Validation of a Serum Screen for Alzheimer's Disease Across Assay Platforms, Species, and Tissues

Abstract

Background There is a significant need for rapid and cost-effective biomarkers of Alzheimer’s disease (AD) for advancement of clinical practice and therapeutic trials. Objective The aim of the current study was to cross-validate our previously published serum-based algorithm on an independent assay platform as well as validate across tissues and species. Preliminary analyses were conducted to examine the utility in distinguishing AD from non-AD neurological disease (Parkinson’s Disease). Methods Serum proteins from our previously published algorithm were quantified from 150 AD cases and 150 controls on the Meso Scale Discovery (MSD) platform. Serum samples were analyzed from 49 Parkinson’s disease (PD) cases and compared to a random sample of 51 AD cases and 62 controls. Support vector machines (SVM) were used to discriminate PD vs. AD vs. NC. Human and AD mouse model microvessel images were quantified with HAMAMATSU imaging software. Mouse serum biomarkers were assayed via MSD. Results Analysis of 21 serum proteins from 150 AD cases and 150 controls yielded an algorithm with sensitivity and specificity of 0.90 for correctly classifying AD. This multi-marker approach was then validated across species and tissue. Assay of the top proteins in human and AD mouse model brain microvessels correctly classified 90–100% of the samples. SVM analyses were highly accurate at distinguishing PD vs. AD vs. NC. Conclusions This serum-based biomarker panel should be tested in a community-based setting to determine its utility as a first-line screen for AD and non-AD neurological diseases for primary care providers.

Serum-based Protein Profiles of Alzheimer’s Disease and Mild Cognitive Impairment in Elderly Hispanics

“…All samples were assayed in duplicate via a multiplex biomarker assay platform using electrochemiluminescence on the SECTOR Imager 2400A from Meso Scale Discovery (MSD) [31] per our previously published methods [18]. The MSD platform has been used extensively to assay biomarkers associated with a range of human diseases including AD and have wellestablished properties of being more sensitive and requiring less sample volume than conventional ELISAs.…”

“…The MSD platform has been used extensively to assay biomarkers associated with a range of human diseases including AD and have wellestablished properties of being more sensitive and requiring less sample volume than conventional ELISAs. The markers assayed were from our previously generated and cross-validated AD algorithm [17,18] and included: FABP, B2M, PPY, CA-125, CRP, sVCAM, THPO, A2M, TNF-α, TN-C, IL-5, IL-6, IL-7, IL-10, IL-18, I309, FVII, TARC, SAA, sICAM, eotaxin 3, adinoponectin, IL-1β and MIP-1α. To be included in the study, analyte concentrations should exceed the limit of detection in >75% of all the samples for each respective analyte.…”

“…Chronic kidney disease-related dysfunction and presence of low-grade inflammation in older adults; upregulated in AD and MCI [33,34] PPY Increased levels in plasma and CSF in subjects with AD and MCI [35] CA-125 Mainly upregulated in ovarian cancer, in older women it is associated with worse functional status; to date no association has been found with AD [36] CRP Associated with neurofibrillary tangles and senile plaques in AD brain tissue [37] sVCAM1 Member of the immunoglobulin superfamily, elevated levels have been found in plasma of AD subjects [38] THPO Induces the proliferation and maturation of megakaryocytes; plasma levels show difference in subjects with dementia compared with controls [39] A2M Mediates Aβ toxicity, clearance and degradation; upregulated in AD subjects [40,41] Eotaxin3 Together with ApoE it discriminates between control and AD; increased levels have been found in plasma and CSF in AD subjects [35,42] TNF-α Involved in neuronal apoptosis and formation of intracellular neurofibrillary tangles and amyloid plaques; higher blood concentrations were detected in AD subjects [43,44] TN-C Involved in the development of neurons and axons, neuronal regeneration, microglial activation and inflammatory responses; increased levels have been found in plasma and CSF in AD subjects [34,35,45] IL-5 Specific hematopoietic growth factor that together with other proteins forms a biomarker profile that can distinguish AD from controls [18] IL-6 Involved in neuronal apoptosis and formation of intracellular neurofibrillary tangles and amyloid plaques; elevated levels have been found in blood but not in CSF in AD subjects [43,44] IL-7 Induces the synthesis of inflammatory mediators such as IL-1 and IL-6; in plasma there is no difference between AD subjects compared with controls, but in CSF there is a downregulation in MCI subjects [46] IL-10 Anti-inflammatory properties and may play a role in schizophrenia pathogenesis; concentrations did not differ between AD subjects and controls [44,47] IL-18 Probable mediator of cerebral pathogenic processes, microglia activator, involved in neuroendocrine and neuroimmune functions; higher blood concentrations were detected in AD subjects [43,44,48] …”

“…Evidence shows that brain imaging and cerebrospinal fluid biomarkers are highly accurate in detecting disease presence; however, these methods are not cost-and time-effective strategies for primary care clinical settings particularly in low-resource settings. In this regard, recent research into blood-based biomarkers of AD has produced encouraging results that suggest that blood-based screening is a viable approach to early diagnosis [13][14][15][16][17][18][19][20]. Together, the results of these studies provide strong evidence that a blood-based screening approach can be useful in discriminating AD from healthy controls as well as from other dementias [21].…”

“…We examined 24 proteins from our previously established algorithm in serum across diagnostic groups and generated a biomarker profile of disease status. The battery of proteins from our previously generated and cross-validated AD algorithm [17,18] consists of various proteins responsible for a wide range of physiological characteristics including chemotactic activity, hematopoiesis, growth factors, tissue markers (receptor and stimulation of B cells and T cells), including proteins that promote the expression and secretion of chemokines, that function as hormones, proteins of acute phase mechanism, proteins involved in inflammatory responses and disease markers and others involved in the development of neurons and axons. Based on previous reports, we hypothesized that our previously generated blood-based profile would yield an algorithm highly accurate in detecting AD.…”