Anticancer Cationic Ruthenium Nanovectors: From Rational Molecular Design to Cellular Uptake and Bioactivity

Abstract

An efficient drug delivery strategy is presented for novel anticancer amphiphilic ruthenium anionic complexes, based on the formation of stable nanoparticles with the cationic lipid 1,2-dioleyl-3-trimethylammoniumpropane chloride (DOTAP). This strategy is aimed at ensuring high ruthenium content within the formulation, long half-life in physiological media, and enhanced cell uptake. An in-depth microstructural characterization of the aggregates obtained mixing the ruthenium complex and the phospholipid carrier at 50/50 molar ratio is realized by combining a variety of techniques, including dynamic light scattering (DLS), small angle neutron scattering (SANS), neutron reflectivity (NR), electron paramagnetic resonance (EPR), and zeta potential measurements. The in vitro bioactivity profile of the Ru-loaded nanoparticles is investigated on human and non-human cancer cell lines, showing IC(50) values in the low μM range against MCF-7 and WiDr cells, that is, proving to be 10-20-fold more active than AziRu, a previously synthesized NAMI-A analog, used for control. Fluorescence microscopy studies demonstrate that the amphiphilic Ru-complex/DOTAP formulations, added with rhodamine-B, are efficiently and rapidly incorporated in human MCF-7 breast adenocarcinoma cells. The intracellular fate of the amphiphilic Ru-complexes was investigated in the same in vitro model by means of an ad hoc designed fluorescently tagged analog, which exhibited a marked tendency to accumulate within or in proximity of the nuclei.

Antiproliferative effects of ruthenium-based nucleolipidic nanoaggregates in human models of breast cancer in vitro: insights into their mode of action

“…According to the IC 50 values (see Table 1), the ToThyRu/DOTAP and DoHuRu/DOTAP formulations are the most effective in reducing the proliferation of all the used cell lines. This result, consistent with our previous reports, is likely associated to the positive charge of DOTAP formulations which can promote the interaction with the plasma membranes allowing a faster and quantitative drug cellular uptake19. Moreover, in relation to their effectiveness in vitro , there are no significant differences between the ToThyRu and DoHuRu nucleolipidic Ru(III) complexes used in our liposomial formulations, consistently with the rationale that the ruthenium center is the bioactive species, and the nucleolipids are simply carrier molecules24.…”

“…, the DoHuRu/DOTAP ones) with respect to the zwitterionic POPC ones. This scenario is fully consistent with our previous studies1927. As a result of different cellular uptake kinetics, ruthenium-containing cationic liposomes (DoHuRu/DOTAP) induce faster biological effects by way of cellular apoptosis activation than those dependent by the neutral liposomes (DoHuRu/POPC).…”

“…Current research efforts are focused on a deeper understanding of the cellular response and/or resistance to anticancer treatments, including the role of cell death pathways activation, such as apoptosis and autophagy, by chemotherapeutics32. Recently, we have developed new biocompatible ruthenium-based nanosystems, proved to be particularly effective against specific cell lines derived from human solid tumours192224252627. Starting from these encouraging data, in the present work we have first confirmed their efficacy focusing on a panel of human tumour cells arising from breast cancer.…”

“…In addition to interfering with the internalization process into targeted cancer cells, degradation events can also limit the therapeutic effectiveness, thereby becoming a central issue that has claimed a reconsideration of the efficacy of low molecular weight ruthenium complexes currently in preclinical or clinical studies19. Additionally, the lack of convincing evidences on their biological mode of action, as well as on targets identification, has significantly limited their further development2021.…”

“…Of special interest have been the results obtained on estrogen and progesterone receptor positive MCF-7 adenocarcinoma cells, an in vitro model reflecting to a large extent the features of luminal breast cancer cells in vivo , worldwide the most common invasive cancer in women. Most remarkably, liposomes loaded with the Ru(III)-containing nucleolipids have shown a promising selectivity towards cancer cells in bioscreens in vitro 1922252627.…”