The endocrine system comprises of a network of glands and organs that produce and release hormones to control various bodily functions. Endocrine disruptors, either natural or man-made, can affect the endocrine system by interfering with the normal actions of hormones. This can lead to serious health problems such as cancer, birth defects and developmental disorders. Many of the traditional endocrine disruption testing approaches use radiolabelled material or involve animal models and so have high cost and low efficiency, in addition to ethical concerns. Accurate low-cost screening models are required to flag up chemicals which are potential endocrine disruptors at an early stage.
At the Society of Toxicology (SOT) conference on March 10-14, 2024, Cyprotex presented a poster titled, ‘Establishment of a High Throughput Endocrine Disruptor Screening Panel of Assays for Rapid Screening of Chemicals’. The research developed a panel of 384 well high throughput cell-based hormone activation assays and in vitro hormone receptor binding assays with a 24-48 hr turnaround time. A range of chemicals with different potencies for the endocrine receptors were assessed.
Cell-based endocrine receptor activation assays expressing the human androgen receptor (AR), the human estrogen receptor (ERα or ERβ) or the the human thyroid hormone receptor (TR) were used to assess the chemicals at a range of concentrations at 37°C over 24 hr. Upon ligand activation, the hormone receptor binds to the promoter sequence linked to a luciferase reporter gene and the activation is quantified by luminescence. Cell viability was also assessed in parallel.
Hormone receptor competitor binding assays were also used to assess potential AR or ER ligands by measuring fluorescence polarization. A fluorescently-tagged ligand is added to the AR or ER in the presence of the competitor test chemical. If the test chemical binds then it prevents the formation of the fluorescent ligand/receptor complex and a decrease in polarisation is observed. The extent of the shift in polarization was used to determine the relative affinity of the test chemical for the hormone receptor.
The results observed for the known set of chemicals were consistent with literature values and data corresponded well between the two assays. Additional steroidogenesis assays are being developed to detect hormone levels of AR and ER in H295R cell culture supernatants following exposure to endocrine disruptors.