CDU Contributing Editor

CHICAGO, Illinois – In vitro diagnostics is becoming increasingly important in the practice of medicine today, not only in the hospital setting, but also in primary care and at the consumer level as the role of preventive medicine expands. There is growing demand for tests used to assess disease risk, both to identify individuals who require frequent monitoring to detect serious diseases at an early stage, as well as to identify individuals who will benefit from preventative therapy.

Assessment of risk for cardiovascular disease is one of the most high-profile applications, but new developments in genomics and proteomics promise to allow improved risk assessment for a variety of other diseases, and to provide more personalized determination of an individual's disease predisposition. In cardiovascular disease, the role of lipoprotein markers is expanding due to new guidelines on acceptable levels of cholesterol, particularly LDL and HDL cholesterol, and a recommendation for use of a complete lipoprotein profile (total cholesterol, LDL cholesterol, HDL cholesterol and triglycerides) as an initial screening test rather than screening for total cholesterol and HDL cholesterol alone. Furthermore, the growing range of lipid-lowering drugs now available provides a new impetus for identification of high-risk individuals who will benefit from therapy.

As described by numerous experts at the late-July meeting of the American Association for Clinical Chemistry (AACC; Washington) in the sprawling McCormick Place convention center along Chicago's Lake Michigan shoreline, cardiovascular risk assessment is expanding to include the use of a variety of additional markers, such as troponin tests, as well as new markers for conditions such as congestive heart failure (CHF) and stroke. By using such markers, clinicians believe that they will be better able to determine the most appropriate course of therapy for patients with acute coronary syndromes and cerebrovascular disorders, and more accurately identify patients with chronic conditions such as CHF. The market opportunity for such tests is substantial and builds on the existing strong and growing demand for cardiac marker tests used in the initial diagnosis of myocardial infarction.

New disease risk assessment tests

The expanded demand for lipoprotein testing has attracted numerous companies, with products now available for a wide range of provider settings. As shown in Table 1 below, there are at least a dozen suppliers of lipoprotein testing kits, not including a large number of suppliers of total cholesterol assays for use on clinical chemistry analyzers. Sales in the lipoprotein testing products market are being driven by rapid growth in the use of lipid-lowering agents such as Lipitor, a drug manufactured by Pfizer/ Warner Lambert (New York) that generated over $5 billion in worldwide sales in 2000, an increase of 33% over 1999. Other lipid lowering agents include Zocor and Mevacor (lovastatin) from Merck & Co. (Whitehouse Station, New Jersey) and Pravachol from Bristol-Myers Squibb (Princeton, New Jersey).

Table 1: Suppliers and Product Offerings for Lipoprotein Testing
Company Products Key Features

Atherotech
(Birmingham, Alabama)
VAP Cholesterol Test Vertical Auto Profile directly measures LDL, HDL and total cholesterol, and also identifies risk factors including Lp(a), VLDL and Intermediate Density Lipoprotein (IDL). Unique ability to determine LDL particle size and density.

Chematics
(North Webster, Indiana)
Chemcard Total cholesterol screening test. List price of $4.25 per test. Classified as Moderately Complex under CLIA. Uses fingerstick whole blood sample and provides results in 3 minutes. Visual readout shows patient risk level.

Cholestech
(Hayward, California)
LDX System Recent addition of ALT assay now allows office-based physicians to perform all required screening tests using a single instrument. Adding non-HDL cholesterol (calculated parameter) based on new guidelines, and hemoglobin A1c. System also provides cardiac risk assessment profile using physician-provided inputs on patient status along with measured lipoprotein values.

Diagnostic Chemicals
(Oxford, Connecticut)
Cholesterol 1,2,3 Three-minute, noninvasive test measures skin cholesterol levels by sampling from patient's palm.

Diasorin
(Stillwater, Minnesota)
Apolipoprotein A1, B, Lp(a) and fibrinogen assays Risk profile developed for use on Dade Behring Dimension XL.

Helena Laboratories
(Beaumont, Texas)
SPIFE Cholesterol Profile Direct measurement of HDL, LDL, VLDL and Lp(a)-C in a single assay at a cost of $30 per sample.

International Reagents
(Kobe, Japan)
Lipoprotein assay reagents for Total, HDL, LDL, RLP cholesterol and triglycerides High-sensitivity enzyme cycling assay for Remnant-Like Particle (RLP) cholesterol developed for use on Toshiba chemistry analyzers. Homogeneous HDL and LDL assays developed for use on Hitachi chemistry analyzers; no interference from high triglyceride levels or bilirubin.

LifeStream Technologies
(Post Falls, Idaho)
LSP 3100 Cholesterol
Monitor
Home-use meter and test strips for patient measurement of total cholesterol from fingerstick samples. Meter and Smart Card for test result storage priced at $129.95. Test strips are priced at $19.95 for package of six (sold through Quick Medical, North Bend, Washington).

Pointe Scientific
(Lincoln Park, Michigan)
Total cholesterol, Direct LDL and HDL, triglycerides, Apolipoprotein A-1 and B Liquid and dry powder reagents for use on automated chemistry analyzers

Polymedco
(Cortlandt Manor,
New York)
Lipi+Plus Direct HDL-C and LDL-C Direct assay designed to run on most automated chemistry analyzers, including Hitachi series, Dade Dimension and Beckman Synchron.

Roche Diagnostics
(Indianapolis, Indiana)
Accu-Chek Instant Plus Hand-held meter providing measurement of glucose and cholesterol at point of care. 12 seconds to result.

Sigma Diagnostics
(St. Louis, Missouri)
EZ LDL, EZ HDL, Infinity total cholesterol and triglycerides, ApoTek Lp(a) No sample pretreatment required. Direct measurement of LDL cholesterol. Kits designed to run on most automated systems.
Source: Cardiovascular Device Update

The tests are not only used to identify patients who are candidates for therapy, but also may have an important role in monitoring of patients who are undergoing treatment. For example, Atherotech (Birmingham, Alabama) is involved in a study with a major pharmaceutical company to determine if the availability of more detailed information on lipoprotein levels will influence physician prescribing behavior. Based on Atherotech's experience, most physicians fail to prescribe the appropriate drug or drug dose using existing tests. The company believes that its comprehensive panel of lipid markers will allow physicians to improve their ability to match drug treatment to disease status. Favorable reimbursement is another factor driving increased use of lipid tests. Cost per test from suppliers ranges from less than 10 cents to about $2.50 per assay, whereas reimbursement from Medicare and private insurers ranges from $13 to $30.

Another rapidly growing segment of the cardiovascular disease testing market is cardiac markers for diagnosis and risk assessment of patients with acute coronary syndromes. The use of troponin markers for initial diagnosis and triage of patients with chest pain is now well-established, and has driven rapid growth in sales in the cardiac marker segment. For example, sales of the Triage Cardiac and BNP product lines for Biosite Diagnostics (San Diego, California), used for point-of-care cardiac and heart failure marker testing, doubled between 1999 and 2000, reaching $13.3 million worldwide. Most of the sales were attributable to the Triage Cardiac line, which consists of a panel of assays for Troponin I, myoglobin, and CK-MB, since marketing of the BNP test did not begin until December 2000.

The troponin assays in particular have, in the space of only a few years, become the primary in vitro diagnostic tool for the diagnosis of myocardial infarction. New studies have demonstrated that troponin markers also are valuable in essentially any situation where it is important to determine the extent of damage to cardiac tissue. As discussed by E. Magnus Ohman, MD, of Duke Clinical Research Institute (Durham, North Carolina), at the AACC conference, troponin I is proving useful in assessing cardiac damage in patients undergoing chemotherapy, and in differentiation of cardiac tissue necrosis vs. ischemic injury. Troponin markers also are proving very powerful in assessing prognosis after myocardial infarction, with the trend in elevation of the marker following an initial MI showing a strong correlation with risk of subsequent adverse events.

The emerging trend for use of troponin markers is to test chest pain patients once upon initial presentation, a second time three to six hours later, again at six to nine hours, and a fourth time at 12 hours. According to Dr. Fred Apple of Hennepin County Medical Center (Minneapolis, Minnesota), the risk prognosis is related to the level of troponin elevation observed during that period. The troponin level can be used to distinguish between unstable angina and MI for non-ST segment elevation patients, and can also help to determine which MI patients will benefit from the use of high-priced platelet function inhibitors, such as ReoPro, a GPIIb/IIIa inhibitor from the Centocor (Malvern, Pennsylvania) unit of Johnson & Johnson (New Brunswick, New Jersey).

The use of point-of-care testing for cardiac markers is now recommended if the turnaround time for testing performed in the central lab cannot be reduced to under one hour. However, according to data presented by Apple, existing POC cardiac marker systems do not have adequate sensitivity and precision for detailed risk assessment. As shown in Table 2 below, Apple presented data on sensitivity of various troponin assays demonstrating that only a few assays designed for use in the central lab, including the Access troponin I assay from Beckman Coulter (Brea, California); the Centaur troponin I from Bayer Diagnostics (Tarrytown, New York); the Elecsys troponin T assay from Roche Diagnostics (Indianapolis, Indiana); and the ECi troponin I assay from Ortho Clinical Diagnostics (Raritan, New Jersey) have the sensitivity needed to discriminate certain high-risk groups of patients.

Table 2: Detection Sensitivity Comparison for Various Troponin Assays
Supplier Assay System Lower Limit
of Detection (ug/L)

Abbott Diagnostics
(Abbott Park, Illinois)
AxSYM 0.14

Bayer Diagnostics
(Tarrytown, New York)
Immuno
ACS-180
Centaur
1 0.10
0.03
0.02

Beckman Coulter
(Brea, California)
Access* 0.01

Biosite
(San Diego, California)
Triage Cardiac 0.19

Dade Behring
(Deerfield, Illinois)
Stratus II
Dimension RxL*
Stratus CS*
OPUS*
0.35
0.04
0.03
0.10

Diagnostic Products
(Los Angeles, California)
Immulite
Turbo
0.20
0.50

Roche Diagnostics
(Indianapolis, Indiana)
Elecsys* 0.01

Ortho-Clinical Diagnostics
(Raritan, New Jersey)
ECi 0.02

Sigma Diagnostics/ First Medical
(St. Louis, Missouri)
Alpha Dx 0.09

Tosoh Medics
(South San Francisco, California)
AIA 0.45

* Second- or third-generation assay; number supplied by manufacturer
Source: Dr. Fred Apple, Hennepin County Medical Center, presentation at Bayer Diagnostics workshop in association with American Association for Clinical Chemistry annual meeting, July 31, 2001

Among the systems designed for point-of-care use, the Stratus CS from Dade Behring (Deerfield, Illinois) is the most sensitive, based on Apple's data, followed by the Alpha Dx system from First Medical/Sigma Diagnostics (St. Louis, Missouri), and the Biosite Cardiac Triage. The sensitivity data indicate that POC systems are best employed for initial diagnosis, to obtain a rapid indication of the presence of MI, while systems in the central lab must be used if trends in troponin level are to be used to decide upon subsequent treatment plans based on the severity of disease.

BNP joins diagnostic lineup

The newest marker in the cardiovascular disease segment is brain natriuretic peptide (BNP), a test introduced by Biosite Diagnostics that is proving valuable in the management of congestive heart failure (CHF). Improved management of CHF is a high priority for disease management providers, as the costs for hospitalization are higher for that disease than for cancer or heart attack. BNP is a neurohormone that, unlike other neurohormones such as norepinephrine, is highly specific for cardiac vessels and exhibits a predictable level between patients. Initial evaluations of the marker indicated a poor correlation with New York Heart Association (NYHA) class ratings, the traditional classification scheme for CHF disease, but, as discussed at AACC by Alan Maisel, MD, of San Diego Veterans Affairs Healthcare (San Diego, California), the problem was due to the subjective nature of the NYHA scheme, rather than lack of correlation of BNP levels with disease. In fact, Maisel no longer uses the NYHA score to classify patients in his CHF clinic, preferring to use BNP levels instead.

CHF historically has been a difficult disease to diagnose accurately. Conventional techniques, including the physical exam and chest X-ray, have been shown to have a poor correlation with cardiac wedge pressures in congestive heart failure. BNP, however, appears to provide a new and highly objective tool for diagnosis and disease stratification, with a high correlation with severity and a very high negative predictive value. According to Maisel, the test can prove life-saving in patients with shortness of breath who do not have CHF, such as patients with chronic obstructive pulmonary disease, since their condition is likely to be an acute one requiring immediate treatment, and knowing that CHF is not causing the symptoms can help the clinician to arrive at the correct diagnosis and initiate appropriate therapy.

In addition to its role in initial diagnosis, BNP appears to have a role in monitoring of treatment for CHF. Until recently, there was no good method to monitor the status of CHF patients, according to Maisel, and also no good therapies. The situation is now changing dramatically, with the introduction of BNP as well as of a number of new, effective drugs. One just-approved drug, Natrecor, from Scios (Sunnyvale, California), is based on BNP itself, and at least 10 other drugs are in various stages of development. As a result, there is now heightened interest in objective methods for monitoring of CHF patients, both in the hospital as well as in outpatient settings such as primary care and heart failure clinics. Maisel said he believes there may be a role for the BNP test as a screening tool in primary care, since the Biosite test is configured for point-of-care use and accurately reflects disease status, being largely unaffected by extraneous factors such as exertion and posture. According to Maisel, some physicians are already beginning to use the BNP test to screen patients for CHF. At present, Biosite is the only supplier of a BNP test. The assay runs on the Cardiac Triage POC analyzer platform, the same one that has already been deployed for cardiac marker testing. As of July, Biosite had a total of 470 Triage Cardiac customers in the U.S. with a goal of reaching 500 to 550 by year-end.

Stroke markers a new market

An emerging area in cardiovascular disease testing is markers for stroke. Biosite also is pursuing new stroke markers, and announced the discovery of a panel of six markers at the AACC gathering that show promise for stroke diagnosis. About 750,000 people suffer a new or recurrent stroke annually, and the disease is the leading cause of adult disability. Although at least one treatment, namely thrombolysis with tissue plasminogen activator (tPA), has been approved by the FDA, less than 4% of all stroke patients receive the drug in time because they usually do not present within the three-hour window needed for effectiveness. In addition, most hospitals are unable to perform adequate stroke treatment, in part because of a lack of adequate diagnostic modalities. Computed tomography scans have a low sensitivity for diagnosis of stroke, while MR imaging, although more effective, is not widely available. Because of the relatively high rate of adverse events (i.e., cerebral hemorrhage) with tPA therapy, physicians are reluctant to give the drug unless they have definitive evidence of occlusive stroke.

Biosite has studied samples from 173 patients, 151 of whom were diagnosed with stroke, plus another 157 normal patient samples. The company investigated a total of 31 potential markers and identified a panel of six proteins that gave a sensitivity of 79% and a specificity of 93% for diagnosis of ischemic stroke. That compares with a sensitivity of only 33% for a CT scan. The same panel is also able to differentiate patients suffering transient ischemic attacks (TIAs) from normals and detects strokes of all types including ischemic strokes and strokes arising from subarachnoid and intracerebral hemorrhage. The company plans to continue its studies to develop additional markers that can distinguish between hemorrhagic and ischemic stroke. The stroke panel will be adapted to the Triage instrument platform, and the company anticipates filing of a premarket approval application by the end of 2002 or early 2003. In the company's most recent filing, for the BNP marker, approval for marketing was obtained about 10 months after filing.

New developments in POC testing

The new Biosite tests not only represent an advance in diagnostic markers for major cardiovascular diseases, but also are configured to take advantage of the increasing migration of testing outside of the central laboratory to a variety of alternate sites both within and outside of the hospital or reference laboratory. In particular, the new BNP test could find widespread application in the primary care or specialist office setting for screening of patients for heart failure, to help determine if more comprehensive and expensive imaging and physiological tests are indicated.

A new cardiac marker system was unveiled at the AACC exhibition by Innotrac Diagnostics Oy (Turku, Finland). Available at present for research use only, the system menu includes troponin I (both complexed and free), CK-MB, and myoglobin, with high-sensitivity CRP, PSA and TSH assays under development. The system initially will be targeted at applications in the emergency department and cardiac care, similar to the Stratus CS system from Dade Behring. All reagents are contained in individual test cups, and 12 cups are supplied in a pen cartridge that is loaded onto the Innotrac analyzer. Sample types include whole blood, plasma, and serum, and sampling can be performed directly from a primary specimen tube, with a volume of from five to 20 microliters depending on the test. Up to 88 samples can be loaded on the system, along with reagents to perform up to 360 tests. Turnaround time is 18 minutes, and the company plans to sell to distributors at a price of under $2 per test, perhaps setting the stage for price erosion in the cardiac marker segment.

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