Pharmacokinetic and pharmacodynamic characterization of an oral lysophosphatidic acid type 1 receptor-selective antagonist.

Lysophosphatidic acid (LPA) is a bioactive phospholipid that signals through a family of at least six G protein-coupled receptors designated LPA1₋6. LPA type 1 receptor (LPA1) exhibits widespread tissue distribution and regulates a variety of physiological and pathological cellular functions. Here, we evaluated the in vitro pharmacology, pharmacokinetic, and pharmacodynamic properties of the LPA1-selective antagonist AM095 (sodium, {4'-[3-methyl-4-((R)-1-phenyl-ethoxycarbonylamino)-isoxazol-5-yl]-biphenyl-4-yl}-acetate) and assessed the effects of AM095 in rodent models of lung and kidney fibrosis and dermal wound healing. In vitro, AM095 was a potent LPA1 receptor antagonist because it inhibited GTPγS binding to Chinese hamster ovary (CHO) cell membranes overexpressing recombinant human or mouse LPA1 with IC50 values of 0.98 and 0.73 µM, respectively, and exhibited no LPA1 agonism. In functional assays, AM095 inhibited LPA-driven chemotaxis of CHO cells overexpressing mouse LPA1 (IC50= 778 nM) and human A2058 melanoma cells (IC50 = 233 nM). In vivo, we demonstrated that AM095: 1) had high oral bioavailability and a moderate half-life and was well tolerated at the doses tested in rats and dogs after oral and intravenous dosing, 2) dose-dependently reduced LPA-stimulated histamine release, 3) attenuated bleomycin-induced increases in collagen, protein, and inflammatory cell infiltration in bronchalveolar lavage fluid, and 4) decreased kidney fibrosis in a mouse unilateral ureteral obstruction model. Despite its antifibrotic activity, AM095 had no effect on normal wound healing after incisional and excisional wounding in rats. These data demonstrate that AM095 is an LPA1 receptor antagonist with good oral exposure and antifibrotic activity in rodent models.

A novel, orally active LPA(1) receptor antagonist inhibits lung fibrosis in the mouse bleomycin model.

BACKGROUND AND PURPOSE: The aim of this study was to assess the potential of an antagonist selective for the lysophosphatidic acid receptor, LPA(1), in treating lung fibrosis We evaluated the in vitro and in vivo pharmacological properties of the high affinity, selective, oral LPA(1)-antagonist (4'-{4-[(R)-1-(2-chloro-phenyl)-ethoxycarbonylamino]-3-methyl-isoxazol-5-yl}-biphenyl-4-yl)-acetic acid (AM966). EXPERIMENTAL APPROACH: The potency and selectivity of AM966 for LPA(1) receptors was determined in vitro by calcium flux and cell chemotaxis assays using recombinant and native cell cultures. The in vivo efficacy of AM966 to reduce tissue injury, vascular leakage, inflammation and fibrosis was assessed at several time points in the mouse bleomycin model. KEY RESULTS: AM966 was a potent antagonist of LPA(1) receptors, with selectivity for this receptor over the other LPA receptors. In vitro, AM966 inhibited LPA-stimulated intracellular calcium release (IC(50)= 17 nM) from Chinese hamster ovary cells stably expressing human LPA(1) receptors and inhibited LPA-induced chemotaxis (IC(50)= 181 nM) of human IMR-90 lung fibroblasts expressing LPA(1) receptors. AM966 demonstrated a good pharmacokinetic profile following oral dosing in mice. In the mouse, AM966 reduced lung injury, vascular leakage, inflammation and fibrosis at multiple time points following intratracheal bleomycin instillation. AM966 also decreased lactate dehydrogenase activity and tissue inhibitor of metalloproteinase-1, transforming growth factor beta1, hyaluronan and matrix metalloproteinase-7, in bronchoalveolar lavage fluid. CONCLUSIONS AND IMPLICATIONS: These findings demonstrate that AM966 is a potent, selective, orally bioavailable LPA(1) receptor antagonist that may be beneficial in treating lung injury and fibrosis, as well as other diseases that are characterized by pathological inflammation, oedema and fibrosis.

Oral mucosal screening as an integral part of routine dental care.

The principal aim of this study was to assess the feasibility of conducting a systematic examination of the oral mucosa, as an integral part of the routine dental check-up and in conditions comparable with those in NHS dental practice. A total of 1949 individuals, who were already registered as patients with an industrial dental clinic, were invited to attend for an oral screen as part of their dental examination. Of these, 1947 patients agreed to participate and were also asked to complete a 'habits' questionnaire relating to their tobacco and alcohol use. A systematic examination of the oral mucosa was undertaken as part of the routine dental inspection and mucosal lesions were recorded as either a positive or negative screening result. Lesions included as a positive result were those which may be associated with early cancer or precancer. Four patients (0.2%) were considered to have a positive screening result and these were referred for specialist evaluation. Of these, two had tobacco-related leukoplakia, one had oral lichen planus and the other had an early squamous cell carcinoma. This study has confirmed that a systematic and thorough examination of the oral mucosa can realistically be carried out as part of the routine dental inspection in NHS dental practice.

The effect of lesion characteristic on remineralization and model sensitivity.

A major criterion for assessing the value of any experimental model in scientific research is the degree of correspondence between its results and data from the real-life process it is designed to model. Intra-oral models aimed at predicting the anti-caries efficacy of toothpastes or other topical treatments should therefore be calibrated against treatments proven to be effective in a caries clinical trial. For this to be achieved, it is necessary that a model with high sensitivity be designed, while at the same time retaining relevance to the process to be modeled. This means that the effects of the various experimental conditions and parameters of the model on its performance must be understood. The purpose of this paper was to assess the influence of two specific factors on the performance of an in situ enamel remineralization model, which is based on human enamel slabs attached to partial dentures. The two factors are initial lesion severity and origin of enamel sample. The results indicated that initial lesion size affected whether net remineralization or net demineralization occurred during in situ treatment. Samples with an initial range of from 1500 to 2500 (delta Z) tended more toward demineralization than did samples with delta Z greater than 3500. This means that treatment groups must be well-balanced with respect to initial lesion size. Differences in initial demineralization severity between different tooth locations must also be considered so that systematic treatment bias can be avoided. The solution used in the model discussed here is based on a balanced experimental design, which allows this effect to be taken into account in the data analysis.

The effect of study design on in situ treatment of carious enamel lesions.

The aim of this exploratory study was to investigate the influence of several factors on changes in the mineral content of carious enamel lesions treated in situ. The study involved 36 adult volunteers who used either a non-fluoride toothpaste or one of two fluoride toothpastes (1000 or 1500 ppm F). Human enamel specimens were prepared and attached to partial dentures as described previously (Schäfer, 1989) and treated in situ for between three and six weeks. The mineral content of lesions was determined by microradiography and computerized densitometry. The factors investigated in this study included study length, frequency of treatment, trial design, patient compliance, patient diet, and previous caries experience of the patient. The effects observed were small, relative to that of the treatment, and not statistically significant (p greater than 0.05). However, the trends in the data were all as would be intuitively predicted. Study duration correlated positively with observed lesion mineral content. Lesions worn by panelists using a fluoride toothpaste for six weeks contained greater levels of mineral with respect to placebo than did those in panelists on a three-week study. The residual variations in the three phases of the study were found to be similar, suggesting that there is no advantage in these studies having a cross-over design. Brushing frequency also correlated positively with observed lesion mineral content, with panelists brushing three times per day with a fluoridated dentifrice having lesions with greater levels of mineral, with respect to placebo, than those brushing twice per day. Overall, no clear relationship between reported diet and changes in lesion mineral levels was apparent.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparison between evaluation methods for the anti-caries efficacy of monofluorophosphate-containing dentifrices.

A comparison is described of three methods of preclinical assessment of potential anti-caries efficacy for topical fluoride treatments. Methods are compared using dentifrices containing 1,000, 1,500 and 2,500 ppm F as sodium monofluorophosphate (SMFP). These formulations have been shown elsewhere to give a statistically significant dose response of increasing anti-caries efficacy [Stephen et al., 1988]. An in situ enamel insert model, oral fluoride pharmacokinetics and F uptake to demineralised enamel are all shown to correlate with clinical efficacy for the test dentifrices studied.