Abstract 4459: Evaluating imaging biomarkers of acquired resistance to targeted EGFR therapy in xenograft models of human squamous cell carcinoma of the head and neck (SCCHN).
Authors: Baker Lauren CJ , Box Carol , Sikka Arti , Box Gary , Eccles Suzanne A. , Robinson Simon P. 
Published: August 2014

Abstract
Overexpression of EGFR has been identified as a negative prognostic factor in squamous cell carcinoma of the head and neck (SCCHN). Patients with SCCHN who respond to EGFR-targeted tyrosine kinase inhibitors (TKIs) eventually develop acquired resistance. Colocalization of EGFR expression and tumor hypoxia in SCCHN is associated with poor outcome, suggesting a role for hypoxia in drug resistance. Non-invasive imaging strategies to accurately identify and monitor patients whose tumors have become resistant to EGFR-TKI therapy would have clinical value. As part of a pre-clinical multiparametric imaging study, we have investigated EGFR-TKI resistance using intrinsic susceptibility magnetic resonance imaging (IS-MRI), which relies on the dependence of the MRI transverse relaxation rate R2* on the ratio of oxy- to deoxyhemoglobin in blood. Given its relationship to blood oxygen saturation and pO2 in and around blood vessels, R2* and hyperoxia-induced ΔR2* are being investigated as imaging biomarkers of tumor oxygenation.
IS-MRI was performed on size-matched xenografts derived from the EGFR TKI sensitive SCCHN cell line CAL27 and an isogenic subline resistant to multiple TKIs (gefitinib, erlotinib, and afatinib). Tumor-bearing mice were administered the hypoxia marker pimonidazole, and baseline R2* quantified from resistant (n=8) or sensitive (n=7) tumors whilst the host breathed air. Gas delivery was then switched to 100% O2, and tumour R2* measured again. Finally, mice were administered the perfusion marker Hoechst 33342. The extent of functional tumor vasculature and hypoxia were quantified ex vivo using fluorescence microscopy. Resistant tumors revealed regions of fast R2* (functional, deoxygenated vasculature) restricted primarily to the tumor periphery, while sensitive tumors exhibited a more heterogeneous distribution of fast R2* throughout. There was no significant difference in baseline R2* between the two cohorts (64 ± 4s-1 and 76 ± 5s-1, p=0.07). Hyperoxia resulted in a significantly smaller ΔR2* in the resistant tumors (-2.4 ± 1.4s-1) compared to sensitive tumors (-9.1 ± 2s-1, p&lt;0.05). Resistant tumors had significantly lower Hoechst 33342 uptake (9.5 ± 1.3%, p&lt;0.01) and increased pimonidazole adducts (21 ± 2%, p&lt;0.01) compared to sensitive tumors (19.2 ± 3% and 10.4 ± 2% respectively). Tumor regions of fast R2* and hyperoxic-induced ΔR2* were spatially associated with Hoechst 33342 uptake.
Tumors with resistance to EGFR-TKIs exhibited a reduced hemodynamic MRI response that was associated with decreased vessel perfusion and increased tumor hypoxia. IS-MRI informs on phenotypic differences in tumor blood vessel distribution, functionality and oxygenation associated with EGFR-TKI resistance and sensitivity, affording useful non-invasive imaging biomarkers for investigating EGFR drug resistance and tumor hypoxia.
Citation Format: Lauren CJ Baker, Carol Box, Arti Sikka, Gary Box, Suzanne A. Eccles, Simon P. Robinson. Evaluating imaging biomarkers of acquired resistance to targeted EGFR therapy in xenograft models of human squamous cell carcinoma of the head and neck (SCCHN). [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4459. doi:10.1158/1538-7445.AM2013-4459