Efeito da hipóxia sobre o acúmulo de células-tronco tumorais em câncer de cabeça e pescoço

Abstract

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common malignancies worldwide. Antiangiogenic drugs are a conventional treatment, although it has been shown to stimulate hypoxic areas inside the tumor, and hypoxia has been associated with metastasis as well as accumulation of cancer stem cell (CSC) in breast cancer. Objective: To study the effects of hypoxia on HNSCC behavior and accumulation of cancer stem cell. Material and methods: Culture of HN6, HN12 and HN13 tumor cells under hypoxic conditions was carried out for 6hrs, 12hrs, and 24hrs under an O2 concentration of 1%. The content of cancer stem cell was determined using Aldefluor assay according to the manufacturer's (STEMCELL Technologies) to detect ALDH1A1Bright cells in combination with APC mouse anti-human CD44. Tumor invasion under hypoxic conditions was carried out using Millicell Cell Culture Inserts (Millipore, Billerica, MA, USA). Control tumor cells were maintained in DMEM supplemented with 10% FBS and 1% antibiotics at normoxia levels (~21% of O2), while hypoxia group was maintained at 1% of O2. We used a PTEN inhibitor (BPV) concentrations of 1nM, 5nM, and 10nM. Results: PTEN expression is downregulated under hypoxic conditions. Hypoxia-induced PTEN downregulation triggered an epithelial-mesenchyme transition (EMT) phenotype and accumulation of CSC. Plus, we showed antagonistic expression of PTEN and HIF-1α when targeted disruption of PTEN (shPTEN HN13 cell line) strongly suggesting that loss of PTEN activates the PI3K/mTOR pathway in association with HIF-1αupregulated. Validation of our results in xenograft animal models showed colocalization of ALDH1A1Bright cells and hypoxic areas along with the loss of nuclear localization of PTEN. Conclusion: Thus, we show that hypoxia-driven downregulation of PTEN plays a crucial role in tumor aggressiveness and that the potential use of CSC-targeted therapies should be considered during the administration of anti-angiogenic drugs to minimize tumor resistance.