Leukotriene A4 hydrolase (LTA4H) is a zinc metalloenzyme with an epoxide hydrolase (EH) activity that converts leukotriene A4 (LTA4) to the neutrophil chemoattractant leukotriene B4 (LTB4) [1]. LTB4 has been implicated in a variety of inflammatory diseases such as atopic dermatitis [2], irritable bowel syndrome (IBS) [3], and chronic obstructive pulmonary disease (COPD) [4]. Thus, inhibition of LTB4 production or LTB4 antagonism are being investigated for treatment of such diseases [5,6]. LTA4H possesses a second aminopeptidase (AP) activity that cleaves the tripeptide Pro-Gly-Pro (PGP) [1]. The work of Blalock and coworkers showed PGP can serve as a biomarker for inflammatory diseases such as COPD [7]. Although PGP's relevance towards LTA4H-mediated inflammatory response has recently been called into question [8], there is a body of literature that supports its cleavage by LTA4H as a means of resolving inflammation [9]. Activation of the LTA4H AP activity by 4MDM was previously shown to resolve chronic inflammation in the lungs of murine models for chronic obstructive pulmonary disease (COPD) and acute lung injury [[10], [11], [12]]. ARM1 (1) was later reported as a selective EH inhibitor of LTA4H that spared AP activity [13]. We showed that 1 and ARM1 derivatives activate the AP activity using the alanine-p-nitroanilide (Ala-pNA) reporter group, albeit through a different kinetic mechanism than 4MDM [14]. As an extension of our previous work [14], we hypothesized a potent activator of Ala-pNA hydrolysis by LTA4H would have the ability to resolve inflammation in a Klebsiella pneumoniae (KP) animal model. Therefore, we synthesized a new series of ARM1 analogues with structural features that were expected to improve AP activating potency and metabolic stability (Fig. 1).

The diphenylmethane scaffold of 1 is susceptible to oxidation at the methylene bridge by cytochrome P450 enzymes [15]. In addition, the aminothiazole moiety of 1 is also susceptible to oxidation [16]. Thus, analogues 2–9 replace the methylene bridge of 1 with an oxygen atom, creating a diaryl ether scaffold. In addition, a p-chloro substituent was added to each analogue (except for 3) as a bioisostere for a p-methyl substituent, which was shown to provide potent AP activation in our previous work [14]. Analogue 2 assesses the effect of these structural changes on 1 while maintaining the metabolically susceptible aminothiazole ring. Analogue 3 tests the effect of a bigger p-bromo substituent on AP activation.

As evidenced by the crystal structure of ARM1 derivative 4-OMe-ARM1 (PDB ID: 6O5H), the aminothiazole group provides rigid hydrogen bonding interactions to the LTA4H protein backbone [14]. Analogues 4–9 explore whether alternative heterocycles can enhance this binding through potentially stronger and/or more numerous hydrogen bonding interactions. In addition, replacing the aminothiazole ring with an alternative heterocycle will help determine whether there is potential for improving the metabolic stability of these ARM1 derivatives. Analogue 4 is similar to 2 except it bears a hydrogen bond donating NH group in the heterocycle, potentially increasing the number of hydrogen bonding interactions. Analogue 5 is an isomer of 4 and would assess the effect of the location of hydrogen bonding groups. Analogue 6 is identical to 2 except the sulfur atom in 2 is replaced with a stronger hydrogen bond-accepting oxygen atom. Analogue 7 was inspired by one of the analogues reported by the Novartis group [8]. This analogue would test the importance of the amino group on the heterocycle for AP activation. Analogue 8 features a larger aminopyridine heterocycle. Finally, analogue 9 is a saturated heterocycle that structurally mimics the aforementioned analogues. Saturated heterocycles present attractive features over aromatic heterocycles such as higher water solubility and greater metabolic stability [17].