Peripheral Functionalization of Dendrimers Regulates Internalization and Intracellular Trafficking in Living Cells

GATG (gallic acid-triethylene glycol) dendrimers represent appealing nanostructures for biomedical applications. The incorporation of specific ligands and targeting and imaging agents on their surface has resulted in promising tools in diagnosis and drug delivery. With the aim to further explore the versatility of GATG dendrimers in the biomedical field, in this work we study the effect of peripheral substitution on their uptake and intracellular trafficking in living cells. To this end, peripheral groups with different physicochemical properties and biological relevance have been installed on the surface of GATG dendrimers, and their interactions, uptake efficacy, and specificity for certain cell populations studied by confocal microscopy. Finally, this information was used to design a pH-sensitive drug delivery system for the selective release of cargo molecules inside cells after lysosomal localization. These results along with the easy functionalization and modular architecture of GATG dendrimers reveal these systems as promising nanotools in biomedicineThis work was financially supported by the Spanish MICINN (CTQ2009-10963 and CTQ2009-14146-C02-02) and the Xunta de Galicia (10CSA209021PR and CN2011/037). M.F.-V. thanks the Spanish Ministry of Education for a FPU fellowshipS

Microwave Assisted Organic Synthesis of Benzamides with C3 Symmetry

The 9th International Electronic Conference on Synthetic Organic Chemistry session Symposium on Microwave Assisted SynthesisXUNTA DE GALICIA for financial support: PR405 A 098/59-0 and PGIDIT05PXIB26201P

The Effect of PEGylation on Multivalent Binding: A Surface Plasmon Resonance and Isothermal Titration Calorimetry Study with Structurally Diverse PEG‐Dendritic GATG Copolymers

This is the peer-reviewed version of the following article: Fernandez-Villamarin, M., Sousa-Herves, A., Correa, J., Munoz, E., Taboada, P., Riguera, R., & Fernandez-Megia, E. (2016). The Effect of PEGylation on Multivalent Binding: A Surface Plasmon Resonance and Isothermal Titration Calorimetry Study with Structurally Diverse PEG-Dendritic GATG Copolymers. Chemnanomat, 2(5), 437-446, which has been published in final form at https://doi.org/10.1002/cnma.201600008. This article may be used for non-commercial purposes in accordance with Wiley-VCH Terms and Conditions for Self-ArchivingA general synthetic strategy to polyethylene glycol (PEG)‐dendritic block copolymers of the GATG (gallic acid–triethylene glycol) family is described from commercially available PEG of different molecular weights and architectures. Glycosylation of the resulting azide‐terminated copolymers with fucose by copper‐catalyzed azide–alkyne cycloaddition (CuAAC) afforded a toolbox to study the effect of PEG on the multivalent binding with the lectin UEA‐I by surface plasmon resonance (SPR, on surface) and isothermal titration calorimetry (ITC, in solution). Our results indicate that PEG reduces the affinity of glycodendrimers towards lectins by steric hindrance in a molecular‐weight‐dependent fashion. Great differences were observed as a function of the PEG architecture, with diblock PEG‐dendritic copolymers benefiting from a positive entropic contribution (PEG folding), not seen in the dendritic‐PEG‐dendritic systems. The self‐inflicted steric stabilization of the PEGylated copolymers onto lectin clusters reveals the necessity of additional competitive experiments to fully assess the antiadhesive properties of PEG in biological environmentsSpanish Government. Grant Numbers: CTQ2015-69021-R, CTQ2012-34790, CTQ2014-61470-EXP Xunta de Galicia. Grant Number: GRC2014/040S

A dendrimer–hydrophobic interaction synergy improves the stability of polyion complex micelles

This work was financially supported by the Spanish Ministry of Economy, Industry and Competitiveness (MINECO) (CTQ2015‐ 69021‐R), the Consellería de Cultura, Educación e Ordenación Universitaria (GRC2014/040 and Centro Singular de Investigación de Galicia Accreditation 2016‐2019, ED431G/09), and the European Regional Development Fund (ERDF). M.F.‐V. thanks the Spanish Government for a FPU FellowshipPolyion complex (PIC) micelles incorporating PEG-dendritic copolymers display an unprecedented stability towards ionic strength that is amplified via hydrophobic interactions. The tridimensional orientation of peripheral hydrophobic linkers between charged groups and the globular/rigid dendritic scaffold maximizes this stabilization compared to PIC micelles from linear polymers. As a result, micelles stable at concentrations higher than 3 M NaCl are obtained, which represents the highest saline concentration attained with PIC micelles. Advantages of this stabilizing dendritic effect have been taken for the design of a robust, pH-sensitive micelle for the controlled intracellular release of the anticancer drug doxorubicin. This micelle displays a slightly higher toxicity, and distinctive mechanisms of cell uptake and intracellular trafficking relative to the free drug. The preparation of mixed PIC micelles by combining differently functionalized PEG-dendritic block copolymers has allowed the fine-tuning of their stability, paving the way towards the facile modulation of properties like biodegradability, drug loading, or the response to external stimuliS

Dendrimers reduce toxicity of Aβ 1-28 peptide during aggregation and accelerate fibril formation

The influence of a GATG (gallic acid-triethylene glycol) dendrimer decorated with 27 terminal morpholine groups ([G3]-Mor) on the aggregation process of Alzheimer's peptide has been investigated. Amyloid fibrils were formed from the Aβ 1-28 peptide and the process was monitored by a ThT assay, changes in CD spectra, and transmission electron microscopy. In the presence of [G3]-Mor, more fibrils were built and the process significantly accelerated compared with a control. The cytotoxicity of (1) Aβ and (2) the system [G3]-Mor/Aβ was monitored at different stages of the aggregation process. Prefibrillar species were more toxic than mature fibrils. [G3]-Mor significantly reduced the toxicity of Aβ, probably because of lowering the amount of prefibrillar forms in the system by speeding up the process of fibril formationThis work was funded by the project “Biological Properties and Biomedical Applications of Dendrimers” operated within Foundation for Polish Science TEAM programme, cofinanced by the European Regional Development Fund. Financial support is also acknowledged from the Spanish MICINN (CTQ2009-10963 and CTQ2009-14146-C02-02) and the Xunta de Galicia (10CSA209021PR and CN2011/037). M.F-V. thanks the Spanish Ministry of Education for a FPU fellowship. The research was performed within the frame of the COST Action TD0802 “Dendrimers for biomedical applications.”S

Biodegradable PEG–dendritic block copolymers: synthesis and biofunctionality assessment as vectors of siRNA

One important drawback of most of the currently used dendrimers for biomedical applications is their high stability under physiological conditions that can result in cytotoxicity or complications induced by the accumulation of non-degradable synthetic materials in the organism. Particularly in the gene therapy field, vector stability can further hinder the intracellular release of the nucleic acid from the dendriplex, consequently leading to low transfection efficiencies. Therefore, biodegradable cationic dendritic structures have been eagerly awaited. However, the development of these dendritic nanocarriers is challenging because of the undesired and/or premature degradation observed during their synthesis and/or application. Here, we report new hybrid-biodegradable, biocompatible, non-toxic, and water-soluble azide-terminated PEG–GATGE dendritic block copolymers, based on a gallic acid (GA) core and triethylene glycol (TG) butanoate arms, incorporating ester bonds (E) at the dendritic arms/shell. Their successful functionalization by “click” chemistry with unprotected alkynated amines allowed complexation and delivery of siRNA. The hydrophobic character of the GATGE building unit confers to these hydrolyzable dendritic bionanomaterials a great ability to complex, protect and mediate the cellular internalization of siRNA. Moreover, the localization of the degradation points at the dendritic periphery, close to the complexed siRNA, was found to be important for nucleic acid release from the nanoparticles, rendering a significant improvement of the transfection efficiency compared to their hydrolytically stable PEG–GATG copolymer counterparts. The present study puts forward these biodegradable PEG–dendritic block copolymers not only as suitable vectors for nucleic acids, but also as new avenues for further developments exploring their use in theranosticsThe authors would like to acknowledge the FEDER funds through the Programa Operacional Factores de Competitividade – COMPETE and the Portuguese funds through FCT – Fundação para a Ciência e a Tecnologia (PTDC/CTM-NAN/112428/2009 and PTDC/CTM-NAN/3547/2014) that supported this work and the FCT / MEC through National Funds and, when applicable, co-financed by the FEDER via the PT2020 Partnership Agreement under the 4293 Unit I&D. V. Leiro acknowledges the support by FCT (SFRH/BPD/69110/2010) and by the project NORTE-01-0145-FEDER-000012, financed by Norte Portugal Regional Operational Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF). P.M.D. Moreno acknowledges the support from the Marie Curie Actions of the European Community’s Seventh Framework Program (PIEF-GA-2011-300485) and FCT fellowship (SFRH/BPD/108738/2015). This work was also financially supported by the Spanish Government (MINECO: CTQ2012-34790, CTQ2012-33436) and the Xunta de Galicia (CN2011/037)S

Mannose-modified hyaluronic acid nanocapsules for the targeting of tumor-associated macrophages

Tumor-associated macrophages (TAMs), a class of immune cells that play a key role in tumor immunosuppression, are recognized as important targets to improve cancer prognosis and treatment. Consequently, the engineering of drug delivery nanocarriers that can reach TAMs has acquired special relevance. This work describes the development and biological evaluation of a panel of hyaluronic acid (HA) nanocapsules (NCs), with different compositions and prepared by different techniques, designed to target macrophages. The results showed that plain HA NCs did not significantly influence the polarization of M0 and M2-like macrophages towards an M1-like pro-inflammatory phenotype; however, the chemical functionalization of HA with mannose (HA-Man) led to a significant increase of NCs uptake by M2 macrophages in vitro and to an improved biodistribution in a MN/MNCA1 fibrosarcoma mouse model with high infiltration of TAMs. These functionalized HA-Man NCs showed a higher accumulation in the tumor compared to non-modified HA NCs. Finally, the pre-administration of the liposomal liver occupying agent Nanoprimer™ further increased the accumulation of the HA-Man NCs in the tumor. This work highlights the promise shown by the HA-Man NCs to target TAMs and thus provides new options for the development of nanomedicine and immunotherapy-based cancer treatmentsOpen Access funding provided thanks to the CRUE-CSIC agreement with Springer Nature. This work was supported by the 2^2-INTRATARGET project (A20/00028) funded by the ISCIII under the umbrella of the ERA NET EuroNanoMed GA N 723770 of the EU Horizon 2020 Research and Innovation Programme. This work was also supported by the Xunta de Galicia (ED431C 2018/30, and “Centro singular de investigación de Galicia” accreditation 2019 − 2022, ED431G2019/03), and the European Union (European Regional Development Fund-ERDF)S