Carbon nano-onions for biomedical applications

The biomedical applications of carbon nanomaterials are under intensive investigation for the development of next-generation therapeutics. Although much focus has been placed on carbon nanotubes (CNTs) and graphene, other carbon nanomaterials including carbon nanohorns (CNHs), nanodiamonds (NDs) and fullerenes have emerged as suitable candidates for biomedical applications. Among these multi-shell fullerenes, also known as carbon nano-onions (CNOs), are the less studied carbon nanomaterials in biomedicine. The unique properties of carbon nano-onions, such as high surface area-to-volume ratio, thermal conductivity, electrical conductance, mechanical stiffness and ease of chemical functionalization render them fascinating materials for diverse applications including drug-delivery, diagnostics, biological imaging and tissue engineering. Carbon nanomaterials are emerging as smart nanostructures for biomedicine due to the possibility to incorporate multiple functionalities and moieties internally or externally. They can be modified at a precise physicochemical level to optimize targeting in the complex in vivo environment and also engineered for fluorescence detection, magnetic resonance imaging and ablation of tumor cells. Herein, robust and versatile synthetic strategies for the modification of carbon nano-onions (CNOs) are reported. The development of novel CNO conjugates represent a promising platform for the realization of novel technology scaffold for molecular imaging, photodynamic therapy and molecular transporter of fully synthetic carbohydrate-based vaccines for immunotherapy due to the large specific surface area and unique optical and electrochemical properties of CNOs. Through the methodologies described, these smart nano-materials can envisage the realization of multi stimuli-responsive and dynamic architectures capable of changing their physicochemical behavior upon encountering specific microenvironmental signals becoming relevant for diagnosis, imaging and therapies of specific disease applications

Membrane protein channels equipped with a cleavable linker for inducing catalysis inside nanocompartments

Precisely timed initiation of reactions and stability of the catalysts are fundamental in catalysis. We introduce here an efficient closing-opening method for nanocompartments that contain sensitive catalysts and so achieve a controlled and extended catalytic activity. We developed a chemistry-oriented approach for modifying a pore-forming membrane protein which allows for a stimuli-responsive pore opening within the membrane of polymeric nanocompartments. We synthesized a diol-containing linker that selectively binds to the pores, blocking them completely. In the presence of an external stimulus (periodate), the linker is cleaved allowing the diffusion of substrate through the pores to the nanocompartment interior where it sets off the in situ enzymatic reaction. Besides the precise initiation of catalytic activity by opening of the pores, oxidation by periodate guarantees the cleavage of the linker under mild conditions. Accordingly, this kind of responsive nanocompartment lends itself to harboring a large variety of sensitive catalysts such as proteins and enzymes

Clustering of catalytic nanocompartments for enhancing an extracellular non-native cascade reaction

Compartmentalization is fundamental in nature, where the spatial segregation of biochemical reactions within and between cells ensures optimal conditions for the regulation of cascade reactions. While the distance between compartments or their interaction are essential parameters supporting the efficiency of bio-reactions, so far they have not been exploited to regulate cascade reactions between bioinspired catalytic nanocompartments. Here, we generate individual catalytic nanocompartments (CNCs) by encapsulating within polymersomes or attaching to their surface enzymes involved in a cascade reaction and then, tether the polymersomes together into clusters. By conjugating complementary DNA strands to the polymersomes' surface, DNA hybridization drove the clusterization process of enzyme-loaded polymersomes and controlled the distance between the respective catalytic nanocompartments. Owing to the close proximity of CNCs within clusters and the overall stability of the cluster architecture, the cascade reaction between spatially segregated enzymes was significantly more efficient than when the catalytic nanocompartments were not linked together by DNA duplexes. Additionally, residual DNA single strands that were not engaged in clustering, allowed for an interaction of the clusters with the cell surface as evidenced by A549 cells, where clusters decorating the surface endowed the cells with a non-native enzymatic cascade. The self-organization into clusters of catalytic nanocompartments confining different enzymes of a cascade reaction allows for a distance control of the reaction spaces which opens new avenues for highly efficient applications in domains such as catalysis or nanomedicine

Multicompartment Polymer Vesicles with Artificial Organelles for Signal-Triggered Cascade Reactions Including Cytoskeleton Formation

Abstract Organelles, i.e., internal subcompartments of cells, are fundamental to spatially separate cellular processes, while controlled intercompartment communication is essential for signal transduction. Furthermore, dynamic remodeling of the cytoskeleton provides the mechanical basis for cell shape transformations and mobility. In a quest to develop cell-like smart synthetic materials, exhibiting functional flexibility, a self-assembled vesicular multicompartment system, comprised of a polymeric membrane (giant unilamellar vesicle, GUV) enveloping polymeric artificial organelles (vesicles, nanoparticles), is herein presented. Such multicompartment assemblies respond to an external stimulus that is transduced through a precise sequence. Stimuli-triggered communication between two types of internal artificial organelles induces and localizes an enzymatic reaction and allows ion-channel mediated release from storage vacuoles. Moreover, cytoskeleton formation in the GUVs` lumen can be triggered by addition of ionophores and ions. An additional level of control is achieved by signal-triggered ionophore translocation from organelles to the outer membrane, triggering cytoskeleton formation. This system is further used to study the diffusion of various cytoskeletal drugs across the synthetic outer membrane, demonstrating potential applicability, e.g., anticancer drug screening. Such multicompartment assemblies represent a robust system harboring many different functionalities and are a considerable leap in the application of cell logics to reactive and smart synthetic materials

Inverting glucuronidation of hymecromone in situ by catalytic nanocompartments

Glucuronidation is a metabolic pathway that inactivates many drugs including hymecromone. Adverse effects of glucuronide metabolites include a reduction of half-life circulation times and rapid elimination from the body. Herein, we developed synthetic catalytic nanocompartments able to cleave the glucuronide moiety from the metabolized form of hymecromone in order to convert it to the active drug. By shielding enzymes from their surroundings, catalytic nanocompartments favor prolonged activity and lower immunogenicity as key aspects to improve the therapeutic solution. The catalytic nanocompartments (CNCs) consist of self-assembled poly(dimethylsiloxane)- block -poly(2-methyl-2-oxazoline) diblock copolymer polymersomes encapsulating β-glucuronidase. Insertion of melittin in the synthetic membrane of these polymersomes provided pores for the diffusion of the hydrophilic hymecromone-glucuronide conjugate to the compartment inside where the encapsulated β-glucuronidase catalyzed its conversion to hymecromone. Our system successfully produced hymecromone from its glucuronide conjugate in both, phosphate buffered solution and cell culture medium. CNCs were non-cytotoxic when incubated with HepG2 cells. After being taken up by cells, CNCs produced the drug in situ over 24 hours. Such catalytic platforms that locally revert a drug metabolite into its active form, open new avenues in the design of therapeutics that aim at prolonging the residence time of a drug

Highly surface functionalized carbon nano-onions for bright light bioimaging

Carbon-based nanomaterials functionalized with fluorescent and water-soluble groups have emerged as platforms for biological imaging because of their low toxicity and ability to be internalized by cells. The development of imaging probes based on carbon nanomaterials for biomedical studies requires the understanding of their biological response as well as the efficient and safety exposition of the nanomaterial to the cell compartment where it is designed to operate. Here, we present a fluorescent probe based on surface functionalized carbon nano-onions (CNOs) for biological imaging. The modification of CNOs by chemical oxidation of the defects on the outer shell of these carbon nanoparticles results in an extensive surface functionalization with carboxyl groups. We have obtained fluorescently labelled CNOs by a reaction involving the amide bond formation between fluoresceinamine and the carboxylic acids groups on the surface of the CNOs. The functionalized CNOs display high emission properties and dispersability in water due to the presence of high surface coverage of carboxylic acid groups that translate in an efficient fluorescent probe for in vitro imaging of HeLa cells, without significant cytotoxicity. The resulting nanomaterial represents a promising platform for biological imaging applications due to the high dispersability in water, its efficient internalization by cancer cells and localization in specific cell compartments

Synthetic Cells Revisited: Artificial Cells Construction Using Polymeric Building Blocks

Abstract The exponential growth of research on artificial cells and organelles underscores their potential as tools to advance the understanding of fundamental biological processes. The bottom–up construction from a variety of building blocks at the micro‐ and nanoscale, in combination with biomolecules is key to developing artificial cells. In this review, artificial cells are focused upon based on compartments where polymers are the main constituent of the assembly. Polymers are of particular interest due to their incredible chemical variety and the advantage of tuning the properties and functionality of their assemblies. First, the architectures of micro‐ and nanoscale polymer assemblies are introduced and then their usage as building blocks is elaborated upon. Different membrane‐bound and membrane‐less compartments and supramolecular structures and how they combine into advanced synthetic cells are presented. Then, the functional aspects are explored, addressing how artificial organelles in giant compartments mimic cellular processes. Finally, how artificial cells communicate with their surrounding and each other such as to adapt to an ever‐changing environment and achieve collective behavior as a steppingstone toward artificial tissues, is taken a look at. Engineering artificial cells with highly controllable and programmable features open new avenues for the development of sophisticated multifunctional systems

Photocatalytic Initiation of Radical Thiol-ene Reactions Using Carbon-Bi2O3 Nanocomposites

A mild, inexpensive, and general photocatalytic initiation protocol for anti-Markovnikov hydrothiolation of olefins using carbon nanomaterial/metal oxide (carbon NM-MO) composites is reported. Graphene oxide (GO), nanodiamonds (ND), and carbon nano-onions (CNO) displaying bismuth or tungsten oxide nanoparticles adhered to the surface and function as highly efficient photocatalysts for thiol?ene ligation under both UV and visible-light-mediated conditions. The straightforward catalyst preparation, excellent overall yields, ease of purification, and broad substrate scope render this a highly versatile method for bioconjugation

Advancing the Design of Artificial Nano-organelles for Targeted Cellular Detoxification of Reactive Oxygen Species

Artificial organelles (AnOs) are in the spotlight as systems to supplement biochemical pathways in cells. While polymersome-based artificial organelles containing enzymes to reduce reactive oxygen species (ROS) are known, applications requiring control of their enzymatic activity and cell-targeting to promote intracellular ROS detoxification are underexplored. Here, we introduce advanced AnOs where the chemical composition of the membrane supports the insertion of pore-forming melittin, enabling molecular exchange between the AnO cavity and the environment, while the encapsulated lactoperoxidase (LPO) maintains its catalytic function. We show that H₂O₂ outside AnOs penetrates through the melittin pores and is rapidly degraded by the encapsulated enzyme. As surface attachment of cell-penetrating peptides facilitates AnOs uptake by cells, electron spin resonance revealed a remarkable enhancement in intracellular ROS detoxification by these cell-targeted AnOs compared to nontargeted AnOs, thereby opening new avenues for a significant reduction of oxidative stress in cells.ISSN:1530-6984ISSN:1530-699

Multicompartment Polymer Vesicles with Artificial Organelles for Signal‐Triggered Cascade Reactions Including Cytoskeleton Formation

Abstract Organelles, i.e., internal subcompartments of cells, are fundamental to spatially separate cellular processes, while controlled intercompartment communication is essential for signal transduction. Furthermore, dynamic remodeling of the cytoskeleton provides the mechanical basis for cell shape transformations and mobility. In a quest to develop cell-like smart synthetic materials, exhibiting functional flexibility, a self-assembled vesicular multicompartment system, comprised of a polymeric membrane (giant unilamellar vesicle, GUV) enveloping polymeric artificial organelles (vesicles, nanoparticles), is herein presented. Such multicompartment assemblies respond to an external stimulus that is transduced through a precise sequence. Stimuli-triggered communication between two types of internal artificial organelles induces and localizes an enzymatic reaction and allows ion-channel mediated release from storage vacuoles. Moreover, cytoskeleton formation in the GUVs` lumen can be triggered by addition of ionophores and ions. An additional level of control is achieved by signal-triggered ionophore translocation from organelles to the outer membrane, triggering cytoskeleton formation. This system is further used to study the diffusion of various cytoskeletal drugs across the synthetic outer membrane, demonstrating potential applicability, e.g., anticancer drug screening. Such multicompartment assemblies represent a robust system harboring many different functionalities and are a considerable leap in the application of cell logics to reactive and smart synthetic materials