Development of a Novel NGS Methodology for Ultrasensitive Circulating Tumor DNA Detection as a Tool for Early-Stage Breast Cancer Diagnosis
Begoña Jiménez-Rodríguez, Alfonso Alba-Bernal, Esperanza López-López, María Elena Quirós-Ortega, Guillermo Carbajosa, Alicia Garrido-Aranda, Martina Álvarez, Ana Godoy-Ortíz, María Isabel Queipo‐Ortuño, Luís Vicioso, Gema Díaz-Córdoba, María Dunia Roldán-Díaz, Jesús Velasco-Suelto, Cristina Hernándo, Begoña Bermejo, Ana Julve, Aňa Lluch, Javier Pascual, Iñaki Comino‐Méndez, Emilio Alba
Abstract
Breast cancer (BC) is the most prevalent cancer in women. While usually detected when localized, invasive procedures are still required for diagnosis. Herein, we developed a novel ultrasensitive pipeline to detect circulating tumor DNA (ctDNA) in a series of 75 plasma samples from localized BC patients prior to any medical intervention. We first performed a tumor-informed analysis to correlate the mutations found in tumor tissue and plasma. Disregarding the tumor data next, we developed an approach to detect tumor mutations in plasma. We observed a mutation concordance between the tumor and plasma of 29.50% with a sensitivity down to 0.03% in mutant variant allele frequency (VAF). We detected mutations in 33.78% of the samples, identifying eight patients with plasma-only mutations. Altogether, we determined a specificity of 86.36% and a positive predictive value of 88.46% for BC detection. We demonstrated an association between higher ctDNA median VAF and higher tumor grade, multiple plasma mutations with a likelihood of relapse and more frequent TP53 plasma mutations in hormone receptor-negative tumors. Overall, we have developed a unique ultra-sensitive sequencing workflow with a technology not previously employed in early BC, paving the way for its application in BC screening.