Abstract
Mounting evidence suggests that the urokinase plasminogen activator (uPA) and its receptor (uPAR) play a central role in tumor progression. The goal of this study was to develop an 89Zr-labeled, antibody-based positron emission tomography (PET) tracer for quantitative imaging of the uPA/uPAR system. An anti-uPA monoclonal antibody (ATN-291) was conjugated with a deferoxamine (Df) derivative and subsequently labeled with 89Zr. Flow cytometry, microscopy studies, and competitive binding assays were conducted to validate the binding specificity of Df-ATN-291 against uPA. PET imaging with 89Zr-Df-ATN-291 was carried out in different tumors with distinct expression levels of uPA. Biodistribution, histology examination, and Western blotting were performed to correlate tumor uptake with uPA or uPAR expression. ATN-291 retained uPA binding affinity and specificity after Df conjugation. 89Zr-labeling of ATN-291 was achieved in good radiochemical yield and high specific activity. Serial PET imaging demonstrated that, in most tumors studied (except uPA- LNCaP), the uptake of 89Zr-Df-ATN-291 was higher compared to major organs at 120 h post-injection, providing excellent tumor contrast. The tumor-to-muscle ratio of 89Zr-Df-ATN-291 in U87MG was as high as 45.2 ± 9.0 at 120 h p.i. In vivo uPA specificity of 89Zr-Df-ATN-291 was confirmed by successful pharmacological blocking of tumor uptake with ATN-291 in U87MG tumors. Although the detailed mechanisms behind in vivo 89Zr-Df-ATN-291 tumor uptake remained to be further elucidated, quantitative PET imaging with 89Zr-Df-ATN-291 in tumors can facilitate oncologists to adopt more relevant cancer treatment planning.
Highlights
Numerous cellular events orchestrate the initiation, progression, and metastasis of cancer [1]
Most important of all, accumulating evidence suggests a central role of urokinase plasminogen activator (uPA)/uPA www.impactjournals.com/oncotarget receptors (uPAR) in cancer progression especially in cancer metastasis [3]. uPA along with its “partners” (e.g. PAI1) can serve as useful biomarkers to forecast disease progression as their expression levels in tumor tissues correlates with prognosis in different types of cancer [4]
The structure of uPA is formed by three distinct regions: a growth factor-like domain (GFD), a kringle domain, and a serine protease domain [6]. uPA possesses very specific proteolytic activity against its preferred substrate, plasminogen, and catalyzes its activation to plasmin
Summary
Numerous cellular events (e.g. cell migration, invasion, angiogenesis etc.) orchestrate the initiation, progression, and metastasis of cancer [1]. Over the past several decades, a multitude of biomarkers have been identified with the goal of elucidating these events for use as diagnostic/therapeutic targets Among these markers, the urokinase plasminogen activator (uPA) system, which is primarily composed of uPA (a serine protease), uPA www.impactjournals.com/oncotarget receptors (uPAR), and several inhibitors/modulators of uPA (especially plasminogen activator inhibitor 1 [PAI-1]), have attracted special attention as they serve as fundamental mediators of signaling pathways contributing to tumor microenvironment regulation [2]. UPA possesses very specific proteolytic activity against its preferred substrate, plasminogen, and catalyzes its activation to plasmin This transformation triggers the activation of various proteases and causes the degradation of the extracellular matrix (ECM) [7], eventually increasing cell motility, invasion, and survival [8]. The interaction between uPA and uPAR recruits uPA to the cell surface and is known to enhance its catalytic efficiency for plasminogen activation by up to two orders of magnitude [9]
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