Yilmaz Capan

Influence of formulation parameters on the characteristics of poly(D, L-lactide-co-glycolide) microspheres containing poly(L-lysine) complexed plasmid DNA.

This study describes the influence of polymer type, surfactant type/concentration, and target drug loading on the particle size, plasmid DNA (pDNA) structure, drug loading efficiency, in vitro release, and protection from DNase I degradation of poly(D, L-lactide-co-glycolide) (PLGA) microspheres containing poly(L-lysine) (PLL) complexed pDNA. PLGA microspheres containing pDNA-PLL were prepared using the water-in-oil-in-water (w-o-w) technique with poly(vinyl alcohol) (PVA) and poly(vinyl pyrrolidone) (PVP) as surfactants in the external aqueous phase. A complex ratio of 1:0.33 (pDNA-PLL, w/w) enhanced the stability of pDNA during microsphere preparation. Higher pDNA-PLL loading efficiency (46.2%) and supercoiled structure (64.9%) of pDNA were obtained from hydrophobic PLGA (M(w) 31000) microspheres compared with hydrophilic PLGA or low-molecular-weight PLGA microspheres. The particle size decreased from 6.6 to 2.2 microm when the concentration of PVA was increased from 1 to 7%. At the same concentration of surfactant, PVA stabilized microspheres showed higher pDNA-PLL loading efficiency (46.2%) than PVP stabilized microspheres (24.1%). Encapsulated pDNA in PLGA microspheres was protected from enzymatic degradation and maintained in the supercoiled form. The pDNA-PLL microspheres showed in vitro release of 95.9 and 84.9% within 38 days from the low-molecular-weight PLGA and hydrophilic PLGA microspheres, respectively, compared to 54.2% release from the hydrophobic, higher-molecular-weight PLGA microspheres. The results suggest loading and release of pDNA-PLL complex can be influenced by surfactant concentration and polymer type.

A Nanomedicine Transports a Peptide Caspase-3 Inhibitor across the Blood–Brain Barrier and Provides Neuroprotection

Caspases play an important role as mediators of cell death in acute and chronic neurological disorders. Although peptide inhibitors of caspases provide neuroprotection, they have to be administered intracerebroventricularly because they cannot cross the blood–brain barrier (BBB). Herein, we present a nanocarrier system that can transfer chitosan nanospheres loaded with N-benzyloxycarbonyl-Asp(OMe)-Glu(OMe)-Val-Asp(OMe)-fluoromethyl ketone (Z-DEVD-FMK), a relatively specific caspase-3 inhibitor, across BBB. Caspase-3 was chosen as a pharmacological target because of its central role in cell death. Polyethylene glycol-coated nanospheres were conjugated to an anti-mouse transferrin receptor monoclonal antibody (TfRMAb) that selectively recognizes the TfR type 1 on the cerebral vasculature. We demonstrate with intravital microscopy that this nanomedicine is rapidly transported across the BBB without being measurably taken up by liver and spleen. Pre- or post-treatment (2 h) with intravenously injected Z-DEVD-FMK-loaded nanospheres dose dependently decreased the infarct volume, neurological deficit, and ischemia-induced caspase-3 activity in mice subjected to 2 h of MCA occlusion and 24 h of reperfusion, suggesting that they released an amount of peptide sufficient to inhibit caspase activity. Similarly, nanospheres inhibited physiological caspase-3 activity during development in the neonatal mouse cerebellum on postnatal day 17 after closure of the BBB. Neither nanospheres functionalized with TfRMAb but not loaded with Z-DEVD-FMK nor nanospheres lacking TfRMAb but loaded with Z-DEVD-FMK had any effect on either paradigm, suggesting that inhibition of caspase activity and subsequent neuroprotection were due to efficient penetration of the peptide into brain. Thus, chitosan nanospheres open new and exciting opportunities for brain delivery of biologically active peptides that are useful for the treatment of CNS disorders.

Preparation and characterization of poly(D,L-lactide-co-glycolide) microspheres for controlled release of human growth hormone

The purpose of this research was to assess the physicochemical properties of a controlled release formulation of recombinant human growth hormone (rHGH) encapsulated in poly(D,L-lactide-co-glycolide) (PLGA) composite microspheres. rHGH was loaded in poly(acryloyl hydroxyethyl) starch (acHES) microparticles, and then the protein-containing microparticles were encapsulated in the PLGA matrix by a solvent extraction/evaporation method. rHGH-loaded PLGA microspheres were also prepared using mannitol without the starch hydrogel microparticle microspheres for comparison. The detection of secondary structure changes in protein was investigated by using a Fourier Transfer Infrared (FTIR) technique. The composite microspheres were spherical in shape (44.6±2.47 μm), and the PLGA-mannitol microspheres were 39.7±2.50 μm. Drug-loading efficiency varied from 93.2% to 104%. The composite microspheres showed higher overall drug release than the PLGA/mannitol microspheres. FTIR analyses indicated good stability and structural integrity of HGH localized in the microspheres. The PLGA-acHES composite microsphere system could be useful for the controlled delivery of protein drugs.

Design and evaluation of sustained-release and buccal adhesive propranolol hydrochloride tablets

Abstract The release of propranolol hydrochloride incorporated into sustained-release and buccal adhesive tablets was studied in vitro. The formulation containing 20% hydroxypropyl methylcellulose (HPMC) yielded good sustained-release matrix tablets. Buccal adhesive controlled-release tablets were prepared by compression of HPMC with polycarbophil (PAA), which served as the bioactive adhesive compound. The release behaviour of buccal adhesive tablets was found to be non-Fickian. The adhesion force was significantly affected by the mixing ratio of HPMC and PAA in the tablet and the weakest adhesion force was observed at the ratio of 1:1 (HPMC:PAA). Interpolymer complex formation was confirmed between HPMC and PAA in acidic medium by turbidity, viscosity and FT-IR measurements. The kinetics of sustained-release and buccal adhesive tablets of propranolol were examined in nine healthy volunteers. Conventional propranolol (Dideral®) was also studied for comparison purposes. As compared to conventional propranolol (40 mg), a single dose of 20% HPMC (160 mg) produced a smoother plasma level profile, with lower and delayed peak times. Dose corrected AUC0–8 values were greater after Dideral® than after 20% HPMC (168.7 ± 80.3 vs 97.3 ± 36.1 ng h ml−1 p 0.05) in the AUC0–4 values between 20% HPMC and buccal adhesive tablets.

Formulation and in vitro-in vivo evaluation of buccoadhesive morphine sulfate tablets

Buccoadhesive controlled-release systems for the delivery of morphine sulfate were prepared by compression of hydroxypropyl methylcellulose (HPMC) with carbomer (CP), which served as the bioactive adhesive compound. The release behavior of systems containing 30 mg of morphine sulfate and various amounts of the two polymers was found to be non-Fickian. The adhesion force was significantly affected by the mixing ratio of HPMC and CP in the tablet, and the weakest adhesion force was observed at a ratio of 1:1 (HPMC:CP). Interpolymer complex formation was confirmed between HPMC and CP in acidic medium by turbidity, viscosity, and FT-IR measurements. The amount absorbed (percentage of the drug loaded) of the controlled-release buccoadhesive tablets in six healthy volunteers and was 30 ± 5%.

Squalenoyl adenosine nanoparticles provide neuroprotection after stroke and spinal cord injury.

There is an urgent need to develop new therapeutic approaches for the treatment of severe neurological trauma, such as stroke and spinal cord injuries. However, many drugs with potential neuropharmacological activity, like adenosine, are inefficient upon systemic administration because of their fast metabolisation and rapid clearance from the bloodstream. Here, we show that the conjugation of adenosine to the lipid squalene and the subsequent formation of nanoassemblies allow a prolonged circulation of this nucleoside, to provide neuroprotection in mouse stroke and rat spinal cord injury models. The animals receiving systemic administration of squalenoyl adenosine nanoassemblies showed a significant improvement of their neurologic deficit score in the case of cerebral ischaemia, and an early motor recovery of the hindlimbs in the case of spinal cord injury. Moreover, in vitro and in vivo studies demonstrated that the nanoassemblies were able to extend adenosine circulation and its interaction with the neurovascular unit. This paper shows, for the first time, that a hydrophilic and rapidly metabolised molecule like adenosine may become pharmacologically efficient owing to a single conjugation with the lipid squalene.

Enhancement of transbuccal permeation of morphine sulfate by sodium glycodeoxycholate in vitro

Abstract In this study, enhancement of transbuccal permeation of morphine sulfate was studied in the presence of sodium glycodeoxycholate (GDC). The permeability of the bovine buccal mucosa to morphine sulfate was determined in vitro in the absence and presence of GDC at 10 mM and 100 mM concentrations. Light and electron microscopy studies were performed to determine the histological and ultrastructural changes resulting from transepithelial permeation enhancement. In addition, infrared spectroscopy (IR) was used to investigate the interaction of GDC with the epithelial lipids of bovine buccal mucosa. The permeation of morphine sulfate across the bovine buccal epithelium was enhanced in the presence of 100 mM GDC by a factor of five whereas at lower concentrations, no significant enhancement was obtained. After 4 h treatment with 100 mM (5% w/v) GDC, significant changes were observed in the epithelium at histological and ultrastructural levels which can be defined as formation of vacuoles, swelling of the cells and a possible increase in intercellular space. Furthermore, by means of IR spectroscopy, it was possible to show the effect of GDC on bovine buccal epithelial lipid domains which was in good correlation with the permeation results. In the light of the results obtained by permeation, histological and IR spectroscopy studies, it is concluded that GDC at 100 mM concentration significantly enhances the permeation of MS across the buccal epithelium and the mechanism of this action appears to involve an interaction with the epithelial lipids.

Development of a peptide-containing chewing gum as a sustained release antiplaque antimicrobial delivery system.

The objective of this study was to characterize the stability of KSL-W, an antimicrobial decapeptide shown to inhibit the growth of oral bacterial strains associated with caries development and plaque formation, and its potential as an antiplaque agent in a chewing gum formulation. KSL-W formulations with or without the commercial antibacterial agent cetylpyridinium chloride (CPC) were prepared. The release of KSL-W from the gums was assessed in vitro using a chewing gum apparatus and in vivo by a chew-out method. A reverse-phase high-performance liquid chromatography method was developed for assaying KSL-W. Raw material stability and temperature and pH effects on the stability of KSL-W solutions and interactions of KSL-W with tooth-like material, hydroxyapatite discs, were investigated.KSL-W was most stable in acidic aqueous solutions and underwent rapid hydrolysis in base. It was stable to enzymatic degradation in human saliva for 1 hour but was degraded by pancreatic serine proteases. KSL-W readily adsorbed to hydroxyapatite, suggesting that it will also adsorb to the teeth when delivered to the oral cavity. The inclusion of CPC caused a large increase in the rate and extent of KSL-W released from the gums. The gum formulations displayed promising in vitro/ in vivo release profiles, wherein as much as 90% of the KSL-W was released in a sustained manner within 30 minutes in vivo. These results suggest that KSL-W possesses the stability, adsorption, and release characteristics necessary for local delivery to the oral cavity in a chewing gum formulation, there-by serving as a novel antiplaque agent.

In vitro studies on enhancing effect of sodium glycocholate on transbuccal permeation of morphine hydrochloride

During the perioperative period, gastric emptying rate and first-pass metabolism limit the use of peroral morphine. Buccal mucosa appears to be a potential site for delivery of morphine as it provides direct entry into the system circulation thereby avoiding the hepatic first-pass effect. However, the low permeability of the buccal epithelium results in a low flux of the drug. The use of a penetration enhancer is required to improve the bioavailability of the drug via buccal route. In this study, the enhancing effect of sodium glycocholate (GC) used at 10 mM and 100 mM concentrations on permeation of morphine hydrochloride (MPH) across the porcine buccal mucosa was studied in vitro. Furthermore, in conjunction with its permeation, accumulation of GC in the tissue with time was also studied in order to elucidate the relationship between GC and enhanced mucosal permeation of the drug. Franz diffusion cells were used in the experiments. Permeation of MPH was increased in the presence of 100 mM GC with an enhancement factor of 9.3 whereas no enhancement was obtained with 10 mM GC. The calculated permeability coefficient for MPH in the presence of 100 mM GC was 2.35 x 10(-5) cm/s. Accumulation of GC at 100 mM in the tissue appears to be more significant at 100 mM concentration which correlated well with the increased permeation of the drug. GC was diffused through the buccal epithelium significantly at 100 mM concentration. Interaction of GC with the tissue appears to be more significant at 100 mM concentration compared to 10 mM concentration, thus resulting in a significant enhancing effect.

Systemically Administered Brain-Targeted Nanoparticles Transport Peptides across the Blood—Brain Barrier and Provide Neuroprotection

Although growth factors and anti-apoptotic peptides have been shown to be neuroprotective in stroke models, translation of these experimental findings to clinic is hampered by limited penetration of peptides to the brain. Here, we show that a large peptide like the basic fibroblast growth factor (bFGF) and a small peptide inhibitor of caspase-3 (z-DEVD-FMK) can effectively be transported to the brain after systemic administration by incorporating these peptides to brain-targeted nanoparticles (NPs). Chitosan NPs were loaded with peptides and then functionalized by conjugating with antibodies directed against the transferrin receptor-1 on brain endothelia to induce receptor-mediated transcytosis across the blood—brain barrier (BBB). Pre-ischemic systemic administration of bFGF- or z-DEVD-FMK-loaded NPs significantly decreased the infarct volume after 2-hour middle cerebral artery occlusion and 22-hour reperfusion in mice. Co-administration of bFGF- or z-DEVD-FMK-loaded NPs reduced the infarct volume further and provided a 3-hour therapeutic window. bFGF-loaded NPs were histologically detected in the brain parenchyma and also restored ischemia-induced Akt dephosphorylation. The neuroprotection was not observed when receptor-mediated transcytosis was inhibited with imatinib or when bFGF-loaded NPs were not conjugated with the targeting antibody, which enables them to cross the BBB. Nanoparticles targeted to brain are promising drug carriers to transport large as well as small BBB-impermeable therapeutics for neuroprotection against stroke.