Structural Characterization and Analyses of the Biological Properties of New Amphiphilic Block Copolymers

La necesidad de encontrar nuevos métodos diagnósticos y terapéuticos para diversas dolencias como el cáncer, las enfermedades cardiovasculares, la diabetes o enfermedades neurodegenerativas para las que no existen tratamientos definitivos, ha generado el progresivo aumento del número de investigaciones en Nanomedicina. Uno de sus grandes retos consiste en el desarrollo de “nanoterapias”, terapias basadas en materiales nanométricos que se puedan dirigir de forma selectiva a los tejidos y órganos enfermos, evitando así los efectos secundarios inevitables en los tratamientos actuales. Entre ellas caben destacar los sistemas de liberación controlada de fármacos, que consisten en utilizar nanoestructuras que transporten el fármaco hasta la zona dañada, y sólo cuando la han reconocido, lo liberen. Para ello, es necesario la previa encapsulación o protección del fármaco para que este sea inerte en su recorrido por el cuerpo y mantenga intactas sus propiedades. Una vez que el nanosistema ha llegado a su destino, debe liberar el fármaco a una velocidad apropiada para que sea efectivo, y luego permitir la expulsión del nanotransportador del cuerpo humano

Inhibiting HER3 Hyperphosphorylation in HER2‐Overexpressing Breast Cancer through Multimodal Therapy with Branched Gold Nanoshells

Treatment failure in breast cancers overexpressing human epidermal growth factor receptor 2 (HER2) is associated mainly to the upregulation of human epidermal growth factor receptor 3 (HER3) oncoprotein linked to chemoresitence. Therefore, to increase patient survival, here a multimodal theranostic nanoplatform targeting both HER2 and HER3 is developed. This consists of doxorubicin-loaded branched gold nanoshells functionalized with the near-infrared (NIR) fluorescent dye indocyanine green, a small interfering RNA (siRNA) against HER3, and the HER2-specific antibody Transtuzumab, able to provide a combined therapeutic outcome (chemo- and photothermal activities, RNA silencing, and immune response). In vitro assays in HER2+/HER3+ SKBR-3 breast cancer cells have shown an effective silencing of HER3 by the released siRNA and an inhibition of HER2 oncoproteins provided by Trastuzumab, along with a decrease of the serine/threonine protein kinase Akt (p-AKT) typically associated with cell survival and proliferation, which helps to overcome doxorubicin chemoresistance. Conversely, adding the NIR light therapy, an increment in p-AKT concentration is observed, although HER2/HER3 inhibitions are maintained for 72 h. Finally, in vivo studies in a tumor-bearing mice model display a significant progressively decrease of the tumor volume after nanoparticle administration and subsequent NIR light irradiation, confirming the potential efficacy of the hybrid nanocarrierE.V.-A. and I.G.-C. contributed equally to this work. This work was sup ported by the Agencia Estatal de Investigación (AEI) through Project No. PID2019-109517RB-I00) and from the Xunta de Galicia, Project No. ED431C2022/18. European Regional Development Funds are also ac knowledged. A.A.-M. and P.T. also thank the International Scientific Part nership Program ISSP at King Saud University for additional funding of this research through Grant No. ISPP-144. This work also received fi nancial support from the ISCIII, Ministerio de Economía y Competitivi dad (Grant No. PI15/01129; J.A.C.), and the AEI (Grant No. PID2020- 113501RB-I00; J.A.C.). I.-G.C. thanks for financial support through Grant No. PRE/2011/131, and the Centro Singular de Investigación de Galicia accreditation Grant No. 2016–2019 ED431G/05)S

Polymer stabilized, phenytoin-loaded nanomicelles as promising nanocarriers: In silico and in vitro evaluations

Many drugs currently used in clinics to treat different diseases show low solubilities in aqueous solutions and must be then administered with the aid of organic solvents within the designed formulations leading to harmful health effects. For such reason, drug delivery using nanocarriers appears as an excellent alternative since it improves the solubilization and controlled release of many different pharmaceuticals, favoring the decrease of the overall administered therapeutic doses and by diminishing the risk of potential associated adverse side effects. Among the different types of nanocarriers, those based on polymeric micelles are an excellent option since the polymer structure and composition may be tuned in order to regulate the characteristics and properties of the resulting nanoassemblies to maximize drug solubilization and release profiles. For this reason, in this paper a drug delivery nanosystem based on oil-in-water polymeric-based microemulsion used to load the anti-epileptic phenytoin drug (PHT) was developed in the oily phase. Two different biocompatible polymers were evaluated to form the nanomicelles and maximize encapsulation efficiencies and colloidal stability: The amphiphilic triblock copolymer Pluronic F127, which poses the ability to cross the blood brain barrier (BBB), and the natural lipophilic lignin, which bears antipathogenic properties. Density functional theory (DFT) calculations were performed with the 6-31G(d) basis set to elucidate the interactions of PHT with F127 and lignin monomers. Simulation data showed that hydrogen bonding (HB) interactions between PHT, F127 and lignin are the predominant force to allow for drug solubilization and stability. Atoms in molecules (AIM) and natural bond orbital (NBO) analyses were performed to evaluate the strength of such HB. and their drug encapsulation efficiency, release profiles, and antibacterial susceptibility were determined. Moreover, the cytotoxicity of the developed nanoformulations together with a morphological examination of PC12 and NIH cells after drug-loaded nanocarrier administration and subsequent uptake were also investigated. In this manner, the obtained nanocarriers were also characterized by dynamic light scattering (DLS) and zeta potential, showing a nanometer size (between ca. 16 and 22 nm) and surface negative charge. PHT loading into F127 and lignin nanomicelles were ca. 96.7 ± 1.5% and 68.2 ± 3.5%, respectively, and the drug release profile kinetics of F127/PHT-loaded nanomicelles was rather slower compared to that of lignin/PHT-loaded ones. On the other hand, in vitro cytotoxicity data confirmed the lack of any significant cytotoxicity of PHT-loaded nanomicelles in both NIH/3T3 and PC12 cell lines, but a slightly higher cell viability and well-preserved cell morphology was observed for PC12 cells compared to NIH/3T3 ones. After culturing in chocolate blood agar medium inoculated with F127/PHT-loaded and lignin/ PHT-loaded nanomicelles, pathogenic bacteria did not grow despite confirming certain antimicrobial character of the encapsulated drug. Hence, thanks to their excellent encapsulation and biocompatibility properties, these nanocarriers appear as an excellent option to configure new drug delivery nanocarriers of hydrophobic drugsThe authors would like to thank Dr. Maryam Abolhasani (Associate Professor of Pathology, Iran University of Medical Sciences). P.T. also thanks Agencia Estatal de Investigación (AEI) by project PID2019-109517RB-I00. ERDF funds are also acknowledgedS

New Approach for Cardiovascular Diseases (CVD) Treatment

Cardiovascular disease (CVD) is a general term that includes diseases that affect the circulatory system and/or the heart. Their underlying pathology is atherosclerosis, an inflammatory disease characterized by the accumulation of lipids, inflammatory cells and fibrous tissue in the arteries\u27 internal wall, provoking to some extent their obstruction. Atherosclerosis is still addressed as a simple disease instead of the complex interplay of different types of cells and cascade signaling pathways, so the use of any single imaging or therapeutic agent alone is unlikely to provide a satisfactory outcome. Hence, other treatment strategies need to be implemented, in particular, those using new nanomaterials able to target the plaque and to efficiently treat it, and that can be easily released by the body without provoking adverse effects. With this background, we have designed a biocompatible drug delivery vehicle that efficiently loads and protects the drug Atorvastatin (ATO reduces the LDL levels) while a folate receptor in the external shell targets inflamed areas. To avoid the common toxic effects of folic acid (FA) or ATO in the body at certain concentrations, the vehicle will provide covalent attachment for the FA on the surface and cage structure for ATO protection. To complement the treatment, genetic material will be included in a separate compartment to actively influence the regulation of immune responses and inflammatory disorders

Light excitation of gold Nanorod-Based hybrid nanoplatforms for simultaneous bimodal phototherapy

In this work, we developed a hybrid nanosystem able to simultaneously combine its potential as a photodynamic (PDT) and plasmonic photothermal (PPTT) therapeutic agent to kill malignant cells. To do that, AuNRs were functionalized by means of the layer-by-layer (LbL) assembly technique using alternating layers of anionic poly(styrene sulfonate) (PSS) and cationic poly-L-lysine (PLL) as polyelectrolytes (PEs), and an outer final layer of hyaluronic acid (HA) to provide the hybrid particles with both sufficient colloidal stability and targeting ability to tumoral cells overexpressing CD44 receptors. To provide the nanoplatform with PDT capabilities, the near-infrared (NIR) sensitive photosensitizer (PS) indocyanine green (ICG) was previously grafted to the PLL PE and assembled on the particle surface coating; in this manner, PSS/PLL-ICG/HA-coated AuNRs hybrid particles were obtained. Then, the conditions for optimized reactive oxygen species (ROS) production under NIR light excitation were elucidated, reaching ca. 80% after 5 min irradiation at 2.0 W/cm2 compared to positive control (H2O2). Since ICG can also absorb near infrared (NIR) light and transforms it by internal conversion into heat under suitable irradiation conditions, which adds to the photothermal plasmonic effect provided by the metallic NP, the heating profiles provided by the present hybrid nanoparticles (NPs) were measured, being observed temperature increments ranging from 9 to 22 °C. The potential contribution of its different components to both PDT and PPTT were deeply analyzed in vitro for intended cancer therapeutics at several power intensities but also at different temperatures with the aim of elucidating the role played by both phototherapies on cell cytotoxicity, the optimal illumination conditions for effective bimodal phototherapy as well as the elucidation of the main cell death mechanism involved. Under optimized conditions and after the administration of 2.5 ·1010 NPs/mL, cell cytotoxicites of up to ca. 70% were determined by combination of PDT + PPTT therapeutic effect.This research was funded by Agencia Estatal de Investigación (AEI), project number PID2019-109517RB-I00 and Xunta de Galicia ED431C 2022/18. ERDF funds are also acknowledged. A.C. also thanks Xunta de Galicia for grant number ED481D-2021–011. L.A. is grateful to CONACYT for funding her PhD fellowship.Peer reviewe

Hybrid Gold Nanorod-Based Nanoplatform with Chemo and Photothermal Activities for Bimodal Cancer Therapy

Metal nanoparticles (NPs), particularly gold nanorods (AuNRs), appear as excellent platforms not only to transport and deliver bioactive cargoes but also to provide additional therapeutic responses for diseased cells and tissues and/or to complement the action of the carried molecules. In this manner, here, we optimized a previous developed metal-based nanoplatform composed of an AuNR core surrounded by a polymeric shell constructed by means of the layer-by-layer approach, and in which very large amounts of the antineoplasic drug doxorubicin (DOXO) in a single loading step and targeting capability thanks to an outer hyaluronic acid layer were incorporated by means of an optimized fabrication process (PSS/DOXO/PLL/HA-coated AuNRs). The platform retained its nanometer size with a negative surface charge and was colloidally stable in a range of physiological conditions, in which only in some of them some particle clustering was noted with no precipitation. In addition, the dual stimuli-responsiveness of the designed nanoplatform to both endogenous proteases and external applied light stimuli allows to perfectly manipulate the chemodrug release rates and profiles to achieve suitable pharmacodynamics. It was observed that the inherent active targeting abilities of the nanoplatfom allow the achievement of specific cell toxicity in tumoral cervical HeLa cells, whilst healthy ones such as 3T3-Balb fibroblast remain safe and alive in agreement with the detected levels of internalization in each cell line. In addition, the bimodal action of simultaneous chemo- and photothermal bioactivity provided by the platform largely enhances the therapeutic outcomes. Finally, it was observed that our PSS/DOXO/PLL/HA-coated AuNRs induced cell mortality mainly through apoptosis in HeLa cells even in the presence of NIR light irradiation, which agrees with the idea of the chemo-activity of DOXO predominating over the photothermal effect to induce cell death, favoring an apoptotic pathway over necrosis for cell death