State of the art and future perspectives of new radinuclides in Nuclear Medicine: Part III

Abstract

In this third installment of the continuing education series, the clinical and therapeutic applications of zirconium, astatine and thorium are analyzed in depth. Although they are not described as theragnostic pairs, each of these radionuclides plays a fundamental role in precision medicine, which is rapidly advancing within Nuclear Medicine.

We begin by analyzing zirconium-89, a positron emitter whose long half-life makes it particularly suitable for labeling large molecules with slow kinetics, such as antibodies, playing a crucial role in immunotherapy. The use of astatine-211, an alpha-emitting radionuclide with a simple decay scheme and chemical behavior similar to iodine, is also discussed. Its main challenge lies in its production, as it requires cyclotrons capable of generating highly energetic alpha particle beams. Furthermore, thorium-227, a 100% alpha emitter, is reviewed. This radionuclide exhibits excellent chelation properties, enabling its conjugation with tumor-targeting molecules to produce thorium-labeled conjugates. While this technique is yielding promising preclinical results, the use of thorium faces challenges, including the potential separation of radium-223 from the molecule and the dependence of activity measurements on the time of production. Since it takes 100 days to reach equilibrium, activity assessment is based on photons emitted by its daughter radionuclides.

Despite these challenges, these radionuclides are driving the evolution of precision medicine, expanding therapeutic and diagnostic possibilities within Nuclear Medicine.

Resumen

En esta tercera entrega de la serie de formación continuada, se analizan en profundidad las aplicaciones clínicas y terapéuticas del zirconio, astato y torio. Aunque no se describen como pares teragnósticos, pero cado uno de los radionúclidos descritos está jugando un papel fundamental dentro de la medicina de precisión que evoluciona dando pasos agigantados en la Medicina Nuclear. Empezamos analizando el zirconio-89, emisor de positrones, cuya vida media larga le hace interesante para unirlo a moléculas grandes con cinética lenta, como anticuerpos, teniendo un papel fundamental en la inmunoterapia. Se describe el uso del radioisótopo astato-211, que presenta un esquema de desintegración alfa sencillo y comportamiento químico similar al yodo, el principal reto es la producción necesitándose ciclotrones que produzcan haces de partículas alfa muy energéticos. Asimismo, se revisa el torio-227, un emisor alfa (100%) que, tiene gran facilidad para ser quelado y permite el radiomarcaje de partes dirigidas al tumor para producir conjugados de torio dirigidos. Esta técnica está mostrando resultados preclínicos muy interesantes, pero el uso del torio tiene la limitación de la posible separación del radio-223 de la molécula y la medida en el activímetro depende del momento de producción, ya que tarda 100 días en llegar al equilibrio y las medidas de actividad se basan en fotones provenientes de los radionúclidos hijo.

A pesar de los retos a los que nos enfrentamos, estos radionúclidos están impulsando la evolución de la medicina de precisión y ampliando las posibilidades terapéuticas y diagnósticas en Medicina Nuclear.

Introduction

The great advances in the development of new molecules that are increasingly more specific and selective for determined tumors has led to the search for optimal radioisotopes in regard to their chemical (improvement in binding) and radioactive characteristics (improvement in doses) to achieve radiopharmaceuticals that are more adapted to the diagnosis and treatment of different diseases. These advances have inspired this review, which has been prepared in three parts and discusses the most relevant theragnostic pairs (first part), a family of radionuclides, such as terbium, and radionuclides emitting alpha particles with high clinical potential (second part), and in the third part, we discuss zirconium (which is the most relevant diagnostic radionuclide for the study of immunotherapy) and we finish with the alpha-emitting particles (astatine and thorium). As in the first and second parts, we will focus on providing the most relevant details from a physical and chemical point of view as well as how to obtain these radionuclides and the possible difficulties that emerge with their use in the routine clinical setting. We end with each radionuclide, including the most relevant clinical and preclinical studies and clinical trials in which these radionuclides have been used.

As an example of the current relevance of targeted treatments with alpha particles, of which we have shown a number of radionuclides, we complete the introduction of this last part by referencing a very recent review that provides a map of alpha-emitting particle production,¹ that is shown in Fig. 1.

Figure 1.

Distribution of the centers around the world producing alpha particle emitters.¹

This series has been made with the aim of providing this information to investigators, clinicians, and the industry to facilitate and initiate the collaborations necessary to integrate treatments with alpha particles.

Zirconium-89

Physical characteristics and processes for obtaining zirconium-89 radionuclides for clinical application

Zirconium-89 (⁸⁹Zr) has a half-life of 3.3 days and it decays via β+ (22.3%) and electron capture (76.6%) to yttrium-89 m (^89mY),² In turn, this decays, emitting a gamma photon to stable ^89mY (Fig. 2). Its long half-life allows its use for acquiring positron emission tomography (PET) images using large molecules with slow kinetics, such as antibodies. The acquisitions can be made for up to one week with conventional PET equipment or for up to 30 days with whole body PET equipment.^3,4 The photon emitted in the gamma decay of ^89mY is of 909 keV, and thus, does not overlap with those produced by the annihilation of the positron (511 keV) and decays about 16 s later,⁵ Thanks to the low energy of the positrons emitted by zirconium-89 (E avg = 396.9 keV), the average range of the positron before annihilation is 1.2 mm, reducing its effect in the spatial resolution of the image and producing images of high spatial resolution.⁶ Studies in patients and in animals have shown its capacity for reproducing the distribution of activity, and thus the dose of treatments with lutetium-177 and ⁹⁰Y combined with antibodies.

Figure 2.

Decay scheme of zirconium-89.

In contrast to other PET radiometal emitters, the availability of this radionuclide is quite attainable. The production and the method for radiochemical isolation are well established, simple and not overly expensive. The production of this radionuclide can be carried out in cyclotrons of medical use from the ^89mY (p,n) ⁸⁹Zr reaction.^7,8

⁸⁹Zr is generally found in its ionic form Zr(IV) and presents high affinity for certain chelators that can coordinate with the metal ion. The radiolabeling process involves the formation of a stable complex between ⁸⁹Zr and an adequate chelating agent. This chelation is one of the main advantages of the radiopharmaceuticals labeled with ⁸⁹Zr, since it facilitates the stability of the biomolecule and avoids non controlled release from the organism.

The most common chelators for ⁸⁹Zr are those containing groups that may be strongly coordinated with the ion Zr(IV). Following the introduction of two of the most commonly used bifunctional chelators (ester Fe-DFO-N-suc-TFP and p-NCS-Bz-DFO),⁹ interest in combining the high affinity and specificity of the monoclonal antibodies (mAbs) with the favorable properties of PET imaging of ⁸⁹Zr ([⁸⁹Zr]Zr-immuno-PET) grew, resulting in a increasing number of publications. The first clinical study was performed in 2006 with a mAb (U36) labeled with ⁸⁹Zr for detecting metastasis in the lymph nodes in patients with head and neck cancer.¹⁰ An exhaustive review published by De Feo et al.¹⁰ in 2022, reported the publication of 820 articles related to ⁸⁹Zr, 74 of which were studies in humans, demonstrating the rapid increase in interest for this radionuclide. Fig. 3 shows a time line with the selection of the most important milestones.

Figure 3.

Chronology of a PET image with zirconium 89, highlighting the most important milestones. Adapted from Wuensche et al.⁵

A family of new bifunctional chelators with improved stabilities has recently been developed to overcome the problem of free ⁸⁹Zr uptake in bone. One of these chelators, the tetrahydroxamate chelator called DFO* and its derivative DFO*pPhe-NCS, were successfully synthesized as well as other hydroxamate groups, such as p-SCN-Bn-H6phospa, 3,4,3-(LI-1,2-HOPO), p-SCN-Bn-HOPO, among others (Fig. 4).

Figure 4.

Chemical structures of the chelators used for the labeling of antibodies with ⁸⁹Zr. Adapted from Weijun et al.¹¹

The radioactive labeling of the hydroxamate chelators is preferably performed with [⁸⁹Zr]Zr-oxalate in oxalic acid 1 M rather than [⁸⁹Zr]ZrCl₄ due to its greater stability versus hydrolysis. Several protocols describe similar reaction conditions at room temperature within a pH range of 6.8–7.2, using HEPES buffer at concentrations between 0.25 and 1 M, and Na₂CO₃ 2 M to adjust the pH before mixing it with the conjugated antibody (Fig. 5).

Figure 5.

General procedures of radiolabeling of ⁸⁹Zr to DFO/DFO* antibody conjugates. Adapted from Wuensche et al.⁵

Main radiopharmaceuticals labeled with zirconium-89

⁸⁹Zr is mainly used in the labeling of mAbs, fragments of antibodies, peptides and other small molecules.

Labeling of monoclonal antibodies (immunoPET)

In the last years, the high target affinity and specificity of mAbs has led to their becoming the most investigated class of drugs studied in antitumoral treatment. Following their binding to the antigen, acting as either agonists or inhibitors, mAbs can modulate the function of the receptors associated with the cell membrane, directly or by recruiting the immune system of the body. In either case, the result is cell death.¹²

The PET image using mAbs and analogs as imaging agents (immunoPET) can provide this information non-invasively, which is crucial for the success of immune therapies and antibody drug conjugates and allows predicting and optimizing the dosimetry for radioimmunotherapies.⁶⁸⁹Zr has a half-life of 78.4 h, which makes it adequate for the imaging of antibodies, since it coincides with the pharmacokinetics of its biodistribution.

The potential use of ⁸⁹Zr immunoPET for monitoring and predicting the effectiveness of therapy was demonstrated by Nagengast et al. in ovarian cancer using [⁸⁹Zr]Zr-bevacizumab and the inhibitor of HSP90; NVP-AUY922.¹³ At present, a great variety of mAbs labeled with ⁸⁹Zr have been developed for a wide range of antigens associated with tumors (i.e., EGFR, HER2, CD44v6, PSMA, CD20, VEGF-A, PD-1 and PDL1) (Table 1).

Table 1.

List of the main radiopharmaceuticals labeled with 89Zr, including the ligand, target and corresponding associated disease.

Antibody	Target	Radiopharmaceutical	Pathology	Ref
Certuximab	Epidermal growth factor receptor (EGFR)	[^89Zr][Zr(DFO-certuximab)]	Colorectal cancer Head and neck cancer	^17,18
Panitumumab		[^89Zr][Zr(DFO-panitumumab)]	Colorectal cancer	^19
Nimotuzumab		[^89Zr][Zr(DFO-nimotuzumab)]	Squamous cell carcinomas and gliomas	^20
Bevacizumab		[^89Zr][Zr(Df-bevacizumab)]	Breast cancer Neuroendocrine tumors Renal carcinoma Gliomas	^21,22
Trastuzumab Pertuzumab	Human epidermal growth factor receptor 2 (HER2)	[^89Zr][Zr(Df-trastuzumab)]	Breast cancer	^23
Onartuzumab	Receptors of tyrosine kinase inhibitors (TKI)	[^89Zr][Zr(Df-onartuzumab)]	Pancreatic adenocarcinoma	^23
Rilotumumab	Hepatocyte growth factor (HGF)	[^89Zr][Zr(DFO-Rilotumumab)] [^89Zr][Zr(DFO-AMG102)]	Gastric and lung cancer	^24
Ranibizumab	Vascular endothelial growth factor A (VEGF-A)	[^89Zr] [Zr(DFO-ranibizumab)]	Tumoral angiogenesis	^25
Lumretuzumab	Human epidermal growth factor receptor 3 (HER3)	[^89Zr] [Zr(DFO-lumretuzumab)]	Breast cancer Lung cancer	^26
Rituximab	CD20 antigen	[^89Zr] [Zr(DFO-rituximab)]	Lymphomas	^27
Girentuximab	Carbonic anhydrase IX (CAIX)	[^89Zr][Zr(DFO-girentuximab)]	Renal carcinoma	^28
Fresolimumab	Tumor growth factor (TGF-β)	[^89Zr][Zr(DFO-fresolimumab)]	Gliomas	^29
PROTEIN	TARGET	RADIOPHARMACEUTICAL	PATHOLOGY	REF
huJ591	Prostate-specific membrane antigen (PSMA)	[^89Zr][Zr(DFO-huJ591)]	Prostate cancer	^30,31
IAB2M		[^89Zr][Zr(Df-IAB2M)]		^32,33
RGD	αvβ3 integrins	[^89Zr][Zr(DFO-RGD)]	Gliomas, melanoma, breast cancer	^34
		[^89Zr][Zr(DFO-RGDfK)] [^89Zr][Zr(DFO-RGDyK)]		^35
BBN	Bombesin receptor	[^89Zr][Zr(NOTA-Aca-BBN)]	Prostate cancer, breast cancer and gliomas	^36

Labeling of peptides

PSMA-617 and PSMA-I&T

Prostate-specific membrane antigens (PSMA)-617 and PSMA-I&T are compounds labeled with ⁸⁹Zr which specifically bind to PSMA and are used in PET imaging for the detection of metastatic prostate cancer. These compounds are based on structures that bind to PSMA but are not antibodies. They are labeled with ⁸⁹Zr to improve the image and obtain information on the distribution of tumoral lesions over time.^14–16

Octreotide (TOC) and somatostatin analogs

⁸⁹Zr has also been used for labeling somatostatin analogs such as octreotide (TOC), [⁸⁹Zr][Zr(DOTA-TOC)] and [⁸⁹Zr][Zr(DOTATATE)] for the detection of neuroendocrine tumors. These tumors overexpress the somatostatin receptors (SSTRs) in their membranes, allowing accurate detection by the use of somatostatin analogs labeled with ⁸⁹Zr. With this approach, SSTR + tumors can be effectively visualized with PET, facilitating the diagnosis, localization and the treatment of these tumors (Table 1).

Integrins (αvβ3)

Integrins are cell adhesion proteins that are overexpressed in several types of cancer, including tumoral cells in the case of angiogenesis. The compounds the bind to the integrins, such as cyclic peptide derivatives that bind to αvβ3, have been labeled with ⁸⁹Zr for PET imaging of vascular tumors, being of great utility in monitoring the growth of tumors and the effectiveness of antiangiogenic therapies¹⁶ (Table 1).

Bombesin (GRP-R)

Bombesin is a peptide that binds to the bombesin receptor (gastrin-releasing peptide receptor [GRP-R], which is overexpressed in several tumors, including those of the prostate, breast and others. The use of bombesin analogs labeled with ⁸⁹Zr has been explored to detect tumors that express this receptor. These compounds allow the visualization of these tumors by PET and the evaluation of the efficacy of targeted therapies (Table 1).

Benefits of the use of zirconium-89

• •

  Adequate half-life: The half-life of ⁸⁹Zr (3.27 days) allows the performance of PET images in more delayed times, facilitating the visualization of the biodistribution of the radiopharmaceuticals over time, which is especially useful for dosimetry procedures and treatment evaluation.

• •

  High image quality: Its emission energy is adequate for obtaining high quality images, improving diagnostic accuracy.

• •

  Chemical versatility: It can bind to different types of ligands, allowing its use in a wide range of applications.

In summary, although most of the developments of radiopharmaceuticals with ⁸⁹Zr have been focused on mAbs, there are also very promising applications with small molecules, peptides and other targeted agents that offer an alternative focus for PET imaging and the treatment of different diseases.

Key points

• •

  Zirconium-89 is mainly used in the labeling of monoclonal antibodies (immunoPET), providing key information for the planning and follow-up of oncological therapies.

• •

  The half-life of zirconium-89 is 78.4 h, which allows the obtention of PET images in more prolonged times, facilitating studies of biodistribution and optimization of the dosimetry in immunotherapy.

Astatine-211

Physical characteristics and obtention processes of the radionuclides of astatine with clinical application

Astatine is known as “the rarest element of the Earth” since in its natural form, it can only be found in the form of short-life isotopes in equilibrium with uranium³⁷ and it is estimated that there are only 0.07 g of this element simultaneously in the earth crust. Precisely due to its rarity, its properties are not known with certainty. All the astatine isotopes are unstable and of short life (with periods from 125 ns to 8.1 h), which justifies the origin of its name from the Greek word “astatos” that means unstable.

Astatine-211 is a 100% alpha emitter, with a half-life of 7.2 h. It decays stable lead-207 by two different pathways. It decays to polonium-211 by electron capture with a probability of 58.2%, which, in turn, decays to lead-207 emitting an alpha particle of 7.5 MeV. The second pathway (42%) implies the emission of alpha particles of 5.9 MeV for transition to bismuth-207, which, in turn, decays through electron capture to lead-207.

Astatine-211 is of note due to its simple decay scheme, with the emission of a single alpha particle per decay, in contrast to other available alpha emitters that generate large decay chains, the daughter isotopes of which, can separate from the radiopharmaceutical and release energy in undesired localizations.

The pathway most frequently used for the production of astatine-211 (Fig. 6) is irradiation in a cyclotron with alpha particles of a solid target of natural bismuth, a very inexpensive and abundant material.³⁸ The energy of particle acceleration should be around 28 or 29 MeV to maximize the production of astatine-211 and, at the same time, avoid the production of significant levels of astatine-210, since this decays to polonium-210 (Fig. 6), an alpha emitter with elevated toxicity.³⁹ It is usually obtained in the form of [²¹¹At]NaAt after its extraction from the solid target by wet chemistry. The availability of astatine-211 is compromised by the limited number of cyclotrons able to generate a beam with these characteristics.

Figure 6.

Decay scheme of astatine-211.

Main radiopharmaceutical labeled with atatine-211

[²¹¹At]NaAt

(²¹¹At) sodium astatide may be a useful radiopharmaceutical with therapeutic applications due to its chemical similarity with iodine (both are halogens and possess a similar behavior). This means that astatine, in the form of [²¹¹At]NaAt, may be absorbed by the thyroid cells thorugh the Na+/I-symporter (NIS) transporter in the treatment of thyroid cancer. At present, its possible toxicity is being studied in a preclinical study, with promising results indicating that it may be an alternative candidate to [¹³¹I]NaI.⁴⁰

[²¹¹At]NaAt is obained by the trapping of the [²¹¹At] produced in the cyclotron in a solution of 1% ascorbic acid and 2.3% of sodium bicarbonate for 1 h at room temperature and posterior sterilization.⁴¹

In 2022, Watabe et al.⁴² demonstrated that [²¹¹At]NaAt showed effective induction of a double-strand DNA break with greater cellular toxicity and was more effective than [¹³¹I]NaI in thyroid cancer that expresses NIS in a murine model.

Prior to its use in humans, preclinical studies of toxicity were carried out, demonstrating that the administration of high doses of [²¹¹At]NaAt produces a transitory reduction of white blood cells and lymphocytes, without presenting severe toxicity in either case.⁴³

The clinical trial (CT) NCT05275946 is a phase I study developed by the University of Osaka to evaluate the safety, pharmacokinetics and efficacy of [²¹¹At]NaAt in patients with treatment-resistant differentiated thyroid cancer who did not show adequate response to standard treatment (ClinicalTrials.gov. (2024). Available online at: https://clinicaltrials.gov/ (accessed December 11, 2024).

Meta-[²¹¹At] astatobenzylguanidine ([²¹¹At]MABG)

[²¹¹At]MABG is an analog of norepinephrine that is uptaken by cells that express the norepinephrine transporter which is usually overexpressed in tumors of neuroendocrine origin, such as pheochromocytoma or neuroblastoma.

Synthesis is performed by the reaction of [²¹¹At] dissolved in chloroform with the precursor meta-trimethylsilylbenzylguanidine hemisulfate and N-chlorosuccinimide dissolved in trifluoroacetic acid for 10 min at a temperature of 70 °C. Then, [²¹¹At]MABG is purified with an extraction column in solid phase Sep-Pak tC18 Plus and extracted with an aqueous solution of ethanol at 5% and is formulated with physiological solution 0.9% and ascorbic acid, obtaining yields greater than 36%.⁴⁴

This radiopharmaceutical has undergone preclinical studies showing a greater survival versus [131I]MIBG in models of disseminated disease in neuroblastoma and pheochromocytoma, with a low toxicity but higher dosimetry, making the performance of more studies necessary.^44–46 At present, a phase I clinical trial is under development at the Fukushima University Hospital to evaluate different doses of [²¹¹At]MABG in patients with malignant pheochromocytoma or paraganglioma with the aim of establishing the optimal dose. For this trial, the evaluation criteria are the maximum dose tolerated, pharmacokinetics, rate of urinary activity, rate of general response and disease-free survival.⁴⁷

In 2022, this same group published guidelines for the clinical management of [²¹¹At]MABG in which recommendations are given regarding the management of the radiopharmaceutical and protection against exposure to radiation.⁴⁸

[²¹¹At][At (BC8-B10)]

BC8 is a conjugated murine IgG₁ anti-CD45 mAb. The CD45 protein belongs to the SRC family that expresses all the hematopoietic cells except erythrocytes. It behaves like a tyrosine-phosphatase, which is essential in the transduction of signals originating in the receptor for the antigen of the T lymphocytes during cellular activation.⁴⁹ This antibody conjugates with B10, that is responsible for the chelation of the radionuclide.

The final radiopharmaceutical is obtained by the reaction of [²¹¹At]NaAt with 8 mg of the BC8-B210 antibody dissolved in ammonium acetate (pH 5.5) at room temperature for 2 min, followed by purification by size exclusion chromatography obtaining a yield of 53% and a radiochemical purity greater than 97%.⁵⁰

In 20 patients with acute advanced leukemia and myelodysplastic syndrome, Sanmaier et al. performed a study with this antibody in combination with fluoradabine and 2.3 Gy of whole body irradiation followed by allogenic hematopoietic stem cell transplantation. They observed that with a median follow-up of 1.6 years after treatment, the overall survival and disease-free survival at 1 year were 43% and 35%, respectively.⁵¹

A current ongoing clinical trial (NCT04083183) is aiming to establish the optimal dose of the antibody [²¹¹At][At(BC8-B10)] for the treatment of patients with non-malignant diseases undergoing hematopoietic cell transplantation (ClinicalTrials.gov. 2024. Available online at: https://clinicaltrials.gov/ accessed December 09, 2024).

[²¹¹At][At(OKT10-B10)]

OKT10 is a murine IgG₁ anti-CD-38 mAb, which is a type II transmembrane glycoprotein that is expressed in normal conditions on the surface of several types of immune system, including CD₄⁺, CD₈+, plasma cells and natural killer cells. This antigen is overexpressed in malignant neoplastic hematologic cells such as multiple myeloma. Its overexpression has been correlated with a bad prognosis.⁵²

[²¹¹At][At(OKT10-B10)] is obtained through the nuclear reaction of [²¹¹At]NaAt with the OKT10-B10 mAB at room temperature for 2 min, followed by purification by size exclusion chromatography.

O´Steen et al. performed a preclinical study with [²¹¹At][At(OKT10-B10)] in a mouse model with residual disease of multiple myeloma and found that after administering a single dose in one of the groups, the survival increased four-fold compared to the control group (untreated), offering a potential cure with minimal treatment toxicity.⁵³

At present, this compound has reached phase I of a clinical trial. The NCT04466475 clinical trial which was evaluating the cohort and the secondary effects of [²¹¹At][At(OKT10-B10)] in combined therapy with melphalan prior to stem cell transplantation in patients with multiple myeloma has recently been completed. This trial recruited 24 patients who had received at least three previous lines of therapy, with the principle criterion of evaluation being the maximum dose tolerated, and as secondary criteria, the rate of response, length of response, overall survival and disease-free survival.

The aim of another ongoing trial (NCT04579523) is to assess increasing doses of [²¹¹At][At(OKT10-B10)] when administered together with fludarabine along or in combination with cyclophosphamide and low doses of whole body irradiation before haploidentical stem cell transplantation in 30 patients with multiple myeloma with high risk of recurrence or who do not respond to treatment.

[²¹¹At][At(m-MeATE-Trastuzumab)]

Trastuzumab is a humanized IgG1 mAB that binds with high affinity and specificity to subdomain IV of the human epidermal growth factor receptor 2 (HER2). HER2 is a transmembrane oncoprotein that physiologically participates in cell development. Some types of cancer cells produce abnormal quantities of HER2, inducing more rapid multiplication and dissemination,⁵⁴ and thus, radiopharmaceuticals based on this antibody can diminish these processes.

[²¹¹At][At(m-MeATE-Trastuzumab)] is synthetized in several steps. First, the antibody trastuzumab is conjugated with N-succinimidyl-3-3 trimethylstannyl-benzoate. Then, 450 ul of the immunoconjugate is reacted with the radioisotope after dissolving the radionuclide precursor in 40 ul of a solution of N-iodosuccinimide in methanol with 1% acetic acid for 1 min. Once the reaction has been carried out, the [²¹¹At][At(m-MeATE-Trastuzumab)] is purified by filtration in gel or a size exclusion column, obtaining yields of 75%.⁵⁵

In 2007, Palm et al.⁵⁶ performed a cohort study to determine the efficacy of the radiopharmaceutical in combination with trastuzumab without radiolabeling in a mouse model with radioresistant ovarian carcinoma. Five cohorts with different quantities of antibody (5, 10, 15, 50 and 500 mg) and different activities of the radiopharmaceutical (100, 200, 300, 400 and 500 kBq) were defined with 10 mice/cohort. Each animal received a single dose containing trastuzumab and [²¹¹At][At(m-MeATE-Trastuzumab)]. The results obtained were very promising, achieving complete eradication of the tumors.

In 2019, Fujiki et al. demonstrated that intratumoral administration of [²¹¹At][At(m-MeATE-Trastuzumab)] improves therapeutic efficacy, significantly reducing treatment-associated toxicity.⁵⁶

In 2021, Li et al. demonstrated the efficacy of [²¹¹At][At(m-MeATE-Trastuzumab)] in mice with liver metastasis of gastric carcinomas overexpressing HER2, achieving total eradication of the disease in 75% of the animals treated and with a significant increase in overall survival.⁵⁷

[²¹¹At][At(PSMA5)]

[²¹¹At][At(PSMA5)] specifically binds to the active site of the extracellular domain of PSMA, which is a type II membrane glycoprotein, that is significantly overexpressed in the tumoral cells of different types of cancer.⁵⁸

It is obtained by the reaction of 20 µl of a PSMA5 0.1 mg/mL peptide solution dissolved in sodium bicarbonate (7%) with 1–40 MBq of an aqueous solution of [²¹¹At]NaAt, in the presence of potassium iodide for 45 min at a temperature of 80 °C. [²¹¹At][At(PSMA5)] is purified with an Oasis HLB cartridge and eluted with 1 mL of ethanol at 20%, obtaining yields greater than 60% and a radiochemical purity greater than 96%.⁵⁹

[²¹¹At][At(PSMA5)] shows elevated tumoral uptake in mice with xenografted prostate tumors, observing excellent suppression of tumoral growth with minimum secondary effects.⁵⁹

Key points

• •

  Astatine-211 is an ideal isotope for therapy since it presents the emission of a single alpha particle, avoiding the formation of undesired isotopes.

• •

  The production of astatine-211 is complicated due to the limited number of cyclotrons with the necessary beam.

• •

  Astatine-211 allows the labeling of a range of various molecules.

• •

  [²¹¹At]NaAt may be a candidate for substituting [211 At]NaI in the treatment of thyroid cancer.

Thorium-227

Physical characteristics and processes for obtaining the radionuclides of thorium with clinical application

Thorium is a metal of the actinide series, the isotopes of which are all radioactive, the most abundant being thorium-232. Most thorium isotopes present medium or very short half-lives (few μs or ms) or very long half-lives (a few years) and only two exhibit potential for medical application due to their alpha emission (thorium-226 (30.6 min) and thorium-227 (18.7 days)). The difficult production process of thorium-226, and thus its availability, led to throium-227 being the most promising candidate.

As a radionuclide, thorium-227 can be obtained by a generator system. This radionuclide is part of the decay chain of actinium-227 (half-life of 21.8 years). Actinium-227 can be produced in nuclear reactors by thermal irradiation of target neutrons of radium-226 after the ²²⁶Ra (n,γ)²²⁷Ra-²²⁷Ac reaction (in a nuclear reactor) or by irradiation of natural solid targets of thorium-232 for 10 days at 800 MeV (²³²Th (p,x)²²⁷Ac) or irradiation with protons at a lower energy (90−135 MeV). This latter method, in which the irradiations are with less energy, is still in developmental phases.^60,61

The main limitation that thorium-227 production has is the production and accumulation of radium-223 (Fig. 7A), a product of its decay chain. In fact, the generator is called ²²⁷Ac/²²⁷Th/²²³Ra, and is a documented method for the production of radium-223. Different methods of chemical purification have been described, which allow isolation of both thorium-227 and radium-223 for their potential clinical use.^61–63

Figure 7.

(A) Scheme of thorium-227 decay. (B) graph of the generation of radium-223 from thorium-227.

The generator is eluted with a dissolution of 0.5 M of HCL, obtaining thorium chloride (²²⁷Th) as a radionuclide precursor.

Thorium-227 is a 100% alpha emitter, which, in turn, decays to stable lead-207 and to polonium-211 passing by different radioisotopes which, in addition, are different chemical elements.⁶⁴ This means that this alpha emitter with such a complex decay chain present two main limitations:

• •

  The measurement of the activity and calibration of the equipment. The first daughter radionuclide to which thorium-227 decays is radium-223. Both present half-lives of the same order (18.7 days and 11.4 days, respectively). It could be said that thorium-227 behaves as a “generator” of radium-223 (Fig. 7B). In the generators used in clinical practice, the parent radioisotope is usually not eluted. Moreover, it is necessary to take into account that in any generator there is a time at which both radioisotopes are in chemical equilibrium, but in this case the equilibrium is achieved more quickly because the difference between the half-life of the parent and the daughter is very small (ratio<10). This presents a limitation in the measurement of the radionuclide and not only represents a challenge for the measurement of the activity of a radiopharmaceutical labeled with thorium-227, but also for calibrating the activimeters and the gamma counters. Some articles have proposed that any practical measurement of the activity should be carried out some time before the equilibrium between the two radioisotopes is reached. These measurements are difficult because the response of the equipment increases with the growth of the progeny but also decreases as the source decays.⁶⁵

• •

  The capacity of the radionuclide to remain bound to the vector molecule. During its decay, thorium-227 transmutes to radioisotopes of a different chemical nature. This change in nature, similar to what actinium undergoes and what was discussed in part I, can generate the separation of the radionuclide from the vehicular molecule, and may lead to the radionuclide having its own biodistribution and accumulating in tissues different from the target tissues; this accumulation is especially relevant in tissues such as the red bone marrow, brain, kidneys, among others. The fact that the first daughter in the decay is radium-223 has been a limitation for years since the chelators most commonly used (NODAGA, HBEEC, etc.) do not bind to radium-223 in a stable manner, being the reason why the development of new radiopharmaceuticals with the radioisotopes of radium has been blocked along the history of Radiopharmacy.⁶⁶

• •

  Due to its state of oxidation +4, thorium may form stable complexes with DOTA. However, to achieve stable radiolabeling with this chelator, the complexation process should be performed as a two-step process or directly at elevated temperatures. The difficult conditions used for this type of labeling are not always compatible with complex biological macromolecules, such as antibodies, since they are thermolabile. In addition, slower speeds of complex formation affect radiolabeling, yield, efficiency and specific activity.⁶⁷ Thus, the search for an adequate chelator has been the main objective of the research lines of the radiochemical groups of thorium-227 studied. In 2016, a new type of chelator was proposed: 2,3-HOPO.⁶⁸ This chelator is able to rapidly bind to thorium-227, even at low concentrations, and also form stable complexes with its progeny. The Radiochemistry Laboratory of Berkley peformed experients of serum stability and biodistribution that confirmed that the ligand has excellent stability in vitro and in vivo. This new ligand can be synthetized in a small number of steps from easily available materials, thereby making it more synthetically accessible than the last generation ligands for radiolabeling.^66,69

• •

  The complexes that use thorium-227 are called targeted thorium conjugates (TTC) in the literature. It is common to find the compounds named as TTC followed by the target to which they are directed.⁷⁰

Main radiopharmaceuticals labeled with thorium-227

[²²⁷Th][Th(HOPO-Epratuzumab] (CD22-TTC)

Epratuzumab is a humanized anti-CD22 mAb. CD22 is a transmembrane glycoprotein of the superfamily of immunoglobulins that are expressed on the surface of mature immune B cells. It is often expressed with leukemia and lymphoma cells. In radiolabeled mAbs it has demonstrated a cytotoxic effect at nanomolar concentrations in CD22-positive cancer cell lines. Studies in preclinical animal models have shown very promising results, lengthening the overall survival of the animals treated with this radiopharmaceutical in comparison with those that had received the control treatment.⁷¹

This compound has already undergone an open phase I clinical study with an increase in dose to evaluate the safety, tolerability, maximum dose tolerated, the biodistribution, dosimetry of radiation and the pharmacokinetics of the radiopharmaceutical in subjects with CD22-positive non-Hodgkin lymphoma in relapse or refractory to treatment. The results obtained showed an objective rate of response of 24% (5/21 patients: 1 complete and 4 partial responses), with the greatest response (30%) in patients with recurrence of low-grade lymphomas (3/10 patients).⁷²

[²²⁷Th][Th(HOPO-CD70] (CD70-TTC)

The CD70 cell surface receptor is considered to be a promising target for some neoplasias such as renal cell cancer or B cell lymphoma. Hageman demonstrated effective in vitro activity of CD70-TTC as well as significant inhibition of tumoral growth in cell lines of xenograft models with an increase in the mean survival (doi: 10.18632/oncotarget.16910).

[²²⁷Th][Th(HOPO-Trastazumab] (HER2-TTC)

As mentioned previously, the radiopharmaceuticals based on this mAb present great potential. This radiopharmaceutical potential has achieved phase I of a clinical study (NCT04147819), after passing the preclinical phase with very promising results.^67,73 The CT is aimed at patients with advanced carcinomas that overexpress HER2, and the main objective is to define the safest and most effective cohort for the patients with a schedule of a maximum of 6 doses every 6 weeks as well as observe how the body absorbs, distributes and excretes the drug.⁷⁴ The post-treatment follow-up will be of up to 3 years.

[²²⁷Th][Th (mAb-FGRF2)] (FGRF2-TCC)

Different specific mABs target fibroblast growth factor receptor 2 (FGFR2). This is a tyrosine kinase surface receptor that participates in physiological processes, such as proliferation or differentiation, through signaling pathways for tissue repair. Its elevation within a context of oncologic pathology is associated with more aggressive tumors and with greater resistance to conventional lines of therapy. While the physiological expression is low, the increased expression of FGFR2 in different types of cancer suggests that [²²⁷Th][Th(mAb-FGRF2)] may be a possible therapeutic option. In contrast to other radiopharmaceuticals discussed in the articles of continuing education, in this case, mAb-FGRF2 is generally indicated because the literature does not include the antibody and usually refers to it with a different alphanumerical code (BAY12345). Since 2016 the Bayer preclinical study team has been developing several lines of investigation with different compounds (BAY 1187982, BAY 1895344 and BAY 2304058) in preclinical in vitro and in vivo models of cell lines of breast, colorectal and gastric cancer, obtaining a reduction in cell viability, good tolerance and diminished tumoral growth.^72,75,76

[²²⁷Th][Th (HOPO-Anetumab)] (MSLN-TTC)

Anetumab is a human IgG1 mAb aimed at the mesothelin cell surface glycoprotein. Mesothelin (MSLN) is a glycoprotein that mediates cell adhesion, but when it is dysregulated in cancer, it promotes proliferation, migration and invasion. It is overexpressed in all the mesotheliomas as well as in many ovarian and pancreatic cancers, while it is minimally expressed in normal tissue.

After internalization, this antibody produces the inhibition of cell division and the growth of the tumoral cells that express MSLN, converting it into an attractive target for therapy. Along this line, the compound [²²⁷Th][Th (HOPO-Anetumab)] has demonstrated potent cytotoxic effects in a mesothelin-positive cell line and with a single dose. In preclinical studies that have been carried out, the administration was well tolerated and prolonged the survival of the treated group of mice with disseminated lung cancer (doi: 10.1158/1078-0432.CCR-18-347656). The results of the first trial in humans with advanced MSLN-positive cancer (mesothelioma, ovarian and pancreatic cancer) are pending.⁷⁷

The combination of MSLN-TTC with inhibitors of response to cell damage has been studied in MSLN-positive ovarian cancer by Wickstroem et al.⁷⁸ with the hypothesis of preventing the activation of the mechanisms of DNA repair to maximize the damage induced by MSLN-TTC, leading to the detention of the cell cycles, obtaining synergic antitumoral activity both in vitro and in vivo in xenograft models in mice. Lejeune et al.⁷⁹ investigated the immunostimulatory effects of MSLN-TTC in vitro and in vivo in monotherapy and in combination with the inhibitor of PD-L1 immune checkpoints in immunocompetent mice, obtaining the inhibition of tumoral growth by MSLN-TTC and anti-PD-L1 individually, increasing its benefit if used in combination. They concluded that the TTC can increase the expression of immunomodulator markers on the cell surface that prepare the tumors for combined treatment with immune checkpoint inhibitors, and guide future combination strategies.

Zitzmann-Kolbe et al.⁸⁰ recently demonstrated that the efficacy of MSLN-TTC is independent of the expression of glycoprotein P and may surpass mechanisms of resistance to the existing therapies in ovarian cancer. In addition, they showed that MSLN-TTC presents additive in vivo antitumoral activity in xenografts derived from patients when combined with chemotherapy pharmaceuticals, such as docetaxel and doxorubicin, or with inhibitors of angiogenesis, such as regorafenib or bevacizumab.

[²²⁷Th][Th(Pelgifatamab )] (PSMA-TTC)

Pelgifatamab is a humanized mAb bound to the HOPO chelator that targets the PSMA. The first preclinical results, obtained in 2020, indicated that PSMA-TTC can be produced with high yields and with a stability of 48 h, which facilitates industrial production and distribution. In regard to competitivity, PSMA-TTC showed rapid internalization and selective binding with the target as well as potent induction of DNA damage in vitro. In vivo, the same compound was tested in different xenograft models that simulated different stages of prostate cancer, including some with resistance to androgen inhibitors. Inhibition of tumoral growth was obtained, and in addition, in the model representing metastatic bone involvement, the morphological bone changes induced by the tumor significantly decreased with a single therapeutic administration and the PSA levels decreased with respect to the control group not treated with PSMA-TTC.⁸⁰ It should be highlighted that the myelosuppression observed with PSMA-TTC was dose-dependent and showed signs of recovery.

Moreover, it has been demonstrated that the androgen receptor inhibitor darolutamide induces PSMA expression in cell lines of prostate cancer and xenografts, providing a justification for its combination with PSMA-TTC. In xenograft models of prostate cancer, this fact facilitated the tumoral uptake of PSMA-TTC and darolutamide also altered the induction of genes involved in DNA repair mediated by PSMA-TTC15. In addition, in xenografts, the combination of PSMA-TTC and darolutamide demonstrated synergic inhibition of tumoral growth, and its inhibitory tumoral activity was greater than each independently in enzalutamide-resistant or hormone-independent xenograft models.⁸⁰ At present, the results of a phase I trial of PSMA-TTC alone or in combination with darolutamide in patients with metastatic castration-resistant prostate cancer are pending (NCT03724747).⁸¹ In relation to inhibitors of the DNA repair pathway, PSMA-TTC in combination with olaparib, demonstrated greater antitumoral activity than PSMA-TTC alone and none for olaparib in BRCA2-mutated prostate cancer xenograft models.⁸² Similar results were obtained on combining PSMA-TTC with the DNA-dependent protein kinase in mice carrying prostate tumors, with combined therapy being more effective.⁸³

Key points

• •

  Thorium-227 (Th227) allows the development of targeted thorium conjugates (TTC), facilitating their distribution in different types of tumors.

• •

  TTC have demonstrated synergic effects on combination with inhibitors of DNA repair, immunotherapy or hormone therapies