Anna Jagusiak

Carbon nanotubes for delivery of small molecule drugs.

In the realm of drug delivery, carbon nanotubes (CNTs) have gained tremendous attention as promising nanocarriers, owing to their distinct characteristics, such as high surface area, enhanced cellular uptake and the possibility to be easily conjugated with many therapeutics, including both small molecules and biologics, displaying superior efficacy, enhanced specificity and diminished side effects. While most CNT-based drug delivery system (DDS) had been engineered to combat cancers, there are also emerging reports that employ CNTs as either the main carrier or adjunct material for the delivery of various non-anticancer drugs. In this review, the delivery of small molecule drugs is expounded, with special attention paid to the current progress of in vitro and in vivo research involving CNT-based DDSs, before finally concluding with some consideration on inevitable complications that hamper successful disease intervention with CNTs.

Research Article: The Use of Rigid, Fibrillar Congo Red Nanostructures for Scaffolding Protein Assemblies and Inducing the Formation of Amyloid‐like Arrangement of Molecules

The ordered amyloid‐like organization of protein aggregates was obtained using for their formation the rigid fibrillar nanostructures of Congo red as the scaffolding. The higher rigidity of used dye nanoparticles resulted from the stronger stacking of molecules at low pH (near the pK of the dye amino group) because of the decreased charge repulsion. The polylysine, human globin, and immunoglobulin L chain were arranged in this way to form deposits of amyloid properties. The scaffolding was introduced simply by mixing the dye and proteins at a low pH or the dye was used in the preorganized form by maintaining it in the electric field before and during protein addition. The polarization and electron microscopy studies confirmed the unidirectional organization of the complex. The precipitate of the complex was used for studies directly or after the partial or complete removal of the dye. The results suggest that the process of formation of amyloid‐like deposits may bypass the nucleation step. It is possible if the protein aggregation occurs in unidirectionally organized (because of scaffolding) assembly of molecules, arranged prior to self‐association. The recognition of the structure of amphoteric Congo red nanoparticles used for the scaffolding was based on the molecular dynamics simulation.

Molecular dynamics study of Congo red interaction with carbon nanotubes

This work deals with molecular dynamics simulations of Congo red (CR) interaction with carbon nanotubes (CNT). We studied several combinations of systems parameters in order to assess how the nanotube diameter and Congo red density affect the structure and stability of CNT–CR conjugates at various pH conditions. We found that, at the considered conditions, the CR binds strongly to the CNT surfaces and the CNT–CR conjugates are thermodynamically stable according to the determined values of the free energies. Adsorption on wider nanotubes is stronger than on the narrow ones and larger densities of CR on the CNT surfaces lead to weakening of binding energy per single CR molecule. Changes of pH, that is varying concentration of protonated and deprotonated forms of CR, lead to significant changes in binding energies as well as to qualitative changes of the structure of the adsorbed CR. It was found that at pH > 5.5 the CR molecules readily occupy inner cavities of the nanotubes. Upon lowering pH the occupation of the inner space of CNTs is strongly reduced and the preferred configuration is formation of a densely packed CR layer on the sidewalls of the CNT. This effect can potentially be utilized in pH controlled corking/uncorking of carbon nanotubes in water solutions.

The use of supramolecular structures as protein ligands

Congo red dye as well as other eagerly self-assembling organic molecules which form rod-like or ribbon-like supramolecular structures in water solutions, appears to represent a new class of protein ligands with possible wide-ranging medical applications. Such molecules associate with proteins as integral clusters and preferentially penetrate into areas of low molecular stability. Abnormal, partly unfolded proteins are the main binding target for such ligands, while well packed molecules are generally inaccessible. Of particular interest is the observation that local susceptibility for binding supramolecular ligands may be promoted in some proteins as a consequence of function-derived structural changes, and that such complexation may alter the activity profile of target proteins. Examples are presented in this paper.

Influence of the electric field on supramolecular structure and properties of amyloid-specific reagent Congo red

Among specific amyloid ligands, Congo red and its analogues are often considered potential therapeutic compounds. However, the results of the studies so far have not been univocal because the properties of this dye, derived mostly from its supramolecular nature, are still poorly understood. The supramolecular structure of Congo red, formed by π–π stacking of dye molecules, is susceptible to the influence of the electric field, which may significantly facilitate electron delocalization. Consequently, the electric field may generate altered physico-chemical properties of the dye. Enhanced electron delocalization, induced by the electric field, alters the total charge of Congo red, making the dye more acidic (negatively charged). This is a consequence of withdrawing electrons from polar substituents of aromatic rings—sulfonic and amino groups—thus increasing their tendency to dissociate protons. The electric field-induced charge alteration observed in electrophoresis depends on dye concentration. This concentration-dependent charge alteration effect disappears when the supramolecular structure disintegrates in DMSO. Dipoles formed from supramolecular fibrillar species in the electric field become ordered in the solution, introducing the modified arrangement to liquid crystalline phase. Experimental results and theoretical studies provide evidence confirming predictions that the supramolecular character of Congo red is the main reason for its specific properties and reactivity.

Intramolecular Immunological Signal Hypothesis Revived - Structural Background of Signalling Revealed by Using Congo Red as a Specific Tool

Micellar structures formed by self-assembling Congo red molecules bind to proteins penetrating into functionrelated unstable packing areas. Here, we have used Congo red - a supramolecular protein ligand to investigate how the intramolecular structural changes that take place in antibodies following antigen binding lead to complement activation. According to our findings, Congo red binding significantly enhances the formation of antigen-antibody complexes. As a result, even low-affinity transiently binding antibodies can participate in immune complexes in the presence of Congo red, although immune complexes formed by these antibodies fail to trigger the complement cascade. This indicates that binding of antibodies to the antigen may not, by itself, fulfill the necessary conditions to generate the signal which triggers effector activity. These findings, together with the results of molecular dynamics simulation studies, enable us to conclude that, apart from the necessary assembling of antibodies, intramolecular structural changes generated by strains which associate high- affinity bivalent antibody fitting to antigen determinants are also required to cross the complement activation threshold.

The use of Titan yellow dye as a metal ion binding marker for studies on the formation of specific complexes by supramolecular Congo red

Abstract Congo red (CR) and other self-assembling compounds creating supramolecular structures of rod- or ribbon-like architecture form specific complexes with cellulose and also with many proteins, including antibodies bound to the antigen and amyloids in particular. The mechanism of complexation and structure of these complexes are still poorly recognized despite the importance of the problem for medicine. This work proposes the progress in electron microscopy studies of amyloid-dye complexes by labeling supramolecular ligand CR with silver ions as a marker. Silver ions are introduced to CR carried by the strongly binding silver dye Titan yellow, which in addition form comicellar structures with CR. Silver carried by self-assembled dye molecules forms in the resulting metal nanoparticles, making the specific amyloid ligand CR perceptible in EM studies.

Formation of amyloid-like aggregates through the attachment of protein molecules to a Congo red scaffolding framework ordered under the influence of an electric field

This study describes a technique which makes it possible to introduce the amyloid-like order to protein aggregates by using the scaffolding framework built from supramolecular, fibrillar Congo red structures arranged in an electric field. The electric field was used not only to obtain a uniform orientation of the charged dye fibrils, but also to make the fibrils long, compact and rigid due to the delocalization of pi electrons, which favors ring stacking and, as a consequence, results in an increased tendency to self-assemble. The protein molecules (immunoglobulin L chain lambda, ferritin) attached to this easily adsorbing dye framework assume its ordered structure. The complex precipitating as plate-like fragments shows birefringence in polarized light. The parallel organization of fibrils can be observed with an electron microscope. The dye framework may be removed via reduction with sodium dithionite, leaving the aggregated protein molecules in the ordered state, as confirmed by X-ray diffraction studies.

Shortening and dispersion of single-walled carbon nanotubes upon interaction with mixed supramolecular compounds

Abstract Congo red (CR) dye molecules self-associate in water solutions creating ribbon-like supramolecular structures that can bind various aromatic compounds by intercalation, forming mixed supramolecular systems. Mixed supramolecular systems, such as CR-doxorubicin and CR-Evans blue, interact with the surface of carbon nanotubes, leading to their stiffening and ultimately to their breaking and shortening. This work presents a simple method of obtaining short and straight carbon nanotubes with significantly better dispersion in aqueous solutions and consequently improved usability in biological systems.

Erratum to: Supramolecular Structures as Carrier Systems Enabling the Use of Metal Ions in Antibacterial Therapy

The antimicrobial activity of metal ions, especially silver ions, has been known since ancient times. Consequently, finding an accessible, cheap and efficient carrier of metal ions remains an important challenge in molecular biology. The supramolecular system presented in this chapter consists of a mixture of Congo red and Titan yellow molecules forming a supramolecular ligand which is a potent complexing agent of silver ions. Delivery of ions in complex with supramolecular dye is advantageous due to the reduced toxicity. In addition, the use of Congo red provides selective action and – thanks to increased solubility – facilitates efficient dispersion of the carrier dye and excretion from the organism.