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Background: <br>Optical imaging is an emerging modality for the detection of lesions without ionizing radiation. Optical imaging probes consisting of a targeting moiety and a fluorescent dye allow molecular imaging approaches, e.g. the specific staining of overexpressed proteins. Fluorescent dyes absorbing and emitting light in the near-infrared (NIR) range of the spectrum (700-900 nm) have proven to be advantageous because of the relative transparency of tissue and blood for this light compared to visible light of shorter wavelengths.
Collagen-induced arthritis is a model for acute arthritis including synovitis and the formation of an aggressive pannus. The inflammatory hypoxic environment leads to angiogenesis and expression of molecular markers thereof.
Objectives: <br>We investigated a molecular imaging approach for the detection of collagen-induced arthritis (CIA) in rats by targeting the Extra Domain B (EDB) of the extracellular matrix protein fibronectin. EDB is a highly conserved domain (identical in humans and rats) that is produced by alternative splicing during embryonic development and conditions of vascular remodeling like angiogenesis.
Methods:<br> We have confirmed the expression of EDB in the hyperplastic synovium of CIA rats by immunohistochemistry. For in vivo diagnostics, the EDB-binding single-chain antibody fragment AP39 was used as a targeting probe (Berndorff et al., 2006). It was covalently linked to the near-infrared (NIR) dye Tetrasulfocyanine (TSC) and visualized by fluorescence reflectance imaging using a 740 nm NIR laser source for excitation and a cooled iCCD-camera for detection. A conjugate consisting of TSC and ovalbumin, having a similar molecular weight as the spontaneously forming AP39-TSC dimer, was used as a control for targeting specificity.
Results: <br>AP39-TSC induced a significantly enhanced fluorescence contrast in arthritic joints that allowed discrimination from healthy control joints in the CIA-rat model up to 24 h after i.v. application. With ovalbumin-TSC, only a minor accumulation of the conjugate was observed in affected joints. This indicates that the enrichment is due to the specific binding of the probe to EDB.
Conclusion:<br> Optical imaging using the EDB-binding single-chain antibody fragment AP39 conjugated to the NIR-dye TSC resulted in clear enhancement of fluorescence intensity in affected joints compared to control joints. Since AP39 is a fully human single-chain antibody fragment, a translation of this molecular imaging approach for arthritis detection in man and possibly a repeated use for therapy response monitoring seems feasible.
References:<br> Berndorff, D., Borkowski, S., Moosmayer, D., Viti, F., Muller-Tiemann, B., Sieger, S., Friebe, M., Hilger, C.S., Zardi, L., Neri, D. and Dinkelborg, L.M. (2006) Imaging of tumor angiogenesis using 99mTc-labeled human recombinant anti-ED-B fibronectin antibody fragments. J Nucl Med, 47, 1707-1716.