Immuno-PET Imaging of Atherosclerotic Plaques with [89Zr]Zr-Anti-CD40 mAb-Proof of Concept.

Kikkie Poels,Guus A M S Van Dongen,Maxime Schreurs,Matthijs Jansen,Danielle J Vugts,Esther Lutgens,Wissam Beaino,Tom T P Seijkens

doi:10.3390/biology11030408

Abstract

Simple SummaryAtherosclerosis is characterized by gradual plaque build-up in the middle and large arteries and is the major cause of cardiovascular disease. Determining which plaques are prone to rupture and cause potential lethal effects (e.g., myocardial infarction) could greatly reduce the potential bad outcomes. CD40 is a co-stimulatory molecule present in various cells in the plaque and has been shown to correlate with plaque vulnerability. In this manuscript, we have combined a murine monoclonal antibody against CD40 (a costimulatory molecule directly linked to plaque progression and present on many cells within the lesion) with Zirconium-89 to test its applicability to detect lesions in a mouse model of atherosclerosis using PET/CT. We show that this Zirconium-89 labeled antibody can detect CD40 in atherosclerotic lesions in a mouse model of atherosclerosis. In wild type mice without plaques, no signal was found. Our results suggest that CD40 could be a potential marker for PET imaging of plaque inflammation and vulnerability.Non-invasive imaging of atherosclerosis can help in the identification of vulnerable plaque lesions. CD40 is a co-stimulatory molecule present on various immune and non-immune cells in the plaques and is linked to inflammation and plaque instability. We hypothesize that a 89Zr-labeled anti-CD40 monoclonal antibody (mAb) tracer has the potential to bind to cells present in atherosclerotic lesions and that CD40 Positron Emission Tomography (PET) can contribute to the detection of vulnerable atherosclerotic plaque lesions. To study this, wild-type (WT) and ApoE−/− mice were fed a high cholesterol diet for 14 weeks to develop atherosclerosis. Mice were injected with [89Zr]Zr-anti-CD40 mAb and the aortic uptake was evaluated and quantified using PET/Computed Tomography (CT) imaging. Ex vivo biodistribution was performed post-PET imaging and the uptake in the aorta was assessed with autoradiography and compared with Oil red O staining to determine the tracer potential to detect atherosclerotic plaques. On day 3 and 7 post injection, analysis of [89Zr]Zr-anti-CD40 mAb PET/CT scans showed a more pronounced aortic signal in ApoE−/− compared to WT mice with an increased aorta-to-blood uptake ratio. Autoradiography revealed [89Zr]Zr-anti-CD40 mAb uptake in atherosclerotic plaque areas in ApoE−/− mice, while no signal was found in WT mice. Clear overlap was observed between plaque areas as identified by Oil red O staining and autoradiography signal of [89Zr]Zr-anti-CD40 mAb in ApoE−/− mice. In this proof of concept study, we showed that PET/CT with [89Zr]Zr-anti-CD40 mAb can detect atherosclerotic plaques. As CD40 is associated with plaque vulnerability, [89Zr]Zr-anti-CD40 mAb has the potential to become a tracer to detect vulnerable atherosclerotic plaques.

Highlights

Atherosclerosis is a chronic inflammatory disease of the middle- and large-sized arteries that results in the establishment of plaques in the arterial wall
Positron Emission Tomography (PET)/Computed Tomography (CT) imaging was performed at day 3 and 7 p.i. to evaluate the potential of [89Zr]Zr-antiCD40 monoclonal antibody (mAb) (“low dose” of 100 μg) for in vivo targeting and detection of aortic plaques
Focal uptake of [89Zr]Zr-anti-CD40 mAb was observed in the aortic arch of the ApoE−/− mice (Figure 2B), which was not observed in the aortic arch of WT mice (Figure 2D), indicating binding to the atherosclerotic plaques that are present in the aorta of ApoE−/− mice

Summary

Introduction

Atherosclerosis is a chronic inflammatory disease of the middle- and large-sized arteries that results in the establishment of plaques in the arterial wall. While atherosclerosis often starts from an early age, its detrimental effects do not become apparent until plaques become advanced and lead to potentially lethal consequences such as myocardial infarction (MI) and ischemic stroke [1]. In 2016, 84.9% of cardiovascular deaths were caused by MIs and strokes, both the result of atherosclerosis, amounting to 15.1 million deaths worldwide per year [2]. Detection and stabilization of unstable plaques could potentially eliminate these lethal ramifications and reduce cardiovascular death. Many factors contribute to a vulnerable plaque phenotype, and while advancements have been made in the latest years, it is still unclear what is the determining factor that drives plaques to rupture and cause cardiovascular complications [3]. Plaques with a vulnerable phenotype still have the potential for stabilization, either spontaneously or drug-induced [4,5], so their identification has major therapeutic impact. While remodeling and restabilization of plaques greatly decreases the chance of rupture of inflammatory plaques, patients with such restabilized lesions are often at increased risk for developing vulnerable lesions elsewhere in the vasculature and are still at risk for developing MI [7]

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