[²²⁵Ac]Ac-iPSMA Radiopharmaceuticals for Targeted Alpha Therapy: Design, Synthesis, and Preclinical Evaluation.

Abstract

Prostate-specific membrane antigen (PSMA) is a validated theranostic target with clinical relevance extending beyond prostate cancer to PSMA-expressing neovasculature in solid tumors. Despite the success of ¹⁷⁷Lu-PSMA therapy, a significant proportion of patients experience disease recurrence or partial responses, motivating the development of targeted alpha therapy (TAT) approaches. In this study, novel monomeric and dimeric ²²⁵Ac-labeled PSMA inhibitors, [²²⁵Ac]Ac-DOTA-iPSMA ([²²⁵Ac]Ac-D-iPSMA) and [²²⁵Ac]Ac-MACROPA-iPSMA ([²²⁵Ac]Ac-M-iPSMA), were designed through receptor-based computational screening, synthesized, radiolabeled, and evaluated in vitro and in vivo. Radiolabeling achieved high radiochemical purities (>96%), and saturation binding assays using lanthanum(III) surrogates confirmed cooperative nanomolar PSMA affinity. The binding affinities were Kd = 8.28 ± 0.51 nM (La-D-iPSMA) and Kd = 16.76 ± 2.51 nM (La-M-iPSMA). PSMA-specific uptake was validated in 4T1 breast cancer and HCT116 colorectal cancer cells. D-iPSMA induced superior late apoptosis (41.74% at 2 Gy), associated with reductions in GSK-3 α/β (Glycogen Synthase Kinase-3) and WNK1 (With No Lysine Kinase 1) phosphorylation, β-catenin downregulation, and dose-dependent DNA double-strand breaks (72.08% γ-H2AX [γ-phosphorylated histone]-positive cells at 6 Gy). [²²⁵Ac]Ac-M-iPSMA biodistribution in 4T1 tumor-bearing BALB/c mice revealed rapid blood clearance, a tumor-to-kidney dose ratio of 54:1, and a high tumor-absorbed dose. A combination of [¹⁷⁷Lu]Lu-D-iPSMA with [²²⁵Ac]Ac-M-iPSMA maximized reactive oxygen species generation. These findings support both conjugates as promising TAT candidates for PSMA-expressing tumors.