Leckey, Claire Alexandra Evelyn;
(2023)
Developing translational kinetic biomarkers for neurodegenerative disease.
Doctoral thesis (Ph.D), UCL (University College London).
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Abstract
Neurofilament light chain (NfL), a protein integral to filamentous neurofilaments in the neuronal cytoskeleton, is an established biomarker of neurodegeneration that is elevated in cerebrospinal fluid (CSF) and blood across multiple neurodegenerative diseases. While this biomarker has been extensively measured in patient cohorts, important questions regarding its dynamics in the central nervous system (CNS) remain unanswered. Stable isotope labelling kinetics (SILK), an analytical technique used for capturing production and clearance rates in vivo and in vitro, has previously investigated amyloid-β and tau kinetics in Alzheimer’s disease, confirming and challenging current hypotheses regarding their kinetics. The recent inclusion of NfL as an outcome measure in clinical trials presents an urgent need to investigate the biomarker’s physiology. Therefore, the overall aim of this study was to develop highly sensitive and quantitative analytical methods to study NfL half-life and kinetic rates in induced pluripotent stem cell (iPSC)-derived neurons and patients with neurodegenerative diseases. A rapid, hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS) assay was adapted for quantitation of 13C6-leucine enrichment in plasma and CSF, which showed excellent agreement and correlation (r=0.99) with the current method used for SILK. A targeted immunoprecipitation–tandem mass spectrometry (IP-MS/MS) method for NfL was established and preliminary in vitro and in vivo SILK analysis performed. Analysis of CSF samples from the Tau Tangles SILK study found that NfL turnover in the CNS is exceptionally slow, with a maximum labelled/unlabelled NfL ratio of 0.35% observed after a 120-day chase period. In vitro analysis determined the half-life of intracellular NfL in control iPSC-neurons to be between 8.7 ± 0.8 days and 13.2 ± 0.9 days. In conclusion, this work has established the first targeted method for NfL using immunoprecipitation and triple quadrupole mass spectrometry, and preliminary data indicates, for the first time, turnover of NfL in vivo is exceptionally slow.
| Type: | Thesis (Doctoral) |
|---|---|
| Qualification: | Ph.D |
| Title: | Developing translational kinetic biomarkers for neurodegenerative disease |
| Open access status: | An open access version is available from UCL Discovery |
| Language: | English |
| Additional information: | Copyright © The Author 2023. Original content in this thesis is licensed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International (CC BY-NC 4.0) Licence (https://creativecommons.org/licenses/by-nc/4.0/). Any third-party copyright material present remains the property of its respective owner(s) and is licensed under its existing terms. Access may initially be restricted at the author’s request. |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Brain Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Brain Sciences > UCL Queen Square Institute of Neurology UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Brain Sciences > UCL Queen Square Institute of Neurology > Neurodegenerative Diseases |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10174661 |
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