Colorectal cancer (CRC) is the third world leading cause of cancer death. Cetuximab and panitumumab, two monoclonal antibodies that bind the extracellular domain of epidermal growth factor receptor (EGFR), are effective only in the subset of RAS/BRAF wild-type metastatic colorectal cancers (mCRC). However, not all the RAS/BRAF wild-type patients benefit from anti-EGFR treatments. We hypothesized that primary resistance might be driven by overexpression of different tyrosine kinase (TK) genes and consequent activation of parallel pathways.

Starting from a cetuximab screening of a panel of 151 CRC cell lines, we identified 41/151 CRC RAS/BRAF wild-type cells for which the mechanism of primary resistance to cetuximab is unaccounted for. Then, transcriptional outlier analysis identifies a subset of RAS/BRAF wild type cells, resistant to EGFR blockade, functionally and pharmacologically addicted to kinase genes including ALK, FGFR2, KIT, NTRK1/2 and RET, all of them target of drugs undergoing clinical testing or approved. Outlier expression was confirmed by RNAseq analysis. For ALK, NTRK1 and FGFR2 we found that overexpression is associated to molecular alterations, such as gene translocation (ALK and NTRK1) or gene amplification (FGFR2), described also in cancer patients. An immunohistochemistry (IHC) based screen on 220 CRC FFPE samples identified one NTRK1 outlier positive sample, with genetic rearrangement (detected by FISH).

In order to demonstrate that outlier kinase genes were functionally relevant in the corresponding cell models and cell-specific functional dependencies, we used two complementary approaches: reverse genetics and pharmacological inhibition. siRNA-mediated candidate-specific gene suppression reduced, in all cases, protein expression of the ‘outlier’ TK resulted in significant impairment of cell growth, which was often accompanied by downstream signaling inhibition and apoptosis. Pharmacological inhibition with specific kinase-targeted agents was cell specific and paralleled the expression profiles of individual TKs. Nonetheless, in KIT overexpressing cells the reverse genetic experiment revealed functional dependency, despite relative kinase inhibitor was not effective.

In conclusion our data suggest that overexpression of TK outliers drives primary resistance to EGFR blockade, and could be used to identify patients unlikely to respond to cetuximab or panitumumab. Moreover, the approach described here can be used to pinpoint colorectal cancers with exquisite dependencies to individual kinases for which clinically approved drugs are already available.

Citation Format: Mariangela Russo, Gabriele Picco, Carlotta Cancelliere, Giorgio Corti, Emanuele Valtorta, Silvio Veronese, Marco Beccuti, Francesca Cordero, Federica Di Nicolantonio, Enzo Medico, Alberto Bardelli. The molecular landscape of colorectal cancer cell lines unveils clinically actionable targets. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 916. doi:10.1158/1538-7445.AM2015-916