Engineering a Radiohybrid PSMA Ligand with an Albumin-Binding Moiety and Pharmacokinetic Modulation via an Albumin-Binding Competitor for Radiotheranostics

Abstract

The prostate-specific membrane antigen (PSMA) is a well-established target for radiotheranostics in prostate cancer. We previously demonstrated that 4-(p-astatophenyl)butyric acid (APBA), an albumin-binding moiety (ABM) labeled with astatine-211 (²¹¹At), enables the modulation of pharmacokinetics and enhancement of therapeutic efficacy when combined with the post-administration of an albumin-binding competitor. However, this strategy has not been explored in PSMA-targeting ligands. We designed and synthesized [²¹¹At]6, a novel PSMA ligand structurally analogous to PSMA-617 with APBA. The compound was obtained via a tin-halogen exchange reaction from the corresponding tributylstannyl precursor. Comparative cellular uptake and biodistribution studies were conducted with [²¹¹At]6, its radioiodinated analog [¹²⁵I]5, and [⁶⁷Ga]Ga-PSMA-617. To assess pharmacokinetic modulation, sodium 4-(p-iodophenyl)butanoate (IPBA), an albumin-binding competitor, was administered 1 h postinjection of [¹²⁵I]5 and [²¹¹At]6 at a 10-fold molar excess relative to blood albumin. The synthesis of [²¹¹At]6 gave a radiochemical yield of 15.9 ± 7.7% and a radiochemical purity > 97%. The synthesized [²¹¹At]6 exhibited time-dependent cellular uptake and internalization, with higher uptake levels than [⁶⁷Ga]Ga-PSMA-617. Biodistribution studies of [²¹¹At]6 in normal mice revealed a prolonged blood retention similar to those of [¹²⁵I]5. Notably, post-administration of IPBA significantly reduced blood radioactivity and non-target tissue accumulation of [¹²⁵I]5 and [²¹¹At]6. We found that ABM-mediated pharmacokinetic control was applicable to PSMA-targeted radiotherapeutics, broadening its potential for the optimization of radiotheranostics.

Keywords: PSMA; albumin-binding moiety; astatine-211; radiotheranostics; targeted alpha therapy (TAT).

Figure 1

Chemical structures of (a) PSMA-617, (b) [¹²⁵I]5 and [²¹¹At]6, and (c) [²¹¹At]PSMA5.

Scheme 1

Synthetic scheme of 4, 5, [¹²⁵I]5, and [²¹¹At]6. Reagents: (a) (i) 2-chlorotrityl chloride resin, Glu(OtBu)₂-urea-Lys(Fmoc)-OH, DIPEA, DCM, 2 h, room temperature (rt); (ii) 20% piperidine/DMF, 15 min, rt; (iii) Fmoc-amino acid or Tris-DOTA, HOBt, N,N-diisopropylcarbodiimide (DIC), DMF, 2 h, rt. (b) trifluoroacetic acid (TFA), TIS, H₂O (95/2.5/2.5 v/v/v), 2 h, rt. (c) Ga(NO₃)₃, 3 M ammonium acetate buffer (pH 5), 1 h, 40 °C. (d) 4-(p-iodophenyl)butanoate NHS ester, DIPEA, DMF, 5 h, rt. (e) 4-(p-tributylstannylphenyl)butanoate NHS ester, DIPEA, DMF, 4 h, rt. (f) [¹²⁵I]NaI/[²¹¹At]At⁻, chloramine-T, AcOH, MeOH, 15 min, 60 °C.

Figure 2

HPLC chromatograms of (a) 5, (b) [¹²⁵I]5, (c) [²¹¹At]6, and (d) [⁶⁷Ga]5. HPLC condition: a flow rate of 1 mL/min with a gradient mobile phase of 60% methanol in water with 0.1% TFA to 80% methanol in water with 0.1% TFA for 20 min.

Figure 3

Time-dependent cellular uptake of (a) [¹²⁵I]5 and [²¹¹At]6 and (b) [¹²⁵I]5 and [⁶⁷Ga]Ga-PSMA-617, as well as internalization of (c) [¹²⁵I]5 and [²¹¹At]6 and (d) [¹²⁵I]5 and [⁶⁷Ga]Ga-PSMA-617 in LNCaP cells. Specific binding of PSMA is blocked by excess amount (100 µM) of 2-PMPA. Data are presented as mean ± SD for three samples.

Figure 4

Biodistribution of radioactivity after the intravenous injection of (a) [¹²⁵I]5 and (b) [⁶⁷Ga]Ga-PSMA-617 in normal mice. S. Intestine and L. Intestine indicate the small intestine and large intestine, respectively. ^‡ Expressed as %ID. Each value represents the mean (SD) for three animals. The distribution data of [¹²⁵I]5 in Figure 4 and Figure 5 were evaluated for each experiment.

Figure 5

Biodistribution of radioactivity after the intravenous injection of (a) [²¹¹At]6 and (b) [¹²⁵I]5 in normal mice. S. Intestine and L. Intestine indicate the small intestine and large intestine, respectively. ^‡ Expressed as %ID. Each value represents the mean (SD) for three or four animals. The distribution data of [¹²⁵I]5 in Figure 4 and Figure 5 were evaluated for each experiment.

Figure 6

Comparison of the biodistribution in normal mice with or without administering IPBA at 1 h postinjection of [¹²⁵I]5 and [²¹¹At]6.