M.E.M. Cruz

Intravenous administration of superoxide dismutase entrapped in long circulating liposomes. II. In vivo fate in a rat model of adjuvant arthritis.

Rheumatoid arthritis (RA) is a prevalent and debilitating autoimmune disease that affects the joints. RA is characterized by an infiltration of the affected joint by blood-derived cells. In response to activation, these cells generate reactive oxygen species, resulting in an oxidative stress situation. One approach to counteract this oxidative stress situation is the use of antioxidants as therapeutic agents. The free radical scavenger enzyme superoxide dismutase (SOD) may be used as a therapeutic agent in rheumatoid arthritis, but its rapid elimination from the circulation is a major limitation. Targeted delivery of SOD may overcome this limitation. In this study, the utility of PEGylated liposomes (PEG-liposomes) for targeting SOD to arthritic sites was explored. The targeting of SOD to arthritic sites following intravenous administration of both PEG-liposomes and positively charged liposomes lacking PEG but containing stearylamine (SA-liposomes) in rats with adjuvant arthritis was studied. At 24 h post injection, the blood levels of long circulating liposomes with a mean size of 0.11 micrometer and 0.20 micrometer were 8- and 3-fold higher, respectively, as compared to the SA-liposomes. The majority of SOD administered in liposomal form remains within the liposomes when they circulate in the bloodstream. The highest target uptake was observed with PEG-liposomes with a mean size of 0.11 micrometer and the lowest uptake with the SA-liposomes. These results demonstrate that SOD can be targeted to inflamed sites most efficiently via small-sized PEG-liposomes. Small-sized PEG-coated liposomes are to be preferred if prolonged circulation and enhanced localization of SOD at arthritic sites are desired.

Lipid nanoparticles containing oryzalin for the treatment of leishmaniasis

Oryzalin is a dinitroaniline drug that has attracted recent interest for the treatment of leishmaniasis. Its use as an antiparasitic therapeutic agent is limited by the low water solubility associated with an in vivo rapid clearance, leading to the administration of larger and possibly toxic doses in in vivo studies, and the use of solvents that may lead to undesirable side effects. In the present work oryzalin-containing lipid nanoparticles were produced by a emulsion-solvent evaporation technique using a composition suitable for parenteral administration, i.e., tripalmitin (solid lipid) and a complex mixture of three emulsifying agents (soya lecithin, Tween® 20 and sodium deoxycholate). Physicochemical characterization included the determination of mean particle size, polydispersity index, zeta potential, encapsulation efficiency and DSC studies. Final formulations revealed values of <140 nm (PI<0.2) and zeta potential of ≈-35 mV, as well as encapsulation efficiency >75%. The effects of various processing parameters, such as lipid and surfactant and composition and concentration, as well as the stability during the harsh procedures of autoclaving (121°C/15 min) and freeze-drying were also evaluated. Formulations revealed to be stable throughout freeze-drying and moist-heath sterilization without significant variations on physicochemical properties and no significant oryzalin losses. The use of a complex surfactant mixture proved crucial for preserving formulation stability. Particularly, lecithin appears as a key component in the stabilization of tripalmitin-based oryzalin-containing lipid nanoparticles. Finally, cell viability studies demonstrated that the incorporation of oryzalin in nanoparticles decreases cytotoxicity, thus suggesting this strategy may improve tolerability and therapeutic index of dinitroanilines.

Targeted delivery of paromomycin in murine infectious diseases through association to nano lipid systems.

UNLABELLEDnTreatment of intracellular infections such as those caused by Mycobacterium spp. and Leishmania spp. is often hampered by limited access of drugs to infected cells. This is the case of paromomycin (PRM), an antibiotic with broad spectrum in vitro activity against protozoa and mycobacteria. Association of chemotherapeutics to liposomes is a worthy strategy to circumvent poor drug accessibility. Six different PRM liposomal formulations were produced, physicochemically characterized and biologically evaluated in a macrophagic cell line confirming their adequacy for in vivo studies. Biodistribution profiles of PRM liposomes revealed preferential targeting of the antibiotic to the liver, spleen and lungs, relative to free PRM, which translated into an enhanced therapeutic effect in murine models infected with Mycobacterium avium and Leishmania infantum and an absence of toxic effects. Our findings demonstrate the advantages of associating PRM to liposomes indicating their potential as an alternative therapeutic strategy for mycobacterial and parasite infections.nnnFROM THE CLINICAL EDITORnInfections caused by intracellular organisms such as Mycobacterium and Leishmania remain a significant problem worldwide. Although effective drugs are available, their actions are limited by access into the intracellular compartment. In this article, the authors developed different liposomal formulations as drug carriers of paromomycin and investigated their efficacy in a mouse model. The positive should provide another treatment option for these organisms in the near future.

Abstract A129: Targeted delivery of therapeutics to tumor cells and the tumor microenvironment

Human cancer is a complex disease caused by genetic instability and accumulation of multiple molecular alterations. Tumor growth and metastasis depend on the development of a neovasculature by a process called angiogenesis, which is controlled by numerous pro‐ and antiangiogenic factors. Therefore, vascular targeting emerges as an attractive therapeutic approach, since angiogenic blood vessels express distinctive molecular markers with direct access by the therapeutic agent. Moreover, ablation of immature angiogenic vessels causes reduction of the interstitial pressure gradient in the tumor periphery, improving drug delivery through the highly permeabilized vessel wall. Hence, additional therapeutic gains will be expected by “starving” tumor cells to death and by increasing the concentration of the therapeutic agent in the target cell. The present work, is aimed at investigating the ability of a system consisting of a ligand coupled to the extremity of poly(ethylene glycol)‐grafted liposomes, containing doxorubicin (DXR), to simultaneously target human tumor cells and endothelial cells of tumor blood vessels. Cellular association studies were conducted by flow cytometry, fluorimetry and confocal microscopy with rhodamine‐labeled liposomes. Competitive inhibition and mechanistic assays were also performed by fluorimetry. Cytotoxicity was evaluated with the MTT assay and biodistribution performed on Balb/c nude female mice bearing orthotopic implanted tumors was analyzed by radioactivity counts of a lipid and a drug tracer ([3H]CHE and [14C]DXR, respectively). Our results show a substantial increase in the levels of association for ligand‐targeted liposomes, in vitro as well as ex vivo, in tumor cells removed from patients, after mastectomy or tumorectomy. These results suggest that internalization occurs, most likely through the clathrin‐mediated endocytic pathway. Cytotoxicity studies show that ligand‐targeted liposomes are more cytotoxic than the control formulation (4 to 180‐fold), against tumor and endothelial cells, indicating that binding and internalization of the system are contributing to a more efficient delivery of the payload to the target cells. In vivo experiments report a higher tumor accumulation of the radiolabeled targeted system over non‐targeted liposomes (approximately 18‐fold). The observed improved tumor targeting is likely due to a strong vascular targeting component that is taking place in vivo. The data generated so far indicate that the same technological platform is able to target two distinct cell populations within the tumor niche. Overall, these results represent a novel and a valuable contribution for delivery strategies to the tumor and its microenvironment. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):A129.