Chemical design of radiolabeled antibody fragments for low renal radioactivity levels

The renal uptake of radiolabeled antibody fragments presents a problem in t argeted imaging and therapy, We hypothesized that the renal radioactivity l evels of radiolabeled antibody fragments could be reduced if radiolabeled c ompounds of urinary excretion were released from glomerularly filtered anti body fragments before they were incorporated into renal cells by the action of brush border enzymes, present on the lumen of renal tubules, 3'-[I-131] Iodohippuryl N-epsilon-maleoyl-L-lysine ([I-131]HML) was conjugated with a thiolated Fab fragment because the glycyl-lysine sequence in HML is a subst rate for a brush border enzyme and meta-iodohippuric acid is released by cl eavage of the linkage. Fab fragments were also radiolabeled by direct radio iodination (I-125-Fab) or by conjugation with meta-[I-125]-iodohippuric aci d via an amide bond [N-(5-maleimidopentyl) 3'-iodohippuric acid amide ([I-1 25]MPH-Fab)] or an ester bond [maleimidoethy 3'-iodohippurate ([I-125]MIH-F ab)] by procedures similar to those used for [I-131]HML-Fab. In biodistribution experiments in mice, [I-131]HML-Fab demonstrated markedl y low renal radioactivity levels with kidney:blood ratios of radioactivity of 1 from 10 min to 1 h due to rapid release of meta-[I-131]iodohippuric ac id. [I-125]MIH-Fab and I-125-Fab reached their peak ratios of 3.8 and 7.3 a t 1 h, respectively, and [I-125]MPH-Fab showed the maximum ratio of 16.8 at 6 h, In subcellular distribution studies, both [I-125]MIH-Fab and I-125-Fa b showed migration of radioactivity from the membrane to the lysosomal frac tion of the renal cells from 10 to 30 min postinjection, whereas the majori ty of the radioactivity was detected only in the membrane fraction after ad ministration of [I-131]HML-Fab at both time points. In nude mice, [I-131]HM L-Fab showed one-quarter of the renal radioactivity of simultaneously admin istered I-125-Fab without impairing the target radioactivity levels 3 h aft er injection. These findings indicated that HML is a useful reagent for tar geted imaging and therapy using antibody fragments as vehicles, These findi ngs also suggested that the radiochemical design of radiolabeled antibody f ragments that liberate radiometabolites of urinary excretion from antibody fragments by the action of brush border enzymes may constitute a new strate gy for reducing the renal radioactivity levels of antibody fragments.