KLF9-TXNRD2 axis in melanoma progression and metastasis

KLF9-TXNRD2 轴在黑色素瘤进展和转移中的作用

• 批准号: 9108882
• 负责人: Mikhail Nikiforov
• 金额: $ 40.68万
• 依托单位: ROSWELL PARK CANCER INSTITUTE CORP
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-10 至 2020-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9108882
• 关键词: Accounting; Affect; Anchorage-Independent Growth; Anoikis; Antineoplastic Agents; Antioxidants; Antirheumatic Agents; Apoptosis; Auranofin; BRAF gene; Benign; Cell Death; Cell Line; Cell Nucleus; Cell Survival; Cells; Cessation of life; Chemicals; Clinical Trials; Data; Databases; Development; Disease; Down-Regulation; Drug Metabolic Detoxication; Ectopic Expression; Extracellular Matrix; FDA approved; Genes; Genotype; Goals; Health; Human; In Vitro; Knockout Mice; Kruppel-like transcription factors; Lung; Maintenance; Malignant Neoplasms; Melanocytic nevus; Melanoma Cell; Messenger RNA; Metastatic Melanoma; Metastatic Neoplasm to Lymph Nodes; Mitochondria; Modeling; Molecular; Mus; Mutation; NRAS gene; Neoplasm Metastasis; Nevus; Nuclear; Oncogenes; Outcome; Oxidative Phosphorylation; Oxidative Stress; Patients; Phenotype; Process; Production; Proteins; Publishing; Reactive Oxygen Species; Regulation; Reporting; Research; Resistance; Role; Staging; Testing; The Cancer Genome Atlas; Transcription Coactivator; Transgenes; Transgenic Mice; Tumor Suppressor Proteins; Tumorigenicity; Up-Regulation; Xenograft procedure; cancer cell; cancer type; drug development; effective therapy; improved; inhibitor/antagonist; killings; melanocyte; melanoma; mouse model; mutant; novel; novel therapeutics; overexpression; response; senescence; thioredoxin reductase; treatment strategy; tumor; tumor progression; tumorigenesis

 DESCRIPTION (provided by applicant): Malignant melanoma is one of the most aggressive types of human cancer. Its ability to metastasize makes melanoma extremely difficult to cure, and consequently, the median survival of patients with metastatic melanoma is only 8.5 months. The molecular mechanisms underlying melanoma development are not well understood, and effective treatment is not currently available. Melanomas often originate from nevi, benign aggregates of senescent melanocytes harboring activating mutations in BRAF, NRAS genes. The causal role of BRAFV600E or NRASQ61R in induction of senescence (termed oncogene-induced senescence (OIS)) has been demonstrated in normal human melanocytes (NHM) and mouse models. Ectopic expression of these oncogenes in NHM leads to accumulation of reactive oxygen species (ROS), whereas chemical suppression of ROS abrogates senescence. Endogenous mechanisms suppressing OIS in NHM via down-regulation of excessive ROS have not been investigated. Metastasis is the most detrimental feature of melanoma. Detachment of cancer cells from the extracellular matrix often causes a specific form of apoptosis, termed anoikis. Invasion, anchorage-independent growth and abrogation of anoikis are critical steps of metastasis. Several independent studies have suggested that increase in ROS levels promotes anoikis and suppresses other transformed phenotypes. Mechanisms controlling these processes via regulation of intracellular ROS in melanoma cells are largely unknown. Tumors often develop adaptive mechanisms to suppress high levels of ROS. The most general one detected in multiple malignancies is an increase in the amounts of NRF2, a universal regulator of antioxidant response genes. Increase in NRF2 has also been shown in melanomas. Recently, we have discovered that paradoxically under conditions of high oxidative stress NRF2 amplifies ROS. It is well known that increase in ROS causes accumulation of NRF2 in the nucleus and activation of anti-oxidant genes. However, if ROS continue to elevate and exceed a critical threshold, NRF2 induces expression of a transcription factor KLF9 (Kruppel-like factor 9). KLF9 represses several anti-oxidant genes, which are not NRF2 targets, including mitochondrial thioredoxin reductase (TXNRD2), thus resulting in ROS amplification. TXNRD2 protein is critical for maintenance of intracellular red-ox status and ROS detoxification. We demonstrated that TXNRD2 depletion generates ROS, whereas its overexpression lowers KLF9-generated ROS. Accordingly, depletion of KLF9 inhibits oxidative stress in all tested cell lines. Thus, KLF9 downregulation or TXNRD2 upregulation could be beneficial for the malignancies where oxidative stress (that hinders tumor progression) overwhelms NRF2- dependent anti-oxidant defense. According to the TCGA database, high KLF9 or low TXNRD2 mRNA levels correlate with the overall survival of patients with lymph node metastases (major determinants of outcome for melanoma patients), suggesting the importance of both genes for melanoma progression. Furthermore, we confirmed that KLF9 directly regulates TXNRD2 in melanocytic cells. Combinational treatment is critical for overcoming resistance to BRAFV600E inhibitor vemurafenib (VEMU). Several independent reports characterized activation of oxidative phosphorylation (which is accompanied by oxidative stress) as an adaptive response to BRAFV600E inhibition in melanoma cells. Our preliminary data demonstrate that oxidative stress-inducing agent AUR (a FDA-approved antirheumatic drug) synergizes with VEMU in inducing cell death in BRAFV600E-melanoma cells. Additionally, AUR is equally potent in causing cell death as a single agent in wildtype and mutant BRAF melanoma cells. This finding is important since no effective treatment exists for wildtype BRAF melanoma patients. The proposed research will establish the mechanisms by which KLF9-TXNRD2 axis regulates OIS in normal melanocytic cells and multiple transformed phenotypes in melanoma cells. Our findings will have broad scientific appeal since the suppression of OIS is a prerequisite for tumorigenesis, and suppression of anoikis, increase in invasion and anchorage-independent growth are prerequisites for metastasis, the most detrimental feature of cancer. Additionally, repurposing of anti-rheumatic agent auranofin for the treatment of malignant melanomas may significantly improve management of this very aggressive disease.

 描述（由适用提供）：恶性黑色素瘤是人类癌症中最具侵略性的类型之一。它的黑色素瘤能力使黑色素瘤极难治愈，因此，转移性黑色素瘤患者的中位存活率仅为8.5个月。黑色素瘤发育产生的分子机制尚不清楚，目前尚无有效治疗。黑色素瘤通常起源于Nevi，是含有BRAF，NRAS基因激活突变的感觉黑素细胞的良性聚集体。在正常的人类黑色素细胞（NHM）和小鼠模型中已证明了Sectim（称为癌基因诱导的感应（OIS））的BRAFV600E或NRASQ61R诱导（称为癌基因诱导的感应（OIS））的因果作用。这些肿瘤基因在NHM中的异位表达导致活性氧的积累（ROS），而化学抑制ROS会消除敏感性。尚未研究通过下调过量ROS，抑制NHM中OI的内源性机制尚未被研究。转移是黑色素瘤最有害的特征。癌细胞与细胞外基质的分离通常会引起特定形式的凋亡形式，称为厌氧。入侵，独立于锚定的生长和消除Anoikis是转移的关键步骤。几项独立研究表明，ROS水平的升高会促进Anoikis并抑制其他转化的表型。通过调节黑色素瘤细胞中细胞内ROS来控制这些过程的机制在很大程度上尚不清楚。肿瘤通常会发展自适应机制以抑制高水平的ROS。在多个恶性肿瘤中检测到的最通用的是NRF2的量增加，NRF2是抗氧化反应基因的通用调节剂。 NRF2的增加也已在黑色素瘤中显示。最近，我们发现在高氧化应激NRF2放大器ROS的条件下矛盾。众所周知，ROS的增加导致NRF2在细胞核中的积累和抗氧化基因的激活。但是，如果ROS继续升高并超过临界阈值，则NRF2诱导转录因子KLF9（Kruppel样因子9）的表达。 KLF9反映了几种不是NRF2靶标的抗氧化基因，包括线粒体硫氧还蛋白还原（TXNRD2），从而导致ROS扩增。 TXNRD2蛋白对于维持细胞内红色氧气状态和ROS排毒至关重要。我们证明TXNRD2耗竭会产生ROS，而其过表达降低了KLF9生成的ROS。根据以下内容，KLF9的耗竭抑制了所有测试的细胞系中的氧化应激。这是KLF9下调或TXNRD2上调对氧化胁迫（阻碍肿瘤进展）的恶性肿瘤可能是有益的。根据TCGA数据库，较高的KLF9或低TXNRD2 mRNA水平与淋巴结转移患者的总体存活率相关（黑色素瘤患者的预后主要确定），这表明这两个基因对黑色素瘤进展的重要性。此外，我们确认KLF9直接调节黑色素细胞中的TxNRD2。组合治疗对于克服对BRAFV600E抑制剂vemurafenib（VEMU）的抗性至关重要。几份独立的报道将激活氧化磷酸化（通过氧化应激完成）的激活作为对黑色素瘤细胞中BRAFV600E抑制的适应性反应。我们的初步数据表明，氧化应激诱导的AUR（FDA批准的抗肿瘤药物）与VEMU在BRAFV600E-甲状腺瘤细胞中诱导的细胞死亡中与VEMU协同作用。另外，AUR在野生型和突变体Braf黑色素瘤细胞中作为单一药物的细胞死亡同样潜力。这一发现很重要，因为对于野生型BRAF黑色素瘤患者没有有效的治疗方法。拟议的研究将建立KLF9-TXNRD2轴在正常黑素细胞中调节OIS的机制，而黑色素瘤细胞中的多个转化的表型。我们的发现将具有广泛的科学外观，因为OIS的抑制是肿瘤发生的先决条件，抑制Anoikis，侵袭和无锚固的生长是转移的先决条件，这是癌症最有害的特征。此外，对抗雨中剂Auranofin的重新利用用于治疗恶性黑色素瘤，可能会显着改善对这种非常侵略性疾病的管理。