Nobiletin suppresses nasopharyngeal carcinoma by regulating the KEAP1/NRF2/ARE pathway

Abstract

Nasopharyngeal carcinoma (NPC) ranks among the most prevalent malignancies, particularly in East Asia and Southeast Asia. Nobiletin (NOB), an exclusive polymethoxyflavonoid derived from citrus peel, exhibits diverse physiological properties, notably its potent anticancer activity. Kelch-like ECH-associated protein 1 (KEAP1), the repressor protein regulating the nuclear factor erythroid 2-related factor 2 (NRF2) transcription factor, has emerged as a promising strategy for addressing oxidative stress in various diseases. The KEAP1/NRF2/ARE signal is a fundamental pathway within the cellular homeostatic defense system. This study robustly demonstrates the chemopreventive potential of NOB through comprehensive in vitro and in vivo assessments using subcutaneous tumor mouse models. Furthermore, our groundbreaking findings reveal that NOB effectively hinders the migration and invasion capacities of CNE-2 and 5-8F (NPC) cells in a dose- and time-dependent manner. Mechanistically, NOB, a potent KEAP1 activator, significantly disrupts the NRF2/ARE signaling pathway by accelerating the proteasomal degradation of NRF2 and suppressing its nuclear translocation. Consequently, this cascade reduces the expressions of ARE-driven genes and antioxidant enzymes, thereby increasing intracellular reactive oxygen species (ROS) levels and increasing antitumor immunity. Moreover, the sensitivity induced by NOB is markedly diminished in CNE-2 cells following the gene silencing of KEAP1. These findings underscore the pivotal role of NOB in activating KEAP1. Overall, KEAP1 has emerged as a compelling target for potential malignancy treatment in nasopharyngeal carcinoma cell lines. Our results suggest the promising application of NOB as a natural sensitizer in chemotherapy, opening avenues for promising therapeutic interventions.

1. Introduction

Nasopharyngeal carcinoma (NPC) is a type of malignancy originating from the nasopharyngeal mucosal lining and is categorized as head and neck epithelial carcinoma [1]. It has a high incidence among individuals in East Asia and Southeast Asia, especially in southern China [2]. Various factors closely linked to this malignancy include Epstein-Barr virus (EBV) infection, genetic susceptibility genes, environmental factors, and lifestyle choices. Although radiotherapy is the primary treatment modality and is often accompanied by adjuvant chemotherapy and excision, the overall 5-year survival rates remain unsatisfactory. The unavoidable acute and late toxicities induced by radiotherapy and/or chemotherapy, such as oral mucosal inflammation, gastrointestinal dysfunction, and bone marrow depression, contribute to low treatment adherence [3–6]. Despite advancements in diagnosis and treatment techniques [7–11], concerns persist regarding the high risk of tumor recurrence, aggressiveness, drug resistance, and poor prognosis [12–14]. Therefore, there is an urgent need to explore and develop new, improved treatments for NPC. The search for antioncogenic agents with high efficacy and minimal side effects is a primary focus in pharmacological research.

Nobiletin (NOB), a natural polymethoxyflavonoid (PMF) exclusively derived from citrus fruit peel, has gained increasing attention because of its pharmacological activities, including anti-inflammatory, antioxidative, antidiabetic, and organ function protection effects [15–20]. Notably, the antitumor activity of NOB, which is mediated through the regulation of multiple signal transduction pathways and mechanisms, has garnered increased attention in recent years [17,21–23]. The chemopreventive potential of NOB has also been convincingly demonstrated in various in vivo colon carcinogenesis animal models [24]. Furthermore, NOB exhibits anti-non-small cell lung cancer properties by suppressing the migration and invasion of A549 cells, inhibiting tumor evasion by the immune system, and reversing drug resistance to chemotherapeutic agents such as paclitaxel and azithromycin [25,26]. In our previous study, NOB significantly inhibited growth and inhibited apoptosis in human C666-1 NPC cells [27]. Despite its recognized role as a traditional natural medicine with presumed chemopreventive and chemotherapeutic adjuvant properties, the antitumor molecular mechanisms of NOB in NPC remain unclear and warrant further evaluation.