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A Blood Test Accurately Shows an Immunosignature for Alzheimer's Disease, According to Small Study



IMMUNOSIGNATURING: Blood carries antibodies that bind to peptides on an array and can be detected with anti-IgG secondary antibodies (red) conjugated to a fluorescent label (white dots). Blood antibodies reflecting specific diseases bind to unique combinations of peptides.

Investigators described a novel technique involving the detection of unique disease patterns in the interaction with and binding of a subject's antibodies to random-sequence peptides on a microarray. In the current experiment, they detected a pattern they deemed to be Alzheimer's disease-like.

PHILADELPHIA—A blood test through which a patient's immunological “fingerprint” is assessed is highly accurate in determining who has been clinically diagnosed with Alzheimer's disease (AD), according to a small study described here at the AAN Annual Meeting in April.

The technique of using antibodies as biomarkers for AD should be explored further and could become an option that is more practical than other biomarkers requiring expensive imaging or a spinal tap, the researchers said.

The approach involves so-called “immunosignatures,” in which random-sequence peptides are used to look for antibodies to disease. Lucas Restrepo, MD, PhD, an assistant professor of neurology at the University of California, Los Angeles, who presented the new data, described the approach at the meeting.

Antibodies present in a single drop of blood are spread over a glass slide containing an array of some 10,000 random sequence peptides, he explained. The antibodies then selectively bind to these peptides with varying degrees of strength. Through these interactions, an antibody-binding pattern — an immunosignature — emerges that can help distinguish a disease.

“Everybody has a different profile,” Dr. Restrepo said. In the current experiment, a determination that a pattern was “AD-like” was made by assessing the statistical differences between two groups of patients. Each spot on the array gives a number, depending on the strength or weakness of the binding of donor antibodies to specific peptides.

Then researchers compared the top values, looking for statistical differences. They found that 30 to 150 peptides are the best classifiers, depending on the groups of patients being assessed.

Dr. Restrepo said the feasibility of this approach was described in a published paper in The Journal of Neuroimmunology in 2013.


In the current study, researchers tested 44 patients with a clinical diagnosis of AD, as well as 53 cognitively normal, healthy, age-matched controls — participants in the Alzheimer Disease Neuroimaging Initiative (ADNI), a longitudinal study that has been validating the use of biomarkers including blood tests, tests of cerebrospinal fluid, and MRI/ PET imaging for AD clinical trials and diagnosis.

A total of 47 samples had an “AD-like” signature, while 49 had a “normal” signature. The test correctly classified all of the 44 AD samples, but misclassified three controls. That gave the test a sensitivity rate of 1.0, a specificity rate of 0.94, and an overall accuracy greater than 95 percent.


DR. LUCAS RESTREPO said a determination that a pattern is “AD-like” was made by assessing the statistical differences between two groups of patients. Each spot on the array gives a number, depending on the strength or weakness of the binding of donor antibodies to specific peptides.

This approach could help fill a great need in the field of dementia: a simple test to specifically identify AD, he said. With disease-modifying drugs being developed, “diagnostic precision has become a necessity.”

“We asked ourselves why every person has an immunological fingerprint,” Dr. Restrepo said. “We speculated that perhaps this has to do with the things that people have been exposed to in life — an infection here and there, and maybe exposure to misfolded proteins that accumulate in the brain.

“There are many reasons why an antibody would be an ideal biomarker,” he continued. “We have lots of antibodies. They're produced early in disease. They're simple to assay. And they're stable.”

More work to further confirm the reliability of the technique is needed, he acknowledged. “We believe this can be a promising platform,” he said. “We need to test more plasma samples. One of the important things that we're interested in doing is looking at people who have autopsy confirmation of AD.”

If the results are confirmed, “immunosignatures could be utilized to screen dementia patients prior to more expensive or invasive tests such as amyloid imaging and cerebrospinal fluid analysis,” he said.


David S. Knopman, MD, FAAN, a professor of neurology at the Mayo Clinic in Rochester, MN, said that if the approach pans out, it would certainly be welcome in the field. “I think that finding a blood biomarker for some aspect of AD is incredibly important because, as the researchers point out, and as is well-known, imaging biomarkers are expensive and cerebrospinal fluid biomarkers involve an invasive procedure. So finding a blood test would be great,” Dr. Knopman said.

The immunosignature approach is worth exploring, he said, but “the devil is in the details” and he said there are reasons to remain skeptical for now. For example, while patients in the study were described as having a clinical diagnosis of AD, that might be misleading.

“The investigators should have said AD-dementia. When you talk about AD these days, you need to differentiate the pathophysiology from the clinical diagnoses,” he said. “Even in the best of hands, somewhere between 10 and 30 percent [of people with a clinical diagnosis of AD] are going to have non-AD diagnoses pathologically.”

In the recent solanezumab clinical trials, for example, 30 percent of the AD-dementia patients who were enrolled, and underwent amyloid imaging, were actually amyloid-negative, Dr. Knopman noted.

Another important point, Dr. Knopman said, is that the investigators did not report how many of the control patients in the study were amyloid-positive, even though it's known through the Alzheimer's Disease Neuroimaging Initiative that roughly 20 to 30 percent of controls were amyloid-positive and have preclinical AD.

“So what's this blood test picking up?” Dr. Knopman said. “You have to ask, who were the controls?”

He pointed out that the methodology involved in the development of the immunosignatures, described in abstracts and published papers, can be “difficult to follow,” and that you “have to take their methodology on face value as a black box.” He added, though, that the authors have time to further clarify the process.

“Their results are intriguing and I think they should be followed up with a larger study in a longitudinal framework and one in which imaging or CSF biomarkers are available,” he said.

The need for that kind of information is what makes this a difficult area to research.

“To go to that next level to have well-characterized patients and controls is really expensive and time-consuming and it takes a huge amount of infrastructure,” Dr. Knopman said. “It's not that you can just go out and recruit a few people here and there. And so it really means that people like Dr. Restrepo need to be tied into a big clinical research infrastructure to properly test this. And that's a real challenge.”


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VIDEOS ON DEMAND: Is a blood test feasible for the detection of Alzheimer's disease? In a panel discussion, David S. Knopman, MD, FAAN, a professor of neurology at the Mayo Clinic in Rochester, MN, Neurology Today Editor-in-chief Steven P. Ringel, MD, FAAN, and Associate Editor Robert Holloway Jr., MD, FAAN, offer an analysis of why a blood test for Alzheimer's disease is needed and the challenges that remain ahead for ensuring that it is accurate: Read more about the Annual Meeting abstract, which inspired the discussion:


•. AAN Annual Meeting Abstract: High accuracy of a microarray-based blood test for Alzheimer's disease:
    •. Restrepo L, Stafford Johnston, et al. Feasibility of an early Alzheimer's disease immunosignature diagnostic test. J Neuroimmunol 2013;254:154–160
      •. More research on immunosignatures: