BIOLOGICAL ASSESSMENT OF [¹⁶¹TB]TB-PSMA IN HUMAN PROSTATE CANCER CELL LINES

Abstract

Introduction

Prostate-specific membrane antigen (PSMA) is highly overexpressed in prostate cancer and represents a validated target for both diagnostic imaging and radionuclide therapy. Radioligand therapy using PSMA-targeting compounds labeled with beta-emitting radionuclides, such as Lutetium-177, has shown promising clinical results. However, alternative radionuclides with more favorable decay properties may further enhance therapeutic efficacy. Terbium-161 is an attractive candidate due to its emission of low-energy beta particles and conversion and Auger electrons, which may improve tumor cell killing, particularly at the cellular and subcellular levels. Taken together, these characteristics highlight Terbium-161 as an attractive candidate for the treatment of disseminated cancers with multiple metastases, from single tumor cells (∼10 µm) to micrometastatic clusters (<1 mm).

Objective

The objective of this study was to perform a biological assessment of [¹⁶¹Tb]Tb-PSMA in human prostate cancer cell lines, focusing on its cellular uptake, specificity, internalization behavior, and radiobiological effects, in order to evaluate its potential as a targeted radiotherapeutic agent. Also, we provide a cellular comparison of Terbium-161 PSMA radiobiological effects with those induced by Actinium-225 PSMA and Lutetium-177 PSMA.

Materials and Methods

All radiopharmaceuticals were radiolabeled under optimized conditions and its radiochemical purity was confirmed prior to biological evaluation. In vitro experiments were conducted using PSMA-positive (LNCaP) and PSMA-negative (PC3) human prostate cancer cell lines. Cellular uptake assays with [¹⁶¹Tb]Tb-PSMA were performed at different time points to assess PSMA-specific binding, with or without blocking using an excess of unlabeled PSMA ligand. The radiobiological effects of [¹⁶¹Tb]Tb-PSMA were evaluated through cell viability, cell growth and clonogenic survival assays following 24h incubation with increasing activity concentrations (0.01‒10 MBq/mL) plus 7 days in fresh media. Experiments were performed under standardized conditions to allow comparative analysis between Terbium-161 PSMA cell effects and Actinium-225 and Lutetium-177 PSMA. Statistical analysis was performed using GraphPad 9.5.1, with significance set at p = 0.05.

Results

[¹⁶¹Tb]Tb-PSMA showed high and specific uptake in PSMA-expressing prostate cancer cells, which was significantly reduced by PSMA blocking, confirming PSMA-mediated binding. Uptake increased over time in PSMA-positive cells, with negligible uptake in PSMA-negative controls. Increasing activities of [¹^6¹Tb]Tb-PSMA significantly reduced cell viability and clonogenic survival in PSMA-expressing cells, with minimal effects in PSMA-negative lines. Compared with [²²5Ac]Ac-PSMA, which induced stronger cytotoxicity at lower activities, [¹⁶¹Tb]Tb-PSMA produced an activity-dependent radiotoxic effect, comparable to that observed with [¹⁷⁷Lu]Lu-PSMA.

Conclusions

The biological evaluation of [¹⁶¹Tb]Tb-PSMA in human prostate cancer cell lines demonstrates high PSMA specificity, efficient uptake, and significant radiobiological effects in PSMA-expressing cells, supporting its potential for targeted radionuclide therapy of prostate cancer. Further preclinical studies, including in vivo evaluation, are needed to confirm these findings and assess the translational potential of [¹⁶¹Tb]Tb-PSMA.