New Cellular Models Of Gaucher Disease Exploiting CRISPR/Cas9 Technology

PAVAN, ELEONORA; ORMAZABAL, MAXIMILIANO E.; ROZENFELD, PAULA A.; DARDIS, ANDREA

Clermont-Ferrand

Congreso; 13th European Working Group on Gaucher Disease; 2019

European Working Group on Gaucher Disease

Mutations in the acid β-glucosidase (GBA) gene cause Gaucher Disease (GD). In addition, GBA mutations constitute the main genetic risk factor for developing Parkinson's disease (PD), a neurodegenerative disorder characterized by α-synuclein (α-syn) accumulation and aggregation. A lot of research has been focused in the development of cellular models for GD, in order to pursue a better understanding of the disease pathophysiology, to study the relationship between GD a PD and most importantly to develop novel therapeutic approaches. Therefore, our study was directed towards the development and characterization of new cellular models of GD, exploiting CRISPR/Cas9 technology to knock out (KO) the GBA gene in different continuous cell lines, including monocytes (THP-1) and glial (U87) cells. After editing, three THP-1 and three U87 clones were selected since they had altered β-glucosidase (GCase) protein expression. All selected clones displayed mutations in the GBA gene, reduced or absent GCase activity and accumulation of LysoGL1. Residual activity correlated with the levels of LysoGL1 storage. Then, to deepening the GD-PD link, we studied the expression of α-syn in the U87 GBA KO clones. All clones presented alterations in α-syn abundance/distribution. However, significant differences in the intracellular/secreted rate of α-syn were found between cells expressing mutated misfolded inactive GBA protein and those with a complete GBA depletion. In conclusion, we obtained a model of GD using 2 different cell types. This models present several advantages in comparisons with other models already available: being generated from commercial cell lines they provide more reproducible results than primary cultured cells; they grow faster in culture, are easy to transfect and are suitable for highthroughput screening experiments; finally, being isogenic cell lines carrying different GBA mutations they represent an ideal system to study mutation specific phenotypes.