Elisabetta Muntoni

Determination of Ibuprofen in human plasma by high-performance liquid chromatography: validation and application in pharmacokinetic study.

A specific method for the simultaneous determination of S-(+)Ibuprofen and R-(-)Ibuprofen enantiomers in human plasma is described. Adopting a high-performance liquid chromatographic (HPLC) system with spectrofluorometer detector, the compounds were extracted from plasma in alcohol medium and were separated on C18 column, using a solution of acetonitrile-water-acetic acid-triethylamine as mobile phase. The limit of quantitation was 0.1 microg/mL for both compounds. The method was validated by intra-day assays at three concentration levels and was used in a kinetic study in healthy volunteers. During the study we carried out inter-day assays to confirm the feasibility of the method.

Expression of CYP3A isoforms and P-glycoprotein in human stomach, jejunum and ileum.

1 CYP3A isoforms metabolise a diverse array of clinically important drugs and P‐glycoprotein (P‐gp), a transmembrane efflux pump, can extrude a wide variety of drugs from the cell. It has been suggested that the function of CYP3A4 is complementary to that of P‐gp along the gastrointestinal (GI) tract, together forming a coordinated intestinal barrier against xenobiotics. Therefore, the expression of CYP3A4, CYP3A5, CYP3A7 and ABCB1 (P‐gp) genes were quantified in five normal samples from the human stomach, seven from the jejunum and eight from the ileum by real‐time reverse transcription–polymerase chain reaction and western blot analysis. 2 In the tissues examined, measurable mRNA expression of CYP3A was found in almost all samples from the stomach, jejunum and ileum. The rank order for CYP3A mRNA expression was CYP3A4 > CYP3A5 > CYP3A7 in the GI tract studied, whereas median mRNA CYP3A4 expression was highest in the small intestine and lowest in the stomach. Expression of ABCB1 mRNA was found in almost all samples and the median mRNA expression level was comparable in the jejunum and ileum, but lower in the stomach. Our data also show a significant correlation between all mRNA transcripts studied and a wide interindividual variation. 3 At the protein level, CYP3A4 was detected in all stomach and small intestine samples, the levels being substantially higher in the small intestine than in the stomach. P‐Glycoprotein was detected in all GI samples, but no statistically significant difference was found along the GI tract considered. 4 Collectively, these results demonstrate that CYP3A4 is the main CYP3A expressed in the GI tract investigated, an extensive interindividual variability in the expression of the different CYP3A isoforms in all tissues examined and P‐gp apoprotein levels similar in the stomach, jejunum and ileum.

Solid lipid nanoparticles as vehicles of drugs to the brain: current state of the art.

Central nervous system disorders are already prevalent and steadily increasing among populations worldwide. However, most of the pharmaceuticals present on world markets are ineffective in treating cerebral diseases, because they cannot effectively cross the blood brain barrier (BBB). Solid lipid nanoparticles (SLN) are nanospheres made from biocompatible solid lipids, with unique advantages among drug carriers: they can be used as vehicles to cross the BBB. This review examines the main aspects surrounding brain delivery with SLN, and illustrates the principal mechanisms used to enhance brain uptake of the delivered drug.

Solid Lipid Nanoparticles for Potential Doxorubicin Delivery in Glioblastoma Treatment: Preliminary In Vitro Studies

The major obstacle to glioblastoma pharmacological therapy is the overcoming of the blood-brain barrier (BBB). In literature, several strategies have been proposed to overcome the BBB: in this experimental work, solid lipid nanoparticles (SLN), prepared according to fatty acid coacervation technique, are proposed as the vehicle for doxorubicin (Dox), to enhance its permeation through an artificial model of BBB. The in vitro cytotoxicity of Dox-loaded SLN has been measured on three different commercial and patient-derived glioma cell lines. Dox was entrapped within SLN thanks to hydrophobic ion pairing with negatively charged surfactants, used as counterions. Results indicate that Dox entrapped in SLN maintains its cytotoxic activity toward glioma cell lines; moreover, its permeation through hCMEC/D3 cell monolayer, assumed as a model of the BBB, was increased when the drug was entrapped in SLN. In conclusion, SLN proved to be a promising vehicle for the delivery of Dox to the brain in glioblastoma treatment.

Thiopurine S-methyltransferase pharmacogenetics in a large-scale healthy Italian–Caucasian population: differences in enzyme activity

AIMS To investigate the influence of genotype, age and gender on the thiopurine S-methyltransferase (TPMT) phenotype in healthy Italian-Caucasian subjects. MATERIALS & METHODS The study investigated the TPMT genotype and the TPMT phenotype of 943 healthy Italian-Caucasian subjects of different age and gender (age range: 0.08-68 years; 623 males 320 females). TPMT red blood cell activity was measured in all samples and genotype was determined for the TPMT alleles *2, *3A, *3B and *3C. RESULTS TPMT activity levels in our whole population ranged from 1.6 up to 75.2 U/gHb. Significant TPMT activity differences between wild-type and heterozygous subjects were observed. We divided our TPMT activity into four categories according to our frequency distribution: low (0.1%), intermediate (32.9%), normal (60%) and high (7%), with arbitrary cut-off values of 8.0, 19.4 and 37.0 U/gHb, respectively. The whole population had a total of 94.5% of homozygous wild-type subjects, 5.4% heterozygous variants and one (0.1%) compound heterozygous variant TPMT*3B/*3C. The overall concordance rate between TPMT genotypes and phenotypes was 71.6%. The TPMT activity was significantly higher in wild-type children (0.08-17 years) than in wild-type adults (aged 18-68 years). Moreover, it was noted that wild-type infants from 0.08 to 5 years had a 9% higher average TPMT activity than the other wild-type groups, and only in children from 0.08 to 2 years was the TPMT activity higher in males than in females. CONCLUSION The data obtained in this study show that genetic factors seem to be the major aspect in TPMT phenotype variability in adults, whilst, in children, other physiological factors should be taken into consideration when assessing the TPMT phenotype, such as age and gender.

Application of lipid nanoparticles to ocular drug delivery

ABSTRACT Introduction: Although eye drops are widely used as drug delivery systems for the anterior segment of the eye, they are also associated with poor drug bioavailability due to transient contact time and rapid washout by tearing. Moreover, effective drug delivery to the posterior segment of the eye is challenging, and alternative routes of administration (periocular and intravitreal) are generally needed, the blood–retinal barrier being the major obstacle to systemic drug delivery. Areas covered: Nanotechnology, and especially lipid nanoparticles, can improve the therapeutic efficiency, compliance and safety of ocular drugs, administered via different routes, to both the anterior and posterior segment of the eye. This review highlights the main ocular barriers to drug delivery, as well as the most common eye diseases suitable for pharmacological treatment in which lipid nanoparticles have proved efficacious as alternative delivery systems. Expert opinion: Lipid-based nanocarriers are among the most biocompatible and versatile means for ocular delivery. Mucoadhesion with consequent increase in pre-corneal retention time, and enhanced permeation due to cellular uptake by corneal epithelial cells, are the essential goals for topical lipid nanoparticle delivery. Gene delivery to the retina has shown very promising results after intravitreal administration of lipid nanoparticles as non-viral vectors.

Cholesteryl butyrate solid lipid nanoparticles inhibit the adhesion and migration of colon cancer cells

BACKGROUND AND PURPOSE Cholesteryl butyrate solid lipid nanoparticles (cholbut SLN) provide a delivery system for the anti‐cancer drug butyrate. These SLN inhibit the adhesion of polymorphonuclear cells to the endothelium and may act as anti‐inflammatory agents. As cancer cell adhesion to endothelium is crucial for metastasis dissemination, here we have evaluated the effect of cholbut SLN on adhesion and migration of cancer cells.

Baclofen-loaded solid lipid nanoparticles: Preparation, electrophysiological assessment of efficacy, pharmacokinetic and tissue distribution in rats after intraperitoneal administration

Intrathecal baclofen administration is the reference treatment for spasticity of spinal or cerebral origin, but the risk of infection or catheter dysfunctions are important limits. To explore the possibility of alternative administration routes, we studied a new preparation comprising solid lipid nanoparticles (SLN) incorporating baclofen (baclofen-SLN). We used SLN because they are able to give a sustained release and to target the CNS. Wistar rats were injected intraperitoneally with baclofen-SLN or baclofen solution (baclofen-sol group) at increasing dosages. At different times up to 4 h, efficacy was tested by the H-reflex and two scales evaluating sedation and motor symptoms due to spinal lesions. Rats were killed and baclofen concentration determined in blood and tissues. Physiological solution or unloaded SLN was used as controls. After baclofen-SLN injection, the effect, consisting in a greater and earlier reduction of the H/M ratio than baclofen-sol group and controls, was statistically significant from a dose of 5 mg/kg and was inversely correlated with dose. Clinical effect of baclofen-SLN on both the behavioral scales was greater than that of baclofen-sol and lasted until 4th hour. Compared with baclofen-sol, baclofen-SLN produced significantly higher drug concentrations in plasma from 2nd hour until 4th hour with a linear decrement and in the brain at all times. In conclusion, our study demonstrated the efficacy of a novel formulation of baclofen, which exploits the advantages of SLN preparations. However, for clinical purposes, high baclofen concentrations in brain tissue and sedation may be unwanted effects, requiring further studies and optimization of dosages.

Solid lipid nanoparticles as promising tool for intraocular tobramycin delivery: Pharmacokinetic studies on rabbits

Eye drops are widely accepted as formulations for targeting the anterior segment notwithstanding their limitations in terms of bioavailability. The unique structure of the eye requires specially-designed formulations able to favor the pharmacokinetic profile of administered drugs, mainly minimizing the influence of ocular barriers. Nanotechnology-based delivery systems lead to significant technological and therapeutical advantages in ophthalmic therapy. The aim of the present study was to determine whether tobramycin as ion-pair incorporated in mucoadhesive Solid Lipid Nanoparticles (SLN) reaches the inner parts of the eye favoring drug activity. After technological characterization of the tobramycin entrapped SLN formulation (Tobra-SLN), a pharmacokinetic study in rabbits after topical instillation and intravenous administration of the formulation has been carried out. In addition, the intracellular activity of Tobra-SLN formulation against phagocytosed Pseudomonas aeruginosa was investigated. The SLN were spherical in shape, and showed a hydrodynamic diameter of about 80nm, a negative zeta potential (-25.7mV) with a polydispersity index of 0.15, representative of a colloidal dispersion with high quality, characterized by an unimodal relatively narrow size distribution. As demonstrated by FTIR and DSC, tobramycin ion-pair could be concentrated into lipid inner core of SLN, without interaction with the stearic acid, thus promoting a slow and constant drug release profile in the dissolution medium. Surprisingly, the drug concentration was significantly higher in all ocular tissues after ocular and intravenous administration of Tobra-SLN formulation with respect to reference formulations and only Tobra-SLN allowed the penetration of drug into retina. Furthermore, the use of Tobra-SLN resulted in both higher intraphagocytic antibiotic concentrations in polymorphonuclear granulocytes and greater bactericidal activity against intracellular Pseudomonas aeruginosa, probably due to the ability of Tobra-SLN to penetrate either into phagocytic cells, or alternatively to cross bacterial barrier. The present study broadens the knowledge on the use of SLN as carriers for ocular drug delivery to the posterior chamber and might open new avenues for treatment of ocular infections, representing a strategy to overcome the microbial resistance.

Lipid nanoparticles for intranasal administration: application to nose-to-brain delivery

ABSTRACT Introduction: The blood brain barrier is a functional barrier allowing the entry into the brain of only essential nutrients, excluding other molecules. Its structure, although essential to keep the harmful entities out, is also a major roadblock for pharmacological treatment of brain diseases. Several alternative invasive drug delivery approaches, such as transcranial drug delivery and disruption of blood brain barrier have been explored, with limited success and several challenges. Intranasal delivery is a non-invasive methodology, which bypasses the systemic circulation, and, through the intra- and extra- neuronal pathways, provides direct brain drug delivery. Colloidal drug delivery systems, particularly lipidic nanoparticles offer several unique advantages for this goal . Areas covered: This review focuses on key brain diseases such as Alzheimer’s disease, Parkinson’s disease, Huntington’s disease and amyotrophic lateral sclerosis, and provide a detailed overview of the current lipid nanoparticle based treatment options explored thus far. The review also delves into basic preparation, challenges and evaluation methods of lipid drug delivery systems. Expert opinion: Brain diseases present complex pathophysiology, in addition to the practically inaccessible brain tissues, hence according to the authors, a two-pronged approach utilizing new target discovery coupled with new drug delivery systems such as lipid carriers must be adopted.