Biochemische Bestimmung und Charakterisierung des Bindungsverhaltens eines physiologisch vorkommenden Autoantikörpers gegen humanes α-Synuclein

Introduction: PD is one of most common neurodegenerative disorders with a prevalence of 0.3% in the United Stated. Age is the most common risk factor and responsible for the observed increase in prevalence with increasing age. Pathophysiologically, the disease is associated with the deposition of α-synuclein throughout the brain. This spread is associated with progressive neuronal loss, mainly in the substantia nigra. The role of αS in the development of this disorder is strengthened by the fact that αS mutations lead to genetic PD. In addition, αS is the main content of Lewy-bodies, which are characteristic of this disease. Recent work has revealed that naturally occurring antibodies exist against αS. We further elucidated their binding partners as well as their ability to detect αS in immunhistochemistry. Material and Methods: Antibodies against αS were isolated from healthy human donors or from IVIG preparations. The establishment of a protocol to isolate these antibodies was part of this work. The antibodies were then used to detect αS in the neural cell line SH-SY5Y, transgenic animal brain slices (Thy1)-h[A30P] as well as in human brain slices. These were obtained from BrainNet Europe. Next, interaction studies followed to determine the binding epitope. Using immunoblot technique, possible binding partners of these nAbs were identified. We used different synuclein proteins such as αS, βS und γS. Further, we used several point mutations as well as shortened peptides. Finally, we compared nAbs-αS to nAbs against Aβ. Results: This work demonstrates the existence of naturally occurring antibodies against αS. Immunhistochemical analyses of these antibodies reveal binding of the antibody to αS in the human neuroblastoma cell line SH-SY5Y, in transgenic animal models as well as in post mortem brains of PD patients. nAbs-αS strongly bind αS and can be used for reliable detection of αS in all these settings. Further analysis reveals that nAbs-αS mainly detect low molecular oligomers, that are currently discussed as main culprits in many neurodegenerative disorders. In addition, our analyses reveal that nAbs-αS mainly detect a protein conformation rather than a linear epitope. Similar observations can be made for other nAbs [Bach et al. 2017]. Discussion: The presented data reveal the existence of naturally occurring antibodies against αS. Similar to AD, these antibodies may play a role in disease progression and may also serve as a therapeutic instrument. Data by Besong-Agbo et al. demonstrate reduced nAbs-αS in patients suffering from PD [Besong-Agbo et al. 2013]. Since nAbs-αS are part of IVIG, these may also constitute a possible therapy for α-syncleinopathies. However, in AD, many trials so far were not convincing. In order to investigate a possible therapeutic role of nAbs-αS, these need further be tested in animal models of the disease. If these are convincing, then therapeutic approaches in humans may be conducted. Nevertheless, several lessons were learned from similar approaches in AD. Therapy needs to start early, preferentially in prodromal disease. For PD, this may be easy to pursue since REM-sleep movement behaviour disorder is regarded as a prodromal disorder of α-synucleinopathies. Patients at risk can thus be easily identified. In addition, the concentration of IVIG in nAbs-αS may be insufficient. Therefore, B-cells from healthy aged human donors should be isolated and nAbs-αS isolated on a single cell base. These then need to be tested for their binding capabilities and the most suited antibody then should be genetically engineered. This approach was pursued for Aducanumab therapy in AD and Cinpanemab in PD, currently the most convincing candidates.