Upregulation of annexin A1 protein expression in the intratumoral vasculature of human non-small-cell lung carcinoma and rodent tumor models.

Kevin L Allen,M Jack Borrok,Michelle Lazzaro,Norman Peterson,Ping Tsui,Herren Wu,Weiguang Zhao,Qing Li,Melissa M Damschroder,Jennifer Cann,Haihong Zhong,William F Dall’Acqua,Ilya Ulasov

doi:10.1371/journal.pone.0234268

Abstract

Annexin A1 (anxA1) is an immunomodulatory protein that has been proposed as a tumor vascular target for antitumor biologic agents, yet to date the vascular expression of anxA1 in specific tumor indications has not been systematically assessed. Attempts to evaluate vascular anxA1 expression by immunohistochemistry are complicated by a lack of available antibodies that are both specific for anxA1 and bind the N-terminal–truncated form of anxA1 that has previously been identified in tumor vasculature. To study the vascular expression pattern of anxA1 in non–small-cell lung carcinoma (NSCLC), we isolated an antibody capable of binding N-terminal–truncated anxA127-346 and employed it in immunohistochemical studies of human lung specimens. Lung tumor specimens evaluated with this antibody revealed vascular (endothelial) anxA1 expression in five of eight tumor samples studied, but no vascular anxA1 expression was observed in normal lung tissue. Tumor microarray analysis further demonstrated positive vascular staining for anxA1 in 30 of 80 NSCLC samples, and positive staining of neoplastic cells was observed in 54 of 80 samples. No correlation was observed between vascular and parenchymal anxA1 expression. Two rodent tumor models, B16-F10 and Py230, were determined to have upregulated anxA1 expression in the intratumoral vasculature. These data validate anxA1 as a potential vascular anti-tumor target in a subset of human lung tumors and identify rodent models which demonstrate anxA1 expression in tumor vasculature.

Highlights

We generated antibodies that are specific for Annexin A1 (anxA1) and capable of binding its proteolytically cleaved form. Employing these uniquely specific antibodies, we evaluated the expression pattern of anxA1 in human non–small-cell lung carcinoma (NSCLC) tissue samples to determine the prevalence of vascular anxA1 expression across patient samples
These results were similar to the IHC analysis of human tissue in so much as no direct association of anxA1 expression was observed between intratumoral vessel staining and neoplastic cell staining across these models
Published data on anxA1 expression in tumor vasculature has established the presence of a membrane-bound, N-terminal–truncated form of anxA1 on the luminal membrane of tumor-associated endothelium, and anxA1 has been proposed as a vascular antitumor target [19,20,21]

Summary

Introduction

The vasculature of tumor tissue is distinct from that of normal tissue in both morphology and gene expression, and expression of multiple proteins is upregulated in tumor endothelium [1–. It was reported that expression of the immunomodulatory protein annexin A1 (anxA1) is enhanced in tumor-associated endothelium, and an antibody targeting a membrane-associated, proteolytically cleaved form of anxA1 (anxA127-346) was reported to induce rapid tumor uptake in rodent models, including models of lung cancer [19,20,21]. We generated antibodies that are specific for anxA1 and capable of binding its proteolytically cleaved form Employing these uniquely specific antibodies, we evaluated the expression pattern of anxA1 in human NSCLC tissue samples to determine the prevalence of vascular anxA1 expression across patient samples.

Materials and methods

Results

Discussion