III-3. Radiochemistry & Nuclear Chemistry

RIKEN Accel. Prog. Rep. 55 (2022)

RIKEN Accel. Prog. Rep. 55 (2022)

Analysis of complex formation between rhenium and varioushydrophilic ligands using HPLC and preparation of 186 Re-carrying
liposomes
I. O. Umeda,∗1 Y. Hamane,∗1 M. Nagata,∗1 A. Tenzaki,∗1 H. Haba,∗2 A. Nambu,∗2 X. Yin,∗2 H. Fujii,∗3 andT. Takahashi∗1Theranostics is an emerging and expanding medical ﬁeld based on therapeutic interventions after
imaging to verify the presence of a biological target.
Radioisotope-based therapeutics, i.e., radiotheranostics, is perhaps the most clinically advanced application of theranostics, with many developments andemerging opportunities. A key aspect of radiotheranostics is that the selection of patients for radiotargeted treatments is based on imaging of the same target area. Therefore, imaging and therapeutic intervention are closely linked.1) Technetium-99m is themost widely used nuclide in diagnostic nuclear medical imaging, because the energy of the corresponding
gamma-ray emission (140 keV) is ideal for imaging using gamma cameras, and it can be generated on demand at medical facilities using 99 Mo/99m Tc generators. Rhenium, a group 7 homologous element of technetium, is assumed to have chemical properties similar
to those of Tc. 186 Re (half-life T1/2 = 3.72 days) emitsbeta rays of 1.07 MeV, appropriate for radionuclide
therapy. Thus, 186 Re and 99m Tc should be an ideal“theranostics pair.” To this end, 186 Re has been produced by neutron irradiation in nuclear reactors, which
is diﬃcult to realize in Japan. At the RIKEN RI beamfactory (RIBF), we succeeded in producing no-carrieradded, high-purity 186 Re using an accelerator-based
ion beam irradiation. We are aiming to develop new
radiotheranostics using 186 Re.
For radiotheranostics, a common platform for diagnostic and therapeutic nuclides is important. We havebeen investigating the application of liposomes, wellknown drug carriers, as a radiotheranostics platform.
Liposomes are promising tools for radiotheranostics,
because they can in principle encapsulate any radionuclides, and deliver them to the target lesion. In thisstudy, we conducted a basic investigation for eﬃcient
encapsulation of 186 Re into liposomes.
We encapsulated various radionuclides in liposomeswith high eﬃciency using the remote loading method,i.e., a ligand exchange reaction across the liposomal
membrane using lipophilic and hydrophilic ligands.2)
In this method, the complex formation between thenuclide and ligand and their stability are importantfactors. There are few data on the complex formationof 186 Re. We ﬁrst developed an analytical method toevaluate their formation and stability using high per∗1∗2∗3Kavli IPMU, University of Tokyo
RIKEN Nishina Center
EPOC, National Cancer Center- 139 -

formance liquid chromatography (HPLC) and a stable
isotope of rhenium.
Rhenium-186 was produced in the 186 W(d, 2n)186 Rereaction. A 24-MeV deuteron beam delivered fromthe AVF cyclotron was irradiated onto a 186 WO3pellet target (isotope enrichment of 186 W: 99.79%;
thickness: 580 mg/cm2 ). After irradiation, 186 Re
was puriﬁed by chemical separation.3) An aqueous solution of 186 Re-perrhenate in 0.01 M HCl(radioactivity concentration: 83.0 MBq/mL) wassupplied.
Ammonium perrhenate was used asa stable isotope of rhenium. DiethylenetriamineN, N, N  , N, N  -pentaacetic acid (DTPA), dimercaptosuccinic acid (DMSA), hydrazinonicotinamide(HYNIC), and ethylenedicysteine (EC) were examined as hydrophilic ligands. HPLC analysis was conducted using Shimazu Prominence LC-20 equippedwith aerosol-based detector NQAD and gamma-ray radiodetector. Liposomes comprised DSPC and cholesterol, or PEGylated, and preloaded hydrophilic ligands.
The formation of complexes between metal ions andligands is diﬃcult to analyze using conventional HPLC
systems, because these samples have no UV absorption, and are highly water-soluble. We realized thisusing unique columns, combination of cation exchange
column (CapcelPak CR) and unique ODS column(Capcell Core ADME), and a unique detector, NQAD.
Using this system, we were able to detect metal ions,ligands, and the metal ion-ligand complexes as separate peaks. Figure 1 shows HPLC proﬁles of rhenium,technetium, EC, and their complexes.
Using this system, the formation of complexes between rhenium and various hydrophilic ligands wasinvestigated. All the ligands used formed complexes
with 99m Tc. Under the conditions studied, DTPA andHYNIC did not form complexes with rhenium. In caseof DMSA and EC, complex formation with rhenium
was observed. However, unlike 99m Tc, the complexes
were formed only under acidic conditions, high temperature, and required approximately ten times more
reducing agent than in the case of 99m Tc.
Next, we preloaded EC into liposomes and attempted to encapsulate 186 Re and 99m Tc into liposomes using the remote loading method. Incubation
at 40◦ C allowed only 99m Tc to be encapsulated, but
heating to 90◦ C allowed 186 Re to be encapsulated intoliposomes. Since 186 Re radioactivity was present in
the water-soluble fraction of liposomes, it was assumed

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RIKEN Accel. Prog. Rep. 55 (2022)㻜㻜㻜Ȗ-ray㻝㻜㻝㻡㻜㻞㻜㻡㻝㻜㻝㻡㻞㻜㻜㻡㻝㻜㻝㻡㻞㻜

186ReO4- + EC

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Detector:㻡㻌㻡㻚㻢㻢㻠㻌㻜㻡㻜㻜㻞㻡㻜㻜

99mTcO4-㻜㻡㻝㻜㻝㻡㻞㻜

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їRetention time (min)Fig. 1. HPLC separation of metal ions, ligands and metalligand complexes. Stable rhenium was used to determine the analytical conditions. NQAD was used fordetection. 99m Tc and 186 Re were then used to conﬁrm
the peaks.

that the 186 Re-EC complex was formed.
In this study, we have established a means to analyze complex formation between hydrophilic ligands
and rhenium, and evaluated optimal conditions for stable rhenium complex formation. Based on the results,
we have successfully prepared 186 Re-EC/99m Tc-ECencapsulated liposomes. These are expected to be useful radiopharmaceuticals for radiotheranostics. We arefurther investigating the quality control of the drugs
and their pharmacokinetics and tumor accumulation.
References
1) K. Herrmann et al., Lancet Oncol. 21, e146 (2020).
2) I. Ogihara-Umeda et al., J. Nucl. Med. 37, 326 (1996).
3) N. Shigeta et al., J. Radioanal. Nucl. Chem. 205, 85(1996).

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