Blood Patterns: Prowling for Alzheimer’s Clues in Plasma

Series - American Academy of Neurology: 2014 Annual Meeting:

Undeterred by past setbacks, researchers continue to pursue a blood test for Alzheimer’s disease. At the 66th annual meeting of the American Academy of Neurology, held 26 April to 3 May in Philadelphia, one presenter described a microarray technique that picks up a particular antibody signature present in the blood of people with AD, but not in most healthy controls. Researchers will test the method to see if it can predict disease progression. Other groups focused on plasma Aβ, which in past research has correlated poorly with amyloid in the brain. At AAN, however, researchers reported that blood Aβ does associate with vascular disease, a risk factor for AD. Moreover, a highly toxic form of Aβ, pyroglutamate Aβ, abounds in the blood of middle-aged Down’s syndrome patients. Together, the results suggest that the plasma may harbor signs of AD, after all. 

Most attempts to develop a blood test for AD have focused on plasma proteins or lipids (see Oct 2007 news story; Mar 2014 news story; Jun 2013 Webinar). At AAN, Lucas Restrepo of the University of California, Los Angeles, described a different approach. He applied an antibody-based test developed by Stephen Johnston and Neal Woodbury at Arizona State University in Phoenix. In this method, researchers add a drop of blood to a microarray dotted with 10,000 random peptides, then look for antibodies that bind to the chip. Antibodies are more abundant than other blood proteins, making this test more sensitive than typical plasma assays, Johnston told Alzforum.

Detecting Disease-Specific Antibodies:

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. [Image courtesy of Stephen Albert Johnston.]

The researchers used blood samples from 44 AD patients and 53 age-matched controls from the Alzheimer’s Disease Neuroimaging Initiative (ADNI). As previously reported, all AD samples displayed a similar pattern, with antibodies binding to a shared subset of peptides. A small number of these peptides mimic segments of Aβ, but for the others, it is unknown what native antigens they resemble. Blood from AD patients also failed to bind some peptides typically bound by antibodies from healthy controls. Altogether, about 200 peptides are differentially bound by AD and control blood, Johnston said (see image below). Three control samples exhibited the AD antibody binding pattern and the test misclassified them as AD; the rest were negative. This gave the test a sensitivity of 100 percent and specificity of 94 percent for AD diagnosis (see Restrepo et al., 2013). The results agree with previous research on distinctive autoantibodies in AD blood (see Aug 2011 news story). 

AD Signature:

Antibodies in blood from cognitively normal controls (left columns) bind to different peptides than do antibodies from AD patients (right columns). [Image courtesy of Stephen Albert Johnston.]

The authors have now completed a follow-up study in 60 autopsy-confirmed AD patients and 60 age-matched controls from the Rush Memory and Aging Project. The researchers used a slightly different set of peptides in the array, but again saw a distinctive AD signature, with similar sensitivity and specificity to the earlier version. They plan to test the diagnostic in the Alzheimer’s Prevention Initiative’s Colombian study, which enrolls people from families with inherited AD. They will also test blood from people with mild cognitive impairment in other populations to see if their diagnostic can flag people with preclinical Alzheimer’s, Johnston said. 

Johnston uses the same microarray to test for various types of cancer, as well as infectious diseases. In 2010 he co-founded a company, Health Tell, to commercialize the technology. The company has made a new chip that contains 350,000 random peptides and is more sensitive and accurate at detecting disease, Johnston claimed.

Other groups are still trying to base diagnostic tests on Aβ in the blood, though no consistent relationship between it and AD risk has been shown (see Jan 2011 news story; Apr 2012 news story). Instead, researchers led by Sara Kaffashian at INSERM, Paris, looked at the relationship between plasma Aβ and markers of cerebrovascular disease. They followed more than 1,000 non-demented French participants over the age of 65 for four years. 

At AAN, Kaffashian reported that people who had the lowest Aβ40 at baseline and lowest Aβ42 at follow-up accumulated the greatest number of lesions in the white matter of their brain. Considered a marker of cerebrovascular disease, damaged white matter associates with AD risk, cognitive impairment, and brain amyloidosis, although what is cause and what is effect is unclear (see, e.g., Brickman, 2013; Noh et al., 2014). Moreover, the nearly 100 participants who had enlarged perivascular spaces at baseline also had the lowest plasma Aβ42. Perivascular spaces are the small clefts around blood vessels entering the brain that act as drainage pathways for interstitial fluid. Enlarged perivascular spaces correlate with lesions and white-matter damage in the brain and an elevated risk of stroke (see Rouhl et al., 2008; Doubal et al., 2010). 

“Our findings support a relationship between vascular [damage] and neurodegenerative mechanisms in cerebral aging,” Kaffashian wrote to Alzforum. When the brain’s small vessels are diseased, less blood reaches brain tissue, which may reduce the clearance of Aβ along perivascular drainage pathways, she noted. That results in less Aβ in the blood. In future studies, she will look at whether those participants who have markers of cerebrovascular disease along with low-plasma Aβ also deposit more amyloid in brain or blood vessels. If so, it would strengthen the idea that low-plasma Aβ could serve as a biomarker, Kaffashian believes. The vascular contribution to AD has become a hot topic of research (see May 2014 conference story part 1 and part 2). 

By contrast, researchers led by Pankaj Mehta and David Miller at the New York State Institute for Basic Research in Developmental Disabilities, Staten Island, chose to examine pyroglutamate Aβ (pyroGluAβ), a truncated form of the protein that aggregates readily. Previous studies have not found it in blood, perhaps because the peptide clumps together so quickly (see DeMattos et al., 2012). 

Mehta and Miller turned to Down’s syndrome patients, who carry an extra copy of the amyloid precursor protein (APP) gene and have high levels of Aβ products in their blood throughout life. The researchers generated an antibody to pyroGluAβ in rabbits. They reported that it detects as little as 100 pg/ml pyroGluAβ and does not cross-react with Aβ40 or Aβ42. With this tool, the authors screened plasma of 35 Down’s patients whose average age was 47. They found the concentration of pyroGluAβ to be eightfold higher than in 32 age-matched controls (about 2.5 ng/ml versus 0.29 ng/ml). The authors do not know if this plasma pyroGluAβ originates from brain or periphery. At their age, all the Down’s patients in the study would have accumulated Aβ plaques in the brain (see Jun 2011 news story). 

In future studies, the authors will gather longitudinal data to see whether plasma pyroGluAβ levels rise or fall over time as amyloidosis increases, and whether these changes relate to cognitive decline. Mehta noted that some participants with Down’s syndrome in this study had relatively low levels of pyroGluAβ in the blood compared to their peers. If that correlates with dementia severity, this marker could help identify people at the greatest risk for cognitive decline. Though dementia can be difficult to diagnose in people with Down’s syndrome, there are cognitive tests for this purpose (see Jun 2011 news story). Mehta also wants to adapt the method for AD patients. Like other researchers who presented at this annual meeting, Mehta has not given up on finding a blood test for Alzheimer’s.—Madolyn Bowman Rogers

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References

News Citations

  1. A Blood Test for AD?
  2. Do Lipids Hold the Key to Blood-Based Alzheimer’s Test?
  3. "Autoantibody-omics" Yields Potential Blood Biomarkers for AD
  4. In the Blood: What Can Plasma Aβ Tell Us About Alzheimer’s?
  5. Plasma Aβ as Biomarker: Not a Lost Cause, After All?
  6. It’s Not All About You, Neurons. Glia, Blood, Arteries Shine at Symposium
  7. Fluid Markers and Imaging Back Idea of Breached Blood-Brain Barrier
  8. Research Brief: Imaging Shows AD Pathology in Down’s Syndrome Brains

Webinar Citations

  1. Webinar: O Blood-Based Biomarker, Where Art Thou?

Paper Citations

  1. Restrepo L, Stafford P, Johnston SA. Feasibility of an early Alzheimer's disease immunosignature diagnostic test. J Neuroimmunol. 2013 Jan 15;254(1-2):154-60. PubMed.
  2. Brickman AM. Contemplating Alzheimer's disease and the contribution of white matter hyperintensities. Curr Neurol Neurosci Rep. 2013 Dec;13(12):415. PubMed.
  3. Noh Y, Seo SW, Jeon S, Lee JM, Kim JH, Kim GH, Cho H, Yoon CW, Kim HJ, Ye BS, Kim ST, Choe YS, Lee KH, Kim JS, Ewers M, Weiner MW, Lee JH, Werring DJ, Kang DR, Kim CS, Na DL. White matter hyperintensities are associated with amyloid burden in APOE4 non-carriers. J Alzheimers Dis. 2014;40(4):877-86. PubMed.
  4. Rouhl RP, van Oostenbrugge RJ, Knottnerus IL, Staals JE, Lodder J. Virchow-Robin spaces relate to cerebral small vessel disease severity. J Neurol. 2008 May;255(5):692-6. Epub 2008 Feb 21 PubMed.
  5. Doubal FN, MacLullich AM, Ferguson KJ, Dennis MS, Wardlaw JM. Enlarged perivascular spaces on MRI are a feature of cerebral small vessel disease. Stroke. 2010 Mar;41(3):450-4. Epub 2010 Jan 7 PubMed.
  6. Demattos RB, Lu J, Tang Y, Racke MM, DeLong CA, Tzaferis JA, Hole JT, Forster BM, McDonnell PC, Liu F, Kinley RD, Jordan WH, Hutton ML. A plaque-specific antibody clears existing β-amyloid plaques in Alzheimer's disease mice. Neuron. 2012 Dec 6;76(5):908-20. PubMed.

External Citations

  1. Colombian study

Further Reading

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