^203/212Pb theranostic radiopharmaceuticals for image-guided radionuclide therapy for cancer

Abstract

Receptor-targeted image-guided radionuclide therapy (TRT) is increasingly recognized as a promising approach to cancer treatment. In particular, the potential for clinical translation of receptor-targeted alpha-particle therapy is receiving considerable attention as an approach that can improve outcomes for cancer patients. Higher linear-energy transfer (LET) of alpha-particles (compared to beta particles) for this purpose results in an increased incidence of double-strand DNA breaks and improved-localized cancer-cell damage. Recent clinical studies provide compelling evidence that alpha-TRT has the potential to deliver a significantly more potent anti-cancer effect compared with beta-TRT. Generator-produced ²¹²Pb (which decays to alpha emitters ²¹²Bi and ²¹²Po) is a particularly promising radionuclide for receptor-targeted alpha-particle therapy. A second attractive feature that distinguishes ²¹²Pb alpha-TRT from other available radionuclides is the possibility to employ elementally-matched isotope ²⁰³Pb as an imaging surrogate in place of the therapeutic radionuclide. Because direct non-invasive measurement of alpha-particle emissions cannot be conducted using current medical scanner technology, the imaging surrogate allows for a pharmacologically-inactive determination of the pharmacokinetics and biodistribution of TRT candidate ligands in advance of treatment. Thus, elementally-matched ²⁰³Pb labeled radiopharmaceuticals can be used to identify patients who may benefit from ²¹²Pb alpha-TRT and apply appropriate dosimetry and treatment planning in advance of the therapy. In this review, we provide a brief history on the use of these isotopes for cancer therapy; describe the decay and chemical characteristics of ^203/212Pb for their use in cancer theranostics and methologies applied for production and purification of these isotopes for radiopharmaceutical production. In addition, a medical physics and dosimetry perspective is provided that highlights the potential of ²¹²Pb for alpha-TRT and the expected safety for ²⁰³Pb surrogate imaging. Recent and current preclinical and clinical studies are presented. The sum of the findings herein and observations presented provide evidence that the ²⁰³Pb/²¹²Pb theranostic pair has a promising future for use in radiopharmaceutical theranostic therapies for cancer.

Introduction

Receptor-targeted image-guided radionuclide therapy (TRT) is emerging as a highly promising approach to cancer treatment.[1–15] In particular, alpha-particle (α-) therapy is receiving considerable attention; and recent studies highlight the potential advantages of α-therapy relative to β-emitters.[1, 3, 5, 6, 16] The advantage of α-emitters (relative to β-emitters) is largely thought to be a result of a higher linear-energy transfer (LET) (100 keV/μm) and concomitant increase in ionizations (primary and secondary) along the short path length of α-particles in tissue.[3, 5, 11, 12, 15, 17] The deposition of high LET radiation over this short pathlength generates an increase in the incidence of double-strand DNA breaks; improved tumor-cell-specific killing; and improved relative biological effectiveness (RBE). [18–21] [5, 16, 18–36] For example, recent studies in patients with advanced-stage castration-resistant metastatic prostate cancer provide compelling evidence that α-therapy (using [²²⁵Ac]PSMA-617) has the potential to deliver a significantly more potent anti-cancer effect compared with β-therapy (using [¹⁷⁷Lu]PSMA-617).[5, 16] Numerous other studies demonstrate the potential of α-emitters for cancer therapy using radionuclides ²²⁵Ac, ²¹²Bi, ²¹³Bi, and ²¹¹At.[5, 16–36]

Generator-produced ²¹²Pb (t_1/2 = 11 h; 100% β decay to alpha emitters ²¹²Bi and ²¹²Po) is a particularly promising radionuclide for receptor-targeted α-particle therapy for metastatic melanoma, neuroendocrine tumors, and other cancers (Figure 1).[10, 33, 37–39] However, because α-decay cannot be directly observed via traditional molecular imaging modalities, surrogate imaging is generally carried out to develop an understanding of the pharmacokinetics and biodistribution of receptor-targeted candidate ligands. These studies are intended to assist in identifying tumors that are positive for the target receptor of interest and in developing appropriate dosimetry and treatment plans in advance of the radionuclide therapy. For example, non-invasive imaging of the extent of disease has been conducted using [⁶⁸Ga]DOTATOC and [⁶⁸Ga]PSMA-11 in advance of therapy via [²¹³Bi]DOTATOC and[²²⁵Ac]PSMA-617 respectively. While these studies reflect the potential of the theranostic approach, using isotopes of different elements in the imaging and therapeutic procedures introduces some ambiguity in the prediction of the in vivo pharmacokinetics of the radiotherapeutic ligands. For example, recent comparisons of tumor and normal organ uptake of [⁶⁸Ga]DOTATOC vs [⁹⁰Y]DOTATOC revealed small, but measurable, differences in the temporal biodistribution that was attributed to differences in the chemistry of the ligands when labeled with Ga and Y.[40] In the case of ²¹²Pb for α-theranostics, the cyclotron-produced gamma(γ)-emitting radionuclide ²⁰³Pb can be used as an elementally-identical imaging surrogate for the therapeutic nuclide.[10, 33, 37, 38] This property provides confidence that predictions made using ²⁰³Pb SPECT and SPECT/CT imaging accurately represent the expected pharmacokinetics/biodistribution of the therapeutic ligand. In this review, we provide (1) a brief overview of Pb chemistry with regard to isolation and purification for radiopharmaceutical production; (2) an overview of isotope production for 203Pb and generators used for producing ²¹²Pb; (3) a summary and discussion preclinical and clinical studies that have been conducted using ²⁰³Pb and ²¹²Pb; as well as a review of the relevant medical physics of ²¹²Pb as it relates to introduction of these isotopes for image-guided radionuclide therapy for cancer.

Figure 1.

Decay series for ²²⁴Ra/²¹²Pb generators.

Production, Chemistry, and Purification of Lead (Pb) Isotopes

Cyclotron production of ²⁰³Pb:

The supply of radiochemical ²⁰³Pb has been primarily been made available by a single commercial supplier (Lantheus Medical Imaging, Inc., Billerica, MA USA). This is largely due to the need for a high-energy cyclotron for production and a relatively low market demand. The company operates an IBA Cyclone-30 (14–30 MeV) for the production of isotopes ⁵⁷Co, ⁶⁷Ga, ⁶⁵Zn, ⁶⁸Ge, ²⁰¹Tl, ²⁰¹Pb, and ²⁰³Pb. Lead-203 can be produced through the proton irradiation of natural Tl (29.5% ²⁰³Tl and 70.5% ²⁰⁵Tl), or enriched ²⁰⁵Tl at energies above 17 MeV (σ_max = 1244 mb @ 26.0 MeV) via the ²⁰⁵Tl(p, 3n)²⁰³Pb nuclear reaction (Figure 2).[41] Ideally, an irradiation would utilize enriched ²⁰⁵Tl in order to produce the most ²⁰³Pb, while reducing the production of the ²⁰¹Pb (σ_max = 1300.2 mb @ 28.2 MeV) by-product from the ²⁰³Tl(p, 3n)²⁰¹Pb nuclear reaction (Table 1). However, cost and availability of the enriched isotope have hindered adoption for this purpose. In 2019, the cost of enriched ²⁰⁵Tl metal (99.8%) is approximately $1000 US per gram,[41] representing a substantial investment in comparison to ^natTl metal ($12.4 gram⁻¹) or gold ($40–50 gram⁻¹ in 2019). However, a comparison to the cost of enriched ⁶⁴Ni metal (95%; $27,500 gram⁻¹) used for ⁶⁴Cu production suggests that optimization of target materials could be on the horizon as demand for ²⁰³Pb/²¹²Pb expands. A summary of the parameters and approach to production of ²⁰³Pb as conducted currently are supplied here. Few publications that describe these parameters in detail have been introduced into the academic literature. For further details, see Li et al., Appl. Rad. Isot., 2017 (Supplemental Information).[42]

Figure 2:

Excitation function of the ²⁰⁵Tl(p,3n)²⁰³Pb nuclear reaction for production of ²⁰³Pb.

Table 1.

Nuclear reactions and daughter products of interest for the production of radioisotopes of lead from a ²⁰³Tl and ²⁰⁵Tl, as provided by current ²⁰³Pb supplier Lantheus Medical Imaging.⁴³

Nuclear Reactions
205Tl(p, 3n)203Pb	t1/2 = 52 h
203Tl(p, 3n)201Pb	t1/2 = 9 h
205Tl(p, 2n)204mPb	t1/2 = 1 h
Daughter Products
203Pb → 203Tl	Stable
201Pb → 201Tl	t1/2 = 73 h
204mPb → 204Pb	Stable

Target design and irradiation parameters:

Tl targets are electroplated onto a Ni flashed Cu substrate (surface area ≈ 100 cm²) using trace metal grade ^natTl (99.999% TlNO₃; Sigma Aldrich—St. Louis, MO) and a mixture of proprietary plating additives using a graphite anode and a constant current of 1A. Electroplating continues until the electroplating bath is depleted. After drying, the Tl targets are weighed, and visually inspected to ensure they do not contain any visual damage or voids. A typical target possesses 2.5–4.0 grams of Tl metal. Irradiations are carried out at a 7–15° incidence angle up to 165 μA for 6–24 hours on an IBA Cyclone 30 or C-28 at a nominal energy of 25–26 MeV. During irradiation targets are rotated at ~350 rpm and cooled with water at a flow rate of ~10 gpm. A selection of ^natTl targets irradiated at Lantheus are shown in Table 2.[43] Prior to processing, irradiated targets are transferred pneumatically to high energy cells and held for ~90 hours to allow for decay of radiocontaminates. After processing, radiochemical grade ²⁰³Pb in quantities ≥300 mCi, with a specific activity of ≥7,000 Ci g⁻¹ is obtained routinely (Table 3).[43]

Table 2.

Irradiation parameters of ^natTl targets irradiated as provided by current ²⁰³Pb supplier Lan-theus Medical Imaging [43].

Target ID	Mass natTI (g)	I (μA)	Tirr(h)
natTl-l	2.94	147	14
natTl-2	2.91	152	13
natTl-3	3.12	149	9
natTl-4	2.88	146	16
natTl-5	2.88	159	14
natTl-6	3.18	154	14

Table 3.

Activity and specific activities of ²⁰³Pb from ^natTl targets irradiated as reported by Lantheus Medical Imaging.⁴³

Target ID	AEOP GBq (Ci)*	203Pb Specific Activity GBq g−1 (Ci g−1)
natTl-1	21 (0.57)	3.46 × 105 (9361)
natTl-2	23 (0.61)	3.28 × 105 (11564)
natTl-3	10 (0.27)	3.01 × 105 (8132)
natTl-4	19 (0.51)	5.88 × 105 (15891)
natTl-5	19 (0.52)	4.86 × 105 (13140)
natTl-6	20 (0.55)	2.79 × 105 (7546)

^*End of production (EOP) is 5 days post EOB

Generator production of ²¹²Pb:

²¹²Pb for radiopharmaceuticals is produced using shielded ion-exchange based ²²⁴Ra/²¹²Pb system referred to as a “generator” or “²²⁴Ra/²¹²Pb generator”. The principle underlying the use of the ²²⁴Ra/²¹²Pb generators for production of ²¹²Pb is the same as that employed for other generator-based technologies that are used in nuclear medicine daily (Figure 3). For example, ^99mTc produced for single photon emission computed tomography (SPECT) or SPECT combined with computed tomography (CT) imaging of cancer and other diseases is based on a ⁹⁹Mo/^99mTc generator system, in which a longer-lived parent isotope (i.e., ⁹⁹Mo; t_1/2 2.75 days) decays to a shorter lived isotope (i.e., ^99mTc; t_1/2 6.0 hours) that is suited for incorporation into targeting ligands for molecular imaging.[44–49] Similarly, the use of ⁶⁸Ge/⁶⁸Ga generators for the preparation of ⁶⁸Ga-labeled positron emission tomography (PET) and PET/CT agents has expanded significantly in the recent past.[39, 50–56] In the case of ²¹²Pb generator technologies, the principle is based on parent radionuclide ²²⁴Ra (t_1/2 3.63 days), which decays through a series of short lived radionuclides to ²¹²Pb (t_1/2 11 h) (Figure 1). The transient radioactive equilibrium relationship between ²²⁴Ra and its decay to ²¹²Pb provides for a similar scenario in which the life of the generator is based on the half life of the parent. Thus, (similar to ⁹⁹Mo/^99mTc; ⁹⁹Mo t_1/2 2.75 days) a ²²⁴Ra/²¹²Pb generator can be delivered for use and ²¹²Pb eluted for preparation of radiopharmaceuticals daily for 1–2 weeks, at which time the generator is discarded or returned to the manufacturer for re-loading of the parent radionuclide, which is regenerated based on the same radioactive decay/ingrowth principles (Figure 4). Ultimate parent ²²⁸Th has a half life of approximately 2 y (Figure 1), which means that a stable supply of ²²⁴Ra to produce generators can be envisioned with ²²⁸Th solutions used to regenerate ²²⁴Ra on a schedule of approximately every 3–4 weeks for each ²²⁸Th source (Figure 4) and can be obtained through the US Department of Energy and may be available through a single commercial company (Orano Med) with operations in France and the United States.

Figure 3.

Radioactivity relationship for ²²⁴Ra/²¹²Pb generators. ²²⁴Ra level set to 740 MBq. Generator is eluted on at t₀ to prepare patient doses. Generator can be eluted again after sufficient ingrowth of ²¹²Pb. At this level generators are estimated to be useful for 1–2 weeks.

Figure 4.

Radioactivity relationship that forms the basis for production of ²²⁴Ra using ²²⁸Th as feedstock. The level here is set to 740 MBq. ²²⁴Ra is isolated on Day 1 (t₀) to prepare generators. ²²⁴Ra levels return to max ²²⁸Th levels within about 3 weeks and ²²⁴Ra can be isolated again. ²²⁸Th feedstock is a waste material available from the US Department of Energy Isotopes Program catalog on-line in abundant supply.

^203/212Pb chemistry and purifications:

Whether the interest is obtaining ²⁰³Pb produced via cyclotron or ²¹²Pb produced via generator, Pb isotopes generally (see section on liquid ²¹²Pb generators) must be purified and isolated for use in the production of radiopharmaceuticals. Elementally, lead (Pb; atomic number 82) is a heavy metal and member of group 14 of the periodic table. Stable isotopes of Pb include ²⁰⁶Pb (24.1%), ²⁰⁷Pb (22.1%), and ²⁰⁸Pb (52.4%), which are the endpoints of the ²³⁸U, ²³⁵U and ²³²Th decay chains, and ²⁰⁴Pb (1.4%), which is the sole primordial nuclide of Pb. Isotopic abundances of Pb deposits therefore may vary due to coincident uranium or thorium found in the deposit. Metallic Pb is considered soft and malleable (with a relatively low melting point of 327.5°C) and is readily dissolved in dilute nitric acid. Soluble salts of Pb include nitrite, nitrate, perchlorate, citrate and acetate. Insoluble Pb salts include the sulfate, phosphate, and carbonate. Lead chloride (PbCl₂) is modestly soluble in dilute hydrochloric acid (HCl) and increases in solubility at higher HCl concentrations due to the formation of anionic Pb-chloride complexes, such as [PbCl₄]^2-. Lead may also be carried on precipitates including ferric hydroxide, calcium carbonate, and barium sulfate.[57]

In aqueous solution, the other members of group 14 (carbon, silicon, germanium, and tin) may exist in in the +4 oxidation state, while Pb exists almost exclusively in the +2 oxidation state.[57] The immediate neighbors of lead in the periodic table, Tl (Tl) and bismuth (Bi), exist in the Tl(I), Tl(III), and Bi(III) oxidation states, forming the basis for many separation methods of Pb from Tl and Bi. Of interest in nuclear medicine are the isotopes ²⁰¹Pb (t_1/2 = 9.33 hours) and ²⁰³Pb (t_1/2 = 51.92 hours) produced by proton irradiation of isotopically enriched or natural abundance Tl targets in cyclotrons and ²¹²Pb (t_1/2 = 10.64 hours) produced by radioactive decay of ²²⁴Ra (daughter in the ²²⁸Th decay series, Figure 1).

Extraction data for Pb, Tl and Ra on anion exchange resin (AG1×8),[58, 59] cation exchange resin (AG50Wx8),[60, 61]macroporous cation exchange resin (AGMP-50),[62] and Sr resin[63, 64] (extraction chromatographic resin (EXC) containing 4,4′(5′)-di-t-butylcyclohexano 18-crown-6 in 1-octanol on an inert polymeric support) is presented in Figure 5. The cation exchange materials do not offer the selectivity from nitrate or chloride media to efficiently separate Pb from the large mass of Tl target material. However, the macroporous cation exchange material AGMP-50 efficiently separates ²¹²Pb and ²¹²Bi from ²²⁴Ra source material. The anion exchange resin AG1×8 exhibits high selectivity for Tl(III), Bi(III), and Fe(III) over Pb(II) from HCl. However, due to the large columns that would be required to extract the large mass of Tl target, anion exchange is typically only used to purify the radiolead once the bulk of the Tl has been removed via another process. The Sr Resin exhibits very high selectivity for the extraction of Pb(II) over Tl and Cu from nitric acid, with tens of grams per liter of Tl and Cu having minimal impact on the retention of lead.

Figure 5.

Extraction of selected metal ions on AG50Wx8, AGMP-50, AG1×8, and Sr.

Separation of ²⁰³Pb from Tl target materials:

The are a number of methods for the separation of radiolead from irradiated Tl targets in the literature. Methods begin with the dissolution of up to 4–5 grams of irradiated metallic Tl or Tl oxide. Nitric acid is normally employed for target dissolution, forming soluble Tl nitrate salts. Tl can also be dissolved in sulfuric acid (H₂SO₄), but dissolves very slowly in HCl due to the formation of less soluble Tl chloride. Significant amounts of Cu and Ni may also be co-dissolved with the Tl from the target backing and cooling mechanism. Therefore, separation methods must also isolate lead from Cu and Ni as well as Fe, Co and Zn impurities and irradiation byproducts. An early method for the separation of ²⁰³Pb, dissolved the irradiated Tl target in 1 M HNO₃ from a water-cooled Cu plate. The Tl was then reduced to Tl(I) by bubbling sulfur dioxide gas through the solution, and an Fe(OH)₃ precipitate was used to selectively carry the radiolead, leaving the Tl(I) in the supernatant. The precipitate, containing 10 mg of Fe(III) and the radiolead was washed twice with dilute NH₄OH, discarding the supernatant, and dissolved in 1 M HNO₃. The Fe(OH)₃ precipitation and washing process was repeated after the addition of 5 mg of Tl(III) carrier, which further purified Fe(OH)₃ precipitate containing the ²⁰³Pb dissolved in 6 M HCl. The Fe carrier was then removed by four successive extractions with equal volumes of diethylether.[57] A similar method replaced the final ether extractions with removal of the Fe carrier with anion exchange chromatography.[65]

A separation method currently utilized for commercially available ²⁰³Pb begins with the dissolution of up to 4.0 grams of irradiated Tl from a Ni flashed Cu substrate with hot HNO₃. The dissolved target is filtered through glass wool, cooled, adjusted to pH 5–6 with concentrated NH₄OH, and loaded onto a 14 mL column of an iminodiacetate chelating ion exchange resin (Chelex-100). The Pb is selectively retained, while Tl is not adsorbed. Rinsing the column with 12.5 bed volumes of ammonium nitrate removes additional Tl, and the Pb is recovered in 50 mL of 0.5 M NaOH. Nitric acid is then used to adjust to pH 5–6 and the Pb fraction is purified again on a second 14 mL column of Chelex-100 resin. After adjustment of the Pb fraction to pH 5–6, the Chelex-100 separation is then performed a third time, recovering the ²⁰³Pb in 100 mL of 1M HNO₃.[65, 66]

An extraction chromatography separation of radiolead from irradiated Tl targets has been proposed, where up to 4.0 grams of irradiated Tl target material is dissolved in hot nitric acid. The solution is cooled, filtered and passed through a 2–4 mL column of Sr Resin. The Pb is retained while the Tl, Cu, Fe, Zn, and Co and most other potential impurities are not adsorbed. Rinsing with 25 bed volumes of 4 M HNO₃, 0.1 M HNO₃, and 0.1 M HCl further purifies the radiolead, reduces the acid concentration, and converts the system to chloride. The radiolead can then be recovered from the Sr resin with 6–8 M HCl, or dilute acetate, tartrate or citrate buffer.[63, 64] Potential leaching of small amounts of the crown ether extractant and 1-octanol diluent from the EXC resin has been a potential concern and in this case the recovered radiolead could be further purified via cation exchange on a 0.25 mL column of Chelex-100, Bio-Rex 70, or weak cation exchange (CM) bonded silica. More quantitative research is needed for fully understand the potential for this leaching to be an issue for radiopharmaceuticals.

Isolation of ²¹²Pb on ²²⁴Ra generators:

Early ²¹²Pb generators immobilized ²²⁸Th (t_1/2 = 1.9 years) on columns of Na₂TiO₃, eluting ²²⁰Rn (t_1/2 = 55.6 seconds) with water.[67] After a period of decay, ²¹²Pb was isolated on a cation exchange column. This generator suffered from loss of Na₂TiO₃ fines, issues with scale up to millicurie levels, and required continuous elution for 21 hours at greater than 1 mL/min to achieve 75% yield of ²¹²Pb. An improvement on the ²²⁸Th based generator using the ²²⁴Ra daughter (t_1/2 = 3.6 days) adsorbed to a column of AGMP-50 sulfonic acid macroporous cation exchange resin. ²¹²Pb/²¹²Bi can be coeluted from the generator with a small volume of 2.0 M HCl, and ²¹²Pb free of ²¹²Bi by first eluting ²¹²Bi from the generator with a small volume of 0.5 M HCl. The ²¹²Pb can then be further purified on a 50 mg column of Pb Resin (EXC resin containing 4,4′(5′)-di-t-butylcyclohexano 18-crown-6 in isodecanol) prior to labeling to a biolocalization agent.[42]

Liquid generator system for in situ labeling with ²¹²Pb.

A special case of the ²¹²Pb generator was introduced recently. This system considers the considerable effort needed to manage ion-exchange based generators in the clinical setting. Notably, the half-life of ²¹²Pb of 11 hours is a limitation to its use and fast and safe production and purification procedures are required. An alternative liquid-generator solution was developed and tested base on ²²⁴Ra dissolved in a buffer suitable for radiolabeleing of biomolecules.[68] In this system the solution level ²¹²Pb increases during the first day after dissolving pure ²²⁴Ra; and thereafter, is maintained according to the half-life of ²²⁴Ra. The liquid generator method was tested and described for the in situ labeling of a monoclonal antibody with ²¹²Pb in ²²⁴Ra solution and subsequent removal of ²²⁴Ra as an alternative strategy for preparing ²¹²Pb based radioimmunoconjugate. It was demonstrated that a chelator-modified monoclonal antibody could be efficiently labeled with ²¹²Pb in solution (²²⁴Ra in equilibrium with ²¹²Pb). Subsequently, the ²¹²Pb labeled conjugate was separated from cationic ²²⁴Ra using a desalting gel exclusion separation. As a result, a ready to use ²²⁴Ra/²¹²Pb solution can be shipped from a centralized supplier to the end user. The potential advantages of using ²²⁴Ra in solution is considered two-fold by these authors: (1) it is less time consuming to perform the procedures because the acid extraction step is avoided; and (2) it is less laborious and does not require evaporation of acids etc. Importantly, these authors further suggest potential disadvantages: (1) the generator can only be used once because the ²²⁴Ra is lost in the purification step and not regenerated; and (2) the purified ²¹²Pb product may contain traces of ²²⁴Ra. It should be pointed out that it is known from studies of using ²²⁴Ra in the treatment of ankylosing spondylitis, that modest amounts (typically less than 1.0 MBq) can be administered to patients without considerable bone marrow toxicity, indicating a 1–2% content in a ²¹²Pb based product (e.g, of 100 MBq), would be acceptable as long as the ²¹²Pb product does not produce a high degree of bone marrow toxicity. Using this system, if a purer product in terms of ²²⁴Ra is needed, repeated purification on a second PD-10 column could accomplish this. In summary, the liquid generator method represents advantages and potential disadvantages depending upon the application and more research is needed to further develop this approach.

Preclinical and Clinical Application Studies of ²⁰³Pb/²¹²Pb Radiopharmaceuticals

In the clinical setting, in addition to ²²³Ra used in the form of ²²³RaCl₂ for treatment of bone metastases in metastatic castration resistant prostate cancer (mCRPC) [69], ²¹³Bi and ²²⁵Ac are the commonly used α-emitters that have been applied. The α-emitters are showing promise in patients with neuroendocrine tumors,[30] acute myeloid leukemia, [70] and metastatic castration-resistant prostate cancer.[71–74] However, the short-lived α-emitter, ²¹³Bi has the practical disadvantage of a relatively short half life (t_1/2, 45.6 min), substantially limits the potential of this radionuclide for continued development. In some early clinical case studies of ²²⁵Ac radiopharmaceuticals remarkable near-complete responses have been reported. [1, 3, 5, 6, 16] One potential disadvantage of the use of long-lived α-emitter, ²²⁵Ac (t_1/2, 10 days) is the potential to elicit a more pronounced off-target normal organ toxicity due to the recoil and redistribution of five decay daughters (²²¹Fr, ²¹⁷At, ²¹³Bi, ²¹³Po, ²⁰⁹Tl, and ²⁰⁹Pb) post administration of ²²⁵Ac in the in vivo setting. While off-target toxicities arising from the administration of all radiopharmaceutical therapies can be anticipated to be a clinical challenge, these may be accentuated by the relatively long half life of ²²⁵Ac and the number of decay daughters in the decay chain. ²¹¹At has the relatively appropriate half-life of 7.2 h for radiopharmceutical production and administration and has been evaluated clinically for brain cancers[75] and recurrent ovarian cancers.[76–78] Potential instability of radio-astatine radiopharmaceuticals in vivo has been considered a challenge to the potential of ²¹¹At in clinical applications.[79] ²²⁷Th has long half-life (18.7 days) and is also under clinical investigation especially for radioimmunotherapy. As is the case for ²²⁵Ac, the long half-life becomes an asset when matching relatively slow pharmacokinetics of antibodies in vivo.[80] A phase 1 clinical trial using ²²⁷Th-epratuzumab targeting CD22 in Non-Hodgkin’s lymphoma (NCT02581878) was completed in November 2019, but the clinical results have not been published. In another ongoing trial (NCT03507452), treatment of advanced recurrent epithelioid mesothelioma (a serous ovarian cancer) or (optionally) advanced pancreatic ductal adenocarcinoma with BAY2287411 (²²⁷Th-labeled mesothelin-targeting monoclonal antibody) are under investigation. ¹⁴⁹Tb (t_1/2, 4.1 h) is another emerging-potential α-emitter that combines a single low-energy α-particle (3.97 MeV) with a positron emission that could enable concomitant PET imaging of therapy administration.[81] In addition, paired imaging surrogates ¹⁵²Tb (PET) and ¹⁵⁵Tb (SPECT) are available for non-destructive imaging.[82] However, these radionuclides have not made significant progress toward clinical evaluation at the time of this writing. Here we present on the preclinical and clinical investigations that have been conducted to evaluate the use of ²¹²Pb and imaging surrogate ²⁰³Pb as an elementally-matched isotope pair for image-guided therapy for cancer. These studies suggest that ²¹²Pb is a promising radionuclide for cancer treatment – and that imaging isotope ²⁰³Pb has potential to serve as an effective imaging agent for patient selection and dosimetry. Importantly, for dosimetry purposes, the half life of ²⁰³Pb (52 h) is sufficiently long to enable serial imaging for precise assessment of the pharmacokinetics of ²¹²Pb-labeled radiopharmaceuticals.

^203/212Pb without conjugation:

²⁰³PbCl₂ was initially employed as imaging tracer for bone scanning due to its accumulation in skeletal bones.[83] In the renal cortical brush border, uptake of ²⁰³PbCl₂ was time- and concentration-dependent.[84] Interestingly, accumulation was inhibited by specific metals, such as Sn²⁺, Sn⁺, La²⁺, Fe²⁺ and Cu²⁺, but highly resistant to other metals. Complete blockade was found upon the inclusion of cysteine, glutathione, EDTA and EGTA. ²⁰³PbCl₂ was also used as radioactive tracer to study the transport of Pb across blood-brain barrier.[85] Rapid influx of Pb into brain was a passive-transport, concentration-dependent process that was saturated at 4 μM, as monitored by ²⁰³PbCl₂. This transport was not influenced by Ca²⁺, Mg²⁺ or bicarbonate, but completely blocked by albumin, 200 μM cysteine, or 1 mM EDTA. Lever et al. reported the whole-body distribution of ²⁰³PbCl₂ in mice.[86] The clearance of ²⁰³PbCl₂ was slow: 68% of injected dose remained in the body at 48 hours post-injection. Major organs of ²⁰³PbCl₂ retention observed in this study were kidney and liver at earlier time points (0.5–1 hour), and bone at later time points after 24 hours. Ando et al. reported the use of ²⁰³PbCl₂ for the imaging of tumors.[87] In this study, accumulation of ²⁰³PbCl₂ in tumor and normal organs in male Donryu rats bearing Yoshida carcinoma xenografts was compared with ⁶⁷Ga citrate, an FDA-approved imaging agent for tumor and inflammatory tissue. Compared with ⁶⁷Ga citrate, lower accumulation of ²⁰³PbCl₂ in Yoshida carcinoma tumor was observed, whereas the accumulation of ²⁰³PbCl₂ in kidney and bone were significantly higher than ⁶⁷Ga citrate. On the other hand, ²⁰³PbCl₂ was shown to have a lower affinity to inflammatory tissues compared with ⁶⁷Ga citrate. In a separate study, Taylor et al. compared the affinity of ²⁰³Pb acetate and ⁶⁷Ga citrate to abscesses in male Sprague-Dawley rats.[88] Although clear visualization of abscesses using ²⁰³Pb acetate was reported, the concentration of ⁶⁷Ga citrate in the abscesses was more than 10-fold higher than that of ²⁰³Pb acetate after 24 hours post-injection. These data highlighted the general biodistribution profile of ²⁰³PbCl₂: moderate affinity to tumor, high retention in kidney and skeletal bones, and low affinity to necrotic and inflammatory tissues. Rotmensch et al. used a ²¹²Pb colloid complex for the treatment of ovarian carcinoma in mice.[89, 90] In this study, ferrous hydroxide ²¹²Pb colloid was employed due to its long retention in the peritoneal cavity than other ²¹²Pb colloid species (e.g., sulfur ²¹²Pb colloid or ferric hydroxide ²¹²Pb colloid). A dose-dependent survival relationship was observed in mice after treatment with 0–50 μCi of ²¹²Pb ferrous colloid via i.p. injection. 50 μCi of ²¹²Pb ferrous colloid extended mean survival from 16 days to 81 days post tumor implantation. The bone-seeking nature of ²¹²Pb-chelates, as well as the ²¹²Bi-chelates, provides a potential route for the treatment of bone metastases. As reported by Hassfjell et al.,[91] a 490 femur/blood ratio was observed at 2 hours post-injection of ²¹²Bi-DOTMP in balb/c mice. ²¹²Pb/²¹²Bi-chelates showed similar specific accumulation in the femur, but with higher retention of decay daughter ²¹²Bi in kidney due to the recoil of ²¹²Bi from ²¹²Pb decay. Ethylenediamine tetra methylene phosphonic acid (EDTMP) were added to ²²⁴Ra solution to enhance the bone-targeting for the treatment of breast cancer with bone metastases in nude mice, as reported by Juzeniene et al.[92]

Human epidermal growth factor receptor 2 targeting:

Human epidermal growth factor receptor 2 (HER2; aka Receptor tyrosine-protein kinase erbB-2, CD340) is a member of epidermal growth factor receptor family and has been found to be overexpressed in several cancer cell lines and tumors.[93, 94] Trastuzumab is a recombinant DNA-derived humanized monoclonal antibody that selectively binds to the extracellular domain of HER2 with high affinity Kd = 5 nM and inhibits the proliferation of cancer cells.[93] Several studies have reported the conjugation of ²¹²Pb to trastuzumab to direct alpha-particle radiation to the tumor microenvironment. One often-used chelator that has been conjugated to trastuzumab was referred to as TCMC [(1,4,7,10-tetra-(2-carbamoyl methyl)-cyclododecane] and was originally introduced by Slamon et al.[95] Milenic et al. reported the i.p. injection of ²¹²Pb-TCMC-trastuzumab in female athymic nu/nu mice bearing HER2 positive human colon carcinoma (LS-174T) xenografts.[96] The median survival was improved from 19 days in control animals to 57 days in animals treated with 20 μCi ²¹²Pb-TCMC-trastuzumab (n=4). The authors also reported acute toxicity and treatment-related death within 9 days after initiation of therapy in animals treated with 40 μCi ²¹²Pb-TCMC-trastuzumab. The overexpression of HER2 in 20–30% of prostate cancer gave rise to interest in targeting HER2 in prostate cancer cells.[97] Tan et al. reported that 20 μCi of i.v. injected ²¹²Pb-TCMC- trastuzumab suppressed tumor growth and prolonged median survival by 50% compared with control, in an orthotopic human prostate cancer xenograft model.[98] Specifically in this study, the authors used PC-3MM2 prostate cancer cells, which expressed HER2 at much lower levels compared with other prostate cancer cells in support of the hypothesis that ²¹²Pb-TCMC- trastuzumab α-particle radiation could suppress tumor growth even with limited expression of HER2 target. Of note, blood urine nitrogen (BUN) was used to assess kidney damage within first 72 hours and at the conclusion of the study at 3 weeks after the initiation of the therapies. Similarly, BUN was used to determine the kidney damage from injection of career-free ²¹²Pb in normal Balb/c mice at 7 days and 90 days post-injection. However, related studies suggest that BUN may not be the most appropriate biomarker to assess kidney damage. For example, Jaggi et al. showed that BUN served as biomarker for kidney dysfunction at late stages, but that elevated BUN level was only observed 20 weeks after injection of ²²⁵Ac-HuM191 radioimmunotherapy.[99, 100] Boudousq et al. compared the internalized [²¹²Pb]TCMC-trastuzumab and non-internalized [²¹²Pb]TCMC-37A7 (targeting carcinoembryonic antigen CEA) in HER+/CEA+ squamous carcinoma A-431 tumors in mice.[101] Despite the difference in internalization, both ²¹²Pb-immunoconjugates treatments reduced platelets and white blood cells (WBC) within 3 weeks of the initiation of the study. Interestingly, in biodistribution/dosimetry experiments, 1.48 MBq of [²¹²Pb]TCMC-37A7 delivered 35.5 Gy to tumors, whereas same amount of [²¹²Pb]TCMC-trastuzumab delivered only 27.6 Gy. However, therapy studies produced contradictory results where [²¹²Pb]TCMC-trastuzumab improved outcomes in terms of inhibition of tumor growth and improving survival. These data indicated that the internalization and cellular level dosimetry was crucial to accurately assess the outcome of α-radiotherapy. Of note, the method employed to radiolabel TCMC-trastuzumab with ²¹²Pb in the previously mentioned studies included digestion using strong acid and evaporation.[102] Westrom et al. recently reported a new method for radiolabeling of TCMC-trastuzumab using ²²⁴Ra solution in equilibrium with daughter nuclides.[68] Greater than 90% radiolabeling efficiency was achieved when more than 0.15 mg/mL of TCMC-trastuzumab was added in reaction mixture. The author also found that desalting (i.e. PD-10 column) achieved better separation of free ²²⁴Ra from [²¹²Pb]TCMC-trastuzumab than using centrifugal concentrator. A Phase 1 clinical study using [²¹²Pb]TCMC-trastuzumab was conducted for initial safety and preliminary evaluation of the investigational agent in human subjects with ovarian cancer.[103] The initial study was conducted with three ovarian cancer patients who had known metastatic disease in the abdominal cavity and had failed standard therapy. A single i.p. injection of 7.4 MBq m⁻² was administered to subjects to evaluate distribution, pharmacokinetics, and safety. Whole body gamma scans suggested low levels of redistribution and uptake in normal organs. The study was followed by a dose escalation trial in which six escalating doses (7.4 – 27.4 MBq/m²) with a 30% increase for each subsequent dose in eighteen patients with HER2 positive cancer (16 female with recurrent ovarian cancer, 2 male with colon cancer).[104, 105] This ascending dose strategy did not produce significant investigational drug-related toxicity, which was consistent with dosimetry analysis. At six weeks post administration, 10/18 patients were reported to have stable diseases and 8/16 patients had progressive disease. Stable disease were reported in 9 of 10 patients receiving administered activities of 12.6 MBq m⁻² or higher. I.p. injection of [²¹²Pb]TCMC-trastuzumab up to 27.4 MBq m⁻² was well tolerated and studies with further dose escalation seemed feasible for improved tumor response.

Prostate-specific membrane antigen targeting:

Prostate-specific membrane antigen (PSMA) is a type II transmembrane glycoprotein that is expressed and localized in normal human prostate.[106, 107] Overexpression of PSMA was found in over 90% of prostate cancer cells at the apical membrane to the luminal surface of the ducts, but relatively absent in normal tissues.[106–108] The major class of synthetic PSMA ligands include a urea-based small peptide (i.e. glutamate-urea-lysine) as the binding moiety connected to a chelator via a linker of various composition.[109] Compared with large molecules such as antibodies or nanoparticles, these small-molecule PSMA ligands display rapid blood clearance and accumulation in the tumor microenvironment. Numerous studies have investigated the structure-activity relationships of these small-molecule ligands with receptor binding.[110–112] Generally, the binding between PSMA and Glu-Urea-Lys backbone is sensitive and can be impaired by macrocyclic chelators such as DOTA, whereas acyclic chelators such as HBCD-CC in PSMA-11 have less impact on binding. On the other hand, insertion of an aromatic linker between DOTA and binding motif was shown to restore binding affinity.[111, 112] Kuo et al. also used different aromatic linkers (e.g., 2-naphthylalanine, 2-indanylglycine, and 3,3-diphenylalanine) to manipulate binding affinity and accumulation of ⁶⁸Ga-PMSA ligands in tumors in mice.[113] Thus, ^203/212Pb-labeled PSMA ligands have emerged as promising theranostic candidates that employ ²⁰³Pb imaging to provide dosimetry information for ²¹²Pb-based α-radiotherapy in PMSA-positive prostate cancer. Banerjee et al. recently described a series of DOTA- or TCMC-conjugated PSMA ligands that were radiolabeled with ²⁰³Pb.[114] These ligands shared the same Glu-Urea-Lys binding motif with PMSA-617, but with modified linkers. These authors investigated the use of lengthy aliphatic 1,4-butanediamine and octanedioic acid linkers, instead of the 2-naphthylalanine linker included in the PSMA-617 compound. Based on reported biodistribution data, kidney is the primary dose-limiting organ for these ²¹²Pb PSMA-ligands. Both 4-carboxylic acid armed DOTA and the 3-arm DOTA-monoamide variant conjugated to PSMA targeted ligands demonstrated higher retention in tumor and kidney than TCMC-ligands, but interestingly TCMC-ligands generally are reported to impart a higher therapeutic index (tumor/organ ratio) in that the significantly lower kidney accumulation offset the slightly impaired tumor targeting. 3.7 MBq ²¹²Pb-L2 was superior to 37 MBq [¹⁷⁷Lu]PSMA-617 in suppressing tumor growth, but also induced significant kidney toxicity and weight loss. Dos Santos et al.[115] reported on imaging of two metastatic prostate cancer patients with ²⁰³Pb-CA012, a TCMC-conjugated PSMA ligand that shared the same binding motif as PSMA-617. In this first-in-human study of the radiotracer with no clinical reference data, 250–300 MBq [²⁰³Pb]CA012 was cautiously chosen for the administered activity. This activity level, however, resulted in low count rates that required a prolonged scanning time for SPECT, which subjects found difficult to tolerate. Thus, planar scintigraphy was conducted at 0.4, 4, 18, 28 and 42 h post injection. Dosimetry was performed for ²¹²Pb using the data extrapolated from ²⁰³Pb kinetics and biodistribution. The preliminary dosimetry work suggested that the injection activity can be safely scaled up to 750 MBq for future studies, at which an 18 mSv effective dose would be delivered.

Melanocortin subtype 1 receptor targeting:

Melanocortin subtype 1 receptor (MC1R) is a seven-transmembrane G-protein coupled receptor. MC1R plays an important role in UV-resistance, inflammation, pigmentation. The expression of MC1R is primarily localized in melanocytes and melanoma cells.[116] MC1R has been investigated as a potential target for drug delivery for melanoma because of its high expression in malignancy vs normal organs and tissues.[117–119] One particular avenue for introduction of MC1R-targeted therapy is the use of radiolabeled analogs of the native-cognate peptide ligand of MC1R (i.e., α-melanocyte-stimulating hormone; α-MSH). Numerous studies described derivatization of the peptide sequence, linker, and cyclization method to enhance tumor targeting as well as decreasing off-targeting dose (i.e. kidney), which are beyond the scope of this review. Theranostic ^203/212Pb labeled α-MSH analogues were first reported based on a DOTA-conjugated rhenium-cyclized α-MSH analogue (DOTA-ReCCMSH).[33] Three cysteines (Cys) were inserted into the α-MSH analogue that allowed for cyclization based on the coordination between rhenium and the sulfhydryl side chains. 7.4 MBq of [²¹²Pb]DOTA-ReCCMSH delivered 122.4 Gy to B16F0 murine melanoma tumors and completely eradicated 40% of tumors in mice, while introducing 72.2 Gy to kidney and 9.1 Gy to blood, according to the dosimetry.[33] The pharmacokinetic profile of [²⁰³Pb]DOTA-ReCCMSH was similar to that of [²¹²Pb]DOTA-ReCCMSH, and B16F0 xenograft was visualized under SPECT/CT at 2 hours post-injection.[37] The tomographic spatial resolution of ²⁰³Pb was similar to ^99mTc in imaging phantom studies. Our group also observed similar pharmacokinetics between [²⁰³Pb]DOTA-ReCCMSH and [²¹²Pb]DOTA-ReCCMSH,[120] indicating the lead isotopes ^203/212Pb are a promising theranostic pair for image-guided α-radiotherapy. Specifically, in this study, we used Pb-resin (Eichrom Technologies Inc, Lisle, IL) based method to rapidly pre-concentrate and purify ^203/212Pb for radiolabeling of DOTA-conjugated peptides.[120] Recently, Yang et al. reported a first-in-human clinical study using [⁶⁸Ga]DOTA-GGNle-CycMSHhex, a lactam bridge cyclized α-MSH analogue, that targets MC1R for the imaging of metastatic melanoma.[121] The authors also demonstrated that [²⁰³Pb]DOTA-GGNle-CycMSHhex can be used to image B16/F10 xenograft and pulmonary metastases in C57BL/6 mice.[122] Together, these data supported the hypothesis that ^203/212Pb can be used for image-guided α-radiotherapy in patients with MC1R-positive melanoma.

Somatostatin receptor 2 targeting:

Somatostatin receptor subtype 2 (SSTR2) is one of five subtypes of somatostatin receptors (SSTR1–5), a group of G-coupled protein transmembrane receptors. Somatostain-14 and somatostatin-28 are two natural active somatostatin peptides that bind non-selectively to the 5 SSTRs in the family.[123] Structurally-modified peptides “octreotide” and “octreotate” were developed based on the somatostatin native ligands and selectively bind with SSTR2 (with reduced affinity to other subtypes).[124] Stallon et al. radiolabeled DOTAMTATE, a TCMC-conjugated octreotate analogue, with ²¹²Pb and evaluated [²¹²Pb]DOTAMTATE in SSTR2 positive AR42J xenograft model.[125] A safety study indicated that a single dose of 40 μCi [²¹²Pb]DOTAMTATE caused acute toxicity and death within 1 week, whereas 3 cycles of 15 μCi [²¹²Pb]DOTAMTATE was well tolerated. These studies demonstrated that 3 cycles of 10 μCi [²¹²Pb]DOTAMTATE at 2-week intervals extended the median survival from 3 weeks in controls to 12 weeks in treated mice. A Phase 1 clinical trial using AlphaMedix ([²¹²Pb]DOTAMTATE) is ongoing in human subjects with SSTR2 positive neuroendocrine tumors (NET; clinicalTrials.gov; NCT03466216). The trial is non-randomized, open-label, dose-escalating, single-centered study to determine the biodistribution, safety, and preliminary effectiveness of AlphaMedix in adult SSTR2 positive NET patients. A maximum of 50 adult NET subjects are anticipated in the trial to delineate the dose-limiting toxicity (DLT) and maximum tolerated dose (MTD). According to the study design, a single initial dose administered via i.v. for the first cohort of the subjects, and escalating doses (in 30% increments) are administered to subsequent cohorts until tumor response or DLT is observed. If the tumor response is observed, the study is designed to redirect to a multiple-escalating dose strategy with 3 treatments of AlphaMedix at eight-week intervals. The initial result of the trial in 10 patients (6 females and 4 males) with metastatic NETs has been recently reported.[126] The treatments appear to be well-tolerated at levels administered. Mild adverse events such as nausea, mild hair loss, abdominal pain, diarrhea, and fatigue were reported; and no dose-limiting toxicity has been observed. In regard to efficacy of the investigational drug, stable diseases were observed in all patients. Notably, a decreased maximum standardized uptake value (SUV) by PET imaging assessed by [¹⁸F]Fluorodeoxyglucose (FDG) and/or [⁶⁸Ga]DOTATATE scans has been reported after 2 cycles of treatment in the multiple dose cohort. The complete evaluation of the investigational drug regarding safety and efficacy are in progress. Despite the availability of ²⁰³Pb as the imaging and dosimetry surrogate for ²¹²Pb via SPECT, the application of this imaging agent was not included in the study design of this trial. Instead, the subjects were confirmed histologically or imaged by other FDA-approved SSTR2 imaging modalities (i.e., [⁶⁸Ga]DOTATATE).

Vascular cell adhesion molecule 1 targeting:

Vascular cell adhesion molecule 1 (VCAM-1) is a cell adhesion molecule that has been shown to be expressed on the surface of endothelial cells (in the vessel lumen).[127] As such, VCAM-1 has been investigated as a target for imaging of inflammation in atherosclerosis, as well as in early human brain metastases. Radiolabeled small antibody fragments and peptides that bind to VCAM-1 have been used to for imaging of atherosclerosis, including a ^99mTc-labeled single-chain variable fragment (scFv),[128] ¹⁸F- and ^99mTc-nanobodies,[129] as well as ¹⁸F- and ^99mTc-labeled peptides.[130–132] Zhang et al. reported using [⁶⁸Ga]NOTA-VCAM-1scFv to monitor tumor growth in a VCAM-1 positive B16F10 tumor model.[133] Corroyer-Dulmont et al. demonstrated successful radiolabeling of a TCMC-conjugated antibody against VCAM-1 (i.e. αVCAM-1) with ²¹²Pb.[134] The efficacy of [²¹²Pb]αVCAM-1 was examined in athymic nu/nu mice bearing brain metastasis developed by injection of MDA-231-Br-GFP cell via left cardiac ventricle. The authors reported 6 times higher dose deposition from [²¹²Pb]αVCAM-1 in brains bearing metastasis relative to healthy brains in these mice. Compared with external beam radiation therapy, reduced tumor burden was observed after [²¹²Pb]αVCAM-1 treatment. Of note, due to relatively large molecular weight and slower clearance compared with small peptides, [²¹²Pb]αVCAM-1 displayed relatively slow blood clearance, and accumulation/retention in liver, kidney and spleen within 24 hours post-injection. More recently, Frelin-Labalme et al. pursued precise dosimetry for [²¹²Pb]αVCAM-1, by determining the spatial distribution of [²¹²Pb]αVCAM-1 in cell culture media and calculating the average absorbed dose per α-particle emission.[135] The measurements of α-particles were taken either from the bottom of cell culture well or from the above. Interestingly, the measured spatial distribution of [²¹²Pb]αVCAM-1 displayed lower distribution in the middle, but higher concentration at the two extremes, resulting in 1.75 times higher dose deposition to cells than uniform distribution of [²¹²Pb]αVCAM-1, indicating that accurate dosimetry is needed to truly understand the dose deposition of ²¹²Pb-based α-radiotherapy.

Gastrin releasing peptide receptor targeting:

Gastrin-releasing peptide, also known as bombesin, binds with gastrin releasing peptide receptor (GRPR), which has been shown to be overexpressed on the extracellular membrane of prostate and breast cancer cells.[136, 137] Bifunctional bombesin-chelator conjugates have been used in preclinical and clinical studies for imaging of GRPR-positive tumors. These bombesin-chelator conjugates are sensitive to structural modifications to the C-terminus, N-terminus and linker.[138–143] Okoye et al. evaluated ²⁰³Pb radiolabeled bombesin peptide RM2 (DOTA- 4-amino-1-carboxymethyl-piperidine-D-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH₂) in ICR-SCID male mice bearing PC-3 human prostate cancer xenografts.[144, 145] Despite accumulation in kidney and pancreas at early timepoints (<24 h), this [²⁰³Pb]RM2 displayed rapid accumulation and sustained retention in tumors (7.7 ± 2.3 %ID/g at 24 hours post-injection).[144] Due to the prolonged retention [²⁰³Pb]RM2 in tumors, SPECT/CT imaging provided excellent tumor to background contrast at 24 hours post-injection.[145] Some discrepancies were observed in the maximum uptake values between [²¹²Pb]Pb-RM2 and [²⁰³Pb]Pb-RM2 in tumor, kidney, and pancreas.[145] These discrepancies were attributed to the difference in molar activity (MBq nmole⁻¹) of [²¹²Pb]Pb-RM2 and [²⁰³Pb]Pb-RM2 used in the biodistribution studies. Studies are underway to determine the in vivo therapeutic efficacy and maximum tolerated dose of [²¹²Pb]Pb-RM2 in SCID mice bearing PC3 prostate tumor xenografts.

B7-H3 (CD276) targeting:

The co-stimulatory protein B7-H3 (CD276) is a member of the B7 family of immune regulatory ligands that has been shown to be overexpressed in various cancer types, including pancreatic cancer, ovarian cancer, prostate cancer etc.[146–149] Human B7-H3 was reported to be a co-stimulator that promoted T cell production and induced CD8⁺ cytotoxic T cells.[150–152] On the contrary, B7-H3 has also been reported as inhibitor of T cell activity.[153–156] Several ²¹²Pb labeled antibodies against B7-H3 have been investigated in preclinical studies. Kasten et al. reported using ²¹²Pb-TCMC-conjugated mAb 376.96 in i.p. xenografts of ES-2 and A2780cp20 human ovarian cancer.[157] Significant accumulation of [²¹²Pb]376.96 in i.p. tumor xenograft was observed at 24 hours post-injection (29 ± 6% ID/g in ES-2, 73 ± 33% ID/g), but no significant difference was found between [²¹²Pb]376.96 and non-specific [²¹²Pb]F3-C25 in both tumor xenografts, due to high accumulation of the non-specific antibody [²¹²Pb]F3-C25 in tumor samples. Of note in this study, although A2870cp20 cells expressed 10-fold higher binding sites compared to ES-2 cells, no significant improvement in median survival was observed after the treatment of [²¹²Pb]376.96 in these two tumor models, highlighting the need for better understand of the difference between in vitro and in vivo experiments. The same group reported an expanded application of [²¹²Pb]376.96 in a subcutaneously-implanted patient derived xenograft (PDX) panc039 model.[158] Similar to previous studies, significant accumulation of [²¹²Pb]376.96 was found in spleen, liver, kidneys and blood, with a tumor/organ ratio less than 1 for spleen and kidney.

Other targets:

Chondroitin sulfate proteoglycan 4 (CSPG4) is highly expressed on the cell surface membrane of several cancers, including melanoma and triple negative breast cancer.[159, 160] Kasten et al. reported 125,000 binding sites in SUM159 human breast cancer cell line and 9,100 binding sites in another human breast cancer cell line 2LMP.[161, 162] A CSPG4-specific mAb (225.28) was labeled with ²¹²Pb for α-radiotherapy for these studies. Differences in accumulation of [²¹²Pb]mAb 225.28 were observed in large SUM 159 xenografts vs small xenograft tumor models, indicating that tumor size and vascularity within s.c. tumor xenografts plays important role in dose delivery and pharmacokinetics in these mouse models. Tumor membrane antigens IL-2 and CD38 have also been assessed as targets for the delivery of ²¹²Pb-labeled antibodies for α-particle immunotherapy in blood cancers. For example, ²¹²Bi was used to radiolabel anti-Tac, a DPTA-conjugated monoclonal murine IgG_2a antibody that targets human IL-2 receptor.[163] The ²¹²Bi-anti-Tac was evaluated in IL-2 receptor-positive adult T-cell leukemia cells HUT-102B2 by protein synthesis inhibition and clonogenic assays. Hartmann et al. conjugated anti-Tac antibody to a DTPA derivative chelator 2-(p-isothiocyantobenzyl)-trans-cyclohexyldiethylenetriaminepentaacetic acid, and labeled the immunoconjugate with ²¹²Bi.[164] The [²¹²Bi]anti-Tac was evaluated in nude mice bearing i.p. SP2/Tac tumors. In this tumor model, i.p. injection of 7.4 MBq [²¹²Bi]anti-Tac completely eradicated 75% SP2/Tac tumors. Recently, Quelven et al. reported ²¹²Pb labeled TCMC-conjugated daratumumab, a human mAb targeting CD38 that targets CD38 in multiple myeloma.[165] This [²¹²Pb]daratumumab was evaluated in mice bearing s.c. RPMI8226 xenografts. Acute toxicity was observed when animals were dosed with 277.5 kBq [²¹²Pb]daratumumab. On the other hand, 277.5 kBq of [²¹²Pb]daratumumab extended median survival from 11 days in controls to 55 days in treated mice.

Combination therapies:

Combination of radioligand therapy with other anti-cancer drugs takes advantage of the ionizing radiation from radionuclides, as well as the cytotoxicity from anti-cancer drugs with the potential to achieve synergistic tumor-killing effect that are tolerable. Generally, the rationale for combining radioligand therapy with other anti-cancer drugs involves: (1) additive cytotoxicities; (2) sensitizing cancer cells to ionizing radiation; and (3) the potential to pharmacologically upregulate the receptor number to enhance total binding sites per cell for the radioligand. In this context, enhanced therapeutic outcomes were observed with [²¹²Pb]trastuzumab when combined with chemotherapy drugs gemcitabine and paclitaxel.[166–168] Similar to previously reported combinations of chemotherapy drugs and radioligands,[169–172] gemcitabine and paclitaxel not only induced cytotoxicity by interfering with DNA synthesis and arresting cell cycle as monotherapies, but also sensitized tumor cells to the high LET radiation from [²¹²Pb]trastuzumab. These studies suggested that the radiosensitizing effect of paclitaxel on [²¹²Pb]trastuzumab might be associated with perturbation of the mitotic spindle checkpoint, leading to increased mitotic catastrophe and cell death.[167] Another example is the combination of [²¹²Pb]DOTAMTATE with 5-fluorouracil in preclinical SST2R positive tumor models.[125] Upregulation of receptor density on the cell surface results in more binding sites for the radioligand, and therefore has the potential for synergy in tumor-cell killing. Histone deacetylase inhibitor (HDAC_i) vorinostat, in addition to its anti-cancer activity, was used to induce the expression of human norepinephrine transporter and enhanced the number of binding sites for [¹³¹I]metaiodobenzylguanidine (MIBG) in neuroblastoma.[173] The combination of vorinostat and [¹³¹I]MIBG enhanced the therapeutic effecacy by upregulating protein density in a SH-SY-5Y neuroblastoma xenograft model. This combination therapy continued into a phase 1 safety trial that identified a relatively tolerable dose including 180 mg m⁻² dose⁻¹ vorinostat with 666 MBq kg⁻¹ [¹³¹I]MIBG.[174] Our group also reported the use of pharmacologically enhanced radionuclide therapy in combination with HDAC_i and inhibitors of the MAPK pathway in a metastatic melanoma model in mice. MAPK_i and HDAC_i were shown to significantly upregulate MC1R expression that provided for increased delivery of [²¹²Pb]DOTA-MC1L to A2058 and MEWO melanoma xenograft.[175]

Image-guided dosimetry for ²⁰³Pb/²¹²Pb Radiopharmaceuticals

Utility of radiation dosimetry:

Ultimately, the value of the image-guided dosimetry philosophe for ²⁰³Pb/²¹²Pb radiopharmaceuticals depends on the use of imaging information to make dosimetry predictions that can guided therapeutic radioactivity to be administered. Thus, a review of the state of ²⁰³Pb/²¹²Pb radiopharmaceuticals requires a discussion of the state of radiation dosimetry with regard to α-particle radionuclide therapies. In particular, it is important to point out the differences between traditional external beam radiation approaches to dosimetry vs dosimetry for α-particle therapy, as well as MIRD based and voxel-based approaches. The success of radiation-based therapies depends on delivering enough radiation dose to malignant tissues for disease control, without delivering so much radiation dose to normal tissues that toxicity out-weighs the therapeutic benefit. Specific at-risk healthy tissues vary by radiation modality and patient anatomy. As an example, in the external-beam radiation treatment of a neck cancer, the following tissues may be at risk for radiation-induced toxicity: spinal cord, esophagus, skin, mandible, parotids, pharyngeal constrictors, lens of the eye, cochleae, brainstem, oral mucosa, submandibular glands, and the cricopharyngeal muscle. Likewise for targeted [¹³¹I]NaI treatment of thyroid carcinoma, the following tissues may be at risk for radiation-induced toxicity: stomach, upper gastrointestinal tract, bone marrow, and lungs. These normal tissues have generally well-characterized radiation dose limits for photon- and electron-emitting sources of radiation,[176, 177] and therefore patient-specific radiation dose calculation, referred to as ‘dosimetry,’ is prudent for optimization of therapeutic outcome.

Dosimetry fundamentally relies on careful characterization of the radiation source – either a linear accelerator in the case of conventional external beam radiotherapy or a radioisotope in the case of radiopharmaceutical therapy – and an understanding of how that radiation source interacts with the human tissues that are being exposed. Characterization of linear accelerators is typically accomplished through careful collection of ionization data within a large water tank at various field sizes and energies in order to form a cohesive model of radiation output.[178] In the case of radiopharmaceuticals, most radioisotope decay schema have been characterized with sufficient accuracy and precision for clinical dosimetry.[179] Radiation interacts with tissue in accordance with fundamental physical principles. In the case of therapy-relevant photons, these interactions are dominated by photoelectric absorption, Compton scattering, and pair production. Charged particles transfer energy to tissue primarily through Coulomb interactions and subsequent ionization of atoms within the medium. Sophisticated tools have been developed to predict external beam radiotherapy dosimetry, given three inputs: (1) the radiation beam model, (2) the beam configuration relative to patient anatomy, and (3) a 3D map of tissue densities, inferred from computed tomography (CT) imaging. These dosimetry tools utilize a number of dose calculation algorithms including stochastic methods such as Monte Carlo simulation[180] and deterministic methods such as collapsed cone convolution superposition[181] and grid-based Boltzmann solvers.[182] Availability of these dosimetry tools and techniques has increased the complexity and improved the efficacy of radiation therapies over the past three decades.[183]

Radiopharmaceutical dosimetry techniques:

Radiopharmaceutical dosimetry differs from external-beam dosimetry in two major ways: (1) the radiation dose delivered depends on the patient’s individual biology and anatomy rather than just anatomy and (2) determining the distribution of energy released in matter is measurement-based rather than calculation-based. For a given quantity of radiopharmaceutical, patient-specific pharmacokinetics determine the residence duration of radioactivity in normal and target tissues; this is not the case with external-beam irradiation where the radiation delivered is biology-invariant. Determining activity residence for each tissue of interest requires patient-specific measurements, in the form of quantitative 1D, 2D, and/or 3D imaging. Quantitative imaging for ²⁰³Pb/²¹²Pb is addressed in a following section. However, given sufficient measurement data two main approaches to patient-specific radiopharmaceutical dosimetry have emerged: the “MIRD method” and voxel-based techniques.

MIRD Method:

A method for determining whole-organ mean doses has been colloquially named after the Committee on Medical Internal Radiation Dose (MIRD) of the Society of Nuclear Medicine and Molecular Imaging, although the MIRD committee has also provided guidance on voxel-based techniques.[184] The conventional MIRD method[185, 186] relies on determining whole-organ activity residence times from planar gamma imaging, 3D gamma imaging, or blood sampling. From these measured residence times, mean-absorbed dose to each organ is calculated by applying inter- and intra-organ energy transfer coefficients, referred to as ‘s-values.’ These energy transfer coefficients are generated for each radioisotope through Monte Carlo simulation of radiation transport through in silico anthropomorphic phantoms. Originally these anthropomorphic phantoms were relatively rudimentary,[187] given that their primary purpose was dose estimation for radiation protection. However, more sophisticated phantoms have been developed with age- and sex-dependent anthropomorphisms.[187, 188] Several softwares have obtained United States FDA clearance for clinical implementation of the MIRD method. Most notable of these, the Organ Level Internal Dose Assessment (OLINDA) platform,[189, 190] which has superseded the earlier MIRDDOSE platform.[191]

The primary advantage of the MIRD method for radiopharmaceutical dosimetry is the relatively simple calculation procedure, and reasonably accurate mean organ doses in a majority of cases. Limitations of the approach include an inability to exactly replicate a patient’s organ geometry, which introduces some amount of error. Additionally, the method cannot provide sub-organ dose information, so-called “dose volume histograms,” which are often used for correlation with toxicity in radiotherapy. Furthermore, the MIRD method cannot accurately predict dose to normal tissues in cases where proximal and avid tumors exist, i.e. dose to bone marrow when there are extensive bone metastasis.

Voxel-based Dosimetry:

Voxel-based dosimetry techniques for radiopharmaceuticals have recently emerged and show promise for addressing some drawbacks of the MIRD method.[184, 192–195] These methods utilize patient-specific 3D anatomy and activity distribution information for dose calculation, with dose calculation algorithms that closely resemble those used for external-beam radiotherapy. These techniques are referred to as ‘voxel-based’ due to their ability to calculate dose for each voxel within a patient image volume. As of this writing, two software tools have received United States FDA clearance for general-purpose radiopharmaceutical voxelwise dosimetry: SurePlan MRT (MIM Software Inc.) and PLANET Dose (Dosisoft). Early stage software products that are emerging include Torch (Voximetry, LLC) and further developments in addressing the differences between external beam radiation and ligand directed radionuclide therapy are anticipated to anticipated to continue.

Strengths of voxelwise dosimetry include accurate representation of the patient geometry, including tumors and normal tissues. Drawbacks include increased computational complexity and a need for 3D imaging. The most notable weakness of voxelwise dosimetry though is the fundamental inaccuracy of reproducing the underlying activity distribution through single photon emission computed tomography (SPECT) imaging, which introduces a spatial blurring on the order of 5 – 20 mm, depending on the patient size, radioisotope, and reconstruction technique.[192] This blurring of the underlying activity distribution introduces significant dosimetric inaccuracies in small-volume source tissues, such as tumors. One must therefore be cautious with how these voxel-based dosimetry techniques are applied in practice.

Quantitative Imaging of ²⁰³Pb/²¹²Pb:

Imaging of ²⁰³Pb-labeled surrogates of ²¹²Pb-labeled therapeutics is attractive for multiple reasons. The half-life of ²⁰³Pb (t_1/2: 51.9 h) is greater than that of ²¹²Pb (t_1/2: 10.6 h) which allows for imaging over the entire duration of dose delivery during therapy. Furthermore, the chemically identical theranostic pair should minimize differences in pharmacokinetics between the imaging and therapeutic agents. The half-life of ²⁰³Pb is also convenient from a production and distribution logistics standpoint, as shorter half-lives (t_1/2 < 12 h) tend limit translatability.

Although the pharmacokinetics of ²⁰³Pb-labeled compounds would be expected to mirror that of ²¹²Pb-labeled compounds, it is notable that in approximately 30% of ²¹²Pb β⁻ decays, the daughter ²¹²Bi atom is not retained within the chelator.[196] Evidence of this effect in literature is limited to the DOTA chelator, and therefore more studies are needed regarding release of daughter ²¹²Bi atoms from other chelators. Nonetheless, release of daughter ²¹²Bi atoms prior to emission of an alpha particle introduces uncertainty in predicting dosimetry from ²⁰³Pb-based imaging. In particular, radioactive ions of bismuth are known to accumulate within the kidneys.[197–199] The degree to which ²⁰³Pb imaging would underestimate renal dose depends on route of clearance of the primary radiopharmaceutical, the specific uptake of radiopharmaceutical in target tissues, and the fraction of internalized activity at the target. Radiolabeled peptides for example tend to primarily clear through the kidneys, have modest uptake in target tissues, and have partial internalization.[200] Thus, biodistribution studies that compare ²¹²Pb and ²⁰³Pb labeled candidate pharmacokinetic properties are required in the development phase of new radiopharmaceuticals.

For imaging, ²⁰³Pb produces a 279 keV gamma ray in 81% of decays. Confounding photon emissions are minimal, with a 3.4% abundant 402 keV photon and several lower-abundance k-shell x-rays below 100 keV. Conventional nuclear medicine imaging techniques, such as SPECT, are therefore applicable and well-suited to quantitative imaging of in vivo ²⁰³Pb distributions. Significant work has recently been done on determining optimal methods for quantitative imaging of ¹⁷⁷Lu,[201, 202] which decays with a 208 keV photon, and these techniques are largely applicable to ²⁰³Pb imaging. One notable difference is that the use of a high-energy (HE) collimator will likely be needed to reduce septal-penetration inaccuracies, whereas a medium energy (ME) collimator is typically used for ¹⁷⁷Lu imaging. Further studies of ²⁰³Pb imaging using the latest in SPECT imaging systems is needed to fully understand the state of the art in SPECT imaging for this isotope.

Special Considerations for Alpha-Emitting Radiopharmaceuticals:

Alpha radiation differs from conventional radiation types – photons, electrons, protons – in that alpha radiation exhibits significantly increased linear energy transfer (LET), which is the rate that a primary or secondary particle transfers energy to the material. Higher LETs are associated with increased ionization density along the particle track, which has significant biological implications at the cellular and subcellular level.[203] As ionization density increases, the propensity for radiation to cause double-strand DNA breaks increases, which tends to increase lethality as a function of radiation dose. This increase in cell killing is known as the Radiobiological effectiveness (RBE) of a radiation type (Figure 6). Experiments have shown that alpha particles have an RBE of approximately 3 – 7 for cell killing,[204] which indicates that the same degree of cell-killing would require 3 – 7 times less absorbed dose for alpha particles in comparison with low-LET radiation.

Figure 6.

Illustration of the process to connect measured organ dosimetry to normal tissue complication probability (NTCP) literature data for low-LET radiation (β⁻ and γ) and high-LET radiation (α). Dotted lines indicate correction factors that are poorly understood and/or targeting ligand-specific.

The increased propensity for inducing double-stranded DNA breaks also introduces significant differences in the radiation damage repair kinetics, and therefore significant deviations are likely from our traditional models of low-LET dose rate effects.[204, 205] This same trend also minimizes the impact of radiosensitizing and radioprotecting agents, which tend to act by promoting or suppressing free radical-induced single strand breaks.[204]

Another unique feature of alpha particles is that their short range in matter, on the order of 100 μm. This reduced range increases the need to understand the microscopic distribution of radiopharmaceutical, as different biological outcomes may be possible for a particular whole-organ mean dose.[203] This motivation has led to significant progress in developing micro-dosimetric methods and alpha particle histological imaging techniques.[206–208] A summary of the challenges in connecting measured whole-organ dose from alpha emitters to the probability of a toxic event occurring is summarized in Figure 6. In general, more empirical data is needed from human studies before a predictive links between whole-organ radiation doses and toxicity can be established.

Strategies for Translation:

Mean absorbed dose to organs from alpha radiation is not useful for toxicity prediction from current literature data, but it may be that for a given radiopharmaceutical the absorbed dose measure can be predictive of toxicity once sufficient data is gathered. For this reason, a desirable strategy for clinical translation of alpha-emitting radiopharmaceuticals is to escalate absorbed dose to organs, rather than administered activity. This strategy acknowledges that absorbed dose to organs will correlate more strongly with toxicity than will administered activity due to inter-patient pharmacokinetic differences.

It is worth noting that the RBE of alpha-emitters for cell-killing (3 – 7) tends to be less than the RBE of alpha radiation for late stochastic effects (~20),[209] such as the induction of secondary cancer. Care should therefore be taken to monitor the incidence of myelodysplastic syndrome, leukemia, and other malignancies following administration of alpha-emitting radiopharmaceuticals. In the case of younger patients, the probability of a secondary malignancy may be higher, and therefore caution should be used until more is known regarding the late effects of alpha-emitting radiopharmaceuticals.

Summary and Future Outlook

Emerging evidence suggests that receptor targeted radionuclide therapy using α-particle emitters has the potential to develop into a new form of therapy that can be widely available. Known radionuclides with half lives that are practical for radiopharmaceutical production and administration for cancer therapy, ²²⁵Ac (t_1/2 10 days);^{11 211}At (t_1/2 7 hours);^{12 212}Pb (t_1/2 11 hours);¹³ and ²²⁷Th (t_1/2 18 days)¹⁴ are under consideration in a growing number of clinical trials and investigator initiated studies. In this review, ²¹²Pb is identified as an isotope for α-particle therapy for which an elementally-identical radionuclide (i.e., ²⁰³Pb) is available as an imaging surrogate that can be used to identify patients that may be candidates for ligand directed ²¹²Pb therapy and for establishing a treatment plan under a dosimetry guided paradigm of patient care. An elementally-matched imaging surrogate radionuclide is not available for other radionuclides identified for α-particle therapy and the use of ²⁰³Pb for this purpose lends confidence to the use of the companion diagnostic as a predictor of the pharmacokinetics of the radiopharmaceutical when labelled with the therapeutic radionuclide (i.e., ²¹²Pb). The half life of ²⁰³Pb (52 h) is well-matched for image-guided dosimetry determinations, as well as for production and distribution via cyclotron manufacturing approaches using current technology. In practical terms, the half life of ²¹²Pb (11 h) is best suited for on-site ²²⁴Ra/²¹²Pb generator production or regional radiopharmacy distribution (using the generator approach) to the patient-care clinical setting. For preclinical research on alpha-particle therapy for cancer, in which tumors are implanted in laboratory mice and rats to establish the preclinical efficacy of new radiopharmaceuticals, the limited production and expense of ²²⁴Ra/²¹²Pb generators to date has contributed to a relatively low demand. On the other hand, increases in use are anticipated to drive expanded production capability. In contrast, the half-life of ²²⁵Ac (10 days) has the advantage of centralized production and a sufficiently long half life to simplify coordination of preclinical and clinical studies. However, in a practical sense, management of radioactive waste streams arising from the use of an alpha-particle emitter with a 10 day half life presents a different set of logistical challenges. While current production of ²²⁵Ac has largely depended upon significant governmental operations and limited supply, recently commercial suppliers have emerged that are developing a centralized production and distribution based on parent radionuclide ²²⁹Th. Similarly, large inventories of parent radionuclide ²²⁸Th (t_1/2 = 1.9 years) needed for production of ²²⁴Ra/²¹²Pb generators are available through the US Department of Energy National Isotopes Program. This identifies a pathway for a cost-effective ready supply of ²²⁴Ra/²¹²Pb generators for producing the radionuclide that can be expanded to meet the need of clinical trials and commercial radiopharmaceutical development. Current demand for ²¹²Pb is met by monthly production by the US DOE, but would need to be expanded for commercial applications beyond early stage clinical trials. Similar to ²²⁵Ac and ²¹²Pb, an increasing number of clinical trials (see clinicaltrials.gov) have been initiated for ²¹¹At and ²²⁷Th. Similar to ²¹²Pb, ²¹¹At possesses a half life (7 h) that is (given current production cyclotron/targetry requirements) too short for practical-centralized production and distribution. In this sense, the potential for producing an on-demand nimble supply of ²¹²Pb is made possible through the use of the ²²⁴Ra/²¹²Pb generator. The half lives of ²¹¹At and ²¹²Pb are likely to be best suited for the pharmacokinetics of small molecules and low-molecular weight peptides with biological half lives on the order of hours. On the contrary, tumor targeting of radiation with antibodies and other biological targeting ligands with long circulating biological half lives requires the use of radionuclides with longer nuclear half lives such as ²²⁷Th and ²²⁵Ac and several clinical trials are underway using this molecular targeting approach. Similar to ²²⁵Ac, the primary production route for ²²⁷Th requires large scale (neutron irradiation) facilities, but sufficient quantities are being produced to supply several clinical trials as of this writing. Ultimately, the selection of radionuclide for delivery of alpha-particle radiation to cancerous tumors will depend on several factors, including biological circulating half life of the targeting ligand, availability and cost of the radionuclides, effectiveness of chelation moieties and adaptability of the radionuclide chemistry for radiopharmaceutical production. Emerging evidence suggests that Pb radionuclides can be isolated in sufficient quantities for radiopharmaceutical production, as evidenced by several studies in the preclinical and clinical setting that are demonstrating the potential of this theranostic pair for image-guided radiopharmaceutical therapy for cancer against several cell-surface protein targets, including SST2R, MC1R, PSMA, and others. In addition, recent FDA approvals and clinical trials of low molecular weight peptides and peptide-like small molecules may have biological half lives that are well suited to ²¹²Pb. Advances in the software applied for dosimetry that has the potential to best take advantage of the ²⁰³Pb/²¹²Pb theranostic pair are underway. Current and future research relating the interactions of high LET alpha-particles with cancer cells, tumors, and normal tissue (at the cellular and whole organism levels) will determine full potential of the ²⁰³Pb/²¹²Pb theranostic approach to improving outcomes for cancer patients.

Table 4.

Summary of the application of ^203/212Pb-labeled antibodies or peptides in preclinical and clinical studies.

Radionuclide	Supplier	Target	Vector	Chelator	Application
212Pb	212Pb generator from the University of Chicago	HER2	trastuzumab	TCMC	athymic (nu/nu) mice bearing LS-174T colon carcinoma and Shaw pancreatic carcinoma xenograft.95
212Pb	212Pb generator from AREVA Med, LLC	HER2	trastuzumab	TCMC	Nude mice bearing PC-3MM2 prostate cancer xenograft.98
212Pb	212Pb generator from AREVA Med, LLC	HER2 and CEA	Trastuzumab, anti-CEA mAb 37A7	TCMC	Swiss nude mice bearin g squamous carcinoma A-431 xenograft.101
212Pb	212Pb generator from AREVA Med, LLC	HER2	trastuzumab	TCMC	i.p. injection in patients with HER+ cancer.103–105
203/212Pb	212Pb generator from Oak Ridge National Labs; 203Pb from NIH	PSMA	Small-molecule PSMA ligands (L1–5)	TCMC ; DOTA	203Pb-SPECT imaging and 212Pb α-radiotherapy in NSG mice bearing PC3 PIP prostate cancer xenograft.114
203Pb	203Pb from Lantheus Medical Imaging, LLC	PSMA	Small-molecule PSMA ligands (CA-008-CA-012)	TCMC	203Pb-SPECT imaging in balb/c nu/nu mice bearing C4–2 prostate cancer xenograft; 203Pb- planar scan castration-resistant metastasized prostate cancer.115
203/212Pb	203Pb and 212Pb generator from AlphaMed, Inc.	MC1R	α-MSH analogue ReCCMSH	DOTA	203Pb-SPECT/CT image and 212Pb α-radiotherapy in C57BL mice bearing B16F0 melanoma xenograft.33, 37
203Pb	212Pb generator from Oak Ridge National laboratory; 203Pb from Lantheus Medical Imaging, LLC	MC1R	α-MSH analogue ReCCMSH	DOTA	Biodistribution of [203/212Pb]DOTA-ReCCMSH in C57BL mice bearing B16F0 melanoma xenograft.120
203Pb	203Pb from Lantheus Medical Imaging, LLC	MC1R	α-MSH analogue GGNle-CycMSHhex	DOTA	Biodistribution and SPECT/CT image of 203Pb-DOTA-GGNle-CycMSHhex in  C57 mice bearing B16F10 flank xenograft and pulmonary B16F10 lesions.122
212Pb	212Pb generator from AREVA Med, LLC	SSTR2	Octreotate analogue	TCMC	Athymic nude mice were bearing AR42J rat pancreatic xenograft.125
212Pb	212Pb generator from AREVA Med, LLC	SSTR2	Octreotate analogue	TCMC	Patients with SSTR+ neuroendocrine tumors.126
212Pb	212Pb generator from Orano Med	VCAM	αVCAM-1 mAb	TCMC	MDA-231-Br breast cancer brain metastasis model of in nu/nu mice.134
203/212Pb	212Pb generator from Oak Ridge National laboratory; 203Pb from Lantheus Medical Imaging, LLC	GRPR	Bombesin analogue RM2	DOTA	Biodistirbution of 203/212Pb-RM2 in ICR-SCID male mice bearing PC-3 prostate cancer xenograft.144, 145
212Pb	212Pb generator from Oak Ridge National laboratory	B7-H3	mAb 376.96	TCMC	i.p. xenograft of ES-2 and A2780cp20 human ovarian cancer.157, 158
212Pb	212Pb generator from Oak Ridge National laboratory	CSPG4	mAb 225.28	TCMC	SUM159 and 2LMP breast cancer xenograft.161, 162
212Bi	212Pb generator from University of Chicago	IL-2	Anti-Tac	DTPA	T-cell leukemia cells HUT-102B2.163
212Pb	212Pb generator from Orano Med	CD38	daratumumab	TCMC	Mice bearing s.c. RPMI8226 xenograft.165
212Pb	212Pb generator from AlphaMed, Inc.	HER2	Trastuzumab	TCMC	In combination with gemcitabine and paclitaxel in HER2+ tumor xenograft models.166–168
203/212Pb	212Pb generator from Oak Ridge National laboratory; 203Pb from Lantheus Medical Imaging, LLC	MC1R	α-MSH analogue ReCCMSH	DOTA	In combination with MAPK inhibitors and HDAC inhibitors in A2058, MEWO melanoma xenografts.175