FGFR4 is a regulator of tumor subtype differentiation in luminal breast cancer and metastatic disease

Mechanisms driving tumor progression from less aggressive subtypes to more aggressive states represent key targets for breast cancer therapy. We identified a subset of Luminal A primary breast tumors to give rise to HER2-enriched (HER2E) subtype metastases, but remain clinically HER2 negative (cHER2-). By testing the unique genetic and transcriptomic features of these cases, we developed the hypothesis that fibroblast growth factor receptor 4 (FGFR4) drives this subtype switching. To evaluate this, we developed two FGFR4 genomic signatures using a PDX model treated with a FGFR4 inhibitor (BLU9931), which inhibited PDX growth in vivo. Examining patient outcomes in the METABRIC breast cancer cohort showed that the FGFR4-induced and FGFR4-repressed signatures each predicted overall survival (OS) (HR=6.30, P<0.0001; HR=0.33; P<0.0001, respectively). Multivariate analysis showed that the FGFR4-induced signature was also an independent prognostic factor beyond subtype and stage for OS (HR=2.34, P=0.014). Supervised analysis of 77 primary tumors with paired metastasis revealed that the FGFR4-induced signature was significantly higher in luminal/ER+ tumor metastases compared with their primaries. Finally, multivariate analysis demonstrated that the FGFR4-induced signature also predicted site-specific metastasis for lung, liver and brain, but not for bone or lymph nodes. These data identify a link between FGFR4-regulated genes and metastasis, suggesting a treatment options for FGFR4-positive patients, whose high expression is non-genetically determined. Single-Cell RNA-seq profiling WHIM11 PDX (patient-derived xenograft) tumor under three different experimental conditions. For therapeutic testing, tumors were allowed to progress to a diameter of 0.5-0.7 cm and tumor-bearing mice were assigned at random to one of three groups: untreated or control (WHIM11_SG334, WHIM11_SG362), BLU9931 treatment (WHIM11_SG334, WHIM11_SG362) or BLU9931 treatment and drug released group (WHIM11_SG329, WHIM11_SG363). BLU9931 was administered at 0.6g of drug/Kg/day. The untreated group were sacrificed if tumors reached 1.5 mm in any direction and the tumors were collected and processed for single cell RNA sequencing (scRNAseq). Once tumors were established to be growing in the presence of BLU9931 (18 days) the tumors were also collected for scRNAseq. Finally, the third group of BLU9931 treatment was ceased for 14 days and processed in the same way for scRNAseq.