The stomach is a digestive organ that is essential for nutrient intake. Alcohol, drugs, stress, and Helicobacter pylori reduce barrier function against acidic pH, subsequently causing gastritis and gastric ulcer. Moreover, gastric cancer is one of the most common malignant tumors in the world. The mortality due to Japanese gastric cancer was over 50000 deaths annually according to the Center for Cancer Control and Information Services, National Cancer Center, Japan. Surgical resection of gastric cancer without metastasis is a first-line therapy and provides relatively good prognosis compared with that of other cancers. However, recurrence, liver metastasis, and/or peritoneal dissemination in gastric cancer are serious problems associated with high mortality. Advanced gastric cancer does not generally respond to conventional chemotherapy or radiotherapy. 1)Gene delivery to the stomach is a rational approach to treat gastric ulcer and cancer since various genes are correlated with these diseases.2) There are mainly three routes to deliver a transgene to the stomach: mucosal route, 3) direct injection, 4) and serosal route. 5-7) Gene transfer via the mucosal route is hardly possible owing to the barrier function of the mucosal layer of the stomach. Direct injection is effective to transfer a transgene; however, the area of transfection is restricted to the injection site(s). The serosal route, which we have developed, 5-7) is attractive for treatment against serosal invasion of gastric ulcer and cancer. Interestingly, simple instillation of naked plasmid DNA solution onto organ surfaces, including liver, 8) kidney, 9) spleen, 10) and gastric serosal surfaces, 5-7) resulted in effective transgene expression in mice and rats. We already elucidated that the mechanism of naked plasmid DNA transfer was uptake via Rac-mediated macropinocytosis.11) However, we were not sure whether the transfection efficiency was sufficient to treat gastric diseases. Moreover, the duration of transgene expression was relatively short. Thus, we considered it necessary to improve transgene expression.We hypothesized two strategies to improve transgene expression. One is enhancement of Rac signaling pathway controlling macropinocytosis of plasmid DNA. However, protein kinase C enhancer (phorbol 12-myristate 13-acetate, PMA), which is one of the macropinocytosis enhancers, did not affect transgene expression after naked plasmid DNA transfer onto the gastric serosal surface in mice (data not shown). Another strategy is utilization of physical force(s). With regard to physical forces, several methods have been reported, such as electroporation 12) and use of a gene gun. 13) However, both methods require specialized devices. Here, we developed a simple method to enhance transgene expression in gastric serosal surface cells by rubbing the surface using a medical spoon after instillation of naked plasmid DNA in rats and mice. This simple rubbing method improved not only transfection efficiency but also duration of transgene expression. MATERIALS A...

Gene transfer to intraperitoneal organs is thought to be a promising approach to treat such conditions as peritoneal fibrosis and peritoneal dissemination of cancers. We previously discovered that simple instillation of naked plasmid DNA (pDNA) onto intraperitoneal organs such as the liver and stomach could effectively transfer foreign genes in mice. In this study, we developed a novel nonviral method to enhance transfection efficiency of naked pDNA to intraperitoneal organs using a calcium carbonate suspension containing pDNA. Using commercially available calcium carbonate, we successfully transfected pDNA to the stomach. Handling of commercially available calcium carbonate, however, was troublesome owing to rapid precipitation and caking. To obtain slowly settling particles of calcium carbonate, we tried to synthesize novel versions of such particles and succeeded in creating flower-shaped particles, named calcium carbonate microflowers. Sedimentation of calcium carbonate microflowers was sufficiently slow for in vivo experiments. Moreover, the transfection efficiency of the suspension of calcium carbonate microflowers to the stomach was more effective than that of commercially available calcium carbonate, especially at low concentrations. Intraperitoneal injection of the suspension of calcium carbonate microflowers containing pDNA greatly enhanced naked pDNA transfer to whole intraperitoneal organs in mice. Furthermore, lactate dehydrogenase activities in intraperitoneal fluid and plasma were not raised by the suspension of calcium carbonate microflowers.

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We have developed a simple administration method, which is gastric serosal surface instillation of naked plasmid DNA (pDNA) in experimental animals. The purpose of this study was to improve gastric gene transfer efficiency by pre-treatment with a macropinocytosis enhancer, such as fetuin or epidermal growth factor (EGF), in mice. A series of concentrations of fetuin were instilled onto gastric serosal surface prior to instillation of naked pDNA in mice; however, fetuin did not improve transgene expression in the stomach 6 h after administration of pDNA. EGF also did not affect transgene expression in the stomach when pDNA was instilled immediately after EGF instillation. On the other hand, when pDNA was instilled onto gastric serosal surface 24 h after EGF treatment, transgene expression in the stomach was significantly improved by 2.6-fold. In addition, transgene-positive cells were increased 5.3-fold by EGF pre-treatment. High transgene expression in the stomach lasted for 48 h in the EGF pre-treatment group in comparison with that in the no pre-treatment group. These findings are valuable to develop an effective method of in vivo gene transfer to the stomach.

Back ground: Patients with schizophrenia who often have poor medication adherence could easily take gummi drugs daily resulting in improvement of their therapy. Because gummi drugs are chewed, drugs with an unpleasant bitter taste in gummi formulations may result in poor adherence. In this study, we developed a cocoa-flavored gummi drug containing aripiprazole (ARP) from commercially available orally disintegrating tablets (ODTs) of ARP. We aimed to clarify the palatability of gummi drugs of ARP by performing gustatory sensation test in healthy volunteers. Methods: Four types of ARP gummi formulations (6.0 mg of ARP/3.5 g of gummi drug) were prepared as follows: formulation without organoleptic masking agents (ARP-G); with aspartame (A-ARP-G); with cocoa powder and flavor (CF-ARP-G); and with a combination of aspartame, cocoa powder, and flavor (ACF-ARP-G). A gustatory sensation test was performed in 10 healthy adult volunteers (mean age, 23.7 years). The volunteers chewed each gummi drug in their oral cavities, and then spat out the drug. We used the visual analog scale (VAS) to evaluate bitterness and the overall palatability of ARP-G, A-ARP-G, CF-ARP-G, and ACF-ARP-G while chewing and just after spitting out the drugs. VAS scores of 100 corresponding to bitterness and overall palatability meant "very bitter" and "good," respectively. Results: We developed a cocoa-flavored gummi drug of ARP from commercially available ARP-ODTs. In the gustatory sensation test, the VAS scores of bitterness while chewing the gummi drugs did not differ among the different formulations. The VAS scores of bitterness of A-ARP-G and ACF-ARP-G were significantly lower after spitting out the gummi drugs, while the VAS scores of sweetness were significantly higher than that of ARP-G. The VAS scores of overall palatability of ACF-ARP-G both during chewing and after spitting out were the highest among all gummi drugs tested and significantly higher than those of ARP-G. Conclusion: We were able to develop a cocoa-flavored gummi drug of ARP possessing good palatability from commercially available tablets. Therefore, ARP gummi drugs will be acceptable to patients and help improve their medication adherence.