Bone, lymph node, and visceral metastases are frequent in castrate-resistant prostate cancer patients. Since such patients have only a few months’ survival benefit from standard therapies, there is an urgent need for new personalized therapies. The prostate-specific membrane antigen (PSMA) is overexpressed in prostate cancer and is a molecular target for imaging diagnostics and targeted radionuclide therapy (theragnostics). PSMA-targeted α therapies (PSMA-TAT) may deliver potent and local radiation more selectively to cancer cells than PSMA-targeted β⁻ therapies. In this review, we summarize both the recent preclinical and clinical advances made in the development of PSMA-TAT, as well as the availability of therapeutic α-emitting radionuclides, the development of small molecules and antibodies targeting PSMA. Lastly, we discuss the potentials, limitations, and future perspectives of PSMA-TAT.

Prostate‐specific membrane antigen (PSMA) is the most promising target for radioligand therapy of prostate cancer. The aim of this study was to prepare a small molecular ligand p‐SCN‐Bn‐TCMC‐PSMA (NG001) and compare it with the commonly used DOTA‐based PSMA‐617. The PSMA‐targeting ability of the ²¹²Pb‐labelled ligands was evaluated using PSMA‐positive C4‐2 human prostate cancer cells. Lead‐212 is an in vivo generator of alpha particles by its daughter nuclides ²¹²Bi and ²¹²Po. NG001 was synthesized by conjugating the isothiocyanato group of p‐SCN‐Bn‐TCMC to the amino group of a glutamate‐urea‐based PSMA‐binding entity. Molecular size, chelator unit and chelator linking method are different in NG001 and PSMA‐617. Both ligands were efficiently labelled with ²¹²Pb using a ²²⁴Ra/²¹²Pb‐solution generator in transient equilibrium with progeny. Lead‐212‐labelled NG001 was purified with a yield of 85.9±4.7% and with 0.7±0.2% of ²²⁴Ra. Compared with [²¹²Pb]Pb‐PSMA‐617, [²¹²Pb]Pb‐NG001 displayed a similar binding and internalization in C4‐2 cells, with comparable tumour uptake in mice bearing C4‐2 tumours, but almost a 2.5‐fold lower kidney uptake. Due to the rapid normal tissue clearance and tumour cell internalization, any significant translocalization of ²¹²Bi was not detected in mice. In conclusion, the obtained results warrant further preclinical studies to evaluate the therapeutic efficacy of [²¹²Pb]Pb‐NG001.

Radioligand therapy targeting the prostate-specific membrane antigen (PSMA) is rapidly evolving as a promising treatment for metastatic castration-resistant prostate cancer. The PSMA-targeting ligand p-SCN-Bn-TCMC-PSMA (NG001) labelled with ²¹²Pb efficiently targets PSMA-positive cells in vitro and in vivo. The aim of this preclinical study was to evaluate the therapeutic potential of ²¹²Pb-NG001 in multicellular tumour spheroid and mouse models of prostate cancer. The cytotoxic effect of ²¹²Pb-NG001 was tested in human prostate C4-2 spheroids. Biodistribution at various time points and therapeutic effects of different activities of the radioligand were investigated in male athymic nude mice bearing C4-2 tumours, while long-term toxicity was studied in immunocompetent BALB/c mice. The radioligand induced a selective cytotoxic effect in spheroids at activity concentrations of 3–10 kBq/mL. In mice, the radioligand accumulated rapidly in tumours and was retained over 24 h, while it rapidly cleared from nontargeted tissues. Treatment with 0.25, 0.30 or 0.40 MBq of ²¹²Pb-NG001 significantly inhibited tumour growth and improved median survival with therapeutic indexes of 1.5, 2.3 and 2.7, respectively. In BALB/c mice, no signs of long-term radiation toxicity were observed at activities of 0.05 and 0.33 MBq. The obtained results warrant clinical studies to evaluate the biodistribution, therapeutic efficacy and toxicity of ²¹²Pb-NG001.

Background: New treatments combating bone and extraskeletal metastases are needed for patients with metastatic castration-resistant prostate cancer. The majority of metastases overexpress prostate-specific membrane antigen (PSMA), making it an ideal candidate for targeted radionuclide therapy. Objective: The aim of this study was to test a novel liquid 224Ra/212Pb-generator for the rapid preparation of a dual-alpha targeting solution. Here, PSMA-targeting ligands are labelled with 212Pb in the 224Ra-solution in transient equilibrium with daughter nuclides. Thus, natural bone-seeking 224Ra targeting sclerotic bone metastases and 212Pb-chelated PSMA ligands targeting PSMA-expressing tumour cells are obtained. Methods: Two PSMA-targeting ligands, the p-SCN-Bn-TCMC-PSMA ligand (NG001), specifically developed for chelating 212Pb, and the most clinically used DOTA-based PSMA-617 were labelled with 212Pb. Radiolabelling and targeting potential were investigated in situ, in vitro (PSMA-positive C4-2 human prostate cancer cells) and in vivo (athymic mice bearing C4-2 xenografts). Results: NG001 was rapidly labelled with 212Pb (radiochemical purity >94% at concentrations of ≥15 μg/ml) using the liquid 224Ra/212Pb-generator. The high radiochemical purity and stability of [212Pb]Pb- NG001 were demonstrated over 48 hours in the presence of ascorbic acid and albumin. Similar binding abilities of the 212Pb-labelled ligands were observed in C4-2 cells. The PSMA ligands displayed comparable tumour uptake after 2 hours, but NG001 showed a 3.5-fold lower kidney uptake than PSMA- 617. Radium-224 was not chelated and, hence, showed high uptake in bones. Conclusion: A fast method for the labelling of PSMA ligands with 212Pb in the 224Ra/212Pb-solution was developed. Thus, further in vivo studies with dual tumour targeting by alpha-particles are warranted.

This study aimed to determine the influence of cellular PSMA expression, radioligand binding and internalization, and repeated administrations on the therapeutic effects of the PSMA-targeting radioligand ²¹²Pb-NG001. Cellular binding and internalization, cytotoxicity, biodistribution, and the therapeutic efficacy of ²¹²Pb-NG001 were investigated in two human prostate cancer cell lines with different PSMA levels: C4-2 (PSMA+) and PC-3 PIP (PSMA+++). Despite 10-fold higher PSMA expression on PC-3 PIP cells, cytotoxicity and therapeutic efficacy of the radioligand was only 1.8-fold better than for the C4-2 model, possibly explained by lower cellular internalization and less blood-rich stroma in PC-3 PIP xenografts. Mice bearing subcutaneous PC-3 PIP xenografts were treated with 0.2, 0.4, and 0.8 MBq of ²¹²Pb-NG001 that resulted in therapeutic indexes of 2.7, 3.0, and 3.5, respectively. A significant increase in treatment response was observed in mice that received repeated injections compared to the corresponding single dose (therapeutic indexes of 3.6 for 2 × 0.2 MBq and 4.4 for 2 × 0.4 MBq). The results indicate that ²¹²Pb-NG001 can induce therapeutic effects at clinically transferrable doses, both in the C4-2 model that resembles solid tumors and micrometastases with natural PSMA expression and in the PC-3 PIP model that mimics poorly vascularized metastases.

The alpha-emitting radioligand [²¹²Pb]Pb-AB001, targeting prostate-specific membrane antigen (PSMA), is a promising therapy approach for prostate cancer. Bromodomain and extra-terminal (BET) protein inhibitors, such as AZD5153 and JQ1, disrupt oncogenic transcriptional programs by altering chromatin structure. This study evaluated whether BET inhibition enhances the efficacy of radioligand therapy. Cytotoxic effects of [²¹²Pb]Pb-AB001 alone and in combination with BET inhibitors were assessed in 2D monolayers and a 3D spheroid model of PSMA-positive C4-2 prostate cancer cells. Cell viability, cell cycle alterations, and DNA damage were assessed using viability assays and flow cytometry. Spheroid growth and viability were assessed by fluorescence microscopy. AZD5153 was more potent than JQ1 in reducing cell proliferation. [²¹²Pb]Pb-AB001 induced activity- and time-dependent cytotoxicity with a delayed apoptotic response. BET inhibitors induced G1 arrest, while [²¹²Pb]Pb-AB001 caused G2 arrest. Combination treatment reduced cell viability in an additive manner but did not further affect cell cycle distribution or increase apoptosis compared with [²¹²Pb]Pb-AB001 alone. γH2AX staining in 2D models showed an activity- and time-dependent increase in DNA damage at 1, 3 and 6 days post-treatment with [²¹²Pb]Pb-AB001. BET inhibitors alone induced minimal γH2AX, and combination treatments did not enhance DNA damage beyond [²¹²Pb]Pb-AB001 alone. In 3D spheroids, combination treatment led to synergistic growth suppression. In conclusion, these findings indicate that therapeutic inhibition of BET bromodomain in combination with the alpha-emitting radioligand [²¹²Pb]Pb-AB001 could significantly enhance tumour control and should be further evaluated in metastatic prostate cancer models.