Design, synthesis, formulation, and bioevaluation of ZZP-2, a FLT3-ITD inhibitor for the treatment of acute myeloid leukemia

FMS-like tyrosine receptor kinase 3 (FLT3) mutations have been recognized as ideal drug discovery targets for the treatment of acute myeloid leukemia (AML). Starting from the reported inhibitor 13v, we rationally designed and synthesized ZZP-2, a pyridine-pyridazine hybrid that displayed single-digit nanomolar IC₅₀ values against FLT3-ITD-positive AML cell lines (MOLM-13 and MV4-11). ZZP-2 suppressed FLT3 autophosphorylation and downstream STAT5, ERK, and AKT signaling in a concentration-dependent manner, exerting antiproliferative effects through multiple mechanisms, including apoptosis induction and cell cycle arrest. However, ZZP-2's aqueous solubility is < 0.5 μg/mL, a very low value which may affect the rate and extent of drug absorption from suspension formulations throughout the gastrointestinal tract. To overcome this limitation, we developed an optimized nano-self-emulsifying drug-delivery system (SEDDS) that reproducibly formed fine droplets (23.77 ± 0.20 nm) upon dilution and achieved > 95 % drug loading efficiency. After oral administration, the SEDDS formulation increased the ZZP-2 plasma area under the curve (AUC_0-∞) by 3.7-fold relative to a suspension formulation in Sprague-Dawley (SD) rats and significantly prolonged survival in MOLM-13-luciferase-bearing NSG mice compared to positive controls sunitinib and gilteritinib, without noticeable toxicity. Our study presents a novel FLT3-ITD inhibitor with high potency and in vivo stability.