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Melanoma cell viability after PLX4032 treatment and HO-1 inhibition. Cell viability was determined using the MTT assay in MeOV-1 cells seeded into 96-well plates and adapted for 5 days to 18 kPa O2, 5 kPa or 1 kPa O2. Cells were then treated with 10 µM PLX4032, 10 µM SnMP and combined treatment for 24 h. Data denote mean ± S.E.M., n = 5 independent cell cultures, ★ p < 0.005 vs. respective CTR and DMSO; # p < 0.05 vs. respective CTR and DMSO; § p < 0.05 vs. respective PLX4032; §§ p < 0.005 vs. respective PLX4032; §§§ p < 0.001 vs. respective PLX4032.
Source publication
Induction of heme oxygenase 1 (HO-1) favors immune-escape in BRAFV600 melanoma cells treated with Vemurafenib/PLX4032 under standard cell culture conditions. However, the oxygen tension under standard culture conditions (~18 kPa O2) is significantly higher than the physiological oxygen levels encountered in vivo. In addition, cancer cells in vivo a...
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Citations
... In a study by Furfaro et al., MeOV-1 CM cells cultured under 18 kPa O 2 (normoxia), 5 kPa O 2 (physioxia), and 1 kPa O 2 (hypoxia) were treated with BRAFi. Of interest, HIF-1α expression was significantly decreased by the treatment under all O 2 tensions tested [118]. Consistently, BRAFi vemurafenib (PLX4032) suppressed HIF-1α expression at the mRNA and protein levels in a panel of BRAF V600 CM cell lines cultured under standard conditions. ...
Hypoxia is a common feature of solid malignancies, including cutaneous melanoma (CM). Hypoxia-inducible factor (HIF)-1α and HIF-2α orchestrate cellular responses to hypoxia and coordinate a transcriptional program that promote several aggressive features in CM, such as angiogenesis, epithelial-mesenchymal transition, metastasis formation, metabolic rewiring, and immune escape. BRAF V600E , which is the most frequent mutation observed in CM patients, usually increases HIF-α signaling not only in hypoxia, but also in normoxic CM cells, enabling HIF-1α and HIF-2α to continuously activate downstream molecular pathways. In this review, we aim to provide a comprehensive overview of the intricate role and regulation of HIF-1α and HIF-2α in CM, with a brief focus on the complex interactions between HIF-α subunits and non-coding RNAs. We also discuss HIF-α-mediated cellular responses in normoxia along with the mechanisms that allow HIF-α subunits to maintain their stability under normal oxygen conditions. Finally, we resume available evidence on potential therapeutic approaches aimed at targeting HIF-1α and/or HIF-2α.
... In BRAF V600E MeOV-1 melanoma cells, vemurafenib, a BRAF V600E inhibitor, showed limited efficacy because the expression of BACH1 was reduced, and HO-1 (encoded by HOMX1) was subsequently upregulated. HO-1 promoted the immune escape of melanoma cells, while the HO-1 inhibitor tin mesoporphyrin IX synergized with vemurafenib to effectively eradicate the tumor [118]. This suggests that inhibiting HO-1 may have a potential role in treatment. ...
Antioxidants play a pivotal role in neutralizing reactive oxygen species (ROS), which are known to induce oxidative stress. In the context of cancer development, cancer cells adeptly maintain elevated levels of both ROS and antioxidants through a process termed “redox reprogramming”. This balance optimizes the proliferative influence of ROS while simultaneously reducing the potential for ROS to cause damage to the cell. In some cases, the adapted antioxidant machinery can hamper the efficacy of treatments for neoplastic diseases, representing a significant facet of the resistance mechanisms observed in cancer therapy. In this review, we outline the contribution of antioxidant systems to therapeutic resistance. We detail the fundamental constituents of these systems, encompassing the central regulatory mechanisms involving transcription factors (of particular importance is the KEAP1/NRF2 signaling axis), the molecular effectors of antioxidants, and the auxiliary systems responsible for NADPH generation. Furthermore, we present recent clinical trials based on targeted antioxidant systems for the treatment of cancer, assessing the potential as well as challenges of this strategy in cancer therapy. Additionally, we summarize the pressing issues in the field, with the aim of illuminating a path toward the emergence of novel anticancer therapeutic approaches by orchestrating redox signaling.
... All the experiments were performed on MeOV (BRAF V600E ) and MeTA (BRAF V600D ) metastatic melanoma cell lines isolated from the biopsies of TT-untreated patients (17). Melanoma cells were maintained in RPMI 1640 medium (Euroclone Spa, Pavia, Italy) supplemented with 10% Fetal Bovine Serum (FBS, Euroclone Spa, Pavia, Italy), 1% L-Glutamine (Euroclone Spa, Pavia, Italy) and 1% Penicillin/Streptomicin (Euroclone Spa, Pavia, Italy) and grown under standard conditions (37°C humidified incubator with 5% CO2). ...
Background
Malignant melanoma is the most lethal form of skin cancer which shows BRAF mutation in 50% of patients. In this context, the identification of BRAFV600E mutation led to the development of specific inhibitors like PLX4032. Nevertheless, although its initial success, its clinical efficacy is reduced after six-months of therapy leading to cancer relapse due to the onset of drug resistance. Therefore, investigating the mechanisms underlying PLX4032 resistance is fundamental to improve therapy efficacy. In this context, several models of PLX4032 resistance have been developed, but the discrepancy between in vitro and in vivo results often limits their clinical translation.
Methods
The herein reported model has been realized by treating with PLX4032, for six months, patient-derived BRAF-mutated melanoma cells in order to obtain a reliable model of acquired PLX4032 resistance that could be predictive of patient’s treatment responses. Metabolic analyses were performed by evaluating glucose consumption, ATP synthesis, oxygen consumption rate, P/O ratio, ATP/AMP ratio, lactate release, lactate dehydrogenase activity, NAD⁺/NADH ratio and pyruvate dehydrogenase activity in parental and drug resistant melanoma cells. The intracellular oxidative state was analyzed in terms of reactive oxygen species production, glutathione levels and NADPH/NADP⁺ ratio. In addition, a principal component analysis was conducted in order to identify the variables responsible for the acquisition of targeted therapy resistance.
Results
Collectively, our results demonstrate, for the first time in patient-derived melanoma cells, that the rewiring of oxidative phosphorylation and the maintenance of pyruvate dehydrogenase activity and of high glutathione levels contribute to trigger the onset of PLX4032 resistance.
Conclusion
Therefore, it is possible to hypothesize that inhibitors of glutathione biosynthesis and/or pyruvate dehydrogenase activity could be used in combination with PLX4032 to overcome drug resistance of BRAF-mutated melanoma patients. However, the identification of new adjuvant targets related to drug-induced metabolic reprogramming could be crucial to counteract the failure of targeted therapy in metastatic melanoma.
Over the past 55 years, the heme oxygenase (HO) system has emerged as a pivotal player in a myriad of cellular, tissue, and integrative physiological processes [...]
