89Zr anti-CD44 immuno-PET monitors CD44 expression on splenic myeloid cells and HT29 colon cancer cells

Abstract

CD44 is a cell-surface glycoprotein involved in cell-cell interaction, adhesion, and migration. CD44 is found on colon cancer cells and on immune cells. Previous studies of ⁸⁹Zr PET imaging of CD44 have relied on an anti-human antibody (Ab), which can influence biodistribution in murine models. In this study, we used an Ab that cross-reacts with both human and mouse origin CD44 of all isoforms to unveil the type of leukocyte responsible for high splenic anti-CD44 uptake and investigate how its regulation can influence tumor immuno-PET. The Ab was site-specifically labeled with ⁸⁹Zr-deferoxamine on cysteine residues. ⁸⁹Zr-anti-CD44 demonstrated high-specific binding to HT29 human colon cancer cells and monocytic cells that showed CD44 expression. When ⁸⁹Zr-anti-CD44 was administered to Balb/C nude mice, there was remarkably high splenic uptake but low SNU-C5 tumor uptake (1.2 ± 0.7%ID/g). Among cells isolated from Balb/C mouse spleen, there was greater CD44 expression on CD11b positive myeloid cells than lymphocytes. In cultured monocytic and macrophage cells, LPS stimulation upregulated CD44 expression and increased ⁸⁹Zr-anti-CD44 binding. Similarly, normal Balb/C mice that underwent lipopolysaccharide (LPS) stimulation showed a significant upregulation of CD44 expression on splenic myeloid cells. Furthermore, LPS treatment stimulated a 2.44-fold increase of ⁸⁹Zr-anti-CD44 accumulation in the spleen, which was attributable to splenic myeloid cells. Finally, in Balb/C nude mice bearing HT29 tumors, we injected ⁸⁹Zr-anti-CD44 with greater Ab doses to reduce binding to splenic cells. The results showed lower spleen uptake and improved tumor uptake (2.9 ± 1.3%ID/g) with a total of 300 μg of Ab dose, and further reduction of spleen uptake and greater tumor uptake (5.7 ± 0.0%ID/g) with 700 μg Ab dose. Thus, using an ⁸⁹Zr labeled Ab that cross-reacts with both human and mouse CD44, we demonstrate that CD44 immuno-PET has the capacity to monitor CD44 regulation on splenic myeloid cells and may also be useful for imaging colon tumors.