AID	Name	Type	Comment
485316	qHTS Assay for Rab9 Promoter Activators: Summary	summary	Rab9 Summary AID
485315	qHTS Assay for NPC1 Promoter Activators: Summary	summary	NPC1 Summary AID

Niemann Pick Type C (NPC) is a rare neurodegenerative lipidosis that is characterized by lipid storage in the endosomal/lysosomal system. Treatment modalities for this devastating disease are currently non-existent due to the severe obstacles associated with accessing the central nervous system with proteins or genes. The majority of mutations causing NPC disease are missense mutations that are distributed throughout the length of the NPC1 protein, with the most prevalent being the I1061T allele. Although NPC1 mutant proteins may be functional, they are typically trapped in the ER due to misfolding. Furthermore, over-expression of some mutant NPC1 proteins can rescue the disease phenotype, suggesting that up-regulation of the endogenous NPC1 mutant protein is a new drug treatment modality of the disorder.

We have developed and optimized a cell based renilla luciferase reporter assay in 1536 well format for the identification of up-regulators of the mutant Rab9 promoter. This assay presents data on the validation of hits from the primary screen, which used a firefly luciferase read-out.

NIH Chemical Genomics Center [NCGC]
NIH Molecular Libraries Probe Production centers Network [MLPCN]

MLPCN Grant: MH089375
Assay Submitter (PI): Yiannis A. Ioannou

Huh7 cells stably expressing NPC promoter-luciferase will be seeded at 1,500 cells/well in 5 uL in 1,536-well white opaque plates with 23 nL/well of compound in DMSO solution. The assay plates will be incubated at 37 deg C for 24 hours, after which 3 uL/well BriteLite plus luciferase detection reagent mixture (Perkin Elmer) will be added. Luminescence signal will be measured in a ViewLux plate reader (Perkin Elmer) after a 10 minute incubation at the room temperature.

Compound Ranking:

1. Compounds are first classified as having full titration curves, partial modulation, partial curve (weaker actives), single point activity (at highest concentration only), or inactive. See data field "Curve Description". For this assay, apparent activators are ranked higher than compounds that showed apparent inhibition.
2. For all inactive compounds, PUBCHEM_ACTIVITY_SCORE is 0. For all active compounds, a score range was given for each curve class type given above. Active compounds have PUBCHEM_ACTIVITY_SCORE between 40 and 100. Inconclusive compounds have PUBCHEM_ACTIVITY_SCORE between 1 and 39. Fit_LogAC50 was used for determining relative score and was scaled to each curve class' score range.

Grant Number: MH089375