Cytotoxicity and mechanism of action of ruthenium(II)/amino acids/diphosphine complexes were evaluated against breast cancer (MDA-MB-231 cells), as well as their toxic effects in animal models.
Ruthenium (II)/amino acid/diphosphine complexes as apoptosis inducers in breast cancer cells and their toxic effects in
animal models.
Francyelli Mello-Andrade1,2, Clever G. Cardoso3, Paulo Melo-Reis4, Cesar Grisólia5, Carlos Castro6, Carlos Menck7, Alzir A.
Batista8, Elisângela Silveira-Lacerda1,2.
1 Laboratório de Oncologia Experimental, Instituto de Física, UFG;
2 Departamento de Genética, Instituto de Ciências Biológicas, UFG;
3 Departamento de Histologia, Embriologia e Biologia Celular, Instituto de Ciências Biológicas, UFG;
4 Departamento de Biomedicina, PUCGO;
5 Departamento de Genética e Morfologia, UnB;
6 Departamento de Fisiologia, Instituto de Ciências Biológicas, UFG;
7 Departamento de Microbiologia, Instituto de Ciências Biomédicas, USP;
8 Departamento de Química, UFSCar.
Cytotoxicity and mechanism of action of ruthenium(II)/amino acids/diphosphine complexes were evaluated against breast
cancer (MDA-MB-231 cells), as well as their toxic effects in animal models. The cell death type induced in MDA-MB-231
cells by compounds was evaluated measuring Annexin V-positive number, the activated caspases levels, and by morphological
features. In order to clarify which mechanisms are responsible for led breast cancer cells to death, we evaluated whether these
compounds can cause DNA damage, changes in cell cycle kinetic, mitochondrial dysfunction, and ultrastructural alterations in
MDA-MB-231 cells. As toxicity assessment is a required step for preclinical study of novel metal based compounds, the acute
oral toxicity was evaluated, and the genetic toxicity was determined by Micronucleus (MN) and Comet assay protocols on
Swiss mice treated with these complexes. Zebrafish model was used to evaluate toxicity of Ru(II)/amino acids complexes
during embryonic and larval development, more specifically, the mortality and hatching rates of zebrafish were determined.
Ru(II)/amino acids complexes induced apoptosis in MDA-MB-231 cells by increase in number of Annexin V-positive cells,
morphological changes, loss of mitochondrial membrane potential, and caspases-3 and 7 activation. Although these
compounds have a weak interaction to DNA molecule, it was observed DNA damage, probably due to reactive oxygen species
production related to mitochondrial membrane depolarization. As can be seen by ultrastructural analysis, these complexes can
reduce mitochondrial amount in MDA-MB-231 cells. Thus, Ru(II)/amino acids complexes are more active for tumor cells, and
their mechanism of action are related to induction cell cycle block, DNA damage, and mitochondrial dysfunction, lead to
apoptosis involving p53, PARP cleavage and caspases activation. In addition, these compounds were very well tolerated orally,
and it was observed lack of micronuclei formation in bone marrow cells and low DNA damage in peripherical blood cells from
Swiss mice. As regard to toxicity on zebrafish embryo development, the compounds caused low embryotoxicity, being mainly
observed hatching delay and mortality at high concentrations. Altogether our findings suggest that ruthenium (II)/amino
acids/diphosphine complexes induce cellular and molecular responses in breast cancer cells leading to mitochondria-mediated
apoptosis, and they are not harmful, presenting low systemic toxic effects.
Key words: ruthenium, preclinical tests, side-effects, apoptosis, ROS.