Download NMP22® BladderChek® Test: point-of-care technology with life- and money-saving potential Diagnostic Profile

Diagnostic Profile
NMP22® BladderChek® Test:
point-of-care technology with
life- and money-saving potential
Kevin M Tomera
CONTENTS
Urinary bladder cancer
Advent of biomarkers
NMP22® BladderChek® Test
Expert opinion
Conclusions
Five-year view
References
A new, relatively obscure tumor marker assay, the NMP22® BladderChek® Test (Matritech,
Inc.), represents a paradigm shift in the diagnosis and management of urinary bladder
cancer (transitional cell carcinoma). Specifically, BladderChek should be employed
everytime a cystoscopy is performed, with corresponding changes in the diagnostic
protocol and the guidelines of the American Urological Association for the diagnosis and
management of bladder cancer. Currently, cystoscopy is the reference standard and
NMP22 BladderChek Test in combination with cystoscopy improves the performance of
cystoscopy. At every stage of disease, BladderChek provides a higher sensitivity for the
detection of bladder cancer than cytology, which now represents the adjunctive standard
of care. Moreover, BladderChek is four-times more sensitive than cytology and is available
at half the cost. Early detection of bladder cancer improves prognosis, quality of life and
survival. The BladderChek may be analogous to the prostate-specific antigen test and
eventually expand beyond the urologic setting into the primary care setting for the testing
of high-risk patients characterized by smoking history, occupational exposures or age.
Expert Rev. Mol. Diagn. 4(6), (2004)
Affiliation
Alaska Regional Medical Plaza,
1200 Airport Heights Drive,
Suite 101, Anchorage,
Alaska 99508, USA
Tel.: +1 907 276 1455
Fax: +1 907 276 1477
[email protected]
[email protected]
KEYWORDS:
assay, cost of healthcare,
diagnosis, diagnostics,
NMP22® Test Kit,
NMP22®BladderChek®Test,
transitional cell carcinoma, TCC,
urinary bladder cancer,
recurrent bladder cancer,
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The NMP22® BladderChek® Test (Matritech,
Inc.) is a new, simple, point-of-care diagnostic
test for urinary bladder cancer. It is as easy to
use as an at-home pregnancy test and has the
potential to immediately save lives, reduce
morbidity and the cost of urologic healthcare.
The NMP22 BladderChek Test is a rather
phenomenal dipstick technology that can
detect 6 × 10-17 moles of nuclear matrix protein (NMP)-22 within urine. NMP22 is a protein that is essential for cell division. It is normally present at low concentrations, however,
it is elevated up to 80-fold in tumor cells;
tumor cells within the urinary tract shed relatively high concentrations of NMP22 into the
urine. The assay is inexpensive and provides
results within the hour.
The NMP22 BladderChek Test serves as an
adjunctive test to cystoscopy, the visual examination of the lower urinary tract and the
bladder with a cystoscope. In a community
clinical setting, the NMP22 BladderChek
Test changes the clinical and diagnostic
behavior of the attending physician for the
10.1586/14737159.4.6.1
better, providing additional information for
refinement of the diagnostic algorithm and/or
treatment of recurrent bladder cancer.
Given its current lack of prominence, the situation with the BladderChek Test may be
analogous to that of the prostate-specific antigen
(PSA) assay, one whose clinical utility at first was
uncertain, but which eventually became a clinical mainstay. The PSA assay has certainly
reduced both morbidity and mortality from
prostate cancer. The NMP22 BladderChek Test
is the only tumor marker approved by the US
Food and Drug Administration (FDA) as an aid
for the diagnosis of de novo bladder cancer as
well as monitoring for cancer recurrence following treatment. This test is available, easy to use,
inexpensive and can save lives on a daily basis.
Urinary bladder cancer
Pathophysiology
Urinary bladder cancer is a common urologic
malignancy whose diagnosis and management
represents a significant challenge to the practicing urologist. Transitional cell carcinoma
© 2004 Future Drugs Ltd
ISSN 1473-7159
1
Tomera
(TCC), also known as urothelial carcinoma, accounts for
approximately 90% of urothelial carcinomas [1]. TCC is a
malignant neoplasm of the transitional epithelium, the internal
lining of the bladder (i.e., urothelium).
Bladder tumors are graded according to their histologic
appearance relative to normal cell tissue (i.e., their level of differentiation). In essence, staging represents the aggressive propensity of the tumor, its likelihood to invade surrounding tissues. In conjunction, bladder tumors are staged according to
how deeply a tumor has penetrated the bladder wall, as
depicted in FIGURE 1. The common stages of progression are
listed in TABLE 1.
Ta tumors are usually low-grade that have remained within
the urothelium and have not progressed into the lamina propria
or deeper into the underlying muscle tissue. Ta tumors account
for approximately 79–80% of initial urinary bladder cancer
diagnoses [4]. T1 tumors have invaded the lamina propria but
not the underlying muscle. Collectively, Ta and T1 tumors are
referred to as superficial tumors. Tumors staged T2 or above
have invaded the underlying muscle tissue and are collectively
referred to as invasive tumors.
Peritoneum
Perivesical fat
Muscle
Lamia propria
The Tis stage of classification is specific for a certain type of
high-stage tumor known as carcinoma in situ (CIS). Most
tumors form bulges on the bladder wall. However, Tis tumors
exhibit a tendency to remain planar (flat), although they are
likely to be aggressive tumors.
Once diagnosed, superficial tumors are usually removed via
transurethral resection (cystoscopic surgery via the urethra).
Once resected, the tumor tissue undergoes histologic analysis
for determination of grade and stage. Alternatively, laser surgery or radiation can treat the tumor, although a biopsy for histologic analysis must be obtained prior to this. Following
tumor removal, chemotherapeutic or immunotherapeutic
agents may be injected directly into the bladder (intravesical
therapy) in an attempt to prevent tumor recurrence.
Prognosis
Urothelial tumors represent a range of diseases with a range of
prognoses. TCC is known for having high progression rates.
Progression, defined as the development of muscle-invasive disease or metastasis, ranges from approximately 5–30% for T1
tumors up to approximately 60% for CIS [1]. Once tumors have
become muscle invasive (T1 stage or greater), cystectomy (surgical removal of the bladder) with a urinary diversion may be the
only treatment option. This requires major surgery with lifelong follow-up for urinary diversion maintenance. Intravesical
chemo- or immunotherapy may also be attempted.
Treatment and surveillance of bladder cancer requires extensive and prolonged follow-up. Once urinary bladder cancer
develops, tumor recurrence may occur at the same or at any
location, at a similar or more advanced stage. Tumor recurrence
rates reach up to 70% at 5 years [1].
Epidemiology
Ta
Urothelium
T1
T2b
T3b
Figure 1. Layers of the bladder wall and selected stages of cancer
progression.
Adapted with permission from the American Urological Association
Doctor’s Guide for Patients on The Management of Bladder Cancer (1999).
2
According to 2003 estimates, bladder cancer will account for
57,400 of new cases and 12,500 associated deaths annually in
the USA. The incidence of bladder cancer in men is approximately threefold greater than in women and approximately twice
as common in whites as blacks [101]. The overall 5-year relative
survival rate is 82.6% [102]. Survival can be as high as 94% for
superficial cancer but as low as 6% for metastatic disease [103].
Significant risk factors for urinary bladder cancer include a
history of smoking or occupational exposure to chemicals and
dyes. Cigarette smoking accounts for an estimated two-thirds
of bladder cancers in males and one-third in females [5]. A correlation reportedly exists between the number of pack-years and
the risk of developing bladder cancer. Smoking cessation
decreases this risk but not to that of a nonsmoker.
A number of industrial organic chemicals are carcinogens
known to induce bladder cancer, particularly aromatic (aryl)amines such as naphthalenes, benzidine, aniline dyes, and 4-aminobiphenyl. High-risk occupations include petroleum chemical
and rubber workers, hair dressers, painters, textile workers, truck
drivers and aluminum electroplaters. Bladder cancer may also
result from pelvic radiotherapy, phenacetin (analgesic) abuse and
exposure to the anticancer therapeutic agent cyclophosphamide [6].
Expert Rev. Mol. Diagn. 4(6), (2004)
NMP22® BladderChek® Test
Table 1. Staging of primary bladder cancer tumors.
Tumor stage
Substage
Ta: Noninvasive papillary carcinoma
Tis: Carcinoma in situ (anaplastic ‘flat tumor’ confined to urothelium)
T1: Tumor invades lamina propria
T2: Tumor invades muscularis propria
T2a: Invades superficial muscularis propria
T2b: Invades deep muscularis propria
T3: Tumor invades perivesical fat
T3a: Invades microscopic perivesical fat
T3b: Invades macroscopic perivesical fat (extravesical mass)
T4: Tumor invades prostate, uterus, vagina, pelvic wall or abdominal wall
T4a: Invades adjacent organs (uterus, ovaries, prostate stoma)
T4b: Invades pelvic wall and/or abdominal wall
T: Tumor.
In some geologic areas, exposure to arsenic in groundwater is
also a recognized risk for bladder cancer, the contamination of
which is presumably due to pesticide use or industrial contamination [7]. Firefighters and arson investigators are also at high
risk of bladder cancer due to exposure to carcinogenic materials
in smoke, building debris and combustible materials [104].
Current diagnostic algorithm
Historically, initial diagnosis of bladder cancer has been problematic. Hematuria is the most common presenting symptom
of bladder cancer. Most cases are discovered as microscopic
hematuria, which cannot be observed visually, but rather
requires a chemical test for detection. However, hematuria is a
nonspecific finding and requires extensive differential diagnosis
as it may arise from a number of other nonmalignant, urologic
conditions, including urinary tract infection, recent strenuous
exercise, benign enlargmenent of the prostate or simply nonspecific presence of blood. The proportion of hematuria attributable to urinary tract malignancy reportedly ranges from
0.5–13% [8,9], subject to population characteristics. Even in the
event of bladder cancer, the hematuria may be intermittent,
and its degree does not correlate with the severity of underlying
disease [10]. However, unless readily explained by the aforementioned benign conditions, the presence of hematuria mandates
an extensive urologic evaluation, even after a single episode [11].
The current diagnostic algorithm for bladder cancer, whether
de novo or recurrent (postsurgical), is accomplished with the
same diagnostic tools: cytology and cystoscopy. Cytology, also
refereed to as voided urine cytology, involves the microscopic
examination of cells obtained from excreted (voided) urine.
The cells are examined for morphologic changes indicative of
cancerous conditions. Cystoscopy is the visual examination of
the bladder mucosa with a cystoscope.
Each diagnostic procedure has its advantages and shortcomings. For example, cytology is highly specific and morphologic
conditions that indicate a cancerous condition often result in a
correct diagnosis. However, cytology is subjective as it relies on
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the observation, interpretation and skill of the cytopathologist,
thereby rendering it subject to human error and interoperator
bias. Intact cells are required for morphologic examination. As
such, cytology has a low sensitivity for low-grade neoplasms,
which are less likely to spontaneously exfoliate intact cancerous
cells into the urinary tract. The intracellular attachments in lowgrade tumors are preserved in a better state, and the degree of
cellular morphologic change is less than that of high-grade
tumors [12–14]. Diagnostic error may arise from some inflammatory conditions that induce cellular changes resembling those
seen in neoplastic processes, thereby generating vague, atypical
findings. Finally, urine samples must be sent to the laboratory for
evaluation, consequently introducing a time delay of weekdays.
When cytology renders cellular morphology suggestive of
cancerous conditions, direct visualization of the bladder via cystoscopy is indicated. Cystoscopy is usually performed in the
office of the attending urologist. The fiberoptic element of the
cystoscope is inserted into the bladder via the patient’s urethra,
enabling the urologist to visually detect and identify the tumor.
Once the tumor is located, a biopsy is usually obtained during
the cystoscopic procedure with the histologic examination of
the biopsy tissue providing confirmed diagnosis.
Although representing the diagnostic gold standard, the
procedure is expensive, and cystoscopy is subject to the fallibility of human observation and interpretation, as well as to
procedural limitations. Poor visualization may result due to
inflammatory conditions, enlarged prostates, bleeding, and
folds or trabeculations in the bladder tissue. Flat urothelial
lesions such as severe dysplasias and CIS are frequently difficult to distinguish from normal bladder tissue [15,16], and can
even be confused with erythema. Finally, the tumor may
reside in the upper urinary tract (i.e., upstream of the bladder
in ureters or kidneys), which the cystoscope can not traverse.
The actual frequency of false-negative cystoscopy results is
difficult to determine, but estimates range from 10–40%
[17–19]. For these reasons, cytology is frequently used as a
adjunctive or back-up test.
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When cytology indicates the presence of cancer and cystoscopy fails to detect any tumors, an intravenous pyelogram
(IVP) – a radiographic investigation of the upper urinary tract
– may be indicated. During an IVP, a radiopaque contrast dye
is injected into the patient’s vein and radiographic visualization
of the kidneys and ureters is accomplished as the contrast agent
is excreted. Though an effective radiologic evaluation, the procedure is expensive, inconvenient and exposes patients to the
risk of adverse reactions to the contrast medium, including an
infrequent fatal adverse reaction.
Advent of biomarkers
Biotechnology has now provided new biomarkers for the diagnosis and monitoring of bladder cancer. Of the current FDAapproved assays, the NMP22 BladderChek Test provides the
best and broadest clinical utility.
The NMP22 BladderChek Test detects 4.5-times as many
superficial cancers and 2.5-times as many invasive cancers as
cytology [105]. Although cytology detects cancer on a cellular level
and cystoscopy detects tumors on a macroscopic level, BladderChek provides a molecular view of cancerous conditions,
detecting the presence of specific molecules within urine that indicate the presence of cancer. Furthermore, BladderChek provides a
molecular view of the upper as well as the lower urinary tract.
Measures of accuracy
Four terms are often used in reference to the accuracy of diagnostic tests: sensitivity, specificity, positive predictive value
(PPV), and negative predictive value (NPV). In the determination of accuracy, the test in question is usually compared with
an established reference method. For the diagnosis of de novo
tumors and monitoring of recurrent bladder cancer, cystoscopy
stands as the best reference method.
In regards to bladder cancer, sensitivity represents the probability of correctly identifying all the patients with bladder cancer among a given population. The higher the sensitivity, the
fewer the number of undetected cancer patients within this
population. If a diagnostic test were to have 100% sensitivity,
all the patients with bladder cancer within a given population
would be correctly identified.
Specificity represents the probability that a patient who tests
negative does not have cancer (i.e., a test with a high specificity
generates very few false positives). Conversely, when a highly
specific test provides a positive test result, the person most
likely has bladder cancer.
Predictive value expresses the likelihood that a given test result
correlates with the presence or absence of disease. PPV represents the probability that a person with a positive test is a true
positive (i.e., has the disease). The PPV value addresses the question of how likely it is that one really has the disease, given that
the test result was positive. The NPV represents the probability
that the person with a negative test does not have the disease.
Clinically, there is a problem with the use of these terms. These
measures of accuracy are actually academic research issues that
often mislead a practicing physician about the clinical value of a
4
test. For example, PSA has a poor sensitivity, specificity, PPV and
NPV, yet it is universally accepted as a valuable tool that has
detected tens of thousands of prostate cancers that otherwise
would have caused significant morbidity and/or mortality.
NMP22 BladderChek Test
Nuclear matrix proteins
The NMPs are a class of nuclear proteins that provide structural
support for a cell’s nucleus [22]. This matrix serves as an anchor
for the enzymatic apparatus responsible for DNA replication,
transcription, RNA processing and gene expression [20,21].
Various NMPs are organ specific. NMP22, or nuclear
mitotic apparatus (NuMA) protein is found in human epithelial cells. The critical function of NuMAs is to ensure the correct separation of genetic material during mitosis into the
respective daughter cells [22]. NuMA concentrations in urothelial tumor cells are up to 80-fold greater than that of normal tissue [23] (cancer-specific NMPs have also been identified in
colon, breast and bone tissues [23–25]).
Groundwork quantitative studies
The NMP22 BladderChek Test is an in vitro immunoassay
intended for the qualitative detection of NMP22 in urine. It
performs a qualitative analysis (i.e., it determines the presence
or absence of a substance based on a predetermined threshold
concentration). In essence, BladderChek determines whether
NMP22 is present in the urine or not and provides an absolute positive or negative test result in the same manner as a
pregnancy test. The NMP22 BladderChek Test is the only
tumor marker approved by the FDA for both the diagnosis of
de novo bladder cancer and monitoring for recurrent cancer
following surgery.
Prior to developing the NMP22 BladderChek Test,
Matritech, Inc. created a quantitative test, the NMP22® Test
Kit, which is a microplate format immunoassay intended for
the quantitative detection of NMP22 in urine. The NMP22
Test Kit measures the actual quantity of NMP22 in the urine,
providing results in units/ml. One unit represents 110 pg of
protein. The NMP22 Test Kit was initially FDA approved in
1996 for the prognosis and monitoring of recurrent bladder
cancer (following surgery). In 2000, the test was the first tumor
marker to receive FDA approval for the diagnosis of bladder
cancer. This is the quantitative test that provided the groundwork studies which validated the use of NMP22 for detecting
the presence of urinary bladder cancer.
At the outset, it was necessary to determine whether the quantification of NMP22 levels in urine could yield important clinical information. The initial investigative focus was a comparison
of NMP22 to traditional cytology. Investigative goals were:
• Differentiation of NMP22 levels between healthy volunteers
and disease groups
• Evaluation of NMP22 efficacy for monitoring recurrent TCC
• Evaluation of NMP22 clinical utility in the differential
diagnosis of hematuria
Expert Rev. Mol. Diagn. 4(6), (2004)
NMP22® BladderChek® Test
The ability of NMP22 to differentiate between bladder cancer and nonmalignant events also had to be evaluated (in all the
following studies, cystoscopy and the histologic analysis of
biopsy tissue were used as the standard reference method).
In an initial large investigation with over 600 patients,
Carpinito and coworkers demonstrated that patients with
active TCC had significantly higher NMP22 concentrations in
their urine compared with patients who had benign urologic
conditions and healthy volunteers. The difference between
healthy volunteers and patients with visible TCC was highly
significant (p = 0.0001) [26].
Subsequent investigations sought to determine the sensitivity
and specificity of the quantitative NMP22 assay relative to
cytology. Although cytology is highly specific (positive results
are usually rarely false positives), its major drawback resides in
its low sensitivity in identifying low-grade tumors.
In three separate studies collectively involving approximately
400 patients, Soloway and coworkers, Miyanaga and coworkers, and Landman and coworkers demonstrated that the quantitative NMP22 test had an overall sensitivity of 70–81% for
detecting recurrent TCC [27–29]. Comparatively, cytology has a
reported sensitivity of approximately 10–40% [17,18,28,30,105].
A notable finding was that NMP22 had a high sensitivity for
low-grade tumors. Miyanaga and coworkers found that
NMP22 sensitivity increased from 70.0% for superficial
tumors (Tis, Ta, and T1) to 100% for invasive tumors (T2, T3,
and T4). Landman and coworkers reported a sensitivity of 81%
for Ta tumors. The three investigators used NMP22 reference
cut-off values of 6.0–20.0 units/ml. Results for only
6.0–10.0 units/ml are reported in this article.
In comparison with cytology, NMP22 was lacking specificity.
Carpinito and coworkers, Soloway and coworkers, and Landman
and coworkers reported a NMP22 specificity of 70–79%, whereas
cytology in the Carpinito and Landman studies reported a specificity of 90–94%. In actual practice, these findings suggest that
NMP22 generated more false positives than cytology. In clinical
terms, the consequences of these false positives (low specificity) are
that a few patients are inconvenienced by unnecessary cystoscopic
evaluations. Conversely, NMP22 detected tumors that cytology
missed (NMP22 has high sensitivity). Obviously, the consequences
of failing to detect the presence of tumors are detrimental.
Regarding the issue of false positives, two authors, Ponsky
and coworkers and Sharma and coworkers, found that the
reduction to six clinical categories, described below, increased
quantitative specificity to over 95% (i.e., potentially reducing
the number of unnecessary cystoscopies). Also, since cystoscopy, the reference standard for these studies, has a generally
accepted sensitivity in the region of 80–85%, it is likely that a
proportion of positive NMP22 test results classified as false
positive may indeed be true positives overlooked by cystoscopy.
Soloway and coworkers and Miyanaga and coworkers also determined that NMP22 had a high NPV, ranging from 86–94.9%.
The importance of a high NPV is that a negative test result provides a high degree of confidence that no tumor is present, and
that further evaluation (cystoscopy) may not be required.
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In the differential diagnosis of microscopic hematuria, three
studies involving over 700 patients demonstrated the efficacy of
the quantitative NMP22 assay for this clinical application. In
patients presenting with hematuria, Akaza and coworkers, Miyanaga and coworkers, and Carpinito and coworkers found that
NMP22 had a sensitivity ranging from 85.7–90.9% for the
detection of urothelial cancer. In comparison, the sensitivity of
cytology ranged from 50–54.5%.
Cytology again proved to have a greater specificity. Cytologic
specificity ranged from 90–99.7%, while that of the NMP22
assay ranged from 70–78.7%. Again, the NMP22 assay proved
better at detecting the presence of TCC at the cost of generating a higher number of false positives [31–33]. The investigators
in these differential diagnosis studies used a NMP22 reference
cut-off value of 6.0 or 12.0 units/ml.
The indicated and intended use of the NMP22 quantitative
test is the management of bladder cancer (i.e., monitoring the
condition of patients who had been diagnosed and treated for
bladder cancer to determine whether TCC had reccurred).
Evaluating the assay in this regard, Stampfer and coworkers
examined the clinical efficacy of the NMP22 assay for the
detection of recurrent TCC in a population of patients with a
history of TCC [34].
Using a 6.4. units/ml cut-off, Stampfer and coworkers found
that NMP22 levels were significantly higher in patients with
recurrent TCC compared with patients exhibiting no recurrence. In comparison with cytology, NMP22 sensitivity was
67% versus 31% and 40% in two separate cytology groups.
The former included malignant cells only, while the latter
included suspicious and dysplastic cytology. Notably, NMP22
sensitivity was greater with low-grade, superficial tumors (Ta):
NMP22 claimed a sensitivity of 54% versus 14% or 29% for
cytology, respectively. In addition, Stampfer and coworkers
determined that NMP22 sensitivity was stratified by tumor
stage: NMP22 sensitivity increased from 59% for low-grade
tumors (Ta, grade 1 and 2) to 90.0% for high-risk tumors (Ta,
grade 3 and T1).
The shedding of NMP22 into the urine by tumors is the
result of cellular apoptosis and other mechanisms of cell death.
Therefore, in theory, nonmalignant, inflammatory urothelial
conditions could elevate urinary NMP22 values and give rise to
false positives. In clinical evaluation, several studies differed in
their findings. Carpinito and coworkers reported no difference
between patients with benign urologic conditions and healthy
volunteers [26]. Akaza and coworkers reported no overlap of confidence intervals for the malignant and benign condition groups
[31]. Miyanaga and coworkers reported high false-positive rates
for urolithiasis (50%), benign prostatic hypertrophy (15.6%),
benign urologic diseases (25.6%) and renal diseases (25.6%) [28].
Later studies categorized specific nonmalignant conditions
that give rise to false positives. These studies will be addressed
later in this article when the issue of false positives is discussed,
as some findings thought to be NMP22 false positives were in
fact true positives (i.e., the NMP22 Test Kit detected tumors
that cystoscopy overlooked on initial evaluation).
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Exclusion criteria
Several investigations have showed that various clinical exclusion criteria can greatly improve the specificity of NMP22 (and
remove the incidence of true false positives). In separate studies
collectively involving almost 900 patients, Ponsky and
coworkers and Sharma and coworkers discovered that the
reduction to six clinical categories increased quantitative
NMP22 specificity to 99.2% and 95.6%, respectively. These
categories include the presence of [35,36]:
• Benign inflammatory/infectious conditions
• Renal or bladder calculi
• Foreign body (or a history thereof )
• Bowel interposition segment
• Other genitourinary cancer
• A urine sample obtained through instrumentation
NMP22 BladderChek Test qualitative studies
Although the quantitative NMP22 Test Kit proved its clinical
utility in detecting recurrent TCC, it was still a laboratory test:
the quantitative test requires the expertise of a laboratory technician. Clinicians are required to obtain urine samples, refrigerate them, send them to an analytical laboratory and wait for the
test results. Naturally, the NMP22 Test Kit was designed for
this purpose, comprised of a 96-microwell plate to enable
simultaneous testing of samples. The quantitative test is still
used in analytical settings with high throughput (e.g., in analytical laboratories and hospitals), but the point-of-care qualitative
NMP22 BladderChek Test provides the best utility for the
practicing clinical urologist.
With the NMP22 BladderChek Test, Matritech, Inc. made
the same NMP22 technology available to the clinician. The
NMP22 BladderChek Test technology uses a lateral-flow,
immunochromatographic strip encased in a plastic cartridge to
detect NMP22 qualitatively in the patient’s urine. The assay
incorporates two different monoclonal antibodies: one capture
antibody and one reporter antibody. The BladderChek test
device requires four drops of urine at room temperature, and
delivers results in 30 min. The results are as easy to read as an athome pregnancy test: a color band in the test position indicates
a positive finding.
The pivotal trial for the FDA approval of the NMP22 BladderChek Test was a prospective, blinded clinical trial involving
23 sites in the USA [GROSSMAN ET AL, SUBMITTED]. Over 1300
patients with either risk factors or cancer symptoms of urothelial cancer (hematuria, dysuria, urinary frequency or urgency,
back pain, bladder pain, history of smoking and/or exposure to
carcinogens) were evaluated prospectively for bladder cancer.
No patients had a history of bladder malignancy, although 26
patients with active cancers other than bladder were recruited.
A voided urine specimen was obtained from each patient
prior to cystoscopy. Four drops of urine were applied to the
BladderChek test device and the result was recorded after 30
min. A portion of the urine specimen was also sent to a laboratory for cytologic analysis for each respective site’s protocol.
6
Biopsies of suspected tumors were obtained during cystoscopy
and the tissues were sent for histologic analysis. The results of
both BladderChek and cytology were analyzed for agreement
with the final pathologic diagnosis.
The results of these studies demonstrated that the NMP22
BladderChek Test detected a higher percentage of bladder
tumors than cytology for all stages of tumors. Of the 69 biopsyconfirmed tumors detected in the study, BladderChek had a
higher sensitivity than cytology for superficial (50% vs. 17%),
CIS (80% vs. 60%) and invasive tumors (89% vs. 22%). Notably, the sensitivity of the NMP22 BladderChek Test increased
relative to the severity of tumor grade.
Another notable finding was that the combined use of BladderChek and cystoscopy detected more tumors than cystoscopy
alone. With superficial tumors, the combination of BladderChek and cystoscopy detected 93% versus 92% for cystoscopy alone. With CIS tumors, BladderChek and cystoscopy
combined detected 80% versus 60% with cystoscopy alone;
and the combination detected 89% of invasive tumors versus
56% with cystoscopy alone. Subsequent follow-up, radiologic
studies or cytology detected tumors missed on initial evaluation. This is an important finding as the addition of the
NMP22 BladderChek Test to cystoscopy improved the efficacy
of the standard’s best urologic reference method.
Overall, BladderChek identified 32 malignancies that cytology missed while cytology was positive only in two patients
for whom NMP22 was negative. BladderChek also detected
four cancers that were not visualized during initial cystoscopy:
three invasive tumors and one CIS tumor – the CIS disease
was initially reported as benign during initial cystoscopy.
The pivotal study also demonstrated an excellent specificity
for the NMP22 BladderChek Test versus benign urologic condition and/or the absence of urologic disease. The specificity of
BladderChek versus benign urologic conditions was 85%, and
90% specific for findings of no evident urologic disease. Cytology was 99% specific in both these circumstances but, again,
cytology missed many tumors that BladderChek detected.
As a participant in this pivotal trial, the authors’ urologic
practice (Alaska Urologic Associates, AK, USA) detected 18
cases of TCC (17 bladder and one ureteral) in 248 evaluated
patients. Initial cystoscopy identified 15 (83%) cases of TCC.
BladderChek confirmed 11 (63%) of these cases, while cytology confirmed only three (17%) (TABLE 2). The three cases
detected by cytology were also detected by BladderChek. The
combination of initial cystoscopy with BladderChek identified
17 (94%) TCC cases.
Of these 18 detected TCC cases, seven cases were high-grade
TCC. Initial cystoscopy identified five (71%) high-grade
tumors; BladderChek confirmed six (86%) of these, while
cytology confirmed only two (29%). Five cases of invasive TCC
were identified with initial cystoscopy and BladderChek successfully identified all five cases (100%), while cytology only
confirmed one (20%). For specificity, BladderChek correctly
confirmed that 204 of 230 (89%) patients had no cancer, while
cytology confirmed 218 (95%) [37].
Expert Rev. Mol. Diagn. 4(6), (2004)
NMP22® BladderChek® Test
Table 2. Summary of results from Alaska Urologic Associates [41].
Type of cancer
Bladder
Ureteral
Number of patients
17
1
Positive by cystoscopy
15
-
83
Positive by BladderChek
10
1
61
26
2
1
18
0
Positive by cytology
In conclusion, the authors found the NMP22 BladderChek
Test to be more valuable than cytology as an adjunctive test to
cystoscopy. Furthermore, in addition to these reported urothelial cancers, BladderChek results also instigated further
extensive urologic investigations that ultimately revealed three
cases of renal cancers [UNPUBLISHED OBSERVATIONS].
Other biomarker assays for bladder cancer
There are currently two other biomarker assays that were
approved by the FDA for the monitoring of recurrent TCC
following surgery:
• BTA stat® Test (Polymedco, Inc.)
• Vysis® UroVision Bladder Cancer Recurrence Kit (Vysis,
Inc.)
No biomarker assay other than the NMP22 BladderChek
Test is approved for the diagnosis of de novo bladder cancer.
The BTA stat Test has low sensitivity for low-grade tumors
and is somewhat inferior to cytology for the detection of
high-grade tumors and CIS. Its specificity is affected by urinary tract infections, calculi, and instrumentation [38]. The
assay is based on the detection of bladder tumor associated
antigen, a complement factor H related protein (CFHrp).
This protein is a variant of human complement factor H
(FH). FH is known to play an important role in regulating
the complement system [106].
UroVision is a fluorescence in situ hybridization assay that
detects chromosomal abnormalities in bladder cells that are
indicative of cancer. It is a multiple assay that detects aneuploidy
in chromosomes 3, 7, 17 and 9p21. Initial research experiments
indicate that UroVision may provide a higher specificity with
perhaps comparable sensitivity to BladderChek [39]. However,
UroVision requires extensive laboratory preparation and
processing accompanied by a relatively astronomical cost (costs
in Alaska are tenfold – one log – greater for UroVision compared
with the NMP22 BladderChek Test).
Telomerase, a third marker, is only suitable for research applications at this time. Telomerase is a ribonucleoprotein that is
active in tumor cells and inactive in normal epithelial cells. Versions of the telomerase repeat amplification protocol (TRAP)
assay are available from Oncor, Inc. and Boehringer Mannheim. Limited study findings suggest that TRAP may provide
sensitivity comparable with BladderChek [40].
www.future-drugs.com
% detected
Number
230
Expert opinion
BladderChek in clinical utility
As a urologist practicing in a community setting, the author
believes that the NMP22 BladderChek Test has an immediate
and vital clinical utility. Moreover, it is possible that the practicing urologist in the community setting may be able to obtain a
greater clinical utility from BladderChek than suggested by the
pivotal trials conducted in an academic setting. The NMP22
BladderChek Test is the only tumor marker approved by FDA as
an aid in diagnosis of bladder cancer. It is immediately available,
has minimal cost and causes no patient discomfort.
The test is extremely easy to administer and read. Only four
drops of room temperature urine are required. As an adjunctive
test to cystoscopy, the simple point-of-care BladderChek is Clinical Laboratory Improvement Amendment (CLIA) waived,
meaning that a licensed laboratory technician is not required for
the test: a urologist or primary care physician is authorized to
conduct the test – no paperwork involved. In fact, there was a
91–95% concordance between lay readers and professional readers of the device in various evaluative settings: all spiked specimens read correctly by both lay and professional readers [41].
Thus, the test is almost foolproof. The NMP22 BladderChek
Test provides point-of-care results within 30 min and the test
minimizes human error and interoperator biases. Furthermore,
as an adjunctive test to cystoscopy, BladderChek combined with
cystoscopy improves the sensitivity of cystoscopy alone [37,GROSSMAN ET AL, SUBMITTED]. Thus the NMP22 BladderChek Test
actually improves the performance of urology’s standard best reference method of detecting bladder cancer.
BladderChek is particularly efficient at detecting high-grade
and/or invasive disease – the diseases that are life threatening.
In the trial at the author’s clinic, BladderChek detected all five
high-grade tumors while cytology identified only one. Clinically, it has been demonstrated that BladderChek detects 4.5times as many superficial cancers and 2.5-times as many invasive cancers as cytology [GROSSMAN ET AL, SUBMITTED]. Thus
NMP22 demonstrated a strong capability wherein cytology is
weak – the detection of low-grade tumors – while also detecting
the life-threatening diseases that cytology missed.
By providing a molecular view of the lower urinary tract, the
bladder and the upper urinary tract, BladderChek is capable of
detecting tumors along the entire urinary tract. As mentioned
previously, in the author’s clinic, positive BladderChek results
7
Tomera
instigated urologic investigations that revealed three cases of
renal cancers. Such findings were not reportable in the pivotal
or academic studies (the NMP22 BladderChek Test is not
approved for the detection of renal cancer).
Notably, in a small study, Ozer and coworkers determined
that preoperative NMP22 levels were elevated in patients with
renal cell carcinoma than in a matching control group with
benign urologic, gastroenterologic, surgical, and cardiovascular
conditions [42]. Undoubtedly, Ozer’s findings will provoke additional investigations into the use of NMP22 to detect upper
tract carcinoma.
Follow-up on false positives
As demonstrated in the pivotal study, the NMP22 BladderChek
Test detected four cancers that were not visualized during initial
cystoscopy: three invasive tumors and one CIS tumor, the CIS
disease initially reported as benign disease during initial cystoscopy [GROSSMAN ET AL, SUBMITTED]. The author proposes that
some BladderChek findings reported as false positives could be
better classified as prematurely positive, biochemically positive
or perhaps subclinically positive. Again, BladderChek detects
some cancers that cannot be visualized in the bladder due to
poor visibility, perplexing tumor manifestations or the presence
of a tumor in the upper urinary tract.
Within the domain of recurrent TCC, the author is supported
in this conclusion by Poulakis and coworkers, who, in monitoring
TCC patients, observed a high tendency toward tumor recurrence
in patients with false-positive, post-surgical NMP22 results [43].
In a similar vein, this prognostic value of NMP22 for recurrent
TCC was also demonstrated by Soloway and coworkers: patients
who were NMP22 positive 5 days postoperatively had a high
tendency towards recurrent TCC by 6-month follow-up [27].
BladderChek in the community versus the academic setting
In essence, the NMP22 Test Kit was an academic test and the
NMP22 BladderChek Test is a community test that validates
the academic findings. BladderChek proves that the pivotal
trial results are reproducible in the community setting (both the
NMP22 Test Kit and the BladderChek provide identical results
when using a 10 units/ml cut-off.) The results of BladderChek
in the community further validate the findings of early
researchers such as Soloway and coworkers, who suggested that
clinicians should be more aggressive in diagnosis and treatment
with any given NMP22 positive finding.
Consequently, BladderChek has changed our diagnostic algorithm and, in the author’s opinion, BladderChek provides better clinical utility in the community setting than indicated by
the research settings of the academic pivotal trials. First, the
community urologist knows his/her patients intimately,
whereas the academic researchers may not. Second, the community urologist can simply repeat the BladderChek test at
will, a procedure not allowed in the academic research trials.
Regarding the diagnostic algorithm, when cystoscopic findings are negative, a concurrent negative BladderChek result
provides a high level of confidence (~90%) that no cancer is
8
present. However, a negative cystoscope finding combined with
a positive BladderChek result is indication for further immediate urologic evaluation and/or extensive and aggressive followup, given that a significant number of BladderChek false positives are eventually recognized as being preclinically positive. By
the same reasoning, with a positive NMP22 finding, we might
change the diagnostic algorithm to include a contrast study,
whereas a negative result might indicate a more conservative
renal ultrasound.
Another finding the author has had in the community setting, although anecdotal in nature, is that NMP22 BladderChek Test provides a method of molecular staging. The
author has also found that the time required to confirm a positive result can be indicative of the disease grade. For example,
when BladderChek provides a positive result within 5 min,
high-grade, bulky or invasive disease has usually been found.
Such findings were not reported in the pivotal studies, nor
were findings of renal cell carcinoma and upper tract disease
addressed in the pivotal and academic studies, although renal
cell carcinomas have been detected in the authors clinic due
to BladderChek.
Such molecular staging also works in conjunction with cystoscopy. A positive cystoscopic finding in conjunction with a
positive BladderChek indicates a 5.7-times higher risk for invasive cancer [105]. Conversely, a positive cystoscope finding with
a negative BladderChek suggest that the tumor is more likely to
be superficial.
As a final benefit, when patients are reluctant to undergo cystoscopy, a positive BladderChek finding provides a strong psychological motivation that often prompts patients to acquiesce.
Cost effeciency
The NMP22 BladderChek Test has the potential to significantly reduce the cost of diagnosing de novo TCC and the
monitoring of recurrent TCC. Currently, voided urine cytology is used exclusively for the diagnosis of bladder cancer.
Using figures for Alaskan Medicare, each BladderChek assay
costs half as much as cytology while providing four-times the
sensitivity. For example in the author’s clinic, 248 patients were
evaluated to detect 18 cases of TCC during the BladderChek
pivotal trial,. Obviously, the use of BladderChek versus cytology would represent significant savings, particularly on a
national scale.
Lotan and Roehrborn used a decision tree analysis to
determine that urinary bladder cancer markers can be effectively incorporated into a surveillance protocol as long as the
marker cost is less than US$264, assuming a 20–80% yearly
recurrence rate and a 4–40% progression rate [44]. Hence, at
US$25, BladderChek could be used to supplement the
standard surveillance approach.
NMP22 also proffers the possibility of optimizing imaging
studies. BladderChek enables the physician to choose more
wisely between, for example, renal ultrasound, computed
tomography (CT) scan and IVP. For example, a nonsmoker
with hematuria and a negative NMP22 will undergo ultra-
Expert Rev. Mol. Diagn. 4(6), (2004)
NMP22® BladderChek® Test
sound whereas a smoker with a very strong BladderChek
result suggesting high-grade disease will undergo a CT scan,
since invasive and/or high-grade disease is much more likely.
On the other hand, an IVP might be indicated in other questionable patients due to the high cost of a contrast CT scan.
Finally, in terms of healthcare cost, patients with metastatic
genitourinary cancer have an estimated sixfold greater
expense than those with localized disease over the same
period of time [45].
Conclusions
Due to recent findings in pivotal studies and community clinical experience, the author believes that the NMP22 BladderChek Test represents a paradigm shift. Basically, BladderChek should be performed everytime a cystoscopy is
performed, with corresponding changes in the diagnostic protocol. This simple dipstick technology can save lives without
the cost and complexity of other technologies. Clinicians can
go out today and help patients. In the author’s practice, any
patient with a positive BladderChek test is told that ‘We need
to find an explanation for this test result!’
In summary, the NMP22 BladderChek Test in combination
with cystoscopy provides an improvement over cystoscopy
alone. Thus, BladderChek improves the sensitivity of what
now represents the gold standard in the diagnosis of de novo
tumors and/or detection of recurrent TCC. At every stage of
disease, BladderChek provides a higher sensitivity than cytology, which now represents the adjunctive standard of care. In
addition, BladderChek is less likely to miss the more serious,
life-threatening tumors that cytology misses. Finally, NMP22
is not dependent on intact cells, and there is no interference
in the assay from blood, common medications or substances
such as nicotine [107].
The early detection of bladder cancer improves both the
prognosis and the patient’s quality of life. Although the 5-year
survival rate is 94% for tumors detected at the superficial stage,
up to 25% of the 57,400 bladder tumors diagnosed each year
are not detected until they have become invasive or metastatic,
which lowers the 5-year survival rate to 48% and 6%, respectively [103]. Thus, approximately 12,500 Americans die annually from bladder cancer in the USA [102].
In addition, early stage disease requires less aggressive treatment, which impacts both quality of life and survival, as invasive disease often requires cystectomy and lifelong medical
maintenance of the compulsory urinary diversion. In terms of
healthcare cost, over the same period of time, patients with
metastatic genitourinary cancer have an estimated sixfold
greater expense than those with localized disease [46].
The incidence of bladder cancer is higher in men, individuals
over 60 years of age, and those exposed to carcinogens. Smoking is the most common cause, and doubles the risk of bladder
cancer [11]. Notably, in the BladderChek pivotal trial, the PPV
of BladderChek was 37% for those patients with highest risk
for bladder cancer (males over 60 years with a history of smoking), which is higher than the value typically reported for PSA
www.future-drugs.com
and prostate cancer [47].
Current, the guidelines of the American Urological Association for the diagnosis and management bladder cancer do not
include any biomarkers. These guidelines need to be updated to
include NMP22. With the inclusion of NMP22, the diagnostic
algorithm will change in a manner that provides the physician
with greater choice and diagnostic dexterity. Once these new
guidelines are implemented, NMP22 will guide subsequent
work-up and the intensity of monitoring, thereby reducing the
cost of diagnosis and management of bladder cancer, with a
concomitant reduction in morbidity and increase in survival.
Another area that must be addressed is the management of
hematuria in the primary care setting. Currently, there is no
consensus on when to test for microscopic hematuria in primary
care [11]. Given the analogy with PSA, at a cost of approximately
$25 US, this CLIA-waived test may be used by any primary care
physician for single-use or periodic testing of any high-risk
patient, that is, any patient over 40 years of age presenting with
hematuria with a history or histories of smoking, urologic disease or disorder, irritative voiding symptoms, urinary tract infections, analgesic abuse or pelvic irradiation [11]. Patients with
high-risk occupations or those at risk of exposure to carcinogens
might also undergo such testing. This at-risk group include:
•
•
•
•
•
•
•
•
•
Petroleum chemical or rubber workers
Hair dressers
Painters
Textile workers
Truck drivers
Aluminum electroplaters
Fire fighters
Arson investigators
Residents in regions with groundwater contamination of
arsenic, pesticides and/or industrial contamination
Five-year view
When the concept of tumor biomarkers was first introduced, urologists anticipated that biomarkers would replace cystoscopy. However, this expectation was unrealistic. What BladderChek accomplishes is improving the sensitivity of cystoscopy: complementing
the best available method to improve the best standard of patient
care. While cystoscopy is macroscopic, BladderChek provides a
molecular view of the urinary tract. Biomarkers are unlikely to
replace cystoscopy and are best suited as adjunctive tests.
Currently, BladderChek is relatively obscure. A major barrier
to adoption is that many physicians are either unaware of BladderChek or of the clinical utility offered by NMP22. Drivers to
adoption are likly to be:
• The consumer-oriented research and demands of patients
• Actuarial and/or financial executives of healthcare organizations and/or insurance companies
• Physicians (since good medicine is good business)
• Alteration of the American Urological Association and perhaps other national guidelines for the diagnosis and/or
management of bladder cancer
9
Tomera
It is likely that BladderChek will be adopted relatively
quickly for testing high-risk populations in both the urologic
and primary care settings, either due to lifestyle issues (smoking), medical histories, occupational exposure or the presence
of other contributory risk factors.
Eventually, panels of biomarkers, rather than a sole biomarker,
may be applied as adjunctive tests to provide both better sensitivity and specificity. For example, the combination specificity
of NMP22 and TRAP was reportedly 96% [40].
What is likely to replace cystoscopy is another form of
instrument or imaging modality. For example, in initial studies, the use of blue light cystoscopy appears more sensitive
than the current use of white light cystoscopy for the detection of bladder tumors [48]. It is also likely that new
approaches to magnetic resonance imaging, CT scans or ultrasound will provide some either improved imaging, improved
resolution, or some form of virtual cystoscopy. For example, a
research team found that saline hydration significantly
improved images of the proximal urinary tract, and that the
450-second delay extended the amount of the proximal tract
that can be visualized [49].
The NMP22 BladderChek Test will be an integral part of
the management and diagnostic work-up, with increasing
involvement of other imaging technology.
Key issues
• The NMP22® BladderChek® Test (Matritech, Inc.) is a tumor marker assay that represents a paradigm shift in the diagnosis and
management of urinary bladder cancer (transitional cell carcinoma).
• A simple dipstick technology, the NMP22 BladderChek Test should be performed with every cystoscopy.
• Cystoscopy is the gold standard, but the NMP22 BladderChek Test combined with cystoscopy is superior to cystoscopy alone.
• For the practicing urologist in the community setting, the actual clinical utility of BladderChek may exceed that suggested by the
pivotal trials in an academic setting
• At every stage of disease, BladderChek provides a higher sensitivity in the detection of bladder cancer than cytology, which now
represents the adjunctive standard of care, and BladderChek is unlikely to miss high-grade, life-threatening tumors.
• BladderChek provides four times the sensitivity of cytology at half the cost: it has the potential to reduce the healthcare cost on a
national level.
• BladderChek may be analogous to the prostate-specific antigen test, and eventually expand beyond the urologic setting into the
primary care setting for the testing of high-risk patients characterized by smoking history, occupational exposures or age.
• BladderChek is a qualitative second-generation device whose clinical utility was steadfastly proven in groundwork, first-generation
quantitative studies.
• Early detection of bladder cancer improves prognosis, quality of life and survival.
• Clinical diagnostic protocols and the guidelines of the American Urological Association for the diagnosis and management bladder
cancer should be updated to include tumor biomarkers.
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••
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the urinary tract with saline and delaying
imaging improve evaluation. Urology Times
January 1 (2004).
105
510(k) premarket notification, NMP22
BladderChek Test
www.accessdata.fda.gov/scripts/cdrh/cfdocs
/cfClia/Detail.CFM?ID=29453&PAGEN
UM=500
(Viewed October 2004)
106
Bladder Tumor Associated Antigen
www.btastat.com/bladder_tumor_
antigen_1.html
(Viewed October 2004)
107
Premarket approval 940035 S003, NMP22
BladderChek Test
www.accessdata.fda.gov/scripts/cdrh/cfdocs
/cfPMA/PMASimpleSearch.cfm?db=PMA
&id=7317#AOStatement
(Viewed October 2004)
Websites
101
Cancer Facts and Figures 2003, American
Cancer Society
www.cancer.org
(Viewed October 2004)
102
SEER Cancer Statistics Review 1975–2000,
National Cancer Institute
www.nci.nih.gov
(Viewed October 2004)
103
National Cancer Institute. CancerNET
PDQ Cancer Information Summaries
Bladder Cancer Screening.
www.nci.nih.gov/cancer_information/doc_
pdq
(Viewed October 2004)
Affiliation
•
Kevin M Tomera, MD
Alaska Regional Medical Plaza, 1200 Airport
Heights Drive, Suite 101, Anchorage, Alaska
99508, USA
Tel.: +1 907 276 1455
Fax: +1 907 276 1477
[email protected]
[email protected]
104 Michigan
Environmental Science Board.
Fire Fighter And Cancer Investigation Panel
Meeting Summary, MI, USA
www.michigan.gov/documents/ff0107_3717_7.pdf
(Viewed October 2004)
Expert Rev. Mol. Diagn. 4(6), (2004)