Off-Target Activation of CatSper Limits the Utility of a Novel Small Molecule Slo3 Inhibitor and its Derivatives in Human Sperm

Abstract

The sperm-specific K+ channel Slo3 is required for sperm function and fertilization in mice and humans. Due to the lack of selective pharmacological tools, its molecular and cellular physiology in human sperm has remained only ill-defined, though. The compound VU0546110 (1) was recently introduced as the first selective Slo3 inhibitor and is now being used to study the channel’s role in human sperm. Whether the compound may have off-target actions in sperm is however unknown. Here, using kinetic Ca2+ fluorimetry, patch-clamp recordings, and sperm from infertile men with a homozygous deletion of the CATSPER2 gene as experimental model, we show that VU0546110 simultaneously inhibits Slo3 and activates the sperm-specific Ca2+ channel CatSper. This demonstrates that the utility of the drug is limited: it is difficult to decipher if its action on human sperm is due to inhibition of Slo3, activation of CatSper, or a combination of both. We also introduce derivatives of VU0546110 that show a reduced potency and efficacy for CatSper activation, while preserving the inhibitory action on Slo3.