A Molecularly Annotated Model of Patient-Derived Colon Cancer Stem–Like Cells to Assess Genetic and Nongenetic Mechanisms of Resistance to Anti-EGFR Therapy

Abstract

<b>Purpose:</b> Patient-derived xenografts ("<i>xenopatients</i>") of colorectal cancer metastases have been essential to identify genetic determinants of resistance to the anti-EGFR antibody cetuximab and to explore new therapeutic strategies. From xenopatients, a genetically annotated collection of stem-like cultures ("<i>xenospheres</i>") was generated and characterized for response to targeted therapies.<b>Experimental Design:</b> Xenospheres underwent exome-sequencing analysis, gene expression profile, and <i>in vitro</i> targeted treatments to assess genetic, biological, and pharmacologic correspondence with xenopatients, and to investigate nongenetic biomarkers of therapeutic resistance. The outcome of EGFR family inhibition was tested in an <i>NRG1</i>-expressing <i>in vivo</i> model.<b>Results:</b> Xenospheres faithfully retained the genetic make-up of their matched xenopatients over <i>in vitro</i> and <i>in vivo</i> passages. Frequent and rare genetic lesions triggering primary resistance to cetuximab through constitutive activation of the RAS signaling pathway were conserved, as well as the vulnerability to their respective targeted treatments. Xenospheres lacking such alterations (RAS^wt) were highly sensitive to cetuximab, but were protected by ligands activating the EGFR family, mostly NRG1. Upon reconstitution of <i>NRG1</i> expression, xenospheres displayed increased tumorigenic potential <i>in vivo</i> and generated tumors completely resistant to cetuximab, and sensitive only to comprehensive EGFR family inhibition.<b>Conclusions:</b> Xenospheres are a reliable model to identify both genetic and nongenetic mechanisms of response and resistance to targeted therapies in colorectal cancer. In the absence of RAS pathway mutations, NRG1 and other EGFR ligands can play a major role in conferring primary cetuximab resistance, indicating that comprehensive inhibition of the EGFR family is required to achieve a significant therapeutic response. <i>Clin Cancer Res; 24(4); 807-20. ©2017 AACR</i><i>See related commentary by Napolitano and Ciardiello, p. 727</i>.