YAKUGAKU ZASSHI, 127(5),779-779, 2007
YAKUGAKU ZASSHI, 127(5),781-787, 2007
Gene therapy has a potentiality for treatment of cancer and diseases from genomic defects. It is important to select a vector which has good potency in terms of gene transduction efficiency, and is safe and easy to apply. Many researchers have attempted to develop an effective gene delivery carrier. Recently, it was reported that microbubbles, which are ultrasound (US) contrast agents, improved the transfection efficiency by cavitation with US exposure. However, microbubbles had problems with stability and targeting ability. To solve these problems, we focused on liposomes that had many advantages such as being stable and safe in vivo and easily modifying targeting ligand. We succeeded in preparing the liposomes (“Bubble liposomes” (BLs)) entrapping perfluoropropane gas which was utilized for contrast enhancement in ultrasonography. In this study, we assessed the feasibility of BLs as gene delivery carrier utilized cavitation by US exposure. BLs could deliver plasmid DNA to various cell types in vitro by combining with US without cytotoxicity. To evaluate the ability of BLs to in vivo gene delivery, we attempted to deliver plasmid DNA into the femoral artery. The gene expression at this artery treated with BLs and US combination was higher than with US only, BLs without US or Lipofectamine 2000. This result suggested that Bubble liposomes could quickly deliver plasmid DNA into the artery even under conditions of short contact time between BLs and the endothelial cells and the existence of the bloodstream and serum. These results suggested that BLs might be a non-invasive and effective carrier for gene delivery.
Key words--liposome; gene therapy; ultrasound; microbubble; cavitation
YAKUGAKU ZASSHI, 127(5),789-796, 2007
Drug delivery system (DDS) research has contributed greatly toward improving chemotherapy efficacy and reducing its adverse effects through the development of approaches to optimize pharmacokinetics, such as controlled release and targeting. On the other hand, the remarkable progress of this latest life science research has altered the concept of what constitutes medical supplies. A change in this concept would allow for the consideration of medical materials that use not only conventional low molecular-weight organic compounds, but also biomacromolecules, including nucleic acids and proteins, that constitute living organisms. Although these biomacromolecular drugs are expected to demonstrate excellent efficacy based on their intrinsic bioactivity, they quickly degrade when administered in vivo and only a limited number have therefore been developed into medicines. In addition, most biomacromolecular drugs are ineffective until they are delivered to particular cells within a tissue or to particular organelles within a cell. To develop effective biomacromolecular medicines, it is necessary to introduce a DDS that is capable of ensuring internal stability as well as precise control of internal and intracellular dynamics, and to establish a new fundamental technology for DDS that can accommodate the material properties and mechanisms of action of the biomacromolecular drugs. In this context, this review introduces our approach to the design and creation of “Intracellular DDS” using fusogenic liposomes for application to gene therapy and tumor peptide vaccines. We suggest that this technology is very important for controlling the intracellular pharmacokinetics of biomacromolecular drugs.
Key words--drug delivery system; fusogenic liposome; nanoparticle; gene therapy; peptide vaccine
YAKUGAKU ZASSHI, 127(5),797-805, 2007
To maximize the beneficial effects and minimize the side effect of drugs, DDS (drug delivery system) has been attracted many researchers in the recent drug development. Especially, the in vivo pinpoint delivery system for drugs is very important and key technology for developing the next generations of anti-cancer drugs and gene therapies. Bio-nanocapsule (BNC) is recombinant yeast-derived hepatitis B virus surface antigen particle, which has been used as a recombinant hepatitis B vaccine for the last 20 years in the world. BNC can incorporate various materials (chemical compounds, proteins, genes, siRNA, etc) by the fusion with liposome, and deliver them to the organs and tissues in vivo specifically by the action of bio-recognition molecules on the BNC's surface. The transfection efficiency is significantly higher than that of liposome, because BNC harbors the complete set of hepatitis B virus infection machinery. Recently, we succeeded in the in vivo retargeting of BNC by displaying either antibody or homing peptide, less than 10 amino acid residues for in vivo targeting. BNC is a hybrid of liposome and virus, and very flexible system for in vivo retargeting. BNC might be very promising carriers in the next generation of DDS.
Key words--bio-nanocapsule; drug delivery system; in vivo pinpoint delivery; gene therapy
YAKUGAKU ZASSHI, 127(5),807-812, 2007
RNAi has rapidly become a powerful tool for drug target discovery and validation in an in vitro culture system and, consequently, interest is rapidly growing for extension of its application to in vivo systems, such as animal disease models and human therapeutics. Novel treatments and drug discovery in pre-clinical studies based on RNAi are currently targeting a wide range of diseases, including viral infections and cancers by the local administration of synthetic small interfering RNA (siRNA) that target local lesions. Recently, specific methods for the systemic administration of siRNAs have been reported to treat non-human primates or a cancer metastasis model. In vivo siRNA-delivery technology is a key hurdle to the successful therapeutic application of RNAi. This article reviews the non-viral delivery system of atelocollagen for siRNA, which could be useful for functional screening of the genes in vitro and in vivo, and will provide a foundation for further development of RNAi therapeutics.
Key words--RNAi; siRNA; atelocollagen; delivery
YAKUGAKU ZASSHI, 127(5),813-823, 2007
Biologically active peptides for therapeutic use have relatively short half-lives in general, requiring appropriate controlled-release systems for better therapy. Controlled release of peptides is, however, not as easy as that of conventional drugs because their large molecular size is much more dramatic in hindering the diffusion and release from polymeric devices. From this perspective, we have been developing two types of microcapsular devices containing new acrylate-based nanogels with a specific solute-permeability for delayed- or thermosensitive-release of peptide drugs. The microcapsule preparation was accomplished by an air suspension coating process. A nanogel-particle of acrylic terpolymer, ethyl acrylate-methyl methacrylate-2-hydroxyethyl methacrylate, was newly synthesized by emulsion polymerization to construct delayed-release microcapsules. By spray-coating the insulin-loaded lactose particles with the acrylic terpolymers, microcapsules showing a pH-independent delayed-release profile can be obtained. Oral administration of the microcapsules with the lag time of 6 hours to beagle dogs resulted in significantly reduced blood glucose concentration, leading to colon-specific insulin delivery with pharmacological availability of 5%. Meanwhile, poly(N-isopropylcarylamide) (p(NIPAAm)) nanogel-particles with a reversible temperature-dependent swelling property were prepared by dispersion polymerization to fabricate microcapsular membranes with thermosensitively changeable permeability. The microcapsules constructed by coating of drug-loaded CaCO3 particles with a blend mixture of the p(NIPAAm) nanogels and ethylcellulose pseudo-latex exhibited an ‘on-off’ positively thermosensitive drug-release; the release rate was remarkably enhanced at higher temperatures possibly due to the formation of voids through the shrinkage of p(NIPAAm) nanogels in the membrane. A possible application of this type of microcapsules can be found in externally temperature-activated pulsatile peptide delivery.
Key words--peptide drug; nanohydrogel; microcapsule; controlled release; coating; acrylic polymer
YAKUGAKU ZASSHI, 127(5),825-837, 2007
Drug delivery systems (DDS) have customarily been developed as part of the technology and methodology used to enhance the in vivo efficacy of therapeutic drugs. However, the DDS concept can also be used for prophylactic and diagnostic drugs to enhance their respective medical efficacy. When applied to biological signaling factors, such as growth factors and genes, which can regulate the proliferation and differentiation of cells, DDS are expected to realize cell-based tissue regeneration therapy. Basic research on biology and medicine which scientifically supports the advanced medical therapies currently available will be advanced by making use of DDS techniques. This paper overviews functional drug carriers indispensable for DDS which are part of the fundamental technology and methodology to achieve such advances.
Key words--functional drug carriers; drug delivery systems; cell-based tissue regeneration; tissue engineering; advanced medical therapy; basic research
YAKUGAKU ZASSHI, 127(5),839-840, 2007
YAKUGAKU ZASSHI, 127(5),841-845, 2007
The regenerative potential of stem cells has recently been under intense investigation. In vitro, stem and progenitor cells have the ability for self-renewal and differentiation into organ-specific cell types. In vivo, transplantation of these cells may reconstitute organ systems, as shown in animal models of disease. In contrast, differentiated cells do not exhibit such characteristics. Human endothelial progenitor cells (EPCs) have been isolated from the peripheral blood of adult individuals, expanded in vitro and committed to an endothelial lineage in culture. The transplantation of these human EPCs has been shown to facilitate successful salvage of limb vasculature and perfusion in athymic nude mice with severe hindlimb ischemia, while differentiated endothelial cells (human microvascular endothelial cells) failed to accomplish limb-saving neovascularization. Future studies will clarify the mechanisms and circumstances that may be responsible for modulating the contribution of vasculogenesis to postnatal neovascularization. Specifically in this regard, it is intriguing to consider the possibility that certain angiogenic growth factors acknowledged to promote both angiogenesis and vasculogenesis in the embryo, but have been assumed to promote neovascularization exclusively by angiogenesis in the adult, may in fact promote migration, proliferation, and mobilization of EPCs from bone marrow. The possibility that modulation of vasculogenesis can be used therapeutically to augment as well as inhibit neovascularization deserves further investigation.
Key words--endothelial progenitor cell; stem cell; vascular regeneration; ischemic disease
YAKUGAKU ZASSHI, 127(5),841-850, 2007
Biodegradable polymers are often used as scaffolds for tissue engineering and these polymers are in class IV under the revised Pharmaceutical Affairs Law. From the point of view of safety and efficacy, recent problems in the development of tissue-engineered products using biodegradable polymers are summarized in this report.
Key words--regenerative medicine; cell/tissue-engineered medical products; biodegradable materials; safety; efficacy; scaffolds
YAKUGAKU ZASSHI, 127(5),851-856, 2007
Several recent studies demonstrated the potential of bioengineering using somatic stem cells in regenerative medicine. Adult human mesenchymal stem cells (hMSCs) derived from bone marrow have the pluripotency to differentiate into cells of mesodermal origin, e.g., bone, cartilage, adipose, and muscle cells; they, therefore, have many potential clinical applications. On the other hand, stem cells possess a self-renewal capability similar to cancer cells. For safety evaluation of tissue engineered medical devices using normal hMSCs, in this study, we investigated the expression levels of several genes that affect cell proliferation in hMSCs during in vitro culture. We focused on the relationship between the hMSC proliferation and their transforming growth factor β (TGFβ) signaling during in vitro culture. The proliferation rate of hMSCs gradually decreased and cellular senescence was observed for about 3 months. The mRNA expressions of TGFβ1, TGFβ2, and TGFβ receptor type I (TGFβRI) in hMSCs increased with the length of cell culture. The mRNA expressions of Smad3 increased, but those of c-myc and nucleostemin decreased with the length hMSCs were in in vitro culture. In addition, the expression profiles of the genes which regulate cellular proliferation in hMSCs were significantly different from those of cancer cells. In conclusion, hMSCs derived from bone marrow seldom underwent spontaneous transformation during 1-2 months in vitro culture for use in clinical applications. In hMSCs as well as in epithelial cells, growth might be controlled by the TGFβ family signaling.
Key words--human mesenchymal stem cells; tissue engineered medical devices; proliferation; transforming growth factor β
YAKUGAKU ZASSHI, 127(5),857-863, 2007
Because the capacity of articular cartilage for repair is limited, defects are a major clinical problem, and there is at present no satisfactory clinical technique to regenerate cartilage defects. Current clinical practice involves the bone stimulation technique, which breaks subchondral bone to facilitate cartilage repair from bone marrow derived cells and cytokines. This consists of multiple perforations, abrasions, and micro-fractures. However, with this procedure, cartilage defects are repaired with fibrocartilage, which is known to be biochemically and biomechanically different from normal hyaline cartilage and degeneration occurs in the reparative tissue. Autologous chondrocyte implantation (ACI) for repair of human articular cartilage was reported in 1994, and approved by the USA Food and Drug Association in1997. This procedure has been performed for more than 20000 people all over the world, but its effectiveness is still controversial. Mosaic plasty was explored in the 1990s. Using this procedure, we can repair defects with hyaline cartilage, but the donor site morbidity is unsolved. To explore a new method for cartilage repair, we transplanted autologous culture-expanded bone marrow mesenchymal cells into articular cartilage defects. Clinical symptoms were improred but the repair cartilage was not hyaline cartilage. Further improvement is required. Many investigations have been made in the search for better means of repair, including gene transduction and the addition of growth factors during cell culture. In addition to bone marrow mesenchymal cells, synovial cells, adipocytes, muscle cells, etc. have been evaluated.
Key words--chondrocyte; progenitor cell; transplantation
YAKUGAKU ZASSHI, 127(5),865-888, 2007
To clarify the marketing status of antimicrobial products, descriptions on the labels of commercially available antimicrobial products were investigated from 1991 through 2005, and the results were analyzed using a database system on antimicrobial deodorant agents. A classification table of household antimicrobial products was prepared and revised, based on which target products were reviewed for any changes in the product type. The number of antimicrobial products markedly increased over 3 years starting from 1996, among which there were many products apparently not requiring antimicrobial processing. More recently, in the 2002 and 2004 surveys, while sales of kitchenware and daily necessities decreased, chemical products, baby articles, and articles for pets increased; this poses new problems. To clarify the use of antimicrobial agents in the target products, a 3-step (large, intermediate, small) classification table of antimicrobial agents was also prepared, based on which antimicrobial agents indicated on the product labels were checked. The rate of identifying the agents increased. However, this is because of the increase of chemical products and baby articles, both of which more frequently indicated the ingredient agents on the labels, and the decrease of kitchenware and daily necessities, which less frequently indicated them on the labels. Therefore there has been little change in the actual identification rate. The agents used are characterized by product types: quaternary ammonium salts, metal salts, and organic antimicrobials are commonly used in textiles, plastics, and chemical products, respectively. Since the use of natural organic agents has recently increased, the safety of these agents should be evaluated.
Key words--antimicrobial products; market research; classification table of products; classification table of antimicrobial agents; investigation on commercially available products; database system on antimicrobial deodorant agents
YAKUGAKU ZASSHI, 127(5),889-896, 2007
Gynostemma pentaphyllum is a popular herbal tea in China and some Asian countries. The modulatory function of G. pentaphyllum total plant extracts on immune cells was evaluated in this study. The extract was intraperitoneally injected into mice for 5 consecutive days. The production of antibodies from B cells or cytokines from T cells was determined mainly with ELISA. After the treatment, serum IgM and IgG2a were significantly enhanced and showed dose-dependent effect. Moreover, serum IgA and IgG1 were also increased when received the extract at the doses of 0.05 or 0.50 g/kg/day. In addition to the serum levels, the injection of the extract enhanced the production of all antibodies from LPS-activated spleen cells. Furthermore, more cytokines were secreted from Con A-stimulated splenocytes of G. pentaphyllum-treated mice. Our results suggest that the extract of G. pentaphyllum might promote immune responses through the activation of T and B cells.
Key words--Gynostemma pentaphyllum; antibodies; cytokines; mice
YAKUGAKU ZASSHI, 127(5),897-901, 2007
Currently in Japan, the preferred method for blood purification in patients with acute renal failure is continuous hemodiafiltration (CHDF). However, CHDF filters out various antifungal drugs such as fluconazole through large pores in the membrane used. Systemic fungal infection is still one of the main causes of death and complications in critically ill patients in intensive care units (ICUs). Therefore it is important to determine the appropriate use of antifungal agents. This study was designed to evaluate the influence of CHDF on the pharmacokinetics of the antifungal agent micafungin in ICU patients. The pharmacokinetics of micafungin were studied in four ICU patients receiving CHDF and in nine ICU patients not receiving CHDF. To evaluate the pharmacokinetics, the ratio of serum micafungin concentration to dose per body weight (C/D) was used in this study. There was no progressive accumulation or exclusion of micafungin in patients receiving CHDF. The mean (±S.D.) extraction rate (%) for micafungin during CHDF was 3.6±3.9. There was no significant difference in the serum micafungin C/D-time profiles between the patients receiving and not receiving CHDF. These results show that CHDF does not affect the pharmacokinetics of micafungin. Therefore it is not necessary to adjust the micafungin dose in patients receiving CHDF.
Key words--continuous hemodiafiltration; micafungin; critical care; pharmacokinetics
YAKUGAKU ZASSHI, 127(5),903-904, 2007
We determined the in vitro antibacterial activity of ciprofloxacin against Vibrio cholerae O139 recently isolated from cholera patients in India. Ciprofloxacin showed excellent antibacterial activity against the O139 strains, and ciprofloxacin-resistant O139 strains were not observed. The lack of incidence of ciprofloxacin resistance in O139 strains may be because O139 strains appeared comparatively recently and have not been extensively treated with antibacterial agents including fluoroquinolones.
Key words--antibacterial activity; ciprofloxacin; Vibrio cholerae O139
YAKUGAKU ZASSHI, 127(5),905-917, 2007
The aim of this study was to analyze students, achievement rate and contents of assessment judged by instructors in objective structured clinical examination (OSCE) attempted at the Faculty of Pharmaceutical Science, Showa University. The OSCE was carried out for fourth-year students in May 28, 2005. In this trial, there were two stations, i.e., counting/measurement dispensing and subsequent audit of dispensed drugs, and 218 students and 31 instructors (as evaluators) participated. We developed a checklist to test students attitudes and skills (two stages) and overall evaluation (five stages). Each student was evaluated by two instructors. Examination time was 8 minutes for drug dispensing, and 4 minuets for the audit of dispensed drug. After the OSCE trial, we analyzed validity of examination time, contents of assessment, and differences in scores between different evaluators. More than half of the students could not finish the examination within the limit of time for dispensing the liquid and cream and audit for dispensed powder. The number of items that 60% of the students achieved was 48 (82.8%). Moreover, 20% of the assessment items did not agree among the evaluators with a disagreement rate of 20% or more. Thus, we distinguished between the items based on the extent of disagreement rates. It was suggested that most of the students achieved such a level to actually perform clinical training in pharmacies. From these results, it is necessary to set up an assignment to finish the within the time limit to extent the time limit depending upon examination contents, to standardize the evaluation to increase the agreement rate among evaluators, and to more clearly identify assessment criteria.
Key words--objective structured clinical examination (OSCE); pharmacy education; evaluation
YAKUGAKU ZASSHI, 127(5),919-922, 2007
In a previous paper we reported a new function of tissue kallikrein: rat urinary kallikrein (RUK) has a marked growth-stimulatory effect on neural stem cells prepared from brains of prenatal rats. We here report that conspicuous differentiation of neural stem cells to neurons and/or glial cells did not occur during stimulation by kallikrein, and this growth-stimulating effect of kallikrein is considerably specific for neural stem cells, i.e., RUK showed no detectable stimulatory effect on rat glial, PC12, GH3, and HeLa cells. Thus this new function of kallikrein suggests the potential involvement of this enzyme on brain physiology. In addition, this effect of kallikrein for neural stem cells may have value in the treatment of cerebral ischemic stroke-induced injuries etc., i.e., kallikrein administered into the body stimulates neural stem cell proliferation and new neurons may be generated from them.
Key words--kallikrein; brain; neural stem cell; neuron; proliferation; cerebrum