Effects of the EGFR Inhibitor Erlotinib on Tumor Microenvironment: Implications for Therapy

Abstract

Purpose/Objective(s): A recently published randomized study showed that combining an EGFR inhibitor with radiation increases survival and local control in patients with locally advanced head and neck cancer. We were interested in understanding the effects of EGFR inhibition on the tumor microenvironment in a head and neck cancer xenograft model, as this could provide insight into its effectiveness in combination with radiation and chemotherapy. Materials/Methods: We used SQ20B head and neck squamous cell carcinoma cells either in vitro or grown as subcutaneous xenografts in nude mice. Both in vitro (Western blotting, radiation cell survival assays) and in vivo studies (tumor regrowth assay, EF5 staining) were conducted. Results: We saw a minimal effect of erlotinib oninvitro radiosensitization. However, mice with SQ20B xenografts treated with the drug had prolonged time to tumor regrowth after 6 Gy of radiation compared to mice given radiation only. Because the in vivo effects appeared greater than the in vitro effects, we investigated the effects of erlotinib on the tumor microenvironment. Erlotinib decreased both vascular endothelial growth factor (VEGF) and hypoxia-inducible factor (HIF)-1a expression in vitro and in tumor xenografts grown in vivo. We measured vascular permeability within the xenografts using Evan’s blue dye and found a 75% reduction in permeability in the tumors of mice treated with the drug. Consistent with these results, tomato lectin staining of the tumor vasculature showed that erlotinib treatment led to a dramatic change in vessel morphology. Using Power Doppler we found that there was a greater than 3-fold increase in tumor perfusion within 48 hours of starting erlotinib treatment. This increased vascular perfusion was evident even at 96 hours, in spite of the fact that the tumors showed a 50% decrease in size by this time. Using the hypoxia marker EF5 we found that the pattern of hypoxia was heterogeneous in these tumors but that erlotinib treatment led to improved oxygenation. Conclusions: Erlotinib decreases HIF-1a/VEGF expression in SQ20B xenografts but paradoxically increases vascular perfusion. A potential explanation for this is the idea of ‘‘vascular normalization’’ as proposed by Jain (Science 2005;307:58). This could lead to increased delivery of chemotherapy to tumors in patients responding to erlotinib. Improved vascular perfusion may also lead to increased oxygenation that could play a role in increased in vivo radiosensitization.