ImmunoPET of tissue factor expression in triple-negative breast cancer with a radiolabeled antibody Fab fragment

Abstract

Purpose To date, there is no effective therapy for triple-negative breast cancer (TNBC), which has dismal clinical outcome. Upregulation of tissue factor (TF) expression leads to increased patient morbidity and mortality in many solid tumor types, including TNBC. Our goal is to employ the Fab fragment of ALT-836, a chimeric anti-human TF mAb, for PET imaging of TNBC, which can be used to guide future TNBC therapy. Methods ALT-836-Fab was generated by enzymatic papain digestion. SDS-PAGE and FACS studies were performed to evaluate the integrity and TF binding affinity of ALT-836-Fab before NOTA conjugation and 64Cu-labeling. Serial PET imaging and biodistribution studies were carried out to evaluate the tumor targeting efficacy and pharmacokinetics in MDA-MB-231 TNBC model, which expresses high level of TF on the tumor cells. Blocking studies, histological assessment, as well as RT-PCR was performed to confirm TF specificity of 64Cu-NOTA-ALT-836-Fab. Results ALT-836-Fab was produced with high purity, which exhibited superb TF binding affinity and specificity. Serial PET imaging revealed rapid and persistent tumor uptake of 64Cu-NOTA-ALT-836-Fab (5.1 ± 0.5 %ID/g at 24 h post-injection; n = 4) and high tumor/muscle ratio (7.0 ± 1.2 at 24 h post-injection; n = 4), several-fold higher than that of the blocking group and tumor models that do not express significant level of TF, which was confirmed by biodistribution studies. TF specificity of the tracer was also validated by histology and RT-PCR. Conclusions 64Cu-NOTA-ALT-836-Fab exhibited prominent tissue factor targeting efficiency in MDA-MB-231 TNBC model. The use of a Fab fragment led to fast tumor uptake and good tissue/muscle ratio, which may be translated into same-day immunoPET imaging in the clinical setting to improve TNBC patient management.

PET Imaging of Tissue Factor in Pancreatic Cancer Using 64Cu-Labeled Active Site–Inhibited Factor VII

“…Despite the impressive tumor uptake, the tumor-to-muscle ratio was similar to what we report because of the long circulation of full-length antibodies. The tumor uptake reported for the antibody Fab fragment was slightly higher, whereas the tumor-to-muscle ratio was about 50% lower than our results (29).…”

“…PET imaging of TF has previously been investigated using a 64 Cu-labeled TF antibody and its Fab fragment (28,29). A superior tumor uptake of 16.5 6 2.6 %ID/g 48 h after injection of the 64 Culabeled full-length antibody was reported (28).…”

Quantitative PET Imaging of Tissue Factor Expression Using 18F-Labeled Active Site–Inhibited Factor VII

“…The faster pharmacokinetics possibly limit 18 F-FVIIai tumor binding because of low availability in the circulating blood. Recently, Shi et al published the results on the antibody Fab fragment of ALT-836 labeled with 64 Cu for PET imaging of TF in a TF-positive triple-negative breast cancer xenograft mouse model (24). The reported maximum uptake in TF-positive tumors was 5.1 %ID/g, which was, hence, significantly lower than previously reported with the full-length antibody.…”

In Vivo Molecular Imaging for Biomedical Analysis and Therapies

“…32,33 Antibody fragments are used for specific imaging over short time periods since the blood half-lives are shorter for the fragments. 34 Shorter half-life radionuclides are used for this purpose, such as 64 Cu (half-life: 12.7 h) for PET 35,36 and 99m Tc (half-life: 6.02 h) for SPECT. 37,38 Recently, antibody imaging has been applied for prediction of the therapeutic effect of immune checkpoint inhibitors.…”