Discovery and implementation of transcriptional biomarkers of synthetic LXR agonists in peripheral blood cells

<p>Abstract</p> <p>Background</p> <p>LXRs (Liver X Receptor α and β) are nuclear receptors that act as ligand-activated transcription factors. LXR activation causes upregulation of genes involved in reverse cholesterol transport (RCT), including ABCA1 and ABCG1 transporters, in macrophage and intestine. Anti-atherosclerotic effects of synthetic LXR agonists in murine models suggest clinical utility for such compounds.</p> <p>Objective</p> <p>Blood markers of LXR agonist exposure/activity were sought to support clinical development of novel synthetic LXR modulators.</p> <p>Methods</p> <p>Transcript levels of LXR target genes ABCA1 and ABCG1 were measured using quantitative reverse transcriptase/polymerase chain reaction assays (qRT-PCR) in peripheral blood from mice and rats (following a single oral dose) and monkeys (following 7 daily oral doses) of synthetic LXR agonists. LXRα, LXRβ, ABCA1, and ABCG1 mRNA were measured by qRT-PCR in human peripheral blood mononuclear cells (PBMC), monocytes, T- and B-cells treated <it>ex vivo </it>with WAY-252623 (LXR-623), and protein levels in human PBMC were measured by Western blotting. ABCA1/G1 transcript levels in whole-blood RNA were measured using analytically validated assays in human subjects participating in a Phase 1 SAD (Single Ascending Dose) clinical study of LXR-623.</p> <p>Results</p> <p>A single oral dose of LXR agonists induced ABCA1 and ABCG1 transcription in rodent peripheral blood in a dose- and time-dependent manner. Induction of gene expression in rat peripheral blood correlated with spleen expression, suggesting LXR gene regulation in blood has the potential to function as a marker of tissue gene regulation. Transcriptional response to LXR agonist was confirmed in primates, where peripheral blood ABCA1 and ABCG1 levels increased in a dose-dependent manner following oral treatment with LXR-623. Human PBMC, monocytes, T- and B cells all expressed both LXRα and LXRβ, and all cell types significantly increased ABCA1 and ABCG1 expression upon <it>ex vivo </it>LXR-623 treatment. Peripheral blood from a representative human subject receiving a single oral dose of LXR-623 showed significant time-dependent increases in ABCA1 and ABCG1 transcription.</p> <p>Conclusion</p> <p>Peripheral blood cells express LXRα and LXRβ, and respond to LXR agonist treatment by time- and dose-dependently inducing LXR target genes. Transcript levels of LXR target genes in peripheral blood are relevant and useful biological indicators for clinical development of synthetic LXR modulators.</p

Properties of adenylate cyclase in skeletal muscle

Bibliography: p. 121-132

Nambi P. Reversal of postischemic acute renal failure with a selective endothelin A receptor antagonist in the rat

Studies were designed to examine the effect of a selective endothelinA (ETA) receptor antagonist, BQ123, on severe postischemic acute renal failure (ARF) in Sprague-Dawley rats. Severe ARF was induced in uninephectomized, chronically instrumented rats by 45-min renal artery occlusion. BQ123 (0.1 mg/kg. min) or vehicle was infused intravenously for 3 h on the day after ischemia. Measurements before infusion (24 h control) showed a 98 % decrease in glomerular filtration rate (GFR), increase in fractional excretion of sodium from 0.6 to 39%, and in plasma K+ from 4.3 to 6.5 mEq/liter. All vehicletreated rats died in 4 d because of continuous deterioration of renal function, resulting in an increase of plasma K+ to fatal levels (&gt; 8 mEq/liter). Infusion of BQ123 significantly improved survival rate (75%) by markedly improving tubular reabsorption of Na &apos; and moderately increasing GFR and K+ excretion. Plasma K+ returned to basal levels by the 5th d after ischemia. Improved tubular function was followed by gradual recovery in GFR and urinary concentrating mechanism. Additional data from renal clearance studies in rats with moderate ARF (30-min ischemia) and in normal rats with intact kidneys showed that ETA receptor blockade increases Nan reabsorption and has no effect on renal hemodynamics. These results indicate that in the rat, the ETA receptor subtype mediates tubular epithelial function, and it plays a significant role in the pathogenesis of ischemia-induced ARF. Treatment with the selective ETA receptor antagonist reverses deteriorating tubular function in established ARF, an effect of possible therapeuti

Nonpeptide Endothelin Receptor Antagonists. XI. Pharmacological Characterization of SB 234551, a High-Affinity and Selective Nonpeptide ET A Receptor Antagonist

ABSTRACT The present study describes the pharmacological profile of ((E)-alpha- [[1-butyl-5-[2-[(2-carboxyphenyl)methoxy]-4-methoxy-phenyl]-1H-pyrazol-4-yl]methlene]-6-methoxy-1,3-benzodioxole-5-propanoic acid) (SB 234551), a high-affinity, nonpeptide endothelin type A (ET A )-selective receptor antagonist. In human cloned ET A and endothelin type B (ET B ) receptors, SB 234551 produced a concentration-dependent displacement of [ 125 I]-endothelin-1 with K i values of 0.13 and 500 nM, respectively. SB 234551 elicited concentration-dependent, rightward competitive shifts in the endothelin-1 concentration-response curves in isolated rat aorta and isolated human pulmonary artery (ET A receptor-mediated vascular contraction) with K b values of 1.9 and 1.0 nM, respectively. SB 234551 antagonized ET B receptor-mediated vasoconstriction in the isolated rabbit pulmonary artery, as demonstrated by concentration-dependent, rightward shifts in the sarafotoxin S6c concentrationresponse curves (K b ϭ 555 nM). SB 234551 produced weak functional inhibition of sarafotoxin S6c-mediated endotheliumdependent relaxation (IC 50 ϭ 7 M). SB 234551 (10 M) had no significant effect against contraction produced by several other vasoactive agents and did not significantly influence radioligand binding to a number of diverse receptors. SB 234551 (0.1-1.0 mg/kg i.v.) dose-dependently inhibited the pressor response to exogenous endothelin-1 in conscious rats. In vivo pharmacokinetic analysis in the rat demonstrated that SB 234551 was rapidly absorbed from the GI tract with a bioavailability of 30%. SB 234551 had a plasma half-life of 125 min and a systemic clearance of 25.0 ml/min/kg. The present study demonstrates that SB 234551 is an antagonist with high affinity for the ET A receptor, while sparing the ET B receptor. SB 234551 is a new pharmacological tool that should assist in the elucidation of the role of endothelin in pathophysiology. A critical need in the search for endothelin-based therapeutics is clarification of the physiological and pathophysiological functions of the endothelin receptor subtypes. Effects associated with ET A receptor activation include vasoconstriction, mitogenic activity, electrolyte excretion, microvascular permeability and release of biological mediator