Submitted on August 26, 2005
Revised on November 8, 2005
Accepted on November 15, 2005
Automated microscopy screening for compounds that partially revert cholesterol accumulation in Niemann-pick C cells
Nina H. Pipalia, Amy Y. Huang, Harold Ralph, Madalina Rujoi, and Frederick R. Maxfield
Biochemistry Dept., Weill Medical College of Cornell University, New York, NY 10021
Corresponding Author: frmaxfie{at}med.cornell.edu
Niemann-Pick type C disease is an autosomal recessive genetic disorder manifested by abnormal accumulation of unesterified cholesterol and other lipids. We screened combinatorially synthesized chemical libraries to identify compounds that would partially revert cholesterol accumulation. Cultured CHO cells with Niemann-Pick type C phenotypes (CT60 and CT43) were used for screening along with normal CHO cells as a control. We developed an automated microscopy assay based on imaging of filipin fluorescence for estimating cholesterol accumulation in lysosomal storage organelles. Our primary screen of 14,956 compounds identified 14 hit compounds that caused significant reduction in cellular cholesterol accumulation at 10 mM. We then screened a secondary library of 3,962 compounds selected based on chemical similarity to the initial hits and identified 7 compounds that demonstrated greater efficacy and lower toxicity than the original hits. These compounds are effective at concentrations of 123 nM to 3 mM in reducing the cholesterol accumulation in cells with a Niemann-Pick type C1 phenotype.
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