Abstract 4990: A discovery of novel methylation profiles in head and neck squamous cell carcinoma in relation to pretreatment pain by cancer subtype

Abstract Pain is the initial symptom reported by nearly 75% of individuals diagnosed with head and neck squamous cell carcinoma (HNSCC), yet effective management remains elusive, with up to 80% of patients reporting inadequate pain management despite opioid use. Pretreatment pain is a key predictor of survival and recurrence, but the molecular mechanisms underlying pain in HNSCC are not well understood. This pilot study aims to explore the epigenetic mechanisms underlying pain perception in HNSCC patients by cancer subtype and identify potential biomarkers to improve pain management strategies. DNA methylation levels were generated for individuals newly diagnosed with HNSCC of oral cavity (OCSCC; n=18) or oropharynx (OPSCC; n=8) prior to definitive treatment using the Illumina Infinium MethylationEPIC v2.0 array. Perceived worst pain score was measured using the Brief Pain Inventory-Worst Pain item prior to treatment, categorized according to NCCN guidelines, with patients reporting no pain serving as the reference group. After quality control, M-values for 26 samples and 913, 962 methylation probes were evaluated. Differentially methylated positions (DMPs) associated with pain severity were identified combining subtypes, with hypo- (fold change (FC) < -2) and hypermethylated (FC > 2) sites determined after adjusting for age. Stratified analyses were performed by cancer type. All analyses set a significance level with a false discovery rate < 5%. Pain phenotype and blood collection occurred on the same day, after diagnosis but prior to definitive treatment. The cohort had a mean age of 65.1 ± 11.3 years, 42.3% of those reporting any level of pain were male. Pre-treatment pain was reported by 94.4% (N=17) of OCSCC and 25% (N=2) of OPSCC patients. DMP analysis revealed probe cg08573204_TC21 in LMO1 gene was hypermethylated in OCSCC patients with mild pain versus no pain (p=0.02, Log2FC=1.11). While LMO1 has not been previously linked to pain, its role in adrenergic cell identity suggests potential involvement in pain pathways. In patients with moderate/severe pain, cg18566479_BC21 located in CHDH gene was hypermethylated in OCSCC (p=1.09E-04, Log2FC=1.98). CHDH, associated with metabolic signatures in HNSCC and neuroinflammatory activation in neuropathic pain, showed significant hypermethylation, suggesting its potential as a biomarker for pain management in HNSCC. Differential DNA methylation may be associated and predictive of the development and severity of pain in HNSCC. With future work in larger samples, this research may provide insight into the biological underpinnings of pain and guide cancer symptom management by using DNA methylation profiles to identify patients at risk for pain who may benefit from preventative measures. Citation Format: Monica A. Wagner, Yanning Wu, Naji Ayyash, Fredrick R. Schumacher, Quintin Pan. A discovery of novel methylation profiles in head and neck squamous cell carcinoma in relation to pretreatment pain by cancer subtype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4990.