Confirmation of high-throughput screening data and novel mechanistic insights into VDR-xenobiotic interactions by orthogonal assays

Abstract

High throughput screening (HTS) programs have demonstrated that the Vitamin D receptor (VDR) is activated and/or antagonized by a wide range of structurally diverse chemicals. In this study, we examined the Tox21 qHTS data set generated against VDR for reproducibility and concordance and elucidated functional insights into VDR-xenobiotic interactions. Twenty-one potential VDR agonists and 19 VDR antagonists were identified from a subset of >400 compounds with putative VDR activity and examined for VDR functionality utilizing select orthogonal assays. Transient transactivation assay (TT) using a human VDR plasmid and Cyp24 luciferase reporter construct revealed 20/21 active VDR agonists and 18/19 active VDR antagonists. Mammalian-2-hybrid assay (M2H) was then used to evaluate VDR interactions with co-activators and co-regulators. With the exception of a select few compounds, VDR agonists exhibited significant recruitment of co-regulators and co-activators whereas antagonists exhibited considerable attenuation of recruitment by VDR. A unique set of compounds exhibiting synergistic activity in antagonist mode and no activity in agonist mode was identified. Cheminformatics modeling of VDR-ligand interactions were conducted and revealed selective ligand VDR interaction. Overall, data emphasizes the molecular complexity of ligand-mediated interactions with VDR and suggest that VDR transactivation may be a target site of action for diverse xenobiotics.

Confirmation of high-throughput screening data and novel mechanistic insights into FXR-xenobiotic interactions by orthogonal assays

“… Hsu et al (2016) observed as similar phenomena where select FXRα antagonists enhanced humanFXRα:coregulator interactions in the presence of CDCA. These results are also similar to that observed with vitamin D receptor antagonists ( Mahapatra et al, 2018 ) suggesting that this process is not unique to human or medaka FXRα. Rather, this finding suggests that alternate mechanisms may be associated with receptor inhibition in lieu of coactivator recruitment.…”

Zebrafish show long-term behavioral impairments resulting from developmental vitamin D deficiency

“…In addition, because of the low cost of the zebrafish model, it could be key in providing a functional screen for determining which environmental chemicals and drugs run the risk of disrupting neurobehavioral function with developmental exposure. This is advantageous particularly coupled with high-throughput in vitro screens such as those already utilized to identify compounds with the potential for interfering with VDR signaling [24].. The results of the current study demonstrate the behavioral impairments in adults that result from dietary vitamin D deficiency throughout development.…”

Vitamin D and Toxic Metals in Pregnancy - a Biological Perspective

“…For example, Cd 2+ has been observed to substitute for Zn 2+ in the zinc finger DNA-binding domain of the estrogen receptors [9]. It has been demonstrated through high throughput screening that the VDR is activated or antagonised by a wide range of chemicals, including metals (Cd) [102]. If downstream effects are blocked or altered this could potentially increase the metabolism of vitamin D and reduce 25OHD.…”