Although the troponins remain the cornerstone of risk
stratification in acute coronary syndrome patients, investigators continue
to search for cardiac markers that can help identify a select group. That
is the 20 percent of cardiac troponin-negative unstable angina/non-ST
segment elevation ACS patients who will visit U.S. emergency departments
with chest pain and go on to experience a cardiac event within six months
of being diagnosed. Under investigation—alone and in conjunction
with one another—are a plethora of biomarkers involved in the atherothrombotic
process, including markers of hemostasis, platelet function, hemodynamic
stress, inflammation, ischemia, and necrosis.
A multi-marker strategy for risk stratification of ACS patients is not
recommended by cardiac marker guidelines, such as those being drafted
by the National Academy of Clinical Biochemistry, or NACB. However, the
academy’s draft laboratory medicine practice guideline on the use of biomarkers
in ACS and heart failure does allow for the possibility that multiple-marker
strategies could in the future yield better information for diagnosis
and risk stratification than the measurement of a single cardiac marker.
The new draft practice guideline, posted on the NACB Web site, www.nacb.org,
addresses the use of biomarkers across the spectrum of ACS and heart failure,
discussing analytical issues, diagnosis, risk stratification, and more.
The draft guideline says that for risk stratification of ACS patients,
cardiac troponin is the preferred marker and, if available, should be
measured in all patients with suspected ACS. In patients with a clinical
syndrome consistent with ACS, the draft guideline says, a maximal concentration
exceeding the 99th percentile of values for a reference control group
(with acceptable precision) should be considered indicative of increased
risk of death or myocardial infarction. (Using the 99th percentile implies
that there will be at least a one percent value above the 99th percentile
producing a false-positive result.)
This is the only class I recommendation made in the document, which divides
its advice into class I, class IIa, class IIb, or class III, based on
the recommendation classes of the American College of Cardiology and American
Heart Association. Class I represents conditions or recommendations for
which there is general agreement or evidence, or both, that a procedure
is useful or effective, while class IIa and IIb recommendations represent
conditions in which opinions diverge, with class IIa carrying a weight
of evidence or opinion in favor of the usefulness or effectiveness of
a procedure, and class IIb carrying a weight of evidence in which the
usefulness or efficacy of the procedure is less well established. Class
III signifies conditions for which a procedure is not useful or effective
and, in some cases, may be harmful.
In its recommendations regarding whether a multi-marker approach should
be used for risk stratification, the draft guideline says there is less
evidence for measuring hs-CRP, BNP, and NT-proBNP in addition to a cardiac
troponin for assessing risk in patients with a clinical syndrome consistent
with ACS than using cardiac troponin alone, but the document gives the
recommendation a class IIa nod. Measurement of markers of myocardial ischemia
in addition to cardiac troponin and electrocardiogram to aid in the short-term
risk stratification of patients with suspected ACS, and in excluding ACS
in patients with low clinical probability of ischemia, carries less evidence,
and was categorized as class IIb. Using a multi-marker strategy that includes
measuring two or more path o bio logi cal ly diverse biomarkers in addition
to cardiac troponin for risk stratification is also now a class IIb recommendation.
Yet the evidence base for adding markers to troponin
for risk assessment continues to grow. "The natriuretic peptides appear
to be strong markers for risk stratification in known acute coronary syndrome
patients and, given the evidence, might have even merit ed a class I recommendation.
However, the committee felt that the benefits of therapy based on elevated
BNP or NT-proBNP levels in the ACS population remain uncertain, and therefore
we felt that a class IIa recommendation was appropriate," explains Robert
Christenson, PhD, professor of pathology and director of rapid response
laboratories at the University of Maryland Medical Systems, Baltimore,
and chair of the group writing the practice guideline on the use of biomarkers
in ACS and heart failure.
"Generally speaking, in the ED right now, most people do not measure
BNP or NT-proBNP for chest pain unless there is associated atrial fibrillation
or symptoms of shortness of breath," says Alan Maisel, MD, professor of
medicine at the University of California, San Diego, and director of the
critical care unit in the heart failure program at the San Diego VA Healthcare
System. "However, before a patient presents with acute coronary syndrome,
when you have suspicion for underlying disease but plaque has not yet
been ruptured, BNP or NT-proBNP could potentially be part of the evaluation."
Recent data from the Platelet Receptor Inhibition in Ischemic Syndrome
Management (PRISM) trial published in the November issue of Circulation
(2004;110:3206-3212) showed that when measured at baseline and at
48 and 72 hours in 1,791 patients with non-ST-segment-elevation ACS, NT-proBNP
levels greater than 250 ng/L were associated with higher event rates at
30 days. In troponin T-negative patients, NT-proBNP identified a subgroup
of high-risk patients whose risk was comparable to that of troponin T-positive
patients. In addition, the researchers found that clinical stabilization
without refractory ischemia was associated with a rapid and significant
decline in NT-proBNP levels, while the lack of a rapid decline in NT-proBNP
levels in patients with baseline levels that were 250 ng/L or higher was
linked to an adverse short-term prognosis. Rising NT-proBNP levels were
also associated with an adverse 30-day prognosis.
A study underway at the SanDiego VA is demonstrating similar findings.
"In our experience, if we see BNP levels rise over a six- to 12-hour period,
we know that something is going wrong with the patient, even if the troponins
are negative," Dr. Maisel says. "In the first couple of hundred patients
that we’ve looked at in the study we are now doing, in the setting of
acute coronary syndrome, about 34 percent ended up developing clinical
CHF [congestive heart failure] in the hospital, and virtually every one
of those patients had BNP concentrations that rose in the first 12 hours
Biosite, the first company to bring BNP to market, is examining how the
assay can be used as part of a multi-marker strategy, but it has taken
a slightly different approach than most of the researchers who are looking
at such strategies. The company has been evaluating an index response
panel, called the CardioProfiler, which uses BNP in conjunction with traditional
cardiac markers for ACS diagnosis and prognosis.
Using an algorithm derived from thousands of values from ACS and noncardiac
chest pain patients, the CardioProfiler will provide a single index value
after analyzing a blood sample using BNP, CK-MB, troponin I and complexes,
and myoglobin assays. "What we have found," says Ken Buechler, PhD, president
and chief scientific officer at Biosite, "is that if we use a multi-marker
index, which is an algorithm that uses each one of the concentrations
of the analytes involved and calculates a single value, the multi-marker
index approach does a much better job at predicting early MI than individual
markers alone." Dr. Maisel, who is doing research with the CardioProfiler,
adds, "Within two hours of the onset of chest pain, data suggests that
the index could diagnose MI with about 94 percent sensitivity, whereas
troponin alone is only about 30 percent sensitive within two hours of
the onset of chest pain."
In addition to the natriuretic peptides, the National Academy of Clinical
Biochemistry includes high-sensitivity C-reactive protein as a potential
additional marker for risk stratification under a class IIa recommendation,
but Dr. Christenson has concerns about CRP’s lack of specificity when
used in the general ED population. "If you look at an isolated population
of ACS patients, hs-CRP will be predictive. However, when you start to
look at the entire population in which ACS is suspected, the test is nonspecific.
In addition, it’s not like D-dimer or IMA [ischemia-modified albumin],
where a negative value predicts low risk of disease. The data strongly
indicate that specificity is going to be a problem with general use of
hs-CRP," he says.
This doesn’t mean, however, that CRP would not add value to a multi-marker
panel for risk-stratifying ACS patients. "If you take troponin, CRP, and
BNP and use them in a multi-marker strategy," Dr. Maisel says, "each one
will give individual additional prognostic risk information."
While BNP, NT-proBNP, the troponins, and hs-CRP have
a growing body of literature that can be used to determine whether they
are ready for use in a multi-marker risk-stratification strategy, there
are other tests that show promise but need additional studies. Myeloperoxidase,
or MPO, a biomarker that has been reported to be useful in identifying
inflammation in the walls of coronary arteries and therefore risk for
heart disease or MI, has seemingly found a number of champions and may
be the next new cardiac marker to gain Food and Drug Administration approval.
Prognostix Inc., of Cleveland, has submitted an MPO test to the FDA for
review, while Abbott Diagnostics, Dade Behring, and Biosite all have MPO
assays in development.
Stanley Hazen, MD, PhD, section head of preventive cardiology and cardiac
rehabilitation at the Cleveland Clinic Foundation, and his colleagues
first published data on the prognostic value of MPO in 2003 (N Engl
J Med. 2003; 349:1595-604), which showed that the marker may be particularly
beneficial in chest pain patients who exhibit low initial troponin levels.
"We looked at initial MPO levels in more than 600 patients immediately
upon presentation to the emergency room with the complaint of chest pain,"
Dr. Hazen says. "By using MPO testing in addition to current laboratory-based
risk assessments, we increased our ability to identify subjects who experienced
a major adverse cardiac event over the next 30 days to six months from
54 percent to 85 percent of the time." An even larger study performed
in Europe (Circulation. 2003;108:e9034-9035) has confirmed these
"Myeloperoxidase is strongly and independently correlated with measures
of endothelial dysfunction in humans, which is consistent with it playing
a deleterious role by consuming nitric oxide in the artery wall," explains
Dr. Hazen. "MPO is markedly enriched in culprit lesions in subjects with
sudden cardiac death at sites of fissure or rupture of plaque and at superficial
erosions. Erosions as a site for intracoronary thrombus formation occur
to a greater extent in women and smokers, where something makes endothelial
cells on the surface of vulnerable plaque die and slough off, exposing
a prothrombotic surface. MPO-induced apoptosis of coronary artery endothelial
cells appears to play a role in that."
In recent published studies by Peter Libby, MD, PhD, and colleagues from
Brigham and Women’s Hospital, Harvard Medical School (Arterioscler
Thromb Vasc Biol. 2004:24;1309-1314), very low levels of MPO such
as those found within vulnerable atherosclerotic plaque, resulted in endothelial
cell apoptosis and increased tissue-factor expression. Tissue factor is
seen as key in the prothrombotic phenotype. Based on what researchers
are seeing, Dr. Hazen thinks MPO testing may have two potential uses in
ACS subjects. "One will be in identifying higher-risk groups, meaning
that if your MPO level is above a certain cutoff, it denotes that you
have a substantial increase in risk for having a heart attack, needing
bypass surgery, or dying in the near-term," he says. The other scenario
is to look at what low levels of MPO mean. "We are seeing a definite population
that has a low MPO concentration and an initial negative troponin, and
the negative predictive value for cardiac events is markedly improved
in this population versus those in whom only the troponins are measured,"
Dr. Hazen says, noting that the addition of a low MPO value to a negative
troponin cuts in half the number of missed cases of cardiac events over
the subsequent six-month interval after a patient is diagnosed with unstable
angina/non-ST-segment elevation ACS.
Says Biosite’s Dr. Buechler: "It’s clearly been demonstrated in the literature
that MPO is an important prognostic marker. It appears to predict bad
outcomes such as death or MI, and we are studying that in our discovery
process right now." Biosite licensed the MPO technology from the Cleveland
Clinic Foundation and its affiliate, Prognostix, about a year ago.
Abbott Diagnostics and Dade Behring recently announced that they have
agreements with the Cleveland Clinic Foundation and Prognostix to develop
MPO assays and are looking to bolster their cardiac marker offerings with
other emerging cardiac markers as well. "I think clinical research is
focused right now on plaque destabilization and rupture, and consequently,
we’re learning more about the role of emerging markers in these processes,"
says John Shaw, PhD, vice president of technology assessment at Dade Behring.
The markers they are pursuing for secondary risk stratification and specifically
for plaque vulnerability are MPO, placental growth factor, and sCD40L.
Though published studies examining sCD40L’s usefulness in risk stratification
have been few, those that have made it into the peer-reviewed literature
indicate it’s a promising marker, and it’s on the radar screen of many
cardiac marker researchers. "I really believe that sCD40L is going to
be a big player in risk stratification in the future," Dr. Christenson
says, "mainly because it gives you more information about platelets, which
many of the other emerging markers do not."
A recent study performed by Christopher Heeschen, MD, and his colleagues
at the University of Frankfurt in Germany, who have performed a number
of multi-marker studies in the risk stratification arena, demonstrated
the potential value of sCD40L when used in conjunction with yet another
emerging marker for risk stratification, pregnancy-associated plasma protein-A,
or PAPP-A, a zinc-binding matrix metalloproteinase that appears to be
expressed abundantly in eroded and ruptured plaques in ACS patients.
The study, published in the Journal of the American College of Cardiology
(2005;45:229-237), examined the predictive value of PAPP-A in 547
patients with angiographically validated ACS and in an emergency department
population of 644 patients with acute chest pain, in addition to troponin
T, vascular endothelial growth factor (VEGF), hs-CRP, interleukin-10,
and sCD40L. Patients were followed for the occurrence of death or myocardial
infarction. In patients with ACS, elevated PAPP-A levels (>12.6 mIU/L)
indicated an increased risk. When the analysis was restricted to TnT-negative
patients, PAPP-A still identified a subgroup of high-risk patients. In
a multivariable model, PAPP-A, sCD40L, IL-10, and VEGF were independent
predictors. Patients negative for TnT, sCD40L, and PAPP-A were at very
low cardiac risk.
Placental growth factor is just one of a variety of pro-inflammatory
markers, including sCD40L, Il-6, PAPP-A, MPO, and monocyte chemoattractant
protein-1, as well as anti-inflammatory markers, including IL-10 and activin
A, that have been suggested to provide relevant prognostic information
in ACS patients. "In the future," says Dade Behring’s Dr. Shaw, "we see
risk stratification going toward a panel of markers of inflammation combined
with troponin and NT-proBNP, and that’s why we’re looking at markers such
as placental growth factor, MPO, and sCD40L."
Abbott is investigating whole-blood choline as a potential new cardiac
marker. Few studies of whole-blood choline’s use in risk assessment have
been published, but an article in the American Journal of Cardiology
(2003;91:1060-1067) demonstrated that the biomarker could be useful
in risk stratification based on a prospective study of 327 patients with
suspected ACS that focused on the analysis of troponin-negative patients.
In the study, choline was measured using high-pressure liquid chromatography
mass spectrometry in whole blood, and patients were followed for 30 days.
In patients with negative troponin I test results at admission, choline
was a predictor of cardiac death and nonfatal cardiac arrest, life-threatening
arrhythmias, heart failure, and coronary angioplasty. In multivariate
analysis of troponin-negative patients, choline was the strongest predictor
of cardiac death or arrest.
Jessie Shih, PhD, associate research fellow for cardiovascular at Abbott
Diagnostics, says Abbott is looking at other new cardiac markers as well.
"Cardiac is a growing area for us, and we are actively investigating new
markers as they are identified in press or as people contact us," she
says. Adds Robert Olson, Abbott’s scientific affairs director for cardiac,
"While troponin is very good, there is still a need to find an early ischemia
marker, and that’s a major focus for us as well."
The only marker for ischemia on the market is ischemia-modified albumin
from Ischemia Technologies, which Dr. Christenson says is another marker
that can be explored for risk stratification. Its most efficacious use
at the moment appears to be in ruling out ACS. "There’s compelling data
that IMA’s negative predictive value is very good, and that the test will
probably be clinically useful. The literature is still developing, however,
on how exactly to interpret a positive test," Dr. Christenson says.
For risk stratification, Drs. Christenson and Maisel
believe that the future is in multi-marker panels. "They’ll have to be
markers that reflect the spectrum of the physiology of acute coronary
syndrome, and we’ll have to find the best mix," Dr. Christenson says.
In the meantime, he adds, it’s important to continue to standardize troponin
I assays and to continue to move toward troponin I tests that can dem
on strate acceptable precision at the 99th percentile of values for a
reference control group.
The American Association of Clinical Chemistry’s troponin I standardization
subcommittee is collecting data from a third round-robin event that will
aid standardization. "I would expect in the next year or so that there
will be a big push to standardize troponin I assays," Dr. Christenson
As for the status of the NACB’s guideline on markers of ACS and heart
failure, a second draft is scheduled to be placed on the academy’s Web
site early this spring, and the guideline probably will be finalized this
summer. To date, at least a dozen groups have signed on to the guideline,
with 25 or so others being represented by the National Heart Attack Alert
What does the immediate future hold for the multitude of emerging risk-stratification
markers for ACS? "The data are out there for using multiple markers, and
I think we are seeing different utilities for each of the markers," says
Abbott’s Dr. Shih. "Some of them may be more useful for shorter-term risk
stratification, and some are more useful further out. But what we need
to do is to get more robust assays out there and understand their clinical
utility better with larger multi-site studies."
Yes, the plethora of emerging markers for ACS risk stratification is
exciting, but the true test will come, Dr. Christenson says, when clinicians
begin using them in the clinical setting. "A multi-marker strategy is
definitely coming. But no matter what stage we get to," he cautions, "there
will still be an art to using cardiac markers for risk stratification,
because the markers will always have to be interpreted in the clinical
context of the patient."
Sue Parham is a writer in Edgewater, Md.