Chemical and Biomolecular Engineering - Research Publications
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ItemTracking the Endosomal Escape of Nanoparticles in Live Cells Using a Triplex-Forming Oligonucleotide(Wiley, 2024)Nanoparticle-mediated intracellular delivery of oligonucleotides is a complex phenomenon that depends on the architecture and the intracellular trafficking of the engineered nanoparticles. Unravelling the molecular arrangements of oligonucleotides within the nanoparticles as well as their intracellular behavior are essential for designing effective nucleic acid delivery systems. Herein, a simple and general strategy for probing the endosomal escape of nanoparticles carrying oligonucleotides in live cells is reported. A triplex-forming oligonucleotide probe is designed to target the transcription factor, kappa-light-chain-enhancer of activated B cells (NF-κB), in the cytosol of cells and to transduce the binding into a fluorescent Förster resonance energy transfer (FRET) signal. The combined use of the triplex-forming oligonucleotide probe and super-resolution microscopy enables the elucidation of the morphology, intracellular localization, and endosomal escape of the oligonucleotide-loaded nanoparticles on a molecular level and with nanoscale resolution. The co-delivery of the FRET probe and mRNA in cells via lipid- and polymer- based nanoparticles allow simultaneous correlation of the endosomal escape properties of nanoparticles and gene expression efficiency.
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ItemEmerging Strategies for Immunotherapy of Solid Tumors Using Lipid-Based Nanoparticles(WILEY, 2024-02)The application of lipid-based nanoparticles for COVID-19 vaccines and transthyretin-mediated amyloidosis treatment have highlighted their potential for translation to cancer therapy. However, their use in delivering drugs to solid tumors is limited by ineffective targeting, heterogeneous organ distribution, systemic inflammatory responses, and insufficient drug accumulation at the tumor. Instead, the use of lipid-based nanoparticles to remotely activate immune system responses is an emerging effective strategy. Despite this approach showing potential for treating hematological cancers, its application to treat solid tumors is hampered by the selection of eligible targets, tumor heterogeneity, and ineffective penetration of activated T cells within the tumor. Notwithstanding, the use of lipid-based nanoparticles for immunotherapy is projected to revolutionize cancer therapy, with the ultimate goal of rendering cancer a chronic disease. However, the translational success is likely to depend on the use of predictive tumor models in preclinical studies, simulating the complexity of the tumor microenvironment (e.g., the fibrotic extracellular matrix that impairs therapeutic outcomes) and stimulating tumor progression. This review compiles recent advances in the field of antitumor lipid-based nanoparticles and highlights emerging therapeutic approaches (e.g., mechanotherapy) to modulate tumor stiffness and improve T cell infiltration, and the use of organoids to better guide therapeutic outcomes.
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ItemYAP Signaling Regulates the Cellular Uptake and Therapeutic Effect of Nanoparticles(Wiley, 2023-11-09)Interactions between living cells and nanoparticles are extensively studied to enhance the delivery of therapeutics. Nanoparticles size, shape, stiffness, and surface charge are regarded as the main features able to control the fate of cell-nanoparticle interactions. However, the clinical translation of nanotherapies has so far been limited, and there is a need to better understand the biology of cell-nanoparticle interactions. This study investigates the role of cellular mechanosensitive components in cell-nanoparticle interactions. It is demonstrated that the genetic and pharmacologic inhibition of yes-associated protein (YAP), a key component of cancer cell mechanosensing apparatus and Hippo pathway effector, improves nanoparticle internalization in triple-negative breast cancer cells regardless of nanoparticle properties or substrate characteristics. This process occurs through YAP-dependent regulation of endocytic pathways, cell mechanics, and membrane organization. Hence, the study proposes targeting YAP may sensitize triple-negative breast cancer cells to chemotherapy and increase the selectivity of nanotherapy.
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ItemSupramolecular Assembly of Polyphenols and Nucleic Acids by Thermal Cycling for Immune Cell Activation(American Chemical Society (ACS), 2023)Supramolecular assembly of polyphenols and biomacromolecules (proteins and nucleic acids) has emerged as a versatile and simple strategy to construct nanomaterials with biological activity. Here, we report a strategy to finely control the supramolecular assembly of tannic acid and oligonucleotides into uniform and stable nanoparticles by exploiting the thermal cycling of tannic acid. The equilibrium of complexation is investigated, and individual nanoparticles are resolved with nanoscale resolution by using stochastic optical reconstruction microscopy. The nanoparticles incorporating cytosine phosphoguanine (CpG) oligonucleotides are efficiently taken up by cells and trafficked via endo/lysosomal compartments and induce up to a 7-fold increase in tumor necrosis factor secretion in RAW 264.7 macrophage cells compared with naked CpG oligonucleotides. This work highlights the potential of this simple approach to engineer two-component tannic acid–oligonucleotide nanoparticles for the intracellular delivery of therapeutic nucleic acids.
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ItemGeneration and maturation of human iPSC-derived 3D organotypic cardiac microtissues in long-term culture(NATURE PORTFOLIO, 2022-10-18)Cardiovascular diseases remain the leading cause of death worldwide; hence there is an increasing focus on developing physiologically relevant in vitro cardiovascular tissue models suitable for studying personalized medicine and pre-clinical tests. Despite recent advances, models that reproduce both tissue complexity and maturation are still limited. We have established a scaffold-free protocol to generate multicellular, beating human cardiac microtissues in vitro from hiPSCs-namely human organotypic cardiac microtissues (hOCMTs)-that show some degree of self-organization and can be cultured for long term. This is achieved by the differentiation of hiPSC in 2D monolayer culture towards cardiovascular lineage, followed by further aggregation on low-attachment culture dishes in 3D. The generated hOCMTs contain multiple cell types that physiologically compose the heart and beat without external stimuli for more than 100 days. We have shown that 3D hOCMTs display improved cardiac specification, survival and metabolic maturation as compared to standard monolayer cardiac differentiation. We also confirmed the functionality of hOCMTs by their response to cardioactive drugs in long-term culture. Furthermore, we demonstrated that they could be used to study chemotherapy-induced cardiotoxicity. Due to showing a tendency for self-organization, cellular heterogeneity, and functionality in our 3D microtissues over extended culture time, we could also confirm these constructs as human cardiac organoids (hCOs). This study could help to develop more physiologically-relevant cardiac tissue models, and represent a powerful platform for future translational research in cardiovascular biology.
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ItemChemoenzymatic surface decoration of Nisin-shelled nanoemulsions: Novel targeted drug-nanocarriers for cancer applications(ELSEVIER, 2022-11)Nisin, a peptide used as a natural food preservative, is employed in this work for the development of a novel nanocarrier system. Stable and uniform nisin-shelled nanoemulsions (NSNE) with a diameter of 100 ± 20 nm were successfully prepared using 20 kHz flow-through ultrasonication technique. The NSNE showed limited toxicity, high bactericidal activity and high drug loading capacity (EE 65 % w/w). In addition, the nisin shell was exploited for the site-specific attachment of a recombinantly produced cancer targeting ligand (αHER2LPETG IgG). Employing a unique two phases (bio-click) approach which involved both Sortase A mediated Azide Bioconjugation (SMAB) and Strain Promoted Azide Alkyne Cycloaddition (SPAAC) reactions, targeted NSNE (NSNEDOX-αHER2 IgG) were successfully assembled and loaded with the chemotherapeutic drug Doxorubicin (DOX). Finally, NSNEDOX-αHER2 IgG showed cancer-specific binding and augmented cytotoxicity to HER2 expressing tumour cells.
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ItemNo Preview AvailableSupramolecular Polyphenol‐DNA Microparticles for In Vivo Adjuvant and Antigen Co‐Delivery and Immune Stimulation(Wiley, 2023-03-13)Abstract DNA‐based materials have attracted interest due to the tunable structure and encoded biological functionality of nucleic acids. A simple and general approach to synthesize DNA‐based materials with fine control over morphology and bioactivity is important to expand their applications. Here, we report the synthesis of DNA‐based particles via the supramolecular assembly of tannic acid (TA) and DNA. Uniform particles with different morphologies are obtained using a variety of DNA building blocks. The particles enable the co‐delivery of cytosine‐guanine adjuvant sequences and the antigen ovalbumin in model cells. Intramuscular injection of the particles in mice induces antigen‐specific antibody production and T cell responses with no apparent toxicity. Protein expression in cells is shown using capsules assembled from TA and plasmid DNA. This work highlights the potential of TA as a universal material for directing the supramolecular assembly of DNA into gene and vaccine delivery platforms.
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ItemAn Engineered Nanosugar Enables Rapid and Sustained Glucose-Responsive Insulin Delivery in Diabetic Mice(WILEY-V C H VERLAG GMBH, 2023-05)Glucose-responsive insulin-delivery platforms that are sensitive to dynamic glucose concentration fluctuations and provide both rapid and prolonged insulin release have great potential to control hyperglycemia and avoid hypoglycemia diabetes. Here, biodegradable and charge-switchable phytoglycogen nanoparticles capable of glucose-stimulated insulin release are engineered. The nanoparticles are "nanosugars" bearing glucose-sensitive phenylboronic acid groups and amine moieties that allow effective complexation with insulin (≈95% loading capacity) to form nanocomplexes. A single subcutaneous injection of nanocomplexes shows a rapid and efficient response to a glucose challenge in two distinct diabetic mouse models, resulting in optimal blood glucose levels (below 200 mg dL-1 ) for up to 13 h. The morphology of the nanocomplexes is found to be key to controlling rapid and extended glucose-regulated insulin delivery in vivo. These studies reveal that the injected nanocomplexes enabled efficient insulin release in the mouse, with optimal bioavailability, pharmacokinetics, and safety profiles. These results highlight a promising strategy for the development of a glucose-responsive insulin delivery system based on a natural and biodegradable nanosugar.
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ItemSupramolecular Polyphenol-DNA Microparticles for In Vivo Adjuvant and Antigen Co-Delivery and Immune Stimulation(WILEY-V C H VERLAG GMBH, 2023-03-13)DNA-based materials have attracted interest due to the tunable structure and encoded biological functionality of nucleic acids. A simple and general approach to synthesize DNA-based materials with fine control over morphology and bioactivity is important to expand their applications. Here, we report the synthesis of DNA-based particles via the supramolecular assembly of tannic acid (TA) and DNA. Uniform particles with different morphologies are obtained using a variety of DNA building blocks. The particles enable the co-delivery of cytosine-guanine adjuvant sequences and the antigen ovalbumin in model cells. Intramuscular injection of the particles in mice induces antigen-specific antibody production and T cell responses with no apparent toxicity. Protein expression in cells is shown using capsules assembled from TA and plasmid DNA. This work highlights the potential of TA as a universal material for directing the supramolecular assembly of DNA into gene and vaccine delivery platforms.