Pharmacophore analysis of the nuclear oxysterol receptor LXRalpha.
A cell-free assay was developed for the orphan nuclear receptor LXRalpha that measures the ligand-dependent recruitment of a peptide from the steroid receptor coactivator 1 (SRC1) to the nuclear receptor. Using this ligand-sensing assay (LiSA), the structural requirements for activation of the receptor by oxysterols and related compounds were studied. The minimal pharmacophore for receptor activation was shown to be a sterol with a hydrogen bond acceptor at C24. 24(S),25-Epoxycholesterol (1), which meets this criterion, is among the most efficacious of the oxysterols and is an attractive candidate as the LXRalpha natural hormone. Cholenic acid dimethylamide (14) showed increased efficacy compared to 1, whereas the unnatural oxysterol 22(S)-hydroxycholesterol (4) was shown to be an antagonist of 1 in the LiSA. The structural requirements for SRC1 recruitment in the LiSA correlated with the transcriptional activity of compounds in a cell-based reporter assay employing LXRalpha-GAL4 chimeric receptors. Site-directed mutagenesis identified Trp(443) as an amino acid critical for activation of LXRalpha by oxysterol ligands. This information was combined with the structure-activity relationship developed from the LiSA to develop a 3D homology model of LXRalpha. This model may aid the design of synthetic drugs targeted at this transcriptional regulator of cholesterol homeostasis.
Spencer
T A
TA
Dartmouth College, Hanover, New Hampshire 03755, USA.
Li
D
D
Russel
J S
JS
Collins
J L
JL
Bledsoe
R K
RK
Consler
T G
TG
Moore
L B
LB
Galardi
C M
CM
McKee
D D
DD
Moore
J T
JT
Watson
M A
MA
Parks
D J
DJ
Lambert
M H
MH
Willson
T M
TM
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D015153:Blotting, Western; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002474:Cell-Free System; D002522:Chlorocebus aethiops; D002784:Cholesterol / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D002793:Cholic Acids / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D004268:DNA-Binding Proteins; D004735:Energy Transfer; D005453:Fluorescence; D051548:Histone Acetyltransferases; D006888:Hydroxycholesterols / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D007653:Ketocholesterols / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D000071518:Liver X Receptors; D008958:Models, Molecular; D008969:Molecular Sequence Data; D056920:Nuclear Receptor Coactivator 1; D057093:Orphan Nuclear Receptors; D018160:Receptors, Cytoplasmic and Nuclear / Q000819:agonists* / Q000037:antagonists & inhibitors; D011987:Receptors, Steroid / Q000819:agonists* / Q000037:antagonists & inhibitors; D013237:Stereoisomerism; D013261:Sterols / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D013329:Structure-Activity Relationship; D014157:Transcription Factors / Q000378:metabolism; D014364:Tryptophan / Q000737:chemistry
D002793:Cholic Acids; D004268:DNA-Binding Proteins; D006888:Hydroxycholesterols; D007653:Ketocholesterols; D000071518:Liver X Receptors; D057093:Orphan Nuclear Receptors; D018160:Receptors, Cytoplasmic and Nuclear; D011987:Receptors, Steroid; D013261:Sterols; D014157:Transcription Factors; C424433:cholenic acid dimethylamide; C003585:22-hydroxycholesterol; C028358:24,25-epoxycholesterol; D014364:Tryptophan; D002784:Cholesterol; D051548:Histone Acetyltransferases; D056920:Nuclear Receptor Coactivator 1