Yoshiki Kojo

Inhalable dry-emulsion formulation of cyclosporine A with improved anti-inflammatory effects in experimental asthma/COPD-model rats

The main purpose of the present study was to develop a novel respirable powder (RP) formulation of cyclosporine A (CsA) using a spray-dried O/W-emulsion (DE) system. DE formulation of CsA (DE/CsA) was prepared by spray-drying a mixture of erythritol and liquid O/W emulsion containing CsA, polyvinylpyrrolidone, and glyceryl monooleate as emulsifying agent. The DE/CsA powders were mixed with lactose carriers to obtain an RP formulation of DE/CsA (DE/CsA-RP), and its physicochemical, pharmacological, and pharmacokinetic properties were evaluated. Spray-dried DE/CsA exhibited significant improvement in dissolution behavior with ca. 4500-fold increase of dissolution rate, and then, nanoemulsified particles were reconstituted with a mean diameter of 317 nm. Laser diffraction analysis on the DE/CsA-RP suggested high dispersion of DE/CsA on the surface of the lactose carrier. Anti-inflammatory properties of the inhaled DE/CsA-RP were characterized in antigen-sensitized asthma/COPD-model rats, in which the DE/CsA-RP was more potent than the RP formulation of physical mixture containing CsA and erythritol in inhibiting inflammatory responses, possibly due to the improved dissolution behavior. Pharmacokinetic studies demonstrated that systemic exposure of CsA after intratracheal administration of the DE/CsA-RP at a pharmacologically effective dose (100 μg-CsA/rat) was 50-fold less than that of the oral CsA dosage form at a toxic dose (10 mg/kg). From these findings, use of inhalable DE formulation of CsA might be a promising approach for the treatment of airway inflammatory diseases with improved pharmacodynamics and lower systemic exposure.

Development of novel solid dispersion of tranilast using amphiphilic block copolymer for improved oral bioavailability

The present study aimed to develop novel solid dispersion (SD) of tranilast (TL) using amphiphilic block copolymer, poly[MPC-co-BMA] (pMB), to improve the dissolution and pharmacokinetic behavior of TL. pMB-based SD of TL (pMB-SD/TL) with drug loading of 50% (w/w) was prepared by wet-mill technology, and the physicochemical properties were characterized in terms of morphology, crystallinity, dissolution, and hygroscopicity. Powder X-ray diffraction and polarized light microscopic experiments demonstrated high crystallinity of TL in pMB-SD/TL. The pMB-SD/TL exhibited immediate micellization when introduced to aqueous media, forming fine droplets with a mean diameter of ca. 122 nm. There was marked improvement in the dissolution behavior for the pMB-SD/TL even under acidic conditions, although the supersaturated TL concentration gradually decreased. NMR analyses demonstrated interaction between TL and pMB, as evidenced by the chemical shift drifting and line broadening. Pharmacokinetic behaviors of orally dosed TL formulations were evaluated in rats using UPLC/ESI-MS. After oral administration of pMB-SD/TL (10mg TL/kg) in rats, enhanced TL exposure was observed with increases of Cmax and AUC by 125- and 52-fold, respectively, compared with those of crystalline TL. From these findings, pMB-based SD formulation approach might be an efficacious approach for enhancing the therapeutic potential of TL.

Self-micellizing solid dispersion of cyclosporine A with improved dissolution and oral bioavailability.

The present study aimed to develop a self-micellizing solid dispersion (SMSD) of cyclosporine A (CsA) using an amphiphilic block copolymer, poly[MPC-co-BMA], to improve the biopharmaceutical properties of CsA. The cytotoxicity of poly[MPC-co-BMA] was assessed in rat intestinal IEC-6 cells, and the pMB was less cytotoxic than polysorbate 80, a non-ionic surfactant with a wide safety margin. SMSD/CsA was prepared using a wet-milling system, and its physicochemical properties were characterized in terms of morphology, crystallinity, dissolution, particle size distribution, and stability. The SMSD/CsA exhibited immediate formation of fine micelles with a mean diameter of ca. 180 nm when introduced into aqueous media. There was marked improvement in the dissolution behavior of the SMSD/CsA compared with amorphous CsA. Even after storage at 40°C/75% relative humidity, the dissolution behavior of aged SMSD/CsA seemed to be almost identical to that of its freshly prepared equivalent, and CsA in aged SMSD/CsA was still in amorphous form. After oral administration of SMSD/CsA (10 mg CsA/kg) in rats, enhanced CsA exposure was observed with increases of Cmax and BA by ca. 11- and 42-fold, respectively, compared with those of amorphous CsA. The poly[MPC-co-BMA]-based SMSD formulation system might be an efficacious dosage option for CsA to achieve improvements in oral bioavailability.

Inhalable sustained-release formulation of long-acting vasoactive intestinal peptide derivative alleviates acute airway inflammation

The present study was undertaken to develop a respirable sustained-release powder (RP) formulation of long-acting VIP derivative, [Arg(15, 20, 21), Leu(17)]-VIP-GRR (IK312532), using PLGA nanospheres (NS) with the aim of improving the duration of action. NS formulation of IK312532 (IK312532/NS) was prepared by an emulsion solvent diffusion method in oil, and a mixture of the IK312532/NS and erythritol was jet-milled and mixed with lactose carrier to obtain the IK312532/NS-RP. Physicochemical properties were characterized focusing on appearance, particle size, and drug release, and in vivo pharmacological effects were assessed in antigen-sensitized rats. The IK312532/NS with a diameter of 140 nm showed a biphasic release pattern in distilled water with ca. 20% initial burst for 30 min and a sustained slow release up to ca. 55% for 24h. Laser diffraction analysis demonstrated that IK312532/NS-RP had fine dispersibility and suitable particle size for inhalation. In antigen-sensitized rats, insufflated IK312532/NS-RP (10 μg of IK312532/rat) could suppress increases of granulocyte recruitment and myeloperoxidase in pulmonary tissue for up to 24h after antigen challenge, although IK312532-RP at the same dose was less effective with limited duration of action. From these findings, newly prepared IK312532/NS-RP might be of clinical importance in improving duration of action and medication compliance for treatment of airway inflammatory diseases.

Formulation design and in vivo evaluation of dry powder inhalation system of new vasoactive intestinal peptide derivative ([R15, 20, 21, L17, A24,25, des-N28]-VIP-GRR) in experimental asthma/COPD model rats

Vasoactive intestinal peptide (VIP) has been considered as a promising drug candidate for asthma and COPD because of its potent immunomodulating and anti-inflammatory activities. Recently, our group developed a new VIP derivative, [R(15, 20, 21), L(17), A(24,25), des-N(28)]-VIP-GRR (IK312548), with improved chemical and metabolic stability. In the present study, a dry powder inhaler system of IK312548 was designed for inhalation therapy with minimal systemic side effects, the physicochemical properties of which were also evaluated with a focus on morphology, particle size distribution, inhalation performance, and peptide stability. Laser diffraction and cascade impactor analysis suggested high dispersion and deposition in the respiratory organs with a fine particle fraction of 31.2%. According to UPLC/ESI-MS and circular dichroic spectral analyses, no significant changes in the purity and structure of VIP derivative were observed during preparation of respirable formulation. Anti-inflammatory properties of IK312548 respirable powder (RP) were characterized in antigen-sensitized asthma/COPD-model rats. There were marked inflammatory cells infiltrated into the lung tissues of experimental asthma/COPD-model rats; however, intratracheal administration of IK312548-RP led to significant reductions of recruited inflammatory cells in lung tissues and BALF by 72 and 78%, respectively. Thus, respirable powder formulation of IK312548 might be a promising medication for asthma, COPD, and other airway inflammatory diseases.

Development of cyclosporine A-loaded dry-emulsion formulation using highly purified glycerol monooleate for safe inhalation therapy.

The main objective of this study was to improve the safety and oxidative stability of glycerol monooleate (GMO)-based dry-emulsion (DE) formulation containing cyclosporine A (CsA) for inhalation therapy. GMO or highly purified GMO (hpGMO) was used as surfactant for the DE formulations (GMO/DE or hpGMO/DE), the toxicological and physicochemical properties of which were characterized with a focus on oxidative stability, in vitro/in vivo toxicity, and dissolution property. Incubation of GMO at oxidation accelerating conditions for 10 days at 60°C resulted in the formation of lipid peroxides as evidenced by increased malondialdehyde (111 μmol/mg); however, hpGMO samples exhibited increase of only 20.7 μmol/mg in malondialdehyde level. No significant acute cytotoxicity was observed in rat alveolar L2 cells exposed to hpGMO (0.28mM), and intratracheal administration of hpGMO powder in rats did not cause an increase of the plasma LDH level. The hpGMO/DE exhibited marked improvement in dissolution behavior of CsA, and stable fine micelles with a mean diameter of 320 nm were formed when suspended in water. A respirable powder formulation of hpGMO/DE (hpGMO/DE-RP) was newly prepared, and its in vitro inhalation property and in vivo efficacy were also evaluated. The hpGMO/DE-RP exhibited high dispersibility in laser diffraction analysis and significantly improved potency to attenuate recruitment of inflammatory cells into airway and thickening of airway wall in an animal model. Thus, the strategic use of hpGMO would improve oxidative stability and local toxicity compared with a GMO-based DE formulation, and its application to RP formulation could be a promising approach for effective inhalation therapy.

Improved oral absorption profile of itraconazole in hypochlorhydria by self-micellizing solid dispersion approach

&NA; The present study was aimed to evaluate the applicability of a self‐micellizing solid dispersion (SMSD) system of itraconazole (ITZ) with the use of Soluplus® to achieve improved dissolution and stable oral absorption of ITZ under hypochlorhydric conditions. The SMSD of ITZ (SMSD/ITZ) was prepared by the freeze‐drying method. Physicochemical properties of SMSD/ITZ were assessed in terms of morphology, crystallinity, particle size, thermal behavior, dissolution profile, and stability. The pharmacokinetic profile of SMSD/ITZ was evaluated in both normal rats and omeprazole‐treated rats as a hypochlorhydric model. From the crystallinity assessment, ITZ in SMSD/ITZ might exist in an amorphous state. The dissolution behavior of SMSD/ITZ was markedly improved under both acidic and neutral conditions through the formation of nano‐micelles with a diameter of 127 nm. The degradation of ITZ in SMSD/ITZ was negligible after storage under accelerated conditions at 40 °C or 40 °C/75%RH for 4 weeks. Under light exposure, ca. 33% of ITZ in SMSD/ITZ was degraded, suggesting the need for protection from light. Although the oral absorption of crystalline ITZ was negligible, SMSD/ITZ showed an improved pharmacokinetic profile in normal rats, with an absolute bioavailability (BA) of 2.9%, and even 6.3% in the hypochlorhydric model. From these findings, SMSD technology could be beneficial for improving the absorption profiles of weak basic drugs, even in hypochlorhydric patients. Graphical abstract Figure. No caption available.

Strategic application of self-micellizing solid dispersion technology to respirable powder formulation of tranilast for improved therapeutic potential.

PURPOSE The present study aimed to develop an inhalable self-micellizing solid dispersion of tranilast (SMSD/TL) using poly[MPC-co-BMA] to improve the therapeutic potential and safety. METHODS The safety of poly[MPC-co-BMA] in lungs was assessed using rat lung epithelium-derived L2 cells. SMSD/TL and respirable powder of SMSD/TL (SMSD/TL-RP) were prepared using a wet milling system and jet mill, respectively. The physicochemical properties of TL formulations were characterized in terms of dissolution, morphology, and particle size. Pharmacological and pharmacokinetic studies were also conducted on inhaled SMSD/TL-RP. RESULTS The lactate dehydrogenase level from L2 cells treated with poly[MPC-co-BMA] was lower than that with polysorbate 80, a positive control. SMSD/TL showed enhanced dissolution behavior of TL. The jet milled SMSD/TL particles easily separated from the lactose carrier, and the particle size was suitable for inhalation. Compared with RP of TL, inhaled SMSD/TL-RP (100 μg-TL/rat) could more strongly suppress the inflammatory responses in antigen-sensitized rats. The TL level in plasma after intratracheal administration of SMSD/TL-RP at a pharmacological effective dose (100 μg-TL/rat) was ca. 4.2-fold lower than that after oral administration of TL solution at a clinical dose (1.67 mg/kg). CONCLUSION SMSD/TL-RP might be an attractive dosage form to improve the anti-inflammatory effects and safety of TL.

Respirable Dry Powder Formulation of Bleomycin for Developing a Pulmonary Fibrosis Animal Model

The main purpose of the present study was to develop a respirable powder (RP) formulation of bleomycin (BLM) as a research tool for developing a pulmonary fibrosis animal model. The BLM-RP was prepared with a jet-milling system, the physicochemical properties of which were characterized focusing on morphology, stability, particle size distribution, and inhalation performance. Under an accelerated condition, the BLM-RP was superior to BLM solution in terms of its stability. Cascade impactor analyses demonstrated high inhalation performance with emitted dose and fine particle fraction of approximately 99% and 46%, respectively. Intratracheal administration of the BLM-RP (3 mg BLM/kg) in rats led to significant increases in collagen production and recruitment of inflammatory cells in lung by approximately 1.5- and 29-fold, respectively. The collagen overexpression was consistent with the results from picrosirius red staining of lung tissues in the rats treated with BLM-RP. Inhaled tranilast (TL; 100 μg/rat), an antifibrotic agent, could ameliorate inflammatory/fibrotic responses with reductions of recruited inflammatory cells and collagen content by 32% and 59%, respectively, validating the pulmonary fibrosis animal model. From these findings, the BLM-RP with improved stability could be a beneficial research tool for developing a pulmonary fibrosis model in drug discovery for antifibrotic drug candidates.

Physicochemical Stability Study on Cyclosporine A Loaded Dry-Emulsion Formulation with Enhanced Solubility

The main purpose of the present study was to evaluate the physicochemical stability of cyclosporine A (CsA)-loaded glycerol monooleate-based dry emulsion (DE). DE formulations containing 5-25% CsA (DE5-25) were stored at 25°C/60% relative humidity for 4 weeks, and freeze-dried solid dispersion formulations containing 5-30% CsA (FD5-30) were also prepared as reference formulations. Even after the storage, no significant changes were observed in the appearance of any formulations. In the dissolution study, both DE and FD exhibited marked enhancement of solubility and there was at least 2.0-fold improvement in the initial dissolution rate of DE formulations compared with that of FD formulations. After storage, DE5, DE15 and FD5 maintained relatively high solubility, with 10% reduction compared with the initial state. However, the solubility of DE25 gradually decreased during storage, as evidenced by 76% reduction of the dissolution amount. No significant changes were seen in DE5-25 using powder X-ray diffraction, although thermal analysis revealed moderate changes in crystallinity in DE25 after storage, possibly leading to the decreased dissolution. Furthermore, particle size distributions of micelles in DE5 and DE15 were almost unchanged after storage for 4 weeks. From these findings, it appears that the physicochemical stability of CsA-loaded DE might vary depending on the manufacturing method and that further optimization could improve physical properties and stability.