Dechlorination and demethylation of ochratoxin A enhance blocking activity of PXR activation, suppress PXR expression and reduce cytotoxicity

Abstract

The pregnane X receptor (PXR) has been established to induce chemoresistance and metabolic diseases. Ochratoxin A (OTA), a mycotoxin, decreases the expression of PXR protein in human primary hepatocytes. OTA is chlorinated and has a methylated lactone ring. Both structures are associated with OTA toxicity. The study was to test the hypothesis that structural modifications differentially impact PXR blocking activity over cytotoxicity. To test this hypothesis, OTA-M and OTA-Cl/M were synthesized. OTA-M lacked the methyl group of the lactone-ring, whereas OTA-Cl/M had neither the methyl group nor the chlorine atom. The blocking activity of PXR activation was determined in a stable cell line, harboring both PXR (coding sequence) and its luciferase element reporter. OTA-Cl/M showed the highest blocking activity, followed by OTA-M and OTA. OTA-Cl/M was 60 times as potent as the common PXR blocker ketoconazole based on calculated IC 50 values. OTA-Cl/M decreased by 90% the expression of PXR protein and was the least cytotoxic among the tested compounds. Molecular docking identified that OTA and its derivatives interacted with different sets of residues in PXR, providing a molecular basis for selectivity. Excessive activation of PXR has been implicated in chemoresistance and metabolic diseases. Downregulation of PXR protein expression likely delivers an effective mechanism against structurally diverse PXR agonists.

Influence of Bisphenol Compounds at Nanomolar Concentrations on Chromosome Damage Induced by Metabolically Activated Carcinogens in HepG2 Cells

“…As shown in Figure , coexposure of the cells to ABT (60 μM), a potent suicide inhibitor of various CYP enzymes, blocked the micronucleus formation by all carcinogens; BAY-218, an inhibitor of AhR, in the pretreatment, however, selectively blocked the effect of BaP. When KET, an inhibitor of PXR and CYP3A, , was included, the effect of AFB1 was selectively reduced, and the presence of CINPA1, an inhibitor of CAR, in the pretreatment slightly reduced the effect of NNK. The effect of benzene was nearly unaffected by BAY-218, KET, or CINPA1 (complete data presented in Table S6).…”

“…At 24 h, cells were pretreated with each BP compound at varying concentrations for 48 h. Then, the medium was changed, and after 2 h, each carcinogen (other than benzene) was added for another 48 h. Alternatively, a regime of 6 h exposure/42 h recovery was applied to the treatment with benzene, as persistent exposure to benzene may lead to false-negative results . In some experiments, known inducers of NRs, that is, PCB 126 (40 nM) for AhR, RIF (10 μM) for PXR, and CITCO (1 μM) for CAR, were used for pretreating the cells; while in some other experiments, known inhibitors of each NR, that is, BAY-218 (700 nM) for AhR, KET (10 μM) for PXR and CYP3A enzymes, , and CINPA1 (5 μM) for CAR, were present during both the pretreatment and treatment (98 h in total); alternatively, the CYP inhibitor ABT (60 μM) was present in some cultures immediately after the removal of BPs until the end of treatment (50 h in total). Please see the scheme of the various regimes shown in Figure .…”

In vitro and in vivo evaluation of AFB1 and OTA-toxicity through immunofluorescence and flow cytometry techniques: A systematic review

“…Hepatotoxicity. As shown in Table 5, the only study reviewed was conducted by Shen et al (2020). It has been proved that OTA-dichlorination (OTA Cl/M) and demethylation (OTA M) triggered the downregulation of pregnane X receptor (PXR), whose excessive activation is related to lipid retention, metabolic diseases and toxicological bioactivation of drugs.…”

Pregnane X Receptor (PXR) Polymorphisms and Cancer Treatment

“…Some of these antagonists, ketoconazole, coumestrol and metformin, are reported to inhibit PXR's transactivation either via interfering with PXR's coactivators or via binding in the AF-2 domain independently of PXR LBD, while ketoconazole is reported to be able to bind to two distinct PXR binding pockets either causing allosteric or direct inhibition of coactivator binding [42,43]. Ochatoxin A, a mycotoxin, has also been shown to significantly downregulate PXR activity in human primary hepatocytes [46,47]. Other antagonists are clotrimazole, dabrafenib, SR12813, Nelfinavir and SPA70 [48,49].…”