Pre-clinical evaluation of immunoPET imaging using agonist CD40 monoclonal antibody in pancreatic tumor-bearing mice

Abstract

Background: A novel [⁶⁴Cu]Cu-NOTA-aCD40 immunoPET tracer was developed to image a CD40⁺ pancreatic tumor model in C57BL/6 mice and to study the biodistribution profile of the agonist CD40 (aCD40) monoclonal antibody (mAb) alone or combined with other mAbs.

Procedures: Copper-64 ([⁶⁴Cu]Cu) labeled NOTA-aCD40 and NOTA-IgG (10 μg; 7 MBq) were injected intravenously into C57BL/6 mice with subcutaneous mT4 tumors to assess specificity 48 h post injection (p.i.) through positron emission tomography/computed tomography (PET/CT) imaging and biodistribution studies (n = 5). [⁶⁴Cu]Cu-NOTA-aCD40 was injected alone or simultaneously in combination with a therapeutic mass of cold aCD40 (100 μg), aPD-1 (200 μg) and aCTLA-4 (200 μg) mAbs. A group of mice with or without tumor received the second round of injections 1 or 3 weeks apart, respectively. PET/CT imaging and biodistribution studies were performed at 48 h p.i. The organ dose for [⁶⁴Cu]Cu was estimated based on biodistribution studies with 2 μg [⁶⁴Cu]Cu-NOTA-aCD40 (corresponds to 5 mg patient dose) in non-tumor bearing mice.

Results: [⁶⁴Cu]Cu-NOTA-aCD40 accumulation was 2.3- and 7.8-fold higher than [⁶⁴Cu]Cu-NOTA-IgG in tumors and spleen, respectively, indicating the specificity of aCD40 mAb in a mouse pancreatic tumor model. Tumor accumulation of [⁶⁴Cu]Cu-NOTA-aCD40 was 21.2 ± 7.3%ID/g at 48 h after injection. Co-injection of [⁶⁴Cu]Cu-NOTA-aCD40 with cold aCD40 mAb alone or with PD-1 and CTLA-4 mAbs reduced both spleen and tumor uptake, whereas liver uptake was increased. With the second round of injections, the liver was the only organ with substantial uptake. With a 2 μg administered dose of [⁶⁴Cu]Cu-NOTA-aCD40 in a dosimetry study, the liver to spleen ratio was greater compared to the 10 μg dose (2.8 vs 0.37; respectively). The human equivalent for the highest dose organ (liver) was 198 ± 28.7 μSv/MBq.

Conclusions: A CD40-immunoreactive [⁶⁴Cu]Cu-NOTA-aCD40 probe was developed. The ratio of spleen to liver accumulation exceeded that of the IgG isotype and was greatest with a single small, injected mass. The safety of human patient imaging with [⁶⁴Cu]Cu was established based on extrapolation of the organ specificity to human imaging.

Keywords: CD40 monoclonal antibody; Pancreatic cancer; Positron emission tomography.