Determination of binding affinity of molecular imaging agents for steroid hormone receptors in breast cancer

Kelley Salem; Manoj Kumar; Kyle C Kloepping; Ciara J Michel; Yongjun Yan; Amy M Fowler

Determination of binding affinity of molecular imaging agents for steroid hormone receptors in breast cancer

Am J Nucl Med Mol Imaging. 2018 Apr 25;8(2):119-126. eCollection 2018.

Authors

Kelley Salem¹, Manoj Kumar¹, Kyle C Kloepping^{1

2}, Ciara J Michel¹, Yongjun Yan^{1

3}, Amy M Fowler^{1

4

3}

Affiliations

¹ Department of Radiology, University of Wisconsin School of Medicine and Public Health 600 Highland Avenue, Madison, WI 53792, USA.
² Perkin Elmer Waltham, MA, USA.
³ Department of Medical Physics, University of Wisconsin School of Medicine and Public Health 1111 Highland Avenue, Madison, WI 53705, USA.
⁴ University of Wisconsin Carbone Cancer Center 600 Highland Avenue, Madison, WI 53792, USA.

PMID: 29755845
PMCID: PMC5944827

Abstract

16α-[¹⁸F]Fluoro-17β-estradiol ([¹⁸F]FES) and 21-[¹⁸F]-Fluoro-16α,17α-[(R)-(1'-α-furylmethylidene)dioxyl]-19-norpregn-4-ene-3,20-dione ([¹⁸F]FFNP) are being investigated as imaging biomarkers for breast cancer patients. Quantitative positron emission tomography (PET) reflects both total receptor content and binding affinity. To study factors that may alter radiopharmaceutical binding and impact PET accuracy, assays that can separate receptor amount from binding affinity are needed. The study purpose was to quantify the binding parameters of [¹⁸F]FES and [¹⁸F]FFNP in breast cancer. Estrogen receptor-alpha (ER) and progesterone receptor (PR) positive breast cancer cell lines (MCF-7 and T47D) were used to measure [¹⁸F]FES and [¹⁸F]FFNP binding parameters via saturation and competitive binding curves. The equilibrium dissociation constant (K_d) and total receptor density (B_max) were determined using nonlinear regression of the saturation binding curves. Half-maximal inhibitory concentration (IC50) was determined using nonlinear regression of the competitive binding curves. Linear correlation between increasing cell number and tracer uptake was observed for both [¹⁸F]FES and [¹⁸F]FFNP (R²=0.99 and 0.91, respectively). Using [¹⁸F]FES, the K_d for ER in MCF-7 cells was 0.13±0.02 nM with a B_max of 1901±89.3 fmol/mg protein and IC50 of 0.085 nM (95% CI: 0.069-0.104 nM). Using [¹⁸F]FFNP, the K_d for PR in T47D cells was 0.41±0.05 nM with a B_max of 1984±75.6 fmol/mg protein and IC50 of 2.6 nM (95% CI: 2.0-3.4 nM). The ligand binding function of ER and PR can be quantified using [¹⁸F]FES and [¹⁸F]FFNP and are comparable to previous studies using tritiated radioligands. [¹⁸F]FES and [¹⁸F]FFNP can be used in cell-based assays to quantify receptor-radioligand binding affinity, which cannot be obtained from a single PET examination.

Keywords: Estrogen receptor; Kd; [18F]FES; [18F]FFNP; breast cancer; equilibrium dissociation constant; progesterone receptor; radioligand binding affinity.