DOI: 10.3390/cancers18193104 ISSN: 2072-6694

A ZIP-Active Tumor Cell State Defines an NRF2-Associated Ferroptosis-Resistant and Immune-Cold Transcriptional Program in Head and Neck Squamous Cell Carcinoma

Hideyuki Takahashi, Miho Uchida, Kaoru Takahashi, Hiroyuki Hagiwara, Hiroe Tada, Toshiyuki Matsuyama, Kazuaki Chikamatsu

Background/Objectives: Zinc homeostasis has emerged as an important regulator of cancer biology, yet the biological and clinical significance of zinc transporter-associated transcriptional programs in head and neck squamous cell carcinoma (HNSCC) remains poorly understood. This study aimed to identify zinc transporter-associated epithelial tumor cell states and evaluate their molecular and clinical significance. Methods: Single-cell RNA sequencing data from an HNSCC cohort were analyzed to identify epithelial tumor cell populations associated with zinc transporter expression and to characterize their molecular features. A ZIP-active transcriptional signature was subsequently applied to The Cancer Genome Atlas (TCGA)-HNSCC cohort to evaluate biological characteristics, immune features, survival outcomes, and the influence of NFE2L2/KEAP1/CUL3 mutation status. Results: A distinct epithelial tumor cell population characterized by coordinated upregulation of SLC39A3 and SLC39A4 was identified and designated the ZIP-active tumor cell state. This population showed NRF2-associated oxidative stress adaptation, glutathione metabolism, and ferroptosis suppression, together with increased NRF2 activity. In the TCGA-HNSCC cohort, a high ZIP-active score was associated with oxidative stress adaptation, reduced T-cell-related immune signatures, and poorer overall survival. NFE2L2/KEAP1/CUL3-mutant tumors showed higher ZIP-active and NRF2-related scores than wild-type tumors, while NRF2-related metabolic features remained associated with ZIP activity in both groups. Conclusions: The ZIP-active tumor cell state represents an NRF2-linked metabolic program associated with oxidative stress adaptation, immune alterations, and adverse prognosis in HNSCC. Its persistence across NFE2L2/KEAP1/CUL3 mutation groups suggests that ZIP activity reflects NRF2-related tumor biology beyond genomic alterations alone.