Abstract B7: Use of ponatinib to restrict growth of non-small cell lung cancer (NSCLC) cells overexpressing FGFR1

Abstract

Abstract For advanced stage NSCLC, targeted molecular therapy based on an understanding of the oncogenic driver mutations is emerging as the new front line care replacing chemotherapy where the oncogenic driver mutation is clearly defined e.g EGFR mutation or EML4-ALK translocation. However, the proportion of patients that fall into this category is small and are mostly non-smokers with adeno histology. Identifying new oncogenic drivers in NSCLC is therefore paramount. In our initial Oncomine database search, we found that FGFRs are overexpressed in various NSCLC tissue specimens. Quantitative real time PCR and Western blot analysis subsequently confirmed overexpression of FGFR1 in about one fourth of NSCLC cell lines analyzed. Among these, H520, H1299 and A549 had significantly higher levels of FGFR1 expression compared to H3122 which carries an EML4-ALK fusion protein. Real time PCR also demonstrated FGFR1 overexpression compared to normal human bronchial epithelial cells transformed by Herpes Papillary Virus E6/E7 (HBE4). Furthermore, as a sign of epithelial to mesenchymal transition, these cells switched from expressing the FGFR1 3b to the 3c form. In a colonogenic assay the growth of cells carrying FGFR1 overexpression/activation is markedly suppressed by ponatinib, a pan FGFR inhibitor as expected. Similarly, ponatinib treatment of NSCLC cell lines using 1 μM concentrations effectively suppressed the phosphorylation level of FGFR1 and its known down-stream targets including pSrc and pPLCγ without significant impact on the total protein expression. Treatment of these cells harboring FGFR1 activation with ponatinib arrests cells at the G1 phase as shown in the flow cytometry analysis. To determine whether the observed effects are specifically due to the suppression of FGFR1 by ponatinib, an shRNA knock down of FGFR1 was performed. Consistently, shFGFR1 significantly inhibited the growth of H520 and H1299 cells, whereas knock-down did not significantly affect the growth of H3122. Similar to ponatinib treatment, shFGFR1 caused more cells to arrest in G1 phase. In a MTT assay, ponatinib synergistically induced growth suppression/killing of H520 cells with cisplatin and paclitaxel. We are currently determining whether ponatinib is effective in inhibiting NSCLC cells derived directly from freshly resected lung cancer specimens. In conclusion, the FGFR1 pathway is constitutively active in a proportion of NSCLC cell lines and ponatinib is a potent inhibitor of constitutive FGFR1 activation. Ponatinib is potentially useful as a molecularly targeted agent against this portion of NSCLC when used alone or in combination with traditional chemotherapy regimens.