Targeted Alpha Therapy with Thorium-227

Abstract

Targeted alpha therapy (TAT) can deliver high localized burden of radiation selectively to cancer cells as well as the tumor microenvironment, while minimizing toxicity to normal surrounding cell. Radium-223 (²²³Ra), the first-in-class α-emitter approved for bone metastatic castration-resistant prostate cancer has shown the ability to prolong patient survival. Targeted Thorium-227 (²²⁷Th) conjugates represent a new class of therapeutic radiopharmaceuticals for TAT. They are comprised of the α-emitter ²²⁷Th complexed to a chelator conjugated to a tumor-targeting monoclonal antibody. In this review, the authors will focus out interest on this therapeutic agent. In recent studies ²²⁷Th-labeled radioimmunoconjugates showed a relevant stability both in serum and vivo conditions with a significant antigen-dependent inhibition of cell growth. Unlike ²²³Ra, the parent radionuclide ²²⁷Th can form highly stable chelator complexes and is therefore amenable to targeted radioimmunotherapy. The authors discuss the future potential role of ²²⁷Th TAT in the treatment of several solid as well as hematologic malignancies.

Introduction

Targeted alpha therapy

Targeted thorium-227 conjugates (TTCs) represent a new class of therapeutic radiopharmaceuticals for targeted alpha therapy (TAT). They consist of the α-emitter thorium-227 (²²⁷Th) complexed to a chelator conjugated to a tumor-targeting monoclonal antibody (MAb).

The most common methods of selective treatment are surgery, chemotherapy, and external beam irradiation. Targeted α-emitting radionuclide therapy represents a promising application, characterized by the potential to deliver highly cytotoxic radiation to cancerous cells, sparing normal surrounding tissue. This kind of therapy can thus potentially reduce side effects caused by radiation in normal tissues and increase the destructive radiobiological effects in tumor cells.¹

Physical and biological bases of TAT

An α-particle consist in a nucleus of helium-4 (⁴He) composed of two protons and two neutrons. The α particles have much higher Linear Energy Transfer (LET) than β-particles (50–230 keV/μm vs. 0.1–1.0 keV/μm) with a mean energy deposition of 100 keV/μm.^2,3 The range of α-particles in tissues is short and corresponds to only 5–10 cell diameters (28–100 μm), which restricts the deposition of radiation to the targeted cell and closely neighboring cells. Cellular DNA is the primary molecular target of high LET α-particles due to the efficient induction of DNA double-stranded breaks (DSBs). Moreover, the induction of DSB by α-particles is independent of tissue oxygenation and cellular resistance to photon irradiation and chemotherapy.⁴ The majority of preclinical trials have demonstrated that α-emitters are ideal for treating smaller tumor burdens² and for eradicating circulating malignant cells (e.g., leukemia cells in the blood or bone marrow) or small clusters of cells (e.g., disseminated disease and micrometastases from solid tumors).⁵

TAT is aimed at both macroscopic as well as microscopic residual disease and is therefore configured as a therapeutic procedure with disease-eradicating intent. Several α-particle emitters can theoretically present limitations in their clinical use, as the daughters will detach from the targeting vector due to the elevated recoil energy (100–200 keV). Such free nuclides then diffuse away and can potentially cause toxic effects, however, the studies carried out so far have shown no particular toxicity issues.

Chemical bases of TAT

The growing interest of TAT has led to the development of new chelating agents because the stable sequestration of the radionuclide in vivo is a critical component of targeted radiation therapy permitting the maximum radiation delivery to a tumor while minimizing toxicity.² TAT holds great potential for the treatment of cancer based on the specific delivery of α-particle emitting radionuclides to tumors. Important considerations in selecting an α-emitter for TAT include its availability, chemistry for radiolabeling, physical half-life (t_1/2p), and decay scheme; in particular, the formation of daughter products that emit α- or β⁻-particles that could spread from the tumor potentially causing “off-target” normal tissue toxicity.⁴ There are ∼100 radionuclides known that emit one or more α-particles in the course of the decay to stable nuclides. However, only a few of them have been used in experimental or clinical trials to investigate the efficacy of targeted radionuclide therapy. Table 1 summarizes the physical and chemical characteristics of some radionuclides of interest for TAT.

Table 1.

Physical and Chemical Characteristics of Radionuclides of Interest for Targeted Alpha Therapy

Parent	Daughters		t½	α-Decay (%)	α-Energy (MeV)	Other emissions	Radiochemistry
²¹¹At	²¹¹Po		7.2 h	42	5.87	Kα X-rays	Halogen chemistry, covalent bond with aryl or boron cluster-based prosthetic groups
			0.52 s	100	7.45	77–92 keV
²²⁵Ac			9.9 d	100	5.94		DOTA, DO3A chelator
	²¹³Bi		45.6 min	2.2	5.87	β⁻ 492 keV	CHX-A DTPA, DOTA, NETA
	²¹³Bi		45.6 min	2.2	5.87	γ 440 keV	CHX-A DTPA, DOTA, NETA
		²¹³Po	3.72 μs	100	8.38
²²⁷Th			18.7 d	100	6.14	γ 50 keV	DOTA, Me3-HOPO
²²⁷Th			18.7 d	100	6.14	γ 236 keV	DOTA, Me3-HOPO
	²²³Ra		11.4 d	100	5.71	γ 269 keV
²²⁴Ra			3.63 d	100	5.69	γ 241 keV
	²¹²Bi		60.6 min	36	6.05	β⁻ 834 keV	CHX-A DTPA, DOTA, NETA
						γ 727 keV
						γ 1620 keV
		²¹²Po	0.30 μs	100	8.78
	²¹²Pb		10.6 h			β⁻ 93.5 keV
						γ 238 keV
						γ 300 keV
¹⁴⁹Tb			4.11 h	17	3.97	β⁻ 2432 keV
						γ 817 keV
						γ 853 keV

²¹¹At, Astatine-211; ²²⁵Ac, Actinium-225; DOTA, 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid; DO3A, 1,4,7,10-tetraazacyclododecane-1,4,7-triacetic acid trisodium salt; ²¹³Bi, Bismuth-213; CHX-A DTPA, isothiocyanatophenyl)propyl]-trans-(S,S)-cyclohexane-1,2-diamine-pentaacetic acid; NETA, {4-[2-(bis-carboxymethylamino)-ethyl]-7-carboxymethyl-[1,4,7]triazonan-1-yl}-acetic acid, N4O4; ²²⁷Th, Thorium-227; Me3-HOPO, dimethyl-3-hydroxypyrid-4-one; ²²³Ra, Radium-223; ²²⁴Ra, Radium-224; ²¹²Bi, Bismuth-212; ²¹²Pb, Lead-212; ¹⁴⁹Tb, Terbium-149.

Th-227-based TAT

²²⁷Th represents a particularly promising weapon for therapeutic radiation applications. Moreover, ²²⁷Th is the parent of Radium-223 (²²³Ra), an α-emitter approved for clinical use worldwide, and does not present regulatory issues other than its daughter.

²²⁷Th-labeled radioimmunoconjugate (RIC) has demonstrated stability in serum and in vivo, as well as a significant antigen-dependent inhibition of cell growth. This review will focus on this unique class of compounds and their impact on TAT.

Thorium-227

Th-227 belongs to the actinium series, and the ²²⁷Th decay chain decays by five α-particles to stable lead-207 (Fig. 1). The α-particle emitted from Th-227 has an energy of 5.9 MeV. Retaining Th-227 and all its daughters at a target tissue would deposit ∼34 MeV of energy in a target tissue. The daughter of Th-227 is ²²³Ra, which is the first-in-class α-emitter approved for castration-resistant prostate cancer (CRPC) with symptomatic bone metastases (Xofigo^® prescribing information, 2018; Bayer HealthCare Pharmaceuticals, Inc., Wayne, NJ).^6

–9 The availability of carrier-free α-emitters for radiopharmaceutical use represents the main limiting factor for TAT development. In the past decades, the main source for α-emitters has been natural decay chains or legacy material. New production strategies are focused on using nuclear reactors, charged particle accelerators, and, more recently high-energy accelerators. The production routes employ intense 100, 200, and 800 MeV proton beams and natural thorium targets for the large-scale production of the therapy isotopes ²²³Ra, Actinium-225 (²²⁵Ac), and ²²⁷Th.^10,11 The main advantage of the ²²⁷Th is represented by the possibility of obtaining large quantities by the β⁻ decay of ²²⁷Ac. In turn, ²²⁷Ac can be produced by irradiation of ²²⁶Ra by thermal neutrons in a nuclear reactor. ²²⁷Ac has a half-life of 21.8 years and therefore could allow ²²⁷Th production for decades.

FIG. 1.

The Thorium-227 decay chain.

The 18.7 d half-life of ²²⁷Th easily enables conjugation, administration, and targeting of a ²²⁷Th-labeled RIC before a significant amount of ²²³Ra is generated. Even though a relevant amount of ²²³Ra is accumulated in bones, this may not cause bone marrow toxicity due to the short range of the α-particle.^12,13 Therefore, a therapeutic window allowing treatment with ²²⁷Th with acceptable toxicity may exist. Unlike ²²³Ra, the parent radionuclide ²²⁷Th can form highly stable chelator complexes and is therefore amenable to targeted radioimmunotherapy (RIT),¹⁴ ²²⁷Th exists in the 4+ oxidation state and forms stable complexes with chelators such as 1,4,7,10-tetra-azacycloododecane-N,N′,N″,N′″-tetraacetic acid (DOTA). Labeling of antibodies with ²²⁷Th is done in two steps. In the first step ²²⁷Th-DOTA complexes are obtained by incubation of ²²⁷Th with the chelating agent p-SCN-benzyl-DOTA at 55°C–60°C. In the second step, p-SCN-benzyl-DOTA labeled with ²²⁷Th is covalently conjugated to an antibody at 37°C and a pH of 8–9.⁴ The harsh conditions used for direct labeling are not always compatible with the stability of complex biological macromolecules, such as antibodies and the slower complex formation rates affect radiolabeling yield, efficiency, and specific activity. For this need Ramdahl et al.¹⁵ have developed alternative chelators which complex ²²⁷Th in nearly quantitative yield at ambient temperature in aqueous solutions: octadentate chelates of the 3,2- hydroxypyridinone (3,2-HOPO) class.^16,17

Therapeutic Efficacy of Targeted ²²⁷Th Conjugates

Therapeutic efficacy of ²²⁷Th-immunoconjugates has been investigated in lymphoma, breast cancer, ovarian cancer, acute myeloid leukemia (AML), and renal cell carcinoma on mouse models and in vitro and in vivo studies have evaluated their strengths. The mode of action of TTCs is predominantly linked to the robust induction of irreparable double-strand DNA breaks. Unlike antibody/drug conjugates, TTCs are not strictly dependent on antigen internalization and may obviate the development of cellular resistance.¹⁸ The studies presented below are briefly summarized in Table 2.

Table 2.

Preclinical Studies Discussed in This Article

Article (references)	Topic	Study design	Findings
Mirzadeh⁹	Description of nuclear characteristics and production parameters for α-emitting radioisotopes	Review	Characteristics and production parameters for 7.2 h ²¹¹At, 60.6 min ²¹²Bi, 45.6 min ²¹³Bi, 11 d ²³³Ra, and 20 h ²⁵⁵Fm.
Hagemann et al.¹⁸	Preclinical evaluation of a new TAT for MSLN-positive cancers	Antitumor potency of BAY 2287411 investigated in vitro and in vivo in cell line and patient-derived xenograft models of breast, colorectal, lung, ovarian, and pancreatic cancer	BAY 2287411 was well-tolerated and demonstrated significant antitumor efficacy when administered through single or multiple dosing regimens in vivo. Significant survival benefit was observed in a disseminated lung cancer model.
Dahle et al.¹⁹	Preclinical Evaluation of ²²⁷Th-rituximab in B-lymphoma	In vitro and in vivo, ²²⁷Th-rituximab radioimmunoconjugate effectiveness on human B-lymphoma model	In vitro, ²²⁷Th-rituximab radioimmunoconjugate killed lymphoma cells at very low radioactivity concentration. Complete tumor regression in up to 60% of nude mice human B-lymphoma xenografts without apparent toxicity.
Dahle et al.²⁰	Long-term radiotoxicity of ²²⁷Th-rituximab radiopharmaceutical	Nude mice with Raji xenografts treated with various doses of ²²⁷Th-rituximab	Therapeutically relevant dose levels of ²²⁷Th-rituximab were well tolerated in mice.
Dahle et al.²¹	Determine RBE of -particle radiation from ²²⁷Th-rituximab and of beta-radiation from 90Y-tiuexetan-ibritumomab compared with external beam X-radiation	Nude mice with Raji lymphoma xenografts submitted to different levels of activity	²²⁷Th-rituximab treatment was more effective per absorbed radiation dose unit than the two other treatments.
Heyerdahl et al.²³	Assess the potential of ²²⁷Th-trastuzumab as a therapeutic agent against metastatic cancers that overexpress the HER2 oncogene	Breast cancer and ovarian cancer cells treated with ²²⁷Th-trastuzumab	Dose-dependent inhibition of cell growth and an increase in apoptosis were induced in all HER2-expressing cell lines.
Abbas et al.²⁴	Explore the biodistribution, normal tissue toxicity, and therapeutic efficacy of ²²⁷Th-trastuzumab in mice with HER2-expressing breast cancer xenografts	Biodistribution of ²²⁷Th-trastuzumab and ²²⁷Th-rituximab in nude mice bearing SKBR-3 xenografts were determined at different time points	Internalizing ²²⁷Th-trastuzumab therapy was well tolerated and resulted in a dose-dependent inhibition of breast cancer xenograft growth.
Heyerdahl et al.²⁵	Investigate therapeutic efficacy and normal tissue toxicity of single dosage and fractionated ²²⁷Th-trastuzumab in mice with HER2-expressing breast and ovarian cancer xenografts	Nude mice carrying HER2-overexpression xenografts were treated with 1000 kBq/kg ²²⁷Th-trastuzumab as single injection or four injections of 250 kBq/kg with intervals of 4–5 d, 2 weeks, or 4 weeks	Tumor growth to 10-fold size was delayed and survival with tumor diameter <16 mm was prolonged in all TAT groups compared with the control groups. The same concentration of radioactivity split into several fractions may improve the toxicity of ²²⁷Th-radioimmunotherapy, whereas the therapeutic effect is maintained.
Abbas et al.²⁶	Compare the biodistribution, normal tissue toxicity, and therapeutic effect of ²²⁷Th-trastuzumab and ¹⁷⁷Lu-trastuzumab in mice with HER2-expressing ovarian cancer xenografts	Trastuzumab (Herceptin^®), conjugated to DOTA and radiolabeled with ²²⁷Th or ¹⁷⁷Lu injected into mice bearing SKOV-3 xenografts; the biodistribution was determined at different time points after injection	²²⁷Th-trastuzumab effectively delayed tumor growth and prolonged survival of mice compared with ¹⁷⁷Lu-trastuzumab administered at the same absorbed radiation dose to the tumor.
Abbas et al.²⁷	Compare the biodistribution of the²²⁷Th-trastuzumab and ¹⁷⁷Lu-trastuzumab in mice with HER2-expressing ovarian cancer xenografts; inhibition of tumor growth was measured after single injection of ²²⁷Th-trastuzumab (200, 400, 600, or 1000 kBq/kg), ¹⁷⁷Lu-trastuzumab (40 or 200 MBq/kg)	When compared at the same therapeutic effect the absorbed radiation dose of ²²⁷Th-trastuzumab was three times lower than with ¹⁷⁷Lu-trastuzumab	RBE was higher for ²²⁷Th-trastuzumab than for ¹⁷⁷Lu-trastuzumab, whereas the therapeutic index of ¹⁷⁷Lu-trastuzumab was superior to that of ²²⁷Th-trastuzumab.
Heyerdahl et al.²⁸	The therapeutic effect of ²²⁷Th-trastuzumab on intraperitoneally growing human HER2-positive ovarian cancer cells	Assess biodistribution of intraperitoneally administrated ²²⁷Th-trastuzumab in athymic nude mice; comparison of single ²²⁷Th-trastuzumab doses of 1000, 600 or 400 kBq/kg, or three injections with 400 kBq/kg ²²⁷Th-trastuzumab separated by 4 weeks	Significant therapeutic effect of the ²²⁷Th-trastuzumab treatment both with respect to survival and tumor growth. The maximum tolerated dosage was 600 kBq/kg.
Heyerdahl et al.²⁹	Effect of ²²⁷Th-trastuzumab radioimmunotherapy on tumor vasculature	Human HER2-expressing SKOV-3 ovarian cancer xenograft was injected with ²²⁷Th-trastuzumab at a dose of 1000 kBq/kg body weight; dynamic T1-weighted contrast-enhanced magnetic resonance imaging was used to study the effect of ²²⁷Th-trastuzumab on tumor vasculature	Significant alterations in parameters associated with increased tumor vessel permeability and tumor perfusion after ²²⁷Th-trastuzumab treatment of HER2-expressing ovarian cancer xenografts.
Hagemann et al.³⁰	Characterization and efficacy of CD70, a target for B cell lymphomas and several solid cancers, including renal cell carcinoma, labeled with ²²⁷Th conjugate (CD70-TTC)	In vitro analysis demonstrated that the CD70-TTC retained binding affinity to its target and displayed potent and specific cytotoxicity; biodistribution study in subcutaneous tumor-bearing nude mice using the human renal cell carcinoma cell line 786-O demonstrated significant uptake	Tumor accumulation correlated with a dose-dependent and statistically significant inhibition in tumor growth. CD70-TTC was well tolerated as evidenced by only modest changes in hematology.
Staudacher et al.³¹	There is a need for treatments that can kill hypoxic tumor cells; the murine monoclonal antibody DAB4 (APOMAB), which binds dead tumor cells after DNA-damaging treatment, was radiolabeled with ²²⁷Th	Mice bearing Lewis lung tumors were injected with ²²⁷Th-DAB4 that was examined, as was the effect of these treatments on tumor growth and survival	α-Therapy of necrotic tumor cells with ²²⁷Th-DAB4 had significant and surprising antitumor activity as it would occur only through a crossfire effect.
Washiyama et al.³³	Biodistribution of ²²⁷Th-EDTMP and retention of its daughter nuclide ²²³Ra were examined	²²⁷Th-EDTMP was found to show high uptake and long-term retention in mice bone	²²⁷Th-EDTMP is a potential radiotherapeutic agent for bone metastasis.
Henriksen et al.³⁴	The study explores the use of α-particle-emitting, bone-seeking agents as candidates for targeted radiotherapy	Actinium and thorium-DOTMP and thorium-DTMP were prepared and their biodistribution evaluated in Balb/C mice	²²⁵Ac-DOTMP, ²²⁷Th-DOTMP, and ²²⁷Th-DTMP have promising properties as potential therapeutic bone-seeking agents.
Wickstroem et al.³⁵	Authors hypothesized that blocking the DDR pathway should further sensitize cancer cells by inhibiting DNA repair, thereby increasing the response to TTCs	Evaluation of the MSLN-TTC (BAY 2287411) in combination with several DDR inhibitors; in vitro cytotoxicity experiments were performed on cancer cell lines.Antitumor efficacy of the MSLN-TTC in combination with DDR inhibitors was assessed in human ovarian cancer xenograft models	Effectiveness of the approach by combining the MSLN-TTC with DDR inhibitors as new treatment strategies in MSLN-positive ovarian cancer.
Hammer et al.³⁶	The study presents a PSMA-TTC	In vitro cytotoxicity experiments were carried out on prostate CA cell lines with different PSMA levels; in vivo biodistribution and antitumor efficacy were analyzed after intravenous injection of 100–500 kBq/kg to mice bearing subcutaneous prostate cancer xenograft models	Selective significant antitumor efficacy was shown for PSMA-TTC in subcutaneous prostate CA xenograft models. Significant prevention of tumor growth was observed after treatment with PSMA-TTC at a dose of 100 kBq/kg in an orthotopic bone xenograft model.

211

At, Astatine-211; ²¹²Bi, Bismuth-212; ²¹³Bi, Bismuth-213; ²²³Ra, Radium-223; ²²⁷Th, Thorium-227; RBE, relative biologic effect; HER2, human epidermal growth factor receptor 2; TAT, targeted alpha therapy; DOTA, 1,4,7,10-tetra-azacycloododecane-N,N′,N′′,N′′′- tetraacetic acid; CD70-TTC, CD70-targeted ²²⁷Th conjugate; DOTMP, ²²⁷Th-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetramethylene phosphonic acid; DTMP, diethylenetriamine pentamethylene phosphonic acid; DDR, DNA damage response; TTC, targeted thorium-227 conjugate; MSLN, mesothelin; PSMA, prostate-specific membrane antigen.

²²⁷Th-Rituximab

In the first preclinical study, the therapeutic efficacy of ²²⁷Th-rituximab was investigated in nude mice bearing macroscopic human B-lymphoma xenografts (Raji B-lymphoma). It was demonstrated that ²²⁷Th-DOTA-p-benzyl-Rituximab targets CD20-expressing Raji B-lymphoma tumor cells in vivo and in vitro, successfully inhibits tumor growth and improves survival in mice. Treatment with 200–1000 kBq/kg demonstrated a delay in tumor growth and prolonged mean survival as compared with the control without any serious toxicity. Overall, the low-dose-rate strategy using ²²⁷Th seems to be effective against macroscopic tumors and single tumor cells.¹⁹ To evaluate possible side effects of ²²⁷Th-rituximab, the long-term radiotoxicity of this potential radiopharmaceutical was investigated in BALB/c mice and nude mice with Raji xenografts. The maximum tolerated activity resulted to be between 600 and 1000 kBq/kg. The maximum tolerated dose to bone marrow was estimated in a range between 2.1 and 3.5 Gy. By contrast, 200 kBq/kg of ²²⁷Th-rituximab, an activity proven to be highly efficient in the treatment of B-lymphoma xenografts, was well tolerated. The white blood cell count decrease, used as myelosuppression marker, was the dose-limiting radiotoxicity.²⁰ The relative biologic effect (RBE) of α-radiation from ²²⁷Th-rituximab has been also investigated by Dahle et al. Therapeutic efficacy of α-particle emitting ²²⁷Th-rituximab both at low doses and low-dose rates turned out to be more effective per absorbed radiation dose unit than β⁻-particle emitting ⁹⁰Y-tiuxetan–ibritumomab (Zevalin^®) or external beam X-radiation. In terms of RBE, treatment with ²²⁷Th-rituximab was 2.5–7.2 times more effective in inhibiting tumor growth than external beam radiation, whereas the Zevalin treatment was 1–1.3 times more effective than external beam radiation. Moreover, authors suggest that the best way to get adequate dose is to administer α-emitting radioimmunoconjugates through multiple injections to minimize toxicity and maximize effect per dose unit, reaching a considerable effect even at low doses.^21,22

²²⁷Th-Trastuzumab

Heyerdahl et al. evaluated the cytotoxicity of low-dose-rate ²²⁷Th–DOTA-trastuzumab for treating human epidermal growth factor receptor 2 (HER2)-positive breast and ovarian cancer. Overexpression of HER2 is correlated with poor prognosis and poor treatment response in patients. Trastuzumab (Herceptin^®) is a humanized immunoglobulin G1 (IgG1) MAb that targets HER2 and is currently used as a therapy supplement to surgery, radiotherapy, or chemotherapy. HER2 is a potential target for TAT, especially when used as adjuvant treatment for patients with trastuzumab-resistant disease. Clonogenic survival and cell growth after incubation with 0–20 kBq/mL ²²⁷Th-trastuzumab were studied in human HER2-expressing breast cancer cell lines (BT-474 and SKBR-3) and one ovarian cancer cell line (SKOV-3). BT-474 cells treated with 2.5 kBq/mL ²²⁷Th-trastuzumab showed a mean absorbed radiation dose of 2–2.5 Gy. Cell growth inhibition and apoptosis were induced in all cell lines at the clinically relevant activity concentration. The cytotoxic effect was higher than that of external beam radiation.²³ Therapeutic efficacy of ²²⁷Th-trastuzumab immunoconjugates was also investigated in mouse models with HER2-positive breast cancer xenograft by Abbas et al.²⁴ For all activities of ²²⁷Th-trastuzumab applied, a significant dose-dependent antitumor effect was observed with no serious delayed bone marrow or normal organ toxicity.²⁴ Moreover, another study demonstrated that it might be possible to increase the cumulative absorbed radiation dose to the tumor with acceptable toxicity by the fractionation of the dosage. Splitting the total activity of the RIC ²²⁷Th-trastuzumab into several injections separated in time might reduce toxicity without reducing the therapeutic efficacy. ²²⁷Th TAT may, however, not be convenient for fractionated use in very rapidly growing tumors due to the low-dose rate by which the dose is delivered to the tumor tissue.²⁵ Interestingly, in a subsequent preclinical study, therapeutic efficacy of ²²⁷Th-trastuzumab was compared with ¹⁷⁷Lu-trastuzumab in mice expressing SKOV-3 ovarian cancer xenografts. Therapeutic efficacy of the α-particle-emitting ²²⁷Th-trastuzumab turned out to be significantly better than the β⁻-particle-emitting ¹⁷⁷Lu-trastuzumab.²⁶ The RBE was also studied in mice with breast cancer xenografts, and it was found that it was higher for ²²⁷Th-trastuzumab than for ¹⁷⁷Lu-trastuzumab, infact, to achieve 100% prolonged growth delay the absorbed radiation dose of ²²⁷Th-trastuzumab was three times lower than with ¹⁷⁷Lu-trastuzumab.²⁶ By contrast, the therapeutic index of ¹⁷⁷Lu-trastuzumab was superior to the ²²⁷Th-trastuzumab as when compared with a 50% decrease in the number of white blood cells, the growth delay was three times longer with ¹⁷⁷Lu-trastuzumab than with ²²⁷Th-trastuzumab.²⁷ Efficacy of ²²⁷Th-trastuzumab in treatment of ovarian cancer was also analyzed in athymic nu/nu mice that had been inoculated intraperitoneally with bioluminescent SKOV-3-luc-D3 ovarian cancer cells. The maximum tolerated activity of ²²⁷Th-trastuzumab was found to be 600 kBq/kg. Treatment with ²²⁷Th-trastuzumab significantly reduced tumor growth and prolonged survival compared with animals treated with unlabeled trastuzumab or saline.²⁸ A recent study analyzed the effect of ²²⁷Th-trastuzumab on tumor vasculature after subcutaneous implantation of human HER2-expressing SKOV-3 ovarian cancer tissue fragments in athymic nude mice. Increased vessel permeability, increased perfusion, or both after ²²⁷Th-trastuzumab therapy is an important result: some of the key obstacles to sufficient delivery of targeted cancer therapy with monoclonal antibodies or other large macromolecules are related to the insufficient blood supply and ineffective extravasation, in addition to important nonvascular factors such as binding-site barrier and sequestration, limiting penetration into tumor tissue.²⁹

²²⁷Th-CD70-TTCs

The CD70 cell surface receptor has been reported as a promising target for B cell lymphomas and other malignancies, including renal cell carcinoma. The resulting CD70-targeted ²²⁷Th conjugate (CD70-TTC) showed an effective in vitro activity as well as a significant inhibition of tumor growth in the human renal cancer 786-O cell line derived xenograft model.³⁰

²²⁷Th-CD33-TTCs

Another study supports the feasibility of RIT with CD33-TTCs as a novel radiopharmaceutical for the treatment of AML. A chelator was conjugated to the CD33-targeting antibody lintuzumab through amide bonds, enabling radiolabeling with the ²²⁷Th. In vitro study showed a cytotoxic effect induced by CD33-TTC on CD33-positive cells, independent of multiple drug resistance phenotype. In vivo, the CD33-TTC demonstrated antitumor activity in a subcutaneous xenograft mouse model using HL-60 cells at a single-dose regimen. The dose-dependent significant survival benefit was further demonstrated in a disseminated mouse tumor model after a single-dose injection or administered as a fractionated dose.¹⁵

²²⁷Th-DAB4

Many inoperable solid tumors, such as lung cancer, are at a high risk of local and distant recurrence as well as resistance to standard DNA-damaging therapies, including chemotherapy and external beam radiotherapy. These tumors have a common pathology of necrotic regions lying in close apposition to hypoxic regions, which are much more resistant to conventional therapies. Tumor hypoxia is associated with a poor prognosis and with poor treatment outcomes. Considering that α-particles can directly kill hypoxic tumor cells, the murine monoclonal antibody DAB4 (APOMAB), which binds dead tumor cells after DNA damaging treatment, was conjugated and radiolabeled with the α-particle-emitting radionuclide ²²⁷Th. Mice bearing Lewis lung tumors were administered with ²²⁷Th-DAB4 alone or after chemotherapy. ²²⁷Th-DAB4 accumulated in the tumor particularly after chemotherapy, whereas the distribution in healthy tissues did not change. ²²⁷Th-DAB4 as monotherapy was able to prolong survival, in particular when administered after chemotherapy. Surprisingly, TAT of necrotic tumor cells with ²²⁷Th-DAB4 demonstrated significant antitumor activity, presumably related to a crossfire effect.³¹ Staudacher et al. demonstrated, through targeting of apoptotic and necrotic tumor cells, that cross-dose rather than self-dose of targeted α-therapy exerts significant therapeutic effects. Moreover, given the proximity of DAB4-targeted necrotic areas to hypoxic tumor areas, authors hypothesized that α-particle-mediated lesions are induced preferentially within the hypoxic tumor cells that lie in proximity to ²²⁷Th-DAB4-targeted necrotic tumor cells. Finally, these data indicated that, although α-particles have a path length of no more than 6–10 cell diameters, crossfire antitumor effects could make significant contributions to cancer control by TAT.³¹

²²⁷Th-EDTMP and ²²⁷Th-DOTMP

Washiyama et al. proposed a bone-seeking agent, ²²⁷Th-EDTMP, as a potential therapeutic agent for the treatment of bone metastasis. They examined the biodistribution of ²²⁷Th-EDTMP and the retention of its daughter nuclide ²²³Ra.³² Relatively to the bone, results showed a high uptake and long-term retention in bone and a rapid clearance from blood and soft tissues. The femur-to-tissue uptake ratios at 30 min resulted in more than 100 for all tissues, excluding the kidney. Seven and fourteen days after injection of ²²⁷Th-EDTMP, the retention index of ²²³Ra in bone showed high values, and the differences between these time points were not significant.³³ Henriksen et al. reported uptake in bone of ²²⁷Th-diethylenetriamine pentamethylene phosphonic acid (²²⁷Th-DTMP), and ²²⁷Th-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetramethylene phosphonic acid (²²⁷Th-DOTMP). It was demonstrated that polyphosphonate complexes of ²²⁷Th could have a relevant in vivo stability and be useful to deliver α-particle radiation to primary bone cancer and skeletal metastases from soft tissue cancers.³⁴

²²⁷Th-Mesothelin-TTCs

Mesothelin (MSLN) is a 40 kDa membrane-anchored glycoprotein that is frequently overexpressed in several cancer types, including mesothelioma, ovarian, lung, and pancreatic cancers. It is considered a suitable antigen for targeted therapies because, in contrast, has a limited expression in healthy tissue. BAY2287411, a MSLN-targeting ²²⁷Th conjugate (MSLN-TTC), which is based on the fully human IgG1 antibody BAY86-1903 (Anetumab) covalently attached to a ²²⁷Th complexing 3,2-HOPO chelator, demonstrated potent in vitro and in vivo activity in cellular patient-derived xenograft (PDX) models of breast, lung (mesothelioma), ovarian, and pancreatic cancer. BAY2287411 elicited DSBs, apoptosis, oxidative stress, and reduced cellular viability with an upregulation of immunogenic death markers. BAY2287411 was well tolerated and showed significant antitumor effectiveness once administered through single or multiple dosing regimens in vivo. Additionally, a significant survival benefit was observed in a disseminated lung cancer model. Specific uptake and retention of BAY2287411 in tumors were shown. As a consequence of these encouraging results, it was decided to start a clinical phase I study protocol on the use of BAY2287411 in mesothelioma and ovarian cancer patients (NCT03507452 study protocol).¹⁸

²²⁷Th-MSLN-TTCs with DNA Damage Response Inhibitors

A recent study has explored the therapeutic efficacy of the association of the MSLN-TTCs with DNA damage response (DDR) inhibitor as a new strategy for treating ovarian cancer patients characterized by overexpression of MSLN. In vitro cytotoxicity experiments were performed on cancer cell lines by combining the MSLN-TTC with inhibitors of Ataxia-telangiectasia mutated (ATM), ataxia telangiectasia, Rad3-related protein (ATR), DNA-dependent protein kinase (DNA-PK), and poly [ADP-ribose]polymerase 1/2 (PARP1/2). Utilizing human ovarian cancer xenograft models, a synergistic activity was observed in vitro for all tested inhibitors when combined with MSLN-TTC. ATRi and PARPi appeared to induce the strongest increase in therapeutical strength. Moreover, in vivo antitumor efficacy of the MSLN-TTC and ATRi combination³⁵ demonstrated synergistic antitumor activity.

²²⁷Th-Prostate-Specific Membrane Antigen-TTC

The prostate-specific membrane antigen (PSMA)-targeted ²²⁷Th conjugate PSMA-TTC (BAY2315497) is a human anti-PSMA antibody linked to a chelator moiety (3,2 HOPO). PSMA-TTC has shown strong antitumor activity in PSMA-positive prostate mCRPC cells. Recently, Hammer et al. presented some preliminary data. In mouse cancer models with prostate cancer xenotransplantation, PSMA-TTC showed preferential absorption in tumors compared with other tissues, and dose-dependent suppression of tumor growth, even in a context of bone metastatic model. The association between Androgen Receptor (AR) antagonists and PSMA-TTC has been explored. AR antagonists induced PSMA levels in LNCaP and C4-2 prostate cancer cells in vitro resulting in an increased sensitivity to growth inhibition by PSMA-TTC. Combining PSMA-TTC and enzalutamide, a tumor reduction was achieved in all animals studied, with partial response in 67% and a complete response in 33% of the animals 40 d after the start of treatment. No significant adverse effects on body weight were detected compared with vehicle-treated animals. The combination of PSMA-TTC with darolutamide increased tumor growth inhibition compared with vehicle to 85% and, importantly, 77% of the animals showed stable disease or partial response until 57 d after the start of treatment. Combining PSMA-TTC with AR antagonists shows promising preclinical data in patient-derived prostate cancer models with increased response rates even in an enzalutamide-resistant model.³⁶

Conclusions

Due to the physical properties of the α-emitters TAT appears as a particularly promising and easily suitable therapeutic strategy for eradicating small-volume lesions or to treat minimal residual disease to prevent recurrence or progression, allowing to improve patient outcome. The cytotoxic potential of α-emitters is unique, as they efficiently eradicate hypoxic tumor cells as well as tumor cells that are resistant toward β⁻- and γ-radiation and treatment with cytostatic drugs. Studies described in this review demonstrate how ²²⁷Th has the potential to be an excellent therapeutic candidate in several solid and hematologic tumors. The development of α-immunoconjugates has enabled TAT to progress from in vitro studies, through in vivo experiments and on to clinical trials.²²

To the best of the authors' knowledge at the completion of this review, the following three clinical trials were registered on clinicaltrials.gov:

preclinical results supported the transition of BAY2287411 into a clinical phase I program in mesothelioma and ovarian cancer patients (Trial number NCT03507452);

BAY2315497 is a ²²⁷Th labeled immunoconjugate, specific for the PSMA, which will be evaluated in patients with metastatic CRPC (Trial number NCT03724747);

Epratuzumab ²²⁷Th conjugate is still in phase I trials for non-Hodgkin's lymphoma (Trial number NCT02581878).

The experience coming from the clinical use of ²²³Ra and preclinical and human data on others as an emitter, suggests how it should be the aim to establish TAT as another standard therapeutic option in the treatment of cancers besides surgery, radiotherapy, chemotherapy, and targeted radiotherapy using β⁻-emitters.

Footnotes

Disclosure Statement

No competing financial interests exist.

Funding Information

No funding was received for this article.

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Targeted Alpha Therapy with Thorium-227

Abstract

Introduction