In this report, we describe the initial demonstration of hu3F8-C825 for PRIT of GD2(+)-NB. On the basis of these reports, as well as our aforementioned experience pretargeting GD2(+)-NB, we reasoned that anti–GD2-C825 could offer a viable clinical developmental path for PRIT directed at GD2(+)-NB. The IgG-scFv BsAbs were sufficiently functional and stable in vivo to permit highly efficient tumor targeting of the 177Lu-DOTA hapten in mouse xenograft models of human adenocarcinoma (e.g., tumor-to-tissue uptake ratios of >450 for blood and >20 for kidneys ref. Specifically intended for PRIT, multiple IgG-scFv BsAb were developed consisting of an IgG with specificity to a cancer cell surface target linked to C825 scFv at the C-terminus of the IgG light chains as an IgG-scFv format ( 6). Orcutt and colleagues subsequently affinity matured the 2D12.5 sequence to yield a novel scFv (“C825”) with pmol/L affinity with improved dissociation half-time of the antibody–DOTA complex (from 5.5 minutes to ∼5 hours ref. The mAb 2D12.5 has nmol/L affinity for low molecular weight (MW) 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) complexes with yttrium (Y) and lutetium (Lu), and is well suited for PRIT in vivo ( 4). Over two decades ago, Reardan and colleagues pioneered a PRIT system using antibodies against metal chelates ( 3). ©2014 AACR.Īs an alternative to streptavidin/biotin, other high-affinity antibody–hapten pairs have been developed for PRIT ( 2). We conclude that this novel anti-GD2 PRIT approach has sufficient TI to successfully ablate subcutaneous GD2(+)-NB in mice while sparing kidney and bone marrow. All nontreated mice required sacrifice within 12 days (>1.0-cm 3 tumor volume). Tumor ablation was confirmed histologically in 4 of 5 mice, and normal organs showed minimal overall toxicities. A therapy study ( n = 5/group tumor volume, 240 ± 160 mm 3) with three successive PRIT cycles (total 177Lu: ∼33 MBq tumor dose ∼3,400 cGy), revealed complete tumor response in 5 of 5 animals, with no recurrence up to 28 days after treatment. Absorbed doses for tumor and normal tissues were approximately 85 cGy/MBq and ≤3.7 cGy/MBq, respectively, with therapeutic indices (TI) of 142 for blood and 23 for kidney. A three-step regimen, including hu3F8-C825, a dextran-based clearing agent, and p-aminobenzyl-DOTA radiolabeled with 177Lu (as 177Lu-DOTA-Bn t 1/2 = 6.71 days), was optimized in immunocompromised mice carrying subcutaneous human GD2(+) neuroblastoma (NB) xenografts. For this purpose, an IgG-scFv BsAb was engineered using the sequences for the anti-GD2 humanized monoclonal antibody hu3F8 and C825, a murine scFv antibody with high affinity for the chelator 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) complexed with β-particle–emitting radiometals such as 177Lu and 90Y. We sought to overcome key PRIT limitations such as high renal radiation exposure and immunogenicity (e.g., of streptavidin–antibody fusions), to advance clinical translation of this PRIT strategy for diasialoganglioside GD2-positive tumors. Bispecific antibodies (BsAb) have proven to be useful targeting vectors for pretargeted radioimmunotherapy (PRIT).