RNAi Screening Of Human Kinome Identified Fibroblast Growth Factor Receptor 4 (FGFR4) As Potential Molecular Target In Basal-Like Breast Cancers (Blbc)

Abstract

Global gene expression profiling has uncovered previously unrecognised subsets of human breast cancer, including the so-called “triple-negative” or “basal-like” tumours (BLBC) characterised by oestrogen/progesterone receptor negativity, lack of HER2/Neu amplification and high frequency of p53 mutation. These refractory tumours therefore are insensitive to hormonal or Herceptin-based therapy, and thus confer a markedly poor prognosis relative to other subtypes. To date, little progress has been made to identify specific molecular pathways associated with these refractory cancers that may be effectively targeted for therapeutic purposes. In the present study, I have established a robust high-throughput RNAi screening assay. Through a human kinome RNAi library screen, 37 candidate genes which mediate the proliferation and survival of basal-like MDA-MB-468 breast cancer cells have been identified. Subsequent meta-analyses via Oncomine and public database, and validation by RNAi approach have established the significance of five candidate kinases – AURKB, FGFR4, PIK3CD, RIPK1 and SPHK1 in the proliferation and survival of breast cancers. In particular, knock-down of endogenous FGFR4 induces tumour-specific cell death in basal-like MDA-MB-468 and HCC1937 but not in luminal-like breast cancers (SKBR3, T47D and MCF7). The FGFR4-dependent survival of these BLBC cells is attributed to the constitutive activation of FGFR4 in these cancer cells, as marked by the presence of phosphorylated-FGFR4. Further analysis reveals that FGFR4 mediates the BLBC cells survival through activation of AKT and STAT3 as knock-down of endogenous FGFR4 in MDA-MB-468 and HCC1937 cells significantly reduced AKT and STAT3 phosphorylation but not the ERK1/2 phosphorylation. Ectopic expression of a constitutively active myristoylated AKT completely abrogates the apoptosis induced by FGFR4 knock-down suggesting that AKT is required for the pro-survival effects of FGFR4. Furthermore, we demonstrated that FGF19, a ligand specific for FGFR4, was secreted in the basal-like MDA-MB-468, MDA-MB-231 and HCC1937 but not in luminal-like breast cancers (SKBR3, T47D and MCF-7) and the non-transformed myoepithelial cells (MCF-10A and HMEC). Interestingly, depletion of FGF19 in MDA-MB-468 and HCC1937 also suppresses the tumour growth and reduced AKT and STAT3 phosphorylation. Together, our results demonstrated the existence of a FGFR4-FGF19 autocrine loop that mediates the survival of BLBC cells and could potentially be developed as a therapeutic target for future treatment of BLBC.