≥ 50% ↓ LDL-C
Primary
Alternate
Genest J et al: 2009 Canadian Cardiovascular Society/Canadian guidelines for the diagnosis and
treatment of dyslipidemia and prevention of cardiovascular disease in the adult- 2009
recommendations. Can J Cardiol 2009; 25(10): 567-579..
ApoB < 0.80 g/L
ApoB < 0.80 g/L

TRANSPORTERS
Bailey KM. Cytochrome p450 variants and response to statin therapy following Acute Coronary
Syndrome. [PowerPoint Presentation – LIGHT Research Meeting]. 2007

DRUG TRANSPORTER GENETICS
Influx Transporter
– Organic Anion Transporter Polypeptide B1
(SLCO1B1)
Efflux Transporter
– Breast Cancer Resistant Protein (BCRP)
– Single Nucleotide Polymorphisms (SNP’S)
» SLCO1B1 521 T>C
» BCRP 421 C>A

SPACE ROCKET
Evaluation
– 630 patient clinical trial
Rosuvastatin (10mg X 3mo)
BCRP CC (249) BCRP CA/AA** (75)
LDL-C 1.98mmol/L 1.78 mmol/L * *(p=0.012)
SLCO1B1 TT
SLCO1B1 TC/CC
LDL-C 1.90 mmol/L 2.01 mmol/L
**Note: The AA Genotype demonstrated an average of 20% reduction.
Romaine SP, et al. AHA Abstract 3447: Circulation 2008;118:S_426

Statins
Are effective in lowering cholesterol and
have established a role in both secondary
and primary prevention
The response of an individual to a given
statin can be determined within 6 weeks
The SNP’s for transporter genes can effect
the efficacy for certain statins
The drug and liver metabolite may have
different pharmacokinetic properties