Sayo Horibe

Molecular changes to HeLa cells on continuous exposure to SN-38, an active metabolite of irinotecan hydrochloride

It is important to clarify the molecular characteristics of tumor cells showing multidrug resistance (MDR) and to identify the novel targets or biomarkers for chemotherapy. The aim of this study is to establish resistant HeLa sublines through exposure to SN-38, an active metabolite of irinotecan hydrochloride, and to investigate their molecular changes. HeLa cells were exposed to SN-38 at 1, 10, or 100 nM, and resistant clones were isolated and named HeLa/SN1, HeLa/SN10, and HeLa/SN100, respectively. Their cellular changes were examined based on growth inhibition assays, the function of ABCG2/BCRP, and a RT-PCR analysis of MDR-related protein. The sublines showed a decrease in sensitivity to not only SN-38 but also other chemotherapeutic agents as compared with HeLa cells. mRNA and protein levels of ABCG2/BCRP were increased, and the transport activity of ABCG2/BCRP was enhanced, in the resistant cells. In addition, the expression levels of ABCC1/MRP1, ABCC3/MRP3, and ABCC5/MRP5 were higher than in HeLa cells. The mRNA levels of GGT1 encoding a gamma-glutamyl transferase, but not GCS encoding a gamma-glutamyl cysteine synthetase, were also higher. Other factors examined, i.e., topoisomerase, SLCO1B1, and apoptosis-regulating factors, were comparable among the cells. The overexpression of ABCG2/BCRP was involved in the mechanism of resistance in SN-38-tolerant cells, and ABCC1/MRP1, ABCC3/MRP3, ABCC5/MRP5, and GGT1 may also have participated.

Practical Training in Pharmaceutical Health Care and Sciences for Undergraduate Course Students at Kyoto Pharmaceutical University: Retrospective Evaluation from 2000 to 2002

A retrospective evaluation of practical training in pharmaceutical health care andsciences undergone by third-year undergraduate students at Kyoto Pharmaceutical University was conducted through an anonymous questionnaire survey of students from 2000 to 2002. During these three years, the students had been very conscious of the necessity of the practical training for them. Therefore, the expectations of students of the practical training increased year by year and students satisfaction with the training was greater than their expectations in every year. However, the students did express various opinions and desires regarding the practical training program and system. Based on the results of the survey, we considered ideas for improvement and solutions to problems. Modifications proposed included taking more care in selecting the practical training schedule, shortening of daily practical training sessions, concentration on training in communication skills, and changing from a passive to an active training schedule, including small group discussion, presentations, and role-plays. In 2003, these modifications were applied in the practical training in pharmaceuticalhealth care and sciences at Kyoto Pharmaceutical University.In conclusion, our survey was useful in that it enabled us to understand the needsof our students better and points of the practical training requiring improvement. We also consider that the survey was important because it enabled us to evaluate the practical training program for ourselves. We now hope to make the present practical training more effective for learning professional ethics required of pharmacists as a member of the clinical team.

Distribution of Abcg2 (BCRP) and Abcc2 (MRP2) mRNAs in rat small intestine and liver

The purpose of this study is to examine the distribution of Abcg2 and Abcc2 mRNAs in the intestine and liver of male Wistar rats, and any regional differences. Nine-week-old male rats were used, and their intestine divided equally into nine segments. The expression patterns of Abcg2 and Abcc2 mRNAs in the intestine and liver were examined using a reverse transcription-polymerase chain reaction (RT-PCR). PCR products for Abcg2 mRNA were readily detectable in the intestine, but no bands were detected in the liver. In addition, the level of Abcg2 mRNA increased significantly from the duodenum to ileum, indicating a regional difference in Abcg2 mRNA expression in the intestine. This regional difference was comparable to that of Abcb1/mdr1a as reported previously. On the other hand, Abcc2 mRNA was detected at a higher level in the liver than in the intestine. However, there was no regional difference in Abcc2 mRNA expression in the intestine. Collectively, Abcg2 was predominant in the intestine rather than liver in male Wistar rats, whereas Abcc2 was predominant in the liver. These findings will provide useful information for evaluating the gastrointestinal secretion of drugs as well as drug-drug interactions in rats.

Expression profiles of a drug metabolizing enzyme CYP3A62 mRNA in the intestine of rats

The purpose of this study was to examine the expression profiles of a drug metabolizing enzyme cytochrome P450 (CYP) 3A62 mRNA in the intestine and liver of normal male Wistar rats, and its regional differences in the intestinal tract. Nine-week-old male rats were used, and their intestine were divided equally into nine segments. The expression patterns of CYP3A62 mRNA in the intestine and liver were examined using a reverse transcription-polymerase chain reaction (RT-PCR). The PCR products for CYP3A62 mRNA were readily detectable in the intestine of normal male rats, whereas no bands for CYP3A62 mRNA were detected in the liver using similar PCR conditions as those used in the intestine. The expression of CYP3A62 mRNA was found to be high in the jejunum, and stable until the bottom of the small intestine. Only the decreased expression pattern in the bottom of the intestine was contradictory to our previous findings on CYP3A9 and CYP3A18 mRNAs. Unlike other CYPs of the CYP3A subfamily, CYP3A62 was found to be predominant in the intestine rather than the liver of normal male Wistar rats. This suggests that CYP3A62 also has an important role in the detoxification of xenobiotics similar to endogenous substances as an absorptive barrier in the intestine. These findings may help the evaluation of gastrointestinal drug absorption and drug-drug interaction in rats.