Chemical modification to reduce renal uptake of disulfide-bonded variable region fragment of anti-Tac monoclonal antibody labeled with ^99mTc

The anti-Tac disulfide-bonded variable region fragment (dsFv) is a genetically engineered, 25 kDa, murine monoclonal antibody fragment that recognizes the α subunit of the interleukin-2 receptor (IL-2Rα). The dsFv radiolabeled with the tetrafluorophenyl ester (TFP) of [^99mTc]mercaptoacetyltriglycine ([^99mTc]MAG3-TFP) showed rapid tumor uptake and fast blood clearance in mice, resulting in high tumor-to-nontumor background ratios. However, its high renal uptake was a problem. In this study, we tested the effect of lowering the isoelectric point (pI) of dsFv to <9.3 on renal and tumor uptake. To lower the pI, dsFv was acylated simultaneously with both [^99mTc]MAG3-TFP and TFP-glycolate. The acylation of dsFv decreased its pI and its immunoreactivity inversely proportional to the molar ratio of TFP-glycolate to dsFv, whereas the conjugation of [^99mTc]MAG3-TFP alone did not. When biodistribution studies were performed in nude mice, the effect of the lowered pI was reflected primarily in decreased kidney uptake and whole-body retention, with its highest effect seen at the earliest time point (15 min) after injection. In tumor-bearing nude mice, glycolated [^99mTc]MAG3-dsFv with a pI range of 4.9 to 6.5 accumulated selectively into IL-2 receptor-positive SP2/Tac tumor similar to that of the control [¹²⁵I]dsFv labeled by the Iodo-Gen method, whereas its renal uptake was 25% of [¹²⁵I]dsFv at 15 min. At 90 min, the ratios of tumor to receptor-negative SP2/0 tumor, liver, kidney, stomach, and blood had peaked at 10.9, 8.5, 0.3, 5.0, and 6.2, respectively, for the glycolated [^99mTc]MAG3-dsFv. The corresponding ratios for [¹²⁵I]dsFv were 3.7, 5.0, 0.1, 1.5, and 2.1, respectively.