Absence of anti-hypocretin receptor 2 autoantibodies in post pandemrix narcolepsy cases

Guo Luo, Ling Lin, Louis Jacob, Mélodie Bonvalet, Aditya Ambati, Giuseppe Plazzi, Fabio Pizza, Ryan Leib, Christopher M. Adams, Markku Partinen, Emmanuel Jean Marie Mignot

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Abstract

Background: A recent publication suggested molecular mimicry of a nucleoprotein (NP) sequence from A/Puerto Rico/8/1934 (PR8) strain, the backbone used in the construction of the reassortant strain X-179A that was used in Pandemrix® vaccine, and reported on anti-hypocretin (HCRT) receptor 2 (anti-HCRTR2) autoantibodies in narcolepsy, mostly in post Pandemrix® narcolepsy cases (17 of 20 sera). In this study, we re-examined this hypothesis through mass spectrometry (MS) characterization of Pandemrix®, and two other pandemic H1N1 (pH1N1)-2009 vaccines, Arepanrix® and Focetria®, and analyzed anti-HCRTR2 autoantibodies in narcolepsy patients and controls using three independent strategies. Methods: MS characterization of Pandemrix® (2 batches), Arepanrix® (4 batches) and Focetria® (1 batch) was conducted with mapping of NP 116I or 116M spectrogram. Two sets of narcolepsy cases and controls were used: 40 post Pandemrix® narcolepsy (PP-N) cases and 18 age-matched post Pandemrix® controls (PP-C), and 48 recent (6 months) early onset narcolepsy (EO-N) cases and 70 age-matched other controls (O-C). Anti-HCRTR2 autoantibodies were detected using three strategies: (1) Human embryonic kidney (HEK) 293T cells with transient expression of HCRTR2 were stained with human sera and then analyzed by flow cytometer; (2) In vitro translation of [35S]-radiolabelled HCRTR2 was incubated with human sera and immune complexes of autoantibody and [35S]-radiolabelled HCRTR2 were quantified using a radioligand-binding assay; (3) Optical density (OD) at 450 nm (OD450) of human serum immunoglobulin G (IgG) binding to HCRTR2 stably expressed in Chinese hamster ovary (CHO)-K1 cell line was measured using an in-cell enzyme-linked immunosor-bent assay (ELISA). Results: NP 116M mutations were predominantly present in all batches of Pandemrix®, Arepanrix® and Focetria®. The wild-type NP109-123 (ILYDKEEIRRIWRQA), a mimic to HCRTR234-45 (YDDEEFLRYLWR), was not found to bind to DQ0602. Three or four subjects were found positive for anti-HCRTR2 autoantibodies using two strategies or the third one, respectively. None of the post Pandemrix® narcolepsy cases (0 of 40 sera) was found positive with all three strategies. Conclusion: Anti-HCRTR2 autoantibody is not a significant biological feature of narcolepsy or of post Pandemrix® autoimmune responses.

Original languageEnglish
Article numbere0187305
JournalPLoS One
Volume12
Issue number12
DOIs
Publication statusPublished - Dec 1 2017

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)

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    Luo, G., Lin, L., Jacob, L., Bonvalet, M., Ambati, A., Plazzi, G., Pizza, F., Leib, R., Adams, C. M., Partinen, M., & Mignot, E. J. M. (2017). Absence of anti-hypocretin receptor 2 autoantibodies in post pandemrix narcolepsy cases. PLoS One, 12(12), [e0187305]. https://doi.org/10.1371/journal.pone.0187305