Comparative Analysis of Septin Modifiers, Forchlorfenuron and UR214-9, on Mitochondrial Fragmentation and Lytic Cell Death

Septins are conserved GTP‐binding proteins that play key roles in cell division, mitochondrial dynamics and immune responses. Despite their importance to human health, pharmacological compounds to modify septins remain limited. Forchlorfenuron (FCF) was the first small molecule identified to modify septins, disrupting their organisation and promoting mitochondrial fragmentation. A more potent FCF analog (UR214‐9) has recently been developed, but its effects on mitochondria were unknown. Here, we compare FCF and UR214‐9 in vitro using macrophages and in vivo using zebrafish larvae. We demonstrate that both modifiers induce mitochondrial fragmentation in macrophages without altering mitochondrial mass or SEPT7 expression. Consistent with mitochondrial fragmentation, both modifiers trigger lytic cell death in a dose‐dependent manner following lipopolysaccharide (LPS) priming. In vivo, both modifiers exhibit dose‐dependent effects on the survival of zebrafish larvae, although UR214‐9 was significantly more toxic. In agreement with in vitro results, we observed that FCF induces macrophage cell death and caspase‐1 activity in zebrafish larvae. Together, our findings show that both septin modifiers impact mitochondrial integrity and macrophage survival. Understanding how septin modifiers regulate immune responses may have important implications for inflammatory disease research and could lead to the development of septin‐based medicines for conditions characterised by dysregulated inflammation.