Personalized circulating tumor DNA analysis to detect residual disease after neoadjuvant therapy in breast cancer

A robust personalized ctDNA test, TARDIS, achieves high accuracy for residual disease after completion of neoadjuvant therapy. Early detection, no time travel needed Analysis of tumor DNA shed into a patient’s circulation can provide a noninvasive means of detecting the presence of a tumor and analyzing its DNA for targetable mutations. Unfortunately, it can be difficult to detect small amounts of tumor DNA in the blood, especially in patients who have already undergone initial chemotherapy treatment. To address this problem, McDonald et al. developed a method of targeted digital sequencing (TARDIS), which is customized for each patient but can then be used to monitor the patient over time for and allow early detection of tumor recurrence. Longitudinal analysis of circulating tumor DNA (ctDNA) has shown promise for monitoring treatment response. However, most current methods lack adequate sensitivity for residual disease detection during or after completion of treatment in patients with nonmetastatic cancer. To address this gap and to improve sensitivity for minute quantities of residual tumor DNA in plasma, we have developed targeted digital sequencing (TARDIS) for multiplexed analysis of patient-specific cancer mutations. In reference samples, by simultaneously analyzing 8 to 16 known mutations, TARDIS achieved 91 and 53% sensitivity at mutant allele fractions (AFs) of 3 in 104 and 3 in 105, respectively, with 96% specificity, using input DNA equivalent to a single tube of blood. We successfully analyzed up to 115 mutations per patient in 80 plasma samples from 33 women with stage I to III breast cancer. Before treatment, TARDIS detected ctDNA in all patients with 0.11% median AF. After completion of neoadjuvant therapy, ctDNA concentrations were lower in patients who achieved pathological complete response (pathCR) compared to patients with residual disease (median AFs, 0.003 and 0.017%, respectively, P = 0.0057, AUC = 0.83). In addition, patients with pathCR showed a larger decrease in ctDNA concentrations during neoadjuvant therapy. These results demonstrate high accuracy for assessment of molecular response and residual disease during neoadjuvant therapy using ctDNA analysis. TARDIS has achieved up to 100-fold improvement beyond the current limit of ctDNA detection using clinically relevant blood volumes, demonstrating that personalized ctDNA tracking could enable individualized clinical management of patients with cancer treated with curative intent.