Project description:Background: Up to 40% of patients with estrogen receptor positive (ER+) breast cancer experience treatment resistance and disease recurrence, often caused by the upregulation of growth factor receptors. Understanding the mechanisms of resistance, and the identification of novel therapeutic targets is, therefore, of vital importance if breast cancer prognosis is to be further improved. Fibroblast growth factor receptor 1 (FGFR1) is a potential driver of endocrine resistance; however, clinically successful attempts at targeting FGFR1 activity in breast cancer are rare and may result from an inability to correctly identify patients that could benefit from such treatment. The identification of additional genes associated with an FGFR1-mediated mechanism of resistance will provide a more precise classification scheme for FGFR1-dependency in breast cancer. Methods: Live cell imaging and RNA sequencing of ER+ breast cancer cell lines (CAMA1, T47D, tamoxifen resistant T47D) were used to investigate FGFR1-dependency in breast cancer, mechanisms of endocrine resistance and the effects of treatment with tamoxifen and erdafitinib. An FGFR1-amplified ER+ breast cancer patient-derived xenograft (PDX) model was used to assess the effects of targeting FGFR1 in vivo. Gene expression analysis of human breast cancer from The Cancer Genome Atlas (TCGA) was used for clinical validation. Results: We evaluated the effects of targeting FGFR1 in ER+/HER2- breast cancer with aberrant FGFR1 amplification and/or overexpression. We found that the FGFR tyrosine kinase inhibitor (TKI) erdafitinib inhibited cell proliferation of FGFR1-amplified CAMA1 cells in vitro. Additionally, erdafitinib significantly enhanced the anti-proliferative effect of 4-hydroxytamoxifen (4-OHT) and its anti-tumor effect was also demonstrated in vivo. Further, we demonstrated that the proliferation of FGFR1-overexpressing tamoxifen-resistant T47D (TR-T47D) cells is dependent on FGFR1 activity. Moreover, these TR-T47D cells are re-sensitized to tamoxifen upon treatment with erdafitinib. Finally, we found that upregulation of genes related to epithelial-mesenchymal-transition (EMT) correlates with FGFR1 overexpression, and is reversed by FGFR inhibition. Conclusions: In this study we demonstrated that targeting FGFR1 in ER+/HER2- breast cancer with aberrant FGFR1 activity has the potential to inhibit tumor growth and to re-sensitize resistant tumors to tamoxifen. Furthermore, we identified a functional relationship between EMT, FGFR activity and endocrine resistance in breast tumorigenesis.

Project description:Introduction: The clinical benefit of EGFR tyrosine kinase inhibitor (TKI) treatment in non-small cell lung cancer (NSCLC) patients with activating EGFR mutations is temporary, as virtually all patients develop acquired EGFR TKI resistance that occurs via diverse mechanisms. Here, we identified increased FGFR1 expression as such a resistance mechanism and using pathways analysis and drug combination testing we identified a novel combination treatment to control growth of these resistant tumors. Methods: Novel erlotinib-resistant NSCLC cell lines were generated and analyzed by mass spectrometry-based proteomics to identify altered pathways associated with erlotinib resistance. The altered pathways were further analyzed in gefitinib and osibenib resistant cell lines. Small molecule inhibitor combinations were used to block the altered pathways and investigate growth reduction in vitro and in two xenograft mouse models. FGFR1 mRNA levels were examined in pre- and (post?)- EGFR TKI treatment clinical tumor samples. Results: Proteomic analysis revealed increased expression of FGFR1 and AXL as well as increased Akt and ERK1/2 activation in a panel of novel erlotinib-resistant HCC827 cell lines. Combined treatment with erlotinib or osimertinib and a panel of small molecule inhibitors targeting AXL/MET, FGFRs, Akt, PI3K/mTOR, MEK or ERK1/2 showed that the most prominent re-sensitization to EGFR TKI occurred with the pan-FGFR inhibitor, PD173074. Interestingly, simultaneous blockade of components of the Akt pathway using specific Akt or dual PI3K-mTOR inhibitors combined with inhibitors targeting the FGFR family exhibited the most efficient growth inhibition of FGFR1 overexpressing EGFR TKI-resistant cell lines. Phosphorylation of proteins downstream of Akt, including PRAS40, FOXO and S6 ribosomal protein, were completely abrogated by PD173074 combined with the Akt inhibitor GSK2141795 . Combination treatment with PD173074 and an Akt inhibitor exhibited synergistic growth inhibition in vivo in two FGFR1 overexpressing NSCLC EGFR TKI-resistant animal models. Conclusion: The significant growth inhibition in vitro and in vivo observed with PD173074 combined with Akt compared to either drug alone imply that inhibition of several key targets may be beneficial in controlling erlotinib-resistant NSCLC. The complete abrogation of PRAS40, FOXO and S6 phosphorylations by PD173074 combined with an Akt inhibitor indicates that the Akt pathway is no longer active.

Project description:Epithelial/mesenchymal transition (EMT) is associated with loss of cell adhesion molecules, such as E-cadherin, and increased invasion, migration, and proliferation in epithelial cancers. In non-small cell lung cancer (NSCLC), EMT is associated with greater resistance to EGFR inhibitors. However, its potential to predict response to other targeted drugs or chemotherapy has not been well characterized. The goal of this study was to develop a robust, platform-independent EMT gene expression signature and to investigate the association of EMT and drug response in NSCLC. A 76-gene EMT signature was derived in 54 DNA-fingerprinted NSCLC cell lines and tested in an independent set of cell lines and in NSCLC patients from the BATTLE clinical trial. The signature classified cell lines as epithelial or mesenchymal independent of the microarray platform and correlated strongly with E-cadherin protein levels, as measured by reverse phase protein array. Higher protein expression of Rab25 (in epithelial lines) and Axl (in mesenchymal lines), two signature genes associated with in EMT in other cancer types, was also confirmed. Mesenchymal cell lines demonstrated significantly greater resistance to EGFR inhibition, independent of EGFR mutation status and were more resistant to drugs targeting the PI3K/Akt pathway. We observed no association between EMT and response to cytotoxic chemotherapies, including cisplatin, pemetrexed, and docetaxel monotherapy and/or doublets (p-values ≥0.2). In NSCLC patients, the EMT signature predicted 8-week disease control in the erlotinib arm, but not in other treatment arms. In conclusion, we have developed a robust EMT signature that predicts resistance to EGFR inhibitors and PI3K/Akt pathway inhibitors. Gene expression profiles were measured in 131 core biopsies from patients with refractory non-small cell lung cancer in the Biomarker-integrated Approaches of Targeted Therapy for Lung Cancer Elimination (BATTLE) trial. We used the BATTLE dataset to test an EMT gene expression signature trained in cell lines and independant of the microarray platform.

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Targeting FGFR overcomes EMT-mediated resistance in EGFR mutant non-small cell lung cancer

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