Radiometals in Imaging and Therapy: Highlighting Two Decades of Research

The present article highlights the important progress made in the last two decades in the fields of molecular imaging and radionuclide therapy. Advancements in radiometal-based positron emission tomography, single photon emission computerized tomography, and radionuclide therapy are illustrated in terms of their production routes and ease of radiolabeling. Applications in clinical diagnostic and radionuclide therapy are considered, including human studies under clinical trials; their current stages of clinical translations and findings are summarized. Because the metalloid astatine is used for imaging and radionuclide therapy, it is included in this review. In regard to radionuclide therapy, both beta-minus (β−) and alpha (α)-emitting radionuclides are discussed by highlighting their production routes, targeted radiopharmaceuticals, and current clinical translation stage.


Introduction
External radiation therapy is a common and effective treatment for various cancers.Radiation therapy was first applied over 100 years ago, immediately after the discovery of the X-ray [1].Clinicians used it for the treatment of skin diseases, lupus, and other lesions [2][3][4][5].However, due to the collateral damage and resulting side effects, including hair loss, blurry vision, and dry and itchy skin, its application stalled and triggered the need for an alternative treatment option [6].
In the last two decades, nuclear medicine, including radionuclide therapy, has observed significant growth and development: newer radiopharmaceuticals have been introduced, and their applications in imaging and targeted radionuclide therapy have been diverse [7].Targeted radionuclide therapy (TRT) has great potential to destroy even the smallest clusters of metastatic cancer cells present anywhere in the body, which is hard to achieve with either external beam radiation therapy or surgery.Therefore, TRT has been clinically used to treat numerous malignancies, such as neuroblastoma, along with breast, thyroid, and prostate cancer [7][8][9].Treatments using TRT have been explored with and without additional treatment options, such as surgery and chemotherapy.

Diagnostic Radionuclides
PET and SPECT are radionuclide-based imaging modalities used routinely in nuclear medicine practice that fall under the category of "molecular imaging" because their radiotracers provide information about particular biological processes at the cellular and molecular levels.
(i) PET measures the energy produced by the two gamma photons (511 keV) that result from annihilation of the positron emitted from the PET radionuclide with atomic electron [11].The emitted gamma photons are detected with γ-cameras, also called scintillation detectors, which produce reconstructed three-dimensional images depicting the spatial distribution of radiotracers [11].The common examples of PET probes includes [ 18 F]FDG, [ 13 N]NH 3 , [ 68 Ga]Ga-PSMA, and [ 18 F]NaF.Preclinical animal PET and clinical PET scanners offer spatial resolution of 1-2 mm and 6-10 mm, respectively, with high sensitivity of 10 −11 -10 −12 mol/L.This level of sensitivity is sufficient to detect biological changes in an organ or tissue to identify the onset of a disease before anatomical changes occur [12].(ii) SPECT measures the single gamma photons emitted directly from γ-emitting radionuclides called SPECT radiopharmaceuticals.The conventional clinical SPECT scanners have lower sensitivity (10 −10 -10 −11 mol/L) and lower spatial resolution (7-15 mm) compared to PET scanners due to the limited performance of collimators [12][13][14] .Despite this, SPECT is the most routinely used nuclear imaging procedure in the clinic and is less expensive compared to PET.The most common SPECT isotopes are 111 In, 99m Tc, 123/131/125 I, and 67 Ga.
Recent advances in SPECT γ-cameras, collimators, and reconstruction algorithms have enhanced the spatial resolution and sensitivity of SPECT scanners, allowing for the imaging of a wide range of isotope energy (20-300 keV) [15,16].Nevertheless, both PET and SPECT need either computed tomography (CT) or magnetic resonance imaging (MRI) for accurate anatomical information.Interestingly, PET cannot distinguish between two different PET probes when injected simultaneously because it measures two γ-rays with the same energy (511 keV); meanwhile, SPECT does have multiplexing capabilities because each radionuclide produces different γ-rays, enabling it to image different targets simultaneously [17].
(i) Beta minus emitters can be either of a high energy ( 90 Y, E β − max = 2.28 MeV,) or a low energy ( 177 Lu, E β − max = 496 keV), with tissue penetration ranges between 12 mm and 1.5 mm, respectively [18].Given the long penetration depth of 0.2-12 mm and the moderate linear energy transfer (LET) radiation of ~0.2 keV/µm, β − emitters are more suited to treating large-sized tumors (>0.5 cm), and they are considered the current gold standard in targeted radionuclide therapy [19,20].
(ii) Alpha emitters emit α particles with high LET energies of 50-230 keV/µm and shorter penetration depths of 50-100 µm (i.e., 5-10 cell diameters) [21].Alpha radionuclidebased targeted therapy is called targeted alpha therapy (TAT), and it is well suited for the treatment of hematological disease, small tumors, metastasis, and isolated cancer cells.Alpha emitters are perceived as a better therapeutic alternative to beta emitters due to their high LET and short tissue penetration range.(iii) Meitner−Auger electrons are low-energy electrons that can penetrate up to the subcellular nanometer range (<0.5 µm), resulting in a high LET of 4-26 keV/µm [22].
Given the low tissue penetration range and high LET in an extremely small area, MAE emitters could be highly valuable for treating metastatic cancers if delivered selectively within the nucleus of the cancer cells [22]. Figure 1 explains the difference between LET, pathlength (penetration range), and the usefulness of α and β − radionuclide therapies.
Pharmaceuticals 2023, 16, x FOR PEER REVIEW 3 of 47 (ii) Alpha emitters emit α particles with high LET energies of 50-230 keV/µm and shorter penetration depths of 50-100 µm (i.e., 5-10 cell diameters) [21].Alpha radionuclidebased targeted therapy is called targeted alpha therapy (TAT), and it is well suited for the treatment of hematological disease, small tumors, metastasis, and isolated cancer cells.Alpha emitters are perceived as a better therapeutic alternative to beta emitters due to their high LET and short tissue penetration range.(iii) Meitner−Auger electrons are low-energy electrons that can penetrate up to the subcellular nanometer range (<0.5 µm), resulting in a high LET of 4-26 keV/µm [22].
Given the low tissue penetration range and high LET in an extremely small area, MAE emitters could be highly valuable for treating metastatic cancers if delivered selectively within the nucleus of the cancer cells [22]. Figure 1 explains the difference between LET, pathlength (penetration range), and the usefulness of α and β − radionuclide therapies.

General Information
Among the long list of radioisotopes of copper (Cu), 60 Cu, 61 Cu, 62 Cu, and 64 Cu are used for diagnostic imaging, while 64 Cu and 67 Cu are applied in radionuclide therapy [23]. 64Cu decays by both β + (~17%) and β − (~38%), making it applicable for PET imaging and targeted radionuclide therapy; therefore, it is considered a theranostic radionuclide [24].Among the long list of radioisotopes of copper (Cu), 60 Cu, 61 Cu, 62 Cu, and 64 Cu are used for diagnostic imaging, while 64 Cu and 67 Cu are applied in radionuclide therapy [23]. 64Cu decays by both β + (~17%) and β − (~38%), making it applicable for PET imaging and targeted radionuclide therapy; therefore, it is considered a theranostic radionuclide [24].In addition, 64 Cu also decays by electron capture (EC), which results in a cascade of Auger electrons [22,24].The decay characteristics of Cu radioisotopes are mentioned in    # Data on 60/61/64/67 Cu are from [24] and data on 62 Cu are from [25].Please refer to Scheme 1A.Scheme 1. Decay scheme for various radionuclides.
Considering the longer half-life of 64 Cu, [ 64 Cu]Cu-ATSM was applied as a hypoxia imaging radiotracer in rectal cancer (National Clinical Trial (NCT) 03951337) [32].However, several debatable preclinical studies highlighted its lower uptake in hypoxic tumors [28].The therapeutic potential of [ 64 Cu]Cu-ATSM was first studied in 2001, where it resulted in a six-fold increase in the survival of 50% of hamsters bearing human GW39 colon cancer [33].Later, several additional preclinical studies supported the theranostic potential of [ 64 Cu]Cu-ATSM to treat various colon carcinoma xenografts (Colon-26, HT-29).In addition to preclinical studies, clinical studies are needed to prove its true theranostic value [34][35][36].
In addition to radiolabeled somatostatin-targeting peptides, a series of PSMA ligands have been identified and radiolabeled with Cu radioisotopes for clinical diagnosis and radionuclide therapy applications in PCa [50][51][52].In 2016, 64 Cu-labeled PSMA-617 became the first 64 Cu-labeled ligand for PET imaging of PCa patients and was investigated at two nuclear medicine centers (Vienna, Austria, and Bed Berka, Germany) [53].Even though [ 68 Ga]Ga-PSMA is an excellent tracer to detect PCa and metastatic lesions in the lymph node or bone at low PSA levels [54], the advantage of the lower positron energy of 64 Cu (E β + avg = 278 keV) vs. 68 Ga (E β + avg = 829 keV) and the longer half-life (12.7 h) of 64 Cu allow its distribution and use as [ 64 Cu]Cu-PSMA-617 at various clinical PET centers with no sophisticated onsite radiotracer production facility [53]. 67Cu is one of the most promising radionuclides for radioimmunotherapy (RIT), as its 61.8 h isotopic half-life is well matched with the residence time of a typical Ab on the tumor site.In 1998, DeNardo reported a pilot study of [ 67 Cu]Cu-2IT-BAT-Lym-1 to image and treat chemo-resistive B-cell in non-Hodgkin's lymphoma while employing favorable SPECT imaging and the remarkable radiotherapeutic effects of 67 Cu-labeled 2IT-BAT-Lym-1 [55].The clinical investigation of Cu radiopharmaceuticals is outlined in Table 2.

Production and Availability
At present, the most common method to produce 64 Cu is proton irradiation of enriched 64 Ni via a 64 Ni(p,n) 64 Cu nuclear reaction in a small−medium-energy biomedical cyclotron [66].The main route to produce 67 Cu for decades had been via a 68 Zn(p,2p) 67 Cu nuclear reaction that utilizes enriched 68 Zn and high-energy proton irradiation (up to 40 MeV), which also coproduces 64 Cu [67].Recently, Mou et al. developed and patented the fabrication of multi-layer targets composed of enriched 70 Zn and 68 Zn that could maximize 67 Cu production yield [68].

General Information
Among the many radioisotopes of Gallium (Ga), 66 Ga, 67 Ga, and 68 Ga are predominantly used in medical applications for the radiolabeling of various biomolecules [69]. 67Ga and 68 Ga are predominantly used in nuclear medicine for SPECT and PET imaging, respectively. 66Ga ((t 1/2 = 9.49 h) is an attractive PET radionuclide with a relatively longer half-life than 68 Ga (t 1/2 = 67.71min).Due to its high positron emission energy (E β+avg = 1750 keV), though, along with the co-emission of higher gamma rays than 68 Ga, 66 Ga suffers from poor image resolution and high radiation exposure to workers, limiting its medical application [70]. 67Ga is one of the longer-lived Ga radioisotopes, and it decays by EC (100%) with multiple gamma emissions, with the most common gamma energies emitted as 93 keV (39%), 184 keV (21%), and 300 keV (17%) for the SPECT imaging [71].[ 67 Ga]Ga-citrate is the most popular radiopharmaceutical of 67 Ga.For several decades, it has been used in the diagnosis of osteomyelitis and other bone infections [72,73].To date, [ 67 Ga]Ga-citrate scintigraphy is used worldwide for the diagnosis of lymphomas [74], lung cancer [75,76], and inflammation of the kidneys [77].The nuclear decay properties of Ga radionuclides are displayed in Table 3. # Data on 66/67/68 Ga are from [78].Please refer to Scheme 1A.

General Information
Zirconium-89 ( 89 Zr) is a promising radionuclide for the PET imaging of Abs due to its longer physical half-life (78.4 h), which matches with the blood half-life of most fulllength Abs (days to weeks) [104]. 89Zr has a relatively short penetration range by emitting low-energy positrons (E β + avg = 396 keV), which facilitate high-resolution PET images [104].
However, 89 Zr emits an abundance of high-energy γ-rays of 909 keV, adding radiation exposure to medical staff and patients [104].Table 5 summarizes the decay characteristics of Zr-89.# Data on 89 Zr are from [105].Please refer to decay Scheme 1A.

Clinical Practice
In 2006, the first clinical study of [ 89 Zr]Zr-immuno PET was reported, where 89 Zrlabeled chimeric mAb U36 localized in all primary tumors and lymph node metastasis of head and neck cancer patients with an accuracy as high as 93% [106].Presently, the FDA has approved hundreds of mAbs against various biological targets, such as HER2, CD20, epidermal growth factor receptor (EGFR), vascular endothelial growth factor (VEGF), and PSMA, resulting in several [ 89 Zr]Zr-Immuno PET-based clinical oncology trials [107,108].Currently, [ 89 Zr]Zr-trastuzumab and [ 89 Zr]Zr-pertuzumab are the two common choices for immuno-PET-targeting HER2+ breast cancer.The first-in-human studies using [ 89 Zr]Zrtrastuzumab (37 MBq) and [ 89 Zr]Zr-pertuzumab (74 MBq) on metastatic breast cancer patients were reported in the years 2010 and 2018, respectively [109,110].Currently, both radiopharmaceuticals are registered in clinical trials.
Additional clinical pilot studies have been published using [ 89 Zr]Zr-immuno-PET probes, such as [ 89 Zr]Zr-bevacizumab targeting VEGF-A expression [111], [ 89 Zr]Zr-rituximab targeting B-lymphocyte antigen (CD20) expression [112], and [ 89 Zr]Zr-cetuximab-targeting EGFR [113], in various tumors.These [ 89 Zr]Zr-immuno-PET probes have been shown to be useful for imaging and/or radionuclide therapy applications.Currently, [ 89 Zr]Zr-bevacizumab is registered in an ongoing clinical trial (National Clinical Trial (NCT) 01894451) [114].Considering the long blood circulation time of monoclonal Abs (mAbs), alternative Abs and their fragments were developed in the last five years to significantly shorten their retention time in blood and to rapidly clear the unbound fragments from the body [115].Early examples of Ab fragment application are minibody-based radiopharmaceuticals, such as [ 89 Zr]Zr-Df-IAB2M, which were used to detect PSMA-positive PCa and recurrent cerebral high-grade gliomas [116,117].In addition to minibody-based radiopharmaceuticals, several preclinical studies reported 89 Zr-labeled affibodies, such as [ 89 Zr]Zr-Df-ZEGFR:03115 and [ 89 Zr]Zr-DFO-MAL-Cys-MZ, as targeting EGFR and HER2, respectively [118,119].To fully evaluate the clinical value of affibodies and Ab fragments, additional clinical studies demonstrating their usefulness are paramount.
Besides Abs, cell labeling with 89 Zr has been explored for the imaging of white blood cells and CAR-T cells.Several preclinical studies using [ 89 Zr]Zr-oxine and [ 89 Zr]Zr-Df-aTCRmu-F(ab') 2 were reported to track T cells in glioblastoma and acute myeloid sarcoma, respectively [120,121].Covalent tethering of [ 89 Zr]Zr-DBN to cells is another highly studied methodology to noninvasively track various cell types with PET.Published reports of this method demonstrate that it offers a robust and reliable approach that could be translated in humans for monitoring cell-based therapies [122][123][124][125]. Table 6 summarizes the clinical applications of 89 Zr-based radiopharmaceuticals.

Production and Availability
The production and availability of 89 Zr have been improved significantly in the last two decades.Various methods of 89 Zr production on solid and liquid targets using cyclotron have evolved over the years, resulting in better and simplified methods of purification and radiolabeling [91,93,[126][127][128][129][130][131][132].The main route of production is proton irradiation of yttrium (Y) via a 89 Y(p, n) 89 Zr nuclear reaction [126].At present, 89 Zr is routinely produced at various academic institutions for their own use, including Mayo Clinic Rochester, and for supplying other institutions, including the University of Wisconsin, the University of Alabama, and commercial vendors within the United States.Some European and Asian academic institutions also manufacture 89 Zr routinely and use it predominantly in preclinical studies.In the last 10 years, several groups have come up with alternative solutions that facilitate the GMP-grade production and formulation of 89 Zr.For example, Wooten et al. designed an automated system for routine 89 Zr production and purification at high radioactivity quantities, with >99.9% of radionuclidic purity [133].In terms of purification, Pandey et al. developed a simplified synthesis of hydroxamate resin for trapping of Zr-89 with a trapping efficiency of 93% and its subsequent elution either as oxalate or phosphate in a high elution efficiency (>90%) [134].Recently, the same group designed a new solid target insert and optimized the thickness of 89 Y foil and proton beam energy to improve the production yield of 89 Zr (~129 mCi or 4.77GBq) using medium energy cyclotrons [126].Others in the field have also significantly contributed towards the advancement of Zr-89 production and purification [135,136].[ 89 Zr]Zr-Df-IAB22M2C CD8 + Tlymphocytes NCT05013099 (Phase IIb; ongoing) NCT03107663 Phase I; completed) NCT03802123 (Phase II; completed) Melanoma [139] Renal cell carcinoma [140] Metastatic solid tumors [141] [ 89   43 Sc, 44 Sc, and 47 Sc are the commonly explored radionuclides for PET imaging and targeted radionuclide therapy applications [150]. 44Sc and 43 Sc are promising PET radionuclides, and they are superior alternatives to 68 Ga because of their lower positron energy and almost 3.5-fold longer half-life [150,151].However, 44 Sc also decays via high gamma ray (E γ = 1157 keV; 99.9% abundance) emission and could give a high radiation exposure dose compared to other competing PET radionuclides [152].The decay properties of Sc radionuclides are given in Table 7.

Current Clinical Application of Scandium-44
In 2017, the first clinical study of generator-derived 44 Sc with [ 44 Sc]Sc-DOTATOC was reported for the imaging of a metastatic neuroendocrine neoplasm at Bed Berka [153].Recently, [ 44 Sc]-PSMA-617 was also applied for the imaging of PCa patients [154], and it showed performance comparable to [ 68 Ga]Ga-PSMA-617 in terms of tumor uptake and image quality [154].Given the availability of both imaging ( 43/44 Sc) and therapeutic ( 47 Sc) radionuclides, Sc radiopharmaceuticals are gaining significant interest as an alternative theranostic pair [155]. 43Sc is another PET isotope of the Sc family with similar physical characteristics to 44 Sc, but it is devoid of high-energy gamma emission and lower positron energy (E β + avg = 476 keV), making it a more favorable imaging isotope than 44 Sc [150].However, no preclinical or clinical studies are yet reported with 43 Sc-labeled radiopharmaceuticals. 47 Sc is a radio theragnostic isotope that emits low-energy β − particles (E β − avg = 162 keV) and low-energy γ-radiations (E γ = 159 keV) [150].The decay characteristics of 47 Sc are like 67 Cu (E β-avg = 141 keV, E γ = 184 keV) and 177 Lu (E β − avg = 134 keV, E γ = 113, 208 keV).

Production and Availability
In 2010, Frank Rosch et al. reported the production of 44 Sc (approx.185MBq) using a 44 Ti/ 44 Sc generator for the first time at Bed Berka, Germany [157].However, the production of the parent radionuclide, 44 Ti, and the accessibility of these generators were challenging.Later, in 2015, Van der Meulen et al. reported a cyclotron-based production of 44 Sc via proton irradiation (11 MeV) of an enriched 44 Ca target, which allowed elution of approximately 2 GBq of 44 Sc at the Paul Scherrer Institute (PSI) in Switzerland [158].Later, Szkliniarz et al. reported several cyclotron-based production routes for emerging 43 Sc using either α particles or deuteron beams because the limited availability of high-energy multi-particle cyclotrons restricted the utility of these production routes [159].Van der Meulen et al. demonstrated the production of 480 MBq of 43 Sc using enriched 43 CaCO 3 and targets via a 43 Ca(p,n) 43 Sc reaction; limited purity of 43 Sc was obtained due to the co-produced mixture of 43 Sc (66.2%) and 44 Sc (33.3%) [160]. 47Sc can be produced using a cyclotron [161], neutron flux reactor [162], or electron linear accelerator [163,164].

General Information
Among the various radioisotopes of Terbium (Tb), four ( 149/152/155/161 Tb) are of great interest in nuclear medicine and are commonly referred as a "Swiss army knife" of nuclear medicine [165]. 149Tb has both positron and α-emission properties for both PET and targeted therapy applications [166]. 152Tb is another positron emitter with a relatively longer half-life of 17.5 h that could be utilized for radiolabeling of large biomolecules [166].The decay characteristics of Tb radionuclides are listed in Table 8.In 2012, Muller et al. performed the first radiolabeling of albumin-binding folate conjugates (cm09) with 149/152/155/161 Tb in an FR-positive tumor xenograft mouse model [167].The findings demonstrated excellent tumor visualization through PET/CT using [ 152 Tb]Tb-cm09 and SPECT/CT, using both [ 155 Tb]Tb-cm09 and [ 161 Tb]Tb-cm09 probes at 24 h post administration.On the other hand, α therapy version [ 149 Tb]Tb-cm09 and β − therapy version [ 161 Tb]Tb-cm09 resulted in significantly delayed tumor growth by 33% and 80%, respectively.
In 2004, Beyer et al. demonstrated the first preclinical RIT with [ 149 Tb]Tb-rituximab in a leukemia xenograft mouse model, which resulted in tumor-free survival >120 days among 89% of the [ 149 Tb]Tb-rituximab-treated mice [169].However, nearly 28% of the residual radioactivity of longer-lived daughter nuclides, 149 Eu (t 1/2 = 93 d), 145 Sm (t 1/2 = 340 d), and others were retained mainly in the mice bone marrow.Baum et al. demonstrated the first in-man PET/CT study of SSTR-targeted [ 149 Tb]Tb-DOTANOC on a male patient diagnosed with neuroendocrine ileum.The result was an excellent localization of [ 149 Tb]Tb-DOTANOC in this neuroendocrine neoplasm, in addition to multiple lymph nodes, skeletal metastasis, and SSTR-expressing organs [170].
(ii) 152 Terbium: 152 Tb is a diagnostic radionuclide that decays via positron emission (E β + avg = 1142 keV) and multiple gamma radiations, which could lead to high radiation exposure [170].The relatively long half-life of 152 Tb (t 1/2 = 17.5 h) allows it to be useful in dosimetry estimation.In fact, 152 Tb is an exact diagnostic match for 149 Tb and 161 Tb, as well as other clinically useful therapeutic radionuclides, like 177 Lu, due to their similarities in coordination chemistry and pharmacokinetics.(iii) 155 Terbium: 155 Tb is a suitable SPECT isotope, a promising alternative to the 111 In isotope, and it could be useful for dosimetry estimation of β − emitters, like 177 Lu, 90 Y, and 166 Ho [173].(iv) 161 Terbium: 161 Tb decays by low-energy (E β − avg = 154 keV) (β − ) emission, having a short tissue penetration (0.29 mm) range and a long half-life (t 1/2 ) of 6.8 d [174].The decay characteristics and half-life of 161 Tb are like 177 Lu (E β − avg = 134 keV, t 1/2 = 6.7d) [174], although 161 Tb also emits a substantial number of auger electrons, which could be advantageous for therapeutic applications.However, the clinical superiority of 161 Tb over 77 Lu is yet to be established [175][176][177].In addition to radionuclide therapy, 161 Tb also emits gamma photons enabling SPECT imaging [174].Recently, Baum et al. demonstrated the first-in-human SPECT imaging using [ 161 Tb]Tb-DOTATOC in patients with paraganglioma and NETs and showed high-quality images and visualization of hepatic metastasis as well as multiple osteoblastic skeletal metastasis in patients [178].
The main constraint to the wider application of Tb isotopes is their availability: there are insufficient production quantities.The production of Tb isotopes requires expensive enriched targets and accelerator-based isotope separation on-line technology (ISOLDE), which is not widely available [166].Table 9 summarizes the clinical investigation of Tb radionuclides.

Production and Availability
In 2012, Muller et al. reported on the production of 149/152/155 Tb in a range of ~6-15 MBq activity through a high-energy proton-induced spallation of tantalum foil targets, followed by dissolution and isotope separation [167].Such a high-energy proton accelerator facility and mass separation technology (ISOLDE) are limited to a few centers worldwide, including CERN, Switzerland.Lately, CERN-MEDICIS (Medical isotopes collected from ISOLDE) technology was developed, which allowed for the production of 38 GBq of 149 Tb, 37 GBq of 152 Tb, and 5.3 GBq of 155 Tb [166]. 161Tb (up to 15 GBq) can be produced in a neutron flux reactor using 160 Gd targets, as proposed by Lehenberger et al. at PSI, Switzerland [181].Interestingly, the production concept and the cost of 161 Tb is like a non-carrier added 177 Lu [181].4.6.Radioisotopes of Zinc 4.6.1.General Information Zinc (Zn) exists in three positron-emitting isotopes ( 62/63/65 Zn) that have the potential to be used as PET biomarkers of zinc trafficking in various pathological conditions [182].Among them, 62 Zn has limited use because it decays to another positron-emitting isotope, 62 Cu (β + = 98%; t 1/2 = 9.7 min), which could confound the image interpretation of PET scans [182].Nevertheless, 62 Zn has been used preclinically to image zinc transport in pancreatic exocrine function [183].
Among the Zn PET isotopes, 65 Zn has the longest half-life (t 1/2 = 243.9d), making it unsuitable for diagnostic imaging because it will cause high radiation exposure to patients over time [182]. 63Zn has a favorable decay characteristic (β + = 93%; t 1/2 = 38.47min) for diagnostic imaging and pharmacokinetic studies [184,185].The decay properties of Zn radionuclides are given in Table 10.[185].Although low uptake of [ 63 Zn]Zn-citrate was seen in the brain (SUV ~0.4) compared to other organs, like the liver, pancreas, kidney, and gastrointestinal tract, it was sufficient to study 63 Zn clearance kinetics on a regional basis in those patients.The regions with slower 63 Zn clearance corresponded to the regions of known amyloid-β pathology on [ 11 C]C-PiB PET scans and also the regions of lower uptake on [ 18 F]FDG-PET scans [185].Further imaging studies are warranted, though, to study zinc homeostasis in persons with Alzheimer's disease.

Production and Availability
In 2014, DeGrado et al. developed a cyclotron-based production of 63 Zn via a 63 Cu(p,n) 63 Zn nuclear reaction using a liquid target by irradiating an isotopically enriched solution of [ 63 Cu]Cunitrate [184]. 63Zn was produced with a specific activity of 41.2 + 18.1 MBq/µg (uncorrected) and radionuclidic purity of 99.9% using 1.23 M of [ 63 Cu]-copper nitrate.

SPECT Probes
5.1.Technetium-99m 5.1.1.General Information Technetium-99m ( 99m Tc) is the most widely used medical isotope in nuclear medicine, accounting for more than 80% of all nuclear medicine procedures, including myocardial perfusion imaging, cancer, and infection imaging [186].99m Tc-based agents are a favored choice for cardiac imaging in the U.S [187].99m Tc mainly disintegrates into its other isomeric 99 Tc (which is radioactive) with the release of low-energy monochromatic gamma rays (140.5 keV, 98.6%) that can be detected by any sensitive gamma cameras [188].Despite the advent of superior PET technology and the prevalence of CT or MRI over nuclear medicine, 99m Tc-based radiopharmaceuticals have been continuously supplied in hospitals during routine clinical examinations [188].The advantages behind them are (i) a short/sufficient half-life of 6 h, which offers minimum radiation exposure to patients, (ii) instant kit-based labeling and formulations due to rich coordination chemistry of Tc (multiple oxidation states), (iii) availability of transportable generators ( 99 Mo/ 99m Tc) for production, and (iv) cost-effective SPECT gamma cameras compared to expensive PET technology.These points have solidified the continuous application of 99m Tc-labeled radiopharmaceuticals [188,189].The nuclear decay characteristics of 99m Tc are given in Table 11.# Data on 99m Tc are from [188].Please refer to Scheme 1B.

Production and Availability
99m Tc is a radioactive decay product of 99 Mo (t 1/2 = 66h), which is traditionally made in a large nuclear reactor via fission of high-enriched uranium targets ( 235 U) [189].The production of 99m Tc in the form of pertechnetate [ 99m Tc]TcO 4 − from the parent 99 Mo was achieved using the commercially available and transportable 99 Mo/ 99m Tc generators in nuclear medicine for the preparation of almost all of the 99m Tc-based radiopharmaceuticals. Until 2011, the global requirement for 99 Mo was fulfilled by seven nuclear research reactors.The mandatory shutdowns of these reactors for maintenance or due to breakdowns stopped the global supply in 2009, 2012, and 2013 [189].To overcome such an unavoidable global crunch in the supply of 99 Mo, several research efforts were initiated, including the use of linear accelerators and cyclotrons, which utilize electron beam and proton irradiation of solid 100 Mo targets, respectively [205,206].

General Information
Indium-111 ( 111 In; t 1/2 = 2.8 d) is a SPECT isotope that decays by EC (100%) and lowenergy γ-emission (171 keV, 245 keV) [207].The decay characteristics are summarized in Table 13.Over the decades, 111 In has been used as the reference standard for SPECTimmuno imaging of Abs [207].110m In is a PET radioisotope of In with a short half-life (69 min), and it is suitable for tracking short peptides (e.g., octreotide) having faster kinetics [208].# Data on 110m /111 In are from [78].Please refer to Scheme 1B.

Clinical Practice
In mid-1978, McAffee and Thakur introduced a radiotracer, [ 111 In]In-oxine, which could be used to radiolabel leukocytes (white blood cells (WBC)) for the scintigraphic detection of focal infections [209].In 1985, the FDA approved [ 111 In]In-oxine-tagged WBC scans for clinical imaging of inflammatory disease [210].The reported sensitivity and specificity of these [ 111 In]In-WBC scans ranged from 60-100% to 69-92%, respectively, in detecting osteomyelitis, vascular grafts infection, bone infections, etc. [211].Other than cell labeling, 111 In was also used in radiolabeling of various peptides, proteins, Abs, and drugs.For example, [ 111 In]In-capromab pendetide (ProstaScint ® ) was FDA-approved for immuno-SPECT imaging of PCa [212]; however, the poor tumor-to-background signals limited its routine clinical use [212].Later, another promising PSMA immuno-SPECT tracer, [ 111 In]In-J591 (PSMA-Ab), was developed, and the first clinical trial was reported in 2005 [213].Clinical trials of [ 111 In]In-J591 are underway and associated with the dosimetric projections of RIT with [ 90 Y]Y-J591 [214].Furthermore, in 2018, Heckman et al. demonstrated the firstin-man study using the novel SPECT tracer [ 111 In]In-DOTA-girentuximab for intraoperative guidance of renal cell carcinoma resection in patients [215].Table 14 summarizes the clinical application of 111 In-labeled radiopharmaceuticals.

Clinical Application of 90 Y
The FDA has approved two types of 90 Y microspheres, TheraSphere TM (glass microspheres) and SIR-spheres ® (resin microspheres), to treat unresectable hepatocellular carcinoma and colorectal metastasis, respectively [227,228].
These 90 Y-microspheres have been used in therapies based on the concept of "radioembolization" (also known as selective internal radiation therapy); it is a promising catheterbased liver-directed therapy approved by the FDA for patients with primary/metastatic liver tumors.It was found that the antitumor effect of 90 Y-microspheres (glass microspheres, also known as Thersphere TM ) are related to beta radiations rather than embolization and therefore proven safer/successful for advanced-stage liver cancer [227].The recent phase III trials of radioembolization of 90 Y-resin microspheres in patients with HCC demonstrated significantly higher tumor response with respect to standard first-line treatment with Sorafenib.However, these results did not meet the primary endpoint, such as overall survival or the patient's quality of life.Several Asian guidelines recommend 90 Y-resin microspheres for HCC treatment based on certain considerations, such as patient selection, treatment planning using accurate dosimetry pre/post-radioembolization, and technical aspects [229][230][231][232].
In 2002, the FDA approved the first anti-CD20 radioimmunoconjugate [ 90 Y]Y-Ibritumomab tiuxetan (Zevalin TM ) for the treatment of advanced B-cell lymphoma as a first line of treatment for rituximab-relapsed or refractory low-grade lymphomas; the overall response rate has ranged from 74% to 82% [233].Despite the demonstrated immunotherapy efficacy of Zevalin TM , it failed commercially due to the underutilized practice by hematologist-oncologists for logistic and economic reasons [234].In addition, other competitive RIT drugs, such as rituximab (anti-CD20) and second-generation mAbs, undoubtedly contributed to the limited sale of Zevalin TM [235,236].
The development of second-generation mAbs, particularly bispecific Abs (e.g., biotin, IgG-single chain variable fragment), have been utilized in an alternative approach called multi-step pre-targeted RIT to enhance the therapeutic efficacy and to diminish its toxicities [237].Based on PRIT technology, [ 90 Y]Y-DOTA-biotin was developed, which makes a strong conjugate with Abs (streptavidin, avidin) present on the tumor.In 1999, Paganelli et al. published the first clinical preliminary results of [ 90 Y]Y-DOTA-biotin for the treatment of high-grade gliomas (n = 48) based on biotin-streptavidin chemistry and showed tumor reduction (>25-100%) in 25% of patients; in 16% of these, the response lasted for at least a year [238].In 2000, a phase II clinical trial of [ 90 Y]Y-DOTA-biotin was reported in patients with metastatic colon cancer [239].Despite evaluating the feasibility, safety, and efficacy of [ 90 Y]Y-DOTA-biotin, the immunogenicity of these types of pre-targeting agents have not been addressed, which in turn caused the clinical trials to end in 2005 [240].
Paganelli et al. developed an innovative therapeutic approach called "Intra-operative avidination for radionuclide therapy" (IART ® ) that relies on a biotin-avidin binding system [241].A phase II study of IART ® in 2010 using [ 90 Y]Y-DOTA-biotin on breast cancer patients demonstrated its potential use immediately after breast resection, thereby shortening the time course of external beam radiotherapy [241].In the past decade, several peptide-based 90 Y-tracers were developed for PRRT, and they are currently under clinical trial.Table 16 summarizes the clinical application of 90 Y radiopharmaceuticals.

Production and Availability
90 Y can be produced from the 90 Sr/ 90 Y generator, where the parent isotope is 90 Sr (t 1/2 = 29 y), and it can be generated as a by-product in large quantities in U-based nuclear reactions [242].Commercial availability and the steady supply of Y-90 are advantageous in conducting Y-90-based clinical trials.

Radioisotopes of Rhenium 6.2.1. General Information
Among several radioisotopes of rhenium (Re), 186 Re and 188 Re are recognized for their therapeutic potential, and they were used to develop various therapeutic radiopharmaceuticals.In addition to beta emission, 186 Re and 188 Re also emit low-abundant γ-rays of 137 keV and 155 keV, respectively (Table 17), that permit scintigraphic monitoring and dosimetry calculations via SPECT imaging [251].# Data on 186/188 Re are from [251].Please refer to Scheme 1C.
Given the two distinct tissue penetration ranges of 188 Re (11 mm) and 186 Re (4.5 mm), they can be selectively applied for treating large-sized tumors and small-or mid-sized tumors, respectively [251].Moreover, to better understand the biodistribution, 99m Tc represents a diagnostic match for 186/188 Re radioisotopes, as both Re and Tc exhibit similar chemical properties [251].However, 99m Tc-and 188 Re-labeled radiotracers do not always show the same in vivo biodistribution [251].

Clinical Applications of Rhenium Radioisotopes
188 Re-labeled therapeutic radiopharmaceuticals have been investigated in multiple clinical trials involving primary tumors, bone metastasis, rheumatoid arthritis, and intracoronary β-brachytherapy [252].In 1998, Maxon et al. evaluated phosphonate-based radiotracer [ 188 Re]Re-HEDP for bone pain palliation [253].Bone pain is a major issue in ~50% of women with breast cancer and 80% of men with PCa.A phase III trial comparing [ 188 Re]Re-HEDP with a well-known bone-targeting agent [ 223 Ra]RaCl 2 is ongoing (NCT03458559).
The primary objective of this study is to compare the overall survival in patients with PCa metastatic to bone after treatment with [ 188 Re]Re-HEDP and [ 223 Ra]RaCl 2 .Several Ab fragments have been radiolabeled with 186/188 Re for RIT.These include alemtuzumab (anti-CD66) in leukemia [254], rituximab (anti-CD20) in lymphoma [255], MN-14 (ant-CEA) in gastrointestinal cancer [256], and bivatuzumab in head and neck cancers [257].Among them, the evaluation of [ 186 Re]Re-bivatuzumab in a variety of diseases (NCT02204033) as a phase I clinical trial has been completed.However, the results are not yet published.Recently, 188 Re-colloids-based brachytherapy kit (Rhenium-SCT ® ) became commercially available to treat basal cell carcinoma or squamous cell carcinoma, particularly to the face and neck, where surgery and radiotherapy are either not possible or refused by patients (NCT05135052) [258].Several preliminary clinical reports have demonstrated that this innovative epidermal therapy is effective in 98% of melanoma patients even after a single application [259].The clinical investigations of Re radiopharmaceuticals are summarized in Table 18.
6.3.Holomium-166 166 Ho is not only a β − emitter but also a gamma emitter; it is one of the lanthanide radionuclides that can be imaged using SPECT and MRI [267,268]. 166Ho is a theranostic radionuclide with favorable physical decay characteristics, including a sufficient half-life of 26.6 h, an average emission energy of (E βav ) of 670 keV, a soft tissue penetration range of 8.7 mm, and a low-energy γ-emission (80.5 keV, 6%) for SPECT imaging [267,268].Being a lanthanide with its paramagnetic properties, 166 Ho-labeled drugs enable the visualization and quantification of the biodistribution of drugs in the tumor tissues by means of SPECT and MRI [268].In 1991, Murphy et al. first investigated the potential possibility of 166 Ho microspheres for the internal radiation therapy of hepatic tumors in rabbits [269].In 2010, Smith et al. investigated the first 166 Ho-based liver radioembolization, which was followed by growing interest in this treatment possibility, as evidenced by the increasing number of publications in the last few years [268].In terms of clinical applications, 166 Homicrospheres serve as an alternative to existing 90 Y microspheres to treat liver tumors, with potential advantages of the shorter half-life of 166 Ho (t 1/2 = 26.6 h) compared to 90 Y (t 1/2 = 64 h) along with its quantification by MRI [268].Additional information on 166 Ho-radiopharmaceuticals have been discussed in a recent review by Klaassen et al. [270]; therefore, we have kept the discussion extremely short.

Clinical Applications
Since 2000, 177 Lu-labeled somatostatin analogues have been utilized in PRRT for the treatment of inoperable or metastatic NETs [272]. 177Lu-labeled somatostatin has sixto seven-fold higher affinity for SSTR2 compared with its 90 Y-loaded counterpart [272].Several preclinical and clinical studies have been conducted on the therapeutic effectiveness of 177 Lu-based radiopharmaceuticals in last two decades [273].In 2005, the first-in-human proof-of-concept study was published on endoradiotherapy with [ 177 Lu]Lu-PSMA-I & T, which was found to be promising in patients with castration-resistant and metastatic prostate cancers [274].
In 2017, the results of a clinical phase 3 trial (NETTER-1) involving 229 patients randomized to either PRRT using [ 177 Lu]Lu-DOTATATE (7.4 GBq every 8 weeks) or a long-acting release (LAR) formulation of octreotide (control groups) to treat patients with midgut NET were released [275,276].The groups receiving [ 177 Lu]Lu-DOTATATE had a significantly higher response rate (18%) and longer progression-free survival (65.2%) at 20 months compared to the controls, with 10.8 and 3%, respectively.[ 177 Lu]Lu-DOTATATE treatment yielded a clinically significant improvement in progression-free survival as a primary end point as well as an improvement in the median survival of 11.7 months [276].Overall, the treatment was well tolerated with grade 3 or 4 adverse events, which were similar in both the groups.No evidence of renal toxicities was observed among patients in the [ 177 Lu]Lu-DOTATATE groups [275,276].In 2018, the U.S. Food and Drug Administration (FDA) and the European Medicines Agency (EMA) approved [ 177 Lu]Lu-DOTATATE (Lutathera ® ; Novartis company) for mid gut NET.In the same year, [ 177 Lu]Lu-PSMA-617 was proposed for the treatment of metastatic castration-resistant prostate cancer (mCRPC).The results of a phase 2 trial (TheraP) demonstrated a significant decline (>50%) in PSA in the groups treated with [ 177 Lu]Lu-PSMA-617 compared to the standard treatment using Carbazitaxel, which eventually led to FDA and EMA approvals under the name of Pluvicto ® (Novartis).The clinical use of 177 Lu-radiopharmaceuticals has been increased in the past few years, including [ 177 Lu]Lu-FAPI-46 [277,278] and combination therapy with 90 Y-labeled peptides and chemotherapy.Several clinical trials of 177 Lu-based radio theranostics are underway, as listed in Table 20.

Production and Availability
There are two common independent ways to produce large-scale 177 Lu in nuclear reactors.The first is a direct production route (also known as carrier added or c.a.) based on neutron irradiation of 176 Lu via 176 Lu (n, γ) 177 Lu nuclear reaction in medium-high-energy reactors [271,288].The second approach is an indirect production route (also known as non-carrier added or n.c.a) based on neutron irradiation of 176 Yb target via 176 Yb (n, γ) 177 Yb → 177 Lu in high-energy flux reactors [288].The advantage of the direct production route is that it can create large quantities of 177 Lu (740-1110 GBq) using 176 Lu; however, the major concern is the co-emission of small amounts of long-lived radioactive impurity of 177m Lu along with "useful" 177 Lu.Additionally, only a part of the target matrix (or carrier) of 176 Lu is converted into the desired 177 Lu, which cannot be chemically isolated as they are the isotopes of the same element; this therefore decreases its specific activity [288].
On the other hand, the indirect approach using highly enriched 176 Yb (>98%) produces high specific activity (>2.96TBq/mg) non-carrier-added 177 Lu; however, this process requires a suitable method for the radiochemical separation of 177 Lu from 176 Yb, which is quite challenging, especially in large-scale or industrial settings, to meet the surging demand [271,288]. 177Lu can also be produced in cyclotron using deuteron beams (<6 MeV); however, this is less explored due to the low production yield [288].

Alpha-Particle-Emitting Radiopharmaceuticals
Alpha radiations are better suited for the treatment of small metastasis due to their short tissue penetration range and high LET per micrometer of tissue compared to β −emitting radionuclide via double strand DNA breaks in cancerous cells, while sparing nearby healthy tissues [21].However, due to the early stages of development of alphatargeted radionuclide therapy, most clinical trials continue to use beta-emitting radionuclide therapy rather than alpha-emitting radionuclide therapy.

Clinical Applications of 213 Bismuth
In 2002, Joseph et al. reported the first proof-of-concept phase I study demonstrating the anti-leukemic effect of 213 Bi conjugated with anti-leukemia Ab HuM195 (Lintuzumab) to treat leukemia patients [290].In a subsequent clinical study (phase I/II) in 2010, complete remission was seen in acute myeloid leukemia patients with sequential administration of [ 213 Bi]Bi-HuM195 (37 MBq/Kg) and the chemotherapy drug cytarabine (Table 22) [291].The promising clinical results with [ 212 Bi]Bi-mAb-TAT initiated its use to treat other cancers, including melanoma, NETs, and glioma [292,293].
In the last two decades, research efforts have facilitated the development of 213 Bibased peptide conjugates for PRRT study.In 2014, Kratochwil et al. reported the first and only radiopeptide therapy with [ 213 Bi]Bi-DOTATOC on NET patients, which was refractory to β − therapy with 90 Y/ 177 Lu-DOTATOC [294].The results indicated that TAT could induce considerable and long-lasting remission in both the primary tumor and liver metastases [294].Another tracer, [ 213 Bi]Bi-PSMA-617, was reported in metastatic castrationresistant PCa patients [295].The remarkable drop in prostate-specific antigen levels from 237 µg/L to 43 µg/L after [ 213 Bi]Bi-PSMA-617 treatment showed the great potential of TAT using [ 213 Bi]Bi-PSMA-617 over conventional β − radionuclide therapy.In addition, TAT may be able to break the radioresistant effect of β − emitters [295].In the past few years, different clinical trials have used 213 Bi-carrying radiopharmaceuticals for the treatment of various diseases.Although the outcomes were encouraging, further investigations are needed to ensure its safety and efficacy in the clinic.  21Bi-1400 209 Tl-2000 209 Pb-600 213 Bi-440 (25.9) # Data on 212 Bi are from [296] and data on 213 Bi are from [297].Please refer to Scheme 2. Scheme 2. Decay scheme for various α-emitting radionuclides.

Production and Availability
Many α-emitter radionuclides are produced from naturally occurring heavy α radionuclides, including U, radium (Ra), and actinium (Ac).The clinical amount of 213 Bi is obtained from its parent radionuclide 225 Ac (t1/2 = 9.9 d) as a 225 Ac/ 213 Bi generator [298].The parent isotope 225 Ac is obtained from the decay of 229 Th (t1/2 = 7317 y), which in turn originates from a decay chain of fissile materials of 233 U [298].The relatively long half-life of the parent radionuclide 225 Ac allows shipment of the 225 Ac/ 213 Bi generator to any radiopharmaceutical facility located even long distances away and permits in-house generation of 213 Bi for radiolabeling purposes over weeks to months.However, the limited global production of 229 Th and the concern for the non-proliferation of the fissile product of 233 U restricted the commercial supply of 225 Ac stocks to produce 213 Bi-labeled radiopharmaceuticals [293].An alternate route to producing 225 Ac is using proton irradiation of 226 Ra targets via 226 Ra (p,2n) 225 Ac in a cyclotron; still, the presence of hazardous 222 Rn poses serious limitations in clinical translation and waste disposal [293].

Production and Availability
Many α-emitter radionuclides are produced from naturally occurring heavy α radionuclides, including U, radium (Ra), and actinium (Ac).The clinical amount of 213 Bi is obtained from its parent radionuclide 225 Ac (t 1/2 = 9.9 d) as a 225 Ac/ 213 Bi generator [298].The parent isotope 225 Ac is obtained from the decay of 229 Th (t 1/2 = 7317 y), which in turn originates from a decay chain of fissile materials of 233 U [298].The relatively long half-life of the parent radionuclide 225 Ac allows shipment of the 225 Ac/ 213 Bi generator to any radiopharmaceutical facility located even long distances away and permits in-house generation of 213 Bi for radiolabeling purposes over weeks to months.However, the limited global production of 229 Th and the concern for the non-proliferation of the fissile product of 233 U restricted the commercial supply of 225 Ac stocks to produce 213 Bi-labeled radiopharmaceuticals [293].An alternate route to producing 225 Ac is using proton irradiation of 226 Ra targets via 226 Ra (p,2n) 225 Ac in a cyclotron; still, the presence of hazardous 222 Rn poses serious limitations in clinical translation and waste disposal [293].  21At, 213 Bi, 213 Po, 209 Tl, 209 Pb) before reaching the stable 209 Bi [300].Overall, 225 Ac decay (t 1/2 = 9.9 d) contributes to the emission of four α-, three β − , and two principal γ-emissions (218 keV; 221 Fr, 440 keV; 213 Bi), from which recognizable 225 Ac results as a "nanogenerator." 225Ac is also considered an in vivo generator of 213 Bi and an alternative to 213 Bi-based TAT, presumably because of the four αemissions and its longer half-life compared to 213 Bi (t 1/2 = 45.6 min) [300].The decay characteristics of Bi radionuclides are given in Table 23.  22Ac-5.8 221 Fr-6.3 217 At-7.1 213 Bi-5.9 213 Po-8.4 213 Bi-0.492 209 Tl-0.178 209 Pb-0.198 213 Bi-100 (1) 221 Fr-218 (11.4) 213 Bi-440 (26) 209 Tl-1567 (99.7) # Data on 225 Ac are from [300].Please refer to Scheme 2. 225 Ac-based radiopharmaceuticals are prone to the redistribution of daughter progenies, particularly 213 Bi, which can induce renal toxicity and dose-limiting toxicity to other organs [300].Moreover, dosimetry is essential using an isotope with a similar half-life and chelation chemistry to 225 Ac (e.g., Ln 3+ ) to track the biodistribution of 225 Ac accurately.A handful of clinical trials of 225 Ac are underway.
By 2018, a multicenter phase I study using [ 225 Ac]Ac-FPI-1434 (NCT03746431) was designed to treat solid tumors from non-cell lung, prostate, and breast carcinomas [302].Recently, a clinical study reported by Kratochwil et al. demonstrated the remarkable antitumor effect of [ 225 Ac]Ac-PSMA-617 (100 kBq/kg) in 81% of metastatic castration-resistant PCa patients [303].Additional clinical trials are warranted to further investigate the antitumor potential of [ 225 Ac]Ac-PSMA-617 TAT in men with prostate cancer (NCT04597411).The clinical investigation of 225 Ac radiopharmaceuticals is summarized in Table 24.

Production and Availability
Currently, the clinical supply of 225 Ac is produced from 229 Th generators (t 1/2 = 7340 y), which are obtained from the parent 233 U (t 1/2 = 160,000y) [300]. 229Th generators are available at the Oak Ridge National Laboratory USA, the Institute of Transuranium Elements, Germany, and the Institute of Physics and Power, Russia [300].However, as of 2008, the approximate total worldwide production of 225 Ac accounts for only 68 GBq/year, which can support only several hundred patients per year.Therefore, large-scale production of 225 Ac is needed.Alternative production routes are being explored, including proton irradiation of 226 Ra targets, which could produce sufficient quantities of 225 Ac due to the relatively high reaction cross-section; however, the handling of 226 Ra (t 1/2 = 1600 y) is challenging [300].
To date, the accelerator-based production route involves high-energy proton irradiation (>100 MeV) of natural thorium ( 232 Th), and it could serve as another potential path for the future production of 225 Ac.This method may yield twenty times greater quantities of 225 Ac than the current annual production worldwide [310].

Production and Availability
212 Pb is commonly produced from the decay chain of a 228 Th (t 1/2 = 1.9 y) generator, followed by its elution in 2M HCl using a cation exchange column with a maximum yield of 85% [319].At high radioactivity (>37 MBq), however, the radiolytic damage of the cation exchange resin in the 228 Th generator increases the back pressure and decreases the yield [319].To circumvent this, an alternative generator using 224 Ra (t 1/2 = 3.7 d) was designed, which serves as a source of either 212 Bi or its parent nuclide 212 Pb [319].The 224 Ra/ 212 Pb generator could elute 212 Pb with a radioactivity up to ~600 MBq (16 mCi) [319].Currently, 212 Pb is mainly supplied by OranoMed and Oak Ridge National Laboratory [319]. 203Pb via proton irradiation of either natural thallium (Tl) or enriched 203 Tl in a TR13 (13 MeV) cyclotron to create a 228Th / 212 Pb generator for 212 Pb [312].

Clinical Practice
From mid-1940 to 1990, [ 224 Ra]RaCl 2 of high doses (up to 140 MBq) was used to treat different bone and joint diseases, mainly in Germany, but this practice was abandoned for technical and commercial reasons [329].During 2000-2005, the use of [ 224 Ra]RaCl 2 (low dose up to 10 MBq) was revived to treat ankylosing spondylitis patients, but this was discontinued in 2005 due to the enhanced risk of malignant disease following injection [330,331].One of the potential drawbacks of 224 Ra is the release of progeny β − -emitting 212 Pb with a significant half-life of 10.6 h, which could cause unwanted non-target exposure [330].Therefore, alternative delivery strategies are of considerable interest, which could promote the retention of the daughter nuclides or mitigate their recoiling spread.
During 2007-2015, several preclinical studies investigated brachytherapy using 224 Raloaded diffusing α-emitter radiation therapy (DaRT) wires or seeds, which minimizes the damage to surrounding normal tissues [332].The first-in-human clinical study based on DaRT was reported in 2020 and involved the implantation of 224 Ra seeds to treat squamous cancers of the skin and head [333].Complete response to the 224 Ra-DaRT treatment was observed in 22 of the 28 patients; the remaining 6 patients showed only a partial response (>30% tumor reduction) [333].Like 224 Ra, 223 Ra was also studied for the treatment of bone skeletal metastasis.The first clinical study (phase I) in prostate and breast cancer patients was reported by Nilsson et al. in 2005 [334].Later, the favorable clinical results (phase II/III) of [ 223 Ra]RaCl 2 to treat metastatic PCa led to FDA approval of [ 223 Ra]RaCl 2 (Xofigo ® ; Bayer) in 2013 [335,336].Several clinical trials of 223 Ra-based radionuclide therapy in combination with chemotherapy (docetaxel, paclitaxel), hormonal therapy (abiraterone, enzalutamide), and immunotherapy are ongoing (Table 28).

Production and Availability
223 Ra is mainly produced from 227 Ac/ 227 Th generators, where 223 Ra is separated from 227 Ac/ 227 Th mother radionuclides using separation columns [300,350].On the other hand, 224 Ra is usually produced from a 228 Th generator, where 228 Th is immobilized on actinide resin, which allows regular elution of 224 Ra in 1M HCl [351].

Clinical Practice
There are four clinical trials listed for 227 Th-based TTCs registered in the US National Library of Medicine.These trials are based on 227 Th-labeled anti-PSMA-HOPO (Bay 2315497) and 227 Th-labeled anti-mesothelin-HOPO for the treatment of PCa (NCT03724747) [355] and mesothelioma (Bay 2287411), respectively.The remaining two trials are based on 227 Th-labeled epratuzumab-HOPO (Bay 1862864) and 227 Th-labeled trastuzumab-HOPO (Bay 2701439) to treat CD22-positive non-Hodgkin's lymphoma and HER2-positive breast or gastric cancers, respectively (Table 30). 89Zr-labeled HOPO has the potential to serve as a PET surrogate for TTCs, which could support the clinical development of novel TTCs by providing crucial pharmacokinetic and pharmacodynamic information [356].7.5.3.Production and Availability 227 Th is produced as a decay product of the parent β − emitter 227 Ac (t 1/2 = 21.8 year) [357].The longer half-life of 227 Th (t 1/2 = 18.7 days) allows for the shipment of cGMP-grade 227 Th solution worldwide [357].At) is an α-emitting therapeutic radionuclide that decays into two branches either by α-emission (42%) to 207 Bi (t 1/2 = 33.9y) or by EC (58%) to 211 Po (t 1/2 = 516 ms); both eventually decay to a stable 207 Pb [361].Each decay yields one α particle and the emission of characteristic X-rays (70-90 keV) through the decay of 211 Po and could be used for SPECT imaging and quantification of 211 At [361]. 209At (t 1/2 = 5.4 h) is another isotope that predominantly decays by β + emission (96%) and has been introduced as a theranostic pair to 211 At (Table 31) [361]. 211At is a more attractive radionuclide than other α-emitting radionuclides because of its suitable half-life of 7.2 h, the absence of long-lived and/or toxic progenies, and its feasibility to be produced in decent quantities [361].  21At-687 211 Po-569.7,897.8 # Data on 211 At are from [361].Please refer to Scheme 2.

Clinical Practice
Although 211 At-labeled TAT agents were discovered more than 30 years ago, only a few clinical studies using 211 At-labeled Abs have been published.Zalutsky et al. reported on the application of 211 At-labeled chimeric anti-tenascin mAb 81C6 (71-347 MBq) in recurrent brain tumor patients with an encouraging median survival time of 52 weeks compared to 23 weeks reported for recurrent glioblastoma multiforme patients treated with best care [362].Another clinical study of intraperitoneal α particle therapy was reported using [ 211 At]At-MX35(Fab) in relapsed ovarian cancer patients [363].The results showed that there was no apparent radiation-induced toxicity discovered in patients for up to 12 years and no decreased tolerance to relapse therapy.The clinical investigation of 211 At-labeled radiopharmaceuticals is summarized in Table 32.

Production and Availability
The most common route is the cyclotron/accelerator-based production of 211 At through alpha irradiation of 209 Bi (natural Bi) via a 209 Bi(α,2n) 211 At nuclear reaction [364,365].However, only a limited number of cyclotrons with α-beam and with > 25 MeV energy are available in the field, limiting the overall 211 At availability [364].Other methods include the use of 211 Rn/ 211 At generators [366].One of the potential advantages of using 211 Rn/ 211 At generators is the longer half-life of 211 Rn (t 1/2 = 14.6 h) compared with 211 At (t 1/2 = 7.2 h), facilitating wider distribution of 211 At.

Conclusions
In summary, the development of radiometal-based radiopharmaceuticals, including their production, purification, bifunctional chelating agents, and biomarker discoveries, have significantly advanced the application of various radiometals in medicine in the last two decades.Both radiometal-based imaging and radionuclide therapy are changing the lives of patients on a daily basis due to the advancements made in the last 20 years.The field of α-emitting radiotherapy is emerging.Several clinical trials are currently under investigation.Further advances in the production and availability of these α-emitters along with the management of radioactive progeny should permit the cost-effective clinical adoption of TAT compared to traditional chemotherapeutics.Indeed, the future of the radiometal-based radiopharmaceutical industry appears to be very bright.

Figure 1 .
Figure 1.Comparison of beta and alpha radionuclide therapies.

Table 1 .
Decay characteristics of copper radioisotopes used in radiopharmaceuticals # .

Table 3 .
Decay characteristics of the three main radioisotopes of gallium # .

Table 7 .
Decay characteristics of commonly used radioisotopes of scandium # .

Table 8 .
Decay characteristics of leading radioisotopes of terbium # .

Table 9 .
Clinical applications of terbium-labeled radiopharmaceuticals under investigation.

Table 10 .
Decay characteristics of PET isotopes of zinc # .

Table 13 .
Decay characteristics of radioisotopes of indium # .
# Data on

Table 21 .
Decay characteristics of radioisotopes of Bismuth # .

Table 27 .
Decay characteristics of radioisotopes of Radium # .
# Data on

Table 28 .
Clinical applications of223Ra in combination with other therapies.