DEVELOPMENT OF A DRUG SENSITIVITY ASSAY IN MURINE CELL MODEL FOR CLN3 DISEASE

Neuronal Ceroid Lipofuscinosis (NCL) is a group of neurogenerative disorders that causes visual failure, dementia, behavioral changes, motor difficulties, seizures, and premature death. There are 13 genetic subtypes of this disease, and the most common is CLN3 disease. CLN3 is inherited in an autosomal recessive pattern and is caused by mutations in the CLN3 gene, which provides instructions for making the protein, CLN3. Various phenotypes have been identified in CLN3 disease, such as lysosomal phenotypes, mitochondrial morphology, and autophagy, which show abnormal autophagosome accumulation in mutant Cln3 GFP-LC3 cells. Research has shown that when thapsigargin, a SERCA (sarcoendoplasmic reticulum calcium transport ATPase) inhibitor, is added to both wild-type and mutant cells, there is a significant increase of GFP-LC3 expression in mutant cells as compared to wild-type cells. Also, preliminary evidence expressed that this may be because of Ca2+ influx, ER stress, autophagy inhibition, or SERCA reacting with the CLN3 in some way. The goal of this study was to obtain more knowledge about the role of thapsigargin and to test possible therapeutic compounds that could be used to decrease or revert thapsigargin sensitivity. CDN1163, a SERCA activator, and verapamil, a Ca2+ channel blocker previously known to rescue lysosomal phenotypes and GFP-LC3 accumulation, were tested in various concentrations and times of exposure to thapsigargin. A drug sensitivity assay was successfully created, in which verapamil (10µM), showed a dose-response in preventing thapsigargin sensitivity. Also, CDN1163 (0.1µM) also was able to prevent thapsigargin sensitivity. Future outlooks involve utilizing this assay to test certain drugs for therapeutic benefits.   

For more information: mewoodbu@eckerd.edu

Accessibility | Non-Discrimination | Privacy | Report It