[Drug delivery systems: a real therapeutic advance]. / Les systèmes de délivrance des médicaments: un réel progrès pour la thérapeutique.

Established at the request of the Research Committee of the French National Academy of Pharmacy, this report on drug delivery systems (DDS) is a summary of information gathered by interviewing leaders in the pharmaceutical community and from the international literature. This report includes: a rapid recall of pharmaceutical formulations and changes over the last decades; a definition of DDS, indications on their evolution and a discussion on their contribution to drug administration; information on firms specialized in the elaboration of DDS, their interactions with the drug industry and the current and future market for DDS; a presentation of the potential offered by DDS for the drug industry; a discussion on technical, regulatory, and economic issues which could obstruct drug administration using a DDS; a description of certain DDS selected for their therapeutic contributions and a brief presentation of perspectives; a presentation of certain recommendations for organizations concerned with DDS.

Biodistribution of long-circulating PEG-grafted nanocapsules in mice: effects of PEG chain length and density.

PURPOSE: To study the pharmacokinetics and biodistribution of novel polyethyleneglycol (PEG) surface-modified poly(rac-lactide) (PLA) nanocapsules (NCs) and to investigate the influence of PEG chain length and content. METHODS: The biodistribution and plasma clearance in mice of different NC formulations were studied with [3H]-PLA. PLA-PEG copolymers were used in NC preparations at different chain lengths (5 kDa and 20 kDa) and PEG contents (10% and 30% w/w of total polymer). In vitro and in vivo stability were also checked. RESULTS: Limited [3H]-PLA degradation was observed after incubation in mouse plasma for 1 h, probably because of to the large surface area and thin polymer wall. After injection into mice. NCs prepared with PLA-PEG copolymers showed an altered distribution compared to poloxamer-coated PLA NCs. An increased concentration in plasma was also observed for PLA-PEG NCs. even after 24 h. A dramatic difference in the pharmacokinetic parameters of PLA-PEG 45-20 30% NCs compared to poloxamer-coated NCs indicates that covalent attachment, longer PEG chain lengths, and higher densities are necessary to produce an increased half-life of NCs in vivo. CONCLUSIONS: Covalently attached PEG on the surface of NCs substantially can reduce their clearance from the blood compartment and alter their biodistribution.

[Modulation of in vivo fate of drugs: fundamental biopharmaceutical and pharmacokinetic aspects]. / Peut-on moduler le devenir in vivo des médicaments? (Bases biopharmaceutiques et pharmacocinétiques).

The in vivo fate of drugs is basically regulated by the drug's physico-chemical properties, and thus depends on its chemical structure. Three successive phases describe the fate of exogenous compounds (absorption, distribution, elimination). The first two phases can be modulated by galenic formulation. Controlling the available dose and release rate by administration route and formulation, allows to modulate bioavailability of the active substance and its delivery to the general circulation. For distribution, vectorization techniques aim at improving the benefit/risk ratio. Two approaches are used: targeting the site of action (or avoiding site(s) leading to undesirable effects) and/or specific release rates.

Poly(rac-lactide) nanocapsules containing diclofenac: protection against muscular damage in rats.

The aim of this work was to determine whether encapsulation of a non steroidal antiinflammatory agent within nanocapsules could reduce local toxicity after intramuscular injection. Diclofenac-loaded nanocapsules were prepared by deposition of poly(rac-lactic acid) polymer, and administered intramuscularly to male Wistar rats. Plasma creatine phosphokinase (CPK) activity and histological examination were used to assess local tissue damage. Following a single intramuscular injection of diclofenac (0.8 mg), CPK activity was shown to depend on both the type of dosage form and, in the case of nanocapsules, on the chemical nature of the central oily core. Lower CPK activity was observed with nanocapsules prepared from Miglyol 810, a caprylic/capric triglyceride, while nanocapsules prepared from benzyl benzoate, either empty or containing diclofenac, exhibited the same CPK activity as the drug solution. Histopathological examination performed three days after administration of free diclofenac or nanocapsules containing diclofenac prepared from Miglyol 810 revealed that a much more intense inflammation was obtained with the solution than with nanocapsules. In conclusion, when appropriately formulated, nanocapsules can considerably reduce the muscular damage caused by diclofenac.

[Evaluation of film forming properties of a hydroxypropylmethylcellulose phthalate (HP 55) pseudolatex]. / Evaluation des propriétés filmogènes d'un pseudolatex de phtalate d'hydroxypropylméthyl cellulose (HP55).

An hydroxypropylmethyl cellulose phthalate (HP55) pseudolatex was prepared by a nanoprecipitation method. This allows nanoparticles formation when a solution of the polymer in an organic solvent (phase S1 or organic phase) is introduced in a precipitant medium (phase S2 or aqueous phase) which consists of water containing surfactant. A modification of the solvent nature of phase S1 resulted in an improvement of the yield of the preparation. Then, the obtained pseudolatex was used to prepare films which were evaluated by mechanical tests and coating of tablets containing theophylline.

Preparation and characterization of nanoparticles bearing heparin or dextran covalently-linked to poly(methyl methacrylate).

Nanoparticles have been obtained directly in aqueous media, from amphiphilic copolymers synthesized by radical polymerization of methyl methacrylate (MMA) initiated by Ce(IV) ions in the presence of heparin or dextran. The reaction conditions under which the copolymers spontaneously formed nanoparticles depended on the type of polysaccharide and on the concentrations of the reagents. Fluorescent nanoparticles containing N-vinyl carbazole (NVC), covalently linked to PMMA, were also prepared by random copolymerization of MMA and NVC in similar polymerization systems. The non-fluorescent nanoparticle suspensions were stable for several months without using surfactant. The fluorescent particles were larger and less stable then the unlabelled ones. Since all the particles are monodisperse, and in the submicron range, they can be used as models of drug carriers; the covalently-linked fluorescent species allowing them to be followed in vivo. The average molecular weights of the PMMA blocks of the copolymers and of oxidized heparin and dextran were determined by viscometry and/or gel permeation chromatography. The antithrombic activity of oxidized heparin was measured. The results show that the polysaccharide chains were cleaved by Ce(IV) in aqueous nitric acid, resulting in formation of block copolymers made of one or two blocks of PMMA linked to the ends of one polysaccharide block. Taken together, the results suggest that the particles were organized with the polysaccharidic moieties on the surface of the particles and the more hydrophobic PMMA or P(MMA-co-NVC) in the core, in a brush-like structure. This should confer 'stealth' properties to such particles.

Pulsed estrogen therapy: pharmacokinetics of intranasal 17-beta-estradiol (S21400) in postmenopausal women and comparison with oral and transdermal formulations.

Pharmacokinetics of estradiol and estrone were assessed in postmenopausal women receiving S21400, a novel 17beta-estradiol formulation administered by nasal route; the results were compared with those obtained with oral and transdermal routes. Thirty six women received three treatments: a specified dose of 17beta-estradiol (100, 300 or 450 microg) given once and as 2 doses, 12 h apart, using three parallel dose groups in a randomised, crossover study. Thereafter, a reservoir patch (50 microg/day of 17beta-estradiol) or a tablet of 2 mg micronised 17beta-estradiol were randomly administered. Plasma concentrations of estradiol and estrone were measured by radioimmunoassays. Following intranasal dosing, estradiol was rapidly absorbed with plasma concentrations reaching maximal values (approximately 1400 pg/ml with a single 300 microg dose) after 10-30 min and returning within 12 h to levels of untreated postmenopausal women. Systemic exposure to estradiol was dose proportional and independent of the treatment regimen. Moreover, the dose of 300 microg gave an estimated 24 h systemic exposure to exogenous estradiol close to that of the 50 microg/day reservoir patch or the 2 mg tablet. The mean estrone to estradiol ratio was similar and 4-fold lower than those with the patch and the tablet, respectively. In conclusion, by this new route for estrogen replacement therapy, the nasal route, the pharmacokinetics of estradiol as S21400 were linear and displayed a 'pulsed' kinetic profile, different from those obtained with the usual routes of administration.

Liposomal formulations for oral immunotherapy: in-vitro stability in synthetic intestinal media and in-vivo efficacy in the mouse.

The aim of this work was to develop a liposomal formulation which could act as a carrier for allergens during oral desensitization therapy. A model protein, ovalbumin, was associated with negatively charged, multilamellar vesicles of various compositions and their stability in the presence of synthetic intestinal media (bile salt, pancreatic enzymes and their combination) was investigated. Liposomes containing soya phosphatidylcholine as the main lipid, regardless of their cholesterol content (20-40%), were unable to protect ovalbumin against the combined action of pancreatic enzymes and bile salt. In contrast, liposomes prepared from distearoylphosphatidylcholine and cholesterol (6:3.5 molar ratio) were more stable: about 50% of the lipid remained as liposomes after a 4-h incubation at 37 degrees C and intact ovalbumin could be demonstrated therein by immunoblotting. The immunomodulating properties of liposomes were tested by following changes in serum IgE levels (by passive cutaneous anaphylaxis) in Balb/C mice sensitized to ovalbumin, after feeding various preparations. In this model, free ovalbumin was able to provoke a premature fall in IgE levels, and liposomes, whatever their composition, contributed no further effect.

Long-circulating nanoparticles bearing heparin or dextran covalently bound to poly(methyl methacrylate).

PURPOSE: In a biomimetic approach to the development of drug carriers escaping early capture by phagocytes, nanoparticles made of amphiphilic copolymers of either heparin or dextran and methyl methacrylate were evaluated relative to their in vivo blood circulation time. They were compared to bare PMMA nanoparticles. METHODS: Owing to the fluorescent properties of the covalently attached N-vinyl carbazole, the particles could be detected directly in mouse plasma. Samples were drawn at different time intervals and fluorescence was recorded. RESULTS: After an initial phase of elimination from the blood with a half-life of 5 h, the remaining heparin nanoparticles circulated for more than 48 h and were still detectable in the plasma at 72 h. Dextran nanoparticles were also eliminated very slowly over 48 h. Bare poly (methyl methacrylate) nanoparticles were found to have a half-life of only 3 min. CONCLUSIONS: Both types of nanoparticles proved to be long-circulating. The potent capacity for opsonisation of the poly(methyl methacrylate) core were hidden by the protective effect of either polysaccharide, probably due to a dense brush-like structure. In the case of heparin nanoparticles, the "stealth" effect was probably increased by its inhibiting properties against complement activation.

Interactions of nanoparticles bearing heparin or dextran covalently bound to poly(methyl methacrylate) with the complement system.

The efficient uptake of injected nanoparticles by cells of the mononuclear phagocyte system (MPS) limits the development of long-circulating colloidal drug carriers. The complement system plays a major role in the opsonization and recognition processes of foreign materials. Since heparin is an inhibitor of complement activation, nanoparticles bearing heparin covalently bound to poly(methyl methacrylate) (PMMA) have been prepared and their interactions with complement evaluated. The particles retained the complement-inhibiting properties of soluble heparin. Nanoparticles bearing covalently bound dextran instead of heparin were weak activators of complement as compared with crosslinked dextran (Sephadex) or bare PMMA nanoparticles. In addition to the specific activity of bound heparin, the protective effect of both polysaccharides is hypothesized to be due to the presence of a dense brush-like layer on the surface of the particles. Such properties are expected to reduce the uptake by MPS in vivo.

Modulation of nitric oxide production in RAW 264.7 cells by transforming growth factor-beta and interleukin-10: differential effects on free and encapsulated immunomodulator.

The mouse macrophage cell line RAW 264.7 can be stimulated to produce nitric oxide (NO) by muramyltripeptide cholesterol included within biodegradable poly(D,L-lactide) nanocapsules (NC MTPChol). The aim of this work was to determine whether one or both of the cytokines transforming growth factor-beta (TGF-beta) and interleukin-10 (IL-10) could be responsible for feedback control seen at high concentrations. Activated RAW 264.7 cells produced TGFbeta1. When exogenous TGF-beta1 was added during stimulation, a dose-dependent inhibition of NO production was observed when NC MTP-Chol was used, whereas activation by the soluble muramyl dipeptide (MDP) was not affected. Furthermore, addition of a blocking antibody to TGF-beta arrested the fall in NO production seen at high concentrations of NC MTP-Chol. Addition of IL-10 during RAW 264.7 cell activation also reduced NO production; however, in this case, both NC MTP-Chol and MDP were equally affected. The presence of anti-IL-10 antibody during activation significantly increased NO production.

Influence of sterilization processes on poly(epsilon-caprolactone) nanospheres.

Polymeric vectors and especially poly(epsilon-caprolactone) nanoparticles have already shown promising results in the optimization of the ophthalmic bioavailability of drugs. Any formulation instilled in the eye must be sterile, and preferentially isotonic. Poly(epsilon-caprolactone) nanospheres were thus formulated with Synperonic PE/F68, Synperonic PE/F127, or Cremophor RH40. A tonicity agent, a preservative and, in some cases, a viscosifiant were then added. The pH was finally adjusted to pH 4 or buffered to pH 7. Different sterilization processes were studied to investigate their influence on the physicochemical characteristics of vectors. Autoclaving did not induce any modification on polymer molecular weight or Synperonic nanospheres diameter, but catalysed some reactions with surfactants and tonicity agents. This method could thus be used if the nanosphere excipients are chosen with care. gamma radiation induced preservative degradation and viscosifiant depolymerization. A cross-linking of poly(epsilon-caprolactone) chains was observed, as reflected by a sharp increase of its molecular weight. However, no variation of the mean particle size was detected. Finally, sterile filtration was the only process which ensured the conservation of physicochemical integrity of nanospheres. This process was successfully applied on non-viscosified vectors with a sufficiently small diameter.

Synthesis and immunostimulating properties of lipophilic ester and ether muramyl peptide derivatives.

Macrophages can become cytotoxic toward tumor cells when activated by immunomodulators. Three different muramyl peptides were synthesized: one hydrolyzable lipophilic ester derivative (MTP-Chol) and two nonhydrolyzable lipophilic ether derivatives (MTP-octadecane and MTP-heptadecafluorooctadecane). Activation of the RAW 264.7 cell line was studied by measuring nitrite production as an indication of NO-synthase activity. The lipophilic ester derivative, incorporated within nanocapsules, was as active as free muramyl dipeptide, whereas the lipophilic ether derivatives were unable to activate macrophages. MTP-octadecane in micellar form was not capable of inducing macrophage cytotoxicity either. These results indicate that lipophilic muramyl peptides need to be hydrolyzed to yield a hydrosoluble metabolite in order to activate macrophages.

In-vitro and in-vivo evaluation of a new amphotericin B emulsion-based delivery system.

The in-vitro and in-vivo toxicity and activity of a new emulsion-based delivery system for amphotericin B (AmB-E) and of deoxycholate-amphotericin B (Fungizone) were studied. In vitro, Candida albicans and human red blood cells (RBCs) were treated with either product and dose-response curves for various cellular effects (changes in potassium cell content, haemoglobin leakage from RBCs and colony-forming ability of fungal cells) were obtained. AmB-E was less toxic than Fungizone against human RBCs and equally active against C. albicans cells. In-vivo studies showed that the LD50 of AmB-E and Fungizone in noninfected OF1 mice were 7.24 and 3.46 mg/kg, respectively. The therapeutic efficacy of AmB-E was assessed in murine candidiasis. Firstly, the efficacy of equal doses (0.8 mg/kg) of AmB-E and Fungizone was evaluated in infected mice. Both formulations increased the survival time compared to the control and were equally effective in reducing the cfu counts in the kidney. In the same model of infection, the maximum tolerated doses (MTD) of Fungizone and AmB-E were determined in order to study the efficacies of Fungizone and AmB-E at their respective MTD. AmB-E significantly increased the number of long-term survivors compared with Fungizone (MTD:2 and 1 mg/kg, respectively). Thus, AmB-E was more effective than Fungizone for treatment of systemic mycoses at the MTD.

Relationship between NO-synthase activity and TNF-alpha secretion in mouse macrophage lines stimulated by a muramyl peptide entrapped in nanocapsules.

The present study evaluates the ability of a new drug carrier: nanocapsules of poly(D,L-lactide) containing muramyldipeptide-L-alanyl-cholesterol (MTP-Chol NC) to induce activation of mouse macrophage cell lines. MTP-Chol NC stimulated nitric oxide (NO) expression and tumor necrosis factor-alpha (TNF-alpha) production, these are two important mediators of macrophage-mediated cytotoxicity. The encapsulated form was more effective than free muramyldipeptide, at low immunomodulator concentrations. The dose-response curves were completely different for NO and TNF-alpha, implying different regulatory mechanisms. In RAW 264.7 cells, the addition of anti-TNF-alpha antibodies during the activation period did not affect the level of nitrite induced by MTP-Chol Nc and lipopolysaccharide. Therefore, autocrine stimulation by TNF-alpha did not contribute to NO production. On the other hand, the presence of an NO synthase inhibitor led to an increase in TNF-alpha secretion. In J774.A1 cells, which were activated by MTP-Chol NC and interferon-gamma, TNF-alpha production seemed to act as a second messenger. Thus, under certain conditions, NO can play a role in modulating the cytotoxic activities of mouse macrophages.

Development of an injectable formulation of albendazole and in vivo evaluation of its efficacy against Echinococcus multilocularis metacestode.

The loading of poly (D, L-lactide) nanoparticles with ABZ has led us to evaluate the potential of this new colloidal drug delivery system against E. multilocularis, using a murine model of hepatic alveolar echinococcosis. ABZ-loaded nanoparticles had a monodisperse size distribution between 100 and 150 nm. The efficiency of drug loading to nanoparticles was over 97%. In vitro, at an ABZ concentration of 1.0 microgram ml-1, the formulation had no toxicity for peritoneal macrophages harvested from uninfected mice. In vivo, the ABZ-loaded nanoparticles exhibited no signs of toxicity at any of the doses tested. Intravenous injections of 6 mg kg-1 of bound ABZ to infected mice had an equivalent antiparasitic effect on the metacestode growth to that of a treatment with 1500 mg kg-1 of orally administered free ABZ. The parasite hepatic superficial size as well as the peritoneal metastatic burden was significantly reduced by these 2 courses of treatment, as compared to those of untreated mice. Our results should encourage further study in order to explain the absence of dose-dependent efficacy of ABZ-loaded nanoparticles demonstrated in the present study.

Factors influencing macrophage activation by muramyl peptides: inhibition of NO synthase activity by high levels of NO.

Treatment with muramyldipeptide (MDP) or a lipophilic derivative (MTP-Chol) included in nanocapsules renders macrophages cytostatic towards tumor cells. At the same time, nitric oxide (NO) synthase (EC 1.14.23) activity is induced, as determined by measurement of the two end products of the reaction (nitrite and L-citrulline). The objective of this study was to investigate some factors which might influence this activation and explain the decreased response observed at high nanocapsule concentrations. The glucose content of the medium did not seem to be limiting. Addition of indomethacin decreased nitrite production in the effector phase, suggesting a role for prostaglandins in the maintenance of the activated state. We also tested the hypothesis that NO itself might regulate inducible nitric oxide synthase activity. The addition of NO donors (SIN-1 and nitrosoglutathione) or superoxide dismutase to cultures of activated macrophages inhibited the NO synthase activity. Since these NO donors were non toxic towards macrophages, these observations indicate clearly that the addition of exogenous NO to that formed by the enzymatic reaction can cause inhibition of the inducible NO synthase.

The activity and ultrastructural localization of primaquine-loaded poly (d,l-lactide) nanoparticles in Leishmania donovani infected mice.

The activity of primaquine (PQ) was compared to that of PQ-loaded Poly (D,L-Lactide) (PLA) nanoparticles against Leishmania donovani. The association of PQ with PLA nanoparticles increased the antileishmanial in vitro and in vivo effects of intravenously administered drug while its toxicity was reduced. The effectiveness of PQ-loaded PLA nanoparticles was 3.3 times as high as that of free drug in the suppression of amastigotes in the liver of BALB/c mice. A total dose of 30 mg PQ bound/kg (600 mg PLA/kg) was well tolerated and no sign of acute toxicity was observed. A similar dose of free drug resulted in 15% weight loss. No significant leishmanicidal activity was observed for unloaded PLA nanoparticles. Ultrastructural studies showed the uptake of drug-loaded nanoparticles by Leishmania-infected Kupffer cells. Nanoparticles were identified closed to amastigotes.