Maria Angela Vandelli

NIR-labeled nanoparticles engineered for brain targeting: in vivo optical imaging application and fluorescent microscopy evidences

The presence of the blood–brain barrier (BBB) makes extremely difficult to develop efficacious strategies for targeting contrast agents and delivering drugs inside the Central Nervous System (CNS). To overcome this drawback, several kinds of CNS-targeted nanoparticles (NPs) have been developed. In particular, we proposed poly-lactide-co-glycolide (PLGA) NPs engineered with a simil-opioid glycopeptide (g7), which have already proved to be a promising tool for achieving a successful brain targeting after i.v. administration in rats. In order to obtain CNS-targeted NPs to use for in vivo imaging, we synthesized and administrated in mice PLGA NPs with double coverage: near-infrared (NIR) probe (DY-675) and g7. The optical imaging clearly showed a brain localization of these novel NPs. Thus, a novel kind of NIR-labeled NPs were obtained, providing a new, in vivo detectable nanotechnology tool. Besides, the confocal and fluorescence microscopy evidences allowed to further confirm the ability of g7 to promote not only the rat, but also the mouse BBB crossing.

Gelatin microspheres crosslinked with D,L-glyceraldehyde as a potential drug delivery system: preparation, characterisation, in vitro and in vivo studies.

To overcome the restriction in using crosslinked gelatin in the pharmaceutical field, D,L-glyceraldehyde (GAL), a non-toxic crosslinking agent, was proposed. Gelatin microspheres crosslinked with different concentrations of GAL (0.5, 1 or 2%, w/v) and for different time periods (1 or 24 h) were prepared. The effect of the preparation variables was evaluated analysing the extent of crosslinking, the morphological aspect, the particle size and the swelling behaviour. To evaluate the pharmaceutical properties, an antihypertensive drug, clonidine hydrochloride, was chosen as drug model and loaded into the microspheres. Either the increase of the crosslinker concentration or of the crosslinking time period decreased both the swelling and the in vitro drug release processes of the microspheres. After the subcutaneous injection, the loaded microspheres crosslinked with the lowest GAL concentration (0.5%, w/v) or for the shortest time period (1 h) showed a reduction of systolic blood pressure (SBP) similar to that recorded with a clonidine hydrochloride solution having the same drug concentration. Instead, the microspheres crosslinked for 24 h with concentrations of GAL higher than 0.5% (w/v) produced a more gradual and sustained SBP reduction and the antihypertensive effect was maintained until 52-72 h. The biocompatibility studies showed that the microspheres crosslinked with GAL are well tolerated in vivo. These results suggest the potential application of gelatin microspheres crosslinked with GAL as a suitable drug delivery system for the subcutaneous administration.

Insight on the fate of CNS-targeted nanoparticles. Part I: Rab5-dependent cell-specific uptake and distribution

Nanocarriers can be useful tools for delivering drugs to the central nervous system (CNS). Their distribution within the brain and their interaction with CNS cells must be assessed accurately before they can be proposed for therapeutic use. In this paper, we investigated these issues by employing poly-lactide-co-glycolide nanoparticles (NPs) specifically engineered with a glycopeptide (g7) conferring to NPs the ability to cross the blood brain barrier (BBB) at a concentration of up to 10% of the injected dose. g7-NPs display increased in vitro uptake in neurons and glial cells. Our results show that in vivo administration of g7-NPs leads to a region- and cell type-specific enrichment of NPs within the brain. We provide evidence that g7-NPs are endocytosed in a clathrin-dependent manner and transported into a specific subset of early endosomes positive for Rab5 in vitro and in vivo. The differential Rab5 expression level is strictly correlated with the amount of g7-NP accumulation. These findings show that g7-NPs can cross the BBB and target specific brain cell populations, suggesting that these NPs can be promising carriers for the treatment of neuropsychiatric and neurodegenerative diseases.

Polymeric nanoparticles for the drug delivery to the central nervous system.

Background: Nanoparticulate polymeric systems (nanoparticles [Np]) have been widely studied for the delivery of drugs to a specific target site. This approach has been recently considered for the therapy of brain diseases. The major problem in accessing the CNS is linked to the presence of the blood–brain barrier. Objective: The present review deals with the different strategies that have been developed in order to allow Np drug carriers entry into the CNS parenchyma. Among these, the use of magnetic Np, Np conjugation with ligands for blood–brain barrier receptors, with antibodies, and the use of surfactants have been considered. Methods: All the literature available is reviewed in order to highlight the potential of this drug delivery system to be used as a drug carrier for the treatment of CNS pathologies. Conclusions: Polymeric Np have been shown to be promising carriers for CNS drug delivery due to their potential both in encapsulating drugs, hence protecting them from excretion and metabolism, and in delivering active agents across the blood–brain barrier without inflicting any damage to the barrier. Different polymers have been used and different strategies have been applied; among these, the use of specific ligands to enhance the specificity of drugs delivered to the CNS has recently been considered. At present, clinical trials are being conducted appeared for the use of these drug carriers but none related to the treatment of CNS diseases.

Influence of preparation conditions on acyclovir-loaded poly-d,l-lactic acid nanospheres and effect of PEG coating on ocular drug bioavailability.

AbstractPurpose. The evaluation of nanosphere colloidal suspensions containing acyclovir as potential ophthalmic drug delivery systems was carried out. The influence of polymer molecular weight and type and concentration of various surfactants on nanosphere properties was studied. The ocular pharmacokinetics of acyclovir-loaded nanoparticles was evaluated in vivo and compared with an aqueous suspension of the free drug. Methods. Nanospheres were made up of poly-d,l-lactic acid (PLA). The colloidal suspension was obtained by a nanoprecipitation process. The surface properties of PLA nanospheres were changed by the incorporation of pegylated 1,2-distearoyl-3-phosphatidylethanol- amine. The mean size and zeta potential of the nanospheres were determined by light scattering analysis. The acyclovir loading capacity and release were also determined. In vivo experiments were carried out on male New Zealand rabbits. The ocular tolerability of PLA nanospheres was evaluated by a modified Draize test. The aqueous humor acyclovir levels were monitored for 6 h to determine the drugs ocular bioavailability for the various formulations. Results. A reduction of the mean size and a decrease of the absolute zeta potential of PLA nanospheres resulted from increasing the surfactant concentration. The higher the polymer molecular weight, the smaller the nanosphere mean size. PEG-coated and uncoated PLA nanospheres showed a sustained acyclovir release and were highly tolerated by the eye. Both types of PLA nanospheres were able to increase the aqueous levels of acyclovir and to improve the pharmacokinetics profile, but the efficacy of the PEG-coated nanospheres was significantly higher than that of the simple PLA ones. Conclusions. PEG-coated PLA nanospheres can be proposed as a potential ophthalmic delivery system for the treatment of ocular viral infections.

Doxorubicin-loaded gelatin nanoparticles stabilized by glutaraldehyde: Involvement of the drug in the cross-linking process

Abstract The possible involvement of the primary amino group of doxorubicin (DXR) in the cross-linking process of gelatin nanoparticles stabilized by glutaraldehyde was investigated. Nanoparticles were characterized with regard to particle size, drug content, enzymatic degradation and cross-linking degree. The size of nanoparticles was around 100–200 nm and DXR was loaded with an entrapment efficiency of 42%. Upon the study of crosslinking rate, DXR-loaded nanoparticles showed a greater number of free amino groups than the unloaded ones. This should be due to a competition between the amino group of DXR and the amino groups of the gelatin chains during the cross-linking process. Hence, a binding of a drug fraction to the protein matrix via glutaraldehyde was hypothesized and confirmed by the results of a thin-layer chromatography (TLC) analysis. According to the in vitro study only a little fraction of DXR was released as free drug (8%) when the nanoparticles were put in saline solution. The addition of proteolytic enzymes disrupts the matrix structure producing the release of a further 10–15% of the drug loading which was entrapped in the nanoparticle matrix. The remaining part of the drug corresponds to DXR covalently linked to peptide residues produced by nanoparticle digestion.

Nanoparticle transport across the blood brain barrier.

ABSTRACT While the role of the blood-brain barrier (BBB) is increasingly recognized in the (development of treatments targeting neurodegenerative disorders, to date, few strategies exist that enable drug delivery of non-BBB crossing molecules directly to their site of action, the brain. However, the recent advent of Nanomedicines may provide a potent tool to implement CNS targeted delivery of active compounds. Approaches for BBB crossing are deeply investigated in relation to the pathology: among the main important diseases of the CNS, this review focuses on the application of nanomedicines to neurodegenerative disorders (Alzheimer, Parkinson and Huntingtons Disease) and to other brain pathologies as epilepsy, infectious diseases, multiple sclerosis, lysosomal storage disorders, strokes.

Nanoparticles as drug delivery agents specific for CNS: in vivo biodistribution

UNLABELLED The pharmacological treatment of neurological disorders is often complicated by the inability of drugs to pass the blood-brain barrier. Recently we discovered that polymeric nanoparticles (NPs) made of poly(D,L-lactide-co-glycolide), surface-decorated with the peptide Gly-L-Phe-D-Thr-Gly-L-Phe-L-Leu-L-Ser(O-beta-D-glucose)-CONH2 are able to deliver, after intravenous administration, the model drug loperamide into the central nervous system (CNS). This new drug delivery agent is able to ensure a strong and long-lasting pharmacological effect, far greater than that previously observed with other nanoparticulate carriers. Here we confirmed the effectiveness of this carrier for brain targeting, comparing the effect obtained by the administration of loperamide-loaded NPs with the effect of an intracerebroventricular administration of the drug; moreover, the biodistribution of these NPs showed a localization into the CNS in a quantity about two orders of magnitude greater than that found with the other known NP drug carriers. Thus, a new kind of NPs that target the CNS with very high specificity was discovered. FROM THE CLINICAL EDITOR This paper discusses a nanoparticle-based technique of targeted drug delivery through the blood-brain barrier. The biodistribution of these novel nanoparticles showed two orders of magnitude greater efficiency compared to other known NP drug carriers.

Sialic acid and glycopeptides conjugated PLGA nanoparticles for central nervous system targeting: In vivo pharmacological evidence and biodistribution

Polymeric nanoparticles (Np) have been considered as strategic carriers for brain targeting. Specific ligands on the surface allowed the Np to cross the Blood-Brain Barrier (BBB) carrying model drugs within the brain district after their i.v. administration in experimental animals. It is known that sialic acid receptors are present in several organs, including in the brain parenchyma. Thus, in this paper, we prepared PLGA Np surface modified with a BBB-penetrating peptide (similopioid peptide) for BBB crossing and with a sialic acid residue (SA) for the interaction with brain receptors. This double coverage could allow to obtain novel targeted Np with a prolonged residence within the brain parenchyma, thus letting to reach a long-lasting brain delivery of drugs. The central analgesic activity of Loperamide (opioid drug, unable to cross the BBB) loaded in these novel Np was evaluated in order to point out the capability of the Np to reach and to remain in the brain. The results showed that the pharmacological effect induced by loaded Np administration remained significant over 24h. Using confocal and fluorescent microscopies, the novel Np were localized within the tissue parenchyma (brain, kidney, liver, spleen and lung). Finally, the biodistribution studies showed a localization of the 6% of the injected dose into the CNS over a prolonged time (24h). Notwithstanding an increased accumulation of SA-covered Np in those organs showing SA-receptors (liver, kidney, and lung), the pharmacological and biodistribution results are proofs of the ability of double targeted Np to enter the brain allowing the drug to be released over a prolonged time.

AFM, ESEM, TEM, and CLSM in liposomal characterization: a comparative study

An outstanding aspect of pharmaceutical nanotechnology lies in the characterization of nanocarriers for targeting of drugs and other bioactive agents. The development of microscopic techniques has made the study of the surface and systems architecture more attractive. In the field of pharmaceutical nanosystems, researchers have collected vital information on size, stability, and bilayer organization through the microscopic characterization of liposomes. This paper aims to compare the results obtained by atomic force microscopy, environmental scanning electron microscopy, transmission electron microscopy, and confocal laser scanning microscopy to point out the limits and advantages of these applications in the evaluation of vesicular systems. Besides this comparative aim, our work proposes a simple confocal laser scanning microscopy procedure to rapidly and easily detect the liposomal membrane.