Abstract
Human carboxylesterase 2 (CES2), one of the most abundant hydrolases distributed in the small intestine, has been validated as a key therapeutic target to ameliorate the intestinal toxicity caused by irinotecan. This study aims to discover efficacious CES2 inhibitors from natural products and to characterize the inhibition potentials and inhibitory mechanisms of the newly identified CES2 inhibitors. Following high-throughput screening and evaluation of the inhibition potency of more than 100 natural products against CES2, it was found that the biflavones isolated from Ginkgo biloba displayed extremely potent CES2 inhibition activities and high specificity over CES1 (>1000-fold). Further investigation showed that ginkgetin, bilobetin, sciadopitysin and isoginkgetin potently inhibited CES2-catalyzed hydrolysis of various substrates, including the CES2 substrate-drug irinotecan. Notably, the inhibition potentials of four biflavones against CES2 were more potent than that of loperamide, a marketed anti-diarrhea agent used for alleviating irinotecan-induced intestinal toxicity. Inhibition kinetic analyses demonstrated that ginkgetin, bilobetin, sciadopitysin and isoginkgetin potently inhibited CES2-catalyzed fluorescein diacetate hydrolysis via a reversible and mixed inhibition manner, with K i values of less than 100 nM. Ensemble docking and molecular dynamics revealed that these biflavones could tightly and stably bind on the catalytic cavity of CES2 via hydrogen bonding and π-π stacking interactions, while the interactions with CES1 were awfully poor. Collectively, this study reports that the biflavones isolated from Ginkgo biloba are potent and highly specific CES2 inhibitors, which offers several promising lead compounds for developing novel anti-diarrhea agent to alleviate irinotecan-induced diarrhea.
Highlights
Mammalian carboxylesterases (CES) are key members of serine hydrolases which play key roles in activation of ester prodrugs and hydrolytic metabolism of a variety of xenobiotics bearing ester bond(s) (Hosokawa, 2008; Hatfield et al, 2016)
It should be noted that the inhibition potency of these four biflavones were much more potent than that of some reported naturally occurring carboxylesterase 2 (CES2) inhibitors and the positive inhibitor LPA (IC50 6240 nM, Supplementary Table S1) (Song et al, 2019a; Song et al, 2019b)
This study reported that four biflavones isolated from Ginkgo biloba displayed potent CES2 inhibition potency and high specificity over carboxylesterase 1 (CES1) (>1000-fold)
Summary
Mammalian carboxylesterases (CES) are key members of serine hydrolases which play key roles in activation of ester prodrugs and hydrolytic metabolism of a variety of xenobiotics bearing ester bond(s) (Hosokawa, 2008; Hatfield et al, 2016). It has been reported that CES2 participated in approximately 99% of the total conversion of irinotecan into SN-38 in the human small intestine (Humerickhouse et al, 2000; Charasson et al, 2004). Increasing evidence has indicated that co-administration with potent CES2 inhibitors may ameliorate irinotecan-induced severe diarrhea via blocking the overproduction of SN38 (the cytotoxic product of CPT-11) in human small intestine (Slatter et al, 1997; Hecht, 1998; Hicks et al, 2009). Loperamide (LPA, a marketed anti-diarrhea agent) has been used for alleviating irinotecan-induced intestinal toxicity in clinical settings (Tobin et al, 2005; Baker, 2007). It is urgent and necessary to find more efficacious CES2 inhibitors to reduce the excessive hydrolysis of CPT-11 into its toxic product SN-38 in the intestinal tract
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