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.

The chemistry of contrast agents (CAs) for magnetic resonance imaging (MRI) applications is an active area of research and, in recent work, it was shown that CA-based graphene oxide (GO) has valuable properties for biomedical uses. GO has a potential as MRI CAs thanks to several functionalities, like its ability to penetrate tissues and cell membranes, as well as easy coupling with therapeutic agents, therefore showing the potential for both a diagnostic and therapeutic role. In this study, we performed a thorough cleaning of the GO sample (synthesized using a modified Hummers method), minimizing the amount of residual manganese down to 73 ppm. Using a wide range of physical–chemical methods (morphology, chemical composition, elemental analysis, spectroscopies, and imaging), we characterized the intrinsic longitudinal and transverse relaxivities of highly purified GO nanosheets. X-band electron paramagnetic resonance allowed to recognize the paramagnetic species involved, and 1.0 T MRI was used to disentangle the relative contributions to the MRI contrast of pristine GO nanosheets arising from structural defects and residual paramagnetic manganese impurities embedded in the nanomaterial. Although experiments show that the MRI relaxivity of GO nanosheets arises from the cumulative effect of structural defects and paramagnetic impurities, we conclude that the latter contribution to the longitudinal and transverse relaxivities becomes irrelevant for highly purified (pristine) GO. This novel finding clearly demonstrates that, apart from trivial manganese inclusion, pristine GO produces an inherent MRI response via structural defects, and therefore it is on its own a suitable candidate as MRI contrast agent.

Graphene oxide (GO)-based nanomaterials have emerged as promising platforms in biomedicine, particularly for their potential use as carriers of magnetic resonance imaging (MRI) contrast agents (CAs). In this study, we investigate the MRI CA performance of reduced graphene oxide and its dependence on the degree of reduction. To ensure full control, GO was synthesized in-house via a modified Hummers’ method and subsequently reduced by two distinct approaches: mild thermal reduction (rGO1) and selective chemical reduction with sodium borohydride (rGO2). Comprehensive physicochemical characterization, including Fourier-transform infrared (FTIR) spectroscopy, Raman spectroscopy, X-ray photoelectron spectroscopy, differential scanning calorimetry, and thermogravimetric analysis (TGA), was performed to evaluate the structural changes induced by each reduction process. The MRI performance of GO, rGO1, and rGO2 was assessed at 1.0 T alongside electron paramagnetic resonance (EPR) measurements, while inductively coupled plasma-mass spectrometry (ICP-MS) quantified residual manganese and iron impurities. EPR and ICP-MS demonstrated that these impurities made a negligible contribution to the relaxivity. Our findings reveal a significant enhancement in MRI relaxivities with an increasing degree of reduction. Chemically reduced rGO2 exhibits the highest longitudinal relaxivity (r1), approximately 3.9-fold greater than that of pristine GO, and transverse relaxivity (r2), approximately 4.6-fold greater. Strong linear correlations between relaxivities and XPS-derived structural parameters, including the C/O ratio and C–C/C═C content, indicate that carbon structural defects govern MRI contrast performance. Overall, these results establish a framework for the rational design of rGO-based CAs through controlled reduction of two-dimensional carbon materials, enabling optimized imaging performance and supporting future biomedical translation.