In vitro-in vivo discord: A preclinical study of AZD2716 and its racemate with comparison to varespladib for the development of snake venom sPLA2 inhibitors

We evaluated a family of repurposed sPLA2 inhibitors as novel candidate snakebite envenoming therapeutics. Stereospecific (<i>R</i>)-7 AZD2716 and its racemic mixture were compared to varespladib in an <i>in vitro</i> sPLA2 assay against a sample of 26 venoms from medically important snake species from five continents. All compounds demonstrated potent nano-to picomolar IC₅₀ values, comparable to the benchmark inhibitory profile of varespladib. Surprisingly, however, this <i>in vitro</i> efficacy did not translate to survival in an <i>in vivo</i> mouse model under GLP standard conditions at an independent third party laboratory. In animal rescue studies evaluating both oral and IV dosing against the same four high sPLA2 venoms, varespladib demonstrated more consistent survival duration versus the chirally separated AZD2716 enantiomer and racemate following single-dose intravenous or two-dose oral drug administration. Additionally, the stereospecific AZD2716 did not provide the same survival advantage as the racemic mixture and neither molecule resulted in the same survival advantage as varespladib <i>in vivo</i> (p < 0.05), despite similar <i>in vitro</i> potency. These findings highlight the importance of following <i>in vitro</i> inhibition assays with preclinical studies in drug candidate selection for lead compounds and advancement to clinical development.