Peri‐Operative Detection of Circulating Tumor <scp>DNA</scp> Using a Customized <scp>NGS</scp> Tumor‐Informed Assay in Patients With <scp>HPV</scp> ‐Negative Locally Advanced Head and Neck Squamous Cell Carcinoma

ctDNA is a promising tool to optimize therapeutic strategies in patients treated for locally advanced head and neck cancer that is negative for Human Papilloma Virus (LAHNSCC/HPV-). The objective was to evaluate pre and postoperative detection of circulating tumor DNA (ctDNA) using a customized Next-Generation Sequencing (NGS) tumor-informed assay. We conducted a prospective study including patients treated with initial surgery for a LAHNSCC/HPV-. We developed an NGS assay covering the exons of 11 genes for tumor and ctDNA analysis. DNA alterations were interpreted according to two classes of variants: "pathogenic variants" classified as class 4 or 5 in public databases and "tumor specific variants" corresponding to alterations detected exclusively in tumor DNA after paired analysis of normal cell DNA using the same NGS panel. A total of 40 patients were included, 45% with T4 and 62.5% with N+ disease. Regarding tumor profiles, 41 "pathogenic variants" were detected corresponding to 30/36 (83.3%) of informative tumors with at least one alteration. In contrast, 354 "tumor specific variants" were identified with at least one alteration in 35/35 (100%) informative tumors. For ctDNA, the NGS panel was associated with a positivity rate of 74.3% and 37.1% pre and postoperative, respectively, using tumor specific variants; compared with 20% and 2.5% using interpretation of pathogenic variant. Pretreatment ctDNA positivity according to pathogenic variant was associated with lower specific progression-free survival (p = 0.025). Our customized 11-genes NGS panel tumor-informed assay provides high rates of ctDNA detection and may be used to evaluate new strategies in LA HNSCC/HPV- patients.