Coelenterazine

Details:

Coelenterazine is widely distributed among marine organisms, producing bioluminescence calcium-dependent oxidation mediated by the photoprotein aequorin, with EC₅₀ of 257 nM. IC50 & Target: EC50: 257 nM. In Vitro: On addition of coelenterazine to the buffer, intact KB 3-1 Rluc cells show high bioluminescence, whereas intact KB 8-5-11 Rluc cells showe low bioluminescence. A specific Pgp-mediated reduction in steady-state content of coelenterazine existed in KB 8-5-11 Rluc cells compare to KB 3-1 Rluc cells. Bioluminescence signals from KB 3-1 Rluc cells, both in the absence and presence of GF120918 (300 nM), are readily detected with as little as 1 nM extracellular coelenterazine and increase in a concentration-dependent manner to 1 μM extracellular coelenterazine. In Pgp-expressing cells, there is evidence for a GF120918-reversible concentration-dependent saturation of bioluminescence with an EC50 of 257 nM coelenterazine. The plateau in bioluminescence signal observe in KB 8-5-11 Rluc cells implies hat the capacity of Pgp to limit delivery of coelenterazine to cytosolic Rluc is not exceeded within the concentration range test. In Vivo: We then attempt to examine Pgp transport activity and the pharmacodynamics of inhibitors in vivo with noninvasive bioluminescence imaging. Tumor xenografts of KB 3-1 and KB 8-5-11 cells (control tumors) and KB 3-1 Rluc and KB 8-5-11 Rluc cells are grown in nu/nu mice and image with an in vivo imaging system and a charge-coupled device camera. Before tail vein administration of coelenterazine (4 μg/g) and imaging, mice are either first gavaged with vehicle, or the following day, the same mice are gavaged with GF120918 (250 mg/kg), thus enabling each mouse to serve as its own control. At baseline, KB 3-1 Rluc tumors show a 4-fold higher bioluminescence signal in vivo than KB 8-5-11 Rluc tumors. Normalized to bioluminescence signals arising from KB 3-1 Rluc tumors, signals emitte from KB 8-5-11 Rluc tumors in vivo are consistently reduced by≈75% .