Investigation of 4-piperidinols as novel H3 antagonists.

Compounds containing a substituted 4-piperidinol core have been found to be potent antagonists of the human H(3) receptor. The compounds exhibited up to a 60-fold preference for inhibiting the human H(3) receptor over the mouse and showed a low binding affinity for the hERG channel.

3-Indolyl sultams as selective CRTh2 antagonists.

CRTh2 (DP(2)) is a prostaglandin D(2) receptor implicated in the recruitment of eosinophils and basophils within the asthmatic lung. Here we report the discovery of a novel series of 3-indolyl sultam antagonists with low nM affinity for CRTh2. These compounds proved to be selective over the other primary prostaglandin D(2) receptor (DP1) as well as the thromboxane A(2) receptor (TP).

Parallel functional activity profiling reveals valvulopathogens are potent 5-hydroxytryptamine(2B) receptor agonists: implications for drug safety assessment.

Drug-induced valvular heart disease (VHD) is a serious side effect of a few medications, including some that are on the market. Pharmacological studies of VHD-associated medications (e.g., fenfluramine, pergolide, methysergide, and cabergoline) have revealed that they and/or their metabolites are potent 5-hydroxytryptamine(2B) (5-HT(2B)) receptor agonists. We have shown that activation of 5-HT(2B) receptors on human heart valve interstitial cells in vitro induces a proliferative response reminiscent of the fibrosis that typifies VHD. To identify current or future drugs that might induce VHD, we screened approximately 2200 U.S. Food and Drug Administration (FDA)-approved or investigational medications to identify 5-HT(2B) receptor agonists, using calcium-based high-throughput screening. Of these 2200 compounds, 27 were 5-HT(2B) receptor agonists (hits); 14 of these had previously been identified as 5-HT(2B) receptor agonists, including seven bona fide valvulopathogens. Six of the hits (guanfacine, quinidine, xylometazoline, oxymetazoline, fenoldopam, and ropinirole) are approved medications. Twenty-three of the hits were then "functionally profiled" (i.e., assayed in parallel for 5-HT(2B) receptor agonism using multiple readouts to test for functional selectivity). In these assays, the known valvulopathogens were efficacious at concentrations as low as 30 nM, whereas the other compounds were less so. Hierarchical clustering analysis of the pEC(50) data revealed that ropinirole (which is not associated with valvulopathy) was clearly segregated from known valvulopathogens. Taken together, our data demonstrate that patterns of 5-HT(2B) receptor functional selectivity might be useful for identifying compounds likely to induce valvular heart disease.

Comparative evaluation of AMPLICOR HPV PCR and Linear Array assays on SurePath liquid-based Pap samples for the detection of high-risk HPV genotypes.

BACKGROUND: Persistent cervical infection with high-risk [HR] HPV is a causative factor for cancer. Liquid-based [L-Pap] Pap samples are convenient for HPV testing and SurePath samples have been least studied. Most HPV tests have multiple step protocols and testing laboratories experience large volumes of samples. OBJECTIVES: Using SurePath L-Pap residual samples the objectives were as follows: [1] to test the performance of AMP-HPV and LA-HPV. [2] To perform an agreement study between two laboratories for the AMP-HPV test and [3] to compare agreement of results between AMP-HPV and LA-HPV and HC2. STUDY DESIGN: Samples from 657 women were tested for Pap cytology then assayed for HR-HPV using AMP-HPV and LA-HPV tests. AMP-HPV performance was compared between 2 laboratories and agreement studies were conducted between AMP-HPV, LA-HPV and HC2. RESULTS: HR-HPV genotypes were associated with L-Pap readings as follows: HSIL 92% [23/25], LSIL 73.6% [162/220], ASCUS 70.4% [131/186], normal 31.9% [72/226]. More women less than 30 were infected with HR-HPV and multiple genotypes regardless of the L-Pap reading. AMP-HPV and LA-HPV testing had an overall raw agreement with each other of 84.2% [Kappa 0.66] and each had agreement of 94% with HC2 testing of 133 samples [Kappa 0.86/0.87]. AMP-HPV agreement between two laboratories was better at 93% [Kappa 0.84] compared to 76.1% [Kappa 0.40] when extraction was standardized. CONCLUSION: It is feasible to perform AMP-HPV and LA-HPV on SurePath samples to detect HR-HPV genotypes. HC2, AMP-HPV and LA-HPV showed strong agreement. The extraction component of the AMP-HPV assay needs careful attention to yield consistent results.

A homogeneous fluorescent live-cell assay for measuring 7-transmembrane receptor activity and agonist functional selectivity through beta-arrestin recruitment.

Seven-transmembrane (7TM) receptors play an essential role in the regulation of a wide variety of physiological processes, making them one of the top target classes for pharmaceuticals. 7TM receptor function is mediated and modulated through 2 primary processes: G-protein and beta-arrestin signaling. Classically, it has been recognized that these 2 processes can interact with one another during 7TM receptor desensitization, but it has more recently been recognized that these 2 processes can also act independently of one another and can activate parallel signaling pathways. As such, the methods used to interrogate 7TM receptor signaling, both from a biological and a pharmaceutical perspective, may need to be reevaluated and the question of whether functionally selective compounds (compounds that selectively activate one pathway over another) can be rationally developed must be raised. Although numerous high-throughput screening (HTS) compatible assays exist for studying second messengers arising from G-protein signaling, far fewer HTS compatible assays exist for studying beta-arrestin recruitment. The authors report on the Tango 7TM receptor assay technology, a high-throughput homogeneous assay method for monitoring beta-arrestin recruitment that uses a live-cell fluorescent readout. This assay format is broadly applicable to 7TM receptors, independent of G-protein coupling and, as such, has been used to produce assays for over 70 7TM receptor targets. The authors also show how flow cytometry can be used to select clones with desired pharmacological profiles and how an inducible expression system can increase the assay window for targets with high levels of constitutive activity. Finally, they demonstrate how the Tango system can be used in parallel with assays aimed at second-messenger signaling to enable functional selectivity studies.

Isosteric ramatroban analogs: selective and potent CRTH-2 antagonists.

The chemoattractant receptor-homologous molecule expressed on T(H)2 cells (CRTH-2), also found on eosinophils and basophils, is a prostaglandin D2 receptor involved in the recruitment of these cell types during an inflammatory response. In this report, we describe the synthesis and optimization of a ramatroban isostere that is a selective and potent antagonist of CRTH-2 which may be useful in the treatment of certain diseases.