Radioactive and Fluorescent Dual Modality Cysteine Cathepsin B Activity-Based Probe for Cancer Theranostics

Abstract

Cysteine cathepsin B (CTS-B) is a crucial enzyme that is overexpressed in numerous malignancies and contributes to the invasion and metastasis of cancer. Therefore, this study sets out to develop and evaluate an activity-based multimodality theranostic agent targeting CTS-B for cancer imaging and therapy. A CTS-B activity-based probe, BMX2, was synthesized and labeled efficiently with 68Ga and 90Y to produce 68Ga-BMX2 for multimodality imaging and 90Y-BMX2 for radiation therapy. The affinity and specificity of BMX2 binding with the CTS-B enzyme were determined by fluorescent western blots using recombined active human CTS-B enzyme (rh-CTS-B) and four cancer cell lines including HeLa, HepG2, MCF7, and U87MG, with CA074 as the CTS-B inhibitor for control. Confocal laser scanning microscope imaging and cell uptake measurement were also performed. Then, in vivo PET imaging and fluorescence imaging were acquired on HeLa xenografts. Finally, the therapeutic effect of 90Y-BMX2 was tested. BMX2 could be specifically activated by rh-CTS-B and stably bound to the enzyme. The binding of BMX2 with CTS-B is time-dependent and enzyme concentration-dependent. Although CTS-B expression varied between cell lines, all showed significant uptake of BMX2 and 68Ga-BMX2. In vivo optical and PET imaging showed a high tumor uptake of BMX2 and 68Ga-BMX2 and accumulation for more than 24 h. 90Y-BMX2 could significantly inhibit HeLa tumor growth. The development of 68Ga/90Y-BMX2, a radioactive and fluorescent dual modality theranostic agent, demonstrated an effective theranostic approach for PET diagnostic imaging, fluorescence imaging, and radionuclide therapy of cancers, which may have a potential for clinical translation for cancer theranostics in the future.

Cathepsin B-Activated PET Tracer for In Vivo Tumor Imaging

“…The T/M was consistently low and remained around 1 within 60 min (Figure D). Specially, the uptake of [ 68 Ga]­NOTA-SF-CV in U87 tumors was rapid and sustained within 60 min due to the high expression of cathepsin B, which was comparable to that of 68 Ga-BMX2 . The HCT116 tumor exhibited a medium expression of cathepsin B, resulting in a corresponding decrease in the uptake as expected.…”

“…Specially, the uptake of [ 68 Ga]NOTA-SF-CV in U87 tumors was rapid and sustained within 60 min due to the high expression of cathepsin B, which was comparable to that of 68 Ga-BMX2. 28 The HCT116 tumor exhibited a medium expression of cathepsin B, resulting in a corresponding decrease in the uptake as expected. Moreover, the PC-3 tumor, characterized by low expression of cathepsin B, was hardly observed during scanning.…”

“…However, these methods usually have limitations, such as long cycles, poor sensitivity, and specificity, and the inability to detect cathepsin B activity in living tissue. Noninvasive molecular imaging techniques, such as fluorescence (FL), , PET, MRI, and photoacoustic imaging (PAI) can monitor cathepsin B activity in vivo . Among them, PET imaging has ideal intrinsic properties to image the activity of the enzyme because of its high sensitivity and deep tissue penetration, providing practical information to differentiate changes in cancer .…”