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High throughput screening and cell-based translocation evaluation for small molecular therapy of Pompe disease

Tuesday, October 09, 2012 — Poster Session I

1:00 p.m. – 3:00 p.m

Natcher Conference Center, Building 45




  • RJ Tamargo
  • W Westbroek
  • E Goldin
  • J Xiao
  • JJ Marugan
  • E Sidransky


Pompe disease is a lysosomal storage disorder caused by deficiency of the enzyme acid-alpha-glucosidase (GAA); accumulation of glycogen occurs primarily in lysosomes of cardiac and skeletal muscle. Symptoms include progressive muscle weakness and loss of motor, respiratory, and cardiac function. Currently, Pompe is treated by enzyme replacement therapy, which is expensive, inconvenient, and does not reverse all symptoms. Many disease-causing mutations retain enzyme activity, but enzyme is not translocated to the lysosome. Small-molecule chaperone treatment could improve folding and translocation of mutant enzyme. High throughput screening (HTS) of small molecule libraries using recombinant enzyme has identified chaperones, but not activators. We developed a novel HTS method utilizing tissue homogenate as the enzyme source, creating a more native physiological environment. Utilizing wildtype spleen homogenate to screen our library of 250,000 compounds, we identified novel inhibitor and activator series for GAA, shown to enhance enzymatic activity. A cell-based translocation assay on wildtype and Pompe fibroblasts showed that treatment with the lead compounds increased translocation of GAA to the lysosomes. Currently, translocation assays in muscle cells from Pompe mice are being developed. Our cell-based studies demonstrate that tissue homogenate can be used for screening, leading to the identification of physiologically-relevant compounds.

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