Abstract 5275: Conjugation and radiolabeling of monoclonal antibodies and other proteins with 89Zr or 68Ga for PET imaging using p-isothiocyanatobenzyl-desferrioxamine
Authors: Vosjan Maria Jwd , Perk Lars R. , Visser Gerard Wm , Jurek Paul , Kiefer Garry E. , van Dongen Guus Ams 
Published: June 2012

Presently, hundreds of intact monoclonal antibodies (mAbs) and mAb fragments are under clinical development because of their excellent potential treatment of cancer and other pathological conditions. Positron emission tomography (PET) offers an exciting imaging option to confirm and quantify selective tumor uptake of such targeting molecules.

To enable PET imaging of mAbs (immuno-PET), an appropriate positron emitter, with a half-life (t1/2) that is compatible with the time needed to achieve optimal tumor-to-non tumor ratios (typically 2-4 days for intact mAbs and &lt; 3 h for mAb fragments), has to be securely coupled to the targeting molecule. Zirconium-89 (89Zr; t1/2:78.4 h) was selected for PET imaging of intact mAbs. For this purpose, we developed the large scale production of radionuclidic pure 89Zr, and a strategy for labeling mAbs with 89Zr by a multi-step synthesis using a succinylated-derivative of desferrioxamine B (Df) as bifunctional chelate (BFC). The recent commercialization of 89Zr has made this radionuclide available for research and clinical development. In the past, Df has also been used for coupling of gallium isotopes to mAbs. Gallium-68 (68Ga; t1/2:68 min) is cyclotron-independently available via a 68Ge/68Ga generator system, and as such an attractive isotope for PET imaging of fast-kinetic mAb fragments and non-traditional antibody-like scaffolds (e.g. Nanobodies®).

Since the aforementioned multi-step labeling approach is relatively complicated and time consuming the new BFC p-isothiocyanatobenzyl-desferrioxamine (Df-Bz-NCS; Macrocyclics, TX) was developed. In this study, we present an efficient and more rapid preparation of 89Zr-mAbs and 68Ga-Nanobodies, whereby conjugates are now prepared in 2 instead of 6 steps. In vitro stability measurements were performed, followed by biodistribution and PET studies in nude mice bearing tumor xenografts.

Labeling of mAbs with 89Zr using the Df-Bz-NCS BFC resulted in radiochemical yields &gt;85% after 60 min at room temperature (RT). Labeling of Nanobodies with 68Ga via the same BFC resulted in radiochemical yields of 55-70% (not corrected for decay) after 5 min at RT. All labeled compounds showed preserved integrity and immunoreactivity while the stability in human serum at 37°C was good: &lt;5% radioactivity loss for the 89Zr-mAbs during a 7 days incubation period, and &lt;2% radioactivity loss during a 5 h period for the 68Ga-Nanobodies. In biodistribution and imaging experiments high and selective tumor uptake was observed for all labeled compounds.

The novel Df-Bz-NCS BFC allows efficient, rapid and easy preparation of radiolabeled mAbs or mAb-fragments with high tumor uptake in nude mice bearing human tumor xenografts. These achievements facilitate further exploration of immuno-PET as imaging tool in antibody development, target characterization and patient selection.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5275. doi:10.1158/1538-7445.AM2011-5275