Nuclear receptors regulate metabolic pathways in response to changes in the environment by appropriate alterations in gene expression of key metabolic enzymes. Here, a computational search approach based on iteratively built hidden Markov models of nuclear receptors was used to identify a human nuclear receptor, termed hPAR, that is expressed in liver and intestines. hPAR was found to be efficiently activated by pregnanes and by clinically used drugs including rifampicin, an antibiotic known to selectively induce human but not murine CYP3A expression. The CYP3A drugmetabolizing enzymes are expressed in gut and liver in response to environmental chemicals and clinically used drugs. Interestingly, hPAR is not activated by pregnenolone 16␣-carbonitrile, which is a potent inducer of murine CYP3A genes and an activator of the mouse receptor PXR.1. Furthermore, hPAR was found to bind to and trans-activate through a conserved regulatory sequence present in human but not murine CYP3A genes. These results provide evidence that hPAR and PXR.1 may represent orthologous genes from different species that have evolved to regulate overlapping target genes in response to pharmacologically distinct CYP3A activators, and have potential implications for the in vitro identification of drug interactions important to humans.The recent progress made in determining the full genomic sequences of model organisms as well as the rapid accumulation of sequence data from the human genome has opened up new possibilities to determine the functional organization of genomes by computational approaches (1). Multiple alignments of members of different protein families followed by homology searching are powerful methods to infer gene function from sequence data as well as to identify novel genes within a given gene family. Hidden Markov models (HMMs) are a general statistical modeling technique that can be used as formal, fully probabilistic forms of sequence profiles (2, 3), describing the consensus of a set of sequences. This approach toward the identification and functional characterization of novel genes is particularly amenable to evolutionary conserved gene families consisting of a large number of orthologs and paralogs. The nuclear receptors constitute one such large gene family that is structurally and functionally conserved and represented within different metazoan phylae from cnidarians to vertebrates (4). These receptors are conditionally regulated transcription factors that exert their effects by interacting with small lipophilic ligands followed by sequence-specific binding of the receptor to DNA sequences called hormone response elements (HREs). Binding of the receptor to DNA results in changes in gene expression of specific target genes (5).Steroid hormones were the first group of small, lipophilic molecules identified as nuclear receptor ligands. Today, the number of substances known to regulate the activity of this group of receptors is represented by a large and chemically diverse group of molecules including retinoids, vi...

Alteration of gene expression is a crucial component of adaptive responses to hypoxia. These responses are mediated by hypoxia-inducible transcription factors (HIFs). Here we describe an inhibitory PAS (Per/Arnt/Sim) domain protein, IPAS, which is a basic helix-loop-helix (bHLH)/PAS protein structurally related to HIFs. IPAS contains no endogenous transactivation function but demonstrates dominant negative regulation of HIF-mediated control of gene expression. Ectopic expression of IPAS in hepatoma cells selectively impairs induction of genes involved in adaptation to a hypoxic environment, notably the vascular endothelial growth factor (VEGF) gene, and results in retarded tumour growth and tumour vascular density in vivo. In mice, IPAS was predominantly expressed in Purkinje cells of the cerebellum and in corneal epithelium of the eye. Expression of IPAS in the cornea correlates with low levels of expression of the VEGF gene under hypoxic conditions. Application of an IPAS antisense oligonucleotide to the mouse cornea induced angiogenesis under normal oxygen conditions, and demonstrated hypoxia-dependent induction of VEGF gene expression in hypoxic corneal cells. These results indicate a previously unknown mechanism for negative regulation of angiogenesis and maintenance of an avascular phenotype.

The peroxisome proliferator-activated receptors (PPAR) are members of the nuclear receptor supergene family and are considered as key sensors of both lipid and glucose homeostasis. The role of the PPAR␥ isoform in glucose metabolism is illustrated by the fact that antidiabetic thiazolidinediones have been shown to be bona fide PPAR␥ ligands. Here we report the crystal structure of apo-PPAR␥ ligand binding domain (LBD) determined to 2.9-Å resolution. Although the structure of apo-PPAR␥-LBD retains the overall fold described previously for other nuclear receptor LBDs, three distinct structural differences are evident. 1) The core AF-2 activation domain of apo-PPAR␥ LBD is folded back toward the predicted ligand binding pocket similar to that observed in the holo-forms of other nuclear receptors. 2) The proposed ligand binding pocket of apo-PPAR␥-LBD is larger and more accessible to the surface in contrast to other LBDs. 3) The region of the LBD called the -loop is extended in PPAR␥ and contains additional structural elements. Taken together, the apo-PPAR␥-LBD structure is in several aspects different from previously described LBDs. Given the central role of PPAR␥ as a mediator in glucose regulation, the structure should be an important tool in the development of improved anti-diabetic agents.

Atherosclerotic cardiovascular disease is a major problem despite the availability of drugs that influence major risk factors. New treatments are needed, and there is growing interest in therapies that may have multiple actions. Thyroid hormone modulates several cardiovascular risk factors and delays atherosclerosis progression in humans. However, use of thyroid hormone is limited by side effects, especially in the heart. To overcome this limitation, pharmacologically selective thyromimetics that mimic metabolic effects of thyroid hormone and bypass side effects are under development. In animal models, such thyromimetics have been shown to stimulate cholesterol elimination through LDL and HDL pathways and decrease body weight without eliciting side effects. We report here studies on a selective thyromimetic [KB2115; (3-[[3,5-dibromo-4-[4-hydroxy-3-(1-methylethyl)-phenoxy]-phenyl]-amino]-3-oxopropanoic acid)] in hu-mans. In moderately overweight and hypercholesterolemic subjects KB2115 was found to be safe and well tolerated and elicited up to a 40% lowering of total and LDL cholesterol after 14 days of treatment. Bile acid synthesis was stimulated without evidence of increased cholesterol production, indicating that KB2115 induced net cholesterol excretion. KB2115 did not provoke detectable effects on the heart, suggesting that the pharmacological selectivity observed in animal models translates to humans. Thus, selective thyromimetics deserve further study as agents to treat dyslipidemia and other risk factors for atherosclerosis.