Development and evaluation of smart polymeric and lipidic nanoparticles for theranosis of breast and pancreatic cancer

Resumen

Nanomedicine is playing a growing part in pharmaceutical research and development, primarily in the form of nanoparticle-based delivery systems for drugs and imaging agents. The development of targeted therapies, especially for cancer, is one of the main goals of nanomedicine today. Conventional chemotherapy usually prompts modest tumor response and provokes undesirable side effects due to the nonspecific action of drugs on proliferating tissues. To avoid these and other disadvantages, drug nanocarriers should be formulated to deliver the antitumor drug directly to the cancerous cells. The main objective of the application of nanomedicine in cancer is to have a better therapeutic effect, to increase the bioavailability and to allow the administration of lower doses of drug while obtaining lower toxicity rates and improving the patient's quality of life. Another objective is to overcome the multiple mechanisms of drug resistance that make this treatment ineffective in a high percentage of cancer cases since cancer cells have the ability to evade cytotoxicity. This is a complex interdisciplinary task with too many variables to be properly controlled. These variables include the use of biocompatible materials, with simple but robust processes for biomaterial assembly, usually requiring different conjugation chemistries followed by some purification processes. Therefore, current formulations based on complex nanostructures such as polymer conjugates, polymeric micelles, liposomes, carbon nanotubes, or nanoparticles, must be superficially modified to provide carriers with vectorization properties. On the one hand, our group has developed several strategies for preparing functionalized polymeric cross-linked polystyrene NPs which are then covalently conjugated to cargoes of different nature. On the other hand, in recent years, lipid liquid nanocapsules (LLNCs) have been developed as potential nanocarriers. The inner hydrophobic domain, encased in a hydrophilic outer shell, has been used to encapsulate hydrophobic drugs that are protected during their transport to the target cells. Thus, the main objective of this thesis has been the development and evaluation of two different nanosystems in its composition, for the diagnosis and treatment of breast and pancreatic cancer. One of the nanosystems used has been polystyrene nanoparticles (NPs). A novel chemical-based orthogonal bioconjugation strategy to produce tri-functionalized (NPs) carrying doxorubicin (DOX), near-infrared cyanine dye (Cy7) and a homing peptide CRGDK, a peptide specifically binds to neuropilin-1 (Nrp-1) overexpressed on triple negative breast cancer (TNBC) cells, has been validated. These theranostic NPs have been evaluated in vitro and in vivo using an orthotopic xenotransplant mouse model using TNBC cells. In vitro assays show that theranostic NPs improve the therapeutic index in comparison with free DOX. Remarkably, in vivo studies showed preferred location of theranostic NPs in the tumor area reducing the volume at the same level than free DOX while presenting lower side effects. Another of the nanosystems used have been lipid liquid nanocapsules (LLN), specifically olive oil liquid nanocapsules. These LLNs have suffered several modifications. On the one hand, LLN covered by the protein human serum albumin (HSA) and loaded with curcumin as a hydrophobic model drug have been successfully developed. A cross-linking procedure with glutaraldehyde (GAD) was performed to further strengthen the protective protein layer. Physicochemical properties and release kinetics of the nanocapsules were investigated, and cellular uptake and killing capacity were evaluated on the human breast-cancer line MCF-7. The nanocapsules exhibited a killing capacity (IC50) similar to that of free curcumin, but avoiding the problems associated with excipients, and displayed an outstanding uptake performance, entering cells massively in less than 1 min. On the other hand, olive oil liquid nanocapsules (O2LNC) functionalized by covalent coupling of an anti-CD44-FITC antibody (αCD44) that can specifically target pancreatic CSCs (PCSCs) overexpressing the CD44 receptors have been successfully developed. Firstly, O2LNCs formed by a core of olive oil were surrounded by a shell containing phospholipids (Epikuron®), a non-ionic surfactant (Pluronic® F68) and deoxycholic acid molecules to provide a surface enriched in solvent-exposed free carboxylic acid functional groups. Then, O2LNCs were coated with an αCD44 antibody to the optimized formulation of immuno-nanocapsules (αCD44-O2LNC). The optimization of an αCD44 coating procedure, a complete physico-chemical characterization of these functionalized nanosystems, as well as clear evidence of their efficacy in vitro and in vivo, were demonstrated. Our results indicate the high targeted uptake of these αCD44-O2LNCs and the increased antitumor efficacy (up to four times) of paclitaxel-loaded αCD44-O2LNC compared to free paclitaxel in PCSCs. Also, αCD44-O2LNCs were able to selectively target PCSCs in an orthotopic xenotransplant in vivo model.