Gold Nanosystems Covered with Doxorubicin/DNA Complexes: A Therapeutic Target for Prostate and Liver Cancer.

Different gold nanosystems covered with DNA and doxorubicin (Doxo) were designed and synthesized for cancer therapy, starting from Au@16-Ph-16 cationic nanoparticles and DNA-Doxo complexes prepared under saturation conditions. For the preparation of stable, biocompatible, and small-sized compacted Au@16-Ph-16/DNA-Doxo nanotransporters, the conditions for the DNA-Doxo compaction process induced by gold nanoparticles were first explored using fluorescence spectroscopy, circular dichroism and atomic force microscopy techniques. The reverse process, which is fundamental for Doxo liberation at the site of action, was found to occur at higher CAu@16-Ph-16 concentrations using these techniques. Zeta potential, dynamic light scattering and UV-visible spectroscopy reveal that the prepared compacted nanosystems are stable, highly charged and of adequate size for the effective delivery of Doxo to the cell. This fact is verified by in vitro biocompatibility and internalization studies using two prostate cancer-derived cell lines (LNCaP and DU145) and one hepatocellular carcinoma-derived cell line (SNU-387), as well as a non-tumor prostate (PNT2) cell line and a non-hepatocarcinoma hepatoblastoma cell line (Hep-G2) model used as a control in liver cells. However, the most outstanding results of this work are derived from the use of the CI+NI combined treatments which present strong action in cancer-derived cell lines, while a protective effect is observed in non-tumor cell lines. Hence, novel therapeutic targets based on gold nanoparticles denote high selectivity compared to conventional treatment based on free Doxo at the same concentration. The results obtained show the viability of both the proposed methodology for internalization of compacted nanocomplexes inside the cell and the effectiveness of the possible treatment and minimization of side effects in prostate and liver cancer.

Exploring Factors for the Design of Nanoparticles as Drug Delivery Vectors.

To achieve optimal results when employing nanoparticles in biomedical fields, choosing the right type of nanoparticle and determining the correct procedure for drug loading are key factors. Each type of nanoparticle presents a determined set of characteristics that are, in some cases, unique. In general, their surface charge, geometry or hydrophilic character may be limiting factors, depending on what their intended application is. Once synthesized, additional factors, such as their interaction with biological systems and liberation mechanisms into the target cells, also need to be taken into account. Multiple advantages arise from the use of nanoparticles, such as the capability to solubilize hydrophobic compounds and an increased bioavailability. Those advantages justify the extensive and delicate study that should be undertaken in order to use them as drug delivery agents. One of the most important factors for the design of a drug delivery system with nanoparticles is achieving a high drug-to-nanoparticle ratio. In this Minireview, all of these key factors, both physicochemical and biological, are described, and special emphasis is placed on loading methods employed to introduce drugs into nanoparticles.

Covalent and Non-Covalent DNA-Gold-Nanoparticle Interactions: New Avenues of Research.

The interactions of DNA, whether long, hundred base pair chains or short-chained oligonucleotides, with ligands play a key role in the field of structural biology. Its biological activity not only depends on the thermodynamic properties of DNA-ligand complexes, but can and often is conditioned by the formation kinetics of those complexes. On the other hand, gold nanoparticles have long been known to present excellent biocompatibility with biomolecules and are themselves remarkable for their structural, electronic, magnetic, optical and catalytic properties, radically different from those of their counterpart bulk materials, and which make them an important asset in multiple applications. Therefore, thermodynamic and kinetic studies of the interactions of DNA with nanoparticles acting as small ligands are key for a better understanding of those interactions to allow for their control and modulation and for the opening of new venues of research in nanomedicine, analytic and biologic fields. The interactions of gold nanoparticles with both DNA polymers and their smaller subunits; special focus is placed on those interactions taking place with nonfunctionalized gold nanoparticles are reviewed in the present work.

Understanding and improving aggregated gold nanoparticle/dsDNA interactions by molecular spectroscopy and deconvolution methods.

It is well known that single-stranded DNA (ssDNA) is easily able to adsorb on citrate-capped, non-functionalized gold nanoparticles (AuNPs). However, the affinity of double-stranded DNA (dsDNA) for them is much more limited. The present work demonstrates that long dsDNA suffers from a bending conformational change when anionic nanoparticles are present in solution. A striking decrease in the persistence length of the double helix in the absence of salt is observed through dynamic light scattering (DLS), viscometric, and atomic force microscopy (AFM) methods. Long dsDNA is therefore shown to be able to interact with anionic gold nanoparticles. To date, only ssDNA detection has been described by making use of interparticle cross-linking aggregation mechanisms; however, the data shown in this work allow for the development of new methods for detecting dsDNA in solution by using aggregated AuNPs as a starting point. The aggregation state is induced by the controlled addition of an inert electrolyte. A deconvolution procedure of the experimental plasmon shows how individual bands corresponding to aggregated nanoclusters diminish as the DNA concentration increases in the presence of 0.075 M NaCl.

Lysozyme-AuNPs Interactions: Determination of Binding Free Energy.

Investigation and optimization of lysozyme (Lys) adsorption onto gold nanoparticles, AuNPs, were carried out. The purpose of this study is to determine the magnitude of the AuNPs-lysozyme interaction in aqueous media by simple spectrophotometric means, and to obtain the free energy of binding of the system for the first time. In order to explore the possibilities of gold nanoparticles for sensing lysozyme in aqueous media, the stability of the samples and the influence of the gold and nanoparticle concentrations in the detection limit were studied. ζ potential measurements and the shift of the surface plasmon band showed a state of saturation with an average number of 55 Lys per gold nanoparticle. Lysozyme-AuNPs interactions induce aggregation of citrate-stabilized AuNPs at low concentrations by neutering the negative charges of citrate anions; from those aggregation data, the magnitude of the interactions has been measured by using Benesi-Hildebrand plots. However, at higher protein concentrations aggregation has been found to decrease. Although the nanocluster morphology remains unchanged in the presence of Lys, slight conformational changes of the protein occur. The influence of the size of the nanoclusters was also investigated for 5, 10, and 20 nm AuNPs, and 10 nm AuNPs was found the most appropriate.

Colorimetric, Naked-Eye Detection of Lysozyme in Human Urine with Gold Nanoparticles.

The stabilizing effect of lysozymes to salt addition over a gold colloid are exploited in order to detect lysozymes in human urine samples. The present research is aimed at the development of a fast, naked-eye detection test for urinary lysozymuria, in which direct comparison with a colorimetric reference, allows for the immediate determination of positive/negative cases. CIEL*a*b* parameters were obtained from sample absorbance measurements, and their color difference with respect to a fixed reference point was measured by calculating the ΔE76 parameter, which is a measure of how well the colors can be distinguished by an untrained observer. Results show that a simple and quick test can reliably, in less than 15 min, give a positive colorimetric response in the naked eye for concentrations of a urinary lysozyme over 57.2 µg/mL. This concentration is well within the limits of that observed for leukemia-associated lysozymurias, among other disorders.

Chitosan as a capping agent: Insights on the stabilization of gold nanoparticles.

A theoretical method based on Dissipative Particle Dynamics (DPD) was applied for the study of the interaction between coarse-grained models of chitosan and gold nanoparticles. These models were used to understand the role of chitosan in the stabilization of gold nanoparticles (5 and 7 nm). In addition, this stabilization was also supported by synthesized and characterized chitosan-gold nanoparticles comparing their stabilities with typical citrate-gold nanoparticles. Finally, the role of chitosan as a capping agent through the steric and electrosteric mechanisms in aqueous media with different pH values was also evaluated.

Decorating a single giant DNA with gold nanoparticles.

We decorated a single giant DNA (1.66 × 105 base pairs) with gold nanoparticles through the simple procedure of mild warming, without denaturation of the DNA molecule. Single-molecule observation with fluorescence microscopy revealed that individual decorated DNA molecules stay in the bulk solution by avoiding aggregation and precipitation, and exhibit translational and conformational fluctuation, i.e., Brownian motion. An analysis of the intra-chain fluctuation of single DNA molecules revealed that the apparent spring constant and damping coefficient of a DNA chain increased by ca. 13- and 5-fold, respectively, upon decoration with gold nanoparticles. Observation by transmission electron microscopy revealed that gold nanoparticles were stably attached to the DNA skeleton. UV-visible measurements revealed the absence of any detectable change in surface plasmon resonance, suggesting that the gold nanoparticles assemble without the formation of a densely packed aggregate. CD measurements showed that the secondary structure of decorated DNA is still essentially the B-form.

Spectroscopic study of the ground and excited state prototropic equilibria of 4-azaindole.

The ground and singlet excited state prototropism of 4-azaindole, 4AI, in acid and basic aqueous solutions, inside and outside the pH range, has been systematically studied by using absorption and fluorescence spectroscopic techniques. These studies have thrown light on some interesting aspects on the nature and the photophysics of the 4-AI prototropic species. Thus, the changes of the 4AI absorption spectra reveal the existence of four ground state species; the pyridinic protonated cation, C (pK(a)(C)=7.5±0.1), the neutral molecule, N (pK(a)(N)=15.5±0.5), the pyrrolic deprotonated anion, A, and a previously unnoticed dication, DC (pK(a)(DC)=-4.6±0.4). Besides the emissions of these species, a new fluorescence profile appears in alkaline solutions at around 500nm. This extra band has been ascribed to the neutral phototautomer, NT. What is more relevant to this study is the fact that the position and the intensity of the emission band assigned to the monoprotonated cation are very different from those observed for the normal cation of the 7-azaindole, 7-AI. This together with the fact that for the formation of the DC species a cationic precursor with a quinoid structure must be invoked, have prompted us to assign this cationic emission to the isomeric CI cations. Finally, the excited-state pK(a)s of the prototropic species of 4AI have been theoretically estimated by using the Förster-Weller cycle.