Biocompatible 2D Materials via Liquid Phase Exfoliation.

2D materials (2DMs), such as graphene, transition metal dichalcogenides (TMDs), and black phosphorus (BP), have been proposed for different types of bioapplications, owing to their unique physicochemical, electrical, optical, and mechanical properties. Liquid phase exfoliation (LPE), as one of the most effective up-scalable and size-controllable methods, is becoming the standard process to produce high quantities of various 2DM types as it can benefit from the use of green and biocompatible conditions. The resulting exfoliated layered materials have garnered significant attention because of their biocompatibility and their potential use in biomedicine as new multimodal therapeutics, antimicrobials, and biosensors. This review focuses on the production of LPE-assisted 2DMs in aqueous solutions with or without the aid of surfactants, bioactive, or non-natural molecules, providing insights into the possibilities of applications of such materials in the biological and biomedical fields.

Structural Transformation of Coassembled Fmoc-Protected Aromatic Amino Acids to Nanoparticles.

Materials made of assembled biomolecules such as amino acids have drawn much attention during the past decades. Nevertheless, research on the relationship between the chemical structure of building block molecules, supramolecular interactions, and self-assembled structures is still necessary. Herein, the self-assembly and the coassembly of fluorenylmethoxycarbonyl (Fmoc)-protected aromatic amino acids (tyrosine, tryptophan, and phenylalanine) were studied. The individual self-assembly of Fmoc-Tyr-OH and Fmoc-Phe-OH in water formed nanofibers, while Fmoc-Trp-OH self-assembled into nanoparticles. Moreover, when Fmoc-Tyr-OH or Fmoc-Phe-OH was coassembled with Fmoc-Trp-OH, the nanofibers were transformed into nanoparticles. UV-vis spectroscopy, Fourier transform infrared spectroscopy, and fluorescence spectroscopy were used to investigate the supramolecular interactions leading to the self-assembled architectures. π-π stacking and hydrogen bonding were the main driving forces leading to the self-assembly of Fmoc-Tyr-OH and Fmoc-Phe-OH forming nanofibers. Further, a mechanism involving a two-step coassembly process is proposed based on nucleation and elongation/growth to explain the structural transformation. Fmoc-Trp-OH acted as a fiber inhibitor to alter the molecular interactions in the Fmoc-Tyr-OH or Fmoc-Phe-OH self-assembled structures during the coassembly process, locking the coassembly in the nucleation step and preventing the formation of nanofibers. This structural transformation is useful for extending the application of amino acid self- or coassembled materials in different fields. For example, the amino acids forming nanofibers could be applied for tissue engineering, while they could be exploited as drug nanocarriers when they form nanoparticles.

Unveiling Liquid-Phase Exfoliation of Graphite and Boron Nitride Using Fluorescent Dyes Through Combined Experiments and Simulations.

Liquid-phase exfoliation (LPE) in aqueous solutions provides a simple, scalable, and green approach to produce 2D materials. By combining atomistic simulations with exfoliation experiments, the interaction between a surfactant and a 2D layer at the molecular scale can be better understood. In this work, two different dyes, corresponding to rhodamine B base (Rbb) and to a phenylboronic acid BODIPY (PBA-BODIPY) derivative, are employed as dispersants to exfoliate graphene and hexagonal boron nitride (hBN) through sonication-assisted LPE. The exfoliated 2D sheets, mostly as few-layers, exhibit good quality and high loading of dyes. Using molecular dynamics (MD) simulations, the binding free energies are calculated and the arrangement of both dyes on the layers are predicted. It has been found that the dyes show a higher affinity toward hBN than graphene, which is consistent with the higher yields of exfoliated hBN. Furthermore, it is demonstrated that the adsorption behavior of Rbb molecules on graphene and hBN is quite different compared to PBA-BODIPY.

Carbon nanotubes targeted to the tumor microenvironment inhibit metastasis in a preclinical model of melanoma.

Despite notable progress in cancer therapy, metastatic diseases continue to be the primary cause of cancer-related mortality. Multi-walled carbon nanotubes (MWCNTs) can enter tissues and cells and interfere with the dynamics of the cytoskeletal nanofilaments biomimetically. This endows them with intrinsic anti-tumoral effects comparable to those of microtubule-binding chemotherapies such as Taxol®. In this study, our focus was on exploring the potential of oxidized MWCNTs in selectively targeting the vascular endothelial growth factor receptor (VEGFR). Our objective was to evaluate their effectiveness in inhibiting metastatic growth by inducing anti-proliferative, anti-migratory, and cytotoxic effects on both cancer and tumor microenvironment cells. Our findings demonstrated a significant reduction of over 80 % in malignant melanoma lung metastases and a substantial enhancement in overall animal welfare following intravenous administration of the targeted biodegradable MWCNTs. Furthermore, the combination of these nanomaterials with the conventional chemotherapy agent Taxol® yielded a remarkable 90 % increase in the antimetastatic effect. These results highlight the promising potential of this combined therapeutic approach against metastatic disease and are of paramount importance as metastasis is responsible for nearly 60,000 deaths each year.

Preparation and optimization of agarose or polyacrylamide/amino acid-based double network hydrogels for photocontrolled drug release.

The high water content and biocompatibility of amino-acid-based supramolecular hydrogels have generated growing interest in drug delivery research. Nevertheless, the existing dominant approach of constructing such hydrogels, the exploitation of a single amino acid type, typically comes with several drawbacks such as weak mechanical properties and long gelation times, hindering their applications. Here, we design a near-infrared (NIR) light-responsive double network (DN) structure, containing amino acids and different synthetic or natural polymers, i.e., polyacrylamide, poly(N-isopropylacrylamide), agarose, or low-gelling agarose. The hydrogels displayed high mechanical strength and high drug-loading capacity. Adjusting the ratio of Fmoc-Tyr-OH/Fmoc-Tyr(Bzl)-OH or Fmoc-Phe-OH/Fmoc-Tyr(Bzl)-OH, we could drastically shorten the gelation time of the DN hydrogels at room and body temperatures. Moreover, introducing photothermal agents (graphene oxide, carbon nanotubes, molybdenum disulfide nanosheets, or indocyanine green), we equipped the hydrogels with NIR responsivity. We demonstrated the light-triggered release of the drug baclofen, which is used in severe spasticity treatment. Rheology and stability tests confirmed the positive impact of the polymers on the mechanical strength of the hydrogels, while maintaining good stability under physiological conditions. Overall, our study contributed a novel hydrogel formulation with high mechanical resistance, rapid gel formation, and efficient NIR-controlled drug release, offering new opportunities for biomedical applications.

Photocrosslinked Co-Assembled Amino Acid Nanoparticles for Controlled Chemo/Photothermal Combined Anticancer Therapy.

Nanostructures formed from the self-assembly of amino acids are promising materials in many fields, especially for biomedical applications. However, their low stability resulting from the weak noncovalent interactions between the amino acid building blocks limits their use. In this work, nanoparticles co-assembled by fluorenylmethoxycarbonyl (Fmoc)-protected tyrosine (Fmoc-Tyr-OH) and tryptophan (Fmoc-Trp-OH) are crosslinked by ultraviolet (UV) light irradiation. Two methods are investigated to induce the dimerization of tyrosine, irradiating at 254 nm or at 365 nm in the presence of riboflavin as a photo-initiator. For the crosslinking performed at 254 nm, both Fmoc-Tyr-OH and Fmoc-Trp-OH generate dimers. In contrast, only Fmoc-Tyr-OH participates in the riboflavin-mediated dimerization under irradiation at 365 nm. The participation of both amino acids in forming the dimers leads to more stable crosslinked nanoparticles, allowing also to perform further chemical modifications for cancer applications. The anticancer drug doxorubicin (Dox) is adsorbed onto the crosslinked nanoparticles, subsequently coated by a tannic acid-iron complex, endowing the nanoparticles with glutathione-responsiveness and photothermal properties, allowing to control the release of Dox. A remarkable anticancer efficiency is obtained in vitro and in vivo in tumor-bearing mice thanks to the combined chemo- and photothermal treatment.

Roadmap on multifunctional materials for drug delivery.

This Roadmap on drug delivery aims to cover some of the most recent advances in the field of materials for drug delivery systems (DDSs) and emphasizes the role that multifunctional materials play in advancing the performance of modern DDSs in the context of the most current challenges presented. The Roadmap is comprised of multiple sections, each of which introduces the status of the field, the current and future challenges faced, and a perspective of the required advances necessary for biomaterial science to tackle these challenges. It is our hope that this collective vision will contribute to the initiation of conversation and collaboration across all areas of multifunctional materials for DDSs. We stress that this article is not meant to be a fully comprehensive review but rather an up-to-date snapshot of different areas of research, with a minimal number of references that focus upon the very latest research developments.

Applications of graphene-based nanomaterials in drug design: The good, the bad and the ugly.

INTRODUCTION: Graphene-based materials (GBMs) have unique physicochemical properties that make them extremely attractive as platforms for the design of new drugs. Indeed, their bidimensional (2D) morphology, high surface area, mechanical and optical properties, associated to different possibilities for functionalization of their surface, provides opportunities for their use as nanomedicines for drug delivery and/or phototherapies. AREAS COVERED: This opinion paper provides an overview of the current status of GBMs in drug design, with a focus on their therapeutic applications, potential environmental and health risks, and some controversial results. The authors discuss the chemical modifications of GBMs for the treatment of various diseases. The potential toxicity associated with some GBMs is also presented, along with a safe-by-design approach to minimize the risks. Finally, the authors address some issues associated to the use of GBMs in the biomedical field, such as contradictory antibacterial effects, fluorescence quenching and imprecise chemical functionalization. EXPERT OPINION: GBMs are a promising and exciting area of research in drug delivery. It is however important that responsible and safe use of these materials is ensured to fully exploit their advantages and overcome their drawbacks.

Graphene Oxide-BODIPY Conjugates as Highly Fluorescent Materials.

Covalent functionalization of graphene oxide (GO) with boron dipyrromethenes (BODIPYs) was achieved through a facile synthesis, affording two different GO-BODIPY conjugates where the main difference lies in the nature of the spacer and the type of bonds between the two components. The use of a long but flexible spacer afforded strong electronic GO-BODIPY interactions in the ground state. This drastically altered the light absorption of the BODIPY structure and impeded its selective excitation. In contrast, the utilisation of a short, but rigid spacer based on boronic esters resulted in a perpendicular geometry of the phenyl boronic acid BODIPY (PBA-BODIPY) with respect to the GO plane, which enables only minor electronic GO-BODIPY interactions in the ground state. In this case, selective excitation of PBA-BODIPY was easily achieved, allowing to investigate the excited state interactions. A quantitative ultrafast energy transfer from PBA-BODIPY to GO was observed. Furthermore, due to the reversible dynamic nature of the covalent GO-PBA-BODIPY linkage, some PBA-BODIPY is free in solution and, hence, not quenched from GO. This resulted in a weak, but detectable fluorescence from the PBA-BODIPY that will allow to exploit GO-PBA-BODIPY for slow release and imaging purposes.

Resolution of MoS2 Nanosheets-Induced Pulmonary Inflammation Driven by Nanoscale Intracellular Transformation and Extracellular-Vesicle Shuttles.

Pulmonary exposure to some engineered nanomaterials can cause chronic lesions as a result of unresolved inflammation. Among 2D nanomaterials and graphene, MoS2 has received tremendous attention in optoelectronics and nanomedicine. Here an integrated approach is proposed to follow up the transformation of MoS2 nanosheets at the nanoscale and assesss their impact on lung inflammation status over 1 month after a single inhalation in mice. Analysis of immune cells, alveolar macrophages, extracellular vesicles, and cytokine profiling in bronchoalveolar lavage fluid (BALF) shows that MoS2 nanosheets induced initiation of lung inflammation. However, the inflammation is rapidly resolved despite the persistence of various biotransformed molybdenum-based nanostructures in the alveolar macrophages and the extracellular vesicles for up to 1 month. Using in situ liquid phase transmission electron microscopy experiments, the dynamics of MoS2 nanosheets transformation triggered by reactive oxygen species could be evidenced. Three main transformation mechanisms are observed directly at the nanoscale level: 1) scrolling of the dispersed sheets leading to the formation of nanoscrolls and folded patches, 2) etching releasing soluble MoO4 - , and 3) oxidation generating oxidized sheet fragments. Extracellular vesicles released in BALF are also identified as a potential shuttle of MoS2 nanostructures and their degradation products and more importantly as mediators of inflammation resolution.

Combined Photothermal and Photodynamic Therapy for Cancer Treatment Using a Multifunctional Graphene Oxide.

Graphene oxide (GO) is one of the most studied nanomaterials in many fields, including the biomedical field. Most of the nanomaterials developed for drug delivery and phototherapies are based on noncovalent approaches that lead to an unspecific release of physisorbed molecules in complex biological environments. Therefore, preparing covalently functionalized GO using straightforward and versatile methods is highly valuable. Phototherapies, including photothermal therapy (PTT) and photodynamic therapy (PDT), have shown great potential as effective therapeutic approaches against cancer. To overcome the limits of a single method, the combination of PTT and PDT can lead to a combined effect with a higher therapeutic efficiency. In this work, we prepare a folic acid (FA) and chlorin e6 (Ce6) double-functionalized GO for combined targeted PTT/PDT. This conjugate can penetrate rapidly into cancer cells and macrophages. A combined effect of PTT and PDT is observed, leading to a higher killing efficiency toward different types of cells involved in cancer and other diseases. Our work provides a simple protocol to prepare multifunctional platforms for the treatment of various diseases.

Aromatic Dipeptide Homologue-Based Hydrogels for Photocontrolled Drug Release.

Peptide-based hydrogels are considered of special importance due to their biocompatibility and biodegradability. They have a wide range of applications in the biomedical field, such as drug delivery, tissue engineering, wound healing, cell culture media, and biosensing. Nevertheless, peptide-based hydrogels composed of natural α-amino acids are limited for in vivo applications because of the possible degradation by proteolytic enzymes. To circumvent this issue, the incorporation of extra methylene groups within the peptide sequence and the protection of the terminal amino group can increase the enzymatic stability. In this context, we investigated the self-assembly capacity of aromatic dipeptides (Boc-α-diphenylalanine and Boc-α-dityrosine) and their ß- and γ-homologues and developed stable hydrogels. Surprisingly, only the Boc-diphenylalanine analogues were able to self-assemble and form hydrogels. A model drug, l-ascorbic acid, and oxidized carbon nanotubes (CNTs) or graphene oxide were then incorporated into the hydrogels. Under near-infrared light irradiation, the photothermal effect of the carbon nanomaterials induced the destabilization of the gel structure, which caused the release of a high amount of drug, thus providing opportunities for photocontrolled on-demand drug release.

Self-assembly of amphiphilic amino acid derivatives for biomedical applications.

Amino acids are one of the simplest biomolecules and they play an essential role in many biological processes. They have been extensively used as building blocks for the synthesis of functional nanomaterials, thanks to their self-assembly capacity. In particular, amphiphilic amino acid derivatives can be designed to enrich the diversity of amino acid-based building blocks, endowing them with specific properties and/or promoting self-assembly through hydrophobic interactions, hydrogen bonding, and/or π-stacking. In this review, we focus on the design of various amphiphilic amino acid derivatives able to self-assemble into different types of nanostructures that were exploited for biomedical applications, thanks to their excellent biocompatibility and biodegradability.

Graphene oxide activates B cells with upregulation of granzyme B expression: evidence at the single-cell level for its immune-modulatory properties and anticancer activity.

We recently found by single-cell mass cytometry that ex vivo human B cells internalize graphene oxide (GO). The functional impact of such uptake on B cells remains unexplored. Here, we disclosed the effects of GO and amino-functionalized GO (GONH2) interacting with human B cells in vitro and ex vivo at the protein and gene expression levels. Moreover, our study considered three different subpopulations of B cells and their functionality in terms of: (i) cytokine production, (ii) activation markers, (iii) killing activity towards cancer cells. Single-cell mass cytometry screening revealed the higher impact of GO on cell viability towards naïve, memory, and plasma B cell subsets. Different cytokines such as granzyme B (GrB) and activation markers, like CD69, CD80, CD138, and CD38, were differently regulated by GONH2 compared to GO, supporting possible diverse B cell activation paths. Moreover, co-culture experiments also suggest the functional ability of both GOs to activate B cells and therefore enhance the toxicity towards HeLa cancer cell line. Complete transcriptomic analysis on a B cell line highlighted the distinctive GO and GONH2 elicited responses, inducing pathways such as B cell receptor and CD40 signaling pathways, key players for GrB secretion. B cells were regularly left behind the scenes in graphene biological studies; our results may open new horizons in the development of GO-based immune-modulatory strategies having B cell as main actors.

The impact of graphene oxide sheet lateral dimensions on their pharmacokinetic and tissue distribution profiles in mice.

Although the use of graphene and 2-dimensional (2D) materials in biomedicine has been explored for over a decade now, there are still significant knowledge gaps regarding the fate of these materials upon interaction with living systems. Here, the pharmacokinetic profile of graphene oxide (GO) sheets of three different lateral dimensions was studied. The GO materials were functionalized with a PEGylated DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid), a radiometal chelating agent for radioisotope attachment for single photon emission computed tomography (SPECT/CT) imaging. Our results revealed that GO materials with three distinct size distributions, large (l-GO-DOTA), small (s-GO-DOTA) and ultra-small (us-GO-DOTA), were sequestered by the spleen and liver. Significant accumulation of the large material (l-GO-DOTA) in the lungs was also observed, unlike the other two materials. Interestingly, there was extensive urinary excretion of all three GO nanomaterials indicating that urinary excretion of these structures was not affected by lateral dimensions. Comparing with previous studies, we believe that the thickness of layered nanomaterials is the predominant factor that governs their excretion rather than lateral size. However, the rate of urinary excretion was affected by lateral size, with large GO excreting at slower rates. This study provides better understanding of 2D materials in vivo behaviour with varying structural features.

Nanobiosensor Reports on CDK1 Kinase Activity in Tumor Xenografts in Mice.

Probing the dynamics and quantifying the activities of intracellular protein kinases that coordinate cell growth and division and constitute biomarkers and pharmacological targets in hyperproliferative and pathological disorders remain a challenging task. Here engineering and characterization of a nanobiosensor of the mitotic kinase CDK1, through multifunctionalization of carbon nanotubes with a CDK1-specific fluorescent peptide reporter, are described. This original reporter of CDK1 activity combines the sensitivity of a fluorescent biosensor with the unique physico-chemical and biological properties of nanotubes for multifunctionalization and efficient intracellular penetration. The functional versatility of this nanobiosensor enables implementation to quantify CDK1 activity in a sensitive and dose-dependent fashion in complex biological environments in vitro, to monitor endogenous kinase in living cells and directly within tumor xenografts in mice by fluorescence imaging, thanks to a ratiometric quantification strategy accounting for response relative to concentration in space and in time.

Two-Dimensional Material-Based Biosensors for Virus Detection.

Viral infections are one of the major causes of mortality and economic losses worldwide. Consequently, efficient virus detection methods are crucial to determine the infection prevalence. However, most detection methods face challenges related to false-negative or false-positive results, long response times, high costs, and/or the need for specialized equipment and staff. Such issues can be overcome by access to low-cost and fast response point-of-care detection systems, and two-dimensional materials (2DMs) can play a critical role in this regard. Indeed, the unique and tunable physicochemical properties of 2DMs provide many advantages for developing biosensors for viral infections with high sensitivity and selectivity. Fast, accurate, and reliable detection, even at early infection stages by the virus, can be potentially enabled by highly accessible surface interactions between the 2DMs and the analytes. High selectivity can be obtained by functionalization of the 2DMs with antibodies, nucleic acids, proteins, peptides, or aptamers, allowing for specific binding to a particular virus, viral fingerprints, or proteins released by the host organism. Multiplexed detection and discrimination between different virus strains are also feasible. In this Review, we present a comprehensive overview of the major advances of 2DM-based biosensors for the detection of viruses. We describe the main factors governing the efficient interactions between viruses and 2DMs, making them ideal candidates for the detection of viral infections. We also critically detail their advantages and drawbacks, providing insights for the development of future biosensors for virus detection. Lastly, we provide suggestions to stimulate research in the fast expanding field of 2DMs that could help in designing advanced systems for preventing virus-related pandemics.

Correction: Controlled functionalization of carbon nanodots for targeted intracellular production of reactive oxygen species.

Correction for 'Controlled functionalization of carbon nanodots for targeted intracellular production of reactive oxygen species' by Ding-Kun Ji et al., Nanoscale Horiz., 2020, 5, 1240-1249, DOI: .