High-grade serous ovarian cancer (HGSOC) accounts for 70% of ovarian cancer cases, and the survival rate remains remarkably low due to the lack of effective long-term consolidation therapies. Clinical remission can be temporarily induced by platinum-based chemotherapy, but death subsequently results from the extensive growth of a platinum-resistant component of the tumor. This work explores a novel treatment against HGSOC using the gold complex auranofin (AF). AF primarily functions as a pro-oxidant by inhibiting thioredoxin reductase (TrxR), an antioxidant enzyme overexpressed in ovarian cancer. We investigated the effect of AF on TrxR activity and the various mechanisms of cytotoxicity using HGSOC cells that are clinically sensitive or resistant to platinum. In addition, we studied the interaction between AF and another pro-oxidant, L-buthionine sulfoximine (L-BSO), an anti-glutathione (GSH) compound. We demonstrated that AF potently inhibited TrxR activity and reduced the vitality and viability of HGSOC cells regardless of their sensitivities to platinum. We showed that AF induces the accumulation of reactive oxygen species (ROS), triggers the depolarization of the mitochondrial membrane, and kills HGSOC cells by inducing apoptosis. Notably, AF-induced cell death was abrogated by the ROS-scavenger N-acetyl cysteine (NAC). In addition, the lethality of AF was associated with the activation of caspases-3/7 and the generation of DNA damage, effects that were also prevented by the presence of NAC. Finally, when AF and L-BSO were combined, we observed synergistic lethality against HGSOC cells, which was mediated by a further increase in ROS and a decrease in the levels of the antioxidant GSH. In summary, our results support the concept that AF can be used alone or in combination with L-BSO to kill HGSOC cells regardless of their sensitivity to platinum, suggesting that the depletion of antioxidants is an efficient strategy to mitigate the course of this disease.

In the field of experimental therapeutics for oncology purposes researchers are continuously evaluating the toxicity of novel treatment approaches against cancer cells. Within this topic of research, it is highly critical to define parameters of toxicity that denote when cancer cells are perturbed in their functionality by a new investigational drug. As the goal for these approaches is to achieve cellular demise, then what approaches to use and what do they mean in terms of assessing such cell death is of critical importance. In this comment article we highlight the definition of vitality and differentiate it from viability, and further define clonogenic survival in a chronic fashion. Additionally, we highly recommend the use of the term cytotoxicity as a general descriptor indicating toxicity towards a cell, but within that we encourage to sub-classify it as either cytostasis (i.e., when a treatment does not allow a cell to grow but it does not kill it either), or lethality (when a cell dies in response to the treatment). A more precise use of these terms should help advance the field of experimental therapeutics in oncology towards better defining the mechanisms of action of novel investigational drugs.

Reactive oxygen species (ROS) play a pivotal role in apoptosis. We reported that Blue Light (BL) induced oxidative stress in human retinal pigment epithelial (RPE) cells in vitro and increased drusen deposition and RPE cell apoptosis in human eyes. Here, we investigated the mechanisms underlying BL-induced damage to RPE cells. Cells were exposed to BL with or without the antioxidant N-acetylcysteine. Cells were analyzed for levels of ROS, proliferation, viability, and mitochondria membrane potential (ΔΨ_M) fluctuation. We performed proteomic analyses to search for differentially expressed proteins. ROS levels increased following RPE cell exposure to BL. While ROS production did not affect RPE cell proliferation, it was accompanied by decreased ΔΨ_M and increased cell apoptosis due to the caspase cascade activation in a ROS-dependent manner. Proteomic analyses revealed that BL decreased the levels of ROS detoxifying enzymes in exposed cells. We conclude that BL-induced oxidative stress is cytotoxic to RPE cells. These findings bring new insights into the involvement of BL on RPE cell damage and its role in the progression of age-related macular degeneration. The use of antioxidants is an avenue to block or delay BL-mediated RPE cell apoptosis to counteract the disease progression.

Although numerous drugs have been tested to treat ovarian cancer (OC), survival rates remain low as there has been no major improvement from platinum (Pt)-based therapy and there is a high rate of Pt resistance in these tumors. Following several rounds of chemotherapy, OC cells develop Pt-resistance by increasing DNA repair and antioxidant defense mechanisms. This study aimed to design a treatment to combat recurrent stages of OC by repurposing the anti-rheumatic gold complex auranofin (AF). We demonstrate that AF enhances the efficacy of cisplatin (CDDP) in Pt-resistant epithelial OC (EOC) cells. The drug combination enhanced mitochondrial-dependent apoptosis, PARP cleavage, DNA damage, and ROS overproduction. These results suggest the potential to combine AF with CDDP as a strategy to improve CDDP sensitivity in recurrent EOCs.

Background: Although rare, uveal melanoma (UM) is a life-threatening malignancy. Understanding its biology is necessary to improve disease outcome. Three-dimensional (3D) in vitro culture methods have emerged as tools that incorporate physical and spatial cues that better mimic tumor biology and in turn deliver more predictive preclinical data. Herein, we comprehensively characterize UM cells under different 3D culture settings as a suitable model to study tumor cell behavior and therapeutic intervention. Methods: Six UM cell lines were tested in two-dimensional (2D) and 3D-culture conditions. For 3D cultures, we used anchorage-dependent (AD) methods where cells were embedded or seeded on top of basement membrane extracts and anchorage-free (AF) methods where cells were seeded on agarose pre-coated plates, ultra-low attachment plates, and on hanging drops, with or without methylcellulose. Cultures were analyzed for multicellular tumor structures (MCTs) development by phase contrast and confocal imaging, and cell wellbeing was assessed based on viability, membrane integrity, vitality, apoptotic features, and DNA synthesis. Vascular endothelial growth factor (VEGF) production was evaluated under hypoxic conditions for cell function analysis. Results: UM cells cultured following anchorage-free methods developed MCTs shaped as spheres. Regardless of their sizes and degree of compaction, these spheres displayed an outer ring of viable and proliferating cells, and a core with less proliferating and apoptotic cells. In contrast, UM cells maintained under anchorage-dependent conditions established several morphological adaptations. Some remained isolated and rounded, formed multi-size irregular aggregates, or adopted a 2D-like flat appearance. These cells invariably conserved their metabolic activity and conserved melanocytic markers (i.e., expression of Melan A/Mart-1 and HMB45). Notably, under hypoxia, cells maintained under 3D conditions secrete more VEGF compared to cells cultured under 2D conditions. Conclusions: Under an anchorage-free environment, UM cells form sphere-like MCTs that acquire attributes reminiscent of abnormal vascularized solid tumors. UM cells behavior in anchorage-dependent manner exposed diverse cells populations in response to cues from an enriched extracellular matrix proteins (ECM) environment, highlighting the plasticity of UM cells. This study provides a 3D cell culture platform that is more predictive of the biology of UM. The integration of such platforms to explore mechanisms of ECM-mediated tumor resistance, metastatic abilities, and to test novel therapeutics (i.e., anti-angiogenics and immunomodulators) would benefit UM care.

High-grade serous cancer is the most common type of ovarian cancer and is usually diagnosed at advanced stages with high mortality due to recurrence and eventual resistance to standard platinum therapy. The aim of this study was to compare two-dimensional (2D) versus tridimensional (3D) cell culture as a preclinical model of response to carboplatin, paclitaxel and niraparib using PEO1, PEO4 and PEO6 cell lines, which were generated from the same patient along disease progression. Morphologically, cells formed flat adherent layers versus spheroidal structures with different compaction patterns in 2D and 3D respectively. In 2D, apoptosis was rare whereas in 3D cells formed a multilayered structure with an outer layer of live proliferating cells and an inner core of apoptotic cells. Furthermore, a differential capacity to produce ATP was observed among the cell lines in 3D but not in 2D. While response to carboplatin, paclitaxel and niraparib in both settings followed a similar trend, a lower sensitivity was observed in 3D with respect to 2D. Overall, 3D cell culture is likely more reflective of the in vivo cellular tumor behavior and more suitable of therapeutic evaluation given its added complexity absent in 2D.