Targeted Alpha Therapy: Progress in Radionuclide Production, Radiochemistry, and Applications

This review outlines the accomplishments and potential developments of targeted alpha (α) particle therapy (TAT). It discusses the therapeutic advantages of the short and highly ionizing path of α-particle emissions; the ability of TAT to complement and provide superior efficacy over existing forms of radiotherapy; the physical decay properties and radiochemistry of common α-emitters, including 225Ac, 213Bi, 224Ra, 212Pb, 227Th, 223Ra, 211At, and 149Tb; the production techniques and proper handling of α-emitters in a radiopharmacy; recent preclinical developments; ongoing and completed clinical trials; and an outlook on the future of TAT.


Introduction
Radionuclide therapy has been employed frequently in the past several decades for disease control, curative therapy, and pain management applications [1]. Targeted radionuclide therapy (TRT) is advantageous as it delivers a highly concentrated dose to a tumor site-either directly to the tumor cells or to its microenvironment-while sparing the healthy surrounding tissues. It has been clinically demonstrated using a variety of radionuclides to treat malignancies, including polycythemia, cystic craniopharyngioma, hyperthyroidism, synovitis and arthritis, and numerous cancers, such as thyroid cancer, bone tumors and metastasis, hepatic metastasis, ovarian cancer, neuroendocrine tumors, leukemia, lymphoma, and metastatic prostate cancer [2][3][4]. Since radionuclide therapy targets diseases at the cellular level, it has advantages for treating systemic malignancies such as tumor metastases over other forms of therapy such as external beam therapy, where full body irradiation is impossible. In addition to being minimally invasive, radionuclide therapy can be shorter in duration than chemotherapy [2].
In TRT, therapeutic radionuclides including alpha (α), beta (β − ), and Auger electron emitters are typically conjugated to a targeting vector such as monoclonal antibodies, biomolecules, peptides, nanocarriers, and small-molecule inhibitors. To maximize the therapeutic efficacy of a TRT radiopharmaceutical, its radionuclide decay pathway, particle emission range, and relative biological effectiveness should be matched appropriately for a given tumor mass, size, radiosensitivity, and heterogeneity [1].
This review focuses on targeted alpha therapy (TAT), outlined by the graphical abstract in Figure 1. A detailed overview of α-emitting radionuclides currently employed in radiotherapy is presented and compared to radionuclides with different emissions, including β − particle and auger electron emitters. Production techniques for α-emitters are outlined, including their separation and unique handling requirements, followed by their radiochemistry and targeting characteristics. Preclinical developments and clinical . Key aspects of targeted alpha therapy: (a) radionuclide production via cyclotron, nuclear reactor or generator decay, and shielded automated processing; (b) radiolabeling the alpha-emitting radionuclide to a suitable targeting vector to form a bioconjugate; and (c) targeted alpha radiotherapy precisely destroys tumor cells while sparing surrounding healthy tissue.

Selecting Radionuclides for Radiotherapy
When selecting a radionuclide for clinical application, the physical and biochemical characteristics must be considered. Physical characteristics include physical half-life, type of emissions, energy of the emissions, daughter products, method of production, and radionuclidic purity. Biochemical characteristics include tissue targeting, retention of radioactivity in the tumor, in vivo stability, and toxicity [2]. For radiotherapy, it is desirable to have a high linear energy transfer (LET), where there is a high ionization energy deposited per unit length of travel. Radionuclides with a high LET deposit radioactive emission energy within a small range of tissue, thereby sparing surrounding healthy tissue and keeping the radioactive dose within, as much as possible, the patient's organ to be treated. It Figure 1. Key aspects of targeted alpha therapy: (a) radionuclide production via cyclotron, nuclear reactor or generator decay, and shielded automated processing; (b) radiolabeling the alpha-emitting radionuclide to a suitable targeting vector to form a bioconjugate; and (c) targeted alpha radiotherapy precisely destroys tumor cells while sparing surrounding healthy tissue.

Selecting Radionuclides for Radiotherapy
When selecting a radionuclide for clinical application, the physical and biochemical characteristics must be considered. Physical characteristics include physical half-life, type of emissions, energy of the emissions, daughter products, method of production, and radionuclidic purity. Biochemical characteristics include tissue targeting, retention of radioactivity in the tumor, in vivo stability, and toxicity [2]. For radiotherapy, it is desirable to have a high linear energy transfer (LET), where there is a high ionization energy deposited per unit length of travel. Radionuclides with a high LET deposit radioactive emission energy within a small range of tissue, thereby sparing surrounding healthy tissue and keeping the radioactive dose within, as much as possible, the patient's organ to be treated. It can also be advantageous for the therapeutic radionuclide, or a complementary theranostic radionuclide, to emit positrons (β + ) or gamma (γ) radiation. This enables positron emission tomography (PET) or single photon emission tomography (SPECT) imaging and visualization of radiopharmaceutical distribution within a patient's body, permitting treatment monitoring. Table 1 outlines key characteristics of α, β − , and auger electron emitters, and some clinical applications for cancer TRT that have been explored. β − emitting radioisotopes have a relatively long pathlength (≤12 mm) and a lower LET of~0.2 keV/µm, giving them effectiveness in medium-large tumors [1]. However, they lack success in solid cancers with microscopic tumor burden. This may be attributed to their emissions releasing the majority of their energy along a several millimeter long electron track, irradiating the surrounding healthy tissue instead of depositing their main energy into the micro-metastatic tumor cells where the radionuclide was delivered [6].
Auger electrons have a medium LET (4-26 keV/µm) [1]; however, their short pathlength of 2-500 nm limits the majority of their effects to within single cells, requiring the radionuclide to be transported into the cell and preferably incorporated into DNA to achieve high lethality. They can also kill cancer cells by damaging the cell membrane and kill non-directly targeted cells through a cross-dose or bystander effect [9]. Clinical studies with Auger electrons for cancer therapy have been limited; however, some encouraging results were obtained using [ 111 In]In-DTPA-octreotide in rats with pancreatic tumors, [ 125 I]I-IUdR where tumor remissions were achieved, and [ 125 I]I-mAb 425 where the survival of glioblastoma patients improved [5,10,11]. However, it has also been determined that some Auger electron emitting compounds, such as [ 123 I]I-IUdR and [ 125 I]I-IUdR only kill cells in the S-phase of the cell cycle, highlighting a potential treatment limitation [12].
α-particles have a high LET (80 keV/µm) and a moderate pathlength (50-100 µm), giving them an effective range of less than 10 cell diameters. This makes them suitable for microscopic tumor cell clusters, while sparing normal organs and surrounding healthy tissues. Importantly, α-particle lethality is not dependent on the cell cycle or oxygenation, and the DNA damage is often via double strand and DNA cluster breaks and is therefore much more difficult to repair than β − damage [6]. It has been estimated that to attain a single cell kill probability of 99.99%, tens of thousands of β − decays are required, whereas only a few α-decays at the cell membrane achieves the same kill probability [13]. From this, it has been estimated that one α particle transversal can kill a cell [14]. Most αemitters are conjugated to a wide range of targeting vectors for delivery to their target site, though some have intrinsic targeting properties, such as the affinity of 223 Ra-dichloride for bone [1]. Preclinical and clinical studies using α-emitters have been ongoing for a variety of cancers, some of which include recurrent brain tumors, recurrent ovarian cancers, human Pharmaceutics 2021, 13, 49 4 of 26 epidermal growth factor receptor-2 (HER-2) positive cancers, myelogenous leukemia, non-Hodgkin lymphoma, metastatic melanoma, and skeletal metastases in prostate cancer [6]. Of these, the most theranostic research is performed on prostate and neuroendocrine tumors (NETs). Examples of studies are numerous-one preclinical study using [ 212 Pb]Pbtrastuzumab found a single injection reduced tumor growth by 60-80%, reduced aortic lymph node metastasis, and prolonged survival or tumor-bearing mice [15]. Another study outlined how α-particle radiotherapy for metastatic castration resistant prostate cancer using [ 225 Ac]Ac-PSMA-617 was able to overcome resistance to [ 177 Lu]Lu-PSMA-617 β − -particle therapy [16]. Additionally, a study using [ 213 Bi]Bi-DTPA and [ 213 Bi]Bi-DOTATATE in mice resulted in a factor of six increase in cell killing compared to [ 177 Lu]Lu-DOTATATE [17,18]. These studies highlight the clinical importance and potential of αemitters, and their potential to be more efficient and effective than β − -therapy.

Alpha Emitter Decay Properties
As emissions from radioactive decay, α-particles are naked 4 He nuclei with a +2 charge. They are 7300 times larger than the mass of β − and Auger electrons, giving them significant emission momentum that reduces deflection and results in a near-linear emission path, as opposed to the winding path of β − particles. With an emission kinetic energy between 5-9 MeV, coupled with a particle range of 50-100 µm, this classifies α-particles as high LET. The energy distribution between the alpha particle and the recoiling daughter atom is typically 98% to 2%. Upon decay, energy imparted to the daughter recoil atom can reach 100 keV [19], which is far higher than the binding energy of the strongest chemical bonds, resulting in release of the daughter isotope from its targeting vector. An example is 219 Rn, which has a daughter recoil range of 88 nm in a cellular environment [19]. These daughters often have a serial decay chain with their own α-emitting progeny, leading to untargeted irradiation of surrounding tissues. As a result, only a limited number of α-emitting radioisotopes are suitable for therapy due to their decay characteristics. The half-life of the radionuclide should be reasonable for therapy; it should not be too short to allow sufficient time for production, radiopharmaceutical synthesis, and delivery to the patient, and it, as well as the half-life of any daughter radionuclide, should not be too long to avoid excess patient dose.

Alpha Emitter Production, Separation, and Handling
Common production methods for clinically relevant α-emitters can involve a cyclotron that accelerates and bombards a target using variety of particles, including protons, alpha particles, lithium and carbon ions, and nuclear reactors such as fast breeder reactors [26]. α-emitters are often delivered by a generator, where a parent isotope decays to the desired radionuclide that is then extracted. Current, anticipated, and potential production methods for various α-emitters are outlined in Table 3. 4.1. 225 Ac and 213 Bi 225 Ac and 213 Bi are currently produced from 229 Th generators (t 1/2 = 7397 y). 229 Th is sourced from the decay of fissile 233 U, which was originally produced by neutron irradiation of natural 232 Th. These generators can be milked over a 3-week period, permitting the separation of 225 Ra and 225 Ac. Current sources of 229 Th that allow production of clinically sufficient activities of 225 Ac/ 225 Bi are available at the Directorate for Nuclear Safety and Security of the Joint Research Centre (JRC) of the European Commission in Karlsruhe, Germany, Oak Ridge National Laboratory (ORNL), the Institute of Physics and Power Engineering (IPPE), and recently Canadian Nuclear Laboratories. Since the 1990s, JRC has produced approximately 13 GBq annually, while ORNL has produced approximately 22 GBq/year, with these 225 Ac products found safe for human administration and applied for patient treatment [32]. As of 2018, worldwide production of 225 Ac has been 68 GBq/year, and while this does support preclinical studies, it only supports several hundred patients per year (when labeled with a 4-50 MBq dose) and prevents large scale 225 Ac/ 213 Bi generator production. However, no production technique has emerged as an effective and economic solution, leaving 225 Ac supply as a patchwork of production methods with varying yields and radioisotopic purities [33]. It is clear that alternative methods of 225 Ac production would enhance the supply chain and enable more widespread use.
Alternative production methods of 225 Ac include neutron, proton, and deuteron irradiation of 226 Ra targets, and high-energy proton irradiation of 232 Th targets. Large-scale production of 225 Ac by cyclotron proton irradiation of 226 Ra via the 226 Ra(p,2n) 225 Ac reaction shows promise. There is a significant 710 mb cross-section peak at 16.8 MeV proton energy, making this ideal for high-yield and cost-effective production for cyclotrons with beam energies less than 20 MeV [26]. Additionally, no long-lived actinium byproducts ( 227 Ac, t 1/2 = 21.8 y) are produced, and coproduced 226 Ac (t 1/2 = 29 h) via the 226 Ra(p,n) 226 Ac and 224 Ac (t 1/2 = 2.8 h) via the 226 Ra(p,3n) 224 Ac can be minimized by optimizing proton beam energy, and allowing for a "cool-down" period for decay post-irradiation due to their shorter half-lives compared to 225 Ac. Although handling 226 Ra is technically demanding due to its inherent radioactivity and 1600-year half-life, this method is preferred for large- scale, cost-effective production [32]. Irradiating 232 Th with high-energy protons has been demonstrated to produce several GBq of 225 Ac using an intense proton beam irradiation for 10 days. However, this method produces a variety of radionuclidic impurities that must be removed by chemical separation and significant amounts of long-lived 227 Ac (0.1-0.2% at end of bombardment) [32]. 213 Bi generators have been developed by JRC and demonstrated to reliably prepare up to 2.3 GBq of 213 Bi radiolabeled compounds for therapeutic doses when loaded with up to 4 GBq of 225 Ac [32]. They feature homogenous distribution of the 225 Ac over the generator resin, minimizing radiolytic degradation and permitting operation for several weeks.
However, the availability of 213 Bi generators depends on the availability 225 Ac generators, which are already in short supply [17].

224 Ra, 212 Bi, and 212 Pb
224 Ra, 212 Pb, and 212 Bi can be produced from 228 Th (t 1/2 = 1.92 years) generators; however, radiolytic damage occurred to the Na 2 TiO 3 resin, resulting in diminished yield and posing a radiation safety issue [27]. This resulted in a switch to 224 Ra (t 1/2 = 3.6 d)based generators from which 212 Bi and 212 Pb are obtained by elution. Despite producing high yields of 212 Pb (>90% of expected activity per daily elution) and its daughter 212 Bi, the generator must be replaced after 1-2 weeks due to the short half-life of 224 Ra [34].

227 Th and 223 Ra
227 Th and 223 Ra are available by separation from their parent 227 Ac radionuclide. Production of 223 Ra for clinical applications utilizes 227 Ac/ 227 Th generators, where the parent isotopes are on actinide chromatographic resins, and 223 Ra-chloride solution is obtained after elution and purification on a cation exchange column [35].

211 At
211 At can be produced using an α-particle beam to bombard natural bismuth at 28-29.5 MeV via the 209 Bi(α,2n) 211 At reaction [29]. Despite being a straightforward method of production, the number of accelerators capable of a 28 MeV α beam limits the availability of 211 At. 211 At can also be produced via the 209 Bi( 7 Li,5n) 211 Rn reaction, where 211 Rn (t 1/2 = 14 h) decays to 211 At. This method, currently under development, exploits the longer half-life of 211 Rn to extend the timeframe for effective distribution and use of 211 At [30].

149 Tb
The availability of 149 Tb is currently quite limited primarily due to inaccessible production routes. 149 Tb has been produced for use in preclinical α-therapy studies by the isotope and separation online facility (ISOLDE) at CERN, using high-energy proton irradiation of tantalum targets (1.4 GeV) and separation using a magnetic field. Potential production routes also include irradiation of rare 152 Gd targets with high-energy protons (>50 MeV), or spallation reactions using light ions at >500 MeV; however, these methods of production would require mass separation to avoid radioisotopic impurities [36]. 149 Tb was also produced using the nat Nd( 12 C,xn) 149 Dy -> 149 Tb method by irradiating a thick target of nat Nd 2 O 3 with 12 C ions at 108 MeV. However, the scarcity of 12 C accelerators limits this production avenue. Another method of production includes the 151 Eu( 3 He,5n) 149 Tb reaction, where 151 Eu is bombarded by 3 He particles from 40-70 MeV. This method results in high yield production of 149 Tb (3 GBq for an 8 h irradiation), and is advantageous due to target material availability and relatively simple radiochemical processing. The main drawback is the limited availability of high intensity 3 He beams [25].

Alpha Emitter Handling
Once produced, α-emitters require special handling and equipment beyond the current capabilities of many radiopharmaceutical centers. Since the external protective layer of skin stops α-particles, pure α-emitters are not an external radiation hazard. However, ingesting and internalizing α-emitters can cause serious effects, including cancer, genetic diseases, teratogenesis, and degenerative changes. To handle α-emitters, specialized equipment to detect α-particles should be employed, such as ZnS(Ag) scintillators to complement Geiger-Muller counters [37]. Detecting α-particles is difficult and time consuming due to the short range of α-particles in air, making a method of long-range detection of α-particles highly desirable [38]. This can be achieved by observing the secondary effects of α-emitters, such as the air-radioluminescence caused by ionization, where α-decay yields up to 400 UV photons in air. These photons have a much longer range in air than α-particles, and in areas with low background UV, they can be used to detect α-particles. One such detector under development has the capability to detect α-particles from a distance of about 40 cm [39]. Due to their properties, handling α-emitters requires much lower contamination removal levels than β − -emitting radionuclides [1].
For α-emitters with low energy γ emissions, a well-ventilated fume hood with a glove box is sufficient. For α-emitters with high-energy γ emissions, work should be performed behind 15 cm lead bricks or inside a shielded hot cell with remote manipulator arms [40]. When volatile α-emitters are involved, gas-tight enclosures should be considered, and with all α-emitters, double gloving and γ-counter and liquid scintillation should be employed to prevent and monitor contamination [1]. Additionally, radiopharmacies should have a dedicated space for the clinical production of α-emitters, and dedicated α-emitter waste storage.

Radiochemistry and Chelators
Chelating agents that result in the stable coordination of α-emitting radionuclides are important, since a matching radiometal-chelate chemistry is key to success in targeted therapies and avoiding unintended distribution and toxicity to nontarget organs and tissues. Metallic radionuclides have utilized bifunctional chelating agents, which have a metal-binding moiety function that sequesters the metallic radionuclide, and a chemically reactive functional group that covalently attaches a targeting vector such as small molecule peptides, proteins, or nanoparticles. The chelator vector can be attached to the vector either directly or through a linker that is often used to modify pharmacokinetics, which can be a simple hydrocarbon chain or a small peptide sequence. The loss of dissociation of the radiometal is associated with therapeutic toxicity; thus, key coordination chemistry factors to consider include charge, similar ionic radius and chelating cavity size, as well as providing an optimal number of chemically appropriate donor binding groups. [41].
From this, the stability of radiopharmaceuticals for use in α-therapy depends on different characteristics, including the coordination properties of the parent radionuclide, complexation kinetics including time, temperature, pH, the thermodynamic stability of the radionuclide-chelator complex in solution, and the kinetic inertness when competing with other ions and chelating agents. Additional factors that influence radiopharmaceutical quality include radiolytic effects, the recoil effects of the daughters, and the unique chemistry of the various daughter nuclides. Several commonly employed chelators are depicted in Figure 3, and notable chelators that have been investigated with therapeutic α emitters are listed in Table 4.
Pharmaceutics 2021, 13, x FOR PEER REVIEW 10 of 27 chemistry of the various daughter nuclides. Several commonly employed chelators are depicted in Figure 3, and notable chelators that have been investigated with therapeutic α emitters are listed in Table 4.  In aqueous solution, 225 Ac is most stable in its +3 oxidation state, with a potential possibility of accessing its +2 oxidation state [48]. Notably, Ac 3+ is the largest +3 ion on the periodic table, and with a low charge density, it is the most basic +3 ion. Ac 3+ possesses similar chemical properties to lanthanide Ln 3+ ions, with lanthanum La 3+ being a useful Ac 3+ surrogate for radiochemistry. With its demonstrated ability to stably coordinate hard +3 ions, 225 Ac is stable on coordination by the DOTA chelator and its derivatives [49], and has shown improvements in whole body clearance and decreased organ uptake compared to acyclic ligands such as EDTA and CHX-A" DTPA. Other ligands, such as macropa and macropa-NCS have promise in coordinating 225 Ac, and they showed minimal organ accumulation and remained highly stable in vivo over an extended period of time [50,51]. It has also been suggested that acyclic chelating agents such as linear polyaminocarboxylates are not good ligands for 225 Ac due to high liver uptake and poor whole body clearance resulting from the loss of 225 Ac from these chelating agents [52]. DOTP 8-has been  In aqueous solution, 225 Ac is most stable in its +3 oxidation state, with a potential possibility of accessing its +2 oxidation state [48]. Notably, Ac 3+ is the largest +3 ion on the periodic table, and with a low charge density, it is the most basic +3 ion. Ac 3+ possesses similar chemical properties to lanthanide Ln 3+ ions, with lanthanum La 3+ being a useful Ac 3+ surrogate for radiochemistry. With its demonstrated ability to stably coordinate hard +3 ions, 225 Ac is stable on coordination by the DOTA chelator and its derivatives [49], and has shown improvements in whole body clearance and decreased organ uptake compared to acyclic ligands such as EDTA and CHX-A" DTPA. Other ligands, such as macropa and macropa-NCS have promise in coordinating 225 Ac, and they showed minimal organ accumulation and remained highly stable in vivo over an extended period of time [50,51]. It has also been suggested that acyclic chelating agents such as linear polyaminocarboxylates are not good ligands for 225 Ac due to high liver uptake and poor whole body clearance resulting from the loss of 225 Ac from these chelating agents [52]. DOTP 8has been chelated with 225 Ac, compared with other 3+ cations, including Am, Cm, and La, and verified to be encapsulated within the DOTP 8binding pocket [44]. Recently developed 225 Ac chelators such as H 4 py4pa and H 4 py4pa-phenyl-NCS have demonstrated excellent in vivo stability and tumor specificity [53]. Another novel chelator, 225 Ac-crown, was shown to be stable over an extended period of time, while binding rapidly to 225 Ac at ambient temperature [45].

213 Bi
213 Bi is obtained in its +3 oxidation state from a 225 Ac/ 213 Bi generator, and forms stable complexes with nitrogen-rich chelators such as CHX-A"-DTPA or NETA, and is also stable with DOTA [49]. Optimized protocols for 213 Bi labelling of antibodies in clinical settings permit the synthesis, sterile filtration, and quality control of therapeutic doses within 15 min of elution from a 225 Ac generator [32]. It has also been found that kinetically inert Bi 3+ complex formation is very slow [54]. Notably, phosphorus containing ligands DOTP, DOTP H , DOTP Et , DOTPI were found to match CHX-A"-DTPA and have superior labelling efficiencies to DOTA, with DOTP being the most efficient. They also exhibited excellent stability in human plasma and exhibited a higher stability against demetallation compared to DOTA and CHX-A"-DTPA [46]. Th has a +4 oxidation state, and has been studied with ligands that accumulate in bone, including phosphate ligands such as DTMP, DOTMP, and EDTMP. Employing 227 Th is advantageous for obtaining an additional α-decay, compared to just using its 223 Ra daughter [28]. Although 223 Ra has a +2 oxidation state, it is highly basic and does not form complexes in aqueous solution. There are no known 223 Ra chelators, so there is no way to target its accumulation. 227 Th can also be chelated by hydroxopyridinone coordinating moieties, including N-methyl-3,2-hydroxypyridinone (Me-3,2-HOPO) [55].

211
At 211 At is a halogen, presenting different radiolabeling chemistry than radiometals. In its positive oxidation states, astatine exhibits properties specific to metal ions such as silver in its +1 oxidation state and polonium in its higher oxidation states. In its −1 state, it exhibits characteristics similar to iodine. Slow, low yield electrophilic radiolabeling of nonactivated aromatic rings can lead to 211 At compounds in vivo [28]. Several compounds that have been stable when labeled with 211 At include N-succinimidyl 3-(trimethylstannyl)benzoate and a boron cage compound [56,57]. 211 At can also be radiolabeled by adapting radioiodination chemistry using tin precursors and prosthetic groups [58]. 5.1.6. 149 Tb 149 Tb forms +3 and +4 oxidation states and can be chelated with DOTA conjugates [59], and it is particularly attractive since there are no α-emitting progeny presenting a redistribution issue upon release from the chelator. Radiolabeling using bifunctional chelators such as DOTA and DTPA has been well developed, giving a clear advantage over alpha emitters such as 211 At and 223 Ra [25]. One study using [ 149 Tb]Tb-DOTANOC achieved high-quality PET images of an AR42J tumor-bearing mouse, demonstrating the exceptional potential of 149 Tb to combine α-therapy with PET in "alpha-PET" using a single radionuclide [59].

α Emitter Redistribution
While α emitters with multiple daughters-in vivo generators-can enhance the delivered dose to a tumor site, they can also redistribute and cause unintended toxic effects. Upon α emission, recoil energy (~100 keV) imparted to a daughter nuclide is at least 1000 times higher than any chemical bond, releasing in at least partial release of the daughter nuclide from the targeting molecule. Redistribution depends on the distance traversed by the daughter nuclide upon release, diffusion and active transport processes, the half-life of the daughter, and the affinity of the radionuclide for different organs [1]. With redistribution, radioactive burden can be spread across the body, reducing elimination and leading to radiotoxic effects such as organ dysfunction and secondary tumorigenesis. Dosimetry is essential to understand the contributions of a radiolabeled targeting vector, labeled metabolites, liberated mother nuclides, and the daughters released upon recoil [19]. Radionuclide distribution is often measured in postmortem ex vivo organ analysis, using an alpha camera or a timepix detector [19,[60][61][62].
Some α-emitters are prone to redistribution. Redistribution compromised the continuation of a 224 Ra clinical study, with 8% of the 220 Rn daughter nuclide leaving the body and significant 212 Pb and 212 Bi uptake observed in the red blood cells, kidney, and liver [63]. 225 Ac faces limitations due to the redistribution of the 213 Bi daughter to the kidneys, with one study in mice showing 0.77 Gy·kBq −1 of kidney dose after tissue harvest, 60% of which was attributed to nonequilibrium 213 Bi [64]. Dose-limiting salivary gland toxicity and reduced salivary gland function have also been observed in clinical studies using 225 Ac-labelled PSMA for treating metastatic castration-resistant prostate cancer [65].
Other α emitters such as 223 Ra demonstrated low redistribution in mice and humans [66]. 149 Tb shows promise compared to the previously listed α-emitters since it has only one α emission in its decay chain; thus, once 149 Tb radiopharmaceuticals accumulate in their target, they should be less prone to toxic redistribution effects.
Several theories have been proposed to mitigate the consequences of the daughter recoil effect [19]: (1) Since the spread of daughter radionuclides takes time in the body, their spread depends on their physical half-life. With blood flow in capillaries between 1-3 mm/s, daughter nuclei will not undergo significant translocation if their half-lives are short (several seconds). (2) Daughter recoil can be mitigated by nanoconstructs. Encapsulating a radionuclide within the core of a nanoconstruct with sufficient stopping power can significantly mitigate free daughter spread. (3) Even if a recoil daughter escapes a nanoconstruct, it has a high probability of back-implantation into surrounding nanoconstructs [19].
While α emitters with a shorter t 1/2 are an effective solution to daughter redistribution, the higher cytotoxicity-and therefore therapeutic potential-of radioisotopes with progeny redistribution has motivated developing techniques to control the daughter radionuclides [1]. Nanocarrier encapsulation of α-emitters to contain recoil daughters shows promise, with one method utilizing liposomes. 223 Ra was encapsulated in pegylated liposomal doxorubicin (PLD) and remained relatively stable in vivo with skeletal uptake lower than free 223 Ra [67]. Liposomes have also enhanced the retention of 225 Ac daughters [68]. One study used TiO 2 nanoparticles labeled as a carrier for 225 Ac and functionalized with substance P (5-11), a peptide fragment targeting NK1 glioma cell receptors. Leaching of 30% of 221 Fr, the first decay daughter of 225 Ac, was observed in cerebrospinal fluid after 10 days, with the complex showing high cytotoxic effects in T98G glioma cells [69]. One study employed 225 Ac gold-coated lanthanide phosphate nanoparticles, where the multishell nanoparticles combined the radiation resistance of lanthanide phosphate, magnetic properties of gadolinium phosphate, and gold chemistry for attaching targeting vectors [70]. Another approach involves diffusing alpha-emitters radiation therapy (DaRT), a new form of brachytherapy, where seeds impregnated with radionuclides are embedded in solid tumor tissue where they continually release α-emitters [71].
An additional method to reduce toxic renal effects of 225 Ac daughters involves metal chelation therapy and diuretics. In animals, oral metal chelation with dithiols was shown to reduce renal 213 Bi activity, and furosemide and chlorothiazide significantly reduced 221 Fr renal activity [72].
Pretargeting also shows potential for both PET imaging and for reducing nonspecific toxicity and hematotoxicity in radioimmunotherapy. This is where an unlabeled immunoconjugate capable of binding a tumor specific antigen is injected prior to a small molecular weight payload that then binds to the immunoconjugate [73].

Preclinical Studies and Clinical Applications
There are numerous preclinical and clinical studies of α-emitting radiopharmaceuticals completed or underway for a variety of cancers, which are outlined in Table 5.

Preclinical Studies
Since α-emitters have the potential for high efficacy but also high toxicity, preclinical studies, in vivo and in vitro, are essential for optimizing α-therapy and guiding further clinical trials. Some notable preclinical studies using a variety of α-emitters are outlined below. antigens are found on all immune cells, including hematopoietic stem cells and precursor and mature lymphoid and myeloid cells, giving potential for radiotherapy of leukemia and lymphoma. Anti-CD45 radioconjugates labeled with 225 Ac, such as [ 225 Ac]Ac-BC8, have demonstrated effective tumor control in mice bearing multiple myeloma tumors, with modest uptake in the kidney and significant uptake in the liver [77]. Human epidermal growth factor receptor type 2 (HER2) is overexpressed in carcinomas; thus, HER2 antibodies and nanobodies have been investigated for radioimmunotherapy. 225 Ac-labeled nanobodies such as [ 225 Ac]Ac-DOTA-Nb have shown fast uptake in tumor-bearing mice with HER2-overexpressing tumors, with coinjection of Gelofusine reducing kidney retention by 70% [95]. TAT can also target the tumor microenvironment, such as the vasculature and neovascular endothelium. One study using a [ 225 Ac]Ac-E4G10 antibody conjugate that targets the vascular endothelium of glioblastoma demonstrated tumor growth control and improved survival in a mouse [81]. Figure 4 shows MRI images of [ 225 Ac]Ac-E4G10-treated glioblastoma-bearing mice, demonstrating the clear tumor growth control effected by the treatment. Metastatic prostate cancer is of interest for TAT due to the external domain of the prostate-specific membrane antigen (PSMA). 225 Ac has been used with PSMA compounds such as PSMA-617 for prostate cancer therapy. One study in PC-3/PC-3-PIP-tumor-bearing mice compared the effects of [ 177 Lu]Lu-PSMA617 with [ 225 Ac]Ac-PSMA617, finding improved overall antitumor effectiveness and enhanced therapeutic efficacy for [ 225 Ac]Ac-PSMA-617 [79]. CD-45 antigens are found on all immune cells, including hematopoietic stem cells and precursor and mature lymphoid and myeloid cells, giving potential for radiotherapy of leukemia and lymphoma. Anti-CD45 radioconjugates labeled with 225 Ac, such as [ 225 Ac]Ac-BC8, have demonstrated effective tumor control in mice bearing multiple myeloma tumors, with modest uptake in the kidney and significant uptake in the liver [77]. Human epidermal growth factor receptor type 2 (HER2) is overexpressed in carcinomas; thus, HER2 antibodies and nanobodies have been investigated for radioimmunotherapy. 225 Ac-labeled nanobodies such as [ 225 Ac]Ac-DOTA-Nb have shown fast uptake in tumor-bearing mice with HER2-overexpressing tumors, with coinjection of Gelofusine reducing kidney retention by 70% [95]. TAT can also target the tumor microenvironment, such as the vasculature and neovascular endothelium. One study using a [ 225 Ac]Ac-E4G10 antibody conjugate that targets the vascular endothelium of glioblastoma demonstrated tumor growth control and improved survival in a mouse [81].  213 Bi has been shown to be more effective than 177 Lu in animal models for treatment of multiple myeloma (MM). Using 213 Bi-labeled 9E7.4 anti-CD138 antibodies in mice with 5T33 MM cells, mean survival was significantly increased significantly and a cure was effected in 45% of animals. 177 Lu-labeled 9E7.4 anti-CD138 antibodies increased survival to a lesser degree than the 213 Bi, with no mice cured [96].
[ 213 Bi]Bi-DOTA-biotin has been used to target non-Hodgkin lymphoma when pretargeted with an anti-CD20 fusion protein. Mice with Ramos lymphoma xenografts exhibited significant delayed tumor growth and no treatment-related mortalities [84]. Another study in mice with intramuscular LNCaP xenografts used 213 Bi conjugated to a J591 anti-PSMA monoclonal antibody, which resulted in improved tumor-free survival, effectively  213 Bi has been shown to be more effective than 177 Lu in animal models for treatment of multiple myeloma (MM). Using 213 Bi-labeled 9E7.4 anti-CD138 antibodies in mice with 5T33 MM cells, mean survival was significantly increased significantly and a cure was effected in 45% of animals. 177 Lu-labeled 9E7.4 anti-CD138 antibodies increased survival to a lesser degree than the 213 Bi, with no mice cured [96].
[ 213 Bi]Bi-DOTA-biotin has been used to target non-Hodgkin lymphoma when pretargeted with an anti-CD20 fusion protein. Mice with Ramos lymphoma xenografts exhibited significant delayed tumor growth and no treatment-related mortalities [84]. Another study in mice with intramuscular LNCaP xenografts used 213 Bi conjugated to a J591 anti-PSMA monoclonal antibody, which resulted in improved tumor-free survival, effectively stopped the growth of LNCaP spheroids, and reduced prostate-specific antigen levels [97]. 6.1.3. 223 Ra.
The antitumor effects of 223 Ra have been demonstrated extensively in animal models, leading to commercial use of this radium isotope in men with metastatic prostate cancer, where it has improved overall survival and reduced the time until the first symptomatic skeletal event [98]. 223 Ra was investigated in an experimental skeletal metastases model in nude rats that received human breast cancer cells. The animals treated with 223 Ra demonstrated significant antitumor effects and improved survival rates, with no signs of bone marrow toxicity or body weight loss. This study led to further development of 223 Ra as a bone-marrow-sparing treatment for skeletal cancers [88]. stopped the growth of LNCaP spheroids, and reduced prostate-specific antigen levels [97]. The antitumor effects of 223 Ra have been demonstrated extensively in animal models, leading to commercial use of this radium isotope in men with metastatic prostate cancer, where it has improved overall survival and reduced the time until the first symptomatic skeletal event [98]. 223 Ra was investigated in an experimental skeletal metastases model in nude rats that received human breast cancer cells. The animals treated with 223 Ra demonstrated significant antitumor effects and improved survival rates, with no signs of bone marrow toxicity or body weight loss. This study led to further development of 223 Ra as a bone-marrow-sparing treatment for skeletal cancers [88].

224 Ra
224 Ra is similar to 223 Ra with its tendency to adsorb to bone at sites of active mineral crystallization, and inability to be stably bound to a targeting molecule vector. Calcium carbonate microparticles have been proposed as 224 Ra carriers for treatment of disseminated cancers, such as in those occurring in the peritoneum. One study demonstrated high labelling efficiencies of 224 Ra on the surface of calcium carbonate microparticles with high retention of both 224 Ra and its 212 Pb daughter, and the radioactivity remained primarily localized in the peritoneal cavity [89]. 224 Ra has also been investigated for osteolytic bone metastasis of MDA-MB-231(SA)-GFP-bearing nude mice, decreasing the number and area of tumor foci and prolonging survival [99]. Preclinical diffusing alpha emitters therapy (DaRT) using 224 Ra-loaded wires was tested on mice bearing prostate (PC-3), glioblastoma (GBM, U87-MG), colon (HCT15), squamous cell carcinoma (FaDu), and melanoma (C32) cancer cell tumors. The in vivo study confirmed that DaRT can destroy tumors, with only the C32 cells exhibiting resistance [100].

212 Pb
The 224 Ra daughter 212 Pb has been used in a variety of preclinical trials. 212 Pb-labeled 376.96 monoclonal antibodies were used in mice bearing pancreatic cancer Panc039 xenografts, exhibiting significant uptake and tumor growth inhibition compared to an existing [ 212 Pb]Pb-F3-C25 compound [90]. 212 Pb-labeled 225.28 antibodies were used in immunedeficient mice bearing SUM159 and 2LMP human triple-negative breast cancer cells, showing high cell uptake and effective inhibition of tumor growth for CSPG4-expressing

224 Ra
224 Ra is similar to 223 Ra with its tendency to adsorb to bone at sites of active mineral crystallization, and inability to be stably bound to a targeting molecule vector. Calcium carbonate microparticles have been proposed as 224 Ra carriers for treatment of disseminated cancers, such as in those occurring in the peritoneum. One study demonstrated high labelling efficiencies of 224 Ra on the surface of calcium carbonate microparticles with high retention of both 224 Ra and its 212 Pb daughter, and the radioactivity remained primarily localized in the peritoneal cavity [89]. 224 Ra has also been investigated for osteolytic bone metastasis of MDA-MB-231(SA)-GFP-bearing nude mice, decreasing the number and area of tumor foci and prolonging survival [99]. Preclinical diffusing alpha emitters therapy (DaRT) using 224 Ra-loaded wires was tested on mice bearing prostate (PC-3), glioblastoma (GBM, U87-MG), colon (HCT15), squamous cell carcinoma (FaDu), and melanoma (C32) cancer cell tumors. The in vivo study confirmed that DaRT can destroy tumors, with only the C32 cells exhibiting resistance [100].

212 Pb
The 224 Ra daughter 212 Pb has been used in a variety of preclinical trials. 212 Pblabeled 376.96 monoclonal antibodies were used in mice bearing pancreatic cancer Panc039 xenografts, exhibiting significant uptake and tumor growth inhibition compared to an existing [ 212 Pb]Pb-F3-C25 compound [90]. 212 Pb-labeled 225.28 antibodies were used in immune-deficient mice bearing SUM159 and 2LMP human triple-negative breast cancer cells, showing high cell uptake and effective inhibition of tumor growth for CSPG4expressing tumors [78]. Recently, a variety of low molecular weight ligands were developed and labeled with 212 Pb, and investigated in PSMA(+) PC3 PIP and PSMA(−) PC3 flu flank xenografts. Several of the labeled ligands demonstrated antitumor efficacy, with the kidneys discovered to be the dose-limiting organ [101]. Internalizing and non-internalizing antibodies can have different outcomes. Using internalizing 212 Pb-labeled trastuzumab and non-internalizing [ 212 Pb]Pb-35A7 in mice with intraperitoneal A-431 cell tumor xenografts, the non-internalizing conjugate led to higher tumor dose, while the internalizing led to longer mean survival. This demonstrates the potential advantage of using internalizing TAT [102]. 212 Pb-labeled NG001, a PSMA ligand, was investigated in mice bearing C4-2 tumors, demonstrating a 2.5-fold lower kidney uptake compared to [ 212 Pb]Pb-PSMA-617. This was attributed to the NG001 chelator having all four arms available due to the use of a backbone linker, instead of using a chelator arm for the linking purpose. [103].
6.1.6. 227 Th 227 Th has been conjugated to antibodies including trastuzumab, rituximab, and has demonstrated a significant delay in tumor growth and prolonged survival in breast, ovarian, and lymphoma models [3]. [ 227 Th]Th-rituximab was investigated in mice with human lymphoma Raji xenografts. The maximum tolerated dose was found to be 600-1000 kBq/kg, with a 1000 kBq/kg dose resulting in significant weight loss and temporary drop in white blood cell and platelet counts, with no significant signs of toxicity in examined tissue [85]. Lintuzumab, an anti-CD33 antibody, has been used extensively in TAT for treatment of myeloid leukemia, and has been conjugated to 213 Bi, 225 Ac, and 227 Th. [3]. [ 227 Th]Thlintuzumab induced cytotoxicity in CD33-positive cells and demonstrated antitumor activity in a HL-60 cell mouse model with a single dose regimen [86].
6.1.7. 211 At 211 At has been investigated for non-small cell lung cancer using an 211 At-labeled octreotide somatostatin analogue. In mice, [ 211 At]At-SPC-octreotide exhibited higher uptake in the lung, spleen, stomach, and intestines with rapid clearance 24 h post-injection, while demonstrating tumor cell apoptosis in a dose-dependent manner [91]. 211 At-labeled anti-Frizzled homolog 10 antibody, [ 211 At]At-OTSA101, has been shown to suppress the growth of synovial sarcoma xenografts in mice with greater efficiency than β − emitter [ 90 Y]Y-OTSA101 [92]. Anti-HER2 single domain antibodies and nanobodies have been evaluated, with iso-[ 211 At]At-SAGMB-5F7 demonstrating high intracellular trapping of radioactivity and greater than 10:1 tumor-to-normal organ ratios except for the kidneys and lungs by 2 h post-injection [104]. Another compound, [ 211 At]At-SAGMB-2Rs15d, demonstrated high tumor uptake, low background signals, and fast renal excretion [105]. 211 At has also been conjugated to anti-CD38 and anti-CD45 antibodies. 211 At-CD38 produced a sustained remission and long-term survival (>150 days) for 50 to 80% of mice with multiple myeloma xenografts, compared to untreated mice dying in 20 to 55 days [106].
[ 211 At]At-anti MICA/B antibodies have shown significant reduction in the tumor growth rate of HCT116 p53 −/− xenografts in mice. Systemic cancers such as B-cell lymphoma can be treated by targeting the CD20 surface antigen [107].6.1.8. 149 Tb Figure 6 depicts PET/CT images of an AR42J tumor-bearing mouse after injection with [ 149 Tb]Tb-DOTANOC. 149 Tb was first used in a SCID mouse model of leukemia with 149 Tb-labeled rituximab. In total, 89% of treated mice experienced tumor-free survival of over 4 months, a significant increase compared to other groups [108]. Since then, the 149 Tb-labeled DOTA-folate conjugate has been evaluated in folate receptor (FR)-positive cancer in KB-tumor-bearing mice, with an observed dose-dependent effect, significant tumor growth delay, and no signs of acute toxicity to the kidneys or liver [36]. 149 Tb is also a positron emitter; thus, it was investigated for its utility in PET imaging using [ 149 Tb]Tb-DOTANOC in a mouse with AR42J tumor xenografts, producing distinct visualization of the tumors with residual radioactivity in the kidneys and bladder [59]. The potential for 149 Tb alpha-PET would make it attractive for clinical applications, allowing the direct visualization of therapeutic 149 Tb-radioligands in vivo [24]. There are currently no listed clinical trials for 149 Tb; additional preclinical studies and increased supply sources may enable future clinical trials.

Clinical Studies
In the past decade, there has been a large increase in TAT clinical trials using an array of α-emitters for treating a variety of cancers. Notably, many trials listed on clinical-trials.gov are currently underway, recruiting, or are not yet open for recruitment. A summary of completed trials and those underway or about to begin is given in this section.
Pharmaceutics 2021, 13, x FOR PEER REVIEW 17 of 27 149 Tb alpha-PET would make it attractive for clinical applications, allowing the direct visualization of therapeutic 149 Tb-radioligands in vivo [24]. There are currently no listed clinical trials for 149 Tb; additional preclinical studies and increased supply sources may enable future clinical trials.

Clinical Studies
In the past decade, there has been a large increase in TAT clinical trials using an array of α-emitters for treating a variety of cancers. Notably, many trials listed on clinical-trials.gov are currently underway, recruiting, or are not yet open for recruitment. A summary of completed trials and those underway or about to begin is given in this section. Figure 7 depicts the results from [ 225 Ac]Ac-PSMA-617 therapy in a human patient, which overcame tumor resistance to [ 177 Lu]Lu-PSMA-617. There are nine clinical trials listed on clinical-trials.gov for 225 Ac, one of which is a completed phase 1 trial using 225 Aclabeled humanized anti-CD33 HuM195 antibodies for advanced myeloid malignancies including leukemia and myelodysplastic syndrome. 225 Ac was considered as an alternative to 213 Bi for its greater cytotoxic potential and longer half-life. [ 225 Ac]Ac-HuM195 eliminated peripheral blasts in 63% of patients at doses of 37 kBq/kg or higher, and reduced bone marrow blasts in 67% of patients [87]. 225 Ac salvage therapy overcame βresistance in two metastatic-CRPC patients after a PSMA-positive tumor phenotype was verified by [ 68 Ga]Ga-PSMA-11 PET/CT. Both patients exhibited a complete response, and PSA dropped below the measurable level, with side effects of xerostomia observed in both patients. A further study with 14 metastatic CRPC patients identified 100 kBq/kg to be an appropriate balance between toxicity and antitumor activity [16]. While [ 177 Lu]Lu-PSMA-617 has improved overall survival of CRPC patients, it has been suggested that further studies with 225 Ac PSMA compounds will elucidate their distinct advantages over βtherapy with 177 Lu [109,110]. A pilot study using [ 225 Ac]Ac-PSMA-617 in patients heavily pretreated with chemotherapy led to a greater than 90% decline in PSA serum in 82% of patients, including 41% of patients with undetectable serum PSA who remained in remission 12 months after therapy [111]. A more recent study reported a single cycle of [ 225 Ac]Ac-DOTATOC achieving a partial remission in a patient with refractory metastatic neuroendocrine tumors without any adverse effects after resistance to 10 cycles of the β --emitters There are nine clinical trials listed on clinical-trials.gov for 225 Ac, one of which is a completed phase 1 trial using 225 Aclabeled humanized anti-CD33 HuM195 antibodies for advanced myeloid malignancies including leukemia and myelodysplastic syndrome. 225 Ac was considered as an alternative to 213 Bi for its greater cytotoxic potential and longer half-life. [ 225 Ac]Ac-HuM195 eliminated peripheral blasts in 63% of patients at doses of 37 kBq/kg or higher, and reduced bone marrow blasts in 67% of patients [87]. 225 Ac salvage therapy overcame β − resistance in two metastatic-CRPC patients after a PSMA-positive tumor phenotype was verified by [ 68 Ga]Ga-PSMA-11 PET/CT. Both patients exhibited a complete response, and PSA dropped below the measurable level, with side effects of xerostomia observed in both patients. A further study with 14 metastatic CRPC patients identified 100 kBq/kg to be an appropriate balance between toxicity and antitumor activity [16]. While [ 177 Lu]Lu-PSMA-617 has improved overall survival of CRPC patients, it has been suggested that further studies with 225 Ac PSMA compounds will elucidate their distinct advantages over β − therapy with 177 Lu [109,110]. A pilot study using [ 225 Ac]Ac-PSMA-617 in patients heavily pretreated with chemotherapy led to a greater than 90% decline in PSA serum in 82% of patients, including 41% of patients with undetectable serum PSA who remained in remission 12 months after therapy [111]. A more recent study reported a single cycle of [ 225 Ac]Ac-DOTATOC achieving a partial remission in a patient with refractory metastatic neuroendocrine tumors without any adverse effects after resistance to 10 cycles of the β − -emitters [ 177 Lu]Lu-DOTATATE and [ 90 Y]Y-DOTATOC [76]. Ongoing trials with 225 Ac include a phase 1 study of [ 225 Ac]Ac-lintuzumab in patients with refractory multiple myeloma; an early phase 1 trial of [ 225 Ac]Ac-PSMA radioligand therapy of metastatic castration-resistant prostate cancer; a phase 1 study that targets type I insulin-like growth factor receptor using [ 225 Ac]Ac-FPI-1434 injection for patients with advanced refractory solid tumors; an early phase 1 trial of [ 225 Ac]Ac-PSMA radioligand therapy of metastatic castration-resistant prostate cancer; a phase 1 and 2 trial with [ 225 Ac]Ac-lintuzumab in older acute myeloid leukemia patients; a phase 1 trial using [ 225 Ac]Ac-lintuzumab in combination with CLAG-M chemotherapy in patients with relapsed/refractory acute myeloid leukemia; a phase 1 and 2 trial of venetoclax and [ 225 Ac]Ac-lintuzumab in acute and relapsed myeloid leukemia patients; and a phase 1 and 2 trial venetoclax, azacytidine, and [ 225 Ac]Ac-lintuzumab in acute myeloid leukemia patients.

225 Ac
[ 177 Lu]Lu-DOTATATE and [ 90 Y]Y-DOTATOC [76]. Ongoing trials with 225 Ac include a phase 1 study of [ 225 Ac]Ac-lintuzumab in patients with refractory multiple myeloma; an early phase 1 trial of [ 225 Ac]Ac-PSMA radioligand therapy of metastatic castration-resistant prostate cancer; a phase 1 study that targets type I insulin-like growth factor receptor using [ 225 Ac]Ac-FPI-1434 injection for patients with advanced refractory solid tumors; an early phase 1 trial of [ 225 Ac]Ac-PSMA radioligand therapy of metastatic castration-resistant prostate cancer; a phase 1 and 2 trial with [ 225 Ac]Ac-lintuzumab in older acute myeloid leukemia patients; a phase 1 trial using [ 225 Ac]Ac-lintuzumab in combination with CLAG-M chemotherapy in patients with relapsed/refractory acute myeloid leukemia; a phase 1 and 2 trial of venetoclax and [ 225 Ac]Ac-lintuzumab in acute and relapsed myeloid leukemia patients; and a phase 1 and 2 trial venetoclax, azacytidine, and [ 225 Ac]Ac-lintuzumab in acute myeloid leukemia patients.  [82]. Another study used 213 Bi-labeled anti-EGFR monoclonal antibodies in patients with carcinoma in situ of the bladder, achieving complete remission in 3 of 12 patients with no adverse effects observed and blood and urine parameters remaining in normal ranges [93].  [82]. Another study used 213 Bi-labeled anti-EGFR monoclonal antibodies in patients with carcinoma in situ of the bladder, achieving complete remission in 3 of 12 patients with no adverse effects observed and blood and urine parameters remaining in normal ranges [93].

223 Ra
Having received FDA approval in May 2013 to treat castration-resistant prostate cancer under the name Xofigo, 223 Ra-dichloride has been studied extensively, with 108 listed clinical trials on the clinicaltrials.gov website. 223 Ra-dichloride resulted in pain relief and a reduction in alkaline phosphatase in the first clinical trial in patients with skeletal metastases. Over 50% of patients reporting pain relief 8 weeks after injection, and median survival exceeded 20 months [112]. One 223 Ra-dichloride phase III trial with 921 patients found that the time to first symptomatic skeletal event (15.6 months) was significantly longer compared to the placebo group (9.8 months). The risks of external beam radiation therapy for bone pain and spinal cord compression were also found to be reduced [113]. Another study in CRPC patients found overall survival was significantly longer with 223 Ra-dichloride at 14.9 months compared to 11.3 months with a placebo. 223 Ra treatment was associated with low myelosuppression and reduced adverse events. [114]. These studies collectively found that 223 Ra-dichloride is well tolerated and minimally toxic, and patients have reported an increase in quality of life. Despite these benefits, 223 Ra does not target soft tissue or circulating disease components [3]. However, in 2018, the European Medicines Agency recommended restricting the use of Xofigo to patients who had two previous treatments for metastatic prostate cancer, or who could not receive other treatments. The agency concluded that Xofigo should not be used in combination with Zytiga, prednisone, or prednisolone, since it was observed to reduce survival and lead to additional bone fractures in patients also taking Zytiga [115].

223 Ra
Having received FDA approval in May 2013 to treat castration-resistant prostate cancer under the name Xofigo, 223 Ra-dichloride has been studied extensively, with 108 listed clinical trials on the clinicaltrials.gov website. 223 Ra-dichloride resulted in pain relief and a reduction in alkaline phosphatase in the first clinical trial in patients with skeletal metastases. Over 50% of patients reporting pain relief 8 weeks after injection, and median survival exceeded 20 months [112]. One 223 Ra-dichloride phase III trial with 921 patients found that the time to first symptomatic skeletal event (15.6 months) was significantly longer compared to the placebo group (9.8 months). The risks of external beam radiation therapy for bone pain and spinal cord compression were also found to be reduced [113]. Another study in CRPC patients found overall survival was significantly longer with 223 Ra-dichloride at 14.9 months compared to 11.3 months with a placebo. 223 Ra treatment was associated with low myelosuppression and reduced adverse events. [114]. These studies collectively found that 223 Ra-dichloride is well tolerated and minimally toxic, and patients have reported an increase in quality of life. Despite these benefits, 223 Ra does not target soft tissue or circulating disease components [3]. However, in 2018, the European Medicines Agency recommended restricting the use of Xofigo to patients who had two previous treatments for metastatic prostate cancer, or who could not receive other treatments. The agency concluded that Xofigo should not be used in combination with Zytiga, prednisone, or prednisolone, since it was observed to reduce survival and lead to additional bone fractures in patients also taking Zytiga [115]. Nine of these trials are utilizing diffusing alpha radiation emitters therapy for the treatment of skin cancer, mucosal neoplasms of the oral cavity, soft tissue neoplasms, pancreatic cancer, squamous cell carcinoma, and breast cancer. The other two trials are using Radspherin ® , which consists of a 224 Ra α-emitting calcium carbonate microsphere for patients with peritoneal carcinoma, colorectal carcinoma, and ovarian cancer. A recent DaRT study used 224 Ra seeds to treat squamous cancers of the skin and head and evaluated early tumor responses 30 to 45 days post insertion, with complete response to the treatment observed in 22 of 28 patients [94].

212 Pb
There are two clinical trials listed for 212 Pb, with a phase 1 safety study using [ 212 Pb]Pb-TCMC-trastuzumab in patients with HER2-expressing ovarian cancer confined to the peri-toneal cavity. After intraperitoneal injection, minimal myelosuppression and radiopharmaceutical redistribution outside the peritoneal cavity were observed; however, no patient exhibited a partial response [116]. The other trial is a phase 1 study using AlphaMedix™ ([ 212 Pb]Pb-DOTAMTATE) in patients with metastatic somatostatin-receptor-positive neuroendocrine tumors.

212 Pb
There are two clinical trials listed for 212 Pb, with a phase 1 safety study using [ 212 Pb]Pb-TCMC-trastuzumab in patients with HER2-expressing ovarian cancer confined to the peritoneal cavity. After intraperitoneal injection, minimal myelosuppression and radiopharmaceutical redistribution outside the peritoneal cavity were observed; however, no patient exhibited a partial response [116]. The other trial is a phase 1 study using Al-phaMedix™ ([ 212 Pb]Pb-DOTAMTATE) in patients with metastatic somatostatin-receptorpositive neuroendocrine tumors.

227 Th
There are four clinical trials using 227 Th listed. They include a phase 1 trial using the thorium labeled PSMA-TTC (BAY 2315497) immunoconjugate consisting of a human anti-PSMA antibody covalently linked to the chelator moiety (3,2 HOPO) radiolabeled to 227 Th for use in patients with metastatic castration resistant prostate cancer [80]; a phase 1 trial using the 227 Th-labeled antibody-chelator conjugate MSLN-TTC (BAY 2287411) in patients with mesothelin expressing tumors; and a phase 1 study using the 227 Th-labeled antibody BAY2701439 in patients with breast, gastric, gastroesophageal, and other cancers expressing HER2. The fourth, a now complete phase I trial, used [ 227 Th]Th-epratuzumab (BAY1862864) in patients with relapsed or refractory CD-22-positive non-Hodgkin lymphoma.

211 At
There are multiple 211 At clinical trials, with a phase I and II trial using [ 211 At]At-labeled BC8-B10 monoclonal antibodies for the treatment of nonmalignant diseases in patients undergoing hematopoietic cell transplant, two trials both in phase I and II using [ 211 At]At-labeled BC8-B10 antibodies in patients with high-risk myeloid leukemia, acute lymphoblastic leukemia, myelodysplastic syndrome [117], or mixed-phenotype acute leukemia, and a completed trial with 18 patients using the 211 At-labeled ch81C6 antibody with primary or metastatic brain tumors. In this trial, no patients experienced dose-limiting toxicity, there was no identified maximum tolerated dose, and median survival increased to 54.1 weeks from 23 weeks. Another clinical trial evaluated the absorbed dose and in-

227 Th
There are four clinical trials using 227 Th listed. They include a phase 1 trial using the thorium labeled PSMA-TTC (BAY 2315497) immunoconjugate consisting of a human anti-PSMA antibody covalently linked to the chelator moiety (3,2 HOPO) radiolabeled to 227 Th for use in patients with metastatic castration resistant prostate cancer [80]; a phase 1 trial using the 227 Th-labeled antibody-chelator conjugate MSLN-TTC (BAY 2287411) in patients with mesothelin expressing tumors; and a phase 1 study using the 227 Th-labeled antibody BAY2701439 in patients with breast, gastric, gastroesophageal, and other cancers expressing HER2. The fourth, a now complete phase I trial, used [ 227 Th]Th-epratuzumab (BAY1862864) in patients with relapsed or refractory CD-22-positive non-Hodgkin lymphoma.

211 At
There are multiple 211 At clinical trials, with a phase I and II trial using [ 211 At]Atlabeled BC8-B10 monoclonal antibodies for the treatment of nonmalignant diseases in patients undergoing hematopoietic cell transplant, two trials both in phase I and II using [ 211 At]At-labeled BC8-B10 antibodies in patients with high-risk myeloid leukemia, acute lymphoblastic leukemia, myelodysplastic syndrome [117], or mixed-phenotype acute leukemia, and a completed trial with 18 patients using the 211 At-labeled ch81C6 antibody with primary or metastatic brain tumors. In this trial, no patients experienced doselimiting toxicity, there was no identified maximum tolerated dose, and median survival increased to 54.1 weeks from 23 weeks. Another clinical trial evaluated the absorbed dose and investigated the toxicity of [ 211 At]At-MX35 F(ab') 2 . It targeted the sodiumdependent phosphate transport protein 2b in patients with complete clinical remission after second-line chemotherapy for recurrent ovarian carcinoma. This trial established that intraperitoneal administration of [ 211 At]At-MX35 F(ab') 2 could achieve therapeutic doses without significant toxicity in microscopic tumor clusters [118].

Future of Alpha Therapy
The future of α-therapy holds significant promise for therapeutic clinical applications. Key to its ongoing success is the expansion of robust and high-yield production routes to enhance α-emitter availability, the development of new chelators, linkers, and vectors to enhance efficacy and targeting specificity, and finding solutions to progeny redistribution induced toxicity.
With additional α-emitter production, larger scale preclinical and clinical trials with αemitters should become possible. The construction of new high-energy particle accelerators should also support more preclinical studies and clinical trials of the appealing 149 Tb α-emitter. This could lead to more widespread use of TAT beyond FDA approved 223 Radichloride, enabling the targeting of a wide range of soft tissue and circulatory disease components.
Controlling the progeny is a challenge with existing α-emitters. Despite the ability to deliver significant dose through a nanogenerator approach, the cytotoxic effects of α-emitter progeny in the 225 Ac, 227 Th, 224 Ra, and 211 At decay chains have shown that balancing tolerable dose with cytotoxic effects can be difficult to achieve. Some solutions to control the daughters or hasten their excretion have included radionuclide impregnated wires, cellular internalization, nanocarriers, metal chelation therapy, and diuretics. Alternatively, the absence of α-emitting progeny in the 149 Tb decay chain could warrant its preference over currently employed α-emitters once its availability improves.
TAT would also benefit from more investigation into dosimetry and radiobiology. Given the majority of DNA lesions along an α-particle path are double strand breaks, this differentiates α-therapy from other classes of therapeutic radionuclides that primarily cause single strand breaks and can be rendered ineffective due to cellular adaptative and resistance mechanisms. Determining an effective microdosimetry method and improving the understanding of α-emitter radiobiology is important for optimizing their efficacy and safe clinical implementation.

Conclusions
The short-range high LET characteristics of α-emitter radiotherapy hold promise in being an effective therapy for a variety of cancers. The highly cytotoxic DNA doublestrand breaks and secondary cross-dose and bystander effects give α-emitters a formidable advantage over other forms of radiotherapy, including β − or auger electron therapy. Supported by ongoing in vitro and in vivo advances in radiochemistry and efficacy in preclinical models, existing clinical trials have demonstrated its feasibility in patients with many additional clinical trials underway or scheduled to start soon. To date, αemitting isotopes have been delivered in free form, with targeting vectors or carriers designed to target specific receptors on cells and limit the spread of the daughters, and the more recent DaRT approach of implantable controlled release. Despite uncertainties in α-therapy regarding optimal patient dose and unintended organ toxicity, preclinical animal models have demonstrated marked improvements. Further advances in production and availability, along with management of the radionuclide progeny, should allow for the development and more cost-effective clinical adoption of TAT. There is immense potential for TAT to complement existing forms of therapy and improve the treatment options and quality of life of patients with highly resistant and late-stage diseases. Funding: The authors would like to thank the Dianne and Irving Kipnes Foundation for supporting this work.

Data Availability Statement:
No new data were created or analyzed in this study. Data sharing is not applicable to this article.