Head and neck squamous cell carcinoma (HNSCC) is recognized as the sixth most common cancer globally, characterized by its aggressive behavior and a high tendency to metastasize to adjacent lymph nodes (Johnson et al., 2020; Siegel et al., 2024). Despite treatment options improving, patients with advanced HNSCC continue to face low survival rates (Gavrielatou et al., 2020; Leemans et al., 2018). The outcome of HNSCC patients is predominantly influenced by a range of clinicopathological factors, including the depth of tumor invasion, the presence of cervical lymph node metastases, and the clinical staging of the disease(Mody et al., 2021). The prognostic markers for HNSCC, however, depend heavily on the tumor-node-metastasis (TNM) staging system, which often fails to accurately predict patient survival due to the complexity and variability of tumors among different patients (Johnson et al., 2020). Therefore, identifying novel and reliable prognostic biomarkers and therapeutic targets is essential for enhancing treatment effectiveness in HNSCC.

In order to preserve intracellular equilibrium, cellular mechanisms are meticulously orchestrated in both spatial and temporal dimensions. Liquid-liquid phase separation (LLPS) plays a pivotal role in the generation of biomolecular condensates, which is essential for the regulation of cellular activities (Mehta & Zhang, 2022; Tong et al., 2022). LLPS represents a natural physiological phenomenon that promotes the spontaneous segregation of a uniform solution into distinct dilute and dense phases (Shin & Brangwynne, 2017). Studies show that LLPS is pivotal in controlling tumor initiation, progression, metastasis, and therapeutic resistance across various malignancies (Tong et al., 2022; Zheng et al., 2023). For instance, in breast cancer, protein condensates that arise from LLPS influence the mechanisms of DNA damage repair, thereby increasing the resistance of malignant tumor cells to chemotherapy (Wei et al., 2023). In prostate cancer, LLPS can facilitate the activation of enhancers, promoting increased transcriptional activity of relevant genes (Chen et al., 2023). Besides, recent studies have increasingly acknowledged the significance of LLPS in the advancement and therapeutic approaches for HNSCC, particularly in its effects on mRNA splicing and mitochondrial metabolic function (Zhang et al., 2024).

To date, LLPS-related genes (LRGs) have been utilized to create prognostic signatures with satisfactory predictive performance in several studies (You et al., 2023; Zheng et al., 2022). However, the association between LLPS and HNSCC has yet to be thoroughly investigated. The objective of this research was to create and substantiate a new prognostic signature for patients with HNSCC based on LRGs using an integrative bioinformatics approach. Additionally, a specific gene of interest was chosen for biological validation through an in vitro functional knockdown assay.