New highly sensitive rodent and human tests for soluble amyloid precursor protein alpha quantification: preclinical and clinical applications in Alzheimer's disease

Background: Amyloid precursor protein (APP), a key molecule in Alzheimer's disease (AD), is metabolized in two alternative cleavages, generating either the amyloidogenic peptides involved in AD pathology or the soluble form of APP (sAPP alpha). The level of amyloidogenic peptides in human cerebrospinal fluid (CSF) is considered to be a biomarker of AD, whereas the level of sAPPa in CSF as a biomarker has not been clearly established. sAPPa has neurotrophic and neuroprotective properties. Stimulating its formation and secretion is a promising therapeutic target in AD research. To this end, very sensitive tests for preclinical and clinical research are required. Methods: The tests are based on homogenous time-resolved fluorescence and require no washing steps. Results: We describe two new rapid and sensitive tests for quantifying mouse and human sAPP alpha. These 20 mu l-volume tests quantify the levels of: i) endogenous mouse sAPP alpha in the conditioned medium of mouse neuron primary cultures, as well as in the CSF of wild-type mice, ii) human sAPP alpha in the CSF of AD mouse models, and iii) human sAPP alpha in the CSF of AD and non-AD patients. These tests require only 5 mu l of conditioned medium from 5 x 10(4) mouse primary neurons, 1 mu l of CSF from wild-type and transgenic mice, and 0.5 mu l of human CSF. Conclusions: The high sensitivity of the mouse sAPP alpha test will allow high-throughput investigations of molecules capable of increasing the secretion of endogenous sAPP alpha in primary neurons, as well as the in vivo validation of molecules of interest through the quantification of sAPP alpha in the CSF of treated wild-type mice. Active molecules could then be tested in the AD mouse models by quantifying human sAPPa in the CSF through the progression of the disease. Finally, the human sAPP alpha test could strengthen the biological diagnosis of AD in large clinical investigations. Taken together, these new tests have a wide field of applications in preclinical and clinical studies.