Citation

Junyi Chen, Mengxin Xu, Yu Liu, Dongban Duan, Yuxiang Han, Zhibo Liu. Isolation of ²¹²Pb from natural thorium for targeted alpha-therapy[J]. Chin. Chem. Lett., 2022, 33(7): 3474-3477  

Junyi Chen^a, Mengxin Xu^a, Yu Liu^b, Dongban Duan^a, Yuxiang Han^a, Zhibo Liu^a,b,*     

TAT (Targeted alpha-therapy) treats cancer by using α-particles [1-3], of which the linear energy transfer (LET) is notably higher than that of β-particles [4], resulting in lethal damage in cancer cells. α-Particles often induce DNA double-strand break, which can hardly repair, and are less dependent on the oxygen level in tumors [5]. The short range of α-particles (50–100 µm) confines the killing region to the lesion and reduces the negative effects on normal tissues [6]. α-Emitters exhibit better therapeutic efficacy than that of β-emitters in clinical studies, features with much less therapeutic dose and less radiation resistance [7, 8].

For instance, ²²⁵Ac-labeled radiopharmaceuticals have demonstrated exceptional performance in clinical studies [9-11]. However, ²²⁵Ac is conventionally generated from ²²⁹Th or ²³³U [12, 13], and its annual worldwide manufacturing capacity is less than 3 Ci [14], posing a barrier to the growth of TAT [15]. ²¹²Pb (t_1/2 = 10.6 h) decays to the stable ²⁰⁸Pb, which is also an α-emitting nuclide and can be radiolabeled with DOTA or other TAT chelators [4, 17].

Extracting ²¹²Pb from natural thorium will resolve the current shortage of α-emitters for TAT and provide a means of supplying α-emitters without relying on high-energy accelerators and reactors. Despite the fact that ²¹²Pb has been developed for TAT for many years, the available source of ²¹²Pb is rather insufficient. ²¹²Pb may be separated from the decay of ²²⁸Th, ²³²Th or ²³²U [18, 19]. Compare to ²³²Th, ²²⁸Th and ²³²U are preferred due to the high content of ²¹²Pb [16, 19]. However, both ²²⁸Th and ²³²U are not commercially available and can hardly be obtained by importation [20]. In contrast, ²³²Th is abundant, especially in China [21]. The low ²¹²Pb content and the consequent technical challenges have hampered the investigation of directly extracting ²¹²Pb from natural thorium. Though it may be the only realistic strategy to prepare ²¹²Pb in China [20], the reports about isolating ²¹²Pb from natural thorium are limited [22, 23].

In this work, we attempted to separate ²¹²Pb directly from natural thorium compounds for labeling radiopharmaceuticals. By solid-phase extraction with a Pb-selective resin (Pb-Spec^TM) [24], the ²¹²Pb was isolated directly from the thorium nitrate solution. 2.2 MBq of ²¹²Pb was obtained from 2.5 kg of thorium nitrate hydrate every 48 h, representing 54% of the theoretical maximal activity and 83% of the ²¹²Pb separation efficiency. To validate the quality of the prepared ²¹²Pb, it was applied to label several peptide conjugates of clinical importance, DOTATATE [25], PSMA-617 [26] and FAPI-04 [27, 28], with good radiochemical yields (over 88%).

²³²Th is abundant and often considered as radioactive waste from mining. The ineffective utilization and treatment of thorium-containing slag not only wastes resources, but also causes major environmental issues when radionuclides from ²³²Th enter the atmosphere as dust or enter the land and rivers via groundwater [29]. The decay of ²³²Th produces ²¹²Pb, which is an α-emitter and can be exploited for TAT. The activity of ²¹²Pb in natural thorium is 4.07 kBq/g ²¹²Th at decay equilibrium [30], and it may be extracted directly from thorium compounds for radiopharmaceuticals. In the decay chain, ²¹²Pb was accumulated via ²²⁴Ra (α, 3.7 d) ²²⁰Rn (α, 55.6 s) ²¹⁶Po (α, 0.14 s) ²¹²Pb. According to the aforementioned, shown in Fig. 1a, ²¹²Pb in ^232/228Th can be recovered 77% within 24 h and up to 93% within 48 h following the separation of ²¹²Pb, which satisfies the nuclide generator's requirement.

Figure 1

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Fig. 1. (a) Recovering of 212Pb from 228Th and 224Ra, (b) 232/228Th decay equilibrium curve, and (c) γ-spectroscopy of commercially available thorium compounds.

The decay equilibrium curve of ^232/228Th is shown in Fig. 1b. Right after the isolation of thorium compounds, the activity of ²¹²Pb gradually declines due to the rapid decay of ²²⁸Th, eventually reaching the lowest value of 1.7 kBq/g ^natTh in the fourth year. Following that, as ²³²Th decays to ²²⁸Th, the ²¹²Pb activity grows gradually until it reaches decay equilibrium.

As shown in Fig. 1c, the γ-spectrum of commercial thorium nitrate. As an inexpensive and easily accessible thorium compounds, thorium nitrate contains radionuclides from the thorium decay chain, ensuring its purity as a raw material for ²¹²Pb generators. The acquired thorium compound had not reached decay equilibrium, as the ²¹²Pb activity was only 1.96 kBq/g ^natTh, or 48% of equilibrium activity, resulting in additional thorium compound.

Thorium nitrate hydrate (2.5 kg) was dissolved in 5 L HNO₃ solution (1 mol/L) with a thorium concentration of around 1 mol/L. According to previous results, this solution had an activity concentration of around 386 kBq/L and total activity of 1.93 MBq.

Pb-Spec^TM is a extraction chromatographic resin by sorbing 4, 4′, (5′)-di-(t-butyldicyclohexano)−18-crown-6 (DtBu-CH18C6) in isodecanol solution on an inert substrate for the separation and preconcentration of radiolead [24, 31]. Lead has a good retention on Pb-Spec^TM under a wide range of nitric acid concentration (0.1–8 mol/L). Under 1 mol/L HNO₃, the D_w of the PB-Spec^TM was around 1.5 × 10³ (k' ~ 8 × 10²) [24], indicating that the extraction column with 6 mL resin (2.3 g resin, Φ12.5 mm × 50 mm, 4 mL free column volume) was adequate as the extraction resin for a more efficient ²¹²Pb extraction.

To trap ²¹²Pb with high efficiency, 5 L of a 1 mol/L Th(NO₃)₄/1 mol/L HNO₃ solution was loaded onto the extraction column at a flow rate of 0.5 L/h. The column was then washed with 5 mL 1 mol/L HNO₃ and 5 mL H₂O to remove residual Th⁴⁺ and to reduce the acidity. Finally, Pb²⁺ was eluted with 7 mL 0.5 mol/L (NH₄)₂HCit [31], which is suitable for radiopharmaceutical labeling directly.

As shown in Fig. 2a, the column trapped Pb but not Th, Ra or Ac. The radioactivity of ²¹²Pb fell from 445 Bq/mL to 68 Bq/mL in the elution, with an 85% retention ratio, suggesting that the column can efficiently separate and enrich ²¹²Pb. The generator could rapidly regenerate for the next ²¹²Pb separation. The change of ²¹²Pb activity in the column effluent is described in Fig. 2b. At the beginning of loading, the column retained 100% of the ²¹²Pb, but as the loading volume increased, small amount of ²¹²Pb leakage was observed. The average extraction efficiency of ²¹²Pb was 83%, yielding 1.8 MBq ²¹²Pb. As demonstrated in Fig. 2c, 7 mL 0.5 mol/L (NH₄)₂HCit could efficiently elute 95% ²¹²Pb from the column. We analyzed the samples that had reached decay equilibrium in the next day, and found that the radioactivity of ²¹²Pb was 2.2 MBq at the end of purification.

Figure 2

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Fig. 2. Pb adsorption and elution by Pb-SpecTM extraction column. (a) 212Pb, 224Ra and 228Ac activities in the loading and effluent of the column. (b) 212Pb activity curves in the extraction column loading and effluent. (c) Extraction column 212Pb elution yield in each fraction.

The γ-spectrum of obtained ²¹²Pb was shown in Fig. 3a. All visible peaks were ascribed to ²¹²Pb and its daughter, confirming the greater purity of ²¹²Pb product obtained by this approach. To further evaluate the ²¹²Pb's purity, it was utilized to label a couple of peptides. Although Cit³⁻ can be coordinated with Pb²⁺, it was demonstrated that 0.5 mol/L (NH₄)₂HCit did not affect the labeling of Pb²⁺. Additionally, 0.5 mol/L (NH₄)₂HCit with a pH of 5.5 may be an ideal buffer for ²¹²Pb labeling with DOTA. The ²¹²Pb labeling reaction was carried out for 10 min at 95 ℃, and the reaction's progress and yield were monitored using radio-iTLC (Na₃Cit as mobile phase, Fig. 3b). Table 1 summarizes the labeling conditions and results for [²¹²Pb]Pb-FAPI-04, [²¹²Pb]Pb-PSMA-617, and [²¹²Pb]Pb-DOTATATE (chemical structure shown in Fig. 4). All radiochemical yields (RCY) were greater than 88%, and isolated yields were greater than 83%, indicating that the ²¹²Pb produced using this method was reliable for radiopharmaceutical investigations.

Figure 3

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Fig. 3. (a) γ-spectrum of 212Pb product. (b) Radio-iTLC of free [212Pb]Pb2+, reaction mixture, and purified 212Pb-labeled compound during the 212Pb labeling of FAPI-04.

Table 1

Table 1 Conditions and results of 212Pb labeling.a

Table 1 Conditions and results of 212Pb labeling.a

Figure 4

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Fig. 4. Chemical structure of [212Pb]Pb-FAPI-04, [212Pb]Pb- DOTATATE and [212Pb]Pb- PSMA-617.

In the above experiment, we successfully obtained 2.2 MBq ²¹²Pb from a 5 L thorium nitrate solution. However, to obtain more ²¹²Pb, additional thorium nitrate solution is needed, which may be difficult in the lab. Another solution to obtain ²¹²Pb based on ^natTh is to extract ^228/224Ra from the matrix and enrich them to isolate ²¹²Pb in a limited volume of ^228/224Ra, which would be easier to generate a higher activity of ²¹²Pb. Therefore, the separation and enrichment of ^228/224Ra from natural thorium compounds are ongoing to construct next-generation of ²¹²Pb generator. In addition, in the process of rare earth and uranium mining, 'radium removal residue' is a kind of radium-containing waste residue in the form of Ba(Ra)SO₄, and the amount of ^228/224Ra is considerable. If the waste could be used as a source of ^228/224Ra to prepare ^228/224Ra-²¹²Pb generators, it would be a "one-stone two-bird" strategy that make full use of the hazardous radioactive waste to produce high-value medical radionuclides.

To summarize, this study aims to extract and purify α-emitter ²¹²Pb from natural thorium compounds for use in TAT radiopharmaceuticals. 2.2 MBq of ²¹²Pb was extracted from 5 L of thorium nitrate solution using Pb-Spec^TM with an extraction efficiency of 83% and a high purity. The ²¹²Pb in 0.5 mol/L (NH₄)₂HCit solution produced could be used directly for the radiolabeling of DOTATATE, PSMA-617 and FAPI-04, which all achieved greater than 88% RCY and 83% isolation yield. The aforementioned results demonstrate that modest amounts of ²¹²Pb can be isolated directly from natural thorium compounds for developing ²¹²Pb-labeled radiopharmaceuticals for targeted alpha-therapy.

We declare that we have no financial and personal relationships with other people or organizations that can inappropriately influence our work, and there is no professional or other personal interest of any nature or kind in any product, service and/or company that could be construed as influencing the position presented in, or the review of, the manuscript entitled.

This work was funded by Beijing Municipal Natural Science Foundation (No. Z200018), the Special Foundation of Beijing Municipal Education Commission (No. 3500–12020123), the National Natural Science Foundation of China (Nos. U1867209 and 21778003) and the Ministry of Science and Technology of the People's Republic of China (No. 2017YFA0506300) and Li Ge-Zhao Ning Life Science Youth Research Foundation (No. LGZNQN202004).