99mTc-labeled estradiol as an estrogen receptor probe: Preparation and preclinical evaluation

Introduction: Most breast cancers express estrogen receptors (ERs). Noninvasive imaging of ER expression may be helpful for planning therapy of ER+ tumors. We developed a new ER- binding probe, Tc-99m-labeled estradiol, with diethylenetriaminepentaacetic acid (DTPA) as a chelating ligand, and assessed its targeting ability in vitro and in vivo. Methods: 3-Aminoethyl estradiol was synthesized in two steps from estrone, followed by Tc-99m labeling. Western blotting and immunofluorescence staining were used to detect ER expression in MCF-7 and MDA-MB-231 breast cancer cells. Saturation binding and specific binding were performed by incubating MCF-7 cells with increasing concentrations of Tc-99m-DTPA-estradiol. Cell uptake, efflux, and blocking assays were also performed. To test Tc-99m-DTPA-estradiol in vivo, nude mice bearing either MCF-7- (high ER expression) or MDA-MB-231- derived tumors (low ER expression) were injected with Tc-99m-DTPA-estradiol, and underwent single-photon emission-computed tomography (SPECT). Mice injected with excess unlabeled DTPA-estradiol were used as controls. Ex vivo gamma-counting of tissues from normal and tumor-bearing mice was used to evaluate Tc-99m-DTPA-estradiol biodistribution. Results: The radiochemical purity of Tc-99m-DTPA-estradiol was 98.3% +/- 1.3% with a specific activity of 33.1 +/- 1.5 MBq/mu mol (n = 3). Western blotting and immunofluorescence staining confirmed extensive expression of ERs by the MCF-7 cells, and less extensive expression by MDA-MB-231 cells. There was high binding affinity of 99mTc-DTPA-estradiol to MCF-7 cells with a > 45% specific rate of total cell uptake. SPECT images and the biodistribution study results showed significantly higher uptake by MCF-7 tumors (6.06 +/- 0.38 %ID/g) than by MDA-MB-231 tumors (1.57 +/- 028 %ID/g). Pre-injection of MCF-7 tumor-bearing nude mice with excess unlabeled DTPA-estradiol significantly reduced tumor uptake of Tc-99m-DTPA-estradiol (2.24 +/- 0.28 %ID/g), suggesting that Tc-99m-DTPA-estradiol specifically targets ERs in tumors. Conclusions: Tc-99m-DTPA-estradiol can be synthesized with satisfactory labeling efficiency and stability. Tc-99m-DTPA-estradiol specifically targeted ERs in vitro and in vivo with favorable pharmacokinetics, allowing ER receptor expression assessment with SPECT imaging. (C) 2015 Elsevier Inc. All rights reserved.