The aim of this thesis was to analyze biosamples of seronegative patients suspicious of autoimmune encephalitis (AE) who were screened positively in a brain protein-reactive immunoblot by focusing on detecting potential so far unidentified brain protein-specific autoantibodies (autoABs). In the first step, we optimized the method for identifying brain protein-reactive autoABs by improving critical steps, including an immunoblot confirmation step to verify additional bands. Next, we analyzed biosamples from 387 patients who presented to the University Hospital Bonn, Germany, between 2017 and 2021 with clinical symptoms of AE but without known autoABs to identify potential brain protein-reactive autoABs in serum or cerebrospinal fluid. Mass spectrometry identified three subunits of the intramitochondrial pyruvate dehydrogenase complex (PDHc) as target proteins in serum samples of three different index patients, including pyruvate dehydrogenase E1 component subunit α (E1α), dihydrolipoyllysine-residue acetyltransferase (E2), and dihydrolipoamide dehydrogenase (E3). Subsequently, we identified a total of 12 patients positive for anti-PDHc autoABs through immunoblotting in our cohort, with E2 being the major autoABs target. In a third step we conducted an initial characterization of the identified autoABs and their targets, by using immunohistochemistry on murine brain slices and immunocytochemistry on primary hippocampal neurons. Localization studies revealed that the PDHc is widely distributed in the cerebral cortex, cerebellum, and hippocampus, as well as at dendritic spines and soma of mainly inhibitory but also excitatory neurons. Anti-PDHc autoABs are present in previously seronegative patients with suspected AE and may play a role in this disease spectrum. However, the potential pathological relevance of the autoABs themselves, associated mitochondrial dysfunction, the involved immunologic reactions, and an in-depth clinical implication regarding disease onset, course, and prognosis require further investigation.
Chiara Adina Hummel (Fri,) studied this question.