International Journal of Pharmaceutics (v.277, #1-2)

(EDI BOARD) (iii).

Introductory remarks by A.Atilla Hinçal (1).

Drug efflux by intestinal P-glycoprotein (P-gp) is known to decrease the bioavailability of many CYP3A4 substrates. We have demonstrated that the interplay between P-gp and CYP3A4 at the apical intestinal membrane can increase the opportunity for drug metabolism by determining bidirectional extraction ratios across CYP3A4-transfected Caco-2 cells for two dual P-gp/CYP3A4 substrates, K77 (an experimental cysteine protease inhibitor) and sirolimus, as well as two negative control, CYP3A4 only substrates, midazolam and felodipine. Studies were carried out under control conditions, with a P-gp inhibitor (GG918) and with a dual inhibitor (cyclosporine). Measurement of intracellular concentration changes is an important component in calculating the extraction ratios. We hypothesize that the inverse orientation of P-gp and CYP3A4 in the liver will result in an opposite interactive effect in that organ. In vivo rat intestinal perfusion studies with K77 and rat liver perfusion studies with tacrolimus under control conditions and with inhibitors of CYP3A4 (troleandomycin), P-gp (GG918) and both CYP3A4/P-gp (cyclosporine) lend support to our hypotheses. These results serve as a template for predicting enzyme-transporter (both absorptive and efflux) interactions in the intestine and the liver.
Keywords: P-glycoprotein; CYP3A4; Transport; Metabolism; Caco-2 cells; Tacrolimus; Sirolimus; Midazolam; Felodipine; Sex differences;

The primary goal of bioadhesive controlled drug delivery is to localize a delivery device within the body to enhance the drug absorption process in a site-specific manner. An important contributor to good adhesion is the presence of molecular adhesion promoters such as polymer-tethered structure (e.g., poly(ethylene glycol) chains grafted to crosslinked networks) or even linear chains which are free to diffuse across the gel/gel interface. Recently, we have developed a very promising class of carriers for drug and especially protein delivery. Copolymer networks of poly(methacrylic acid) grafted with poly(ethylene glycol) exhibit reversible, pH-dependent swelling behavior due to the formation of interpolymer complexes between protonated pendant acid groups and the etheric groups on the graft chains. Gels containing equimolar amounts of MAA/EG exhibited the lowest degree of swelling at low pH increased complexation. The average network mesh size or correlation length was dramatically affected by the pH of the swelling solution. The in vitro release of insulin from P(MAA-g-EG) gels containing PEG grafts of molecular weight 1000 indicates a significant release of insulin as the gel decomplexes and insulin is freed through the structure. The results of additional in vitro studies have shown that insulin release rates can be controlled by appropriate adjustment of the structure of the gels.
Keywords: Hydrogels; Protein; Insulin; Mucoadhesion; Poly(ethylene glycol);

Keywords: Magnetic targeting; Liver tumor; Magnetic microspheres; Cell separation; Contrast agent; Chemotherapy;

In order to achieve a sustained pharmacological activity of oligonucleotides (ODNs) and avoid repeated administrations, we have developed a new concept of delivery system that combine sustained release and improved intracellular penetration. These systems are designed for the intravitreal delivery of antisense ODNs. The first concept consisted in using liposomes dispersed in a thermosensitive gel (poloxamer 407). After intravitreal administration in a rabbit model, liposomes and liposomes-gel formulations provided, 1-day postinjection, significantly higher drug levels than the control solution of the oligothymidilate pdT16. In addition, there was no significant difference in the amounts of pdT16 found in the vitreous humor between the liposomes and liposomes-gel. Nevertheless, because of their better stability in the absence of poloxamer, liposomes alone allowed to a larger extent to control the delivery of ODNs as compared to liposome-gel formulations since 37% of the ODNs were still found in the vitreous 15 days after administration. In addition, the ODNs found in the vitreous humor were protected against degradation by their encapsulation within liposomes. The second approach consisted in designing microspheres allowing to release in a controlled fashion pdT16. The ODN was encapsulated within poly(lactide-co-glycolide) microspheres alone or associated with polyethylenimine (PEI) at different nitrogen/phosphate (N/P) ratios. The introduction of PEI in the internal aqueous phase resulted in a strong increase of the ODN encapsulation efficiency. PEI affected microsphere morphology inducing the formation of very porous particles yielding to an accelerated release of pdT16. Porosity and controlled delivery was prevented by introducing sodium chloride in the external preparation medium. When incubated with HeLa cells, microspheres encapsulating pdT16/PEI complexes allowed an improvement of the intracellular penetration of the released ODN. Both liposomes and microspheres are suitable for local delivery of ODNs.
Keywords: Oligonucleotide; Thermosensitive gel; Microparticles; Polyethylenimine;

Active and intelligent inhaler device development by Mike Tobyn; John N Staniforth; David Morton; Quentin Harmer; Mike E Newton (31-37).
The dry powder inhaler, which has traditionally relied on the patient’s inspiratory force to deaggregate and deliver the active agent to the target region of the lung, has been a successful delivery device for the provision of locally active agents for the treatment of conditions such as asthma and chronic obstructive pulmonary disease (COPD). However, such devices can suffer from poor delivery characteristics and/or poor reproducibility. More recently, drugs for systemic delivery and more high value compounds have been put into DPI devices. Regulatory, dosing, manufacturing and economic concerns have demanded that a more efficient and reproducible performance is achieved by these devices. Recently strategies have been put in place to produce a more efficient DPI device/formulation combination. Using one novel device as an example the paper will examine which features are important in such a device and some of the strategies required to implement these features.All of these technological advances are invisible, and may be irrelevant, to the patient. However, their inability to use an inhaler device properly has significant implications for their therapy. Use of active device mechanisms, which reduce the dependence on patient inspiratory flow, and sensible industrial design, which give the patient the right clues to use, are important determinants of performance here.
Keywords: Dry powder inhaler; Formulation; Inspiratory force; Delivery device;

This review is focused on the macromolecular drug carrier systems by the effect of enhanced permeability and retention (EPR) and the mechanism of receptor-mediated endocytosis (RME). The effect of EPR is thought to be useful for the targeting of the macromolecular drugs to the tumor tissues on a vasculolymphatic level. The RME reveals the selective recognition, high affinity binding, and immediate internalization for the ligand on a cellular level. In the receptor, recognizing transferrin, a level of expression on the tumor cells is higher than that on the normal cells. We have used serum albumin and transferrin as drug carriers to deliver mitomycin C (MMC) to the tumor tissues and into the tumor cells. The properties of the conjugates of MMC to serum albumin and transferrin were examined in vitro and in vivo. We concluded that MMC could be delivered to the tumor tissue and cells by the use of albumin and transferrin as drug carriers.
Keywords: Drug delivery system; Tumor targeting; Mitomycin C; Macromolecular conjugate; Serum albumin; Transferrin; Enhanced permeability and retention; Receptor-mediated endocytosis; Intracellular disposition; Antitumor activity;

Polymeric anticancer drugs with pH-controlled activation by K Ulbrich; T Etrych; P Chytil; M Pechar; M Jelinkova; B Rihova (63-72).
The paper is dealing with the synthesis and properties of new, nontargeted or antibody-targeted pH-sensitive polymer–doxorubicin (DOX) conjugates designed as anticancer drugs facilitating site-specific therapy. These conjugates are stable and inactive during transport in the body but activate inside target cells as a result of pH changes outside and inside the cells. Cytotoxicity of the conjugates depends on the detailed structure of the polymer and of the spacer between the drug and polymer carrier. In both protective and therapeutic regimes of drug administration, the in vivo antitumor activity of the pH-sensitive conjugates containing DOX was significantly enhanced (T-cell lymphoma EL 4, C57BL/16 mice) in comparison with the free DOX or classic PK1, the PHPMA–DOX conjugate clinically tested at present.
Keywords: HPMA copolymers; Poly(ethylene glycol); Doxorubicin; Drug targeting; Cytotoxicity; Antitumor activity;

The role of the newly discovered cysteine protease, cathepsin K, in osteoporosis and rheumatoid arthritis is reviewed. The current development of cathepsin K inhibitors and their targeted delivery using synthetic polymer carriers are discussed. Future challenges and possible strategies to improve these delivery systems are addressed.
Keywords: Cathepsin K; Inhibitor–polymer conjugates; Semitelechelic HPMA polymer; mPEG; Antiresorptive; Resorption lacuna;

Preparation and characterization of water-soluble pH-sensitive nanocarriers for drug delivery by M.-H. Dufresne; D.Le Garrec; V. Sant; J.-C. Leroux; M. Ranger (81-90).
pH-sensitive drug delivery systems can be engineered to release their contents or change their physicochemical properties in response to variations in the acidity of the surroundings. The present work describes the preparation and characterization of novel polymeric micelles (PM) composed of amphiphilic pH-responsive poly(N-isopropylacrylamide) (PNIPAM) or poly(alkyl(meth)acrylate) derivatives. On one hand, acidification of the PNIPAM copolymers induces a coil-to-globule transition that can be exploited to destabilize the intracellular vesicle membranes. In this work, PNIPAM-based PM were loaded with either doxorubicin or aluminium chloride phthalocyanine and their cytotoxicity was assessed in murine tumoral models. On the other hand, poly(alkyl(meth)acrylate) copolymers can be designed to interact with either hydrophobic drugs or polyions and release their cargo upon an increase in pH.
Keywords: Block copolymers; pH-controlled release; Polymeric micelles; Anticancer agents; Poorly soluble drugs;

Calcium phosphate-PEG-insulin-casein (CAPIC) particles as oral delivery systems for insulin by T Morçöl; P Nagappan; L Nerenbaum; A Mitchell; S.J.D Bell (91-97).
An oral delivery system for insulin was developed and functional activity was tested in a non-obese diabetic (NOD) mice model. Calcium phosphate particles containing insulin was synthesized in the presence of PEG-3350 and modified by aggregating the particles with caseins to obtain the calcium phosphate-PEG-insulin-casein (CAPIC) oral insulin delivery system. Single doses of CAPIC formulation were tested in NOD mice under fasting or fed conditions to evaluate the glycemic activity. The blood glucose levels were monitored every 1–2 h for 12 h following the treatments using an ACCU CHECK blood glucose monitoring system. Orally administered and subcutaneously injected free insulin solution served as controls in the study. Based on the results obtained we propose that: (1) the biological activity of insulin is preserved in CAPIC formulation; (2) insulin in CAPIC formulations, but not the free insulin, displays a prolonged hypoglycemic effect after oral administration to diabetic mice; (3) CAPIC formulation protects insulin from degradation while passing through the acidic environment of the GI track until it is released in the less acidic environment of the intestines where it can be absorbed in its biologically active form; (4) CAPIC formulation represents a new and unique oral delivery system for insulin and other macromolecules.
Keywords: Oral drug delivery; Insulin; Diabetes; Oral insulin formulation; Sustained release drug delivery; Calcium phosphate particles;

Hydrogels are based on hydrophilic polymers, which are crosslinked to prevent dissolution in water. Because hydrogels can contain large amounts of water, they are interesting devices for the delivery of proteins. In this contribution a biodegradable dextran hydrogel is described which is based on physical interactions and is particularly suitable for the controlled delivery of pharmaceutically active proteins. The unique feature of our system is that the preparation of the hydrogels takes place in an all-aqueous solution, by which the use of organic solvents is avoided. Furthermore, chemical crosslinking agents are not needed to create the hydrogels, since crosslinking is established physically by stereocomplex formation between enantiomeric oligomeric lactic acid chains. The hydrogel is simply obtained after mixing aqueous solutions of dextran(l)-lactate and dextran(d)-lactate. In this contribution, the formation of the hydrogels as well as their protein release properties and degradation behavior are discussed.
Keywords: Biodegradable dextran; Hydrogels; Cross-linking by stereocomplex; Proteins;

Intelligent polymers, also referred as “stimuli-responsive polymers” undergo strong property changes (in shape, surface characteristics, solubility, etc.) when only small changes in their environment (changes in temperature, pH, ionic strength light, electrical and magnetic field, etc.). They have been used in several novel applications, drug delivery systems, tissue engineering scaffolds, bioseparation, biomimetic actuators, etc. The most popular member of these type of polymers is poly(N-isopropylacrylamide) (poly(NIPA)) which exhibits temperature-sensitive character, in which the polymer chains change from water-soluble coils to water-insoluble globules in aqueous solution as temperature increases above the lower critical solution temperature (LCST) of the polymer. Copolymerization of NIPA with acrylic acid (AAc) allows the synthesis of both pH and temperature-responsive copolymers. This paper summarizes some of our related studies in which NIPA and its copolymers were synthesized and used as intelligent carriers in diverse applications.
Keywords: Intelligent polymers; Poly(NIPA) and copolymers; Conjugates with amino acids; Gene therapy; Nonviral vectors;

Polymeric prodrugs by K Hoste; K De Winne; E Schacht (119-131).
In 1975 Prof. H. Ringsdorf proposed a model for rational design of polymeric prodrugs [J. Polym. Sci. Symp. 51 (1975) 135]. The model has been the most important basis for research in the field, since it was the first model that took into account both the chemical and biological aspects needed for the design of polymeric prodrugs. This paper deals with the most important properties that were discovered by designing polymeric prodrugs: prolongation of action of the drug, controlled release of the drug, passive tumor accumulation by the EPR-effect and alteration of body distribution and cell uptake. Over the years, other objectives have been formulated and other properties of polymer–drug conjugates were discovered. One recent example, the immunoprotective ability of polymeric prodrugs, is described in more detail in this paper.
Keywords: Prodrug; Controlled drug release; Conjugate; Tumor-associated enzymes; Targeting;

Most polymeric implants are biodegraded by one of two common chemical degradation mechanisms: (i) hydrolysis and (ii) oxidation. The chemical structure is among the most important factors which affect the biodegradation of polymeric implants. Hydrolytic biodegradations are often accompanied by substantial decrease of pH, whilst oxidative biodegradation processes are usually very slow due to consumption of stoichiometric amounts of oxidising agents. A dramatic acceleration of the biodegradation can be expected, if the biodegradation can be initiated by catalytic amounts of oxidation agents. Poly(ethylene carbonate) (PEC) and poly(trimethylene carbonate) (PTMC) are presumably biodegraded by such catalytic oxidation processes. Their biodegradation shows all the characteristics of surface erosion. Poly(ethylene carbonate) is utilised as a surface eroding biocompatible polymer for controlled delivery of peptide and protein drugs.
Keywords: Chemical structure; Biodegradation; Hydrolysis; Oxidation; Surface erosion; Poly(ethylene carbonate) (PEC); Superoxide anion radical; Controlled release; Functional polycarbonates;

Pharmacokinetic and pharmacodynamic aspects of gastroretentive dosage forms by Amnon Hoffman; David Stepensky; Eran Lavy; Sara Eyal; Eytan Klausner; Michael Friedman (141-153).
Controlled release gastroretentive dosage forms (CR-GRDF) enable prolonged and continuous input of the drug to the upper parts of the gastrointestinal (GI) tract and improve the bioavailability of medications that are characterized by a narrow absorption window. CR-GRDF provide a means to utilize all the pharmacokinetic (PK) and pharmacodynamic (PD) advantages of controlled release dosage forms for such drugs. Thus, CR-GRDF may improve therapy with clinically used medications, as well as enable oral administration of drugs, or drug candidates, that hitherto had to be infused parenterally. This manuscript discusses the complexity of the PK and PD factors that influence the treatment benefits of CR-GRDF and summarizes the results of our recent in vivo investigations in animal models (rats and dogs) and in human subjects. We found that a CR-GRDF formulation was superior to the other modes of administration for levodopa and riboflavin, but not for metformin. The PK and PD rationales of GRDFs for the studied drugs are presented and discussed. We conclude that due to the complexity of the PK and PD factors for a certain drug, the rationale for continuous administration obtained by CR-GRDF should be assessed and established in vivo.
Keywords: Gastroretentive dosage forms; Pharmacokinetics; Pharmacodynamics; Controlled release; Bioavailability; Metformin; Levodopa; Riboflavin;

A new modality of drug targeting to tumors that is under development in our lab is based on the drug encapsulation in polymeric micelles followed by a localized release at the tumor site triggered by focused ultrasound. The rationale behind this approach is that drug encapsulation in micelles decreases systemic concentration of drug and provides for a passive drug targeting to tumors via the enhanced penetration and retention (EPR) effect, thus, reducing unwanted drug interactions with healthy tissues. In addition, polymeric micelles sensitize multidrug resistant (MDR) cells to the action of drugs. Upon the accumulation of drug-loaded micelles at the tumor site, ultrasonic irradiation of the tumor is used to provide for the effective intracellular drug uptake. Ultrasound releases drug from micelles and enhances the intracellular uptake of both released and encapsulated drug. An important advantage of ultrasound is that it is noninvasive, can penetrate deep into the interior of the body, can be focused and carefully controlled. The results of the in vitro application of this technique for delivering anthracyclin drugs to ovarian carcinoma A2780 drug-sensitive and MDR cells are described.
Keywords: Ultrasound; Polymeric micelles; Pluronic; Doxorubicin; Ovarian carcinoma;

The healing effect of TGF-α on gastric ulcer induced by acetylsalicylic acid in rats by G Yetkin; N Çelebi; Ç Özer; B Gönül; C Özoğul (163-172).
The present study was designed to investigate the effects of microemulsion and aqueous solution containing transforming growth factor alpha (TGF-α) and/or aprotinin administered intragastrically (i.g.) on healing of acute gastric ulcers induced by acetylsalicylic acid (ASA). The microemulsion was prepared by modification of the microemulsion formulation described in our previous study. Acute gastric lesions were induced by the application of ASA (150 mg/kg in 1.5 ml of 0.2 N HCl i.g.). TGF-α in solution or microemulsion formulations were administered at a dose of 10 μg/kg per 24 h i.g. for 2 days. The effects of TGF-α on the healing was evaluated with the measurement of ulcer score, basal gastric acid secretion, total protein content of gastric fluid, gastric mucus level and histological analysis. The results indicated that the highest decrease in ulcer area was observed in group treated with microemulsion containing TGF-α plus aprotinin (TA-ME). TGF-α in microemulsion formulation was more effective than TGF-α in solution formulation in the increase of gastric mucus secretion, in the decrease of gastric acid secretions and ulcer scores. Histological evaluation of the gastric mucosa samples revealed that, best recovery was obtained in the TA-ME treated group.
Keywords: Gastric ulcer; Acetylsalicylic acid; Transforming growth factor alpha; Aprotinin; Microemulsion; Intragastric administration;

Development of a buccal bioadhesive nicotine tablet formulation for smoking cessation by G Ìkinci; S Şenel; C.G Wilson; M Şumnu (173-178).
Bioadhesive buccal tablet formulations for delivery of nicotine into the oral cavity were developed. Carbomer (Carbopol®974P NF) (CP) and alginic acid sodium salt (NaAlg) were used as bioadhesive polymers in combination with hydroxypropyl methylcellulose (HPMC) at different ratios. Magnesium carbonate was incorporated into the formulations as a pH increasing agent. In vitro release and bioadhesion studies were performed on the developed tablets. In the formulations containing CP:HPMC, the NHT released increased with the increasing HPMC concentration whereas a decrease was observed with increasing HPMC concentration in formulations containing NaAlg:HPMC. The bioadhesive properties of the tablets containing NaAlg:HPMC was not affected by the concentration of the NaAlg (P>0.05) but increased significantly with the increasing CP concentration (P<0.05). A decrease in pH of the dissolution medium to acidic values was avoided by incorporation of magnesium hydroxide into the formulations. The developed formulations released NHT for 8 h period, and remained intact except for the formulation containing CP:HPMC at 20:80 ratio.
Keywords: Nicotine hydrogen tartrate; Buccal tablets; Bioadhesion;

Notice board (179-181).