Secretory Bioluminescent Biosensor Quantifies Glucocorticoid Activity In Vitro and In Vivo with High Specificity

Accurate assessment of glucocorticoid activity is crucial for understanding endocrine regulation and stress responses. Current methods like immunoassays and electrochemical sensors often struggle to differentiate structurally similar glucocorticoids and their metabolites, leading to potential inaccuracies in functional assessment. While mass spectrometry offers high specificity, it typically provides endpoint measurements and requires extensive sample processing, hindering real-time monitoring. There's a significant gap for a dynamic, functional, and real-time method to evaluate glucocorticoid receptor (GR) activation, which is essential for drug screening, mechanistic studies, and physiological monitoring.

Researchers developed a secretion-based genetically encoded biosensor for glucocorticoid detection. This sensor employs intein-mediated protein splicing triggered by glucocorticoid receptor (GR) activation. Upon ligand binding, GR nuclear translocation brings split intein domains into proximity, initiating protein trans-splicing. This process reconstitutes a split secretory peptide, leading to the secretion of a luciferase reporter into the extracellular space. The sensor was tested in vitro for dose-dependent responses to various glucocorticoids (e.g., corticosterone, dexamethasone), selectivity against inactive analogues, and response time. For in vivo assessment, encapsulated sensor cells were implanted in mice to monitor systemic corticosterone fluctuations under both basal and restraint-stress conditions.

The developed biosensor demonstrated robust dose-dependent luminescent responses to glucocorticoids across a wide dynamic range of 0.1 nM-10 μM. It exhibited a low detection limit of 10 nM and a remarkably rapid response, detectable within 5 min of ligand exposure. Crucially, the sensor selectively distinguished functional GR agonists from structurally related inactive analogues, highlighting its specificity for active glucocorticoid signaling. When encapsulated sensor cells were implanted in mice, the platform successfully provided extracellular luminescent readouts of systemic corticosterone. This enabled the detection of endogenous glucocorticoid fluctuations under both basal physiological states and in response to restraint-stress conditions. This real-time monitoring capability represents a significant advancement.

The sensor's ability to provide dynamic, functionally relevant assessment of glucocorticoid activity in vivo under physiological and stress conditions is a key breakthrough.