Background: A better understanding of the biological changes occurring during metastatic progression of breast cancer is needed to identify new biomarkers, targets and novel treatment strategies. Here, we compared the intrinsic subtype and the expression of a gene panel across a large dataset of paired primary and metastatic tissues.
Methods: Expression profiling of 105 breast cancer-related genes was performed on 254 (127 pairs) formalin-fixed paraffin-embedded tumor tissues using the nCounter platform. Tumor samples were obtained from 3 independent sources (ConvertHER trial [BCRT 2014] and two in-house datasets). Tumors were classified into each intrinsic subtype using the research-based PAM50 classifier (Parker et al. J Clin Oncol 2009). Chi-square tests were performed to determine the differences in the distribution of variables. Paired two-class Significance of Microarrays (SAM) was performed to determine the genes differentially expressed between paired primary and metastatic tissues. In vitro stable transfection of FGFR4-GFP was performed on Luminal B MCF7 cell line. RNA was purified on control vs. transfected cell lines. 7-AAD cell viability was performed following estrogen deprivation for 6 days.
Results: Subtype distribution in primary vs. metastatic disease was 39.0% vs. 26.8% for Luminal A (p=0.012), 26.0% vs. 35.0% for Luminal B (p=0.322), 11.4% vs. 20.3% for HER2-enriched (p=0.115) and 10.6% vs. 13.0% for Basal-like tumors (p=0.843). The rate of subtype conversion was 7.7% in Basal-like, 23.1% in HER2-enriched, 30.0% in Luminal B and 54.3% in Luminal A disease. The majority of subtype conversions in Luminal A disease were to Luminal B (72.0%) and HER2-enriched (24.0%). Overall, 13.2% of primary Luminal A/B tumors progressed to a HER2-E subtype despite 70% of them being clinically HER2-negative. In a paired analysis using all samples, 10- and 12- genes were found up- and down- regulated in metastatic tissues (False Discovery Rate [FDR] <5%). The up-regulated gene list in metastatic disease was composed of FGFR4 (top gene) and proliferation genes (CDC6, CCNB1, CEP55). The down-regulated gene list in metastatic disease was enriched for luminal-related genes (ESR1, PGR, NAT1 and MAPT). A similar paired analysis within Luminal A, Luminal B, HER2-enriched and Basal-like disease revealed 22, 8, 7 and 0 differentially expressed genes (FDR<5%), respectively. Finally, MCF7 cell line transfected with FGFR4 showed a relative increase in the HER2-enriched profile compared with transfected control. In vitro, MCF7-FGFR4 cells showed estrogen independent growth compared to transfected controls.
Conclusions: Metastatic tissues are relatively more proliferative and less luminal compared to primary tumors. This is especially relevant in primary Luminal A disease. In contrast, metastatic tissues from Basal-like primary disease remain largely unchanged. In luminal disease, a significant increase in the HER2-enriched profile is observed in metastatic disease despite most tumors being clinically HER2-negative. A potential driver of the HER2-enriched profile and estrogen independence in clinically HER2-negative metastatic tissues might be FGFR4.
Citation Format: Prat A, Martínez de Dueñas E, Galván P, Garcia S, Burgués O, Paré L, Antolín S, Martinello R, Blancas I, Adamo B, Guerrero Á, Muñoz M, Nuciforo P, Vidal M, Pérez RM, Chacón JI, Caballero R, Gascón P, Carrasco E, Rojo F, Perou CM, Cortés J, Adamo V, Albanell J, Lluch A. Intrinsic subtype and gene expression changes between primary and metastatic breast cancer. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P6-05-02.
