Preparation of ²¹¹At-Labeled Humanized Anti-Tac Using ²¹¹At Produced in Disposable Internal and External Bismuth Targets

Abstract

These studies describe the production and purification of ²¹¹At as well as the procedure for labeling humanized anti-Tac, the antibody to the α-chain of the IL-2 receptor (IL-2Rα), which has been shown to be a useful target for immunotherapy. The optimized protocol combines the advantages of the two-stage dry distillation procedure with the astatination of trialkylstannyl substances as labeling compounds for proteins. The ²¹¹At was produced by bombarding either an external or a recently developed disposable internal bismuth target with α-particles from a Cyclotron Corporation CS-30 cyclotron. The ²¹¹At was found to contain less than 0.01% ²¹⁰At. The production rate for the external target was 0.15 mCi ± 0.056 μA⁻¹ h⁻¹ (n = 9) (5.55 MBq μA⁻¹ h⁻¹). The production rate for the internal target was 0.44 ± 0.14 mCi μA⁻¹ h⁻¹ (n = 16) (16.28 MBq μA⁻¹ h⁻¹).

Introduction

Radioactive-labeled antibodies are attractive cancer therapeutics because of the cytotoxicity of particle emitters at distances of several cell diameters. α-Particles show a much higher linear energy transfer (LET) with a range of only a few cell diameters compared to the lower LET and much longer range of β-particles, which may make α-emitters superior for destroying tumors and minimizing damage to distant normal tissues.

α-Emitting nuclides under investigation for use in radioimmunotherapy include ²¹²Bi, ²¹³Bi and ²¹¹At. This study focuses on ²¹¹At, whose half-life of 7.2 h is long enough to obtain high tumor uptake. The 100% high-energy α-particle emission (58.1% 7.45 MeV, 41.9% 5.9 MeV) produces a very efficient LET. The decay of ²¹¹At produces low amounts of γ-emissions, which enable the use of conventional single position emission computed tomography (SPECT) cameras to follow the in vivo distribution of ²¹¹At-labeled compounds, but does not require extensive shielding as is required for ²¹²Bi. A limiting factor for its use is availability since ²¹¹At is produced by the ²⁰⁹Bi(α,2n)²¹¹At reaction using α-particles from a cyclotron. Since there is no stable direct astatine-labeling procedure for proteins, additional steps to prepare and separate an astatinated compound for indirect labeling are required using a prosthetic group.

This paper describes the production and purification of ²¹¹At as well as the procedure for labeling antibodies. The optimized protocol combines the advantages of the two-stage dry distillation procedure of Doberenz et al. [1] with the astatination of trialkylstannyl substances as labeling compounds for proteins as described by Zalutsky et al. [12].

In previous studies, the α-chain of the interleukin-2 receptor (IL-2Rα) was shown to be a useful target for immunotherapy. The IL-2Rα is overexpressed on T cells from patients with certain lymphoid malignancies, select autoimmune diseases and allograft rejection, but not on normal resting cells 9, 10. Besides the murine antibody (anti-Tac), humanized anti-Tac was produced by genetic engineering combining the complementary-determining region of the murine antibody with constant and framework regions from human antibodies [8]. The product is less immunogenic in cynomolgus monkeys and manifested an antibody-dependent cellular cytotoxicity with human mononuclear cells that was absent with the parental mouse anti-Tac.

Recent experiments demonstrated the effectiveness of this antibody/antigen system in therapy using conjugates armed either with the β-emitter ⁹⁰Y or the α-emitting isotope ²¹²Bi 3, 4, 5 or with toxins [6].