THE EFFECT OF GENETIC VARIATION ON BRANAPLAM SPLICE-MODULATION

Huntington’s Disease (HD) is a neurodegenerative disorder that causes choreatic movements, behavioral and psychiatric disturbances, and dementia. HD is caused by an expansion of repeating CAG nucleotides in the Huntingtin Gene (HTT) beyond 35 repeat units. Therapeutic approaches for HD have focused on lowering HTT mRNA or protein levels to reduce the toxicity from the expanded repeat. Branaplam is a small brain-penetrant molecule that induces the inclusion of a pseudoexon between exons 49 and 50 of HTT. This inclusion lowers HTT gene expression by introducing an early stop codon. Previously, we showed that Branaplam was less effective in lowering HTT in certain cell lines, likely due to genetic variation close to the pseudoexon. This project focused on two other genetic targets of Branaplam, TENT2 and ZFP82, that were predicted by deep learning tools to be affected by variants proximal to their pseudoexons. Cells with predicted variants were dosed with 100 nM Branaplam, reverse transcribed into cDNA, amplified using PCR, visualized using gel electrophoresis, and the densitometry analyzed through ImageJ. We found that Branaplam splice modulation was affected in cell lines carrying the predicted variants. These data highlight the role of genetic variation in causing unintended off-target effects in targeted genetic therapies. 

For more information:  infarnum@eckerd.edu