Hydroxytyrosol Counteracts Triple Negative Breast Cancer Cell Dissemination via Its Copper Complexing Properties

SIMPLE SUMMARY: Several naturally occurring substances, known as polyphenols, may be found in a variety of plant-based foods and drinks, including fruits, vegetables, whole grains, tea, and red wine. Several studies have shown that these molecules have many beneficial effects on human health, particularly in their ability to counteract tumor growth thanks to their capacity to dampen inflammatory processes. Furthermore, polyphenols can form complexes with certain metals, such as copper. This property is of great importance considering that copper is closely involved in both the early stages and progression of cancer. In this study, we investigated the capacity of hydroxytyrosol, a derivative of oleuropein found in the leaves and fruits of the Olea europaea plant, to complex with copper and, consequently, to counteract the progression of triple-negative breast cancer. We chose this type of cancer as it is the most aggressive subtype due to the lack of specific receptors, which results in the absence of targeted oncological therapy. Our results demonstrate that hydroxytyrosol forms a complex with copper, resulting in the reduction of its content within the triple-negative breast cancer cell, consequently reducing its aggressiveness and, eventually, its ability to form metastases. ABSTRACT: Polyphenols have gained increasing attention for their therapeutic potential, particularly in conditions like cancer, due to their established antioxidant and anti-inflammatory properties. Recent research highlights their ability to bind to transition metals, such as copper. This is particularly noteworthy given the key role of copper both in the initiation and progression of cancer. Copper can modulate the activity of kinases required for the epithelial–mesenchymal transition (EMT), a process fundamental to tumor cell dissemination. We have previously demonstrated the copper-binding capacity of oleuropein, a secoiridoid found in Olea europaea. In the present study, we investigated the effect of hydroxytyrosol, the primary oleuropein metabolite, on the metastatic potential of three triple-negative breast cancer cell lines (MDA-MB-231, MDA-MB-468, and SUM159). We found that hydroxytyrosol modulated the intracellular copper levels, influencing both the epithelial and mesenchymal markers, by downregulating copper-dependent AKT phosphorylation, a member of the EMT signaling cascade, through Western blot, RT-qPCR, and immunofluorescence. Indeed, by optical spectra, EPR, and in silico approaches, we found that hydroxytyrosol formed a complex with copper, acting as a chelating agent, thus regulating its homeostasis and affecting the copper-dependent signaling cascades. While our results bring to light the copper-chelating properties of hydroxytyrosol capable of countering tumor progression, they also provide further confirmation of the key role of copper in promoting the aggressiveness of triple-negative breast cancer cells.