## 3 Overview of the Inhibitors Targeting the OxPhos Pathway
### Menaguinone (MK) Biosynthesis
#### 3.2.3 MenA  
Several inhibitors have been identified to target MenA. Aurachin RE is a natural product from _Rhodococcus erythropolis_ JCM6824, and harbours potent activity against a wide range of Gram-positive and Gram-negative bacteria, including _Corynebacterium glutamicum_[121]. Using the aurachin RE scaffold, [PERSON] et al. synthesised a series of more than 400 derivatives that were assessed for their ability to specifically inhibit the \(M\). _tb_ MenA enzyme [49]. Compounds with IC\({}_{50}\) lower than 20 uM against \(M\). _tb_ and \(M\). _smeg_, and without activity against _Staphylococcus aureus_ MenA enzyme, were identified [49]. The lead compound of this series, (R)-13, had a MIC of 2.3 ug/mL against \(M\). _tb_. The compound is also bactericidal against hypoxic, non-replicating mycobacteria, suggesting a high MK turnover rate during persistence.  
Ro 48-8071, a (4-bromophenyl)[2-fluoro-4-[[6-(methyl-2-propenylamino)hexyl]oxy]phenyl]-methanone developed by Hoffman La Roche Inc is a known inhibitor of oxidogalene cyclase, which is involved in cholesterol biosynthesis in mammalians. It is effective against \(M\). _tb_, \(M\). _bovis_ BCG and \(M\). _smeg_[46; 53]. The target of Ro 48-8071 was inferred from metabolic labelling experiment in \(M\). _bovis_ BCG [46]. The synthesis of a neutral, apolar liquid, which was biochemically identified as MK through the incorporation of radiolabelled isoprenoid precursors and methionine, was inhibited in the presence of Ro 48-8071 [46]. Moreover, when \(M\). _smeg_ was treated with Ro 48-8071, a reduction of 2.5-3.3-fold in the concentrations of DMK-9 and MK-9 was observed, suggesting that the compound inhibits the late steps of MK biosynthesis in mycobacteria. Ro 48-8071 decreased the oxygen consumption in \(M\). _smeg_ and \(M\). _tb_, a phenotype that was rescued by MK supplementation. To validate MenA as the direct target of Ro 48-8071, MenA activity was assessed in membranes prepared from a recombinant \(E\)._coli_ strain expressing the \(M\). _tb_ MenA [46]. An eight-fold decrease in MenA activity was observed upon treatment with Ro 48-8071, indicating that MenA is the target of this inhibitor. Structure-activity relationship (SAR) studies led to the derivative CSU-20 which has improved MIC against \(M\). _tb_ and was bactericidal against hypoxic, non-replicating mycobacteria [46]. [PERSON] et al. showed that Ro 48-8071 has dual activity against MenA by behaving as a non-competitive inhibitor of the substrate DHNA, but also as a competitive inhibitor of isoprenyl diphosphate [122].  
The bicyclic long-chain fatty acids 7-methoxy-2-naphthol-based inhibitors also target MenA [117; 123; 124]. [PERSON] et al. tested four compounds of this series (NM-1 to NM-4) for their ability to inhibit MenA and \(M\). _tb_ growth. The most active compound, NM-4, had an MIC\({}_{90}\) of 4.5 uM and was bactericidal against replicating and non-replicating \(M\). _tb_[117]. NM-4 was synergistic with BDQ, CFZ and imidazopyridine ND-10885 in vitro against \(M\). _tb_, with culture sterilisation observed after 21 days of incubation. This new lead is non-cytotoxic, specific for Gram-positive bacteria, and amenable to chemical optimisation, properties that could be exploited to optimise the chemical series [124]. Further characterisation work on the PK/PD properties and in vivo activity remains to be undertaken.