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

摘要

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 (Kd) and total receptor density (Bmax) 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 Kd for ER in MCF-7 cells was 0.13±0.02 nM with a Bmax of 1901±89.3 fmol/mg protein and IC50 of 0.085 nM (95% CI: 0.069-0.104 nM). Using [¹⁸F]FFNP, the Kd for PR in T47D cells was 0.41±0.05 nM with a Bmax 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.