IADR Abstract Archives
Novel Mode of EGFR Inhibition to Overcome Acquired Resistance
Objectives: Epidermal growth factor receptor (EGFR) is often over-expressed or mutated in human tumors, including 50% of oral squamous cell carcinomas (OSCC), and is a valid target for therapy of oral cancers. Currently used direct inhibitors of EGFR are ineffective and patients eventually develop drug resistance to these agents. Activation of other receptor tyrosine kinases (RTKs) such as insulin-like growth factor receptor (IGFR) is a primary mechanism that contributes to the insensitivity to EGFR inhibitors. In a previous study, using immortalized cell lines CHO and CaCO-2 over-expressing EGFR, we showed that the anti-anginal agent fendiline inhibits EGFR by mislocalizing it from the plasma membrane (PM). Goals of this study were to validate EGFR mislocalization in OSCC cell lines and to determine whether fendiline alters cross-activation of IGFR by EGFR.
Methods: Confocal microscopy analysis of EGFR over-expressing OSCC cell line SCC25 stimulated with FITC-tagged EGF and colocalization studies using trafficking compartment markers were conducted to analyze mislocalization of EGFR in OSCC cell lines. Co-immunoprecipitation assays using lysates from EGFR over-expressing OSCC cell line HN5 were done to evaluate IGFR interaction with EGFR. Western blot analysis was utilized to compare the effects of EGFR inhibitors erlotinib and fendiline on cross-activation of IGFR when stimulated with EGF.
Results: Our results show the following important findings: Fendiline causes mislocalization of the EGFR from the PM primarily to the early endosomes with some being localized to late and recycling endosome. Co-immunoprecipitation assays revealed no significant changes in IGFR interaction with EGFR in fendiline-treated cells compared to control. However, cross-activation of IGFR with EGF stimulation was reduced in fendiline-treated cells, evident by reduced pERK levels.
Conclusions: Fendiline, therefore, not only inhibits EGFR-mediated signaling but also inhibits cross-activation of IGFR by EGFR. Fendiline could serve as a novel mode of treatment for OSCC with less likelihood of patients developing acquired resistance.
Methods: Confocal microscopy analysis of EGFR over-expressing OSCC cell line SCC25 stimulated with FITC-tagged EGF and colocalization studies using trafficking compartment markers were conducted to analyze mislocalization of EGFR in OSCC cell lines. Co-immunoprecipitation assays using lysates from EGFR over-expressing OSCC cell line HN5 were done to evaluate IGFR interaction with EGFR. Western blot analysis was utilized to compare the effects of EGFR inhibitors erlotinib and fendiline on cross-activation of IGFR when stimulated with EGF.
Results: Our results show the following important findings: Fendiline causes mislocalization of the EGFR from the PM primarily to the early endosomes with some being localized to late and recycling endosome. Co-immunoprecipitation assays revealed no significant changes in IGFR interaction with EGFR in fendiline-treated cells compared to control. However, cross-activation of IGFR with EGF stimulation was reduced in fendiline-treated cells, evident by reduced pERK levels.
Conclusions: Fendiline, therefore, not only inhibits EGFR-mediated signaling but also inhibits cross-activation of IGFR by EGFR. Fendiline could serve as a novel mode of treatment for OSCC with less likelihood of patients developing acquired resistance.