qHTS screen for small molecules that inhibit ELG1-dependent DNA repair in human embryonic kidney (HEK293T) cells expressing luciferase-tagged ELG1
Cancer cells divide rapidly compared to normal cells in the body. During cell division, cancer cells need to duplicate their genome by DNA replication. The failure of genome duplication leads to cancer cells' death. Therefore, inhibitors of DNA replication could specifically kill cancer cells. Based on this concept, many chemotherapeutic agents were developed and have been used to treat cancer more ..
BioActive Compounds: 7652
Depositor Specified Assays
NIH Molecular Libraries Probe Production Network [MLPCN]
NIH Chemical Genomics Center [NCGC]
MLPCN Grant: 1 R03 MH092164-01
Assay Provider: Kyungjae Myung, NHGRI, NIH
Keywords: MLPCN, qHTS, NCGC, genotoxicity, cancer
Cancer cells divide rapidly compared to normal cells in the body. During cell division, cancer cells need to duplicate their genome by DNA replication. The failure of genome duplication leads to cancer cells' death. Therefore, inhibitors of DNA replication could specifically kill cancer cells. Based on this concept, many chemotherapeutic agents were developed and have been used to treat cancer patients. However, there are a limited number of agents available in the clinical setting, and some of those that do exist have low efficacy and severe side effects. Therefore, if we can find the compounds that effectively block DNA replication either by directly damaging DNA or by inhibiting other cellular mechanisms, the options for cancer treatment will be broadened. With this in mind, we developed a quantitative high-throughput ELG1-luciferase reporter gene assay. Small molecules that specifically block ELG1 function in response to DNA damage can be identified by monitoring the suppression of luciferase activity. Since ELG1 defines a pathway for repairing DNA damage, agents blocking ELG1 function can be used to sensitize cancer cells to chemotherapeutic agents whose mechanism of action involves the induction of DNA damage.
Assay Protocol Summary:
For genotoxicity screening, 2,000 cells in 4 uL/well were dispensed into white, solid-bottom 1536-well plates using a solenoid-based dispenser. Following transfer of 1 uL 5-Fluorouridine and 23 nL compound or DMSO vehicle by a pin tool, the plates were incubated 16 hr at 37oC and 5% CO2. 5 ul ONE-Glo (Promega) was then added to each plate and following a 30 min incubation at room temperature, luciferase activity was quantified on a ViewLux CCD-based plate reader.
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 inhibitors are ranked higher than compounds that showed apparent activation.
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.
* Activity Concentration. ** Test Concentration.
Data Table (Concise)