Exploring cellular uptake, accumulation and mechanism of action of a cationic Ru-based nanosystem in human preclinical models of breast cancer

Abstract

According to WHO, breast cancer incidence is increasing so that the search for novel chemotherapeutic options is nowadays an essential requirement to fight neoplasm subtypes. By exploring new effective metal-based chemotherapeutic strategies, many ruthenium complexes have been recently proposed as antitumour drugs, showing ability to impact on diverse cellular targets. In the framework of different molecular pathways leading to cell death in human models of breast cancer, here we demonstrate autophagy involvement behind the antiproliferative action of a ruthenium(III)-complex incorporated into a cationic nanosystem (HoThyRu/DOTAP), proved to be hitherto one of the most effective within the suite of nucleolipidic formulations we have developed for the in vivo transport of anticancer ruthenium(III)-based drugs. Indeed, evidences are implicating autophagy in both cancer development and therapy, and anticancer interventions endowed with the ability to trigger this biological response are currently considered attractive oncotherapeutic approaches. Moreover, crosstalk between apoptosis and autophagy, regulated by finely tuned metallo-chemotherapeutics, may provide novel opportunities for future improvement of cancer treatment. Following this line, our in vitro and in vivo preclinical investigations suggest that an original strategy based on suitable formulations of ruthenium(III)-complexes, inducing sustained cell death, could open new opportunities for breast cancer treatment, including the highly aggressive triple-negative subtype.

Anticancer Ruthenium(III) Complexes and Ru(III)-Containing Nanoformulations: An Update on the Mechanism of Action and Biological Activity

“…Following the investigations carried out on KP1019 and NAMI-A [72], in a recent study the subcellular accumulation of AziRu was determined by inductively coupled plasma-mass spectrometry (ICP-MS) analysis performed on various biological samples taken from MCF-7 breast cancer cell cultures (culture medium, cellular pellet, cytosolic fraction, nuclear fraction and DNA samples) after 24 h of in vitro ruthenium treatment [100]. About 80% of the administered ruthenium was found in the culture medium, while only a very small amount (less than 10%) was detected in the nuclear fraction [100]. This result corroborated the hypothesis of a massive early drug deactivation in the extracellular environment and/or a poor ability to penetrate the cell membranes, reinforcing the need to protect the metal core in Ru-based drugs in order to obtain more efficient cell internalization.…”

“…In detail, the low molecular weight complex AziRu was decorated using ribo-and deoxyribonucleosides as starting building blocks, functionalized with suitable hydrophilic (oligoethylene glycol moieties) and lipophilic (oleoyl or cholesteroxyacetyl groups) chains, obtaining a mini-library of amphiphilic nanovectors to enhance the Ru(III) delivery in vivo. Within this approach, the nucleolipid-based Ru(III) complexes-named ToThyRu, HoThyRu, DoHuRu [77,106], ToThyCholRu [79,105], the fluorescently-labelled HoThyDansRu [100,106] and the second-generation HoUrRu [107] (Figure 2)-were successfully prepared and evaluated in their physico-chemical properties (for a recent review covering their design, synthesis and characterization, see Riccardi et al [35]). (Figure 2)-were successfully prepared and evaluated in their physico-chemical properties (for a recent review covering their design, synthesis and characterization, see Riccardi et al [35]).…”

“…In order to verify the antiproliferative efficacy of the nucleolipid-or aminoacyl lipid-based Ru(III) complexes inserted in POPC-and DOTAP-based formulations, detailed investigations on their in vitro bioactivity were performed in comparison with naked AziRu [77,79,100,[105][106][107]110,134]. A selected panel of human and non-human cancer cell lines was chosen, particularly focusing on cells of different histopathological origin and widely used in anticancer research due to their replicative potential and malignancy, such as MCF-7 (human breast adenocarcinoma cell line), WiDr (human epithelial colorectal adenocarcinoma cell line), HeLa (human cervical cancer cells) and C6 (tumour rat glioma cells) [134][135][136].…”

“…In line with our expectations, liposomes exclusively composed by POPC or DOTAP did not interfere with cell viability and proliferation [77,105,106]. On the contrary, all the tested Ru(III)-containing POPCand DOTAP-based formulations inhibited cancer cell proliferation at least one order of magnitude more effectively than naked AziRu, with IC 50 values in the low µM range, indicative of a significant activity in reducing cancer cells growth (Table 1) Thus, the insertion of AziRu into highly functionalized scaffolds (nucleolipid or aminoacyl lipid structures), along with the subsequent liposome encapsulation, allowed us converting a very weakly cytotoxic compound into effective antiproliferative agents, with IC 50 values from ∼4 to ∼50 times lower than the parent metal complex [35,77,79,100,[105][106][107]110,134].…”

Induction of Hair Keratins Expression by an Annurca Apple-Based Nutraceutical Formulation in Human Follicular Cells

“…In addition, by using high in vitro concentrations (up to 3 mg/mL in the bioscreen), we have tested the nutraceutical for undesirable cellular responses, to validate its safety, and for complete biocompatibility. In detail, the experimental procedure involved the estimation of a "cell survival index," arising from the combination of cell viability evaluation with cell counting [27]. The cell survival index is calculated as the arithmetic mean between the percentage values derived from the MTT assay and the automated cell count, providing a more consistent indicator of cellular in vitro responses.…”