Abstract P2-02-19: Circulating tumor DNA detection anticipates disease recurrence in early stage breast cancer: A pilot study generating an observational confirmatory trial
Authors: Daidone MG , Di Cosimo S , Veneroni S , Cascone F , De Cecco L , Dugo M , Folli S , Bianchi GV , Tamborini E , Busico A , Appierto V 
Published: February 2018

Sensitive tumor biomarkers able to monitor disease progression would contribute to post-surgical treatment decision making in early breast cancer. We investigated the feasibility of using circulating tumor DNA (ctDNA) to early detect new disease manifestations in serial plasma samples collected during post-surgery follow-up from patients operated for stage I breast cancer from 1992 to 1993 at Istituto Nazionale Tumori in Milan. Forty patients that underwent radical or conservative surgery for T1/T2-N0-M0 breast cancer and that were followed for at least 15 years were included in a pilot study for the retrospective analysis of ctDNA on at least 3 plasma samples obtained during follow-up. To assess the feasibility of ctDNA analysis in archival plasma samples collected in heparin and stored from 10 to 25 years, preliminary experiments demonstrated that ctDNA was not affected by: 1) heparinase I digestion of extracted DNA and 2) DNA pre-amplification step to overcome limitations due to small plasma aliquots. Mutational analysis of breast cancer tissues was performed by Ion Torrent-targeted next generation sequencing and the identified Single Nucleotide Variations (SNV) were first validated and then tracked in plasma samples by using ad hoc digital polymerase chain reaction assays. One or more SNVs were identified in tumor tissue specimens and validated in 27/40 cases. Among those 27 breast cancers, 6 cases relapsed locally, 4 in distant sites, and 17 remained disease-free for the entire follow-up. ctDNA was undetectable during the post-surgical follow-up in 16/17 disease-free women up to 160 months of surgery, while it was detectable in 9/10 patients developing unfavorable events and anticipated the clinical diagnosis of relapse in 7/10 patients with a median lead time of 20 months. Our results are the first to associate mutation tracking to local recurrence and indicate that in patients with early breast cancer ctDNA monitoring during post-operative follow-up can anticipate the diagnosis of new disease manifestations, thus potentially allowing prompt treatments. These findings establish the rational to plan prospective studies to evaluate in the early breast cancer context the potential of ctDNA as a non-invasive and sensitive biomarker for monitoring tumor progression. Based on these results, we activated in 2016 a prospective observational study to confirm the predictive value of ctDNA on local and distant relapse in patients with early and localized triple negative breast cancer. As for May 2017, 145 patients with triple negative tumors were potentially enrolled for a ctDNA-based post-surgical follow-up: 111 cases at first diagnosis and 34 cases at surgery after neo-adjuvant treatment. One hundred-ten women accepted to participate in the study and signed a specific informed consent, whereas 22 patients refused to participate and 13 were lost to follow-up. For 94 patients (66 at initial diagnosis and 28 after neoadjuvant chemotherapy) plasma samples have been already longitudinally collected and DNA sequencing is currently in progress.

Citation Format: Daidone MG, Di Cosimo S, Veneroni S, Cascone F, De Cecco L, Dugo M, Folli S, Bianchi GV, Tamborini E, Busico A, Appierto V. Circulating tumor DNA detection anticipates disease recurrence in early stage breast cancer: A pilot study generating an observational confirmatory trial [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr P2-02-19.