A team of Johns Hopkins biochemists has identified a
mixed bag of five key proteins out of
thousands secreted into blood draining from the heart's
blood vessels that may together or in certain
quantities form the basis of a far more accurate early
warning test than currently in use of impending
heart attack in people with severely reduced blood flow, or
ischemia.
The work, involving more than a dozen scientists and
taking more than a year to perform, is
believed to be the largest protein analysis ever done at
Johns Hopkins. It was based on 76 arterial
blood samples from 19 men and women taken immediately
before and after a period of medically
induced ischemia lasting as long as 45 minutes.
All had ischemia induced through accelerated pacing of
the heart's main chambers. Blood
samples were provided by cardiologists at the University of
Texas Southwestern. Key to the
researchers' selection criteria for which proteins to
analyze from among tens of thousands in the
blood was what they call "a pipeline approach."
"From the start, we knew that we were looking for
rare, almost unique biomarkers that bore
some direct relationship with ischemia," said study senior
investigator
Jennifer Van Eyk, whose first
step was to remove from the analysis common blood proteins,
such as albumin and globulins. That left
batches of 400 proteins for in-depth measure of any changes
before and after ischemia.
The scientists' analysis, presented Nov. 9 at the
American Heart Association's annual Scientific
Sessions in New Orleans, found that only the five proteins
were present in significantly increased
amounts after ischemia occurred, with at least a doubling
in the blood concentration, compared with
those recorded during healthy blood flow. These were
lumican, semenogelin, angiogenin, extracellular
matrix protein and so-called long palate, lung and nasal
epithelium carcinoma-associated protein 1.
All the proteins are believed to originate in the
heart, but they can also be found in other
tissues varying from the corneas of the eyes (lumican) to
semen. Semenogelin, as it is known, has never
before been seen in the heart, while others, such as
angiogenin, are more predictably found in growing
blood vessels and muscle tissue, and are actively involved
in tissue repair. Little is known about the
remaining two, which ironically have the longest names:
extracellular matrix protein, secreted in a rare
inflammatory disease; and long palate, lung and nasal
epithelium carcinoma-associated protein 1,
thought to play a role in innate immunity.
The Johns Hopkins biochemists say that the presence of
all or even a selected set of these
proteins in a simple, rapid blood test could aid emergency
paramedics and physicians during the critical
12- to 24-hour window before ischemia causes substantial
heart tissue damage or death from heart
attack.
A positive reading on a blood test incorporating these
proteins, they add, could provide first
responders with advance warning to take urgent action, such
as using blood thinners like aspirin to
prevent clotting, or performing cardiac catheterization to
check for any more blockages in the blood
vessels feeding the heart, which may in turn prompt more
aggressive treatment. Further actions could
involve angioplasty, in which a balloon device is threaded
into the heart's surrounding blood vessels and
then expanded to widen the arteries, or even surgery.
"Our results lay the foundation for a first-of-a-kind,
early-warning system that could save tens
of thousands of people on the brink of a heart attack,"
said Van Eyk, a professor at the Johns Hopkins
University School of Medicine and its Heart and Vascular
Institute. "People experiencing chest pain
too often come to the emergency room, with subsequent
electrocardiogram, also called EKG, readings
not showing any evidence that a heart attack has occurred
but still leaving open the question of
whether or not a heart attack is imminent and about to
happen or has already happened," said Van Eyk,
director of the
Johns Hopkins NHLBI Proteomics Group and the Proteomics
Center at Johns Hopkins
Bayview Medical Center, where the protein analysis took
place.
More than 8 million American men and women have
suffered at least one heart attack, the cause
of almost 157,000 deaths in 2005 alone. Nearly 6 million
have suffered a stroke, which caused
150,000 deaths in 2005.
Van Eyk says that people frequently have symptoms of
chest pain, shortness of breath and
dizziness, with pale or clammy skin coloring, while
arterial blood is constricted but not yet closed. But
this myriad of complaints can just as easily be mistaken
for the more everyday, less serious problems
of heartburn, stomach cramps or gas. In 2006, the Centers
for Disease Control and Prevention
reported more than 12,000 visits to doctors' offices and
emergency rooms by people complaining of
chest pain.
A new test based on these five proteins, Van Eyk says,
could provide a more definitive answer to
the question "how serious is it?" much earlier than
existing assays for heart attack, such as tests for
troponin proteins I and T.
Van Eyk says that commercially available tests for
cardiac troponin, which is released into the
blood in telltale patterns for heart attack, provide
results "too late to take preventive action" and
"after some damage has already occurred." Troponin lab
tests also depend on the heart muscle dying
first, a process that can take hours to detect, "so a
negative reading is unreliable and can still mean
that an ischemic problem is about to happen or has already
happened," she said.
In the study, the protein analysis was conducted by
mass spectrometry machines that can
measure the presence of proteins in minute amounts. The
machines, operated six days a week for six
months, consumed more than 3,700 hours of spectrometric
analysis.
Researchers next plan to verify the presence of the
five proteins in a larger study with at least
150 participants, and more than 1,000 blood samples.
Simultaneously, they plan further analysis of the
proteins to map their molecular structures so that an
antibody can be identified to bind to one or
several of the proteins, laying the basis for a blood test
for ischemia. And they will conduct tests to
verify that their study findings also apply to ischemia in
stroke.
Funding support for this study was provided by
Inverness. The technology development in the
study and the "pipeline approach" were supported by the
Johns Hopkins Bayview Proteomics Center.
The Johns Hopkins NHLBI Proteomics Group is one of 10
centers funded as part of the United States'
seven-year program dedicated to the study of proteomics and
understanding the functions of proteins
in the development of cells, tissues and organisms, in both
normal and disease processes. Van Eyk has a
patent pending on the protein analysis. Under an option
agreement with The Johns Hopkins University,
Inverness Medical Innovations' Unipath Ltd., of Bedford,
United Kingdom, has the right to negotiate a
license to the patent.
Other Johns Hopkins researchers who took part in this
study were Qin Fu, Simon Sheng,
Steven Elliott and Miroslava Stastna. Additional support
was provided by James de Lemos at the
University of Texas Southwestern.