Designing L-RNA-Based                           Spiegelmers Against Ecto-GPR37: A Novel Approach for Parkinson’s Disease Biomarker Detection

Parkinson’s disease (PD) is a neurological condition that progressively deteriorates the nervous system over time. One potential biomarker associated with this disease is Ecto-GPR37, the extracellular portion of the GPR37 protein. To enable its detection, we propose the use of spiegelmers—synthetic mirror-image oligonucleotides composed of L-ribose instead of the naturally occurring D-ribose found in RNA and DNA—making them inherently resistant to nuclease degradation. The research aimed to design and evaluate L-RNA-based spiegelmers targeting ecto-GPR37 as stable molecular probes for the early detection of PD. We hypothesize that spiegelmers can bind to the predicted binding site of ecto-GPR37 and serve as molecular probes for its detection. Moreover, the conversion of nucleotides from the D-form to the L-form is expected to enhance their stability by preventing enzymatic degradation. To investigate this, we employed AlphaFold 3 to generate a high-confidence three-dimensional structure of the ecto-GPR37 protein. In parallel, RNA sequences were initially modeled using RNAfold and subsequently converted into 3D structures using SimRNA. These D-form RNA structures were then transformed into their L-form counterparts via an in-house script. Following this, the spiegelmers were docked onto both ecto-GPR37 and RNase H1 using the HDOCK software to assess their binding potential and nuclease resistance. The resulting docked complexes were further analyzed using the PLIP web server to characterize the molecular interactions involved. Our analysis revealed that RNA A1 exhibited strong binding affinity to the ecto-GPR37 protein and was therefore selected for downstream evaluation. Overall, the findings from this study support the design and application of nuclease-stable spiegelmers as a novel approach for the detection of PD.