Download Imaging endothelial activation

Imaging endothelial activation
Despite the important role that endothelial activation plays during inflammatory
responses, endothelium is relatively inaccessible to clinical investigation. We have a
long-standing interest in the molecular imaging of endothelial activation, using Eselectin as a lead target. Monoclonal antibody 1.2B6 is a mouse antibody generated
in our laboratory, which primarily reacts with E-selectin but also reacts with Pselectin at about four-fold lower affinity 1, 2 (and unpublished observations). Using
this mAb, we were the first group to obtain images of cytokine-activated endothelium
in vivo. The technique was first evaluated in pig models of phytohaemaggltinininduced and MSU crystal-induced arthritis, in which we found that the images of
synovitis obtained with 111In-labelled mAb 1.2B6 F(ab’)2 were significantly more
focal and intense than those obtained with an isotype matched control mAb or 99mTclabelled leukocytes 3-5. We then went on to show that 111In-labelled mAb 1.2B6
F(ab’)2 could image inflamed joints in patients with RA 6, 7 and inflamed bowel in
Crohn’s Disease and Ulcerative Colitis 8
Radioscintigraphy
In view if the accessibility of inflamed tissue, RA provides a good clinical model with
which to develop inflammation imaging. We have continued to collaborate with Dr
Francois Jamar (University of Louvain, Brussels) to refine radioscintigraphic imaging
with mAb 1.2B6. Over the last five years, Jamar has shown that 99mTc-Fab fragment
of 1.2B6 discriminates better than 99mTc-Fab-HDP (the tracer used for clinical bone
scans) between actively inflamed and silent joints, providing further data supporting
the potential of mAb 1.2B6 as an imaging agent 9.
Magnetic resonance
Exposure to ionising radiation and relatively poor anatomical resolution of nuclear
gamma cameras are limiting factors for the broader application of the use of
radiolabeled antibodies for imaging. Therefore other modalities have been assessed in
vitro for E-selectin targeting potential, including MR 10, 11 and near infra-red optical
imaging 12. Furthermore, sialyl Lewis X, a carbohydrate moiety that binds E-selectin,
has been conjugated to gadolinium, and used to target E-selectin for MR imaging in
preclinical models of focal brain ischaemia 13 and cytokine-mediated inflammation 14.
Ultrasmall superparamagnetic iron oxide (USPIO) nanoparticles are MR imaging
contrast agents that consist of an iron oxide core about 5nm in size, surrounded by a
non-covalently attached low molecular weight dextran which increases the size to
about 30-50nm. USPIO align magnetically in an applied magnetic field and cause
microscopic field gradients that efficiently dephase nearby protons and disrupt the
homogeneity of the magnetic field, thereby strongly enhancing the T2 and T2*
relaxation rate over a length scale much larger than their size. Hence they are called
T2 contrast agents. They are also referred to as “negative” contrast agents since they
decrease the signal intensity in standard imaging sequences 15. Their relaxivity
increases with field strength up to its saturation threshold, with higher MR field
strengths producing higher signal and hence higher contrast-to-noise imaging. Several
USPIO preparations are in advanced stages of clinical trials, with their safety in
humans being increasingly established 16-18. Originally designed for lymph node
imaging 15, USPIO have been used for intracellular labelling 19, with degradation
occurring through normal physiological iron-handling pathways. This gives USPIO
significant potential safety advantages over gadolinium or other heavy metal-based
MR contrast agents, which have no known intracellular excretion pathway.
Conjugation of USPIO with ligands or antibodies offers the possibility of specific
MR imaging of molecular targets. USPIO have previously been modified to bind Eselectin in vitro, as reported by Kang et al. 10. We have extended this approach into an
in vivo application. We have generated an E-selectin binding USPIO and shown that
this allows specific imaging of E-selectin with MR in vivo, using a contact
hypersensitivity mouse model of acute inflammation in the ear 20, 21.
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