Chemists have created molecular machines and switches with specific mechanical responses that were typically demonstrated in solution, where mechanically relevant motion is dissipated in the Brownian storm. The next challenge consists of designing specific mechanisms through which the action of individual molecules is transmitted to a supramolecular architecture, with a sense of directionality. Cellular microtubules are capable of meeting such a challenge. While their capacity to generate pushing forces by ratcheting growth is well known, conversely these versatile machines can also pull microscopic objects apart through a burst of their rigid tubular structure. One essential feature of this disassembling mechanism is the accumulation of strain in the tubules, which develops when tubulin dimers change shape, triggered by a hydrolysis event. We envision a strategy toward supramolecular machines generating directional pulling forces by harnessing the mechanically purposeful motion of molecular switches in supramolecular tubules. Here, we report on wholly synthetic, water-soluble, and chiral tubules that incorporate photoswitchable building blocks in their supramolecular architecture. Under illumination, these tubules display a nonlinear operation mode, by which light is transformed into units of strain by the shape changes of individual switches, until a threshold is reached and the tubules unleash the strain energy. The operation of this wholly synthetic and stripped-down system compares to the conformational wave by which cellular microtubules disassemble. Additionally, atomistic simulations provide molecular insight into how strain accumulates to induce destabilization. Our findings pave the way toward supramolecular machines that would photogenerate pulling forces, at the nanoscale and beyond.artificial molecular switches | supramolecular polymers | supramolecular machines | light

Fully homogeneous facial amphiphiles consisting in a cyclodextrin (CD) platform onto which a polycationic cluster and a multi-tail hydrophobic moiety have been installed (polycationic amphiphilic CDs; paCDs) self-organized in the presence of plasmid DNA to form nanometric complexes (CDplexes) which exhibit broad-range transfection capabilities. We hypothesized that biorecognizable moieties located at the hydrophilic rim in the CD scaffold would be exposed at the surface of the corresponding nanoparticles after DNA-promoted aggregation, endowing the system with molecular recognition abilities towards cell receptors. This concept has been demonstrated by developing an efficient synthetic strategy for the preparation of multivalent polycationic glyco-amphiphilic CDs (pGaCDs). Self-assembled nanoparticles obtained from mannosylated pGaCDs and pDNA (average hydrodynamic diameter 80 nm) have been shown to be specifically recognized by mannose-specific lectins, including concanavalin A (Con A) and the human macrophage mannose receptor (MMR). Further macrophage adhesion studies indicated that unspecific binding, probably due to electrostatic interactions with negatively charged cell membrane components, can also operate. The relative specific versus non-specific internalization is dependent on the pGaCD:pDNA proportion, being optimal at a protonable nitrogen/phosphate (N/P) ratio of 5. The resulting GlycoCDplexes were shown to specifically mediate transfection in Raw 264.7 (murine macrophage) cells expressing the mannose-fucose receptor in vitro. FACS experiments confirmed that transfection using these nanoparticles is mannose-dependent, supporting the potential of the approach towards vectorized gene delivery.

The controlled one-dimensional supramolecular polymerization of synthetic building blocks in confined spaces constitutes a key challenge to simplify the understanding of the fundamental physical principles behind the behavior of more complex encapsulated polymer networks.

Monodisperse facial amphiphiles consisting of a β-cyclodextrin (βCD) platform exposing a multivalent display of cationic groups at the primary rim and bearing hydrophobic chains at the secondary oxygens have been prepared by implementing two very robust "click" methodologies, namely cuprous cation-catalyzed azide-alkyne cycloaddition (CuAAC) and thiourea-forming reaction. Most interestingly, the use of solid-supported Cu(I) catalysts was found to be very well suited for multiple CuAAC while facilitating purification of the C(7)-symmetric macromolecular triazole adducts. The strategy is compatible with molecular diversity-oriented approaches, which has been exploited to generate a small library of click polycationic amphiphilic CDs (paCDs) for assessing the influence of structural modifications in the ability to complex, compact, and protect pDNA and the efficiency of the resulting paCD:pDNA nanocomplexes (CDplexes) to deliver DNA into cells and promote transfection. The results indicate that fine-tuning the hydrophilic/hydrophobic balance is critical to achieve optimal self-assembling properties and stability of the resulting CDplexes in saline- and serum-containing media. Triazole-type paCDs were, in general, less efficient in promoting gene transfection than thiourea-type derivatives. Nevertheless, the current body of results support that the "dual click" approach implying sequential CuAAC and thiourea-forming reactions represents a versatile strategy to optimize the gene delivery capabilities of cyclodextrin-based facial amphiphiles.

Aim: In this study, a set of polycationic amphiphilic cyclodextrins featuring self-assembling capabilities in the presence of nucleic acids have been evaluated as therapeutic gene vectors for in vivo purposes. Materials & Methods: A tetradecacationic structure incorporating 14 primary amino groups and 7 thioureido groups in the primary face of the cyclooligosaccharide core and 14 hexanoyl chains in the secondary face was judged to be optimal for therapeutic gene delivery. Results & Conclusion: This compound efficiently mediated serum-resistant transfection in HeLa and HepG2 cells, comparing favorably with branched poly(ethyleneimine), with a low associated toxicity. Further transfection experiments using an encoding therapeutic gene plasmid (pCMVIL-12) were effected to report expression levels of IL-12. Finally, in vivo gene delivery experiments by systemic injection in mice indicated relatively high transfection levels in the liver, overcoming trapping of the nanoparticles in lung cells, with low toxicity.

T2 complexed and protected pDNA (luciferase-encoding plasmid DNA, pCMVLuc) and efficiently mediated transfection in vitro and in vivo with no associated toxicity. The combination of folic acid with CDplexes afforded ternary nanocomplexes (Fol-CDplexes) that enhanced significantly the transfection activity of pCMVLuc in human cervix adenocarcinoma HeLa cells, especially when formulated with 1 μg FA/μg DNA. The observed transfection enhancement was associated to specific folate receptor (FR)-mediated internalization of Fol-CDplexes, as corroborated by employing a receptor-deficient cell line (HepG2) and an excess of free folic acid. The in vivo studies, including luciferase reporter gene expression and biodistribution, indicated that 24h after intravenous administration of the T2-pDNA nanocomplexes, transfection takes part mainly in the liver and partially in the lung. Interestingly, the corresponding Fol-CDplexes lead to an increase in the transfection activity in the lung and the liver compared to non-targeted CDplexes. Folate-CDplexes developed in this study have improved transfection efficiency and although various methods have been used for the preparation of ligand-DNA-complexes, covalent binding is usually needed and insoluble aggregates are formed unless the concentration of the components is minimized. However, the complexes developed by first time in this work were prepared by simple mixing. The synthetic nature of this formulation provides the potential of flexibility in terms of composition and the capability of inexpensive and large-scale production of the complexes. These nanovectors may be an adequate alternative to viral vectors for gene therapy in the future.

Gene delivery systems based on the beta-cyclodextrin scaffold have been synthesized by combining the copper(I)-catalyzed azide-alkyne coupling ("click chemistry") and an efficient acylation method of the secondary hydroxyls; molecular flexibility, charge density and hydrophobic-hydrophilic balance are critical parameters that can be fine-tuned by the click approach.

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