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Selective targeting of nuclear receptor FXR by avermectin analogues with therapeutic effects on nonalcoholic fatty liver disease.

Jin L, Wang R, Zhu Y, Zheng W, Han Y, Guo F, Ye FB, Li Y - Sci Rep (2015)

Bottom Line: Mechanistically, the avermectin analogues that interact with FXR exhibited features as partial agonists, with distinctive properties in modulating coregulator recruitment.Structural features critical for avermectin analogues to selectively bind to FXR were also revealed.Additionally, the structural features that discriminate the selective binding of FXR by avermectin analogues may provide a unique safe approach to design drugs targeting FXR signaling.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Cellular Stress Biology, Innovation Center for Cell Signaling Network, School of Life Sciences, Xiamen University, Fujian 361005, China.

ABSTRACT
Non-alcoholic fatty liver disease (NAFLD) has become a predictive factor of death from many diseases. Farnesoid X receptor (FXR) is an ideal target for NAFLD drug development due to its crucial roles in lipid metabolism. The aim of this work is to examine the molecular mechanisms and functional roles of FXR modulation by avermectin analogues in regulating metabolic syndromes like NAFLD. We found that among avermectin analogues studied, the analogues that can bind and activate FXR are effective in regulating metabolic parameters tested, including reducing hepatic lipid accumulation, lowering serum cholesterol and glucose levels, and improving insulin sensitivity, in a FXR dependent manner. Mechanistically, the avermectin analogues that interact with FXR exhibited features as partial agonists, with distinctive properties in modulating coregulator recruitment. Structural features critical for avermectin analogues to selectively bind to FXR were also revealed. This study indicated that in addition to antiparasitic activity, avermectin analogues are promising drug candidates to treat metabolism syndrome including NAFLD by directly targeting FXR. Additionally, the structural features that discriminate the selective binding of FXR by avermectin analogues may provide a unique safe approach to design drugs targeting FXR signaling.

No MeSH data available.


Related in: MedlinePlus

The potency of avermectin analogues in regulating metabolism correlates with their FXR binding capabilities.(a) Various coactivator motifs bind to FXR in response to 0.5 μM avermectin analogues or GW4064 by AlphaScreen assay. (b) 1 μM compounds induce FXR activity in reporter assays. (c) Dose responses of compounds in inducing FXR to recruit SRC1-2 coregulator binding motif by AlphaScreen assay. (d) Dose response of compounds in inducing the activity of FXR in reporter assays. (e) The recruitment of NCoR-2 to FXR in response to avermectin analogues by AlphaScreen assay. Values are the means ± SEM of three independent experiments.
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f4: The potency of avermectin analogues in regulating metabolism correlates with their FXR binding capabilities.(a) Various coactivator motifs bind to FXR in response to 0.5 μM avermectin analogues or GW4064 by AlphaScreen assay. (b) 1 μM compounds induce FXR activity in reporter assays. (c) Dose responses of compounds in inducing FXR to recruit SRC1-2 coregulator binding motif by AlphaScreen assay. (d) Dose response of compounds in inducing the activity of FXR in reporter assays. (e) The recruitment of NCoR-2 to FXR in response to avermectin analogues by AlphaScreen assay. Values are the means ± SEM of three independent experiments.

Mentions: To investigate the molecular mechanism of various avermectin analogues in differential regulating metabolism in mice, we tested the binding affinity of these analogues to FXR. Based on the characteristic that ligands induce nuclear receptor to recruit cofactors37, we used his-tagged FXR ligand binding domain (LBD) expressed and purified from BL21 (DE3) and synthetic biotin-labelled coactivator peptides including steroid receptor coactivator (SRC) 1–2 and SRC2–3, to perform the AlphaScreen assay. Interestingly, the data showed that ivermectin, doramectin and abamectin, analogues that reduced lipid accumulation in mice livers, strongly promoted the coactivators recruitment by FXR (Fig. 4a). In contrast, eprinomectin and moxidectin, analogues that did not reduce lipid accumulation in mice livers, failed to induce the recruitment of either biotin-SRC1–2 or biotin-SRC2–3 to FXR LBD (Fig. 4a). Since ligands can regulate the transcriptional activity of nuclear receptors by binding into their ligand binding pocket, we performed cell-based reporter assay to validate the results from AlphaScreen. Eprinomectin and moxidectin failed to activate the transcriptional activity of FXR (Fig. 4b). Similar to ivermectin28, doramectin and abamectin selectively activated the transcriptional activity of FXR using an EcRE reporter, with weaker activity than GW4064, but had no substantial impact on other nuclear receptors tested (Supplementary Fig. S3). It’s reported that LXRα antagonist can attenuate high-fat diet-induced nonalcoholic fatty liver38. To test whether avermectins regulate metabolism by targeting LXRs as antagonists, we performed a competitive reporter assay. The results showed that avermectins had no significant effects on the transcriptional activities of LXRs stimulated by T0901317 (Supplementary Fig. S4), demonstrating that avermectins are not LXRs antagonists.


Selective targeting of nuclear receptor FXR by avermectin analogues with therapeutic effects on nonalcoholic fatty liver disease.

Jin L, Wang R, Zhu Y, Zheng W, Han Y, Guo F, Ye FB, Li Y - Sci Rep (2015)

The potency of avermectin analogues in regulating metabolism correlates with their FXR binding capabilities.(a) Various coactivator motifs bind to FXR in response to 0.5 μM avermectin analogues or GW4064 by AlphaScreen assay. (b) 1 μM compounds induce FXR activity in reporter assays. (c) Dose responses of compounds in inducing FXR to recruit SRC1-2 coregulator binding motif by AlphaScreen assay. (d) Dose response of compounds in inducing the activity of FXR in reporter assays. (e) The recruitment of NCoR-2 to FXR in response to avermectin analogues by AlphaScreen assay. Values are the means ± SEM of three independent experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4664883&req=5

f4: The potency of avermectin analogues in regulating metabolism correlates with their FXR binding capabilities.(a) Various coactivator motifs bind to FXR in response to 0.5 μM avermectin analogues or GW4064 by AlphaScreen assay. (b) 1 μM compounds induce FXR activity in reporter assays. (c) Dose responses of compounds in inducing FXR to recruit SRC1-2 coregulator binding motif by AlphaScreen assay. (d) Dose response of compounds in inducing the activity of FXR in reporter assays. (e) The recruitment of NCoR-2 to FXR in response to avermectin analogues by AlphaScreen assay. Values are the means ± SEM of three independent experiments.
Mentions: To investigate the molecular mechanism of various avermectin analogues in differential regulating metabolism in mice, we tested the binding affinity of these analogues to FXR. Based on the characteristic that ligands induce nuclear receptor to recruit cofactors37, we used his-tagged FXR ligand binding domain (LBD) expressed and purified from BL21 (DE3) and synthetic biotin-labelled coactivator peptides including steroid receptor coactivator (SRC) 1–2 and SRC2–3, to perform the AlphaScreen assay. Interestingly, the data showed that ivermectin, doramectin and abamectin, analogues that reduced lipid accumulation in mice livers, strongly promoted the coactivators recruitment by FXR (Fig. 4a). In contrast, eprinomectin and moxidectin, analogues that did not reduce lipid accumulation in mice livers, failed to induce the recruitment of either biotin-SRC1–2 or biotin-SRC2–3 to FXR LBD (Fig. 4a). Since ligands can regulate the transcriptional activity of nuclear receptors by binding into their ligand binding pocket, we performed cell-based reporter assay to validate the results from AlphaScreen. Eprinomectin and moxidectin failed to activate the transcriptional activity of FXR (Fig. 4b). Similar to ivermectin28, doramectin and abamectin selectively activated the transcriptional activity of FXR using an EcRE reporter, with weaker activity than GW4064, but had no substantial impact on other nuclear receptors tested (Supplementary Fig. S3). It’s reported that LXRα antagonist can attenuate high-fat diet-induced nonalcoholic fatty liver38. To test whether avermectins regulate metabolism by targeting LXRs as antagonists, we performed a competitive reporter assay. The results showed that avermectins had no significant effects on the transcriptional activities of LXRs stimulated by T0901317 (Supplementary Fig. S4), demonstrating that avermectins are not LXRs antagonists.

Bottom Line: Mechanistically, the avermectin analogues that interact with FXR exhibited features as partial agonists, with distinctive properties in modulating coregulator recruitment.Structural features critical for avermectin analogues to selectively bind to FXR were also revealed.Additionally, the structural features that discriminate the selective binding of FXR by avermectin analogues may provide a unique safe approach to design drugs targeting FXR signaling.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Cellular Stress Biology, Innovation Center for Cell Signaling Network, School of Life Sciences, Xiamen University, Fujian 361005, China.

ABSTRACT
Non-alcoholic fatty liver disease (NAFLD) has become a predictive factor of death from many diseases. Farnesoid X receptor (FXR) is an ideal target for NAFLD drug development due to its crucial roles in lipid metabolism. The aim of this work is to examine the molecular mechanisms and functional roles of FXR modulation by avermectin analogues in regulating metabolic syndromes like NAFLD. We found that among avermectin analogues studied, the analogues that can bind and activate FXR are effective in regulating metabolic parameters tested, including reducing hepatic lipid accumulation, lowering serum cholesterol and glucose levels, and improving insulin sensitivity, in a FXR dependent manner. Mechanistically, the avermectin analogues that interact with FXR exhibited features as partial agonists, with distinctive properties in modulating coregulator recruitment. Structural features critical for avermectin analogues to selectively bind to FXR were also revealed. This study indicated that in addition to antiparasitic activity, avermectin analogues are promising drug candidates to treat metabolism syndrome including NAFLD by directly targeting FXR. Additionally, the structural features that discriminate the selective binding of FXR by avermectin analogues may provide a unique safe approach to design drugs targeting FXR signaling.

No MeSH data available.


Related in: MedlinePlus