Persistent URL

http://hdl.handle.net/10950/5155

Document Type

Poster

Publication Date

11-2025

Abstract

Nicotinic acetylcholine receptors (nAChRs) play a crucial role in the pathogenesis of neurodegenerative diseases such as Parkinsonʼs disease (PD). In PD, degeneration of dopaminergic and cholinergic neurons impairs neurotransmitter release, leading to cognitive and motor dysfunction. Current treatments provide only symptomatic relief, highlighting the need for novel therapeutic strategies. The (α4)₃(β2)₂ subtype of nAChRs is expressed on affected neurons, making it a promising druggable target for enhancing neurotransmitter release and providing neuroprotection. To explore the therapeutic potential of positive allosteric modulators (PAMs) of (α4)₃(β2)₂ nAChRs, we evaluated chemical derivatives of CMPI [3-(2-chlorophenyl)-5-(5-methyl-1-(piperidin-4-yl)-1H-pyrazol-4-yl)isoxazole]. We used two-electrode voltage-clamp recordings from Xenopus laevis oocytes expressing nAChRs with defined subunit compositions to screen new derivatives for PAM activity and to construct doseresponse curves for lead compounds. We also evaluated the in vitro cytotoxicity of the CMPI derivatives using the MTT assay. Our results thus far establish that substitutions at the pyrazole ring of CMPI (e.g., to a thiazole in A12a or an imidazole in A12c and A12e) are tolerated within the α4-α4 binding pocket and maintain selectivity for (α4)₃(β2)₂ (EC₅₀ for potentiation < 1 µM) over (α4)₂(β2)₃ nAChRs. In contrast, modifications at the piperidine ring (e.g., to piperazine derivatives in GAT2227-GAT2230) are less tolerated, resulting in reduced or no PAM activity. Cell toxicity assays using HEK cells expressing α4β2 nAChRs showed no toxicity of the CMPI derivatives at concentrations that potentiate (α4)₃(β2)₂ nAChRs. Among these, A12c exhibited no effect on cell viability at concentrations up to 100 µM. The mode of binding for CMPI derivatives was investigated using computer-simulated docking and site-directed mutagenesis, which confirmed the extracellular. α4-α4 subunit interface as a potential binding site and identified key interactions between PAMs and amino acid residues. This study provides further evidence supporting the potential of PAMs acting at the α4-α4 interface as a strategy to selectively target the (α4)₃(β2)₂ nAChR, a major subtype in the cortex. CMPI derivatives are currently being investigated in a PD-like zebrafish model, which is expected to provide evidence of in vivo efficacy and support their therapeutic potential for the treatment of neurodegenerative diseases.

Journal/Conference Details

Presented at the 41st Southern Biomedical Engineering Conference, The University of Texas at Tyler.

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