Effect of Modulating FcRn Binding on Direct and Pretargeted Tumor Uptake of Full-length Antibodies

Abstract

Full-length antibodies lack ideal pharmacokinetic properties for rapid targeted imaging, prompting the pursuit of smaller peptides and fragments. Nevertheless, studying the disposition properties of antibody-based imaging agents can provide critical insight into the pharmacology of their therapeutic counterparts, particularly for those coupled with potent payloads. Here, we evaluate modulation of binding to the neonatal Fc receptor (FcRn) as a protein engineering-based pharmacological strategy to minimize the overall blood pool background with directly labeled antibodies and undesirable systemic click reaction of radiolabeled tetrazine with circulating pretargeted trans-cyclooctene (TCO)-modified antibodies. Noninvasive SPECT imaging of mice bearing HER2-expressing xenografts was performed both directly (111 In-labeled antibody) and indirectly (pretargeted TCO-modified antibody followed by 111 In-labeled tetrazine). Pharmacokinetic modulation of antibodies was achieved by two distinct methods: Fc engineering to reduce binding affinity to FcRn, and delayed administration of an antibody that competes with binding to FcRn. Tumor imaging with directly labeled antibodies was feasible in the absence of FcRn binding, rapidly attaining high tumor-to-blood ratios, but accompanied by moderate liver and spleen uptake. Pretargeted imaging of tumors with non-FcRn-binding antibody was also feasible, but systemic click reaction still occurred, albeit at lower levels than with parental antibody. Our findings demonstrate that FcRn binding impairment of full-length IgG antibodies moderately lowers tumor accumulation of radioactivity, and shifts background activity from blood pool to liver and spleen. Furthermore, reduction of FcRn binding did not eliminate systemic click reaction, but yielded greater improvements in tumor-to-blood ratio when imaging with directly labeled antibodies than with pretargeting.

Click Chemistry and Radiochemistry: An Update

“…In), 203−212 β + -emitters (i.e., 68 Ga, 18 F, 64 Cu, 44 Sc, and 89 Zr), 135,213−231 β − -emitters (i.e., 177 Lu and 67 Cu), 232−237 and α-emitters (i.e., 212 Pb, 211 At, and 225 Ac). 238,239 While the earliest reports on IEDDA-based pretargeting focused primarily on proof-of-concept, a significant body of recent work has been dedicated to optimizing this approach to imaging and therapy.…”

“…Because TCO exhibits greater in vivo stability than Tz, the overwhelming majority of IEDDA-based pretargeting strategies employ a TCO-modified immunoconjugate alongside a Tz-bearing radioligand (Figure ). Since our last review, a remarkable surge in the field has yielded pretargeting systems featuring radioligands labeled with gamma-emitters (i.e., 99m Tc and 111 In), − β + -emitters (i.e., 68 Ga, 18 F, 64 Cu, 44 Sc, and 89 Zr), ,− β – -emitters (i.e., 177 Lu and 67 Cu), − and α-emitters (i.e., 212 Pb, 211 At, and 225 Ac). , …”

“…One particularly exciting way of harnessing the CuAAC ligation for radiolabeling stands apart from both methods discussed thus far. In 2019, Denk et al reported the creation of compounds containing 211 At-labeled 1,2,3-triazole rings by performing CuAAC conjugations in the presence of the α particle-emitting radiohalogen (Figure 6). 80 The investigators hypothesize that 211 At(I) is formed via oxidation by Cu(II) during the CuAAC reaction and that this astatonium cation undergoes electrophilic substitution with the triazole ring to provide the 5-[ 211 At]astato-1,2,3-triazole product.…”

Antibody Engineering for Nuclear Imaging and Radioimmunotherapy

“…Several laboratories have sought to improve the behavior of mAb-based radioimmunoconjugates by attenuating or enhancing Fc receptor binding. Nazarova et al explored immunoSPECT imaging using a HER2-targeting 111 In-labeled mAb bearing a pair of histidine mutations-H310A and H435Q-that abrogated FcRn binding (25). The double mutant displayed a more rapid pharmacokinetic profile and greater tumor-to-blood activity concentration ratios than its wild-type parent but at the cost of reduced tumoral uptake and lower tumor-to-spleen and tumor-to-liver contrast (Fig.…”

In Vivo Pretargeting Based on Cysteine-Selective Antibody Modification with IEDDA Bioorthogonal Handles for Click Chemistry

“…Herein, a novel carbonylacrylic derivative functionalized with a trans -cyclooctene (TCO) moiety was developed and conjugated to THIOMAB LC-V205C, a modified trastuzumab antibody against the human epidermal growth factor receptor 2 (HER2) presenting an engineered cysteine in the light chain . This receptor has already been exploited in clinical imaging of breast and gastric cancer HER2-expressing tumors. ,,,− Pretargeting was first explored in vitro to assess the IEDDA reaction between the antibody bound to HER2-expressing cells and a fluorescent/radiolabeled tetrazine. Then, in vivo studies were performed in a relevant HER2 + mouse model using an 111 In-labeled tetrazine.…”