Positron emission tomography (PET) imaging of neuroblastoma and melanoma with 64Cu-SarAr immunoconjugates

Abstract

The advancement of positron emission tomography (PET) depends on the development of new radiotracers that will complement (18)F-FDG. Copper-64 ((64)Cu) is a promising PET radionuclide, particularly for antibody-targeted imaging, but the high in vivo lability of conventional chelates has limited its clinical application. The objective of this work was to evaluate the novel chelating agent SarAr (1-N-(4-aminobenzyl)-3, 6,10,13,16,19-hexaazabicyclo[6.6.6] eicosane-1,8-diamine) for use in developing a new class of tumor-specific (64)Cu radiopharmaceuticals for imaging neuroblastoma and melanoma. The anti-GD2 monoclonal antibody (mAb) 14.G2a, and its chimeric derivative, ch14.18, target disialogangliosides that are overexpressed on neuroblastoma and melanoma. Both mAbs were conjugated to SarAr using carbodiimide coupling. Radiolabeling with (64)Cu resulted in >95% of the (64)Cu being chelated by the immunoconjugate. Specific activities of at least 10 microCi/microg (1 Ci = 37 GBq) were routinely achieved, and no additional purification was required after (64)Cu labeling. Solid-phase radioimmunoassays and intact cell-binding assays confirmed retention of bioactivity. Biodistribution studies in athymic nude mice bearing s.c. neuroblastoma (IMR-6, NMB-7) and melanoma (M21) xenografts showed that 15-20% of the injected dose per gram accumulated in the tumor at 24 hours after injection, and only 5-10% of the injected dose accumulated in the liver, a lower value than typically seen with other chelators. Uptake by a GD2-negative tumor xenograft was significantly lower (<5% injected dose per gram). MicroPET imaging confirmed significant uptake of the tracer in GD-2-positive tumors, with minimal uptake in GD-2-negative tumors and nontarget tissues such as liver. The (64)Cu-SarAr-mAb system described here is potentially applicable to (64)Cu-PET imaging with a broad range of antibody or peptide-based imaging agents.

Fig. 1.

Structure of SarAr. SarAr is based on the macrobicyclic cage diamsar and was modified to contain the reactive aminobenzyl group.

Fig. 2.

Lindmo plot of GD2 binding data, confirming retention of immunoreactivity. Immunoreactive fraction of 70% was obtained [inverse y intercept (1/b = 1/1.4356)] by using a fixed concentration of labeled ch14.18 and increasing concentrations of GD2.

Fig. 3.

In vivo biodistribution of [⁶⁴Cu]ch14.18 in neuroblastoma and melanoma xenograft-bearing animals. Data from athymic nude mice bearing s.c. M21 melanoma (A) or IMR6 neuroblastoma (B) human tumor xenografts are shown.

Fig. 4.

MicroPET imaging of GD2-positive human tumor xenografts. Athymic nude mice bearing subcutaneous GD2-positive human tumor xenografts IMR6 neuroblastoma and M21 melanoma (A) and M21 melanoma (B) were injected with ⁶⁴Cu-labeled anti-GD2 mAb ch14.18 (A and B) or 14.G2a (B) and imaged. Control GD-2-negative PC-3 prostate carcinoma xenograft, treated and imaged similarly, is shown in a. Arrows point to flank tumors.