Kosuke Kusamori

Transdermal delivery of relatively high molecular weight drugs using novel self-dissolving microneedle arrays fabricated from hyaluronic acid and their characteristics and safety after application to the skin

The purpose of this study was to develop novel dissolving microneedle arrays fabricated from hyaluronic acid (HA) as a material and to improve the transdermal permeability of relatively high molecular weight drugs. In this study, fluorescein isothiocyanate-labeled dextran with an average molecular weight of 4kDa (FD4) was used as a model drug with a relatively high molecular weight. The microneedle arrays significantly increased transepidermal water loss (TEWL) and reduced transcutaneous electrical resistance (TER), indicating that they could puncture the skin and create drug permeation pathways successfully. Both TEWL and TER almost recovered to baseline levels in the microneedle array group, and relatively small pathways created by the microneedles rapidly recovered as compared with those created by a tape stripping treatment. These findings confirmed that the microneedle arrays were quite safe. Furthermore, we found that the transdermal permeability of FD4 using the microneedle arrays was much higher than that of the FD4 solution. Furthermore, we found that the microneedle arrays were much more effective for increasing the amount of FD4 accumulated in the skin. These findings indicated that using novel microneedle arrays fabricated from HA is a very useful and effective strategy to improve the transdermal delivery of drugs, especially relatively high molecular weight drugs without seriously damaging the skin.

Development of a Novel Self-Dissolving Microneedle Array of Alendronate, a Nitrogen-Containing Bisphosphonate: Evaluation of Transdermal Absorption, Safety, and Pharmacological Effects After Application in Rats

Alendronate is a nitrogen-containing bisphosphonate that is widely used for the treatment of osteoporosis. In this study, we developed a novel self-dissolving micron-size needle array (microneedle array) containing alendronate, which was fabricated by micromodeling technologies using hyaluronic acid as a basic material. Micron-scale pores in the skin were seen after the application of the alendronate-loaded microneedle array, verifying establishment of transdermal pathways for alendronate. The absorption of alendronate after the application of alendronate-loaded microneedle array was almost equivalent to that after subcutaneous administration, and the bioavailability of alendronate was approximately 90% in rats. Furthermore, delivery of alendronate via this strategy effectively suppressed the decrease in the width of the growth plate in a rat model of osteoporosis. Although mild cutaneous irritation was observed after the application of the alendronate-loaded microneedle array, it resolved by day 15. These findings indicate that this alendronate-loaded microneedle array is a promising transdermal formulation for the treatment of osteoporosis.

Development of a novel transdermal patch of alendronate, a nitrogen‐containing bisphosphonate, for the treatment of osteoporosis

Bisphosphonates are widely used for the treatment and prevention of bone diseases, including Paget disease, hypercalcemia of malignancy, and postmenopausal osteoporosis. In this study, we developed a novel transdermal patch of alendronate, a nitrogen‐containing bisphosphonate, for the treatment of bone diseases. The maximum permeation fluxes of alendronate through rat and human skin after application of this patch were 1.9 and 0.3 µg/cm2 per hour, respectively. The bioavailability (BA) of alendronate in rats was approximately 8.3% after the application of alendronate patch and approximately 1.7% after oral administration. These results indicated that the transdermal permeation of alendronate using this patch system was sufficient for the treatment of bone diseases. The plasma calcium level was effectively reduced after application of the alendronate patch in 1α‐hydroxyvitamin D3–induced hypercalcemia model rats. The alendronate patch also effectively suppressed the decrease in bone mass in model rats with osteoporosis. Modest alendronate‐induced erythema of rat skin was observed after application of the alendronate patch. Incorporation of butylhydroxytoluene in the alendronate patch almost completely suppressed this alendronate‐induced skin damage while maintaining the transdermal permeation and pharmacologic effects of alendronate. These findings indicate that our novel transdermal delivery system for alendronate is a promising approach to improve compliance and quality of life of patients in the treatment of bone diseases.

Transplantation of insulin-secreting multicellular spheroids for the treatment of type 1 diabetes in mice.

The efficacy of cell-based therapy depends on the function and survival of transplanted cells, which have been suggested to be enhanced by spheroid formation. However, few attempts at spheroid generation from insulin-secreting cells, which may be used to treat type 1 diabetes, have been reported. We therefore developed spheroids from the mouse insulinoma cell line NIT-1 by using polydimethylsiloxane (PDMS)-based microwells with a coating of poly(N-isopropylacrylamide) (PNIPAAm). The prepared NIT-1 spheroids or dissociated NIT-1 cells were transplanted into the subrenal capsule in streptozotocin-induced diabetic mice. NIT-1 spheroids prepared using the PNIPAAm-coated PDMS-based microwells had a uniformly sized spherical structure with a diameter of 200-300μm. The PNIPAAm coating increased cell survival in the spheroids and the recovery of the spheroids from the microwells. In diabetic mice, the transplanted NIT-1 spheroids reduced blood glucose levels to normal values faster than dissociated NIT-1 cells did. Additionally, survival was higher among NIT-1 cells in spheroids than among dissociated NIT-1 cells 24h after transplantation. These results indicate that insulin-secreting NIT-1 spheroids prepared using PNIPAAm-coated PDMS-based microwells are more effective for the treatment of type 1 diabetes than dissociated cells in suspension.

Absorption and safety of alendronate, a nitrogen-containing bisphosphonate, after intrapulmonary administration in rats.

Alendronate, a nitrogen-containing bisphosphonate, has been used as a first-choice drug for the treatment of hypercalcemia and osteoporosis. In the present study, we examined the absorption and safety of alendronate after intrapulmonary administration in rats. The bioavailability (BA) of alendronate after intrapulmonary administration was 47% at a dose of 5 mg/kg, while the BA after oral administration was only 2.9% at a dose of 50 mg/kg in rats. Plasma calcium level, an index of the pharmacological effect of alendronate, was effectively reduced after intrapulmonary administration of alendronate. Furthermore, alendronate continuously reduced the increase in plasma calcium levels for 9 days after a single intrapulmonary administration in rats with 1α-hydroxyvitamin-D(3)-induced hypercalcemia. Intrapulmonary administration of alendronate also effectively suppressed the decrease in bone mass in a rat model of osteoporosis. Alendronate significantly increased the activity of lactate dehydrogenase (LDH) in bronchoalveolar lavage fluid (BALF), indicating that pulmonary mucosal damage was induced by intrapulmonary administration of alendronate. However, co-administration of superoxide dismutase (SOD) with alendronate completely suppressed the alendronate-induced increase in LDH activity in BALF, while maintaining sufficient pulmonary absorption and therapeutic effects of alendronate in rats with 1α-hydroxyvitamin-D(3)-induced hypercalcemia. These findings indicated that the lung is a promising, noninvasive alternative route for the delivery of alendronate in the treatment of bone diseases.

Improvement of Transdermal Delivery of Exendin-4 Using Novel Tip-Loaded Microneedle Arrays Fabricated from Hyaluronic Acid.

The purpose of this study was to evaluate the characteristics of exendin-4 tip-loaded microneedle arrays and to compare their acute efficacy with subcutaneous injections in type 2 diabetic GK/Slc rats. Fluorescein isothiocyanate labeled dextran with an average molecular weight of 4,000 (FD4) was selected as a model drug, and FD4 tip-loaded microneedle arrays were prepared in this study. In addition, intraperitoneal glucose tolerance tests after application of exendin-4 tip-loaded microneedle arrays were also compared with those after subcutaneous injection in type 2 diabetic GK/Slc rats. The release of FD4 from the tip-loaded microneedle arrays was very rapid, particularly in the initial 30 s, and most of the FD4 was released within 5 min. In addition, glucose tolerance was improved and the insulin secretion was enhanced after application of exendin-4 tip-loaded microneedle arrays, and these effects were comparable to those after subcutaneous injection of exendin-4. Similar plasma concentration profiles were seen after application of exendin-4 tip-loaded microneedle arrays, as was the case with subcutaneous injection in type 2 diabetic GK/Slc rats. These findings indicate that exendin-4 tip-loaded microneedle arrays can be used as an alternative to achieve sufficient delivery of exendin-4 for treatment of type 2 diabetes. To our knowledge, this is the first report of transdermal exendin-4 delivery using tip-loaded microneedle arrays.

Improvement of transdermal delivery of sumatriptan succinate using a novel self-dissolving microneedle array fabricated from sodium hyaluronate in rats.

The purpose of the present study was to develop an alternative transdermal formulation containing sumatriptan succinate (SS) for the treatment of migraine. Novel self-dissolving SS-loaded microneedle arrays (MNs) were fabricated from sodium hyaluronate and their efficacy for transdermal delivery of SS was characterized. The resulting MNs maintained their skin piercing abilities for at least 30 min after being placed at a high relative humidity of 75%. Rapid release of SS from the MNs was also observed in vitro. Optical coherence tomography images demonstrated that MNs were able to successfully pierce into rat skin without any bending or cracking, and needles were completely dissolved within 1 h. MNs significantly increased transepidermal water loss; however, skin barrier function gradually recovered to control levels within 24 h, in contrast to the skin damage observed after tape stripping treatment. These findings indicated that the micropores created by MNs quickly resealed, and that the skin damage was reversible. Furthermore, a dose-dependent plasma concentration of SS was obtained after transdermal delivery using SS-loaded MNs in rats. Absorption of SS delivered by MNs was similar to that observed after subcutaneous injection and was associated with high bioavailability (ca. 90%), which was much higher than that produced by oral administration. These findings suggested that application of SS-loaded MNs to the skin provided an effective alternative approach to enhance the transdermal delivery of SS without serious skin damage, and would be likely to improve patient compliance.

Absorption-enhancing effects of gemini surfactant on the intestinal absorption of poorly absorbed hydrophilic drugs including peptide and protein drugs in rats.

In general, the intestinal absorption of small hydrophilic molecules and macromolecules like peptides, after oral administration is very poor. Absorption enhancers are considered to be one of the most promising agents to enhance the intestinal absorption of drugs. In this research, we focused on a gemini surfactant, a new type of absorption enhancer. The intestinal absorption of drugs, with or without sodium dilauramidoglutamide lysine (SLG-30), a gemini surfactant, was examined by an in situ closed-loop method in rats. The intestinal absorption of 5(6)-carboxyfluorescein (CF) and fluorescein isothiocyanate-dextrans (FDs) was significantly enhanced in the presence of SLG-30, such effect being reversible. Furthermore, the calcium levels in the plasma significantly decreased when calcitonin was co-administered with SLG-30, suggestive of the increased intestinal absorption of calcitonin. In addition, no significant increase in the of lactate dehydrogenase (LDH) activity or in protein release from the intestinal epithelium was observed in the presence of SLG-30, suggestive of the safety of this compound. These findings indicate that SLG-30 is an effective absorption-enhancer for improving the intestinal absorption of poorly absorbed drugs, without causing serious damage to the intestinal epithelium.

Development of a drug-coated microneedle array and its application for transdermal delivery of interferon alpha

Interferon alpha (IFNα) is one of the most famous drugs for the treatment of chronic hepatitis C and various types of human malignancy. Protein drugs, including IFNα, are generally administered by subcutaneous or intramuscular injection due to their poor permeability and low stability in the bloodstream or gastrointestinal tract. Therefore, in the present study, novel IFNα-coated polyvinyl alcohol-based microneedle arrays (IFNα-MNs) were fabricated for the transdermal delivery of IFNα without the painful injection. IFNα was rapidly released from MNs in phosphate buffered solution and these MNs presented piercing ability in the rat skin. Slight erythema and irritation were observed when MNs were applied to the rat skin, but these skin damages completely disappeared within 24 h after removing the IFNα-MNs. Furthermore, the pharmacokinetic parameters of IFNα-MNs were similar to those of IFNα subcutaneous administration. Finally, IFNα-MNs showed a significant antitumor effect in tumor bearing mice similar to that of IFNα subcutaneous administration. These results indicate that IFNα-MNs are a useful biomaterial tool for protein drug therapy and can improve the quality of life in patients by avoidance of painful injections.

Improved dissolution and absorption of ketoconazole in the presence of organic acids as pH-modifiers.

Formulation development of poorly water-soluble compounds can be challenging because of incomplete dissolution that causes low and variable bioavailability. Enhancing compound solubility is important and many techniques have been investigated to that end, but they require specific materials and machinery. This study investigates the incorporation of a pH-modifier as a method to increase compound solubility and uses ketoconazole (KZ), which is weakly basic (pKa: 6.5), as a model compound. Organic acids are effective pH-modifiers and are generally used in pharmaceutical industries. We successfully obtained granules containing variable organic acids (KZ/acid granule) using a high-shear mixer. Dissolution tests of the KZ/acid granule resulted in highly enhanced solubility under non-sink conditions. Adding water-soluble acids, such as citric acid (CA) and tartaric acid, resulted in more than 8-fold higher dissolution at pH 6.0 compared to that of KZ only. The granules containing citric acid (KZ/CA granule) improved the dissolution of KZ after oral administration to rats under low gastric acid conditions, where the bioavailability of the KZ/CA granules at elevated gastric pH was comparable with that of KZ only at gastric acidic pH. The incorporation of organic acids would result in effective therapeutic outcomes independent of gastric pH in patients. In addition, higher bioavailability of KZ was observed after oral administration of KZ/CA granules under gastric acidic pH conditions than that of KZ alone. Thus, CA improved the dissolution and absorption rate of KZ after oral administration.