Patients with RAS and BRAF wt mCRC are regarded as the best candidates to receive anti-EGFRs. Nevertheless, almost half of these patients do not achieve response, so that a deeper refinement of candidates’ selection would be highly desirable.

Different molecular alterations, supported by a strong and sound biologic rationale, have been suggested as predictors of primary resistance to anti-EGFRs, but up today their potential impact has been suggested only in preclinical experiences and retrospective series, and their low frequency makes difficult the validation of each single marker.

The present case-control study was conducted to prospectively demonstrate the negative predictive impact of HER-2 amplification or mutations, MET amplification, NTRK/ROS1/ALK/RET rearrangements, and mutations activating MAPKs or PI3K/Akt axis.

Patients with RAS and BRAF wt mCRC clearly resistant (cases) vs. clearly sensitive (controls) to single-agent anti-EGFRs were selected, their archival tissue samples were collected and the prevalence of above-mentioned alterations was prospectively assessed.

HER-2 status was evaluated by immunohistochemistry (IHC) and silver in-situ hybridization (SISH); MET amplification was studied by SISH; gene rearrangements were screened by IHC followed by RNA-based NGS confirmation. Other candidate mutations were investigated by NGS (Hotspot Cancer Panel v2, Ion Torrent Personal Genome platform (Life Technologies®).

Hypothesizing a prevalence of candidate alterations of 0% and 15% among controls and cases, respectively, we needed to include 47 cases and 47 controls to be able to reject the null hypothesis that the prevalence of alterations is equal, with α-error 0.05 and β-error 0.20.

Moreover, we evaluated the predictive impact of microsatellite instability, since hypermutated tumors may hardly rely on a single pathway (i.e. EGFR) for their growth.

Forty-six cases and 47 controls were included. The primary endpoint was met: above-mentioned alterations were reported in 19 (41.3%) cases and 1 (2.1%) control (p<0.001). MSI-high was significantly more frequent among resistant than sensitive patients (6% vs 0%, p=0.012).

This is the first prospective demonstration that the combined assessment of these rare alterations allows to better select patients for treatment with anti-EGFRs, while opening the way to properly tailored therapies.

   
Molecular alteration Cases (Resistant patients) Controls (Sensitive patients) 
 N=46 N=47 
HER-2 amplification 7* 
HER-2 mutations 1 (G776V, exon 20) 
MET amplification 5* 
NTRK rearrangements 2 (SCYL3-NTRK1 and TPM3-NTRK1
ALK rearrangements 
ROS1 rearrangements 
RET rearrangements 1 (CCDC6-RET
PIK3CA mutations (exon 20) 1 (A1035V, exon 20) 1 (H1047R, exon 20) 
PTEN mutations 3 (L247S, R233stop and del P248, exon 7) 
Total n. of patients with candidate alterations 19 1 
Microsatellite instability (MSI-high) 
RAS mutations at low allele fraction ** 2 (KRAS G12V, exon 2, 6%; NRAS Q61R, exon 3, 10%) 
New RAS mutations 3 (2 KRAS L19F, exon 2; KRAS T50I, exon 3) 
   
Molecular alteration Cases (Resistant patients) Controls (Sensitive patients) 
 N=46 N=47 
HER-2 amplification 7* 
HER-2 mutations 1 (G776V, exon 20) 
MET amplification 5* 
NTRK rearrangements 2 (SCYL3-NTRK1 and TPM3-NTRK1
ALK rearrangements 
ROS1 rearrangements 
RET rearrangements 1 (CCDC6-RET
PIK3CA mutations (exon 20) 1 (A1035V, exon 20) 1 (H1047R, exon 20) 
PTEN mutations 3 (L247S, R233stop and del P248, exon 7) 
Total n. of patients with candidate alterations 19 1 
Microsatellite instability (MSI-high) 
RAS mutations at low allele fraction ** 2 (KRAS G12V, exon 2, 6%; NRAS Q61R, exon 3, 10%) 
New RAS mutations 3 (2 KRAS L19F, exon 2; KRAS T50I, exon 3) 

* in 1 case HER-2 and MET co-amplification was found

** previously defined as wt by pyrosequencing

Citation Format: Chiara Cremolini, Rosa Berenato, Federica Morano, Roberto Moretto, Federica Perrone, Elena Tamborini, Daniele Rossini, Annunziata Gloghini, Adele Busico, Giovanni Fucà, Chiara Baratelli, Emiliano Tamburini, Iolanda Capone, Chiara Costanza Volpi, Massimo Milione, Massimo Di Maio, Gabriella Fontanini, Filippo De Braud, Alfredo Falcone, Filippo Pietrantonio. Dissecting primary resistance to anti-EGFR monoclonal antibodies (anti-EGFRs) in RAS and BRAF wild-type (wt) metastatic colorectal cancer (mCRC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr LB-238. doi:10.1158/1538-7445.AM2017-LB-238