October 7th, 2011 - CHORI scientist Michael Oda, PhD, has developed a novel assay for determining a patient’s risk for coronary artery disease. Dr. Oda can assess the quality of a person’s high density lipoprotein (HDL) by probing their blood plasma with paramagnetically labeled apolipoprotein A-I (apoA-I), the primary protein component of HDL. Commonly known as the “good cholesterol,” HDL isn’t cholesterol at all, but the lipoprotein in the blood that is responsible for withdrawing cholesterol from cells and transporting it through the blood stream to the liver, where it is processed.
“What we are able to measure is the ability of a person’s HDL to effectively withdraw cholesterol from the cell,” says Dr. Oda.
|“While traditional measurements can only tell us how much cholesterol is associated with HDL, this assay can tell us how well that HDL is actually functioning.”
Cardiovascular disease is the leading cause of death in the United States, and is poised to become the number one cause worldwide. Although much research has focused on boosting "good" HDL levels, recent drug trials show that increasing HDL cholesterol levels in the bloodstream alone is not sufficient to reduce heart disease risk. Rather, results suggest, it is the quality of the HDL – or its functional capacity – that may be the key to cardiovascular disease risk.
As such, Dr. Oda's innovative new assay is of particular import, providing the basis for a much more accurate clinical diagnosis of cardiovascular disease risk than ever before.
"When apoA-I binds to HDL, however, this amplitude goes up by about 2-orders of magnitude, allowing us to determine the rate and degree of HDL binding.”
|“The current hypothesis is that dysfunctional HDL causes people to be more susceptible to cardiovascular disease. That dysfunctionality is associated with a reduction in the ‘labile pool’ of HDL, a species of HDL that is responsible for withdrawing cholesterol from cells.”
Without enough labile HDL to remove cholesterol from cells, the functional capacity of HDL is drastically reduced. Current HDL measures only quantify the level of cholesterol on HDL, not how much HDL is capable of mobilizing cholesterol. Dr. Oda’s assay, however, can measure the latter by determining the binding capacity of HDL for the paramagnetically-labeled apoA-I.
“Because apoA-I specifically binds to HDL in its labile form and no other blood plasma components, measuring the binding capacity of HDL to the paramagnetically-labeled apoA-I in our assay provides a relative measure of how much labile HDL a person has, and thus, a relative assessment of their HDL functional capacity,” Dr. Oda says.
Using apo-AI as a probe, Dr. Oda then uses a method called an electron paramagnetic resonance spectrometry (EPR) to determine the rate of HDL binding.
“The EPR signal of paramagnetically labeled apoA-I when not bound to HDL is relatively low in amplitude," explains Dr. Oda.
While paramagnetic labels are not often used in blood because the reducing power of blood can destroy them, Dr. Oda was able to choose a binding site for the label within the apoA-I protein that rendered them very stable in both lipid-free and lipid-bound states. This allowed the label to remain stable in the blood for up to four hours – plenty of time to conduct a 30-second assay.
“The results showed that relative to a maximum response control, where in all the apoA-I is lipid bound, healthy patients had an 80 percent response rate, give or take about five percent," Dr. Oda says. "In patients with metabolic syndrome, which is considered a precursor for diabetes and heart disease, the response was about 55 percent, while those who have established diabetes had a response of less than 50 percent.”
The triple-tiered results showing higher to lower responses associated with healthy to less healthy or already ill individuals suggest that the assay can provide a direct measure of HDL function. Even more importantly, by capturing how well HDL actually functions, the new assay can identify people at risk for heart disease who might otherwise be missed, due to having high HDL numbers and thus a “good” cholesterol assessment – even though their HDL functionality might actually be low.
|“This will enable researchers to develop new drugs for cardiovascular disease, assess the nutritional affects of diet on how well particular drugs work, and to determine how to make existing drugs even better.”
While additional validation studies are still needed for FDA approval, Dr. Oda’s assay has the potential to revolutionize heart disease risk assessment by providing a diagnostic tool that both accurately identifies individuals at risk for heart disease, assesses how well a particular treatment is working, and evaluate the impact of diet and exercise on an individual basis.
Tuesday, November 22, 2011 12:20 PM