18F-16α-17β-Fluoroestradiol Binding Specificity in Estrogen Receptor–Positive Breast Cancer

Purpose
To determine the binding specificity of 18F-16α-17β-fluoroestradiol (FES) in estrogen receptor (ER) α–positive breast cancer cells and tumor xenografts.

Materials and Methods
Protocols were approved by the office of biologic safety and institutional animal care and use committee. By using ER-negative MDA-MB-231 breast cancer cells, clonal lines were created that expressed either wild-type (WT; 231 WT ER) or G521R mutant ERα (231 G521R ER), which is defective in estradiol binding. ERα protein levels, subcellular localization, and transcriptional function were confirmed. FES binding was measured by using an in vitro cell uptake assay. In vivo FES uptake was measured in tumor xenografts by using small-animal positron emission tomographic/computed tomographic imaging of 24 mice (17 WT ER tumors, nine mutant G521R ER tumors, eight MDA-MB-231 tumors, and four MCF-7 ER-positive tumors). Statistical significance was determined by using Mann-Whitney (Wilcoxon rank sum) test.

Results
ERα transcriptional function was abolished in the mutated 231 G521R ER cells despite appropriate receptor protein expression and nuclear localization. In vitro FES binding in the 231 G521R ER cells was reduced to that observed in the parental cells. Similarly, there was no significant FES uptake in the 231 G521R ER xenografts (percent injected dose [ID] per gram, 0.49 ± 0.042), which was similar to the negative control MDA-MB-231 xenografts (percent ID per gram, 0.42 ± 0.051; P = .20) and nonspecific muscle uptake (percent ID per gram, 0.41 ± 0.0095; P = .06).

Conclusion
This study showed that FES retention in ER-positive breast cancer is strictly dependent on an intact receptor ligand-binding pocket and that FES binds to ERα with high specificity. These results support the utility of FES imaging for assessing tumor heterogeneity by localizing immunohistochemically ER-positive metastases that lack receptor-binding functionality.

© RSNA, 2017

Online supplemental material is available for this article.

File: Salem-Kumar-et-al-Radiology-FES-binding-specificity-in-ER-positive-breast-cancer.pdf

A Calibrating Procedure for a Bone Loading System

García-Rodríguez et al. Trabecular bone tissue is a three-dimensional structure that is difficult to duplicate with in vitro cell cultures or animal models. In an attempt to better understand the underlying
mechanisms of tissue response to load, a system to load isolated bone preparations was developed. This ex vivo bone culture and loading system, given the name of ZETOS, compressively loads trabecular bone (10 mm diameter, 5.0 mm height) to evaluate its morphological and physiological responses while keeping cells viable. Compliance of the system may change with time, thus requiring recalibration. The purpose of this research
was to develop and validate a recalibration protocol for the ZETOS system. Ten reference bodies (RBs) were designed and machined out of aluminum 7075-T6, with a structural rigidity range representative of trabecular bone (0.628–28.3 N/m, or apparent elastic modulus of 40 MPa–1.80 GPa). Finite element analysis (FEA) was used to calculate the rigidity of each RB and was validated with physical testing in a universal testing machine.
Results from FEA were then used to calibrate the system and relate force, piezoelectric actuator expansion, and specimen compressive deformation through a surface generated by spline interpolation, thus creating a calibration table. Calibration of ZETOS was verified by testing the RBs as well as three custom-made, metal springs and comparing measured rigidity to that calculated by FEA. Mean percent difference of FEA results with respect to those from physical testing was 3.28%. The mean percent difference of RB rigidity found with ZETOS with respect to rigidity found with FEA was 1.12% and for the metal springs, the mean percent difference was 1.74%. The calibration procedure for the ZETOS bone loading system has been successfully applied and verified. The use of RBs and FEA allows users to easily and periodically evaluate and recalibrate
the system. Accuracy in studies of human bone mechanotransduction in a controlled environment can therefore be achieved. The recalibration procedure is relevant for other ZETOS users and may serve as the basis for calibration of other testing systems for small specimens of compliant materials.

File: Garcia_et_al.pdf

Alkylphosphocholine Analogs for Broad Spectrum Cancer Imaging and Therapy

Weichert et al.
Many solid tumors contain an over-abundance of phospholipid ethers relative to normal cells. Capitalizing on this difference, we created cancer-targeted alkylphosphocholine (APC) analogs through structure activity analyses. Depending on the iodine isotope used, radioiodinated APC analog CLR1404 was used as either a PET imaging (124I) or molecular radiotherapeutic (131I) agent. CLR1404 analogs displayed prolonged tumor-selective retention in 55 in vivo rodent and human cancer and cancer stem cell models. 131I-CLR1404 also displayed efficacy (tumor growth suppression, survival extension) in a wide range of human tumor xenograft models. Human PET/CT and SPECT/CT imaging in advanced cancer patients with 124I- or 131I-CLR1404, respectively, demonstrated selective uptake and prolonged retention in both primary and metastatic malignant tumors. Combined application of these chemically identical APC-based radioisosteres will enable personalized dual modality cancer therapy of using molecular 124I-CLR1404 tumor imaging for planning 131I-CLR1404 therapy.

File: Weichert_et_al_Alkylphosphocholine.pdf

Apparent elastic modulus of ex vivo trabecular bovine bone increases with dynamic loading

Vivanco et al.
Although it is widely known that bone tissue responds to mechanical stimuli, the underlying biological control is still not completely understood. The purpose of this study was to validate required methods necessary to maintain active osteocytes and minimize bone tissue injury in an ex vivo three-dimensional model that could mimic in vivo cellular function. The response of 22 bovine trabecular bone cores to uniaxial compressive load was investigated by using the ZETOS bone loading and bioreactor system while perfused with culture medium for 21 days. Two groups were formed, the ‘‘treatment’’ group (n = 12) was stimulated with a physiological compressive strain (4000me) in the form of a ‘‘jump’’ wave, while the ‘‘control’’ group (n = 10) was loaded only during three measurements for apparent elastic modulus on days 3, 10, and 21. At the end of the experiment, apoptosis and active osteocytes were quantified with histological analysis, and bone formation was identified by means of the calcium-binding dye, calcein. It was demonstrated that the treatment group increased the elastic modulus by 61%, whereas the control group increased by 28% (p\0.05). Of the total osteocytes observed at the end of 21 days, 1.7% (60.3%) stained positive for apoptosis in the loaded group, whereas 2.7% (60.4%) stained positive in the control group. Apoptosis in the center of the bone cores of both groups at the end of 21 days was similar to that observed in vivo. Therefore, the three-dimensional model used in this research permitted the investigation of physiological responses to mechanical loads on morphology adaptation of trabecular bone in a controlled defined load and chemical environment.

File: Vivanco_et_al_Apparent_elastic_modulus_of_ex_vivo.pdf

Application of a whole-body pharmacokinetic model for targeted radionuclide therapy to NM404 and FLT

Grudzinski et al. We have previously developed a model that provides relative dosimetry estimates for targeted radionuclide therapy (TRT) agents. The whole-body and tumor pharmacokinetic (PK) parameters of this model can be noninvasively measured with molecular imaging, providing a means of comparing potential TRT agents. Parameter sensitivities and noise will affect the accuracy and precision of the estimated PK values and hence dosimetry estimates. The aim of this work is to apply a PK model for TRT to two agents with different magnitudes of clearance rates, NM404 and FLT, explore parameter sensitivity with respect to time and investigate the effect of noise on parameter precision and accuracy. Twenty-three tumor bearing mice were injected with a ‘slow-clearing’ agent, 124I-NM404 (n = 10), or a ‘fast-clearing’ agent, 18FFLT (3-deoxy-3-fluorothymidine) (n = 13) and imaged via micro-PET/CT pseudo-dynamically or dynamically, respectively. Regions of interest were drawn within the heart and tumor to create time-concentration curves for blood pool and tumor. PK analysis was performed to estimate the mean and standard error of the central compartment efflux-to-influx ratio (k12/k21), central elimination rate constant (kel), and tumor influx-to-efflux ratio (k34/k43), as well as the mean and standard deviation of the dosimetry estimates. NM404 and FLT parameter estimation results were used to analyze model accuracy and parameter sensitivity. The accuracy of the experimental sampling schedule was compared to that of an optimal sampling schedule found using Cramer–Rao lower bounds theory. Accuracy was assessed using correlation coefficient, bias and standard error of the estimate normalized to the mean (SEE/mean). The PK parameter estimation of NM404 yielded a central clearance, kel (0.009 ± 0.003 h−1), normal body retention, k12/k21 (0.69 ± 0.16), tumor retention, k34/k43 (1.44 ± 0.46) and predicted dosimetry, Dtumor (3.47 ± 1.24 Gy). The PK parameter estimation of FLT yielded a central elimination rate constant, kel (0.050 ± 0.025 min−1), normal body retention, k12/k21 (2.21 ± 0.62) and tumor retention, k34/k43 (0.65 ± 0.17), and predicted dosimetry, Dtumor (0.61 ± 0.20 Gy). Compared to experimental sampling, optimal sampling decreases the dosimetry bias and SEE/mean for NM404; however, it increases bias and decreases SEE/mean for FLT. For both NM404 and FLT, central compartment efflux rate constant, k12, and central compartment influx rate constant, k21, possess mirroring sensitivities at relatively early time points. The instantaneous concentration in the blood, C0, was most sensitive at early time points; central elimination, kel, and tumor efflux, k43, are most sensitive at later time points. A PK model for TRT was applied to both a slowclearing, NM404, and a fast-clearing, FLT, agents in a xenograft murine model. NM404 possesses more favorable PK values according to the PK TRT model. The precise and accurate measurement of k12, k21, kel, k34 and k43 will translate into improved and precise dosimetry estimations. This work will guide the future use of this PK model for assessing the relative effectiveness of potential TRT agents.

File: Grudzinski_et_al_Application_of_a_whole-body_pharmacokinetic.pdf