Abstract
BackgroundAt least 50% of triple negative breast cancer (TNBC) overexpress the epidermal growth factor receptor, EGFR, which paved the way for clinical trials investigating its blockade. Outcomes remained dismal stemming from mechanisms of resistance particularly the nuclear cycling of EGFR, which is enhanced by Src activation. Attenuation of Src reversed nuclear translocation, restoring EGFR to the cell surface. Herein, we hypothesize that changes in cellular distribution of EGFR upon Src inhibition with dasatinib can be annotated through the EGFR immunopositron emission tomography (immunoPET) radiotracer, [89Zr]Zr-cetuximab.MethodsNuclear and non-nuclear EGFR levels of dasatinib-treated vs. untreated MDA-MB-231 and MDA-MB-468 cells were analyzed via immunoblots. Both treated and untreated cells were exposed to [89Zr]Zr-cetuximab to assess binding at 4 °C and 37 °C. EGFR-positive MDA-MB-231, MDA-MB-468, and a patient-derived xenograft were treated with dasatinib or vehicle followed by cetuximab PET imaging to compare EGFR levels. After imaging, the treated mice were separated into two groups: one cohort continued with dasatinib with the addition of cetuximab while the other cohort received dasatinib alone. Correlations between the radiotracer uptake vs. changes in tumor growth and EGFR expression from immunoblots were analyzed.ResultsTreated cells displayed higher binding of [89Zr]Zr-cetuximab to the cell membrane at 4 °C and with greater internalized activity at 37 °C vs. untreated cells. In all tumor models, higher accumulation of the radiotracer in dasatinib-treated groups was observed compared to untreated tumors. Treated tumors displayed significantly decreased pSrc (Y416) with retained total Src levels compared to control. In MDA-MB-468 and PDX tumors, the analysis of cetuximab PET vs. changes in tumor volume showed an inverse relationship where high tracer uptake in the tumor demonstrated minimal tumor volume progression. Furthermore, combined cetuximab and dasatinib treatment showed better tumor regression compared to control and dasatinib-only-treated groups. No benefit was achieved in MDA-MB-231 xenografts with the addition of cetuximab, likely due to its KRAS-mutated status.ConclusionsCetuximab PET can monitor effects of dasatinib on EGFR cellular distribution and potentially inform treatment response in wild-type KRAS TNBC.
Highlights
At least 50% of triple negative breast cancer (TNBC) overexpress the epidermal growth factor receptor, EGFR, which paved the way for clinical trials investigating its blockade
We examined our hypothesis in a longitudinal study using established MDA-MB-231, MDA-MB-468 and TNBC patient-derived xenografts (PDX)
In MDA-MB468 cells (Fig. 1a, Table S1), a significant decrease was displayed in both pEGFR (Y845) (2.40 ± 0.08 to 0.53 ± 0.03, p = 0.006) and pSrc (Y416) (1.10 ± 0.06 to 0.50 ± 0.06, p = 0.044) after treatment
Summary
At least 50% of triple negative breast cancer (TNBC) overexpress the epidermal growth factor receptor, EGFR, which paved the way for clinical trials investigating its blockade. While many patients initially respond to chemotherapy, the high rate of recurrence makes it a far more aggressive disease with worse prognosis compared to other subtypes. This created a strong impetus to find other biomarkers for targeting TNBC. Gene expression profiling studies identified the epidermal growth factor receptor (EGFR) as a potential biomarker in at least 50% of patients with TNBC [2], paving the way for clinical trials investigating EGFRtargeted therapies, including monoclonal antibodies (panitumumab and cetuximab) and small molecule inhibitors (gefitinib, erlotinib, and afatinib) within this patient population [3]. One rationale likely stems from the loss of EGFR from the cell surface as a result of its translocation to the nucleus; potentially diminishing drug target and accessibility
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