= Emerging. More than 5 years before clinical availability. (26.55%)
= Expected to be clinically available in 1 to 4 years. (39.66%)
= Clinically available now. (33.79%)
MSACL 2020 US : Heller

MSACL 2020 US Abstract

Topic: Proteomics

Podium Presentation in Room 5 on Wednesday at 16:35 (Chair: Tim Collier)

Measurement of CSF Hypothalamic Peptides in Frontotemporal Dementia Using LC-MS/MS

Carolin Heller (Presenter)
University College London (UCL)

Presenter Bio(s): Carolin received her first degree as BSc (Hons) in Biomedical Science from the University of East London (UK) in 2015, during her final year research project she used a mass spectrometric approach to detect lipid changes within the bacterial membrane exploring treatment effects of various antibiotics. She entered the world of neuroscience working as a Research Assistant at the Institute of Neurology at University College London (UK), investigating rare dementias, in particular frontotemporal dementia (FTD). She started her part-time PhD in the Translational Mass Spectrometry Research Group at the Institute of Child Health, UCL (UK) in February 2018. Carolin is interested in investigating novel fluid biomarkers of neurodegeneration, which are paramount to monitoring treatment response in future clinical trials.

Authors: Heller C, Heywood W, Theodoridi A, Woollacott IOC, Mills K, Rohrer JD
University College London, London, UK


Introduction: Frontotemporal dementia (FTD) is a progressive, neurodegenerative disorder with clinical heterogeneity. The main clinical FTD phenotypes are behavioural variant FTD (bvFTD), semantic dementia (SD) and progressive non-fluent aphasia (PNFA). One key clinical characteristic of bvFTD is disturbance in eating behaviour, which can be helpful in diagnosing bvFTD and differentiating it from Alzheimer’s disease (AD). The aim of this study was to optimise a targeted multiple reaction monitoring (MRM) assay using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) investigating hypothalamic feeding peptides in FTD.

Method: A targeted multiple reaction monitoring (MRM) LC-MS/MS method was set-up to screen for a variety of hypothalamic peptides, including 12 centrally and 10 peripherally produced peptides. Peptide MRM assay was first optimised using Skyline, screening for multiple precursor and product ions in a tryptic digest of synthetic peptides. As the overall aim of the project was to identify endogenous peptide levels in cerebrospinal fluid (CSF), multiple sample processing methods were investigated to test their efficiency in extracting peptides: ultrafiltration, solid phase extraction (SPE) and protein precipitation.

Results: SPE yielded most detectable peptides compared to both protein precipitation and ultrafiltration. Various volumes of CSF were analysed using the optimised MRM method following peptide extraction optimisation, which enabled the detection of multiple endogenous peptides. We were able to detect endogenous peptides in a volume of CSF as little as 300 µL, which shows that LC-MS/MS is a sensitive and specific tool enabling us to detect neuropeptides. Standard calibration curves were created for each peptide and goodness of fit (r2) varied from 0.88 to 0.99 (p <0.001).

In our pilot study, we found a significant difference (p < 0.05) in five of the hypothalamic peptides (expressed as pmol/µl): neuropeptide W levels were significantly higher in all three groups: bvFTD (0.029), SD (0.034) and PNFA (0.039), controls (0.012); cerebellin was decreased in bvFTD (0.586) and SD (0.591) [controls 0.945], and cocaine-amphetamine regulated transcript was decreased in PNFA (0.359) and SD (0.359) [controls 0.575]; corticotropin-releasing hormone was decreased in bvFTD (0.007) [controls 0.011] and galanin was increased in PNFA (0.107) [controls 0.059].

Conclusions: A hypothalamic peptide panel has been set-up using a targeted MRM method with a 10 minute LC-MS/MS run duration that can be used to screen human CSF samples. This pilot study shows changes in concentration of a substantial proportion of hypothalamic peptides within the CSF in FTD groups compared to controls. Further exploration on a larger clinically defined cohort will enable understanding of the differences in hypothalamic peptides in FTD and investigate whether such a panel could be used as a biomarker in disease diagnosis, prognosis or stratification.

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