Measuring LH Pulsatility in Patients with Reproductive Disorders using a Novel Robotic Aptamer-Enabled Electrochemical Reader (RAPTER)

Abstract

Normal reproductive functioning is critically dependent on pulsatile secretion of LH. Assessment of LH pulsatility is important for the clinical diagnosis of patients with reproductive disorders, but this is not routinely available in the clinic because of the need for frequent blood sampling coupled with expensive serial immunochemical analysis. The emerging technology of electrochemical aptamer-based (E-AB) sensing could potentially offer a generalizable approach for LH pulsatility measurement in humans. E-AB sensors take advantage of the intrinsic properties of nucleic acid aptamers to specifically bind to a molecular target and undergo a reversible conformational change. The conformational change can be measured through the electrochemical signal response. Aptamers, unlike antibodies, can facilitate low-cost repeat analytical measurements. However, there is currently no aptamer-based LH sensing technology. Here, we report the development and application of a novel antibody-free DNA aptamer-mediated electrochemical analysis to accurately assess LH pulsatility in patients with reproductive disorders. Through selective evolution of ligands by exponential enrichment (SELEX), single-stranded DNA oligonucleotides (aptamers) were identified that bind specifically to LH (and that do not bind to the closely related FSH). The aptamers were integrated into E-AB sensors on a robotic platform (which we term RAPTER) to enable sensitive, rapid and repeatable detection of LH. We next determined if the RAPTER system could measure LH pulsatility in clinical samples from patients with reproductive disorders. We obtained 441 serum samples from 3 patient cohorts - young females with regular menstrual cycles (normal LH pulsatility), menopausal women (disordered LH pulsatility, high LH), and women with hypothalamic amenorrhoea (suppressed/ absent LH pulsatility, low LH). Blood sampling was performed every 10 minutes for 8 hours in these patients and samples were analysed using both the current gold standard LH immunochemical assay and RAPTER. We performed a Bland-Altman analysis and plotted the linear regression to compare the results obtained. The RAPTER assay showed an almost perfect correlation with the clinical immunochemical assay (R2= 0.94). Bayesian Spectrum Analysis was used to determine the effective LH pulse interval from both datasets (clinical assay and RAPTER assay). We are able to distinguish 3 patient cohorts based on the time interval ranges. Taken together, the RAPTER system provides a new approach for LH pulsatility determination by reliably and immediately calculating varying LH concentrations within distinct patient cohorts. This has the potential to transform the clinical care of patients with reproductive disorders as LH pulsatility could be measured easily in clinical practice to aid correct diagnosis. Unless otherwise noted, all abstracts presented at ENDO are embargoed until the date and time of presentation. For oral presentations, the abstracts are embargoed until the session begins. Abstracts presented at a news conference are embargoed until the date and time of the news conference. The Endocrine Society reserves the right to lift the embargo on specific abstracts that are selected for promotion prior to or during ENDO.