Conclusion
Our findings suggest that the deglycosylation of antibodies represents a facile strategy for improving the quality of immuno-PET in animal models as well as in certain patient populations.
Methods
Since the heavy chain glycans influence the affinity of FcγR for the Fc domain, we set out to investigate whether radioimmunoconjugates with truncated glycans would exhibit altered binding to FcγRI and, in turn, improved in vivo performance. Using the HER2-targeting antibody trastuzumab, we synthesized a series of desferrioxamine-bearing immunoconjugates with differing glycosylation states and interrogated their FcγRI binding via surface plasmon resonance, enzyme-linked immunosorbent assay, and flow cytometry. Furthermore, we labeled these immunoconjugates with 89Zr and explored their biodistribution in athymic nude, NSG, and humanized NSG mice bearing human epidermal growth factor receptor 2-expressing human breast cancer xenografts.
Results
We observed a strong correlation between the impaired in vitro FcγRI binding of deglycosylated immunoconjugates and significant decreases in the in vivo off-target uptake of the corresponding 89Zr-labeled radioimmunoconjugates (i.e., liver activity concentrations are reduced by ∼3.5-fold in humanized NSG mice). These reductions in off-target uptake were accompanied by concomitant increases in the tumoral activity concentrations of the glycoengineered radioimmunoconjugates, ultimately yielding improved tumor-to-healthy organ contrast and higher quality PET images.