Postoperative Lymphatic Exudate is a Proximal Source of ctDNA and Detects Recurrence in HPV-negative Head and Neck Cancer

Relapse is a major cause of failure after surgery in HPV-human papillomavirus (HPV) negative head and neck squamous cell carcinoma (HNSCC),), with up to 50% of patients recurring within 2 years. While clinicopathologic criteria exist for adjuvant treatment, these criteria are imprecise and have not changed for decades. However, no reliable method exists to tailor adjuvant therapy for individual patients based on risk of recurrence after surgery. Using an ultra-sensitive targeted sequencing approach, we demonstrate that circulating tumor DNA (ctDNA) in lymphatic exudate collected via surgical drains (“lymph”) 24 hours after surgery accurately identifies MRD and outperforms plasma in an initial cohort of 36 HNSCC patients. We then applied the lymph ctDNA test to an independent, multi-site cohort of 37 HNSCC patients, replicating the original finding. Lymph performance was particularly enhanced in locoregional relapse in both cohorts and generalized to early stage (I-II) patients. Analysis of matched plasma collected at this early timepoint was not predictive of recurrence. We demonstrate a liquid biopsy approach using a historically overlooked biofluid to potentially enable precision adjuvant therapy and achieve superior oncologic outcomes.