It’s all relative: Genetic dyslipidaemia
CVD risk factors, and especially lipid metabolism, exemplify gene / environment interactions
Mainly genetic
– Co-dominant mutations: Either genetic allele affected – Recessive mutations: Both genetic alleles affected
– Polymorphisms and SNPs: Alleles consistent with “normal” and
markers in proximity to significant genetic effects. Genome-wide
association studies (GWAS). Mainly environmental or secondary to other
disorders.
A missing piece of the genetic puzzle
Clinical Effect
Gene Prevalence
Common genes with small effect
Autosomal dominant
Autosomal recessive
Uncommon genes with large effect
Polymorphisms
Pathogenic mutations
Genetic hyper and hypo - cholesterolaemia
• Dominant Monogenic Hypercholesterolaemia• Familial hypercholesterolemia (FH)• Familial defective apo-100 (FDB-100)• PCSK9 gain-of-function (FH-3)• Hyperalphalipoproteinaemia (CETP deficiency, non-atherogenic?)
• Recessive Monogenic Hypercholesterolaemia• Autosomal recessive hypercholesterolemia (ARH)• Lysosomal acid lipase deficiency: Wolman’s disease and
Cholesterol ester storage disease
• Dominant Monogenic LDL deficiency• PCSK9 loss-of-function
• Recessive Monogenic LDL deficiency• Abeta and hypobeta – lipoproteinaemia• Chyomicron retention disease
Familial Hypercholesterolaemia:What goes wrong?
NORMAL FH
LPL
LIVER
INTESTINE
B
Chy
lom
icro
ns
CII
LPL R
E
B
CII
LDL-R
LRP
-48
IDL
B
E
VLDL
B
ECIII
-100
B
CIII
HL
BOxLDL
O.
AI
AIAII
ESR-B1
HDL
Macrophage
ABCA1
LCAT
OtherTissues
CE
FC
FH
SR-ASR-B1
LDLAI
Metabolic Defect in Familial Hypercholesterolemia
DAVIGNON 2006
LPL
LIVER
INTESTINE
B
Chy
lom
icro
ns
CII
LPL R
E
B
CII
LDL-R
LRP
-48
IDL
B
E
VLDL
B
ECIII
-100
B
CIII
HL
BOxLDL
O.
AI
Macrophage
ABCA1 SR-A
LDLAI
AII
ESR-B1
HDL
LCAT
OtherTissues
CE
FC
SR-B1
AI
Metabolic Defect in Familial Defective ApoB-100
FDB
ApoB
Defective apoB B
DAVIGNON 2006
PCSK9 regulates the surface expression of LDLRs by targeting for lysosomal degradation
1. Qian YW, et al. J Lipid Res. 2007;48:1488-1498.2. Horton JD, et al. J Lipid Res. 2009;50:S172-S177.3. Zhang DW, et al. J Biol Chem. 2007;282:18602-18612.
Gain-of-Function Mutations in PCSK9 Cause Familial Hypercholesterolaemia (FH)
• Associated with:• High serum LDL-C2
• Premature CHD and MI2
• In vitro testing in many identified mutations show decreased levels of LDLRs3
*For a full list of ADH mutations, please see refer to Abifadel reference.
1. Abifadel M, et al. Hum Gen. 2009;30:520-529.2. Horton JD, et al. J Lipid Res. 2009;50:S172-S177.3. Cameron J, et al. Hum Mol Genet. 2006;15:1551-1558.
PCSK9 Variant Population Clinical Characteristics
D374YBritish, Norwegian
families, 1 Utah family
Premature CHD
Tendon xanthomas
Severe hypercholesterolemia
S127R French, South African, Norwegian families
Tendon xanthomas; CHD, early MI, stroke
R215H Norwegian family Brother died at 31 from MI; strong family history of CVD
What is “FH”? What does it cause?
FH is
– Co-dominant mutation of genes affecting formation or function of the LDL-receptor
– This causes metabolic and clinical consequences including precocious cardiovascular disease (CVD)
Metabolic– Increased LDL, – Reduced clearance of remnants including LDL’s
precursor, IDL.– Increased Lp(a)?– Reduced HDL?
Clinical– Dominant: 50% of each generation. Risk 50:50– Premature CHD, CVD and PVD– Aortic stenosis– Tendon xanthomas (11%) specific?– Corneal arcus (27%) non-specific > 40y?– Xanthelasmas (12%) nonspecific – No signs highly sensitive – FH IS NOT JUST HIGH CHOLESTEROL IN A
PATIENT AND THEIR RELATIVE(S)
FH: Why is it important?
0.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1.0
25-3
0
30-3
5
35-4
0
40-4
5
45-5
0
50-5
5
55-6
0
60-6
5
65-7
0
70-7
5
75-8
0
80-8
5
85-9
090
+
Age
Cu
mu
lati
ve P
rob
abili
ty o
f C
linic
al C
AD Non-FH Women
Non-FH Men
FH Women
FH Men
MED PED Registry 2001.
Why FH matters: Prevalence and Impact
Prevalence: – 0.2 – 0.5% (1 : 200-500)
Atherosclerosis 173:55-68
– > 8 x 106 affected world-wide. Seminars in Vasc Med 4:87-92
– > 40,000 Australians
– Equal numbers of unaffected relatives
– Up to 1:60 in local groups with “founder effect”.
– Detection rates 0 – 44%• World’s best 20-40%• World average (including Australia)
< 5%
Impact: – 5 – 10% of CHD events under
age 60. J Lipid Res 34:269-77
– CVD death in >80% of FH cases.
– Absolute CVD risk differs from general population models.
• High risk profile from birth, • Interaction differs (smoking, gender)
Circulation 97:1837-47
• Risk algorithms underestimate risk Eur Heart J 19:A2-11
– Missed and misdiagnosed
Molecular Medicine meets Public Health
Clinical protocol
Laboratory protocol
Cascade Screening
Clinical services
Detecting index cases
Genetic testing
Management
Diagnosis assessment
Diagnostic criteria
Children,Adolescents
Adults
Children,Adolescents
Adults
Process
Optimal components
Model of Care for
FH
Process
LDL-Apheresis
Case detection: Dutch Lipid
Clinic Score?
Severe triglyceride elevation due to recessive impairment of lipoprotein lipase
Chylomicrons persist after fasting, massive levels of TG. Homozygous deficiency of Lipoprotein Lipase (LPL) Homozygous deficiency of the cofactor for L PL, Apo CII Impaired transport of LPL to site of action (endothelium) due to homozygous
defect in ANGPTL or GPIHBP Combined overproduction and undercatabolism of triglyceride-rich lipoproteins
sufficient to saturate LPL, eg in Apo AV mutations.
LIVER
INTESTINE
B
Chy
lom
icro
ns
CII LPL
CRE
BCII LDL-R
LRP
-48
IDL
B
E
VLDL
B
ECIII
-100
BLDL
CIII
HL
BOxLDL
O.
Metabolic Defects affecting Lipoprotein Lipase
MacrophageABCA1 SR-A
-48
Dietaryfat
TG
TG
TG
ApoB-48 R
LPL
VLDLR
AI
DAVIGNON 2006
AIAII
ESR-B1
HDL
LCAT
OtherTissues
CE
FC
SR-B1
AI
FFA + MG
AVINTESTINE
B
Chy
lom
icro
ns
CII
LPL CRE
BCII
LRP
-48
IDL
B
E
TG richVLDL
B
ECIII
-100
B
CIII
HL
BOxLDL
O.
SR-B1
Metabolic defects saturating in LPL, eg Apo AV mutations
Macrophage
ABCA1
-48
Sugarfat
calories
LPL
AI
Overproductionof VLDL
sdLDL
ADIPOSETISSUE
aGP + FFATG
Normalor reduced
VLDLcatabolism
LDLR
LOX-1
CD-36
SR-A
SR-PSOX
HSL
CETPTG CE
AV
AIAII
EHDL
LCAT
OtherTissues
CE
FC
SR-B1
VLDLR
DAVIGNON 2006
AI
■ Normal■ Hypoalpha- lipoproteinemia
■ Complete (FHC) or partial LPL deficiency associated with a secondary factor
■ Complete LPL deficiency (FHC)■ Primary apoC-II deficiency
■ Familial dysbeta- lipoproteinemia (type III)■ Hepatic lipase deficiency■ (Primary cause associated with a secondary factor)
Apo B > 0.75 g/L
Apo B < 0.75 g/L
TC:Apo B > 6.2
TC:Apo B < 6.2
■ Familial hyperTG■ Partial LPL deficiency
■ FH■ Polygenic■ FDB■ PCSK9deficiency■ ARH deficiency■ CYP7A1deficiency■ Hypoalphalipo-proteinemia
■ FCH■ β-Sitosterolemia
Normal Chylo + VLDL Chylo Chylo +
VLDL Remnants
VLDL LDL VLDL + LDL
NormoTG< 1.5
mmol/L
TG:Apo B > 0.12
NormoApo B< 1.2 g/L
TG:Apo B < 0.12
HyperTG> 1.5
mmol/L
NomoTG> 1.5
mmol/L
HyperApo B> 1.2 g/L
Hyper TG> 1.5
mmol/L
PrimaryCauses
Algorithm for Diagnosis of Apo B DyslipoproteinemiasAlgorithm for Diagnosis of Apo B Dyslipoproteinemias
Abbreviations: apo, apolipoprotein; ARH, autosomal recessive hypercholesterolemia; CAPD, continuous ambulatory peritoneal dialysis; Chylo, chylomicrons; CP7A1, cytochrome P450 7A1; DM2, diabetes mellitus type 2; dysbeta; dysbetalipoproteinemia; FCH, familial combined hyperlipidemia; FDB, familial defective apoB; FH, familial hypercholesterolemia; FHC, familial hyperchylomicronemia; HAART, highly active antiretroviral therapy; LPL, lipoprotein lipase; PCOS, polycystic ovary syndrome; SLE, systemic lupus erythematosus; TC, total cholesterol; TG, triglyceride. de Graaf J et al. Nat Clin Pract Endocrinol Metab 2008;4:608-
18.
Lipoproteins
Autosomal recessive disorders have revealed HDL metabolism
A-I
FC
A-I
ABCA1
MacrophageRapid catabolism
LCAT
Nascent HDL
CE
FC
Homozygous (?heterozygous) Apo AI deficiency: AtherogenicTangier’s Disease: ABC-AI deficiency: Atherogenic?LCAT Deficiency: Non-atherogenic?Apo A1 Milano: Anti-atherogenic?
+
100
50
030 40 50 60 70 80
Age ( years)
Eve
nt F
ree
Sur
viva
l ( %
)
apoAI (L178P) carriers(n=54)
family controls(n=147)
p = 0.008
Carriers of the ApoAI Leu178Pro Variant Are at Increased Risk of Developing CAD
Hovingh K et al. J Amer Coll Cardiol 44:1429, 2004
ApoAI 50%HDL-C 63%
18.9 x CAD risk
DAVIGNON 2006
Normal ApoAI and ApoAIMILANO Dimer
1
25
35
66 121 165209 220
243
18714399
Lipid Binding In Vivo Catabolism
LCAT Activation Cholesterol Efflux
“Receptor” Binding
AI
Franceschini G Eur J Clin Invest 26; 733, 1996
AIm/AIm
1 1
243243
173173s s
DAVIGNON 2006
Role of apolipoprotein E
Apo E: ligand for hepatic removal of remnants.
Apo E knockout model is atherogenic
Apo E2 has lower binding affinity (E4>E3>E2).
E2:E2 only critical if lipids increase for other reasons.
Other roles for Apo E (CNS lipid transport and neural repair).
CR
E
E
AI
CHYLOMICRON
CIIITG
TGTG
OOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOO
OOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOO
LPLLRP
AII
C
E
E
E
AI CEHDL
B 48 B 48
CII
Lipolysis products
C
FC AI
SR-B1
CEFC
AI
AII
ECE
AIFC
AI
AII
ECE
HLCE
Kidney
AI
HDL3
HDL2
HL
CE
ECEAI
HDL3
LCAT
LCAT
FC/PL
FC/PL
AII
AI
Metabolic Defects in Remnant DyslipidaemiaApo E2 homozygosity plus apo B overproduction or Hepatic Lipase deficiency
LIVER
CELL
ABCA1
VLDL to
Remn
BE
CE
CETP
Chyloto
ChyloRe
BE
LPL
PLTP
INTESTINE
LDLB
HL
Degradation (catabolic) Formation (anabolic)
CETG
LDLR
TG
OxLDL
B
LRP
Modified fromDeeb SS et al. J Lipid Res 44:1279, 2003
Unspecified hereditary Dyslipoproteinemia
Common:
Lipoptotein (a)
Rare:
Familial Phytosterolemia
DAVIGNON 2006
ABCG5ABCG8Micelle
sPhytosterols
Cholesterol
ProteaseK VK IVTypes 3-10
Apo(a)
LDL Receptor Binding
Site on apoB
K IVType 2
ApoB
LDLparticle
K IVType 1
C
N
-C
-N
DAVIGNON 2006
Genetic dyslipidaemias have motivated treatment discovery
• Clinical abnormalities represent real human problems• Massive yield on research into genetic dyslipidaemia
• Familial Hypercholesterolaemia: Receptor mediated endocytosisStatins, PCSK9
antisense and Abs
• Familial Hyperchylomicronaemia: LPL gene therapy
• Apo A1 Milano: Synthetic HDL
• Deficiency of Apo B or MTP MTP inhibitors, Apo B antisense
• CETP Deficiency CETP inhibitors
• Apo E Valuable animal k/o model
IT’S ALL RELATIVE : MANAGEMENT OF GENETIC DYSLIPIDAEMIA
CASE 1
AAS MASTERCLASS5-6TH OCTOBER 2012
• Referred to Specialist Llipid Clinic
• Hypercholesterolaemia• ‘Yellowish butterfly patterned
lesion’ around her eyes• On Pravastatin 20mg ON• TC 13.7, LDL-c 11.6 mmol/L
July 1997
• Cycling accident at her home town• Sustained minor soft tissue injury• Noted to have xanthelasma around her
eyes• Lipid profile results:
• TC : 15.4 mmol/L • LDL: 13.9 mmol/L
• Referred to Medical Clinic, home town• Started Pravastatin 20 mg/ON
6th August 1998
HISTORY…
Miss KM, 21year old Malay student nurse
Noted ‘yellowish butterfly patterned lesion’ surrounding both eyes since primary school days
Asymptomatic, well No h/o chest pain, palpitation, shortness of breath or poor effort
tolerance No history of intermittent claudication or syncopal attacks. No h/o polyuria, polydipsia No h/o cold intolerance, lethargy or menstrual disturbance No past h/o jaundice, liver or renal diseases Not HT, DM No past surgical history
HISTORY……
- HISTORY…
The youngest out of 11 siblings 3 siblings died :
eldest brother : died @ 43 years – AMI (HC)elder brother (6th) : died @ 23 years – AMI (HC,
xanthelasma)eldest sister (2nd) : died @ 33 years - ?MI, ? SCD
Parents - consanguinous marriage (first cousins), not known to have DM/HPT/CAD
- HISTORY…
Student nurse at a Teaching Hospital, KL Single Non smoker No history of alcohol intake Exercise - once weekly (jogging), 30minutes Normal diet, low fiber intake
Physical ExaminationAnthropometry:• BMI : 22.1• Waist circumference : 61cm (<80cm)• Waist-to-hip ratio: 0.71 (<0.85)
• PR: 72/min, regular, BP : 120/62 mmHg, DRNM• Peripheral pulses – present, carotid & renal bruit :
absent• Other systems: Normal
Xanthelasma
Corneal Arcus grading:0 – no arcus observable1- upper or lower segments affected2- both segments affected3- both segments and just confluent4- heavy confluent arcus
(Wider et al, 1983)
Right Eye
Left Eye
Grade 4 Grade 4
Corneal Arcus
Achilles tendon xanthomata
Digital xanthomata
Test Result Reference range
**FSL Baseline(1997)
1st visit to the SLC
TC 15.4 13.7 < 5.7 mmol/L
HDL-c 1.5 1.6 > 1.3 mmol/L
LDL-c 13.9 11.6 <3.8 mmol/L
Triglycerides 1.0 1.1 < 1.3 mmol/L
On pravastatin 20 mg / ON x 1 year
Summary of KM’s risk factors
Positive risk factors: Negative risk factors:
Markedly elevated TC & LDL levels
Strong family history of premature CAD
HDL > 1.3 mmol/L Non smoker Not hypertensive Not DM Not overweight/ obese Premenopausal female
Differential Diagnosis
1° Hypercholesterolaemia 2° Hypercholesterolaemia - TRO
Familial hypercholesterolaemia (FH)
Hypothyroidism
Familial Defective ApoB100 Cholestasis
PCSK9 gain-of-function mutation (FH-3)
Nephrotic syndrome
Polygenic hypercholesterolaemia
Tests Results Reference Ranges
TFT
fT4 11.78 9.10 – 23.8 nmol/L
TSH 1.22 0.32- 5.00 Iu/L
FPG
Glucose 4.4 <6.1 mmol/L
Renal Profile
Sodium 134 135 – 150 mmol/L
Potassium 4.2 3.5 – 5.0 mmol/L
Urea 3.2 2.5 – 6.4 mmol/L
Creatinine 66 44-80 umol/L
Tests Results Reference Ranges
LFT
Albumin 42 35-50 g/L
Total Protein 84 67-88 g/L
Bilirubin Total 16 < 23 umol/L
ALT 17 <44 U/L
ALP 89 32 – 104 U/L
Creatine Kinase 42 24-135 U/L
Lp(a) 0.58 <0.3g/L
OTHER INVESTIGATIONS….
ECG – normal Exercise stress test – normal ECHO – Normal, no evidence of aortic stenosis Carotid artery IMT – normal, no evidence of atheromatous
plaques
Question 1
What are the various criteria for the diagnosis of FH?
• Dutch Lipid Clinic Network diagnostic scoring• Simon Broome’s criteria• MedPed criteria for FH• NCEP ATPIII criteria
Question 2
According to the Dutch Lipid Clinic Network criteria, scoring for definite FH is:
• > 8 points• 6 – 8 points• 3 – 5 points
In determining the Dutch Lipid Clinic diagnostic scoring for FH, the following are taken into account:
• Baseline LDL-c concentration: Yes/ No• Clinical history of premature CAD: Yes/ No• Clinical history of premature cerebral or PVD: Yes/ No• Tendon xanthoma(ta) in the patient: Yes/ No • Premature corneal arcus in the patient: Yes/ No• Family history of hypercholesterolaemia: Yes/ No• Family history of premature CAD/PVD in 1st degree relatives: Yes/ No• Family history of tendon xanthomata and/or corneal arcus in 1st degree
relatives: Yes/ No
Question 3
Dutch Lipid Clinic – Diagnostic scoring for FH
Criteria:• 8 points - DNA Mutation, or LDL-C
> 8.5mmol/L• 6 points - Tendon xanthomas• 5 points - LDL-C 6.5 – 8.4mmol/L• 4 points - premature corneal arcus
< 45 yrs• 3 points - LDL 5.0 – 6.4mmol/L• 2 points - 1st degree relative with
xanthomas or premature CA or childhood LDL > 95th percentile, or personal premature CAD
• 1 point - 1st deg relative with premature CAD/ vascular dis or LDL > 95th percentile, or personal history of premature cerebral or PVD, or LDL-c 4.0 – 4.9mmol/L
Criteria:Family history:• 1st degree relative with (a) premature CAD or
vascular dis (men<55yrs, women<60yrs) OR (b) LDL > 95th percentile, in - 1 point and / or
• 1st degree relative with tendon xanthomata and/or corneal arcus OR childhood (<18yrs) LDL > 95th percentile - 2 points
Clinical history• Patient with premature CAD (men<55, women
<60yrs) - 2 points• Patient with premature cerebral or PVD
(men<55, women <60yrs) -1 point
Physical Examination - patient• Tendon xanthomas - 6 points• premature arcus - 4 points
Lab Analysis – • LDL-c >8.5mmol/> - 8 points• LDL-c 6.5 – 8.4 - 5• LDL-c 5.0 - 6.4 - 3• LDL-c 4.0 – 4.9 - 1
DNA analysis• Functional DNA Mutation - 8 points
Definite: > 8 points, Probable: 6 – 8 points, Possible FH: 3-5
US MedPed Criteria vs Simon Broome criteria for the dx of FH
• Total cholesterol cutpoints (mmol/L)• 1st-degree vs 2nd-degree vs 3rd-degree relatives with FH vs
General population
• Age (years) 1st-degree 2nd degree 3rd degree General population
• <20 5.7 5.9 6.2 7.0• 20–29 6.2 6.5 6.7 7.5• 30–39 7.0 7.2 7.5 8.8• ≥40 7.5 7.8 8.0 9.3
• FH is diagnosed if TC levels exceed the cutpoint• Key: FH = familial hypercholesterolaemia
Simon Broome Criteria - Diagnosis of FH
Criteria Description• (A) TC > 7.5 mmol/L in adults or TC > 6.7 mmol/L in children <16
years, or LDL-c > 4.9 mmol/L in adults or > 4.0 mmol/L in children• (B) Tendon xanthomas in the patient, or a first-degree or second-
degree relative • (C ) DNA-based evidence of mutation in the LDLR, or apo- B100
or PCSK9 gene• (D) Family history of premature CHD (age <50 years in a
second-degree relative or <60 years in a first-degree relative)• (E) Family history of raised TC >7.5 mmol/L in a first- or second-
degree relative, or >6.7mmol/L in child, brother or sister <16yrs of age
Diagnosis• Definite FH diagnosis requires either A + B or A + C • Possible FH diagnosis requires either A +D or A + E• Key: FH = familial hypercholesterolaemia
• Hypercholesterolaemia (LDL-c 13.9mmol/L)• Xanthomata, corneal arcus, xanthelasma• No evidence of personal CAD • Strong family history of premature CAD• Positive family history of HC – 1st and 2nd
degree relative• Consanguity• Ruled out secondary causes of HC
Summary
CLINICAL DIAGNOSIS
DEFINITE FAMILIAL HYPERCHOLESTROLEMIA
Question 4:
Should risk estimation tools eg Framingham Risk Scoring be used for her?
• YES• NO
CHD Risk Stratification• CHD estimation tools such as those based on the Framingham
risk scoring SHOULD NOT be used because people with FH are already at high risk of premature CHD.
• Heterozygous FH has >50% risk of CHD in men by the age of 50years, >30% in women by the age of 60years
NICE Clinical Guideline 2008- Identification and Mx of FH
• 2.5.1 Individuals with FH are at high CHD risk. The 10-year CHD risk in the FH patient is not adequately predicted by any conventional risk assessment tools. Therefore, assessment of 10-year risk is not recommended.
Management
Statin therapy - continued, titratedAdd-on lipid loweringLifestyle modificationReferred to dieticianReferred to :
- Cardiology
- Plastic surgeryFamily tracing, cascade screening and counselling
Medication TC (mmol/L)
TG HDL LDL
Baseline, statin naive 15.4 1.0 1.5 13.9
Pravastatin 20 mg /ON 13.7 1.1 1.6 11.6
Pravastatin 20 mg/ON 11.9 1.1 1.0 10.4
Simvastatin 30 mg/ON 13.1 1.1 1.4 11.3
Atorvastatin 80 mg/ON 15.3 1.4 1.2 13.5
Atorvastatin 80 mg/ON 11.7 0.7 1.26 10.1
Atorvastatin 80 mg/ON 11.3 0.7 1.1 9.8
Atorvastatin 80 mg/ON 11.9 0.6 1.4 10.2
Atorvastatin 80 mg/ON 11.5 1.1 1.2 9.8
Simvastatin 80mg & Ezetimibe 10mg/ ON
8.9 1.2 1.1 7.3
Alternative or add-on lipid lowering therapy
• Ezetimide• Bile acid binding resin• Fibric acid derivatives• Nicotinic acid• LDL-apharesis
LIFESTYLE MODIFICATION
Lifestyle modification - reduce LDL-c and other coronary RFs Modest, variable degree of LDL reduction N 10%Diet – low fat <30% calories, saturated fat < 10% of calories,
cholesterol < 200mg/day, fibre 10-25g/day, caloric deficit 300-500kcal/day
Plant sterol esters or plant stanol esters 2g/dayPhysical activity – 30min/day, moderate intensity, at least 5days
per week
Ideal weight BMI < 23 (Asian)SmokingAlcohol intakeHT, DM
1 32 4 5 6 7 8 9 10 11
63
62
Father Mother
FAMILY TREE - 1998
43
3340
39 3
8
23
23
32
33
26
21
HCCAD+
SD ?MI HCXanthoma
HCXanthoma
HCXanthoma
HC Xanthelasma
XanthelasmaCAD+
HC
Those found to have HC following family screening
1 32 4 5 6 7 8 9 10 11
TC : 8.1LDL : 5.7
TC : 8.7LDL : 6.7
TC : 10.1LDL : 6.7
TC : 8.7LDL : 6.5
TC : 8.7LDL : 6.5
TC : 7.7LDL : 4.6
TC : 8.0LDL : 6.4
TC : 16.2LDL : 14.8
TC : 6.5LDL : 4.6
TC : 7.0LDL : 5.2TC : 8.1
LDL : 5.7
TC : 4.9LDL : 2.6
TC : 15.3LDL : 13.9
Father Mother
Family Screening ( Lipid Screening) - 1999
Affected with FH
DiedTC : 6.1LDL : 4.4
FH with known mutation
43
33
4039 38 23
33 32 26 23 21
Genetic testing was performed on KM and her family members
Question 5:
What genes have been implicated in FH?
• LDL Receptor• Lipoprotein lipase• Apo B100• Apo CII• PCSK9
Low Density Protein Receptor (LDLR) Gene
• Cytogenetic Location: 19p13.2
• Size: 44,469 bases• Mosaic protein of ~840 amino acids (after removal of
signal peptide); Molecular weight 95,376 Dalton
Made up of a number of functionally distinct domains that can function independently of each other.
Ex 1 contains a signal sequence that localizes the receptor to the ER for transport to the cell surface
Ex 2-6 code the ligand binding region
Ex 7-14 code the EGFP domain
Ex 15 codes the oligosaccharide rich region
Ex 16 (and some of 17) code the membrane spanning region
Ex 18 (with the rest of 17) code the cytosolic domain.
LDLR PROTEIN
DHPLC RESULTS – LDLR Exon 5
g.763T>A
g.763T>A
Homoduplex peak of NC sample (wild type sample)
Heteroduplex peaks of FH patients with variant in exon 5
Heteroduplex peaks of FH patients with variant in exon 5
Mutation Screening by DHPLC
Wash peak
The DHPLC chromatogram profiles of FH patients showed presence of heteroduplex peaks eluted at 5.2 mins and 5.8 mins at denaturing temperature of 62.8 0C. The presence of heteroduplex peaks were suggestive of disease-causing variants and subjected to DNA sequencing to confirm the variants.
DNA SEQUENCING RESULTS
FH Patient ID
Affected LDLR regions
DNA Sequence Description of variant and effect
KM, SFM &
WC (3
members of a
Malay Family)
Exon 5 Reference :GCCGGCAGTGTG
ACCG
Variant :GCCGGCAGAGTG
ACCG
g.763T>A; Cysteine (C) to Serine (S);
Homozygous and Heterozygous
mutation C234S
NYK Exon 9 Reference :
CTTCACCAACCGGCACG
Variant :
CTTCACCAACTGGCACG
g.1216C>T; Arginine (R) to Tryptophan
(W), Heterozygous mutation R385W
DNA SEQUENCING RESULTS
Affected LDLR region
DNA Sequence Description of variant and effect
Exon 5 Reference: GCCGGCAGTGTGACCGVariant : GCCGGCAGAGTGACCG
g.763T>A; Cysteine(C) to Serine (S) at codon 234. Identified as homozygous mutation C234S.
Reference sequence from normal control
DNA sequence of Homozygous FH patient
Homozygous:Substitution T>A at nucleotide position 763 (g.763T>A)
DNA SEQUENCING RESULTS
Affected LDLR region
DNA Sequence Description of variant and effect
Exon 5 Reference: GCCGGCAGTGTGACCGVariant : GCCGGCAGAGTGACCG
g.763T>A; Cysteine(C) to Serine (S) at codon 234. Identified as heterozygous mutation C234S.
Reference sequence from normal control
DNA sequence of Heterozygous FH patient
Heterozygous:Substitution T>A at nucleotide position 763 (g.763T>A)
KM is now married with 2 children
Questions 5:
What is the probablity that her children may have FH with every pregnancy?
• 50%• 25%
Questions 6:When should diagnostic tests be done for her children?• At birth• By the age of 10 years or earliest opportunity
thereafter
• AD, one affected parent – 1:2 (50%) chance of heterozygous with every pregnancy
• In children with one affected parent, the following Dx-tic tests should be done by the age of 10yrs or at the earliest opportunity thereafter:
- DNA test if the family mutation is known- LDL-c if the family mutation is not known. To exclude the Dx of FH,
LDL-c should be repeated after puberty - LDL-c concentration changes at puberty to almost similar to that of adult concentrations
• Cascade testing – combination of DNA testing and LDL-c to identify affected relatives
• In children at risk of homozygous FH (2 affected parents or presence of clinical signs eg cutaneous xanthomata), measure LDL-c before 5yrs of age or at the earliest opportunity thereafter. If LDL >11mmol/L, clinical Dx of homozygous FH should be considered
NICE Clinical Guideline 2008- Identification and Mx of FH
Question 7:Who can be considered for LDL
apheresis treatment?
Adults and children/young people with • Homozygous FH• Heterozygous FH
LDL lowering apheresis
• Treatment of adults and children/ young adults with homozygous FH
- In children with 2 affected parents- Presence of clinical signs eg cutaneous lipid deposits
(xanthomata)- LDL-c should be measured before 5 yrs of age, or earliest
opportunity thereafter- LDL-c > 13mmol/L (adults), > 11mmol/L (children/ young
adults) – consider clinical diagnosis of homozygous FH
• Exceptional cases of heterozygous FH –progressive symptomatic CHD despite maximal tolerated lipid-modifying drug therapy, optimal medical and surgical therapy
• Specialist centre• Cost implication
Question 8:What are the future lipid lowering
therapy?
• Antisense oligonucleotides (ASO) to inhibit Apolipoprotein B production –eg Mipomersen
• PCSK9 targeted therapy –eg PCSK9 inhibitor• Microsomal TG Transfer Protein Inhibitors –eg lomitapide• Thyroid Mimetics – eg eprotirome• Cholesterol Ester Transfer Protein (CETP) Inhibitors -
anacetrapib, dalcetrapib
(1) ANTISENSE OLIGONUCLEOTIDES (ASO) – MIPOMERSEN
• Subcutaneous 200mg 1x/wk – phase 3 CT, LDL reduction lasted for 4/52 after last dose. No clinically relevant interactions wrt clearance of statins and ezetimide - impt as add-on therapy
Yu et al Clin Pharmacokinet 2009;48:39-50
• Homo FH (n=51) - Recent double blind CT, sc mipomersen 200mg/wk vs placebo x26wks – 25% vs 3% LDL reduction, 27% reduction apoB, 21% reduction TC
Raal FJ et al. Lancet 2010;375:998-1006
• Hetero FH (n=124), CAD+ on max tolerated statins: 45% of these high risk subjects achieved target LDL of < 100mg/dL (2.6mmol/L)
EAS Congress 2011
2. PCSK9 TARGETED THERAPY
• Animal studies – beneficial up to 80% LDL lowering, human studies not published yet
Frank-Kamenetsky et al. Proc Natl Acad Sci USA 2008;105:11915-20
Chan JC et al. Proc Natl Acad Sci USA 2009;106:9820-5
• Statins and fibrates induce increase PCSK9 expression, thus PCSK9 inhibition could induce robust LDL reduction as add-on therapy in FH
3. MICROSOMAL TG TRANSFER PROTEIN (MTP) INHIBITORS
• MTP inhibition with BMS-201038 in homozygous FH (n=6), dose 1mg/kg/day - 50% LDL reduction but noted to induce hepatic steatosis
Cuchel et al. New Engl J Med 2007;356:148-56
• MTP inhibition in homozygous FH (n=10) , dose 60mg/D – 44% LDL reduction; less steatosis
Cuchel et al.Circulation 2009;120:S441
• Ezetimibe 10mg vs MTP inhibition by lomitapide (5-10mg/ D x4/52) – 20% vs 20-30% in a dose-dependant manner
• Combined therapy (ezetimide + lomitapide) – similar but larger dose dependent LDL reduction (35-45%); SE: Mild ALT increase, diarrhoea
Samaha et al. Nat Clin Pract Cardivasc Med 2009;2010:906-16
• Potential attractive candidate for lipid lowering in FH patients if administered in lower doses
4. THYROID MIMETICS
• Thyroid hormone analogues – reduce LDL but associated with cardiac and bone related SEs
• More recently differential molecular mechanisms (cpds that act on TR-beta mainly expressed in the liver, do not affect TR-alpha expressed in brain and heart) – eg eprotirome, selective TR beta agonists (under Ix)
• Statin + placebo/ eprotirome for 12/52 – 20-30% additional reduction of LDL; no SE on heart or bone
Ladenson et al. Use of thyroid hormone analoque eprotirome in statin-treated dyslipidaemia. New Engl J Med 2012;362:906-16
5. CETP (CHOLESTEROL ESTER TRANSFER PROTEIN) INHIBITORS
• X2 approaches to inhibit CETP• (a) Vaccine - CETi-1 synthetic CETP peptide vaccine, immune response anti
CETP- Abs• Animal studies of this vaccine – increase HDL, reduce aortic atherosclerosis
but human studies poor response of autoAB productionRittershaus et al. Vaccine induced Abs inhibit CETP activity in vivo and reduce aortic lesions in a rabbit model of atherosclerosis, ATVB 2000;20:2106-12
Davidson et al. The safety and immunogenicity of a CETP vaccine in healthy adults. Atherosclerosis 2003;169:113-20
(b) Small molecule CETP inhibitors – eg Torcetrapib, anacetrapib, dalcetrapib - antagonize CETP activity by binding to it.• RADIANCE I – 800 FH pts, torce + atorva: No reduction in atherosclerosis
(IMT) despite reduction in LDL (20%) and increase HDL (52%)
Kasteleine JJ et al. Effect of torcetrapid on carotid atherosclerosis in FH. NEJM 2007;356: 1620-30
• ILLUMINATE – torce + atorva, prematurely terminated, unexpected increase in M&M – exact mechanisms unclear ?torce induced increase BP
Barter et al. Effects of tercetrapid in patients at high risk of coronary event, NEJM 2007;357:2109-22
Xie et al. Drug discovery using chemical systems biology:identidfication of the protein-ligand binding network to explain the side effects of CETP inhibitors. PLoS Comput Biol 2009;5:e1000387
• 2 other CETP inhibitors have no effect on BP – molecule-specific off-target effect
• Anacetrapid and dalcetrapib – phase 3 clinical trials , mild HC and pts with CVD risk, effective lipid modifiers
• Pending CV outcome trials (DAL-outcomes I and II and HPS3/REVEAL)
• CETP inhibition likely to benefit patients with FH
5. CETP (CHOLESTEROL ESTER TRANSFER PROTEIN) INHIBITORS
Cont…….
Summary
• A 21year old Malay female student nurse who presented with incidental xanthelasma and severe hypercholesterolaemia associated with a strong family h/o premature CAD, xanthomata and grade 4 corneal arcus. There was consanguity between her parents. She is slim, non-smoker, euglycaemic, normotensive and has good HDL-c levels (>1.3mmol/L). Family tracing was performed. Molecular analyses confirmed the presence of homozygous FH with exon 5, position 763 T>A mutation of the LDLR, coding for the LDL binding domain.
• The updates on the present and future management, and the molecular pathogenesis of FH have been discussed.
Learning Issues
• FH criteria for diagnosis• High coronary risk category, should not use risk
assessment tools• LDL Receptor protein and gene• Genetic testing – when to test for children?• Lifestyle modification• Current treatment, targets• Plasma apheresis – when is it indicated?• Future lipid lowering therapy
