*BBSRC EASTBIO Programme* The Role and Utility of Exosomes as a Biomarker of N-Methyl-D-Aspartate Receptor Encephalitis

Introduction

IgG autoantibodies to N-methyl-D-aspartate (NMDA) receptor GluN1/2 subunits stimulate an autoimmune neuroinflammatory state in NMDA receptor encephalitis (NMDARe)1. Patients present insidiously, with changes in behaviour, but subsequently develop psychotic features and movement disorders. Hypoventilation and coma can necessitate critical care support. The first case diagnosed at Ninewells Hospital was in 2011, in a patient with ovarian teratoma (teratomas are present in around 60% of cases).

Exosomes, vesicles shed by cells, contain nucleic acids and proteins from the host cell2. Increasing evidence implies roles in immune modulation and as biomarkers of disease.

Following our preliminary data, this study aims to explore the potential of exosomes as biomarkers of NMDARe.

Previous work

Serum samples from a patient at Ninewells Hospital with NMDARe were examined to assess the concentration and size of nanoparticles by nanosight tracking analysis. Exosomes were then extracted from the serum by immunoisolation with anti-CD63 beads. CD63 is a member of the tetraspanin protein family, known to be enriched in exosomes populations.

The nanoparticle concentration was reduced in samples which were autoantibody positive (from the time of peak symptoms), then increased to pre-disease levels when autoantibody negative (after successful treatment). In contrast, the CD63 positive subpopulation was found to increase in samples which were autoantibody positive, then reduce with treatment. This finding, novel for NMDARe, is consistent with exosome population profiles from work on other autoimmune diseases3.

A changing exosome population profile in NMDARe raises the possibility of utilising serum exosomes as a biomarker of disease activity. The presence of NMDA receptor on these exosomes could allow them to be disease specific.

Currently all UK samples of suspected NMDA receptor encephalitis are sent to Professor Vincent’s laboratory (https://www.ndcn.ox.ac.uk/team/angela-vincent), University of Oxford, for diagnosis. An existing collaboration provides access to an extensive collection of clinical material.

PhD project

We will test the hypothesis that exosomes can be used as a biomarker of NMDARe and whether different aetiologies or syndromes of NMDARe (e.g. teratoma-associated) are associated with distinct exosomal markers.

  1. Nanoparticle population: size and number
    Nanosight tracking analysis will be used to fully assess whether serum nanoparticle (sub)population changes correlate with antibody load and disease severity.
  2. Exosome identification
    Exosome appearance and protein markers will be confirmed by electron microscopy and western blotting.
  3. Protein content
    Exosome protein content will be analysed using quantitative SWATH mass spectrometry, to determine the exact content at each stage of disease.
  4. Nucleic acid content
    Currently, microRNAs are a popular target for potential biomarkers, and the nucleic acid content of these exosomes will be assessed for this purpose.
  5. Assessing the functional status of exosome surface proteins
    The functional role of exosome-surface NMDA receptors will be assessed using bilayer and patch-clamp electrophysiology, taking advantage of recent techniques for fusing exosomes with liposomes.

Aims

  1. Evaluate the potential to use exosomes as a biomarker of disease activity in NMDARe, by characterising the (sub)population profiles and protein/nucleic acid content.
  2. Test whether exosomes associated within NMDARe in individuals with teratomas differ in their characteristics from those associated with NMDARe in the absence of a tumour.
  3. Identify whether exosomes contain functional NMDA receptors on their surface.

APPLY

Supervisor: 
Prof T Hales, Dr S Powis & Dr J Bowness
School: 
Start Date: 
Monday, September 4, 2017
Application Closing Date: 
Sunday, April 30, 2017