IVD Technology Magazine
IVDT Article Index

Originally published March, 1998

Cardiac Markers: Developers enter a new era of progress

Greg Freiherr

Troponin-based diagnostics have revolutionized coronary care, say the experts—but wait till you see what IVD manufacturers come up with next.

Photo Courtesy of Simon Fraser/Science Photo Library

To the medical community's ongoing war against heart disease, the IVD industry brings to bear a legion of biochemical markers designed to speed clinical diagnoses. For physicians, these diagnostics are the essential tools they need to make a difference in their patients' health outcomes.

At the head of the diagnostic ranks are the troponins—proteins that reside on the heart's thin filament of striated muscle fibers, the myocardium, and are released when it is injured. These markers represent the latest advance in the treatment of cardiac patients, vastly increasing specificity and speed of diagnosis. Farther back in the ranks are the traditional markers creatine kinase (CK), CK-MB, myoglobin, and other proteins traditionally associated with myocardial infarction (MI). And on the diagnostic horizon are cardiac markers already known to the research community but whose clinical significance has only begun to be appreciated.

"What we have learned in the past year about the science of acute coronary syndrome represents an exponential growth of knowledge," says Robert Jesse, MD, PhD, director of acute cardiac care at the Virginia Commonwealth University/Medical College of Virginia in Richmond. "We are on the edge of being able to develop biochemical markers that will give us the information we really want to know."

But these same markers are also the ammunition for a struggle within the ranks—a battle among diagnostic corporations to achieve supremacy in a rapidly evolving diagnostic regimen. Three companies—Boehringer Mannheim Corp. (Indianapolis), Dade Behring (Deerfield, IL), and Spectral Diagnostics (Toronto)—are already selling more than a half-dozen products that challenge the position of CK as the standard-bearer of cardiac markers. Even more companies, including Biosite Diagnostics (San Diego), Abbott Laboratories (Abbott Park, IL), Quantech, Ltd. (St. Paul, MN), and Centocor (Malvern, PA) seem poised to enter this market very soon.

At stake are the welfare of some 6 million patients who complain of chest pain each year, a share of some 16 million cardiac tests conducted annually in the United States, and revenues that account for about a half-billion dollars in health-care charges each year.

Troponins

The principal candidates currently vying to capture that revenue are tests based on the troponins. Two of these proteins are standouts—troponin T and troponin I. Both are released after myocardial injury and can therefore serve as specific indicators of myocardial damage.

"Clearly, in the last couple of years, both of the troponins have been very exciting to laboratorians and cardiologists because of their specificity and sensitivity," says Derek P. LeHane, PhD, vice president for clinical and scientific affairs at Dade Behring.

That company, by virtue of the megamerger of Dade International and Behring Diagnostics completed late last year, currently has the widest range of products for assaying cardiac markers. In its sales efforts, the company has placed increasing emphasis on its troponin I tests that run on three different instruments—the Dimension, Stratus, and Opus analyzers.

Figure 1. Boehringer Mannheim's CardiacT assay is small enough to be used at a patient's bedside. The device provides a qualitative measure of troponin T that can be used to rule in or rule out AMI. Photo Courtesy of Boehringer Mannheim

But the pioneer in this area is Boehringer Mannheim, which first commercialized troponin testing in late 1995 with an assay that identifies very small amounts of troponin T. Called CardiacT, the handheld assay can be used at emergency room bedsides to rule in or rule out acute myocardial infarction (AMI) as well as to assist in risk stratification of unstable angina patients (see Figure 1). It can also be used in coronary care units (CCUs) to rapidly identify myocardial damage. The rapid assay requires just a few drops of whole blood dropped into a sample well. The assay is then inserted into a test card and sealed. Minutes later, red lines appear, indicating whether troponin T is present; two lines indicate a positive result, one line a negative.

"Troponin T is not expected to be elevated unless there's some cardiac condition, so a positive or negative, plus or minus, absence or presence answer is sufficient," says Terry Prewitt, product manager for CardiacT at Boehringer Mannheim.

In 1996, the company brought a second-generation CardiacT assay to market for use in its Elecsys immunoassay analyzer. Later developments led to the assay's being adapted for use on other of the company's analyzers.

Despite Boehringer Mannheim's success with troponin T, some competitors, including Dade Behring and Spectral Diagnostics, have chosen to base their assays on troponin I. Spectral Diagnostics offers the Cardiac Status troponin I panel assay, a rapid-format panel test meant for point-of-care use.

Although clinical data show similar effectiveness for the two troponins, a recent study published in the New England Journal of Medicine found that troponin I, assayed by the Spectral Diagnostics test, was more sensitive to both MI and unstable angina than was troponin T, assayed with the Boehringer Mannheim CardiacT.¹ The authors nevertheless concluded that both types of tests are highly sensitive for the early detection of myocardial-cell injury in acute coronary syndromes. They also found that negative test results are associated with low risk for cardiac damage, enabling physicians to discharge patients from the emergency room rapidly and safely.

More Needs, More Markers

Beyond the two front-runners, there are a number of other assays from which clinicians can select. While clinical studies point to the improved effectiveness of both troponin T and I over standard-bearers CK and CK-MB, long years of service by these enzymes have created a familiarity that diagnosticians are not readily inclined to cast off.

But current protocols based on biochemical tests involving CK and CK-MB fail to identify more than half the patients with MI. Conventional cardiac enzyme tests together with ECG results still miss about 25% of MI patients. Even troponin testing, which represents the best near-term answer, is difficult to interpret, because tests made by different vendors deliver widely different readings.

"There is no standardization for troponin assays," says Scott Mader, cardiovascular product director at Biosite Diagnostics, one of the firms close to commercializing a new test for troponin I. "A troponin level of 0.4 for one assay may actually be equivalent to a 1.2 for another assay."

Recognizing the shortcomings of diagnostic tests, physicians have to rely on their acumen at sizing up patients. In the end, physicians tend to err on the side of caution, leading to more admissions than necessary. Of the estimated 700,000 patients hospitalized each year with suspected AMI, about 75,000 are later found not to have suffered this cardiac event. The cost of admitting a patient to a CCU is estimated at about $30,000.

"The CCU is one of the most resource-intensive, high-cost-per-day areas of the hospital," notes Douglas Ball, chairman and CEO of Spectral Diagnostics. "Rapid and accurate diagnosis is where you can avoid cost, because so many of the patients admitted to CCUs are later determined to be experiencing nothing more serious than heartburn or a noncardiac event."

More distressing than the cost incurred by being cautious, however, is the unknown number of patients that are sent home when they should have been hospitalized. Estimates of the percentage of AMI patients inappropriately discharged from the emergency room range from 5 to 13%. Of these, between 10 and 26% subsequently die. These cases account for about 20% of the emergency department costs incurred in the emergency room. "Earlier diagnosis of a cardiac patient should lead to better and earlier treatment and, as a result of earlier treatment, better outcome in terms of saved lives or better quality of life," Ball says.

The diagnostic quandary surrounding cardiac patients has spurred the search for new biochemical markers that promise greater specificity or sensitivity than the gold standards CK and CK-MB.

Figure 2. Levels of key cardiac markers—myoglobin, CK-MB, and troponin I—in the hours following AMI. Source: Dade Behring

Myoglobin, which stores and transports oxygen, is attractive because of its early release into the bloodstream, showing up two to four hours after the onset of chest pain. Speed of treatment is a major concern in the ER (see Figure 2). Dissolution of a clot that is blocking oxygen meant for the myocardium—which can be accomplished with such agents as streptokinase—must be done early to have maximum effectiveness, potentially salvaging tissue that would otherwise be lost. Underscoring the need for speed is the 30% of patients who die within one hour of the first signs of MI. Unfortunately for diagnosticians, myoglobin is not specific to cardiac injury. The protein is present in skeletal muscle and can be released as the result of an injury unrelated to MI. It is very useful, however, in ruling out MI.

Troponin T also offers quick measurement, appearing as early as three hours after the onset of chest pain. However, this regulator of muscle contraction can also be expressed as the result of regenerating muscle tissue such as occurs after a muscle injury, and elevated levels can appear as the result of renal disease. High concentrations of the protein have also been reported in patients with pneumonia. "If a patient has experienced a trauma or condition during which there is a release of skeletal muscle contents, there would be a cross-reaction between the cardiac and the skeletal troponin T," says Joseph H. Keffer, MD, vice president and chief medical officer at Spectral Diagnostics, which has chosen to base its newest diagnostics on troponin I.

Despite its drawbacks, clinicians consider troponin T a major indicator of MI and a useful tool for risk assessment of unstable angina patients. The peer-reviewed literature is replete with clinical documentation of its utility for both applications. Peak elevation occurs about 12 hours after the cardiac event, and the level remains elevated for up to seven days.

Troponin I is very specific to cardiac damage. An isoform of troponin I is found only in myocardial tissue. Primary applications for this protein are as an indicator for myocardial damage and for risk stratification of unstable angina patients. Troponin I appears early—about four hours after the onset of chest pain. Like troponin T, troponin I levels peak in about 12 hours and remain elevated up to seven days. Although troponin I is very specific to cardiac tissue damage, it exhibits a relatively low diagnostic sensitivity.

Cardiac Panels

Concerns over sensitivity and specificity have led some developers of cardiac assays to construct panels that include several biochemical tests. Spectral Diagnostics offers a point-of-care panel test combining CK-MB and myoglobin. Called Cardiac Status Controls, it is designed as a definitive rule-out test to assist in the early diagnosis of an evolving heart attack (see Figure 3). It is, however, a biochemical dinosaur, likely to be made extinct by the growing popularity of troponin testing.

Figure 3. The Cardiac Status troponin I assay by Spectral Diagnostics, a qualitative point-of-care test for elevation of troponin I levels following AMI. Photo Courtesy of Spectral Diagnostics

"Many physicians have to go from the familiar to the unfamiliar in a fairly gradual way," says Keffer. "They can't just leap into troponin; some need CK-MB and some need myoglobin because that is what they are comfortable with." Putting troponin into the mix is a logical next step, although Keffer stops short of saying that Spectral Diagnostics will be doing that. Other companies are in the process of doing so.

Dade Behring is developing the Stratus CS Stat fluorometric analyzer, which will quantitatively measure troponin I, myoglobin, and mass CK-MB. The product is designed for emergency departments and other locations where quick and effective diagnosis is required. It is also suited for low-volume cardiac testing performed in a clinical laboratory.

Biosite Diagnostics may be close to obtaining FDA clearance for its Triage Cardiac System, a handheld point-of-care device about the size of a telephone that quantitatively measures CK-MB, myoglobin, and troponin I (see figure 4). FDA has already cleared one part of the system, the Triage Cardiac Panel. "The philosophy is that there is no perfect marker, and that each one of these has specific value and augments the measures of the others," says Biosite's Mader.

Figure 4. The Triage Cardiac System quantitates CK-MB, myoglobin, and troponin I. Photo courtesy Biosite Diagnostics. Photo Courtesy of Quantech, Inc.

The ultimate in testing flexibility may be the Quantech system, which uses surface plasmon resonance to quantify biochemical markers (see Figure 5). Measurements are made using the interaction of light photons with electrons in a metal. The metal—in this case gold—is part of a disposable mini-tray, with an antibody coated on the surface and a patient sample in a receptacle. When an antigen such as CK-MB binds to the antibody, the wavelength of the light changes in direct correlation to the amount of antigen. The shift in wavelength between the initial and final readings provides a quantitative result that promises extraordinary precision.

Figure 5. Quantech's test system uses surface plasmon resonance to quantify the levels of biochemical markers, including key markers for AMI.

"We believe that the critical-care areas would like a product that gives practitioners quick, quantitative results, because there is a high value to quick diagnosis," says George Dodge, Quantech director of finance. Dodge notes that the company is working on CK-MB, myoglobin, and troponin I. "We believe we will be able to quantify all three."

Faster, Closer

Each of the new tests on the market—as well as the ones nearing commercialization—offer fast processing. No more than 15 to 17 minutes elapse from the time of sampling to result. This immediacy, accompanied by the compactness of the assays—either handheld or the size of a desktop computer—makes point-of-care testing a viable possibility in the ER as well as the CCU. Putting these tests in the hands of physicians, however, changes the conduct of medicine and displaces many of the functions that have traditionally been assigned to clinical laboratories.

"It's always difficult when you deal with point-of-care testing because most of the regulations and inspections are centered around laboratories," notes David Quick, Elecsys marketing manager at Boehringer Mannheim. "That's the biggest challenge for point-of-care systems."

Most companies are meeting this challenge by deferring to the customer's preference, not favoring any locale for use of the test but instead letting the customer decide. The same technology being sold for point-of-care use is applicable to laboratory testing. And because analyzers are becoming more compact and easier to use, some institutions are choosing to place them in or near ERs and CCUs.

According to Spectral Diagnostics chairman and CEO Ball, all of these changes are part of a shifting dynamic in the practice of medicine. "We are seeing a migration of the marketplace. The division between emergency services and the clinical lab is breaking down, and there is increasing evidence that clinical chemistry practitioners are becoming more inclined to embrace point-of-care diagnostics," Ball says.

Even so, the expertise found in clinical labs may still be needed to properly interpret test results, which are becoming increasingly sophisticated. Up to now, companies have sought simply to educate prospective customers about the advantages of troponin and how to read individual tests. But as panels combining several analytes enter the market, practitioners will need to be schooled in how to interpret the measurements in context with one another. "There is no magic bullet," says Quick. "It's still a physician's call."

The Next Wave

The challenges for physicians and laboratorians seem destined to become even greater over time. Already companies are experimenting with the next wave of products to be introduced. These analytes could radically reduce the time needed to make a diagnosis.

Among the most promising near-term products is glycogen phosphorylase isoenzyme BB (GPBB), which appears to be specific to myocardial injury. GPBB demonstrates a rapid rise to detectable levels in patients with infarction—within two to three hours of chest pain onset—according to on-line literature published by Advanced ImmunoChemical Inc. (Long Beach, CA).² The company supplies antibodies, antigens, enzymes, and purified proteins to the IVD industry as well as academia.

Patients and their care-givers have the most to gain from analytes that allow diagnosis closer to the cardiac event. One of these analytes is P-selectin, a marker indicating platelet activation. Biotechnology innovator Centocor has been studying these markers for several years. Citing company policy, executives refused to be quoted for the record, but the company is known to be focusing on activated platelets as the means for both diagnosis and treatment, as in the case of Centocor's latest therapeutic, ReoPro.

The advantage of looking at platelets is that accompanying markers are not dependent on a by-product of necrosis, but rather are intimately involved in the earliest stages of myocardial infarction. P-selectin, which is just one of a class of markers being examined by Centocor, might be detectable within minutes of the beginning of a cardiac event. Another possibility is to find markers of fibrin activation, which is also involved in the early stages of infarction. Either type of marker promises to give physicians the boost they need.

"Quite simply, markers of necrosis—which are the current state of medical practice—develop too late for use on most of the patents we are seeing," says clinical cardiologist Robert Jesse.

But within 5 to 10 years, he believes, clinicians may have markers that will identify not only the activation of platelets and the generation of fibrin rupture of plaques—which indicate that coagulation has begun and that a clot may soon form—but even the presence of so-called "vulnerable plaques," which cause coagulation to occur.

"Today, everyone in clinical medicine is focusing on ruling in or ruling out MIs," Jesse says. "What you really want to know is how to identify the person who has acute coronary syndrome or unstable angina in the absence of necrosis."

In the not-too-distant future, he says, diagnostics manufacturers may make that possible.

References

1. Hamm CW, Goldmann BU, Heeschen C, et al., "Emergency Room Triage of Patients with Acute Chest Pain by Means of Rapid Testing for Cardiac Troponin T or Troponin I," N Engl J Med, 337(23):1648—1653, December 4, 1997.

2. Access via http://www.advimmuno.com.