Monomerized Ectatomin Subunits ET1A and ET1B Combine Antibacterial, Anti-NET and Antinociceptive Activity Without Cytolysis

Venom pore-formers are a rich source of membrane-active chemistry, but cytolysis has kept most out of development. Ectatomin, from the ant Ectatomma tuberculatum, is a disulfide-linked heterodimer whose lysis discouraged therapeutic study for three decades.To test whether the isolated monomers ET1A (37 residues) and ET1B (34) retain activity once the lytic dimer is dismantled, across antibacterial, immunomodulatory and antinociceptive axes. Assays followed previously described protocols. MIC and MBC by broth microdilution against nine strains, three multidrug-resistant; membrane permeabilization and biofilm inhibition; NET release induced by ionomycin and by LPS; PBMC viability; chick-embryo toxicity; and a Drosophila larval chemical-nociception assay with topical delivery across a graded cuticular barrier. Docking, MD and ADMET and protein-protein interactions formed an exploratory in silico layer. Both were bactericidal across the panel (MBC/MIC 1-2), ET1A most potent against Gram-positives (Bacillus subtilis, MIC 8 µM) and ET1B against Gram-negatives; both remained active against vancomycin-resistant Enterococcus faecalis and Acinetobacter baumannii, where amikacin was inactive (MIC > 512 µM). Killing involved membrane permeabilization and leakage, and both inhibited biofilm formation. LPS-induced NET release fell concentration-dependently (3-30 µM), whereas ionomycin-driven NETosis was only partly suppressed. PBMC viability stayed above 98% (CC₅₀ > 100 µM), and neither was embryolethal or teratogenic. In Drosophila larvae both prolonged nocifensive rolling latency over vehicle and inert-protein controls, the delay growing with concentration, exposure time and barrier severity; ET1B was more potent than ET1A. Dismantling a cytolytic heterodimer yielded subunits that are bactericidal, host-directed anti-NET and antinociceptive in vivo, without cytotoxicity to host cells: a miniaturization route for venom-derived leads. Mechanistic assignments from the in silico layer remain predictions.