Novel Hyaluronic Acid-Chitosan Nanoparticles for Ocular Gene Therapy

purpose. Gene therapy offers a promising alternative for the treatment of ocular diseases. However, the implementation of this type of therapy is actually hampered by the lack of an efficient ocular gene delivery carrier. The main objective of the present work was to assess the effectiveness and investigate the mechanism of action of a new type of nanoparticle made of two bioadhesive polysaccharides, hyaluronic acid (HA) and chitosan (CS), intended for the delivery of genes to the cornea and conjunctiva. methods. The nanoparticles were obtained by a very mild ionotropic gelation technique. They were loaded with either the model plasmid pEGFP or pβ-gal. Transfection and toxicological studies were conducted in human corneal epithelial (HCE) and normal human conjunctival (IOBA-NHC) cell lines. The mechanism of internalization of the nanoparticles by the corneal and conjunctival cells was investigated by using fluorescence confocal microscopy. results. The nanoparticles had a size in the range of 100 to 235 nm and a ζ-potential of −30 to +28 mV. The results of the transfection studies showed that HA-CS nanoparticles were able to provide high transfection levels (up to 15% of cells transfected), without affecting cell viability. The confocal images indicated that HA-CS nanoparticles were internalized by fluid endocytosis and that this endocytic process was mediated by the hyaluronan receptor CD44. conclusions. The results give evidence of the potential of HA-CS nanoparticles for the targeting and further transfer of genes to the ocular surfaceS

Nanoengineering of vaccines using natural polysaccharides

Currently, there are over 70 licensed vaccines, which prevent the pathogenesis of around 30 viruses and bacteria. Nevertheless, there are still important challenges in this area, which include the development of more active, non-invasive, and thermo-resistant vaccines. Important biotechnological advances have led to safer subunit antigens, such as proteins, peptides, and nucleic acids. However, their limited immunogenicity has demanded potent adjuvants that can strengthen the immune response. Particulate nanocarriers hold a high potential as adjuvants in vaccination. Due to their pathogen-like size and structure, they can enhance immune responses by mimicking the natural infection process. Additionally, they can be tailored for non-invasive mucosal administration (needle-free vaccination), and control the delivery of the associated antigens to a specific location and for prolonged times, opening room for single-dose vaccination. Moreover, they allow co-association of immunostimulatory molecules to improve the overall adjuvant capacity. The natural and ubiquitous character of polysaccharides, together with their intrinsic immunomodulating properties, their biocompatibility, and biodegradability, justify their interest in the engineering of nanovaccines. In this review, we aim to provide a state-of-the-art overview regarding the application of nanotechnology in vaccine delivery, with a focus on the most recent advances in the development and application of polysaccharide-based antigen nanocarriers

A nano-enabled cancer-specific ITCH RNAi chemotherapy booster for pancreatic cancer

UNLABELLED: Gemcitabine is currently the standard therapy for pancreatic cancer. However, growing concerns over gemcitabine resistance mean that new combinatory therapies are required to prevent loss of efficacy with prolonged treatment. Here, we suggest that this could be achieved through co-administration of RNA interference agents targeting the ubiquitin ligase ITCH. Stable anti-ITCH siRNA and shRNA dendriplexes with a desirable safety profile were prepared using generation 3 poly(propylenimine) dendrimers (DAB-Am16). The complexes were efficiently taken up by human pancreatic cancer cells and produced a 40-60% decrease in ITCH RNA and protein expression in vitro (si/shRNA) and in a xenograft model of pancreatic cancer (shRNA). When co-administered with gemcitabine (100 mg/kg/week) at a subtherapeutic dose, treatment with ITCH-shRNA (3x 50 mg/week) was able to fully suppress tumour growth for 17 days, suggesting that downregulation of ITCH mediated by DAB-Am16/shRNA sensitizes pancreatic cancer to gemcitabine in an efficient and specific manner. FROM THE CLINICAL EDITOR: Gemcitabine delivery to pancreatic cancer often results in the common problem of drug resistance. This team overcame the problem through co-administration of siRNA and shRNA dendriplexes targeting the ubiquitin ligase ITCH

Sphingomyelin nanosystems decorated with TSP-1 derived peptide targeting senescent cells

Senescent cells accumulation can contribute to the development of several age-related diseases, including cancer. Targeting and eliminating senescence cells, would allow the development of new therapeutic approaches for the treatment of different diseases. The 4N1Ks peptide, a 10 amino acid peptide derived from TSP1 protein, combines both features by targeting the CD47 receptor present in the surface of senescent cells and demonstrating senolytic activity, thereby representing a new strategy to take into account. Nonetheless, peptide drugs are known for their biopharmaceutical issues, such as low short half-life and tendency to aggregate, which reduces their bioavailability and limits their therapeutic potential. In order to overcome this problem, herein we propose the use of biodegradable and biocompatible sphingomyelin nanosystems (SNs), decorated with this peptide for the targeting of senescent cells. In order to efficiently associate the 4N1Ks peptide to the nanosystems while exposing it on their surface for an effective targeting of senescent cells, the 4N1Ks peptide was chemically conjugated to a PEGylated hydrophobic chain. The resulting SNs-4N1Ks (SNs-Ks), were extensively characterized for their physicochemical properties, by dynamic light scattering, multiple-angle dynamic light scattering, nanoparticle tracking analysis and atomic force microscopy. The SNs-Ks demonstrated suitable features in terms of size (∼100 nm), association efficiency (87.2 ± 6.9%) and stability in different biorelevant media. Cell toxicity experiments in MCF7 cancer cells indicated an improved cytotoxic effect of SNs-Ks, decreasing cancer cells capacity to form colonies, with respect to free peptide, and an improved hemocompatibility. Lastly, senescence escape preliminary experiments demonstrated the improvement of SNs-Ks senolytic activity of in chemotherapy-induced senescence model of breast cancer cells. Therefore, these results demonstrate for the first time the potential of the combination of SNs with 4N1Ks peptide for the development of innovative senolytic therapies to battle cancerThis work was funded by Instituto de Salud Carlos III (ISCIII) and European Regional Development Fund (FEDER) (PI18/00176), and by the Axencia Galega de Coñecemento en Saúde (GAIN), Xunta de Galicia (IN607B2021/14). NANOMAG group belongs to Galician Competitive Research Group (GRC) (ED431C-2021/16), co-funded by FEDER (EU). R.J. also acknowledges the European financial support in the frame of the NanoFar program, an Erasmus Mundus Joint Doctorate program in nanomedicine and pharmaceutical innovationS

The Potential of Zebrafish as a Model Organism for Improving the Translation of Genetic Anticancer Nanomedicines

In the last few decades, the field of nanomedicine applied to cancer has revolutionized cancer treatment: several nanoformulations have already reached the market and are routinely being used in the clinical practice. In the case of genetic nanomedicines, i.e., designed to deliver gene therapies to cancer cells for therapeutic purposes, advances have been less impressive. This is because of the many barriers that limit the access of the therapeutic nucleic acids to their target site, and the lack of models that would allow for an improvement in the understanding of how nanocarriers can be tailored to overcome them. Zebrafish has important advantages as a model species for the study of anticancer therapies, and have a lot to offer regarding the rational development of efficient delivery of genetic nanomedicines, and hence increasing the chances of their successful translation. This review aims to provide an overview of the recent advances in the development of genetic anticancer nanomedicines, and of the zebrafish models that stand as promising tools to shed light on their mechanisms of action and overall potential in oncologyThis work was supported by the Xunta de Galicia, Spain (Competitive Reference Groups, GRC2014/010), the Carlos III Health Institute/FEDER (PI15/00828), and the Spanish Ministry of Education, Culture and Sport (FPU15/06595)S

Exploring the efficiency of gallic acid-based dendrimers and their block copolymers with PEG as gene carriers

The synthesis of a new family of amino-functionalized gallic acid-triethylene glycol (GATG) dendrimers and their block copolymers with polyethylene glycol (PEG) has recently being disclosed. In addition, these dendrimers have shown potential for gene delivery applications, as they efficiently complex nucleic acids and form small and homogeneous dendriplexes. On this basis, the present study aimed to explore the interaction of the engineered dendriplexes with blood components, as well as their stability, cytotoxicity and ability to enter and transfect mammalian cells. Results show that GATG dendrimers can form stable dendriplexes, protect the associated pDNA from degradation, and are biocompatible with HEK-293T cells and erythrocytes. More importantly, dendriplexes are effectively internalized by HEK-293T cells, which are successfully transfected. Besides, PEGylation has a marked influence on the properties of the resulting dendriplexes. While PEGylated GATG dendrimers have improved biocompatibility, the long PEG chains limit their uptake by HEK-293T cells, and thus, their ability to transfect them. As a consequence, the degree of PEGylation in dendriplexes containing dendrimer/block copolymer mixtures emerges as an important parameter to be modulated in order to obtain an optimized stealth formulation able to effectively induce the expression of the encoded proteinThe authors gratefully acknowledge support from the Spanish Ministry of Science and Innovation (SAF2004‐09230‐004‐01, CTQ2006‐12222/BQU, and CTQ2009‐10963) and the Xunta de Galicia (10CSA209021PR). M. Raviña and A. Sousa‐Herves also acknowledge fellowships from the Spanish Government (FPI and FPU, respectively)S

Selective interaction of PEGylated polyglutamic acid nanocapsules with cancer cells in a 3D model of a metastatic lymph node

Background Metastases are the most common reason of cancer death in patients with solid tumors. Lymph nodes, once invaded by tumor cells, act as reservoirs before cancer cells spread to distant organs. To address the limited access of intravenously infused chemotherapeutics to the lymph nodes, we have developed PEGylated polyglutamic acid nanocapsules (PGA-PEG NCs), which have shown ability to reach and to accumulate in the lymphatic nodes and could therefore act as nanotransporters. Once in the lymphatics, the idea is that these nanocapsules would selectively interact with cancer cells, while avoiding non-specific interactions with immune cells and the appearance of subsequent immunotoxicity. Results The potential of the PGA-PEG NCs, with a mean size of 100 nm and a negative zeta potential, to selectively reach metastatic cancer cells, has been explored in a novel 3D model that mimics an infiltrated lymph node. Our 3D model, a co-culture of cancer cells and lymphocytes, allows performing experiments under dynamic conditions that simulate the lymphatic flow. After perfusion of the nanocarriers, we observe a selective interaction with the tumor cells. Efficacy studies manifest the need to develop specific therapies addressed to treat metastatic cells that can be in a dormant state. Conclusions We provide evidence of the ability of PGA-PEG NCs to selectively interact with the tumor cells in presence of lymphocytes, highlighting their potential in cancer therapeutics. We also state the importance of designing precise in vitro models that allow performing mechanistic assays, to efficiently develop and evaluate specific therapies to confront the formation of metastasisThe authors acknowledge financial support given by the Carlos III Health Institute (ISCIII) and European Regional Development Fund (FEDER) (CP12/03150, PIE13/00024 and PI15/00828), ERA-NET EuroNanoMed 2009 (Lymphotarg PI09/2670) and EuroNanoMed 2013 (053 NICHE). The first author also acknowledges a fellowship received from the Fundación Ramón Domínguez, Spain. Abellan-Pose also acknowledges a fellowship from the Biomedical Sciences and Health Technologies Doctoral School-University of Santiago de Compostela (Spain)S

Hydroxychloroquine is associated with a lower risk of polyautoimmunity: data from the RELESSER Registry

Objectives. This article estimates the frequency of polyautoimmunity and associated factors in a large retrospective cohort of patients with SLE. Methods. RELESSER (Spanish Society of Rheumatology Lupus Registry) is a nationwide multicentre, hospital-based registry of SLE patients. This is a cross-sectional study. The main variable was polyautoimmunity, which was defined as the co-occurrence of SLE and another autoimmune disease, such as autoimmune thyroiditis, RA, scleroderma, inflammatory myopathy and MCTD. We also recorded the presence of multiple autoimmune syndrome, secondary SS, secondary APS and a family history of autoimmune disease. Multiple logistic regression analysis was performed to investigate possible risk factors for polyautoimmunity. Results. Of the 3679 patients who fulfilled the criteria for SLE, 502 (13.6%) had polyautoimmunity. The most frequent types were autoimmune thyroiditis (7.9%), other systemic autoimmune diseases (6.2%), secondary SS (14.1%) and secondary APS (13.7%). Multiple autoimmune syndrome accounted for 10.2% of all cases of polyautoimmunity. A family history was recorded in 11.8%. According to the multivariate analysis, the factors associated with polyautoimmunity were female sex [odds ratio (95% CI), 1.72 (1.07, 2.72)], RP [1.63 (1.29, 2.05)], interstitial lung disease [3.35 (1.84, 6.01)], Jaccoud arthropathy [1.92 (1.40, 2.63)], anti-Ro/SSA and/or anti-La/SSB autoantibodies [2.03 (1.55, 2.67)], anti-RNP antibodies [1.48 (1.16, 1.90)], MTX [1.67 (1.26, 2.18)] and antimalarial drugs [0.50 (0.38, 0.67)]. Conclusion. Patients with SLE frequently present polyautoimmunity. We observed clinical and analytical characteristics associated with polyautoimmunity. Our finding that antimalarial drugs protected against polyautoimmunity should be verified in future studies

Hyperkalemia in Heart Failure Patients in Spain and Its Impact on Guidelines and Recommendations: ESC-EORP-HFA Heart Failure Long-Term Registry

[Abstract] Introduction and objectives: Hyperkalemia is a growing concern in the treatment of patients with heart failure and reduced ejection fraction because it limits the use of effective drugs. We report estimates of the magnitude of this problem in routine clinical practice in Spain, as well as changes in potassium levels during follow-up and associated factors. Methods: This study included patients with acute (n=881) or chronic (n=3587) heart failure recruited in 28 Spanish hospitals of the European heart failure registry of the European Society of Cardiology and followed up for 1 year. Various outcomes were analyzed, including changes in serum potassium levels and their impact on treatment. Results: Hyperkalemia (K+> 5.4 mEq/L) was identified in 4.3% (95%CI, 3.7%-5.0%) and 8.2% (6.5%-10.2%) of patients with chronic and acute heart failure, respectively, and was responsible for 28.9% of all cases of contraindication to mineralocorticoid receptor antagonist use and for 10.8% of all cases of failure to reach the target dose. Serum potassium levels were not recorded in 291 (10.8%) of the 2693 chronic heart failure patients with reduced ejection fraction. During follow-up, potassium levels increased in 179 of 1431 patients (12.5%, 95%CI, 10.8%-14.3%). This increase was directly related to age, diabetes, and history of stroke and was inversely related to history of hyperkalemia. Conclusions: This study highlights the magnitude of the problem of hyperkalemia in patients with heart failure in everyday clinical practice and the need to improve monitoring of this factor in these patients due to its interference with the possibility of receiving optimal treatment.[Resumen] Introducción y objetivos. La hiperpotasemia es una preocupación creciente en el tratamiento de los pacientes con insuficiencia cardiaca y fracción de eyección reducida, pues limita el uso de fármacos eficaces. Este trabajo ofrece estimaciones de la magnitud de este problema en la práctica clínica habitual en España, los cambios en las concentraciones de potasio en el seguimiento y los factores asociados. Métodos. Pacientes con insuficiencia cardiaca aguda (n = 881) y crónica (n = 3.587) seleccionados en 28 hospitales españoles del registro europeo de insuficiencia cardiaca de la European Society of Cardiology y seguidos 1 año para diferentes desenlaces, incluidos cambios en las cifras de potasio y su impacto en el tratamiento. Resultados. La hiperpotasemia (K+ > 5,4 mEq/l) está presente en el 4,3% (IC95%, 3,7-5,0%) y el 8,2% (6,5-10,2%) de los pacientes con insuficiencia cardiaca crónica y aguda; causa el 28,9% de todos los casos en que se contraindica el uso de antagonistas del receptor de mineralocorticoides y el 10,8% de los que no alcanzan la dosis objetivo. Del total de 2.693 pacientes ambulatorios con fracción de eyección reducida, 291 (10,8%) no tenían registrada medición de potasio. Durante el seguimiento, 179 de 1.431 (12,5%, IC95%, 10,8-14,3%) aumentaron su concentración de potasio, aumento relacionado directamente con la edad, la diabetes mellitus y los antecedentes de ictus e inversamente con los antecedentes de hiperpotasemia. Conclusiones. Este trabajo destaca el problema de la hiperpotasemia en pacientes con insuficiencia cardiaca de la práctica clínica habitual y la necesidad de continuar y mejorar la vigilancia de este factor en estos pacientes por su interferencia en el tratamiento óptimo

Genome-wide pathway analysis identifies VEGF pathway association with oral ulceration in systemic lupus erythematosus

Background: Systemic lupus erythematosus (SLE) is a genetically complex rheumatic disease characterized by heterogeneous clinical manifestations of unknown etiology. Recent studies have suggested the existence of a genetic basis for SLE heterogeneity. The objective of the present study was to identify new genetic variation associated with the clinically relevant phenotypes in SLE. Methods: A two-stage pathway-based approach was used to identify the genetic variation associated with the main clinical phenotypes in SLE. In the discovery stage, 482 SLE patients were genotyped using Illumina Human Quad610 microarrays. Association between 798 reference genetic pathways from the Molecular Signatures Database and 11 SLE phenotypes was tested using the set-based method implemented in PLINK software. Pathways significantly associated after multiple test correction were subsequently tested for replication in an independent cohort of 425 SLE patients. Using an in silico approach, we analyzed the functional effects of common SLE therapies on the replicated genetic pathways. The association of known SLE risk variants with the development of the clinical phenotypes was also analyzed. Results: In the discovery stage, we found a significant association between the vascular endothelial growth factor (VEGF) pathway and oral ulceration (P value for false discovery rate (P FDR) < 0.05), and between the negative regulation signaling pathway of retinoic acid inducible gene-I/melanoma differentiation associated gene 5 and the production of antinuclear antibodies (P FDR < 0.05). In the replication stage, we validated the association between the VEGF pathway and oral ulceration. Therapies commonly used to treat mucocutaneous phenotypes in SLE were found to strongly influence VEGF pathway gene expression (P = 4.60e-4 to 5.38e-14). Analysis of known SLE risk loci identified a strong association between PTPN22 and the risk of hematologic disorder and with the development of antinuclear antibodies. Conclusions: The present study has identified VEGF genetic pathway association with the risk of oral ulceration in SLE. New therapies targeting the VEGF pathway could be more effective in reducing the severity of this phenotype. These findings represent a first step towards the understanding of the genetic basis of phenotype heterogeneity in SLE