Detecting GHRH Analogue Tesamorelin and Its Metabolites in Post‐Administration Human Urine and Serum Using LC‐HRMS/MS

ABSTRACT Tesamorelin is a growth hormone‐releasing hormone (GHRH) analogue used for reducing excess abdominal fat in HIV‐infected patients suffering from lipodystrophy. Together with other GHRH analogues, it is classified as a doping agent and is prohibited at all times according to the World Anti‐Doping Agency (WADA). To investigate whether current testing methodologies are adequate for detecting tesamorelin abuse, an administration study was performed involving six healthy volunteers and 10 consecutive days of Egrifta (tesamorelin) injections, mimicking a doping protocol. A validated solid‐phase extraction method was used to analyse the samples, reaching limits of detection of 13 pg/mL in urine and 41 pg/mL in serum. To account for the generation of metabolites by proteolytic degradation, tesamorelin was incubated in fresh human serum. Five previously unreported metabolites were found and identified, extending the detectability of tesamorelin abuse. Oxidation of the methionine residue was observed in tesamorelin and metabolites, hence ions to monitor the sulfoxide and sulfone forms were added to the methods. Despite a limit of detection 70 times lower than the WADA minimum required performance level and the inclusion of several metabolites, tesamorelin use could not be detected in post‐administration urine. In serum, tesamorelin metabolites were detected in all six volunteers for up to 2 h after injection, whereas intact tesamorelin was detected in just two volunteers, up to 30 min post‐administration. The outcome of this study shows that tesamorelin has a short detection window and testing methods should focus on metabolites in serum, rather than intact compounds in urine.