Targeted degradation of ⍺-synuclein aggregates in Parkinson's disease using the AUTOTAC technology.

BACKGROUND: There are currently no disease-modifying therapeutics for Parkinson's disease (PD). Although extensive efforts were undertaken to develop therapeutic approaches to delay the symptoms of PD, untreated α-synuclein (α-syn) aggregates cause cellular toxicity and stimulate further disease progression. PROTAC (Proteolysis-Targeting Chimera) has drawn attention as a therapeutic modality to target α-syn. However, no PROTACs have yet shown to selectively degrade α-syn aggregates mainly owing to the limited capacity of the proteasome to degrade aggregates, necessitating the development of novel approaches to fundamentally eliminate α-syn aggregates. METHODS: We employed AUTOTAC (Autophagy-Targeting Chimera), a macroautophagy-based targeted protein degradation (TPD) platform developed in our earlier studies. A series of AUTOTAC chemicals was synthesized as chimeras that bind both α-syn aggregates and p62/SQSTM1/Sequestosome-1, an autophagic receptor. The efficacy of Autotacs was evaluated to target α-syn aggregates to phagophores and subsequently lysosomes for hydrolysis via p62-dependent macroautophagy. The target engagement was monitored by oligomerization and localization of p62 and autophagic markers. The therapeutic efficacy to rescue PD symptoms was characterized in cultured cells and mice. The PK/PD (pharmacokinetics/pharmacodynamics) profiles were investigated to develop an oral drug for PD. RESULTS: ATC161 induced selective degradation of α-syn aggregates at DC50 of ~ 100 nM. No apparent degradation was observed with monomeric α-syn. ATC161 mediated the targeting of α-syn aggregates to p62 by binding the ZZ domain and accelerating p62 self-polymerization. These p62-cargo complexes were delivered to autophagic membranes for lysosomal degradation. In PD cellular models, ATC161 exhibited therapeutic efficacy to reduce cell-to-cell transmission of α-syn and to rescue cells from the damages in DNA and mitochondria. In PD mice established by injecting α-syn preformed fibrils (PFFs) into brain striata via stereotaxic surgery, oral administration of ATC161 at 10 mg/kg induced the degradation of α-syn aggregates and reduced their propagation. ATC161 also mitigated the associated glial inflammatory response and improved muscle strength and locomotive activity. CONCLUSION: AUTOTAC provides a platform to develop drugs for PD. ATC161, an oral drug with excellent PK/PD profiles, induces selective degradation of α-syn aggregates in vitro and in vivo. We suggest that ATC161 is a disease-modifying drug that degrades the pathogenic cause of PD.

TNF-α increases the intrinsic excitability of cerebellar Purkinje cells through elevating glutamate release in Bergmann Glia.

For decades, the glial function has been highlighted not only as the 'structural glue', but also as an 'active participant' in neural circuits. Here, we suggest that tumor necrosis factor α (TNF-α), a key inflammatory cytokine, alters the neural activity of the cerebellar Purkinje cells (PCs) by facilitating gliotransmission in the juvenile male rat cerebellum. A bath application of TNF-α (100 ng/ml) in acute cerebellar slices elevates spiking activity of PCs with no alterations in the regularity of PC firings. Interestingly, the effect of TNF-α on the intrinsic excitability of PCs was abolished under a condition in which the type1 TNF receptor (TNFR1) in Bergmann glia (BG) was genetically suppressed by viral delivery of an adeno-associated virus (AAV) containing TNFR1-shRNA. In addition, we measured the concentration of glutamate derived from dissociated cerebellar cortical astrocyte cultures treated with TNF-α and observed a progressive increase of glutamate in a time-dependent manner. We hypothesised that TNF-α-induced elevation of glutamate from BGs enveloping the synaptic cleft may directly activate metabotropic glutamate receptor1 (mGluR1). Pharmacological inhibition of mGluR1, indeed, prevented the TNF-α-mediated elevation of the intrinsic excitability in PCs. Taken together, our study reveals that TNF-α triggers glutamate release in BG, thereby increasing the intrinsic excitability of cerebellar PCs in a mGluR1-dependent manner.

Glia and gliotransmitters on carbon nanotubes.

Introduction: Functionalised carbon nanotubes (CNTs) have been shown to be promising biomaterials in neural systems, such as CNT -based nerve scaffolds to drive nerve regeneration. CNTs have been shown to modulate neuronal growth and improve electrical conductivity of neurons. Methods: Cultured astrocytes on the functionalized CNTs (PEG, caroboxyl group) were assessed for distribution of GABA, glutamate uptake assay using isotope and change of conductance of CNTs by ATP. Immunostaining of GABA using anti-GABA (red), anti-GFAP (green) antibody in primary cortical astrocytes on MW-CNT and PDL coverslips. Results: The functionalization of CNTs has improved their solubility and biocompatibility and alters their cellular interaction pathways. Recently, CNTs have been shown to modulate morphofunctional characteristics of glia as well as neurons. Among the various types of glia, astrocytes express diverse receptors for corresponding neurotransmitters and release gliotransmitters, including glutamate, adenosine triphosphate, and γ-amino butyric acid. Gliotransmitters are primarily released from astrocytes and play important roles in glia-neuron crosstalk. Conclusion: This review focuses on the effects of CNTs on glial cells and discusses how functionalized CNTs can modulate morphology and gliotransmitters of glial cells. Based on exciting new findings, they look to be a promising material for use in brain disease therapy or neuroprosthetics.