Characterization of dementia with Lewy bodies: Identificacion of a specific bio-signature from blood for the differential diagnosis of dementia with Lewy bodies

Ana Gámez Valero

Dr Katrin Beyer Dr Francesc E Borràs

Dementia with Lewy bodies (DLB), one of the most common neurodegenerative diseases along with Alzheimer's disease (AD) and Parkinson's disease (PD), was described about 30 years ago, but even today its correct diagnosis remains a challenge. Due to the clinical, neuropathological and genetic characteristics that it shares with AD and PD, many cases of DLB ​​are still misdiagnosed. To improve this diagnosis, we set two objectives: On the one hand, to better characterize the genetic risk factors related to DLB, analyzing the genes involved in the pathology; and, in parallel, to define new non-invasive biomarkers obtained from peripheral blood for a better differential diagnosis of DLB ​​versus AD. To study the genetic risk factors related to MCI, the glucocerebrosidase (GBA) gene was examined for variants in MCI, MP, patients with idiopathic REM sleep behavior disorder (RSBD) and healthy controls. A higher rate of variants was observed in MCI -compared to control individuals-, which were especially associated with male sex and with a precipitation of the onset of the disease. The different genotypes of the main genes involved in the development of synucleinopathies, SNCA and SNCB, were also analyzed. Specifically, we observed that the SNCB gene contributes differently in the development of the pure form and/or the common form of MCI as well as in PD. On the other hand, to define non-invasive biomarkers in peripheral blood, extracellular vesicles (EVs) obtained from plasma and isolated by molecular exclusion chromatography were analyzed to determine their proteomic and transcriptomic content. Thus, we first identified the protein “gelsolin” as a possible biomarker, given its reduced expression in DCL compared to EA or healthy controls. In this objective, we also analyzed the content of microRNAs (miRNAs) in the VEs and, additionally, in the platelets obtained from the blood samples. In the case of the VEs, we identified 9 different miRNAs that were dysregulated in vesicles of EA compared to DCL and controls. In the case of the platelet content, several miRNAs were identified that were dysregulated in DCL compared to controls. Among all, the miRNA hsa-miR-150-5p should be highlighted, whose expression was significantly decreased in DCL compared to controls and EA. These results, which have been validated in different samples and experiments, included a final validation in which MP samples were also analyzed, which showed the same decreased expression of hsa-miR-150-5p as DCL. This could indicate the participation of this miRNA in the specific development of synucleinopathies. Finally, the diagnostic potential of the new possible biomarkers described was determined by calculating their values ​​in ROC curves, which showed high sensitivity and specificity to distinguish between DCL and AD patients. The results, protected by a patent, of this thesis are the first step towards a better and more precise diagnosis for DCL and synucleinopathies.

for the determination of platelet hsa-miR-150-5p levels, blood would be collected in tubes pre-treated with sodium citrate and processed within 2 hours following collection, avoiding coagulation and platelet activation. Platelets would be isolated by differential centrifugation, also at room temperature, (if needed, the pellet could be characterized by flow cytometry for the presence of CD61 platelet marker and CD45 leukocyte marker as contamination measurement) and miRNA isolation would be performed using a standard commercial kit followed by a standard qPCR assay determining if the patient suffers from AD or a synucleinopathy. For synucleinopathy determination (PD or DLB), SNCA and SNCB analysis, collection of blood in EDTA tubes followed by DNA extraction would be needed. We then propose the analysis by PCR and restriction fragment length polymorphism (RFLP) analysis with specific restriction enzymes for the genotype determination of SNCA rs356219. SNCB polyC sequence would be amplified using specific primers, and analysed using a standard 36-cycle PCR program

Dementia with Lewy bodies (DLB); Neurodegenerative Diseases; Alzheimer's Disease (AD); Parkinson's Disease (PD); Correct Diagnosis; Misdiagnosis; Genetic Risk Factors; Genes; Non-Invasive Biomarkers; Peripheral Blood; Differential Diagnosis; Glucocerebrosidase (GBA) gene; Variants; MCI; MP; Idiopathic REM Sleep Behavior Disorder (RSBD); Healthy Controls; Male Sex; Disease Onset Precipitation; Synucleinopathies; SNCA; SNCB; Pure Form of MCI; Common Form of MCI; Extracellular Vesicles (EVs); Plasma; Molecular Exclusion Chromatography; Proteomic Content; Transcriptomic Content; Gelsolin; Biomarker; Reduced Expression; MicroRNAs (miRNAs); Platelets; Dysregulated miRNAs; Hsa-miR-150-5p; Decreased Expression; Validation; Diagnostic Potential; ROC Curves; Sensitivity; Specificity; Patent Protection.