Increased expression of both FGF proteins and their receptors observed in many cancers is often associated with the development of chemoresistance, limiting the effectiveness of currently used anti-cancer therapies. Malfunctioning of the FGF/FGFR axis in cancer cells generates a number of molecular mechanisms that may affect the sensitivity of tumors to the applied drugs. Of key importance is the deregulation of cell signaling, which can lead to increased cell proliferation, survival, and motility, and ultimately to malignancy. Signaling pathways activated by FGFRs inhibit apoptosis, reducing the cytotoxic effect of some anti-cancer drugs. FGFRs-dependent signaling may also initiate angiogenesis and EMT, which facilitates metastasis and also correlates with drug resistance. Therefore, treatment strategies based on FGF/FGFR inhibition (using receptor inhibitors, ligand traps, monoclonal antibodies, or microRNAs) appear to be extremely promising. However, this approach may lead to further development of resistance through acquisition of specific mutations, metabolism switching, and molecular cross-talks. This review brings together information on the mechanisms underlying the involvement of the FGF/FGFR axis in the generation of drug resistance in cancer and highlights the need for further research to overcome this serious problem with novel therapeutic strategies.

Fibroblast growth factors (FGFs) via their receptors (FGFRs) transduce signals from the extracellular space to the cell interior, modulating pivotal cellular processes such as cell proliferation, motility, metabolism and death. FGF superfamily includes a group of fibroblast growth factor homologous factors (FHFs), proteins whose function is still largely unknown. Since FHFs lack the signal sequence for secretion and are unable to induce FGFR-dependent cell proliferation, these proteins were considered as intracellular proteins that are not involved in signal transduction via FGFRs. Here we demonstrate for the first time that FHF1 directly interacts with all four major FGFRs. FHF1 binding causes efficient FGFR activation and initiation of receptor-dependent signaling cascades. However, the biological effect of FHF1 differs from the one elicited by canonical FGFs, as extracellular FHF1 protects cells from apoptosis, but is unable to stimulate cell division. Our data define FHF1 as a FGFR ligand, emphasizing much greater similarity between FHFs and canonical FGFs than previously indicated. Graphical

Fibroblast growth factor receptor 1 (FGFR1) is a plasma membrane protein that transmits signals from the extracellular environment, regulating cell homeostasis and function. Dysregulation of FGFR1 leads to the development of human cancers and noncancerous diseases. Numerous tumors overproduce FGFR1, making this receptor a perspective target for cancer therapies. Antibody-drug conjugates (ADCs) are highly potent and selective anticancer agents. ADCs are composed of antibodies (targeting factors) fused to highly cytotoxic drugs (warheads). The efficiency of ADC strategy largely depends on the internalization of cytotoxic conjugate into cancer cells. Here, we have studied an interplay between affinity of anti-FGFR1 antibodies and efficiency of their cellular uptake. We have developed a unique set of engineered anti-FGFR1 antibodies that bind the same epitope in the extracellular part of FGFR1, but with different affinities. We have demonstrated that these antibodies are effectively taken up by cancer cells in the FGFR1-dependent manner. Interestingly, we have found that efficiency, defined as rate and level of antibody internalization, largely depends on the affinity of engineered antibodies towards FGFR1, as high affinity antibody displays fastest internalization kinetics. Our data may facilitate design of therapeutically relevant targeting molecules for selective treatment of FGFR1 overproducing cancers.

Background: Antibody drug conjugates (ADCs) represent one of the most promising approaches in the current immuno-oncology research. The precise delivery of cytotoxic drugs to the cancer cells using ADCs specific for tumor-associated antigens enables sparing the healthy cells and thereby reduces unwanted side effects. Overexpression of fibroblast growth factor receptor 1 (FGFR1) has been demonstrated in numerous tumors and thereby constitutes a convenient molecular target for selective cancer treatment. We have recently engineered tetravalent anti-FGFR1 antibody, T-Fc, and have demonstrated that it displays extremely efficient internalization into FGFR1 producing cells, a feature highly desirable in the ADC approach. We have revealed that T-Fc mediates clustering of FGFR1, largely enhancing the uptake of FGFR1-T-Fc complexes by induction of clathrin-independent endocytic routes. The aim of this study was to obtain highly internalizing cytotoxic conjugate of the T-Fc for specific delivery of drugs into FGFR1-positive cancer cells. Methods: Conjugation of the T-Fc to a cytotoxic payload, vcMMAE, was carried out via maleimide chemistry, yielding the T-Fc-vcMMAE. The specific binding of the T-Fc-vcMMAE conjugate to FGFR1 was confirmed in vitro with BLI technique. Confocal microscopy and flow cytometry were applied to determine FGFR1-dependence of the T-Fc-vcMMAE internalization. Western blot analyses of FGFR1-dependent signaling were conducted to assess the impact of the T-Fc-vcMMAE on FGFR1 activation and initiation of downstream signaling cascades. Finally, using FGFR1-negative and FGFR1-possitive cell lines, the cytotoxic potential of the T-Fc-vcMMAE was evaluated. Results: We have performed the efficient conjugation of the tetravalent engineered antibody with a cytotoxic drug and generated FGFR1-specific ADC molecule, T-Fc-vcMMAE. We have demonstrated that T-Fc-vcMMAE conjugate exhibits high selectivity and affinity for FGFR1, similarly to T-Fc. Furthermore, we have shown that T-Fc constitutes an effective drug delivery vehicle as T-Fc-vcMMAE was efficiently and selectively internalized by FGFR1-producing cells leading to their death. Interestingly, we show that the efficiency of the uptake of T-Fc-vcMMAE corresponds well with the cytotoxicity of the conjugate, but doesn’t correlate with the FGFR1expression level. Conclusion: Our results show that T-Fc-vcMMAE fulfills the key criteria for the successful cytotoxic drug carrier in a targeted approach against FGFR1-positive cancer cells. Furthermore, our data implicate that not solely expression level of the receptor, but rather its cellular trafficking should be taken into account for selection of suitable molecular targets and cancer models for successful ADC approach.

Breast cancer is a widespread and complex disease characterized by abnormal signaling pathways that promote tumor growth and progression. Despite significant medical advances and the development of increasingly effective therapies for breast cancer, drug resistance and reduced sensitivity to prior therapies remain persistent challenges. Dysregulation of growth factors such as FGFs and EGF and their receptors is a contributing factor to reduced response to treatment, promoting cell survival and proliferation, metastasis, EMT or increased expression of ABC transporters. Our study demonstrates a protective role for FGF1 in MCF-7 breast cancer cells against taltobulin-induced cytotoxicity, mediated by activation of its receptors and compares its activity to EGF, another growth factor involved in breast cancer development and progression. The mechanisms of action of these two proteins are different: FGF1 exerts its effects through the activation of both ERKs and AKT, whereas EGF acts only through ERKs. FGF1 action in the presence of the drug promotes cell viability, reduces apoptosis and increases cell migration. Although EGF and its receptors have received more attention in breast cancer research to date, our findings highlight the key role played by FGFs and their receptors in promoting drug resistance to tubulin polymerization inhibitors in FGFR-positive tumors.

Fibroblast growth factor 1 (FGF1) acts by activating specific tyrosine kinase receptors on the cell surface. In addition to this classical mode of action, FGF1 also exhibits intracellular activity. Recently, we found that FGF1 translocated into the cell interior exhibits anti-apoptotic activity independent of receptor activation and downstream signaling. Here, we show that expression of FGF1 increases the survival of cells treated with various apoptosis inducers, but only when wild-type p53 is present. The p53-negative cells were not protected by either ectopically expressed or translocated FGF1. We also confirmed the requirement of p53 for the anti-apoptotic intracellular activity of FGF1 by silencing p53, resulting in loss of the protective effect of FGF1. In contrast, in p53-negative cells, intracellular FGF1 regained its anti-apoptotic properties after transfection with wild-type p53. We also found that FGF1 directly interacts with p53 in cells and that the binding region is located in the DBD domain of p53. We therefore postulate that intracellular FGF1 protects cells from apoptosis by directly interacting with p53.