The sponsor developed a novel antibody fragment against a tumor-specific target that is present in several cancer types. They aimed to develop a radiopharmaceutical for the treatment of these tumors. TRACER assisted in the preclinical development by labeling the antibody fragment for imaging in tumor-bearing mice. One batch was labeled with Indium-111 and another with Zirconium-89 for Single Photon Emission Computed Tomography (SPECT) and Positron Emission Tomography (PET), respectively. In vitro evaluation showed binding of the antibody fragment to the target-expressing tumor cells. The rodent study resulted in clear images of biodistribution and target binding in vivo for both modalities.
Labeling
As a first development step, a labeling strategy was developed, and the antibody fragment was conjugated with a chelator to enable radiometal labeling. Various ratios of chelator:antibody were tested in vitro, and the optimal conjugate was selected for in vivo studies. The radiolabeling with 111In and 89Zr was set up and optimized, followed by quality control, purification, and in vitro stability studies. Two nuclides were selected for comparison of imaging quality and stability in subsequent research.
111In and 89Zr labeling and biodistribution studies
In preclinical studies, 111In labeling is often used as a starting point because it is faster and less expensive. It delivers biodistribution data early. 89Zr labeling can be more difficult and time-consuming. In this project, the preferred 89Zr labeling was developed in parallel, and after the first 111In biodistribution study was completed, the 89Zr biodistribution study could commence together with the second 111In study.
Preclinical studies
TRACER performed preclinical in vitro testing in tumor cell lines in binding and internalization assays, and in vivo in tumor-bearing mice.
- In vitro stability in plasma.
- In vitro binding characteristic and internalization assays.
- Biodistribution by ex vivo gamma counting in tumor-bearing mice.
- In vivo small animal PET and SPECT study (image quality, biodistribution, and tumor targeting and binding).
Results
The antibody fragment was successfully labeled with 111In and 89Zr, resulting in a good yield, purity, and molar activity. In vitro, the radiolabeled antibody fragment was stable and showed efficient binding. The compound allowed PET and SPECT imaging in rodents. Biodistribution studies showed high uptake in tumors and low accumulation in non-target organs. PET and SPECT imaging confirmed the previously generated biodistribution results. Additionally, the images showed accumulation in salivary glands, which was not included in the sponsor’s original biodistribution study.
Without PET and SPECT imaging, uptake in organs that are not included for excision in routine biodistribution could be missed. Imaging data adds significant insights into the path to clinical translation.
Future prospects
From this study, the sponsor was able to compare PET vs SPECT as imaging methods. The results were used for insights into the potential development of the antibody fragment as an antibody drug conjugate (ADC) or radiopharmaceutical, which the study data showed are both viable options.