Paul Auger (Presenter)
Genentech
Authorship: Paul Auger, Stephen Schauer, William Mathews, Lee Honigberg, Kristin Wildsmith
Genentech, South San Francisco, Ca
| Short Abstract The microtubule-associated protein tau has been implicated in the pathogenesis of Alzheimer’s disease (AD) and other neurodegenerative disorders. Tau represents not only an attractive therapeutic target for AD, but also a candidate biomarker for AD and other neurodegenerative disorders. Tau is a very heterogenous protein with multiple splice forms and numerous post-translational modifications. Specific tau isoforms have been hypothesized to play a role in the onset and progression of AD. An ideal tau biomarker assay would be capable of measuring distinct tau isoforms. We have set out to identify what soluble isoforms are measureable in AD and control patients using an immunoprecipitation mass spectrometry (IPMS) based assay. |
Long Abstract
The microtubule-associated protein tau has been implicated in the pathogenesis of Alzheimer’s disease (AD) and other neurodegenerative disorders. Tau represents not only an attractive therapeutic target for AD, but also a candidate biomarker for AD and other neurodegenerative disorders. Tau is a very heterogenous protein with multiple splice forms and numerous post-translational modifications. Specific tau isoforms have been hypothesized to play a role in the onset and progression of AD. An ideal tau biomarker assay would be capable of measuring distinct tau isoforms. We have set out to identify what soluble isoforms are measureable in AD and control patients using an immunoprecipitation mass spectrometry (IPMS) based assay. This assay will be useful in informing and guiding further affinity based assay development The IPMS assay we have constructed is a quantitative assay to measure tau isoforms in cerebrospinal fluid as well as plasma. We have selected 14 peptides to monitor by parallel reaction monitoring mass spectrometry for quantitation of tau in matrix. The peptides were identified using recombinant isoform proteins. Concurrently, stable isotope labeled peptides were synthesized for the generation of standard curves for quantitation. Full length (2N4R) heavy protein was used as a process control as well as an internal quantitative reference. We compared several commercial antibodies for the immunoprecipitaiton of tau from the matrices to enrich for our target protein and quantify the isoforms present. By developing this assay we will gain insight into what portions of the total tau measurement are related to specific isoforms in CSF and plasma by comparing the data from the isoform-specific peptides with the total tau peptide measurements. The complex nature of tau biology and the many isoforms of tau that are present make MS a powerful tool to help properly characterize tau as a biomarker.
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