Radiolabeled methotrexate as a diagnostic agent of inflammatory target sites: A proof-of-concept study

Methotrexate (MTX), as a pharmaceutical, is frequently used in tumor chemotherapy and is also a part of the established treatment of a number of autoimmune inflammatory disorders. Radiolabeled MTX has been studied as a tumor-diagnostic agent in a number of published studies. In the present study, the potential use of technetium-99m-labelled MTX ((99m)Tc-MTX) as a radiotracer was investigated for the identification of inflammatory target sites. The labelling of MTX was carried out via a (99m)Tc-gluconate precursor. Evaluation studies included in vitro stability, plasma protein binding assessment, partition-coefficient estimation, in vivo scintigraphic imaging and ex vivo animal experiments in an animal inflammation model. MTX was successfully labelled with (99m)Tc, with a radiochemical purity of >95%. Stability was assessed in plasma, where it remained intact up to 85% at 4 h post-incubation, while protein binding of the radiotracer was observed to be ~50% at 4 h. These preclinical ex vivo and in vivo studies indicated that (99m)Tc-MTX accumulates in inflamed tissue, as well as in the spinal cord, joints and bones; all areas with relatively high remodeling activity. The results are promising, and set the stage for further work on the development and application of (99m)Tc-MTX as a radiotracer for inflammation associated with rheumatoid arthritis.