Compounds enhancing human sperm motility identified using a high-throughput phenotypic screening platform

Abstract

Study question: Can a high-throughput screening platform facilitate male fertility drug discovery? Summary answer: A high-throughput screening platform identified a large number of compounds that enhanced sperm motility. What is known already: Several efforts to find small molecules modulating sperm function have been performed but not using high-throughput technology. Study design, size, duration: Healthy donor semen samples were used and samples were pooled (3-5 donors per pool). Primary screening was performed in singlicate; dose-response screening was performed in duplicate (independent donor pools). Participants/materials, setting, methods: Spermatozoa isolated from healthy donors were prepared by density gradient centrifugation and incubated in 384-well plates with compounds (6.25 uM) to identify those compounds with enhancing effects on motility. A total of ~17,000 compounds from the following libraries: ReFRAME, Prestwick, Tocris, LOPAC, CLOUD and MMV Pathogen Box were screened. Dose response experiments of screening hits were performed to confirm the enhancing effect on sperm motility. Experiments were performed in a University setting. Main results and the role of chance: From our primary single concentration screening, 105 compounds elicited an enhancing effect on sperm motility compared to DMSO treated wells. Confirmed enhancing compounds were grouped based on their annotated targets/target classes. A major target class, phosphodiesterase inhibitors, were identified in particular PDE10A inhibitors as well as number of compounds not previously identified/known to enhance human sperm motility such as those related to GABA signaling. Limitations, reasons for caution: Compounds have been tested with prepared donor spermatozoa and only incubated for a short period of time. Therefore, the effect of compounds on whole semen or with longer incubation time may be different. All experiments were performed in vitro. Wider implications of the findings: This phenotypic screening assay identified a large number of compounds that increased sperm motility. In addition to furthering our understanding of human sperm function, for example identifying new avenues for discovery, we highlight potential inhibitors as promising start-point for a medicinal chemistry programme for potential enhancement of male infertility. Moreover, with disclosure of the results of screening we present a substantial resource to inform further work in the field.