Preferential Cleavage of a Tripeptide Linkage by Enzymes on Renal Brush Border Membrane To Reduce Renal Radioactivity Levels of Radiolabeled Antibody Fragments

Abstract

The obstructive renal radioactivity after injection of antibody fragments/constructs labeled with metallic radionuclides would be improved by liberating a radiometal chelate of urinary excretion from the antibody molecules by enzymes on the renal brush border membrane (BBM). A tripeptide GFK sequence was newly evaluated as an enzyme-cleavable linkage and conjugated to a Tc chelate of an isonicotinic acid derivative of 2-picolylglycine (Tc-IPG). Tc-IPG-glycine was liberated fromTc-IPG-GFK by the enzymes, while Tc-IPG-GK (where the tripeptide GFK was substituted with a dipeptide GK) did not. When injected into mice,Tc-IPG-GFK-conjugated Fab exhibited lower renal radioactivity levels than directly radioiodinated Fab shortly after injection without reducing the tumor radioactivity levels, due to a release and excretion of Tc-IPG-glycine by enzymes present on the renal BBM. These findings would provide insights to develop antibody fragments/constructs labeled with metallic radionuclides of the clinical relevance for improved renal radioactivity levels.

Development and characterization of a 68Ga-labeled A20FMDV2 peptide probe for the PET imaging of αvβ6 integrin-positive pancreatic ductal adenocarcinoma

“…In contrast, the accumulation of [ 68 Ga]Ac-CG6 in the kidneys remained at a level greater than 75 %ID/g at 2 h post-injection. Therefore, further structural modification, such as the incorporation of renal brush border enzyme-cleavable linkers, 34 will be required to reduce the renal retention of [ 68 Ga]Ac-CG6.…”

Bismuth-213 for Targeted Radionuclide Therapy: From Atom to Bedside

“…A study has demonstrated that derivatives of [ 68 Ga]Ga-DOTA-AmBz-MVK-OH could be cleaved specifically by neutral endopeptidase (NEP), and therefore, the amino acids linker MVK could reduce kidney uptake of radiolabeled DOTA-conjugated peptides and peptidomimetics [133]. These findings would allow to develop radiometal-based radiopharmaceuticals with clinically relevant lower renal radioactivity levels [134].…”

An Efficient Method for Labeling Single Domain Antibody Fragments with 18F Using Tetrazine-Trans-Cyclooctene Ligation and a Renal Brush Border Enzyme-Cleavable Linker

“…18–19 Incorporation of renal brush border enzyme (BBE)-cleavable linkers such as glycine-lysine (GK) or glycine-tyrosine dipeptide between the radiolabeled moiety and the targeting vector significantly reduced renal activity levels from radiolabeled antibody fragments without concomitant reduction in their tumor uptake. 20–21…”

“…High and persistent localization of activity from radiolabeled peptides and low molecular weight antibody fragments in the kidneys has been attributed to several factors. − These include electrostatic interaction between the negatively charged proximal tubular cell surface and positively charged molecules and endocytosis mediated by receptors such as megalin and cubulin. Strategies to reduce renal accumulation of activity by countering these factors have included administration of basic amino acids and the plasma expander gelofusine, increasing the negative charge of the labeled entity via chemical modification, and insertion of cleavable linkers. , Incorporation of renal brush border enzyme (BBE)-cleavable linkers such as glycine-lysine (GK) or glycine-tyrosine dipeptide between the radiolabeled moiety and the targeting vector significantly reduced renal activity levels from radiolabeled antibody fragments without concomitant reduction in their tumor uptake. , …”