New paradigm for expediting drug development in Asia.

Some Asian regulators currently require Phase I data in Asians before joining global Phase II/III trials. Here, we discuss inherent limitations of Phase I ethnic sensitivity studies (ESS) to identify potential interethnic differences. We review recent new drug applications (NDAs) for Japan and China to critically assess the value of separate ESSs in Asian populations. Given that the observed value of ESS was limited, we propose a new global drug development paradigm: if relevant safety, pharmacokinetic (PK), and pharmacogenetic (PG) data are available from the original Phase I study population, it might be possible to extrapolate those data to Asian populations for their inclusion in Phase II/III trials, without an ESS. This could help to streamline drug development in Asia while still addressing regulatory requirements.

[Effect of occlusal form in buccal cusps of molars on masticatory function and intraoral food flow].

PURPOSE: The degree of crushing and intraoral food flow was evaluated using peanuts by the occlusal forms of a removable bridge as indices to clarify the occlusal formative effect of maxillary molars on masticatory function. METHODS: The subject was a 29-year-old male without any gnathostomatic disorders and his removable bridge ([7] 6 [5]) was so made in interocclusal distance as to be 0.5, 1.0 or 1.5 mm from 0 mm standard having occlusal contact to antagonist. Then the subject was asked to chew 3 g of peanuts in a series of unilateral chewing schemes consisting of 5, 10, and 20 masticatory strokes each. The pool rate in the buccal oral vestibule and lingual proper oral cavity as an index of food flow was calculated as the weight of peanut particles accumulated in each side as a percentage of the total volume of recovered peanut particles. Also, the degree of crushing was calculated as the weight of peanut particles that passed through 10-mesh sieves as a percentage of the total weight in each side. RESULTS: As the interocclusal distance to the mandibular cusp increased with abrasion on the inside at maxillary buccal cusps, the pool rate of the buccal oral vestibule was significantly increased and correspondingly that of the lingual proper oral cavity was decreased. The degree of crushing indicated non-significances statistically among 0, 0.5, and 1.0 mm and significantly decreased in 1.5 mm as compared with 0 mm. CONCLUSION: As the interocclusal distance to mandibular cusps from the maxillary inside of buccal cusps was increased, the pool rate of the crushed peanuts in the buccal side increased and the pool rate of crushed peanuts in the lingual side decreased, and the particles showed a comparatively low degree of crushing. It is clear that maxillary buccal occlusion has an effect on the crushing function and food flow.

Discovery of a Novel Piperidine-Based Inhibitor of Cholesteryl Ester Transfer Protein (CETP) That Retains Activity in Hypertriglyceridemic Plasma.

Herein we describe the discovery and characterization of a novel, piperidine-based inhibitor of cholesteryl ester transfer protein (CETP) with a core structure distinct from other reported CETP inhibitors. A versatile synthesis starting from 4-methoxypyridine enabled an efficient exploration of the SAR, giving a lead molecule with potent CETP inhibition in human plasma. The subsequent optimization focused on improvement of pharmacokinetics and mitigation of off-target liabilities, such as CYP inhibition, whose improvement correlated with increased lipophilic efficiency. The effort led to the identification of an achiral, carboxylic acid-bearing compound 16 (TAP311) with excellent pharmacokinetics in rats and robust efficacy in hamsters. Compared to anacetrapib, the compound showed substantially reduced lipophilicity, had only modest distribution into adipose tissue, and retained potency in hypertriglyceridemic plasma in vitro and in vivo. Furthermore, in contrast to torcetrapib, the compound did not increase aldosterone secretion in human adrenocortical carcinoma cells nor in chronically cannulated rats. On the basis of its preclinical efficacy and safety profile, the compound was advanced into clinical trials.

[Evaluation of prosthetic treatment using number of chewing strokes until swallowing of water absorbing rice crackers].

PURPOSE: Various methods of evaluating the ability of mastication have been reported, most of which involve evaluating of the ability to comminute foods. In this study, a new method of evaluating the ability of mastication focusing on bolus formation was applied to denture wearers. METHODS: One hundred and twenty-five subjects (71 males and 54 females) using removable dentures and receiving follow-up in the Niigata University Hospital were selected. This group was categorized by Eichner's classification, and investigated for the effect of prosthetic treatment by counting the number of chewing strokes until the first swallow of rice crackers. RESULTS: It was clarified that the number of chewing strokes until the first swallow counted by visual observation agreed well with the number of chewing strokes counted by electromyogram wave of masseter muscle, laryngeal motion, and swallowing sound, as a preparatory experiment. Using the method of counting chewing strokes by visual observation, the effect of prosthetic treatment was evaluated, and the results were as follows. 1)There is a significant difference in the number of chewing strokes without dentures between the Eichner A group, B group, and C group. 2)There was no significant difference in the number of chewing strokes with dentures among the three groups. 3)A significant decrease in the number of chewing strokes was proven by the use of removable dentures. 4)Both the improvement rate of chewing strokes (the ratio of the number of chewing strokes with dentures until the first swallow to that without dentures) and the difference of chewing strokes increased significantly in the order of Eichner A, B, and C. CONCLUSION: It was clarified that the effect of prosthetic treatment can be quantitatively evaluated by counting the chewing strokes of rice cracker. Moreover, it was suggested that the condition of prosthetic treatment can be evaluated by analyzing the improvement rate of chewing strokes.

Ethnic sensitivity assessment of pharmacokinetics and pharmacodynamics of omalizumab with dosing table expansion.

A three-part license expansion for omalizumab (Xolair(®)), humanized anti-IgE antibody, was recently made in Japan for paediatric use, additional higher doses and revised dosing frequency in allergic asthma. The dosing level and frequency of omalizumab are guided by a dosing table based on the total serum IgE and bodyweight. Nonlinear mixed-effect pharmacokinetic (PK) and pharmacodynamic (PD) modeling and simulation techniques described the binding between omalizumab and its target IgE. The population PKPD analysis was conducted using data from the nine studies included originally in the European application of dosing table expansion together with three Japanese clinical studies to assess the influence of the ethnicity. Statistically significant differences between the ethnic groups were detected. These were small, within or close to bioequivalence criteria. The model described the primary pharmacology in Caucasian and Japanese patients, both adult and paediatric, with simulations showing that the interplay between the clearance, volume and binding affinity parameters was such that there was no clinical impact of the Japanese ethnic differences on either drug PK or free IgE suppression and hence the required posology.

[A study on the differences in function of free-sided and unilateral chewing].

PURPOSE: Unilateral chewing requires a greater number of chewing strokes up to the point of the first swallowing. The rate of storage of ground food particles in the oral vestibule is said to be high during unilateral chewing. However, influence of the masticatory method, that is free-sided or unilateral chewing, involved in masticatory efficiency has not been elucidated. On that point, in order to clearly represent the effects that free-sided (bilateral) chewing and unilateral chewing have on the chewing efficiency, our purpose was to examine the difference of the functions of both free-sided chewing and unilateral chewing. METHODS: We used 116 dentate subjects with normal occlusal relations, and compared the number of chewing strokes for peanuts up to the point of the first swallowing (abbreviated as "chewing strokes") for both free-sided chewing and unilateral chewing. Following this, we made calculations regarding the amount of saliva secretion, occlusal force, and contact points of posterior teeth. Based on the averages of three parameters (saliva secretion, occlusal force and contact points of posterior teeth), all the subjects were classified into two groups (high score group and low score group). In the two groups, we counted the difference in the number of free-sided chewing and unilateral chewing. And also in two groups, the amount of remaining coarse particles immediately prior to the first swallowing was investigated between free-sided chewing and unilateral chewing. RESULTS: The number of chewing strokes by free-sided chewing for all subjects was 10% less than that of unilateral chewing. Furthermore, the numbers of chewing strokes by free-sided chewing were all less than those of unilateral chewing in both groups on three parameters. The amount of remaining coarse particles by free-sided chewing was significantly smaller than that of unilateral chewing in the low score group of saliva secretion and occlusal force. CONCLUSIONS: The number of chewing strokes by free-sided chewing was less than that of unilateral chewing in both of the high score group and low score group on the amount of saliva secretion, occlusal force, and contact points of posterior teeth.

A hydroxysteroid sulfotransferase, st2b2, is a skin cholesterol sulfotransferase in mice.

The mRNA of a sulfotransferase (St2b2) mediating cholesterol sulfation was detected in mouse skin. Recombinant St2b2 also mediated the sulfation of pregnenolone, 3beta-hydroxy-5-cholen-24-oic acid, and dehydroepiandrosterone. St2b2 protein was detected in skin cytosols on Western blotting. The addition of 10 nM TPA to skin epidermal cells from newborn mice resulted in a twofold increase in cholesterol sulfation and concomitantly enhanced the St2b2 content after 40 h. Other candidate cholesterol sulfotransferases, St2a4 and St2a9, were not detected in skin by RT-PCR. These results indicate that St2b2 is a cholesterol sulfotransferase in mouse skin.

Functional role of residues in the helix B' region of cytochrome P450 2B1.

Comparison of several recently determined X-ray crystal structures of mammalian cytochrome P450 family 2 enzymes suggests considerable movement of helix B' when ligands bind. To investigate the functional role of helix B' in P450 2B1, residues 100-109 were substituted with alanine and phenylalanine. Kinetic properties were examined with the typical 2B substrates 7-benzyloxyresorufin, 7-ethoxy-4-trifluoromethylcoumarin, benzphetamine, and testosterone. Several mutants showed 2- to 3-fold changes in k(cat) values and significant differences in catalytic efficiencies among the substrates examined, consistent with structural information suggesting that the helix B' region can adopt multiple conformations with different contact residues depending on the substrate. Homology modeling of P450 2B1 was performed based on an inhibitor-bound P450 2B4 structure, and the docking analyses were consistent with experimental results. The findings suggest that residues in the helix B' region affect regio- and stereoselective oxidation in P450 family 2 enzymes as well as substrate entry.

Unique properties of a renal sulfotransferase, St1d1, in dopamine metabolism.

Although catecholamine sulfation is higher in the kidney than in the liver of mice, no detectable amounts of previously reported sulfotransferases (STs) such as St1a, St1b, St1c, and St1e were expressed in mouse kidney cytosols. A new sulfotransferase (St1d1) cDNA was isolated from kidney cDNA library of BALB/c strain by reverse transcription-polymerase chain reaction (RTPCR) using information from expressed sequence tags (EST) database. The cDNA sequence resembled that of cDNA reported previously (AA238910) (Sakakibara et al., 1998) but differed in two amino acids, (206)Q/K and (216)Y/F, in the deduced amino acid sequence. The St1d1 expressed had unique substrate specificities for catecholamine derivatives, which preferred their deaminated metabolites rather than their parent amines. St1d1 showed the highest activity toward 3,4-dihydroxyphenylacetic acid (230.2 +/- 2.69 nmol/mg/min) among the examined substrate. St1d1 protein was abundant in kidney, followed by liver, lung, and uterus. Furthermore, an addition of anti-St1d1 serum in the cytosolic reaction mixture resulted in complete inhibition of the sulfotransferase activity suggesting a major role of St1d1 on catecholamine sulfations. No human ST1D ortholog was detected at both mRNA and protein levels, although ST1A5 selectively catalyzing parent amine sulfation was detected in human kidney. These results indicate the functional basis of sulfation and the clear species difference on renal catecholamine metabolisms in mice and humans.

Phenol sulfotransferase, ST1A3, as the main enzyme catalyzing sulfation of troglitazone in human liver.

Since sulfation is the main metabolic pathway of troglitazone, accounting for about 70% of the metabolites detected in human plasma, we have aimed to identify human cytosolic sulfotransferases catalyzing the sulfation of troglitazone and to examine a possible role of the sulfation in the cytotoxicity observed in cell lines of human origin (HepG2 and Hep3B). Experiments using the recombinant sulfotransferases and human liver cytosols indicated that phenol sulfotransferase (ST1A3) and estrogen sulfotransferase (ST1E4) were the sulfotransferases most active toward troglitazone. Immunoblot analyses indicated that hepatic content of ST1A3 is about 13 times higher than that of ST1E4, suggesting that ST1A3 is mainly responsible for the sulfation of troglitazone in the liver. Lactate dehydrogenase (LDH) leakage was elicited by troglitazone in a concentration-dependent manner in the hepatoma cells. The troglitazone metabolites (the sulfate, glucuronide, and quinone forms) caused negligible LDH leakage. These findings suggest that accumulation of unmetabolized troglitazone causes the cytotoxicity in the hepatoma cells and may be responsible for toxicity in human liver.

Protective role of hydroxysteroid sulfotransferase in lithocholic acid-induced liver toxicity.

Supplement of 1% lithocholic acid (LCA) in the diet for 5-9 days resulted in elevated levels of the marker for liver damage aspartate aminotransferase and alkaline phosphatase activities in both farnesoid X receptor (FXR)-null and wild-type female mice. The levels were clearly higher in wild-type mice than in FXR-null mice, despite the diminished expression of a bile salt export pump in the latter. Consistent with liver toxicity marker activities, serum and liver levels of bile acids, particularly LCA and taurolithocholic acid, were clearly higher in wild-type mice than in FXR-null mice after 1% LCA supplement. Marked increases in hepatic sulfating activity for LCA (5.5-fold) and hydroxysteroid sulfotransferase (St) 2a (5.8-fold) were detected in liver of FXR-null mice. A 7.4-fold higher 3alpha-sulfated bile acid concentration was observed in bile of FXR-null mice fed an LCA diet compared with that of wild-type mice. Liver St2a content was inversely correlated with levels of alkaline phosphatase. In contrast, microsomal LCA 6beta-hydroxylation was not increased and was in fact lower in FXR-null mice compared in wild-type mice. Clear decreases in mRNA encoding sodium taurocholate cotransporting polypeptide, organic anion transporting polypeptide 1, and liver-specific organic anion transporter-1 function in bile acid import were detected in LCA-fed mice. These transporter levels are higher in FXR-null mice than wild-type mice after 1% LCA supplement. No obvious changes were detected in the Mrp2, Mrp3, and Mrp4 mRNAs. These results indicate hydroxysteroid sulfotransferase-mediated LCA sulfation as a major pathway for protection against LCA-induced liver damage. Furthermore, Northern blot analysis using FXR-null, pregnane X receptor-null, and FXR-pregnane X receptor double-null mice suggests a repressive role of these nuclear receptors on basal St2a expression.