Preliminary Therapy Evaluation of (225)Ac-DOTA-c(RGDyK) Demonstrates that Cerenkov Radiation Derived from (225)Ac Daughter Decay Can Be Detected by Optical Imaging for In Vivo Tumor Visualization

Abstract

The theranostic potential of (225)Ac-based radiopharmaceuticals continues to increase as researchers seek innovative ways to harness the nuclear decay of this radioisotope for therapeutic and imaging applications. This communication describes the evaluation of (225)Ac-DOTA-c(RGDyK) in both biodistribution and Cerenkov luminescence imaging (CLI) studies. Initially, La-DOTA-c(RGDyK) was prepared as a non-radioactive surrogate to evaluate methodologies that would contribute to an optimized radiochemical synthetic strategy and estimate the radioactive conjugate's affinity for αvβ3, using surface plasmon resonance spectroscopy. Surface plasmon resonance spectroscopy studies revealed the IC50 and Ki of La-DOTA-c(RGDyK) to be 33 ± 13 nM and 26 ± 11 nM, respectively, and suggest that the complexation of the La(3+) ion to the conjugate did not significantly alter integrin binding. Furthermore, use of this surrogate allowed optimization of radiochemical synthesis strategies to prepare (225)Ac-DOTA-c(RGDyK) with high radiochemical purity and specific activity similar to other (225)Ac-based radiopharmaceuticals. This radiopharmaceutical was highly stable in vitro. In vivo biodistribution studies confirmed the radiotracer's ability to target αvβ3 integrin with specificity; specificity was detected in tumor-bearing animals using Cerenkov luminescence imaging. Furthermore, tumor growth control was achieved using non-toxic doses of the radiopharmaceutical in U87mg tumor-bearing nude mice. To our knowledge, this is the first report to describe the CLI of αvβ3 (+) tumors in live animals using the daughter products derived from (225)Ac decay in situ. This concept holds promise to further enhance development of targeted alpha particle therapy.

Keywords: Actinium-225; Cerenkov Luminescence Imaging; Targeted Alpha Particle Therapy; αvβ3 integrin..

Figure 1

Quality control of ²²⁵Ac-DOTA-c(RGDyK) using Radio-HPLC. UV-HPLC chromatogram (220 nm) of DOTA-c(RGDyK) (A) and nonradioactive La-DOTA-c(RGDyK) (B) compared with radio-HPLC chromatogram of ²²⁵Ac-DOTA-c(RGDyK) (C).

Figure 2

Biodistribution of ²²⁵Ac-DOTA-c(RGDyK). The radiotracer was injected into U87MG tumor bearing nude mice (n = 6/time point). While the radiotracer clears efficiently from the blood (A), clearance from the liver (B) and kidney (C) are more modest over time. (D) Tumor uptake is also relatively high.

Figure 3

In vivo blockade demonstrates specific targeting of α_vβ₃ integrin. α_vβ₃⁺ tumors demonstrate significantly more uptake of ²²⁵Ac-DOTA-c(RGDyK) when compared to α_vβ₃⁺ tumors in animals receiving the radiotracer and excess c(RGDyK) as blockade.

Figure 4

Cerenkov luminescence imaging with ²²⁵Ac-DOTA-c(RGDyK). Images were acquired 24 h after intravenous injection to allow for equilibrium to occur between ²²⁵Ac and its daughter products, which generate Cerenkov luminescence. (A) Efficient tumor localization was observed in U87mg tumor bearing animals at 24 h p.i. (B) Localization was reduced upon administration of blockade demonstrating in vivo specificity. (C, D) In vivo (C) and ex vivo (D) quantification based upon ROI analysis demonstrates that Cerenkov radiation in the tumor is significantly reduced upon the administration of blockade. (★) denotes tumor.