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Houston Researchers Looking for Salivary Markers for Lupus Diagnosis, Monitoring

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NEW YORK (360Dx) – University of Houston researchers are exploring the potential of salivary protein markers to diagnose and monitor systemic lupus erythematosus (SLE).

Chandra Mohan, professor of biomedical engineering at the university, has received a $386,599 grant from the National Institutes of Health that he is using to evaluate the usefulness of measuring in saliva anti-double-stranded-DNA (anti-dsDNA) antibodies, a known lupus marker.

Levels of these antibodies are elevated in the blood of lupus patients, and researchers have determined they are also detectable in saliva.

"So, the question is, is the diagnostic potential of salivary anti-dsDNA antibodies as good as it is in blood or serum," Mohan said. He also plans to use the grant to investigate whether other lupus markers for purposes like predicting the severity and path of the disease are present in saliva.

"Lupus is a very heterogenous disease," he said, raising as an example the renal complications that often result from the condition. "At the very least, you would like to try to distinguish between patients with and without [lupus-related] renal disease and predict who might die of renal disease [in the near future]."

"One of the additional aims of our proposal is to see whether we can [identify markers] to subtype patients based on [which] organs are involved and to predict whether or not their disease is going to turn severe," he said.

Mohan said that in preliminary work supporting the grant proposal, he and his colleagues demonstrated that they could distinguish between SLE patients and healthy controls based on levels of anti-dsDNA antibodies in their saliva. They also identified two additional protein markers that were elevated in the saliva of SLE patients.

"Based on that we are expanding the study to look at increased numbers of patient samples and controls, including both healthy controls and controls with systemic autoimmune diseases," Mohan said. In all, they plan to look at 90 SLE patients, 30 healthy controls, and 30 disease controls.

The researchers will also collect blood samples of these patients to compare the performance of the markers in blood and saliva.

Mohan said he is pursuing saliva markers due to the ease of sampling, particularly in resource-constrained areas where diagnosing SLE can still be a challenge. He added that beyond sampling considerations, there are reasons to think saliva could be a more effective source for biomarker discovery than blood.

Blood, Mohan noted, is an extremely complex matrix with a large number of proteins expressed across a wide range of abundances. Saliva, by comparison, "is relatively clean," he said. "There are relatively few proteins in it."

This obviously limits its potential as a source of biomarkers, but it also means that markers that are present can be more easily detected.

Mohan said that one other factor that could aid saliva-based protein marker detection is the presence in saliva of mucin, a substance that has been shown to preserve and stabilize proteins. "So, there's a chance that protein biomarkers may be more stable in saliva than in blood," he said.

The researchers will also collect blood and urine samples from the study subjects for use in biomarker discovery, which they will pursue using targeted immunoassays as well as Somalogic's aptamer-based SomaScan platform, which Mohan said will allow them to assess the levels of around 1,300 proteins in these samples.

He said that his group has used mass spec-based biomarker discovery approaches in the past but that they had found the SomaScan platform to be more effective, particularly for assessing the potential of low-abundance markers.

While Mohan and his colleagues hope to demonstrate the utility of salivary anti-dsDNA antibody levels for diagnosing SLE, this is not the area of greatest clinical need, he said.

"Diagnosis is pretty straightforward these days," he said. "The real problem is trying to predict which patients are going to dial up, let's say, to renal disease or cardiovascular disease. The real challenge is trying to determine which patients are stable and which patients are going to die of the disease down the line. That's where the real need for biomarkers is."

Currently, doctors rely on procedures like biopsies for assessing complications like kidney disease in SLE patients. "But biopsies can't be repeated frequently," Mohan said. "It can be done once, a couple of times at the most.

Additionally, he noted, "by the time the kidney is affected, it's already too late. What we want to have is a biomarker that can predict that someone's kidney is going to become [damaged], say, a month from now. That would be very useful."

The hope, Mohan said, is that he and his colleagues will identify promising markers in this initial study that they can then test in a longitudinal trial to assess their predictive value.

"We want to have really solid data before we propose doing it longitudinally," he said.

Mohan is working under a separate, four-year $1.4 million NIH grant to develop a point-of-care device for measuring protein biomarkers for early detection of kidney flare-ups in SLE patients. In this work he is collaborating with his University of Houston colleague Richard Willson, a professor of chemical and biomolecular engineering to develop a smartphone-based test.

The test uses the basic lateral flow approach featured in home pregnancy tests but uses nanoparticles fixed with antibodies to the proteins of interest for detection. When the markers bind to the antibodies, the nanoparticles emit a detectable signal. Because the candidate markers are present in lower levels in healthy individuals, the test needs not only to detect the markers but also provide a relatively quantitative measurement of their expression levels. This is done via a smartphone camera and an app that calculates biomarker levels based on the brightness of the nanoparticle emissions.

Willson published on the technology underlying the test in a 2014 paper in Analytical Chemistry.