Abstract P4-01-09: Combined analysis of tissue and blood biopsies by NGS in patients with advanced breast cancer identifies targetable molecular alterations
Authors: Salvador JF , Pinto JA , Araujo JM , Tirado-Hurtado I , Flores CJ , Chirinos LA , Requena MC , Carpio S , Finzel Pérez A , Aguilar A , Demol F , Schwarz LJ 
Published: May 2019

Abstract
Background: Targeted-next generation sequencing (t-NGS) analysis in tissue and peripheral blood is increasingly performed in patients with advanced cancer. Liquid biopsy is a non-invasive method that allows to understand the molecular changes occurring in the tumor in real-time, derived from intratumor heterogeneity and/or therapeutic pressure. In the present work we sought to elucidate the ability of liquid biopsy to detect mutations related with acquired resistance.
Methods: 190 formalin-fixed paraffin-embedded (FFPE) samples from primary and metastatic tumors of patients with advanced breast cancer (ABC) were analyzed by OncoDEEP® (1000X depth of coverage of 75 gene alterations including point mutations, insertions/deletions, gene fusions and copy number variations), and comparison between targeted-NGS results from matched primary/metastatic tumors (FFPE) and cell-free DNA (cfDNA) recovery from plasma analyzed by OncoSTRAT&GO® in 34 patients with ABC (193 genes with 1000x and 40 genes with 10000x depth coverage for FFPE and plasma, respectively).
Results: Of the 190 FFPEs analyzed by OncoDEEP®, 41% were triple negative breast cancer (TNBC), 49% hormone receptor positive (HR+) and 11% were HER2+. Mutations in TP53 (44.2%), PIK3CA (41.1%) and ERBB2 (9.47%) were the most frequent. As expected, PIK3CA and TP53 mutations were significantly higher in HR+ and TNBC tumors, respectively (PIK3CA: 53.8% for HR+ vs 28.6% and 30% for TNBC and HER2+, respectively; TP53 68.8% for TNBC vs 26.9 and 30% in HR+ and HER2+, respectively; in both cases p&lt;0.001). Interestingly, the alteration c.1459-7C&gt;T in JAK1 (unknown significance) was detected in 8% of patients (9.7% and 6.5% in HR+ and TNBC, respectively). Also, in contrast to other reports, high frequencies of APC alterations were found exclusively in HR+ tumors (previously reported as germline mutations). From the OncoSTRAT&GO® analysis (tumor tissue and cfDNA), ESR1 and FGFR mutations in HR+ ABC were significantly more present in cfDNA (9/20 mutations found [45%]] compared to tumor tissue analysis (18/93 mutations found [19.3%]) (p=0.02), and in three patients, mutations in ESR1 and FGRF were only detected in cfDNA. Finally, 11/34 cases (32.3%) showed gene mutations only in cfDNA with predominant alterations in MAPK signaling pathway (54.5%), TP53 (36.4%) and ESR1 (18.2%), the latest as previously described.
Conclusions: JAK1 and APC mutations need further evaluation to establish their clinical significance in ABC. The combination of tumor tissue and cfDNA t-NGS analyses into clinical routine enables detailed and comprehensive evaluation of tumor heterogeneity under therapeutic pressure; in our cohort, known mutations for acquired resistance to endocrine therapy (ESR1 and FGFR2) were better detected by cfDNA compared to tumor tissue (45% vs 19.3%) in HR+ ABC patients, increasing the probability to identify early drug resistance to prioritize patients' selection into clinical trials with targeted agents.
Citation Format: Salvador JF, Pinto JA, Araujo JM, Tirado-Hurtado I, Flores CJ, Chirinos LA, Requena MC, Carpio S, Finzel Pérez A, Aguilar A, Demol F, Schwarz LJ. Combined analysis of tissue and blood biopsies by NGS in patients with advanced breast cancer identifies targetable molecular alterations [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P4-01-09.