Removal of Fc Glycans from [89Zr]Zr-DFO-Anti-CD8 Prevents Peripheral Depletion of CD8+ T Cells.

摘要: The N-linked biantennary glycans on the heavy chain of immunoglobulin G (IgG) antibodies (mAbs) are instrumental in recognition Fc region by Fc-gamma receptors (FcγR). In case full-length mAb-based imaging tracers targeting immune cell populations, these Fc:FcγR interactions can potentially deplete effector cells responsible for tumor clearance. To bypass this problem, we hypothesize that enzymatic removal will disrupt and spare tracer-targeted from depletion during immunopositron emission tomography (immunoPET) imaging. Herein, compared vitro vivo properties 89Zr-radiolabeled CD8-specific murine mAb (anti-CD8wt, clone 2.43), a well-known depleting mAb, its deglycosylated counterpart (anti-CD8degly). Deglycosylation was achieved via treatment with peptide: N-glycosidase F (PNGaseF). Both anti-CD8wt anti-CD8degly mAbs were conjugated to p-SCN-Bn-desferrioxamine (DFO) labeled 89Zr. Bindings both DFO-conjugated FcγR CD8+ splenocytes compared. distribution studies conducted examine specificity pharmacokinetics radioimmunoconjugates tumor-naive CT26 colorectal tumor-bearing mice. Ex analysis T population spleens tumors obtained postimaging measured flow cytometry qRT-PCR. confirmed SDS-PAGE. A reduction interaction exhibited DFO-anti-CD8degly, while binding CD8 remained unchanged. Tissue showed similar [89Zr]Zr-DFO-anti-CD8degly wt radioimmunoconjugate. blocking further demonstrated retained radiotracer CD8. From studies, no difference accumulation observed between radiotracers. Results mice administered DFO-anti-CD8wt, whereas an increase shown DFO-anti-CD8degly. No statistically significant infiltrating cohorts probes when control unmodulated mRNA levels excised increased transcripts antigen imaged [89Zr]Zr-DFO-anti-CD8wt. conclusion, offers straightforward approach develop full length antibody-based specifically detecting molecules consequential their target peripheral tissues.