Screening method for nonsteroidal antiinflammatory drugs based on the cyclooxygenase 2 pathway activated by serum-free stimulation in A549 cells.
Cyclooxygenase 2 (COX-2) pathway inhibitors were regarded as promising nonsteroidal antiinflammatory drugs (NSAIDs). We discovered that the COX-2 pathway in A549 cells, a human lung cancer cell line, was activated by serum-free stimulation, and a drug screening model for NSAIDs was established based on this principle with simple performance and sufficient reliability. The COX-2 pathway was activated by treating with serum-free medium for 12 h. The activated cells were incubated with NS398 (selective COX-2 inhibitor), SC560 (selective COX-1 inhibitor), acetyl salicylic acid (ASA) (nonselective COX inhibitor) at 37 degrees C for 15 min. Then the cells were incubated with 10 microM of arachidonic acid (AA) for another 30 min prostaglandin E2 and 6-keto-prostaglandin F(1alpha) were assayed in an enzyme immunoassay (EIA). The results showed that the COX-2 pathway was dominant in A549 cells whether activated by serum-free medium or not, and the COX-1 pathway could be ignored. The model accepted the positive inhibition threshold as NS398 2 microM; if a compound (10 microM) inhibited COX-2 pathway more than NS398 (2 microM), it was regarded as a hit. The COX-2 pathway inhibition experiment showed that the Z;-factor of the screening model was 0.62, which suggests that the model is suitable for COX-2 pathway inhibitor screening.
Yao
Jin Cheng
JC
National Center of New Drug Screening, China Pharmaceutical University, Nanjing, PR China.
Duan
Wei Gang
WG
Yun
Yu
Y
Liu
De Quan
DQ
Yan
Ming
M
Jiang
Zhen Zhou
ZZ
Zhang
Lu Yong
LY
D015121:6-Ketoprostaglandin F1 alpha / Q000032:analysis; D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology*; D001241:Aspirin / Q000494:pharmacology; D016895:Culture Media, Serum-Free; D052246:Cyclooxygenase 2 Inhibitors / Q000494:pharmacology*; D015232:Dinoprostone / Q000032:analysis; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D006801:Humans; D007124:Immunoenzyme Techniques; D009578:Nitrobenzenes / Q000494:pharmacology*; D011720:Pyrazoles / Q000494:pharmacology*; D013449:Sulfonamides / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D000894:Anti-Inflammatory Agents, Non-Steroidal; D016895:Culture Media, Serum-Free; D052246:Cyclooxygenase 2 Inhibitors; D009578:Nitrobenzenes; D011720:Pyrazoles; C115461:SC 560; D013449:Sulfonamides; C080955:N-(2-cyclohexyloxy-4-nitrophenyl)methanesulfonamide; D015121:6-Ketoprostaglandin F1 alpha; D015232:Dinoprostone; D001241:Aspirin