Self-assembly of a 5-fluorouracil and camptothecin dual drug dipeptide conjugate.

Nano-formulated, combinatory therapeutics that control the spatiotemporal aspects of drug release have potential to overcome many of the challenges faced in cancer therapy. Herein, we describe a peptide nanotube functionalized with two anticancer drugs, 5-fluoruracil (5-FU) and camptothecin (CPT). The nanotube was formed via peptide self-assembly, which positioned 5-FU on the surface at the aqueous interface; whereas, CPT was sequestered within the hydrophobic walls. Thus, two different release profiles were observed: rapid release of 5-FU, followed by slower, sustained production of CPT. This profile emerged from the rapid hydrolytic cleavage of 5-FU at the aqueous/nanotube interface, which produced a smaller nanotube comprised of the peptide fragment.

Enhanced Stability of Peptide Nanofibers Coated with a Conformal Layer of Polydopamine.

The susceptibility of self-assembled materials to changes of environmental conditions and mechanical forces often limits their utility for many applications. In this work, the surface of nanofibers formed by ß-sheet peptide self-assemblies were coated by polydopamine (PDA) deposition. This conformal coating process rendered the nanofiber dimensions and internal π-stacking chirality impervious to changes in pH, temperature, and physical processing by spin-coating onto a silicon wafer. Whereas sonication-induced shearing of the dopamine/naphthalenediimide-dilysine (DA/NDI-KK) composite irreversibly shortened the nanofibers into 100-200 nm segments, the uncoated nanofibers unraveled into single strands upon similar treatment. Additionally, the PDA-coated nanofibers could be wrapped by an additional layer comprised of a positively charged polyelectrolyte polymer.

pH-Controlled Chiral Packing and Self-Assembly of a Coumarin Tetrapeptide.

A coumarin-tetrapeptide conjugate, EFEK(DAC)-NH2 (1), is reported to undergo a pH-dependent interconversion between nanotubes and nanoribbons. An examination of zeta potential measurements, circular dichroism (CD) spectra, and microscopy imaging (transmission electron microscopy and atomic force microscopy) identified three different self-assembly regimes based on pH: (1) pH 2-5, positively charged, left-handed helical nanotubes; (2) pH 6-8, negatively charged, right-handed helical nanoribbons; and (3) pH ≥ 9.0, a monomeric/disassembled peptide. The nanotubes exhibited uniform diameters of 41 ± 5 nm and wall thicknesses of 4.8 ± 0.8 nm, whereas the nanoribbons existed as either flat or twisted sheets ranging in width from 11 to 60 nm with heights of 8 ± 1 nm. The UV-vis and CD spectra of the most common antiparallel, ß-sheet conformation of 1-dimer were simulated at the B3LYP/def2svpd level of theory in implicit water. These studies indicated that the transition from nanotubes to nanoribbons was coupled to an M → P helical inversion of the coumarin packing orientation, respectively, within the nanostructures. The assembly process was driven by ß-sheet aggregation and π-π interactions, leading to the formation of nanoribbons, which progressively wound into helical ribbons and laterally grew into smooth nanotubes as the pH decreased.

Strategy for the Co-Assembly of Co-Axial Nanotube-Polymer Hybrids.

Nanostructured materials having multiple, discrete domains of sorted components are particularly important to create efficient optoelectronics. The construction of multicomponent nanostructures from self-assembled components is exceptionally challenging due to the propensity of noncovalent materials to undergo structural reorganization in the presence of excipient polymers. This work demonstrates that polymer-nanotube composites comprised of a self-assembled nanotube wrapped with two conjugated polymers could be assembled using a layer-by-layer approach. The polymer-nanotube nanostructures arrange polymer layers coaxially on the nanotube surface. Femtosecond transient absorption (TA) studies indicated that the polymer-nanotube composites undergo photoinduced charge separation upon excitation of the NDI chromophore within the nanotube.

The self-assembly of a camptothecin-lysine nanotube.

A simple, low molecular weight camptothecin-lysine conjugate is reported to self-assemble into nanotubes with diameters of 70-100nm and a drug loading level of 60.5%. The nanotubes exhibited promising in vitro cytotoxicity against cancer cell lines A549, NCI-H460 and NCI-H23. The release of active camptothecin was highly dependent on conjugate concentration, temperature and pH of the solution.

The self-assembly of anticancer camptothecin-dipeptide nanotubes: a minimalistic and high drug loading approach to increased efficacy.

20-(S)-Camptothecin (CPT)-conjugated dipeptides are reported that preassemble into nanotubes with diameters ranging from 80-120 nm. These nanoassemblies maintain a high (∼47 %) drug loading and exhibit greater drug stability (i.e., resistance to lactone hydrolysis), and consequently greater efficacy against several human cancer cells (HT-29, A549, H460, and H23) in vitro compared with the clinically used prodrug irinotecan. A key and defining feature of this system is the use of the CPT-conjugated dipeptide as both the drug and precursor to the nanostructured carrier, which simplifies the overall fabrication process.

Electrostatic assembly of a multicomponent peptide/amphiphile nanotube.

The ability to integrate the elements of a multicomponent nanostructure with nanoscale precision by co-assembly provides a versatile strategy to create novel materials with tunable properties. The search for function in these materials will require new strategies to be developed that control the assembly process, especially for structurally dissimilar components, which often have a propensity to self-sort into non-integrated nanostructures. In this work, two components, a peptide (1) and an amphiphile (2), were integratively co-assembled into a multicomponent nanotube. The interaction between the two components at the supramolecular level was driven by the electrostatic complementarity of the components, which was controlled by the pH-dependent charge of 1. Characterization of the co-assembled nanotube, 1-2NT, was achieved using a combination of TEM, AFM, CLSM and SIM techniques, which showed that both components were colocalized within the nanotube. These studies, in conjunction with CD, IR and fluorescence studies, suggested that 1 and 2 were arranged in partially reorganized, self-sorted domains, which were integrated as laminated nanoribbons that coiled together into the final co-assembled nanotube.

Molecular bowls for inclusion complexation of toxic anticancer drug methotrexate.

We describe the preparation and study of novel cavitands, molecular bowls 16+ and 26+, as good binders of the anticancer drug methotrexate (MTX). Molecular bowls are comprised of a curved tribenzotriquinacene (TBTQ) core conjugated to three macrocyclic pyridinium units at the top. The cavitands are easily accessible via two synthetic steps from hexabromo-tribenzotriquinacene in 25% yield. As amphiphilic molecules, bowls 16+ and 26+ self-associate in water by the nucleation-to-aggregation pathway (NMR). The bowls are preorganized, having a semi-rigid framework comprising a fixed bottom with a wobbling pyridinium rim (VT NMR and MD). Further studies, both experimental (NMR) and computational (DFT and MCMM), suggested that a folded MTX occupies the cavity of bowls wherein it forms π-π, C-H-π, and ion pairing intermolecular contacts but also undergoes desolvation to give stable binary complexes (µM) in water. Moreover, a computational protocol is introduced to identify docking pose(s) of MTX inside molecular bowls from NMR shielding data. Both molecular bowls have shown in vitro biocompatibility with liver and kidney cell lines (MTS assay). As bowl 26+ is the strongest binder of MTX reported to date, we envision it as an excellent candidate for further studies on the way toward developing an antidote capable of removing MTX from overdosed cancer patients.

Proton-coupled self-assembly of a porphyrin-naphthalenediimide dyad.

The construction of an n-p heterojunction through the self-assembly of a dyad based on tetraphenylporphyrin (TPP) and 1,4,5,8-naphthalenedimide (NDI) (1) is described. Proton transfer from the lysine head group of 1 to the porphyrin ring occurs concomitantly with self-assembly into 1D nanorods in CHCl3. TEM and AFM studies showed that the nanorods are formed by the lateral and vertical fusion of multilameller vesicles into networks and hollow ribbons, respectively. These intermediate structures transitioned to nanorods over the course of 4-6 days. Time-resolved spectroscopy revealed that photoinduced charge separation occurs with rate constants that depend on the nature of the aggregation.

Intramolecular chiral communication in peptide-dendron hybrids.

The conformational properties of a series of peptide-dendron hybrids progressively incorporating 1-4 dendritic side chains were investigated by circular dichroism. Although the presence of multiple adjacent dendrons along the peptide backbone precluded the formation of α-helical or ß-sheet secondary structure, intramolecular packing of the dendrons mediated efficient peptide → dendron chirality transfer in both organic and aqueous media.

Dissipative self-assembly of a proline catalyst for temporal regulation of the aldol reaction.

The spatiotemporal regulation of chemical reactivity in biological systems permits a network of metabolic reactions to take place within the same cellular environment. The exquisite control of reactivity is often mediated by out-of-equilibrium structures that remain functional only as long as fuel is present to maintain the higher energy, active state. An important goal in supramolecular chemistry aims to develop functional, energy dissipating systems that approach the sophistication of biological machinery. The challenge is to create strategies that couple the energy consumption needed to promote a molecule to a higher energy, assembled state to a functional property such as catalytic activity. In this work, we demonstrated that the assembly of a spiropyran (SP) dipeptide (1) transiently promoted the proline-catalyzed aldol reaction in water when visible light was present as fuel. The transient catalytic activity emerged from 1 under light illumination due to the photoisomerization of the monomeric, O-protonated (1-MCH+) merocyanine form to the spiropyran (1-SP) state, which rapidly assembled into nanosheets capable of catalyzing the aldol reaction in water. When the light source was removed, thermal isomerization to the more stable MCH+ form caused the nanosheets to dissociate into a catalytically inactive, monomeric state. Under these conditions, the aldol reaction could be repeatedly activated and deactivated by switching the light source on and off.

Co-assembly of a multicomponent network of nanofiber-wrapped nanotubes.

Strategies to create organized multicomponent nanostructures composed of discrete, self-sorted domains are important for developing materials that mimic the complexity and multifunctionality found in biological systems. These structures can be challenging to achieve due to the required balance of molecular self-recognition and supramolecular attraction needed between the components. Herein, we report a strategy to construct a two-component nanostructure via a hierarchical assembly process whereby two monomeric building blocks undergo self-sorting assembly at the molecular level followed by a supramolecular association to form a nanofiber-wrapped nanotube. The two molecules self-sorted into respective nanofiber and nanotube assemblies, yet assembly of the nanofibers in the presence of the nanotube template allowed for directed integration into a hierarchical multilayer structure via electrostatic interactions. The fiber-wrapped nanotube co-assembly was characterized using transmission electron microscopy (TEM), atomic force microscopy (AFM) and Förster resonance energy transfer (FRET) between the components. Strategies to co-assemble multicomponent nanostructures composed of discrete, spatially sorted domains with controllable higher level interactions will be critical for the development of novel, functionally competent nanomaterials.

Self-assembly of a donor-acceptor nanotube. A strategy to create bicontinuous arrays.

The self-assembly of bolaamphiphile 1 into nanotubes containing a nanostructured electron donor/acceptor heterojunction is reported. In 10% MeOH/H(2)O, the tetraphenylporphyrin (TPP) and 1,4,5,8-naphthalenetetracarboxylic acid diimide chromophores engage in strong J-type π-π interactions within monolayer rings that further stack into the nanotube assemblies. In 10% MeOH/H(2)O at pH 1 or 11 or in pure MeOH, assembly is driven exclusively by the TPP ring, leading to the formation of nonspecific, unstructured aggregates. Steady-state, time-resolved fluorescence and femtosecond transient absorption spectroscopy revealed a strong dependence of the fluorescence decay and electron-transfer/charge-recombination time constants on the nature of the assemblies. These studies highlight the importance of local nanostructure in determining the photophysical properties of optoelectronic materials.

Light-driven dissipative self-assembly of a peptide hydrogel.

Light energy provides an attractive fuel source for energy dissipating systems because of the lack of waste production, wavelength tunability and the potential for spatial and temporal resolution. In this work, we describe a peptide-spiropyran conjugate that assembled into a transient nanofiber hydrogel in the presence of visible light, and dissociated when the light source was removed.

Amino Acid Nanofibers Improve Glycemia and Confer Cognitive Therapeutic Efficacy to Bound Insulin.

Diabetes poses a high risk for debilitating complications in neural tissues, regulating glucose uptake through insulin-dependent and predominantly insulin-independent pathways. Supramolecular nanostructures provide a flexible strategy for combinatorial regulation of glycemia. Here, we compare the effects of free insulin to insulin bound to positively charged nanofibers comprised of self-assembling amino acid compounds (AACs) with an antioxidant-modified side chain moiety (AAC2) in both in vitro and in vivo models of type 1 diabetes. Free AAC2, free human insulin (hINS) and AAC2-bound-human insulin (AAC2-hINS) were tested in streptozotocin (STZ)-induced mouse model of type 1 diabetes. AAC2-hINS acted as a complex and exhibited different properties compared to free AAC2 or hINS. Mice treated with the AAC2-hINS complex were devoid of hypoglycemic episodes, had improved levels of insulin in circulation and in the brain, and increased expression of neurotransmitter taurine transporter, Slc6a6. Consequently, treatment with AAC2-hINS markedly advanced both physical and cognitive performance in mice with STZ-induced and genetic type 1 diabetes compared to treatments with free AAC2 or hINS. This study demonstrates that the flexible nanofiber AAC2 can serve as a therapeutic platform for the combinatorial treatment of diabetes and its complications.

The impact of metal coordination on the assembly of bis(indolyl)methane-naphthalene-diimide amphiphiles.

The self-assembly and coordination of amphiphiles comprised of naphthalenediimide (NDI) and bis(indolyl)methane (BIM) chromophores were investigated as a function of pH and metal. As observed by TEM, SEM and AFM imaging, the self-assembly of NDI-BIM 1 produced irregular nanostructures at neutral pH in CH3CN-H2O (1 : 1); whereas, well-defined nanotubes were observed at pH 2. Conversely, Fmoc-protected, NDI-BIM 2 formed nanotubes at neutral pH and nonspecific aggregates at pH 2. Upon coordination of Cu2+ ions to the bis(indoyl)methane moiety, a reorganization from nanotubes to vesicular structures was observed.

Self-assembly of a robust, reduction-sensitive camptothecin nanotube.

The self-assembly and covalent crosslinking of a camptothecin (CPT) tetrapeptide nanotube is reported. Intermolecular disulfide bond formation of a self-assembled CPT-peptide reversibly stabilized the nanotubes toward dissociation at low concentrations, resulting in inhibited release of CPT. In the presence of dithiothreitol (DTT), the release of CPT was significantly accelerated. The crosslinked nanotubes also exhibited in vitro cytotoxicity against human non-small cell lung cancer cell lines A549 and H460.

Light-controlled self-assembly of a dithienylethene bolaamphiphile in water.

The self-assembly of bolaamphiphiles comprised of a central photochromic dithienylethene (DTE) chromophore was investigated in aqueous media. Irradiation at 254 nm induced a conversion from the open to closed states of the DTE chromophores. Whereas, in water, irradiation produced a photostationary state of 20 : 80 (open/closed), in methanol the ratio improved to 10 : 90 (open/closed). The open → closed transition was accompanied by the formation of 1D nanofibers during incubation in darkness.

Amino acid-based compound activates atypical PKC and leptin receptor pathways to improve glycemia and anxiety like behavior in diabetic mice.

Differences in glucose uptake in peripheral and neural tissues account for the reduced efficacy of insulin in nervous tissues. Herein, we report the design of short peptides, referred as amino acid compounds (AAC) with and without a modified side chain moiety. At nanomolar concentrations, a candidate therapeutic molecule, AAC2, containing a 7-(diethylamino) coumarin-3-carboxamide side-chain improved glucose control in human peripheral adipocytes and the endothelial brain barrier cells by activation of insulin-insensitive glucose transporter 1 (GLUT1). AAC2 interacted specifically with the leptin receptor (LepR) and activated atypical protein kinase C zeta (PKCς) to increase glucose uptake. The effects induced by AAC2 were absent in leptin receptor-deficient predipocytes and in Leprdb mice. In contrast, AAC2 established glycemic control altering food intake in leptin-deficient Lepob mice. Therefore, AAC2 activated the LepR and acted in a cytokine-like manner distinct from leptin. In a monogenic Ins2Akita mouse model for the phenotypes associated with type 1 diabetes, AAC2 rescued systemic glucose uptake in these mice without an increase in insulin levels and adiposity, as seen in insulin-treated Ins2Akita mice. In contrast to insulin, AAC2 treatment increased brain mass and reduced anxiety-related behavior in Ins2Akita mice. Our data suggests that the unique mechanism of action for AAC2, activating LepR/PKCς/GLUT1 axis, offers an effective strategy to broaden glycemic control for the prevention of diabetic complications of the nervous system and, possibly, other insulin insensitive or resistant tissues.

Self-assembly of 1-D n-type nanostructures based on naphthalene diimide-appended dipeptides.

n-Type 1D nanostructures are formed from the beta-sheet assembly of dipeptides bearing a 1,4,5,8-naphthalenetetracarboxylic acid diimide (NDI) side chain into either helical nanofibers or twisted nanoribbons. Amyloid-like 1-D helical nanofibers and twisted nanoribbons assemble in an aqueous solution depending on the placement of the NDI group. beta-Sheet-type hydrogen bonding and pi-pi association play important roles in directing the assembly process. A delicate balance between electrostatic repulsion and hydrophobic interactions is critical for self-assembly. Fluorescence lifetime and anisotropy experiments indicate that the nature of the intermolecular organization and packing within the nanostructures critically impacts intermolecular energy migration pi-electron delocalization.