BitterDB database analysis plus cell stiffness screening identify flufenamic acid as the most potent TAS2R14-based relaxant of airway smooth muscle cells for therapeutic bronchodilation

FFA as a potent TAS2R14 agonist to relax ASMCs while suppressing cytokine release might be a favorite drug agent for further development of TAS2R-based novel dual functional medication for bronchodilation and anti-inflammation in asthma therapy.

Bitter compounds in the BitterDB database were retrieved and analyzed for their working subtype of TAS2R and effective concentration. Compounds activating TAS2R5, 10, and 14 at < 100 μM effective concentration were identified and subsequently screened by cell stiffness assay using optical magnetic twisting cytometry (OMTC) to identify the most potent to relax ASMCs. Then the compound identified was further characterized for efficacy on various aspects related to relaxation of ASMCs, incl. but not limited to traction force by Fourier transform traction force microscopy (FTTFM), [Ca2+]i signaling by Fluo-4/AM intensity, cell migration by scratch wound healing, mRNA expression by qPCR, and protein expressing by ELISA. The compound identified was also compared to conventional β-agonist (isoproterenol and salbutamol) for efficacy in reducing cell stiffness of cultured ASMCs and airway resistance of ovalbumin-treated mice.

BitterDB analysis found 18 compounds activating TAS2R5, 10, and 14 at < 100 μM effective concentration. Cell stiffness screening of these compounds eventually identified flufenamic acid (FFA) as the most potent compound to rapidly reduce cell stiffness at 1 μM. The efficacy of FFA to relax ASMCs in vitro and abrogate airway resistance in vivo was equivalent to that of conventional β-agonists. The FFA-induced effect on ASMCs was mediated by TAS2R14 activation, endoplasmic reticulum Ca2+ release, and large-conductance Ca2+-activated K+ (BKCa) channel opening. FFA also attenuated lipopolysaccharide-induced inflammatory response in cultured ASMCs. Conclusions: FFA as a potent TAS2R14 agonist to relax ASMCs while suppressing cytokine release might be a favorite drug agent for further development of TAS2R-based novel dual functional medication for bronchodilation and anti-inflammation in asthma therapy.