Synthetic vesicles have been assembled and coassembled from phospholipids, their modified versions, and other single amphiphiles into liposomes, and from block copolymers into polymersomes. Their time-consuming synthesis and preparation as stable, monodisperse, and biocompatible liposomes and polymersomes called for the elaboration of new synthetic methodologies. Amphiphilic Janus dendrimers (JDs) and glycodendrimers (JGDs) represent the most recent self-assembling amphiphiles capable of forming monodisperse, stable, and multifunctional unilamellar and multilamellar onion-like vesicles denoted dendrimersomes (DSs) and glycodendrimersomes (GDSs), dendrimercubosomes (DCs), glycodendrimercubosomes (GDCs), and other complex architectures. Amphiphilic JDs consist of hydrophobic dendrons connected to hydrophilic dendrons and can be thought of as monodisperse oligomers of a single amphiphile. They can be functionalized with a variety of molecules such as dyes, and, in the case of JGDs, with carbohydrates. Their iterative modular synthesis provides efficient access to sequence control at the molecular level, resulting in topologies with specific epitope sequence and density. DSs, GDSs, and other architectures from JDs and JGDs serve as powerful tools for mimicking biological membranes and for biomedical applications such as targeted drug and gene delivery and theranostics. This Review covers all aspects of the synthesis of JDs and JGDs and their biological activity and applications after assembly in aqueous media.

A library of eight amphiphilic Janus glycodendrimers (GDs) with D-mannose (Man) headgroups, a known routing signal for lectinmediated transport processes, was constructed via an iterative modular methodology. Sequence-defined variations of the Janus GD modulate the surface density and sequence of Man after selfassembly into multilamellar glycodendrimersomes (GDSs). The spatial mode of Man presentation is decisive for formation of either unilamellar or onion-like GDS vesicles. Man presentation and Janus GD concentration determine GDS size and number of bilayers. Beyond vesicle architecture, Man topological display affects kinetics and plateau level of GDS aggregation by a tetravalent model lectin: the leguminous agglutinin Con A, which is structurally related to endogenous cargo transporters. The agglutination process was rapid, efficient, and readily reversible for onion-like GDSs, demonstrating their value as versatile tools to explore the nature of physiologically relevant glycan/lectin pairing.self-assembly | synthetic multilamellar vesicles | glycolipid mimics S upramolecular chemistry has enormous potential to help resolve fundamental questions in the realm of cell biology. One of the key challenges is the design of programmable models for vesicles and cells and their surfaces as a means of establishing a chemical platform that mimics natural features in size and shape, and also allows customized implementation of bioactive epitopes, in structural and topological terms. Natural complexity can conveniently be reduced to simple systems, whose degree of diversity can then be rationally reconstituted in a stepwise process. Focusing on surface properties, the recently gained access to uniform populations of stable glycodendrimersomes (GDSs) by a simple injection method using a solution of amphiphilic Janus glycodendrimers (GDs) as building blocks for self-assembly in a water-soluble aprotic solvent (1-3), has afforded a promising opportunity to realize this concept. Interestingly, the resulting GDSs, which have tunable surface features, can cover the size range of naturally occurring vesicles such as endo-and exosomes.

A library of eight amphiphilic Janus glycodendrimers (Janus-GDs) presenting D-lactose (Lac) and a combination of Lac with up to eight methoxytriethoxy (3EO) units in a sequence-defined arrangement was synthesized via an iterative modular methodology. The length of the linker between Lac and the hydrophobic part of the Janus-GDs was also varied. Self-assembly by injection from THF solution into phosphate-buffered saline led to unilamellar, monodisperse glycodendrimersomes (GDSs) with dimensions predicted by Janus-GD concentration. These GDSs provided a toolbox to measure bioactivity profiles in agglutination assays with sugar-binding proteins (lectins). Three naturally occurring forms of the human adhesion/growth-regulatory lectin galectin-8, Gal-8S and Gal-8L, which differ by the length of linker connecting their two active domains, and a single amino acid mutant (F19Y), were used as probes to study activity and sensor capacity. Unpredictably, the sequence of Lac on the Janus-GDs was demonstrated to determine bioactivity, with the highest level revealed for a Janus-GD with six 3EO groups and one Lac. A further increase in Lac density was invariably accompanied by a substantial decrease in agglutination, whereas a decrease in Lac density resulted in similar or lower bioactivity and sensor capacity. Both changes in topology of Lac presentation of the GDSs and seemingly subtle alterations in protein structure resulted in different levels of bioactivity, demonstrating the presence of regulation on both GDS surface and lectin. These results illustrate the applicability of Janus-GDs to dissect structure-activity relationships between programmable cell surface models and human lectins in a highly sensitive and physiologically relevant manner.

A library of amphiphilic Janus dendrimers including two that are fluorescent and one glycodendrimer presenting lactose were used to construct giant dendrimersomes and glycodendrimersomes. Coassembly with the components of bacterial membrane vesicles by a dehydration-rehydration process generated giant cell-like hybrid vesicles, whereas the injection of their ethanol solution into PBS produced monodisperse nanometer size assemblies. These hybrid vesicles contain transmembrane proteins including a small membrane protein, MgrB, tagged with a red fluorescent protein, lipopolysaccharides, and glycoproteins from the bacterium Escherichia coli. Incorporation of two colored fluorescent probes in each of the components allowed fluorescence microscopy to visualize and demonstrate coassembly and the incorporation of functional membrane channels. Importantly, the hybrid vesicles bind a human galectin, consistent with the display of sugar moieties from lipopolysaccharides or possibly glycosylated membrane proteins. The present coassembly method is likely to create cell-like hybrids from any biological membrane including human cells and thus may enable practical application in nanomedicine.E. coli | transmembrane protein | lipopolysaccharides | galectin N aturally occurring (1), chemically modified (2, 3), and synthetic (4, 5) lipids, amphiphilic block copolymers (6, 7), polypeptides (8), Janus dendrimers (JDs) (9), and Janus glycodendrimers (JGDs) (10, 11) self-assemble into vesicles denoted as liposomes, polymersomes, dendrimersomes (DSs), and glycodendrimersomes (GDSs), respectively. These vesicles provide models for primitive (12) and contemporary (13, 14) cell membranes and drug-delivery devices (15)(16)(17). Recently, hybrid vesicles coassembled from naturally occurring phospholipids and amphiphilic block copolymers (18-20) have been described; these vesicles eliminated some of the deficiencies of liposomes, such as limited stability under oxidative conditions and general instability over time, and the deficiencies of polymersomes, which possess wide membrane thickness [8-50 nm (20)], exhibit toxicity, and can be tedious to synthesize. These hybrid vesicles combined the desirable feature of liposomes-specifically, their biologically suitable membrane thickness of 4 nm-with that of polymersomes, which are known for their stability. In addition, transmembrane proteins (21-23) could be incorporated into the phospholipid fragments of planar membranes derived from these assemblies. However, the variability in the extent of miscibility between the hydrophobic fragments of the phospholipid and the block copolymer (20) generates a complex morphology of the hybrid membrane that requires further characterization to enable practical applications both as drug-delivery devices and cell membrane models. Here, we report the coassembly of the components of DSs and GDSs with those of the bacterial membrane vesicles (BMVs) to generate functional hybrid vesicles. DSs, GDSs, and liposomes have hydrophobic fragments with similar chemical st...

SignificanceCells are decorated with charged and uncharged carbohydrate ligands known as glycans, which are responsible for several key functions, including their interactions with proteins known as lectins. Here, a platform consisting of synthetic nanoscale vesicles, known as glycodendrimersomes, which can be programmed with cell surface-like structural and topological complexity, is employed to dissect design aspects of glycan presentation, with specificity for lectin-mediated bridging. Aggregation assays reveal the extent of cross-linking of these biomimetic nanoscale vesicles—presenting both anionic and neutral ligands in a bioactive manner—with disease-related human and other galectins, thus offering the possibility of unraveling the nature of these fundamental interactions.

Self-assembling dendrimers have facilitated the discovery of periodic and quasiperiodic arrays of supramolecular architectures and the diverse functions derived from them. Examples are liquid quasicrystals and their approximants plus helical columns and spheres, including some that disregard chirality. The same periodic and quasiperiodic arrays were subsequently found in block copolymers, surfactants, lipids, glycolipids, and other complex molecules. Here we report the discovery of lamellar and hexagonal periodic arrays on the surface of vesicles generated from sequence-defined bicomponent monodisperse oligomers containing lipid and glycolipid mimics. These vesicles, known as glycodendrimersomes, act as cell-membrane mimics with hierarchical morphologies resembling bicomponent rafts. These nanosegregated morphologies diminish sugar–sugar interactions enabling stronger binding to sugar-binding proteins than densely packed arrangements of sugars. Importantly, this provides a mechanism to encode the reactivity of sugars via their interaction with sugar-binding proteins. The observed sugar phase-separated hierarchical arrays with lamellar and hexagonal morphologies that encode biological recognition are among the most complex architectures yet discovered in soft matter. The enhanced reactivity of the sugar displays likely has applications in material science and nanomedicine, with potential to evolve into related technologies.

A three-component system of Janus dendrimers (JDs) including hydrogenated, fluorinated, and hybrid hydrogenated-fluorinated JDs are reported to coassemble by film hydration at specific ratios into an unprecedented class of supramolecular Janus particles (JPs) denoted Janus dendrimersomes (JDSs). They consist of a dumbbellshaped structure composed of an onion-like hydrogenated vesicle and an onion-like fluorinated vesicle tethered together. The synthesis of dye-tagged analogs of each JD component enabled characterization of JDS architectures with confocal fluorescence microscopy. Additionally, a simple injection method was used to prepare submicron JDSs, which were imaged with cryogenic transmission electron microscopy (cryo-TEM). As reported previously, different ratios of the same three-component system yielded a variety of structures including homogenous onion-like vesicles, core-shell structures, and completely self-sorted hydrogenated and fluorinated vesicles. Taken together with the JDSs reported herein, a self-sorting pathway is revealed as a function of the relative concentration of the hybrid JD, which may serve to stabilize the interface between hydrogenated and fluorinated bilayers. The fission-like pathway suggests the possibility of fusion and fission processes in biological systems that do not require the assistance of proteins but instead may result from alterations in the ratios of membrane composition.Janus particles | onion-like vesicles | self-sorting | fusion mechanism J anus particles (JPs) are nanoscale or microscale objects that exhibit two-faced asymmetry, enabling the presentation of vastly different chemical or physical properties localized at distinct parts of the same structure. They have been prepared as unimolecular systems in the form of dendrimers, dendrimer-like polymers, heterografted polymers, and polyhedral oligomeric silsesquioxanes; as supramolecular assemblies from lipids, block copolymers, and terpolymers and other polymers; as liquid crystals; and as inorganic nanoparticles. JPs have gained increasing attention due to their application in a variety of fields including as emulsion stabilizers, facilitators for the development of novel liquid/liquid and liquid/air interfaces among other interfacial applications, optical nanoprobes, electronic inks, self-propelling beads, targeted stealth drug delivery systems, MRI contrast agents, and theranostics agents, among other biomedical applications (1).Asymmetric molecules such as phospholipids, block copolymers, and Janus dendrimers (JDs) can be thought of as the molecularlevel equivalent of JPs. Phospholipids are the major building block of the bilayers of biological membranes. Synthetic vesicles, which mimic biological membranes, have been prepared from phospholipids and are denoted liposomes. As the result of their instability due to oxidation, phospholipids must be coassembled with cholesterol and polyethylene glycol-conjugated phospholipids to form stable liposomes (2, 3). Furthermore, time-consuming fractionation and extrusio...

Nonlamellar lipid arrangements, including cubosomes, appear in unhealthy cells, e.g., when they are subject to stress, starvation, or viral infection. The bioactivity of cubosomes—nanoscale particles exhibiting bicontinuous cubic structures—versus more common vesicles is an unexplored area due to lack of suitable model systems. Here, glycodendrimercubosomes (GDCs)—sugar-presenting cubosomes assembled from Janus glycodendrimers by simple injection into buffer—are proposed as mimics of biological cubic membranes. The bicontinuous cubic GDC architecture has been demonstrated by electron tomography. The stability of these GDCs in buffer enabled studies on lectin-dependent agglutination, revealing significant differences compared with the vesicular glycodendrimersome (GDS) counterpart. In particular, GDCs showed an increased activity toward concanavalin A, as well as an increased sensitivity and selectivity toward two variants of banana lectins, a wild-type and a genetically modified variant, which is not exhibited by GDSs. These results suggest that cells may adapt under unhealthy conditions by undergoing a transformation from lamellar to cubic membranes as a method of defense.