Multicolor Photoluminescent Carbon Dots à La Carte for Biomedical Applications.

Dual-emission fluorescence probes that provide high sensitivity are key for biomedical diagnostic applications. Nontoxic carbon dots (CDs) are an emerging alternative to traditional fluorescent probes; however, robust and reproducible synthetic strategies are still needed to access materials with controlled emission profiles and improved fluorescence quantum yields (FQYs). Herein, we report a practical and general synthetic strategy to access dual-emission CDs with FQYs as high as 0.67 and green/blue, yellow/blue, or red/blue excitation-dependent emission profiles using common starting materials such as citric acid, cysteine, and co-dopants to bias the synthetic pathway. Structural and physicochemical analysis using nuclear magnetic resonance, absorbance and fluorescence spectroscopy, Fourier-transform infrared spectroscopy, and X-ray photoelectron spectroscopy in addition to transmission electron and atomic force microscopy (TEM and AFM) is used to elucidate the material's composition which is responsible for the unique observed photoluminescence properties. Moreover, the utility of the probes is demonstrated in the clinical setting by the synthesis of green/blue emitting antibody-CD conjugates which are used for the immunohistochemical staining of human brain tissues of glioblastoma patients, showing detection under two different emission channels.

Carbon-based glyco-nanoplatforms: towards the next generation of glycan-based multivalent probes.

Cell surface carbohydrates mediate a wide range of carbohydrate-protein interactions key to healthy and disease mechanisms. Many of such interactions are multivalent in nature and in order to study these processes at a molecular level, many glycan-presenting platforms have been developed over the years. Among those, carbon nanoforms such as graphene and their derivatives, carbon nanotubes, carbon dots and fullerenes, have become very attractive as biocompatible platforms that can mimic the multivalent presentation of biologically relevant glycosides. The most recent examples of carbon-based nanoplatforms and their applications developed over the last few years to study carbohydrate-mediate interactions in the context of cancer, bacterial and viral infections, among others, are highlighted in this review.

A 3D Peptide/[60]Fullerene Hybrid for Multivalent Recognition.

Fully substituted peptide/[60]fullerene hexakis-adducts offer an excellent opportunity for multivalent protein recognition. In contrast to monofunctionalized fullerene hybrids, peptide/[60]fullerene hexakis-adducts display multiple copies of a peptide in close spatial proximity and in the three dimensions of space. High affinity peptide binders for almost any target can be currently identified by in vitro evolution techniques, often providing synthetically simpler alternatives to natural ligands. However, despite the potential of peptide/[60]fullerene hexakis-adducts, these promising conjugates have not been reported to date. Here we present a synthetic strategy for the construction of 3D multivalent hybrids that are able to bind with high affinity the E-selectin. The here synthesized fully substituted peptide/[60]fullerene hybrids and their multivalent recognition of natural receptors constitute a proof of principle for their future application as functional biocompatible materials.

Structure-activity relationship studies on divalent naphthalene diimide G quadruplex ligands with anticancer and antiparasitic activity.

Naphthalene diimide (NDI) is a central scaffold that has been commonly used in the design of G-quadruplex (G4) ligands. Previous work revealed notable anticancer activity of a disubstituted N-methylpiperazine propyl NDI G4 ligand. Here, we explored structure-activity relationship studies around ligand bis-N,N-2,7-(3-(4-methylpiperazin-1-yl)propyl)-1,4,5,8-naphthalenetetracarboxylic diimide, maintaining the central NDI core whilst modifying the spacer and the nature of the cationic groups. We prepared new disubstituted NDI derivatives of the original compound and examined their in vitro antiproliferative and antiparasitic activity. Several N-methylpiperazine propyl NDIs showed sub-micromolar activity against Trypanosoma brucei and Leishmania major parasites with up to 30 fold selectivity versus MRC-5 cells. The best compound was a dimorpholino NDI with an IC50 of 0.17 µM against T.brucei and 40 fold selectivity versus MRC-5 cells. However, no clear correlation between G4 binding of the new NDI derivatives and antiproliferative or antiparasitic activity was observed, indicating that other mechanisms of action may be responsible for the observed biological activity.

Carbon dot-based fluorescent antibody nanoprobes as brain tumour glioblastoma diagnostics.

The development of efficient and sensitive tools for the detection of brain cancer in patients is of the utmost importance particularly because many of these tumours go undiagnosed until the disease has advanced and when treatment is less effective. Current strategies employ antibodies (Abs) to detect Glial Fibrillary Acid Protein (GFAP) in tissue samples, since GFAP is unique to the brain and not present in normal peripheral blood, and it relies on fluorescent reporters. Herein we describe a low cost, practical and general method for the labelling of proteins and antibodies with fluorescent carbon dots (CD) to generate diagnostic probes that are robust, photostable and applicable to the clinical setting. The two-step protocol relies on the conjugation of a dibenzocyclooctyne (DBCO)-functionalised CD with azide functionalised proteins by combining amide conjugation and strain promoted alkyne-azide cycloaddition (SPAAC) ligation chemistry. The new class of Ab-CD conjugates developed using this strategy was successfully used for the immunohistochemical staining of human brain tissues of patients with glioblastoma (GBM) validating the approach. Overall, these novel fluorescent probes offer a promising and versatile strategy in terms of costs, photostability and applicability which can be extended to other Abs and protein systems.

Immunomodulatory Response of Toll-like Receptor Ligand-Peptide Conjugates in Food Allergy.

Covalent conjugation of allergens to toll-like receptor (TLR) agonists appears to be a powerful strategy for the development of safety compounds for allergen-specific immunomodulatory response toward tolerance in allergy. In this work, we have synthesized two family of ligands, an 8-oxoadenine derivative as a ligand for TLR7 and a pyrimido[5,4-b]indole as a ligand for TLR4, both conjugated with a T-cell peptide of Pru p 3 allergen, the lipid transfer protein (LTP) responsible for LTP-dependent food allergy. These conjugates interact with dendritic cells, inducing their specific maturation, T-cell proliferation, and cytokine production in peach allergic patients. Moreover, they increased the Treg-cell frequencies in these patients and could induce the IL-10 production. These outcomes were remarkable in the case of the TLR7 ligand conjugated with Pru p 3, opening the door for the potential application of these allergen-adjuvant systems in food allergy immunotherapy.

Photoresponsive Control of G-Quadruplex DNA Systems.

G-quadruplex (G4) oligonucleotide secondary structures have recently attracted significant attention as therapeutic targets owing to their occurrence in human oncogene promoter sequences and the genome of pathogenic organisms. G4s also demonstrate interesting catalytic activities in their own right, as well as the ability to act as scaffolds for the development of DNA-based materials and nanodevices. Owing to this diverse range of opportunities to exploit G4 in a variety of applications, several strategies to control G4 structure and function have emerged. Interrogating the role of G4s in biology requires the delivery of small-molecule ligands that promote its formation under physiological conditions, while exploiting G4 in the development of responsive nanodevices is normally achieved by the addition and sequestration of the metal ions required for the stabilization of the folded structure. Although these strategies prove successful, neither allows the system in question to be controlled externally. Meanwhile, light has proven to be an attractive means for the control of DNA-based systems as it is noninvasive, can be delivered with high spatiotemporal precision, and is orthogonal to many chemical and biological processes. A plethora of photoresponsive DNA systems have been reported to date; however, the vast majority deploy photoreactive moieties to control the stability and assembly of duplex DNA hybrids. Despite the unique opportunities afforded by the regulation of G-quadruplex formation in biology, catalysis, and nanotechnology, comparatively little attention has been devoted to the design of photoresponsive G4-based systems. In this Perspective, we consider the potential of photoresponsive G4 assemblies and examine the strategies that may be used to engineer these systems toward a variety of applications. Through an overview of the main developments in the field to date, we highlight recent progress made toward this exciting goal and the emerging opportunities that remain ripe for further exploration in the coming years.

Visible-light photoswitching of ligand binding mode suggests G-quadruplex DNA as a target for photopharmacology.

We report the selective targeting of telomeric G4 DNA with a dithienylethene ligand and demonstrate the robust visible-light mediated switching of the G4 ligand binding mode and G-tetrad structure in physiologically-relevant conditions. The toxicity of the ligand to cervical cancer cells is modulated by the photoisomeric state of the ligand, indicating for the first time the potential of G4 to serve as a target for photopharmacological strategies.

Peptide Glycodendrimers as Potential Vaccines for Olive Pollen Allergy.

Olive pollen is one of the most important causes of respiratory allergy, with Ole e 1 being the most clinically relevant sensitizing allergen. Peptide-based vaccines represent promising therapeutic approaches, but the use of adjuvants is required to strengthen the weak immunogenicity of small peptides. We propose the use of dendrimeric scaffolds conjugated to the T cell immunodominant epitope of Ole e 1 (OE109-130) for the development of novel vaccines against olive pollen allergy. Four dendrimeric scaffolds containing an ester/ether with nine mannoses, an ester succinimidyl linker with nine N-acetyl-glucosamine units or nine ethylene glycol units conjugated to OE109-130 peptide were designed, and their cytotoxicity, internalization pattern, and immunomodulatory properties were analyzed in vitro. None of the dendrimers exhibited cytotoxicity in humanized rat basophil (RBL-2H3), human bronchial epithelial Calu-3, and human mast LAD2 cell lines. Confocal images indicated that mannosylated glycodendropeptides exhibited lower colocalization with a lysosomal marker. Moreover, mannosylated glycodendropeptides showed higher transport tendency through the epithelial barrier formed by Calu-3 cells cultured at the air-liquid interface. Finally, mannosylated glycodendropeptides promoted Treg and IL10+Treg proliferation and IL-10 secretion by peripheral blood mononuclear cells from allergic patients. Mannosylated dendrimers conjugated with OE109-130 peptide from Ole e 1 have been identified as suitable candidates for the development of novel vaccines of olive pollen allergy.