Angela Lopedota

A comparative study of chitosan and chitosan/cyclodextrin nanoparticles as potential carriers for the oral delivery of small peptides ☆

The aim of this study was to characterize new nanoparticles (NPs) containing chitosan (CS), or CS/cyclodextrin (CDs), and evaluate their potential for the oral delivery of the peptide glutathione (GSH). More precisely, NP formulations composed of CS, CS/alpha-CD and CS/sulphobutyl ether-beta-cyclodextrin (SBE(7m)-beta-CD) were investigated for this application. CS/CD NPs showed particle sizes ranging from 200 to 500nm. GSH was loaded more efficiently in CS/SBE(7m)-beta-CD NPs by forming a complex between the tripeptide and the CD. X-ray Photoelectron Spectroscopy (XPS) analysis suggested that GSH is located in the core of CS/SBE(7m)-beta-CD NPs and that it is almost absent from the NP surface. Release studies performed in vitro at pH 1.2 and pH 6.8 showed that NP release properties can be modulated by selecting an appropriate CD. Transport studies performed in the frog intestine model confirmed that both CS and CS/CD nanoparticles could induce permeabilization of the intestinal epithelia. However, CS/SBE(7m)-beta-CD NPs provided absorption-enhancing properties in all segments of the duodenum, whereas CS NPs effect was restricted to the first segment of the duodenum. From the data obtained, we believe that CS/CD nanoparticles might represent an interesting technological platform for the oral administration of small peptides.

Recent Advances in Medicinal Chemistry and Pharmaceutical Technology- Strategies for Drug Delivery to the Brain

This paper provides a mini-review of some recent approaches for the treatment of brain pathologies examining both medicinal chemistry and pharmaceutical technology contributions. Medicinal chemistry-based strategies are essentially aimed at the chemical modification of low molecular weight drugs in order to increase their lipophilicity or the design of appropriate prodrugs, although this review will focus primarily on the use of prodrugs and not analog development. Recently, interest has been focused on the design and evaluation of prodrugs that are capable of exploiting one or more of the various endogenous transport systems at the level of the blood brain barrier (BBB). The technological strategies are essentially non-invasive methods of drug delivery to malignancies of the central nervous system (CNS) and are based on the use of nanosystems (colloidal carriers) such as liposomes, polymeric nanoparticles, solid lipid nanoparticles, polymeric micelles and dendrimers. The biodistribution of these nanocarriers can be manipulated by modifying their surface physico-chemical properties or by coating them with surfactants and polyethylene-glycols (PEGs). Liposomes, surfactant coated polymeric nanoparticles, and solid lipid nanoparticles are promising systems for delivery of drugs to tumors of the CNS. This mini-review discusses issues concerning the scope and limitations of both the medicinal chemistry and technological approaches. Based on the current findings, it can be concluded that crossing of the BBB and drug delivery to CNS is extremely complex and requires a multidisciplinary approach such as a close collaboration and common efforts among researchers of several scientific areas, particularly medicinal chemists, biologists and pharmaceutical technologists.

The use of Eudragit RS 100/cyclodextrin nanoparticles for the transmucosal administration of glutathione

The aim of this work was to develop and characterize new nanoparticle systems based on Eudragit RS 100 and cyclodextrins (CDs) for the transmucosal administration of glutathione (GSH). For this purpose, nanoparticles (NPs) with the mucoadhesive properties of Eudragit RS 100 and the penetration enhancing and peptide protective properties of CDs were prepared and evaluated. The quasi-emulsion solvent diffusion technique was used to prepare the NPs with natural and chemically modified (HP-beta-CD and Me-beta-CD) CDs. The NPs prepared showed homogeneous size distribution, mean diameters between 99 and 156nm, a positive net charge and spherical morphology. Solid state FT-IR, thermal analysis (DSC), and X-ray diffraction studies suggest that the nanoencapsulation process produces a marked decrease in crystallinity of GSH. The encapsulation efficiency of the peptide was found to be between 14.8% and 24%. The results indicate that mean diameters, surface charges and drug-loaded NPs were not markedly affected by the CD, whereas the presence of the latter influences drug release and to some extent peptide stability and absorption. Finally, it has been shown that CD/Eudragit RS 100 NPs may be used for transmucosal absorption of GSH without any cytotoxicity using the epithelial human HaCaT and murine monocyte macrophage RAW264.7 cell lines.

Translocator protein ligand-PLGA conjugated nanoparticles for 5-fluorouracil delivery to glioma cancer cells.

Translocator protein 18 kDa (TSPO) is a promising target for molecular imaging and for targeted drug delivery to tumors overexpressing TSPO. In our previous work, new macromolecular conjugates with a high affinity and selectivity for TSPO were prepared by conjugating the biodegradable poly(d,l-lactic-co-glycolic acid) (PLGA) polymer with two potent and selective TSPO ligands, namely, compounds 1 and 2. Based on this, nanoparticle delivery systems (NPs), employing TSPO ligand-PLGA conjugated (PLGA-TSPO) polymers, were prepared. Furthermore, to evaluate the ability of the new NPs to be used as a drug delivery systems for anticancer therapy, PLGA-TSPO NPs were loaded with 5-fluorouracil (5-FU), chosen as a model hydrophilic anticancer drug. The main goal of this work was to investigate the synergistic potential of using NP conjugates PLGA-TSPO, TSPO ligands being pro-apoptotic agents, to simultaneously deliver a cytotoxic anticancer drug. To better highlight the occurrence of synergistic effects, dual drug loaded PLGA NPs (PLGA NPs/5-FU/1) and dual drug loaded PLGA-TSPO NPs (PLGA-TSPO NPs/5-FU/1), with 5-FU and TSPO ligand 1 physically incorporated together, were also prepared and characterized. The particle size and size distribution, surface morphology, and drug encapsulation efficiency, as well as the drug release kinetics, were investigated. In vitro cytotoxicity studies were carried out on C6 glioma cells overexpressing TSPO, and to evaluate the potential uptake of these nanoparticulate systems, the internalization of fluorescent labeled PLGA-TSPO NPs (FITC-PLGA-TSPO NPs) was also investigated by fluorescence microscopy. Results demonstrated that PLGA-TSPO NPs/5-FU and dual drug loaded PLGA NPs/5-FU/1 and PLGA-TSPO NPs/5-FU/1 could significantly enhance toxicity against human cancer cells due to the synergistic effect of the TSPO ligand 1 with the anticancer drug 5-FU.

In vitro targeting and imaging the translocator protein TSPO 18-kDa through G(4)-PAMAM-FITC labeled dendrimer

Mitochondria represent an attractive subcellular target due to its function particularly important for oxidative damage, calcium metabolism and apoptosis. However, the concept of mitochondrial targeting has been a neglected area so far. The translocator protein (TSPO) represents an interesting subcellular target not only to image disease states overexpressing this protein, but also for a selective mitochondrial drug targeting. Recently, we have delivered in vitro and in vivo small molecule imaging agents into cells overexpressing TSPO by using a family of high-affinity conjugable ligands characterized by 2-phenyl-imidazo[1,2-a]pyridine acetamide structure. As an extension, in the present work we studied the possibility to target and image TSPO with dendrimers. These nano-platforms have unique features, in fact, are prepared with a level of control not reachable with most linear polymers, leading to nearly monodisperse, globular macromolecules with a large number of peripheral groups. As a consequence, they are an ideal delivery vehicle candidate for explicit study of the effects of polymer size, charge, composition, and architecture on biologically relevant properties such as lipid bilayer interactions, cytotoxicity, cellular internalization, and subcellular compartments and organelles interactions. Here, we present the synthesis, characterization, cellular internalization, and mitochondria labeling of a TSPO targeted fourth generation [G(4)-PAMAM] dendrimer nanoparticle labeled with the organic fluorescent dye fluorescein. We comprehensively studied the cellular uptake behavior of these dendrimers, into glioma C6 cell line, under the influence of various endocytosis inhibitors. We found that TSPO targeted-G(4)-PAMAM-FITC dendrimer is quickly taken up by these cells by endocytosis pathways, and moreover specifically targets the mitochondria as evidenced from subcellular fractionation experiments and co-localization studies performed with CAT (Confocal-AFM-TIRF) microscopy.

Water-soluble salts of aminoacid esters of the anaesthetic agent Propofol

Abstract The glycinates 4 , 5 , acetates 6 , 7 , 10 , propionate 8 , butyrate 9 and carbonate 11 were synthesized and evaluated as potential water-soluble prodrugs of Propofol (2,6-diisopropylphenol) 1 suitable for parenteral administration. The 4–9 · HCl salts were also prepared and some of them (i.e. 4 · HCl and 6 · HCl) were found sufficiently soluble in aqueous solutions. The kinetics of hydrolysis of the esters 4–11 and 4–9 · HCl salts were studied in 0.05 M phosphate buffer pH 7.4, and a number of derivatives ( 4 , 6 , 7 , and corresponding HCl salts) were examined for their stability in human plasma and brain homogenate. Our results indicated that the salts 4 · HCl and 6 · HCl, sufficiently soluble in water, are relatively stable in physiological media. Most of the examined compounds, in particular compound 6 , were found to inhibit the binding of [ 35 S]- tert -butylbicyclophosphorothionate ([ 35 S]TBPS) demonstrating to possess affinity for the Propofol recognition site on GABA A receptors.

Translocator Protein (TSPO) Ligand-Ara-C (Cytarabine) Conjugates as a Strategy To Deliver Antineoplastic Drugs and To Enhance Drug Clinical Potential

The aim of this work was to evaluate TSPO ligand-Ara-C conjugation as an approach for the selective delivery of the antineoplastic agent to brain tumors as well as for overcome P-gp resistance induction observed for the majority of cytotoxic agents, enhancing the drug clinical potential. To this end, the novel N-imidazopyridinacetyl-Ara-C conjugates 3a-c, 10 and 15 have been prepared and evaluated for their cytotoxicity against glioma cell lines. In contrast to that observed for 3a-c and 10, the conjugate 15 resulted stable in both phosphate buffer and physiological medium. In all cases, the release of free Ara-C from hydrolyzed conjugates was checked by HPLC and ESI-MS analysis. Conjugates 10 and 15 displayed very high in vitro TSPO affinity and selectivity, and, hence, they may possess potential for targeted brain delivery. Due to the favorable features displayed by the conjugate 15, it was further evaluated on glioma cell lines, expressing high levels of TSPO, in the presence and in the absence of specific nucleoside transport (NT) inhibitors. In contrast to that observed for the free Ara-C, the presence of NT inhibitors did not reduce the cytotoxic activity of 15. Moreover, conjugate 15, as N(4)-acyl derivative of Ara-C, should be resistant to inactivation by cytidine deaminase, and it may possess enhanced propensity to target brain tumor cells characterized by a reduced expression of NTs. In addition, this conjugate behaves as a clear P-gp modulator and thereby may be useful to reverse MDR. Transport studies across the MDCKII-MDR1 monolayer indicated that conjugate 15 should overcome the BBB by transcellular pathway. All these features may be useful for enhancing the clinical potential of the nucleoside drug Ara-C.

Novel L-Dopa and Dopamine Prodrugs Containing a 2-Phenyl-imidazopyridine Moiety

PurposeThe aim of this study was to gain insight into the feasibility of enhancing the delivery of L-Dopa and dopamine to the brain by linking these neurotransmitters and L-Dopa ethyl ester to 2-phenyl-3-carboxymethyl-imidazopyridine compounds giving rise to the so-called Dopimid compounds.Materials and MethodsA number of Dopimid compounds were synthesized and both stability and binding studies to dopaminergic and benzodiazepine receptors were performed. To evaluate whether Dopimid compounds are P-gp substrates, [3H]ritonavir uptake experiments and bi-directional transport studies on confluent MDCKII-MDR1 monolayers were carried out. The brain penetration properties of Dopimid compounds were estimated by the Clark’s computational model and evaluated by investigation of their transport across BBMECs monolayers. The dopamine levels following the intraperitoneal administration of the selected Dopimid compounds were measured in vivo by using brain microdialysis in rat.ResultsTested compounds were adequately stable in solution buffered at pH 7.4 but undergo faster cleavage in dilute rat serum at 37°C. Receptor binding studies showed that Dopimid compounds are essentially devoid of affinity for dopaminergic and benzodiazepine receptors. [3H]ritonavir uptake experiments indicated that selected Dopimid compounds, like L-Dopa and dopamine hydrochloride, are not substrates of P-gp and it was also confirmed by bi-directional transport experiments across MDCKII-MDR1 monolayers. By Clark’s model a significant brain penetration was deduced for L-Dopa ethyl ester and dopamine derivatives. Transport studies involving BBMECs monolayers indicated that some of these compounds should be able to cross the BBB. Interestingly, the rank order of apparent permeability (Papp) values observed in these assays parallels that calculated by the computational approach. Brain microdialysis experiments in rat showed that intraperitoneal acute administration of some Dopimid compounds induced a dose-dependent increase in cortical dopamine output.ConclusionBased on these results, it may be concluded that some Dopimid compounds can be proposed as novel L-Dopa and dopamine prodrugs.

A New Complex of Curcumin with Sulfobutylether‐β‐Cyclodextrin: Characterization Studies and In Vitro Evaluation of Cytotoxic and Antioxidant Activity on HepG‐2 Cells

Curcumin (CR) is a natural polyphenol with antioxidative, anti-inflammatory, and anticancer properties but its therapeutic potential is substantially hindered by the rather low-water solubility and bioavailability. Thus, in this work, a new soluble inclusion complex of CR with sulfobutylether-β-cyclodextrin (SBE-β-CD) was prepared in solution and at the solid state using different preparation techniques and characterized by Fourier transform infrared, nuclear magnetic resonance, differential scanning calorimetry, scanning electron microscopy, phase solubility studies, and Jobs plot method. Results clearly indicate that CR reacts with SBE-β-CD to form a host-guest complex with an apparent formation constant of 1455 M(-1) . Moreover, SBE-β-CD strongly increases water solubility of CR (from 0.56 to 102.78 μg/mL, at 25°C), and lyophilization method seems to be the best preparation technique to obtain the complex at the solid state. Finally, an in vitro test on a human hepatic cancer cell line (HepG-2) shows that complexation positively influences CR anticancer and antioxidant activity.

Effect of 2-hydroxypropyl-β-cyclodextrin on the aqueous solubility of the anaesthetic agent propofol (2,6-diisopropylphenol)

The inclusion complexation of Propofol (2,6-diisopropylphenol), an intravenous anaesthetic agent in clinical use, with 2-hydroxypropyl-β-cyclodextrin (HPβCyD) was investigated for the purpose of improving the low aqueous solubility and developing a potentially acceptable aqueous formulation. The aqueous solubility of propofol was increased as a function of HPβCyD concentration affording a linear phase-solubility profile.