Molecular Imaging with Macrophage VSIG4-targeting Nanobodies for Diagnosis and Prognosis in a mouse model of Rheumatoid Arthritis.

An accurate and noninvasive tracer able to detect molecular events underlying the development of rheumatoid arthritis (RA) would be of great help during RA diagnosis and to assess drug efficacy. The V-set and Ig domain-containing 4 (VSIG4) receptor is expressed on synovial macrophages of RA patients and is an interesting target for molecular imaging of RA using radiotracers. Here we report the visualization of VSIG4 in a mouse model for RA using a radiolabeled single domain variable antibody VHH fragment (also called Nanobody). A highly selective Nanobody (Nb) with nanomolar affinity, NbV4m119, was selected after screening a VSIG4-specific VHH-phage library. Flow cytometric analysis confirmed binding of the Nb to VSIG4+ transfected cells as well as to liver macrophage suspensions. In naive mice, single-photon emission computed tomography (SPECT/CT) imaging revealed that 99mTc-NbV4m119 exclusively targeted VSIG4+ liver macrophages, whereas no 99mTc-NbV4m119 tissue accumulation was found in VSIG4-/- mice. In collagen-induced arthritis (CIA) mice, arthritic lesions in inflamed paws could be successfully imaged using the 99mTc-NbV4m119 Nb. Phosphorimaging and confocal microscopy further confirmed NbVm119 binding to CD68+ synovial cells. Interestingly, the 99mTc-NbV4m119 signal in arthritic lesions increased according to the severity of the inflammation. In knees of collagen challenged mice, 99mTc-NbV4m119 was found to accumulate even before the onset of macroscopic clinical symptoms. Thus, imaging of the knees with the Nb could predict which mice will develop inflammation during CIA. Consequently, imaging of joint inflammation with VSIG4-specific Nanobodies offers perspectives for clinical applications in RA patients.