(0679) Metabolomic Profiling of Nodal Metastasis in HPV-Negative Head and Neck Squamous Cell Carcinoma

Introduction: Regional metastasis to the lymph nodes is a key prognostic factor in head and neck squamous cell carcinomas (HNSCC), occurring in over 40% of patients at initial diagnosis. The exact alterations that underly nodal metastasis in HNSCC are incompletely understood, presenting a challenge to effective treatment. The rationale for this study is to improve our understanding of metabolic drivers of nodal metastases and identify prognostic markers by correlating specific profiles to clinical outcomes.

Methods: This single-institution case-control study included n = 42 HPV-negative pT3-4 HNSCC cases from 2019-2022 without neoadjuvant treatment, staged pN0 or pN1+ to delineate the presence (n = 21 cases) or absence (n = 21 controls) of nodal metastasis. Fresh tumor, normal, and lymph node tissue was prospectively collected from surgical cases and snap frozen in liquid N2 immediately after excision. RNAseq and untargeted metabolomics screening using liquid chromatography mass spectrometry (LC–MS) was performed on all samples in the cohort.

Results: Metabolomic analysis revealed multiple metabolites were increased in the metastatic pN1+ samples, including many long chain fatty acids and phospholipids (phosphatidylcholines, phosphatidylethanolamines, and phosphatidylserines). Pathway analysis demonstrated significant differences in the oxidation of branched chain fatty acids; glycerophospholipid metabolism; and butanoate metabolism pathways between the metastatic and non-metastatic groups (p < 0.05). Preliminary gene set enrichment analyses of RNAseq data showed enrichment of hallmark gene sets for fatty acid metabolism (p < 0.01) and cholesterol homeostasis (p < 0.04) in the N0 compared to N1+ metastatic tumor tissue. This relationship was inverted when comparing normal tissue samples.

Conclusions: These preliminary results indicate a potential functional importance of fatty acid and phospholipid metabolism in lymph node metastasis. Further analysis within these pathways is ongoing in the hopes of elucidating a potential mechanism and biomarkers underlying HPV-negative nodal metastasis in HNSCC.