Setsuo Hasegawa

Mobile Genetic Element SCCmec-encoded psm-mec RNA Suppresses Translation of agrA and Attenuates MRSA Virulence

Community acquired-methicillin resistant Staphylococcus aureus (CA-MRSA) is a socially problematic pathogen that infects healthy individuals, causing severe disease. CA-MRSA is more virulent than hospital associated-MRSA (HA-MRSA). The underlying mechanism for the high virulence of CA-MRSA is not known. The transcription product of the psm-mec gene, located in the mobile genetic element SCCmec of HA-MRSA, but not CA-MRSA, suppresses the expression of phenol-soluble modulin α (PSMα), a cytolytic toxin of S. aureus. Here we report that psm-mec RNA inhibits translation of the agrA gene encoding a positive transcription factor for the PSMα gene via specific binding to agrA mRNA. Furthermore, 25% of 325 clinical MRSA isolates had a mutation in the psm-mec promoter that attenuated transcription, and 9% of the strains had no psm-mec. In most of these psm-mec-mutated or psm-mec-deleted HA-MRSAs, PSMα expression was increased compared with strains carrying intact psm-mec, and some mutated strains produced high amounts of PSMα comparable with that of CA-MRSA. Deletion of psm-mec from HA-MRSA strains carrying intact psm-mec increased the expression of AgrA protein and PSMα, and virulence in mice. Thus, psm-mec RNA suppresses MRSA virulence via inhibition of agrA translation and the absence of psm-mec function in CA-MRSA causes its high virulence property.

Copy number variation in sulfotransferase isoform 1A1 (SULT1A1) is significantly associated with enzymatic activity in Japanese subjects.

Sulfotransferase isoform 1A1 (SULT1A1) plays a key role in the metabolism of a variety of endo- and xenobiotics and it’s activity could influence response to drugs. Our previous studies have focused on the impact of genetic variants of SULT1A1 on enzymatic activity in Caucasians and African-Americans. However, the contribution of genetic variants to SULT1A1 activity in Asians has not been explored. In this study, we investigated the collective effects of both SULT1A1 copy number variants (CNVs) and single nucleotide polymorphisms (SNPs) in the promoter region, coding region, and 3′ untranslated region on SULT1A1 activity in Japanese subjects. SNPs in the SULT1A1 promoter and 3′ untranslated region were not associated with SULT1A1 activity (P > 0.05). SULT1A1*1/2 (Arg213His) was marginally associated with SULT1A1 activity (P = 0.037). However, SULT1A1 CNVs were strongly associated with SULT1A1 activity (trend test P = 0.008) and accounted for 10% of the observed variability in activity for Japanese subjects. In conclusion, SULT1A1 CNVs play a pivotal role in determination of SULT1A1 activity in Japanese subjects, highlighting the influence of ethnic differences in SULT1A1 genetic variants on drug metabolism and therapeutic efficacy.

Microdose pharmacogenetic study of 14C-tolbutamide in healthy subjects with accelerator mass spectrometry to examine the effects of CYP2C9∗3 on its pharmacokinetics and metabolism

Microdose study enables us to understand the pharmacokinetic profiles of drugs in humans prior to the conventional clinical trials. The advantage of microdose study is that the unexpected pharmacological/toxicological effects of drugs caused by drug interactions or genetic polymorphisms of metabolic enzymes/transporters can be avoided due to the limited dose. With a combination use of accelerator mass spectrometry (AMS) and (14)C-labaled compounds, the pharmacokinetics of both parent drug and its metabolites can be sensitively monitored. Thus, to demonstrate the usability of microdose study with AMS for the prediction of the impact of genetic polymorphisms of CYP enzyme on the pharmacokinetics of unchanged drugs and metabolites, we performed microdose pharmacogenetic study using tolbutamide as a CYP2C9 probe drug. A microdose of (14)C-tolbutamide (100 μg) was administered orally to healthy volunteers with the CYP2C9(∗)1/(∗)1 or CYP2C9(∗)1/(∗)3 diplotype. Area under the plasma concentration-time curve for the (14)C-radioactivity, determined by AMS, or that for the parent drug, determined by liquid chromatography/mass spectrometry, was about 1.6 times or 1.7 times greater in the CYP2C9(∗)1/(∗)3 than in the CYP2C9(∗)1/(∗)1 group, which was comparable to the previous reports at therapeutic dose. In the plasma and urine, tolbutamide, carboxytolbutamide, and 4-hydroxytolbutamide were detected and practically no other metabolites could be found in both diplotype groups. The fraction of metabolites in plasma radioactivity was slightly lower in the CYP2C9(∗)1/(∗)3 group. Microdose study can be used for the prediction of the effects of genetic polymorphisms of enzymes on the pharmacokinetics and metabolic profiles of drugs with minimal care of their pharmacological/toxicological effects.

Bioequivalence of Rebamipide granules and tablets in healthy adult male volunteers

AbstractObjective: Rebamipide tablets, which are used in the treatment of patients with gastric ulcers or gastritis, can be difficult to administer in subjects with reduced swallowing ability or impaired swallowing. The granule formulation may be more easily administered in these patients. The bioequivalence between rebamipide granules (20%/0.5g) and tablets (100mg) was determined in healthy male adult volunteers, in accordance with the Partially Revised Guidelines for Bioequivalence Studies of Generic Products. Study design: In a randomised, nonblind, crossover design, 28 individuals were allocated into two groups of 14 to receive either rebamipide granules or rebamipide tablets. Each individual, under fasting conditions, was administered a single oral dose of rebamipide 100mg followed by a 7-day washout period. Individuals then received a single oral dose of the other rebamipide formulation. Blood samples were collected at 0.5, 1, 1.5, 2, 3, 4, 6, 8, 12 and 24 hours. Plasma rebamipide concentrations were measured by validated high-performance liquid chromatography with tandem mass spectrometry. Results: The plasma concentration-time profiles and pharmacokinetic parameters of rebamipide after administration of the granule formulation were similar to those of the tablet in 27 healthy male volunteers. Following administration of the granule formulation, the area under the plasma concentration-time curve from time 0–24 hours (AUC24h) was 912.82 μg/L · h, the maximum plasma concentration (Cmax) was 241.82 μg/L, time to maximum plasma concentration (tmax) was 2.5 hours, and plasma elimination half-life (t1/2) was 1.97 hours. Corresponding values for the tablet formulation were 873.55 μg/L · h, 216.19 μg/L, 2.4 hours, and 1.94 hours. The difference in mean log values was 1.01 for AUC24h and 1.09 for Cmax after granule and tablet administration. The 90% confidence interval of this difference in mean log value was 0.93–1.10 for AUC24h, and 0.97–1.21 for Cmax. This satisfies the criteria for bioequivalence in the guidelines [within log (0.8) to log (1.25)]. Conclusions: Rebamipide granules (20%/0.5g) and tablet (100mg) were bioequivalent. Rebamipide granules may therefore be a more practical treatment option in patients with gastric ulcers or gastritis who have difficulty swallowing tablets.