Phosphate-based 2,3-DPG mimetics as Hemoglobin allosteric modulators with potent anti-hypoxic activity

High-altitude hypoxia causes acute mountain sickness due to insufficient tissue oxygen delivery. Herein we report the rational design, synthesis, and pharmacological evaluation of novel 2,3-diphosphoglycerate (2,3-DPG) mimetics based on phosphate ester and amino acid scaffolds, featuring PN and PS bonds. Bioisosteric replacement of labile PO bonds with PN or PS linkages and introduction of lipophilic ester groups was designed to improve metabolic stability and membrane permeability. Seven target compounds were synthesized and characterized. In normobaric hypoxia, ethyl ester derivatives 1e, 2e, 2f, 2h significantly prolonged survival, with compounds 2f and 2h showing the highest efficacy (38.97% prolongation, p < 0.01). In a hypobaric chamber simulating 7000 m, 2f and 2h increased Hb P50 by up to 35% (48.17 ± 1.25 mmHg for 2hM, p < 0.01), shifting the oxygen dissociation curve rightward and improving tissue oxygenation. In a high-altitude field study at 4010 m, 1e and 2e attenuated oxidative stress (GSH restored by 235%, p < 0.01), reduced serum IL-6 and TNF-α (up to 54% reduction), and alleviated histopathological damage. To our knowledge, compounds 2f and 2h represent the first phosphate-based Hb allosteric modulators that enhance tissue oxygenation under extreme hypoxia, offering a promising therapeutic strategy for high-altitude illnesses.