A brief Note on Cardiac Biomarkers past, today and future persepctive

1. CARDIAC BIOMARKER
Dr Anurag Yadav
PostGraduate

2. BIOMARKER
“A biomarker is a substance used as an
indicator of a biologic state”.
Accurate repeated measurements at
reasonable cost
Must provide additional information
Should aid treatment

3. CARDIAC BIOMARKERS
Cardiac biomarkers are protein molecules
released into the blood stream from damaged
heart muscle
Since ECG…… inconclusive ….biomarkers
!!!!!?????
• myocardial injury
These biomarkers have a characteristic rise
and fall pattern

4. CHARACTERISTICS
High cardiac specificity
Pharmacokinetics of cardiac biomarker
Easy diagnosis
Marker should play a designed role in the
treatment and management of clinical subject

5. HISTORY OF CARDIAC BIOMARKERS
1954 - SGOT (AST)
1955 - LDH
1960 - CPK
1972 - CPK isoforms by Electrophoresis
1975 - CK - MB by immunoinhibition
1975 - Myoglobin
1985 - CK - MB Mass immunoassay
1989 - Troponin T
1992 - Troponin I

6. CLASSIFICATION OF CARDIAC
BIOMARKERS
Biomarkers of myocardial injury
• markers of myocardial necrosis
• markers of myocardial ischemia
Biomarkers of haemodynamic stress
Inflammatory and prognostic Biomarkers

7. CLASSIFICATION
OF
CARDIAC BIOMARKERS
ACCORDING TO VARIOUS
STAGES
DURING CARDIAC DISEASE
PROCESS

9. BIOMARKERS OF MYOCARDIAL INJURY
Markers of myocardial necrosis
• Creatine kinase – MB
• Myoglobin
• Cardiac troponins
Markers of myocardial ischemia
• Ischemia Modified Albumin (IMA)
• Heart-type fatty acid binding protein (H-
FABP

10. Marker for
inflammation
hsCRP sCD40L Homocystein
Marker for plaque
Destabilization
PAPP-A LP-PLA2
Marker for
hemodynamic
stress
Natriuretic peptides
• ANP
• BNP
• Pro-BNP
• CNP
• DNP

11. Time of
increase
Peak Return to
normal
CK-MB 4-8 h 12-24h 72-96h (3-4D)
LDH 2-5 D 10 D
Myoglobin 2-4 h 8-10h 24 h
cTnI 4-6 h 12 h 3-10 D
cTnT 4-6 h 12-48 h 7-10 D

12. 0
1
2
3
4
5
6
7
0 4 8 12 16 20 24 28 32 36 40 44 48
myoglobin
CK-MB
cTnT
cTnI
COMPARISON OF CTN, CK-MB , MB
Time after onset of AMI (hours)
Χupperlimitofreferenceinterval

13. OTHER CARDIAC MARKERS

14. Stefan Blankenberg, MD; Renate Schnabel, MD; Edith Lubos, MD, et al., Myeloperoxidase Early Indicator of Acute Coronary Syndrome and
Predictor of Future Cardiovascular Events 2005

17. TROPONIN ASSAYS
TropT (Roche Diagnostics, Germany) ; Trop I (Siemens Healthcare
Diagnostics)
Troponin T
• 99th percentile limits - 0.01 ng/mL
• assay ranges - 0.01-25 ng/mL
(Troponin I)
• 99th percentile limits -0.04 ng/mL
• assay range -0.04-40 ng/mL
Reference limits based on the 99th percentile for a healthy
population are 0.01 ng/mL (Troponin T) and 0.04 ng/mL (Troponin I)

18. Creatine kinase
(CK) is a
cytosolic enzyme
CK is a dimer composed of two subunits B (brain type)
and M (muscle type), resulting in three isoenzyme:
CK-BB (CK1) : is of brain origin, found in blood only
when BBB is damaged.
CK-MB (CK2) : it is relatively specific for myocardial
origin
CK-MM (CK3) : it is found primarily in skeletal muscle
CK-MB

19. CK-MB :
it is a valuable tool for the diagnosis
of MI because of its relative high
specificity for myocardial damage.
Rise : 4-6 hrs after onset of
symptoms
Peak : 12 hrs
Return to normal : 24-36 hrs
Can be used to indicate early re-
infarction if level normalizes and
then increases again.

20. CK-MB
Relative Index = χ 100
Total CK
*The relative index allows the
distinction between increased total
CK due to myocardial damage and
that due to skeletal or neural
damage.
*A relative index exceeding 3 is
indicative of AMI

21. Small-size heme protein found in all tissues mainly assists in oxygen
transport
It is released from all damaged tissues
Its level rises more rapidly than cTn and CK-MB.
Released from damaged tissue within 1 hour
Normal value: 17.4-105.7 ng/ml
Timing:
• Earliest Rise: 1-4 hrs
• Peak 6-9 hrs
• Return to normal: 12 hrs
Myoglobin

22. CONDITIONS FOR MYOGLOBIN
INCREASE :
Acute myocardial infarction
Skeletal muscle damage, muscular
dystrophy, inflammatory myopathies
Renal failure, severe uremia
Shock and trauma

23.  Clinical usefulness of myoglobin :
*if myoglobin concentration remains within the reference range
8 hours after the onset of chest pain, AMI can be ruled out
essentially.
*because of its rapid clearance by the kidney, a persistently
normal Mb concentration will rule out reinfarction in patient
with recurrent chest pain after AMI
*Rapid monitor of success of thrombolytic therapy
DRAWBACKS
 Due to poor specificity, myoglobin levels do not always predict
myocardial injury

24. 0
1
2
3
4
5
6
7
0 4 8 12 16 20 24 28 32 36 40 44 48
myoglobin
CK-MB
cTnT
cTnI
COMPARISON OF CTN, CK-MB , MB
Time after onset of AMI (hours)
Χupperlimitofreferenceinterval

25. Ischemia Modified Albumin (IMA)
A novel marker of ischemia, is produced when circulating serum
albumin contacts ischemic heart tissues
IMA can be measured by the albumin cobalt binding (ACB)
assay that is based on IMA's inability to bind to cobalt
Mechanism- due to structural change in the amino terminal end
of albumin
IMA levels rise within 6 hours
remain elevated for 12 hours

26. Drawbacks
IMA levels raised in non- cardiac ischemia
Modification to n- terminal end may also be induced
by extracellular hypoxia, acidosis etc,
Conclusion
FDA in 2010 has approved a multimarker approach for
using the combination of ECG, the cTnI, and the IMA levels
achieving a sensitivity of 95% for ACS

27. Heart type fatty acid binding protein is a very stable low
molecular weight (14-15kDa) in the cytoplasm of myocardial
cells.
FABPs are involved in active fatty acid metabolism where it
transports fatty acid from cell membrane to mitochondria for
oxidation.
Small size of H-FABP facilitates rapid diffusion through
interstitial space, appearing as early as 1-3 hrs after onset and
peaking within 6hrs. It return to normal levels with in 12-24hrs.
Normal levels : 1.6 – 19 ng/ml
H-FABP

28. NATRIURETIC PEPTIDES
The natriuretic peptides (NP) are a group of
structurally similar but genetically distinct peptides.
NPs are identified as regulatory diuretic-natriuretic
substances responsible for salt and water
homeostasis
Lowers blood pressure.

29. The NP family
includes
ANP : -atrial
natriuretic peptide
(28 a.a.)
N-terminal
proANP (98 a.a.)
BNP : brain
natriuretic peptide
(32 a.a.)
N-terminal
proBNP (76 a.a.)
CNP : C-type
natriuretic peptide
(22 and 53 a.a.)

30. Fig. Schematic representation of the ANP and BNP precursors with
sequence numbering defining low-molecular-mass forms, N-terminal forms
and high-molecular-mass precursors

31. OTHER CARDIAC MARKERS

32. Homocysteine
Intermediary amino acid formed by the conversion of
methionine to cysteine
Moderate hyperhomocysteinemia occurs in 5-7% of the
population
Recognized as an independent risk factor for the
development of atherosclerotic vascular disease and venous
thrombosis
Can result from genetic defects, drugs, vitamin deficiencies,
or smoking

33. Homocysteine implicated directly in
vascular injury including:
• Intimal thickening
• Disruption of elastic lamina
• Smooth muscle hypertrophy
• Platelet aggregation
Vascular injury induced by leukocyte
recruitment, foam cell formation, and
inhibition of NO synthesis
Homocysteine

34. Elevated levels appear to be an independent risk factor,
Screening recommended in patients with premature CV
disease (or unexplained DVT) and absence of other risk
factors
Treatment includes supplementation with folate, B6 and
B12
Homocysteine

35. C-REACTIVE PROTEIN
CRP is an acute-phase protein produced by the liver
Pentameric structure consisting of five 23-kDa identical
subunits
Plasma levels can increase rapidly to 10000x levels
High-sensitivity CRP (hs-CRP) assays

37. CRP previously known to be a marker of high risk in
cardiovascular disease
More recent data may implicate CRP as an actual
mediator of atherogenesis
Mechanism of CRP-mediated atherogenesis:
Once ligand-bound, CRP can:
• Activate the classical compliment pathway
• Stimulate phagocytosis
• Bind to immunoglobulin receptors
• Endothelial dysfunction via ↑ NO synthesis
• ↑LDL deposition in plaque by CRP-stimulated macrophages

40. Clinical Uses
• Screening for cardiovascular risk in otherwise
“healthy” individuals
• Predictive value of CRP levels for disease
severity in pre-existing Coronary artery disease
Elevated levels predictive of
• Long-term risk of first MI
• Ischemic stroke

41. Limitations of CRP
Low specificity
No evidence that lowering CRP levels decreases CV risk
• Industry and FDA staff guidelines 2005 had given clinical cut off value
as less than 1 mg/l as safe levels with hs-CRP tests
CRP Risk for CVD
Less than 1.0 mg/L Low
1.0-2.9 mg/L Intermediate
Greater than 3.0 mg/L High

42.  miRNAs are appx. 20-25 nucleotide long non coding RNAs, that negatively
regulate or inhibit gene expression by binding to sites in the untranslated
regions of targeted messenger RNAs.
miRNA

43.  miRNA are found to be involved in almost every
biological process, from cellular differentiation and
proliferation to cell death and apoptosis
 Many different types of miRNA can be detected in
circulating blood and these miRNA are present in
remarkably stable form that even withstand repetitive
freezing/thawing cycle and are protected against Rnases.
 Thousands of miRNAs have been described in human to
date which ehibits tissue specific pattern of expression.

44.  miRNAs that regulates cardiovascular system can be
divided into 4 groups :
1. miRNA regulating endothelium function and
angiogenesis : miR126, miR17-92 cluster, miR130a,
miR221, miR21
2. cardiac myocyte specific mRNA : miR208a
3. cardiac myocyte and skeletal muscle miRNA : miR1,
miR133a, miR499
4. smooth muscle miRNAs :miR143, miR145
miRNAs hold promise as very specific and accurate
marker of cardiac dysfunction.

45. CYTOKINES
Variety of interleukins (TNF, IL-1, IL-6, IL-8, IL-12, IL-
18) are responsible for the atherosclerosis events.
Regulated through the T cell-mediated and
monocytes.
At present there are no standardized assay,
reference interval studies.

46. MYELOPEROXIDASE (MPO)
Released from aggregated neutrophils.
Increased in CAD, ACS.
Is one of the best prognostic marker
At present no standardized assay method, nor consistency validation
of the test.
Further, Type of specimen collect have shown variations.
Normal level: 640 pmol/L

47. LIPOPROTEIN ASSOCIATE
PHOSPHOLIPASE A2 (Lp-PLA2)
Previously known as PAF ( platelet activating factor)
Synthesised from monocytes and lymphocytes.
Known to form lipid fragments which are most
atherogenic which increase endothelial adhesion.
FDA approved for the assay.
Threshold for high risk: >200μg/L

48. PREGANACY ASSOCIATED
PLASMA PROTEIN A (PAPP-A)
It is an metaloprotimase.
Produced from the plaque which are prone to rupture.
Marker for the adverse CV events.
At present no standardized assay, consistency validation
is present.
By two approach, median value for free PAPP-A is 0.18
mIU/L.

49. CHOLINE
Is released after stimulation by phospholipase D.
Touted as a test of prognosis in pt with chest
discomfort.
Not available

50. UNBOUND FREE FATTY ACID
(FFA-U)
Marker for ischemia.
Ischemia increase the small unbound
fraction.
Studies revealed mixed result.
Not available

51. NOURIN
Small protein released by stressed
myocytes.
Induce changes of inflammatory cytokines
and attract neutrophils.
Preliminary studies have made attempt to
validate its use.

52. 30 a.a glycoprotein.
Prognostic marker in hemorrhagic and sepsis.
Recently, early biomarker for rule out the AMI with
normal cTn.
The test method has to undergo many trials and
head-to-head comparison with cTn.
COPEPTIN