Targeting the heparin-binding domain of fibroblast growth factor receptor 1 as a potential cancer therapy

Abstract

BackgroundAberrant activation of fibroblast growth factor receptors (FGFRs) deregulates cell proliferation and promotes cell survival, and may predispose to tumorigenesis. Therefore, selective inactivation of FGFRs is an important strategy for cancer therapy. Here as a proof-of-concept study, we developed a FGFR1 neutralizing antisera, IMB-R1, employing a novel strategy aimed at preventing the access of essential heparan sulfate (HS) co-receptors to the heparin-binding domain on FGFR1.MethodsThe mRNA and protein expression level of FGFR1 and other FGFRs were examined in several lines of breast cancer and osteosarcoma cells and corresponding normal cells using Taqman real-time quantitative PCR and Western blot analysis. The specificity of IMB-R1 against FGFR1 was assessed with various ELISA-based approaches and Receptor Tyrosine Kinase array. Proliferation assay and apoptosis analysis were performed to assess the effect of IMB-R1 on cancer cell growth and apoptosis, respectively, in comparison with known FGFR1 inhibitors. The IMB-R1 induced alteration of intracellular signaling and gene expression were analysed using Western blot and microarray approaches. Immunohistochemical staining of FGFR1 using IMB-R1 were carried out in different cancer tissues from clinical patients. Throughout the study, statistical differences were determined by Student’s t test where appropriate and reported when a p value was less than 0.05.ResultsWe demonstrate that IMB-R1 is minimally cross-reactive for other FGFRs, and that it potently and specifically inhibits binding of heparin to FGFR1. Furthermore, IMB-R1 blocks the interaction of FGF2 with FGFR1, the kinase activity of FGFR1 and activation of intracellular FGFR signaling. Cancer cells treated with IMB-R1 displayed impaired FGF2 signaling, were unable to grow and instead underwent apoptosis. IMB-R1-induced cell death correlated with a disruption of antioxidative defense networks and increased expression of several tumor suppressors and apoptotic proteins, including p53. Immunostaining with IMB-R1 was stronger in human cancer tissues in which the FGFR1 gene is amplified.ConclusionOur study suggests that blocking HS interaction with the heparin-binding domains of FGFR1 inhibited cancer cell growth, which can be an attractive strategy to inactivate cancer-related heparin-binding proteins.Electronic supplementary materialThe online version of this article (doi:10.1186/s12943-015-0391-4) contains supplementary material, which is available to authorized users.

Affinity Selection of FGF2-Binding Heparan Sulfates for Ex Vivo Expansion of Human Mesenchymal Stem Cells

“…A and B). Using an FGFR1‐neutralizing antibody developed in our laboratory (IMB‐R1) (Ling et al, ) and the commercial FGFR kinase inhibitor SU5402, we show that these proliferative effects were inhibited by FGFR1 inactivation (Fig. A and B).…”

“…Human MSCs (3,000 cells/cm 2 ) were seeded in triplicate wells and allowed to adhere overnight. Cells were treated as indicated and viable cell numbers determined using the GUAVA Flow Cytometry Viacount Program (Merck Millipore, Billerica, MA) as previously described (Ling et al, ).…”

The anti‐proliferative effects of oleanolic acid on A7r5 cells—Role of UCP2 and downstream FGF‐2/p53/TSP‐1

“…Our results confirmed such effect, since pretreatment with UCP2 inhibitor genipin effectively inhibited FGF‐2 expression in ox‐LDL‐exposed A7r5 cells. Additionally, FGF‐2 is known to regulate multiple downstream signal molecules, including p53 (Ling et al, ). Consistently, in the current study, siRNA‐mediated FGF‐2 silencing resulted in increased expression levels of p53, while no remarkable changes were observed in the expression level of UCP2.…”

Bringing Heparan Sulfate Glycomics Together with Proteomics for the Design of Novel Therapeutics: A Historical Perspective

“…In an analogous manner to the proteomic/glycomic analysis we conducted above, Ling et al . developed an antibody, IMB‐R1, in an attempt to block the formation of an active FGF‐HS‐FGFR‐1 signaling complex.…”

“…FGFR‐1 is widely expressed, particularly in breast and bone, in both normal and cancerous tissues . Treatment with IMB‐R1 may alter cell surface FGFR‐1 levels by interfering with receptor endocytosis .…”