Evaluation of 99mTc-MAG3-annexin V: influence of the chelate on in vitro and in vivo properties in mice

We conjugated mercaptoacetyltriglycine (MAG3) to rh-annexin V to permit radiolabeling with 99mTc in an effort to decrease the high kidney and liver accumulation observed for 99mTc-labeled Hynic-annexin V. The 36-kDa protein was conjugated at a 5:1 molar ratio with NHS-MAG3 in HEPES buffer pH 7.8 at room temperature, then quenched with glycine and purified by dialysis. The biopotency of the resulting MAG3-annexin was similar to that of Hynic-annexin as determined by a sensitive red blood cell membrane affinity binding assay and a surface plasmon resonance (SPR) assay. The 99mTc radiolabeling of MAG3-annexin resulted in radiochemical yields of 90% under mildly basic pH conditions. Biodistribution data in normal mice clearly showed a significant decrease in kidney and liver uptake at 1 h postinjection for the 99mTc MAG3-annexin compared to the 99mTc Hynic-annexin (from 24% ID to 4% ID for the liver, and 45% ID to 15% ID for the kidneys, respectively). Autoradiography of the kidneys showed retention of radioactivity in the collecting tubules following administration of both labeled annexins. The 99mTc MAG3-annexin biodistribution was also characterized by a lower retention of radioactivity in the whole body, but with small intestine accumulation over fivefold higher than observed with 99mTc Hynic-annexin. These findings show a definite improvement in renal and hepatic clearance of the MAG3 radioligand. However, due to the increased radioactivity uptake in the small intestines, the early in vivo detection of ongoing apoptosis in the lower abdomen might be more difficult with 99mTc MAG3-annexin. Nevertheless, 99mTc MAG3-annexin may be an attractive alternative to 99mTc Hynic-annexin for the in vivo imaging of phosphatidylserine receptors.