Abstract 4339: Molecular subtypes and novel genetic mechanisms of primary and acquired anti-EGFR resistance in colorectal cancer in the Prospect C biomarker trial
Authors: Khan Khurum Hayat , Woolston Andrew , Spain Georgia , Barber Louise , Patil Yatish , Griffiths Beatrice , GonzalezExposito Reyes , Mansukhani Sonia , Davies Matthew , Rao Sheela , Watkins David , Sclafani Francesco , Suntharanathan Jana , Peckitt Clare , Begum Ruwaida , Rana Isma , Thomas Janet , Oates Jacqui , Bryant Annette , Wotherspoon Andrew , Fotiadis Nicos , Khan Nasir , Guettler Sebastian , Loga Katharina von , Starling Naureen , Chau Ian , Sadanandam Anguraj , Cunningham David , Gerlinger Marco 
Published: August 2018

Abstract
Despite stratification of only RAS wt CRCs for anti-EGFR antibody (aEa) therapy, many patients (pts) do not benefit and the molecular resistance (res) landscape remains incompletely understood. In order to decipher novel res mechanisms, we treated 40 RAS wt CRCs with single-agent aEa in a prospective trial and applied exome- and RNA-Seq to biopsies (bx) taken at baseline (BL) and at progression (PD). Among 20 BL bx from tumors with primary progression, 7 showed BRAF V600E mutations (mut), 3 harbored noncanonical BRAF D594K and/or KRAS L19F/A18D mut or KRAS amplifications (amp), 1 had a MEK1 mut and 1 was ERBB2 amp. Inactivating mut of the NF1 gene, a negative regulator of RAS, were found as novel mechanisms of primary res in 2 pts. No genetic drivers of primary res could be identified in 6/20 bx. Using transcriptomic analysis, 78% of pts with prolonged clinical benefit (≥16wks) displayed the Transit-Amplifying (TA) molecular subtype (Sadanandam et al., 2013). Stem-like (SL), goblet and inflammatory subtypes predominated (75%) among primary progressors. This significant enrichment in aEa sensitive CRCs (p=0.017) validates the TA subtype as a predictive biomarker. Exome- and deep Seq of PD bx from 13 pts who acquired res after prolonged clinical benefit detected KRAS mut or amp in only 2 cases. The FGFR2 ligand FGF10 was amplified in 1 PD bx and was validated in vitro as a novel mechanism of acquired res. No mut/amp of genes to which aEa res is usually attributed (RAS/RAF, EGFR, MET, ERBB2, MEK1) were found in the remaining 10 PD bx. Ultra-sensitive circulating tumor DNA-Seq at PD in 7 of these pts detected EGFR exodomain and RAS mut, including parallel evolution of multiple mut, in 3 cases. However, comparison to truncal TP53/APC mut showed that these res drivers were confined to small subclones and could therefore not explain the bulk of res. Overall, aberrations of RAS/RAF-pathway members and regulators were less abundant as drivers of acquired aEa res in this prospective trial than in reported retrospective series. Hence, we suspected additional novel mechanisms of acquired res. Molecular subtype switching from the aEa sensitive TA subtype to the comparatively insensitive SL subtype occurred in paired BL/PD bx from 3/7 pts who progressed after prolonged aEa benefit without detectable genetic res drivers in PD bx. Subtype switching was not observed in any of 6 analyzed BL/PD pairs from primary progressors. Considering the strong association of aEa sensitivity with TA subtype at BL, these data suggest subtype switching from TA to SL as a novel mechanism of acquired aEa res. This prospective trial revealed novel genetic (NF1 mut, FGF10 amp) and likely transcriptomic (TA to SL subtype switching) mechanisms of aEa resistance in CRC. These results should enable more precise aEa therapy stratification and may open opportunities to prevent res through SL subtype targeting strategies.
Citation Format: Khurum Hayat Khan, Andrew Woolston, Georgia Spain, Louise Barber, Yatish Patil, Beatrice Griffiths, Reyes GonzalezExposito, Sonia Mansukhani, Matthew Davies, Sheela Rao, David Watkins, Francesco Sclafani, Jana Suntharanathan, Clare Peckitt, Ruwaida Begum, Isma Rana, Janet Thomas, Jacqui Oates, Annette Bryant, Andrew Wotherspoon, Nicos Fotiadis, Nasir Khan, Sebastian Guettler, Katharina von Loga, Naureen Starling, Ian Chau, Anguraj Sadanandam, David Cunningham, Marco Gerlinger. Molecular subtypes and novel genetic mechanisms of primary and acquired anti-EGFR resistance in colorectal cancer in the Prospect C biomarker trial [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4339.