Iminoguanidines as Allosteric Inhibitors of the Iron-Regulated Heme Oxygenase (HemO) of <i>Pseudomonas aeruginosa</i>

New therapeutic targets
are required to combat multidrug resistant
infections, such as the iron-regulated heme oxygenase (HemO) of Pseudomonas aeruginosa, due to links between iron and virulence
and dependence on heme as an iron source during infection. Herein
we report the synthesis and activity of a series of iminoguanidine-based
inhibitors of HemO. Compound 23 showed a binding affinity
of 5.7 μM and an MIC50 of 52.3 μg/mL against P. aeruginosa PAO1. An in cellulo activity assay was developed
by coupling HemO activity to a biliverdin-IXα-dependent infrared
fluorescent protein, in which compound 23 showed an EC50 of 11.3 μM. The compounds showed increased activity
against clinical isolates of P. aeruginosa, further
confirming the target pathway. This class of inhibitors acts by binding
to an allosteric site; the novel binding site is proposed in silico
and supported by saturation transfer difference (STD) NMR as well
as by hydrogen exchange mass spectrometry (HXMS).

鉴于铁与病原菌毒力存在相关性，且感染过程中病原菌需以血红素作为铁源，因此亟需开发新型治疗靶点以对抗多重耐药感染，例如铜绿假单胞菌（Pseudomonas aeruginosa）中铁调控型血红素氧合酶（HemO）。本文报道了一系列基于亚氨基胍的HemO抑制剂的合成与活性表征。化合物23对铜绿假单胞菌PAO1的结合亲和力为5.7 μM，半最大抑菌浓度（MIC₅₀）为52.3 μg/mL。本研究通过将HemO活性与胆绿素-IXα依赖型红外荧光蛋白偶联，建立了细胞内活性测定方法，化合物23在该体系中的半最大效应浓度（EC₅₀）为11.3 μM。该系列化合物对铜绿假单胞菌临床分离株的活性显著提升，进一步验证了其作用靶点通路。该类抑制剂通过结合别构位点发挥作用：其全新结合位点经计算机模拟（in silico）提出，并通过饱和转移差谱核磁共振（STD NMR）与氢交换质谱（HXMS）得到证实。