## 3 Overview of the Inhibitors Targeting the OxPhos Pathway
### Inhibitors of the Cyt-bcc-aa3 Complex
Through additional cross-resistance studies, QcrB was implicated as the main target [130]. The mutations identified in QcrB were A179P, M342T, W312C, and W312G, residues of the Qp binding site. Interestingly, while PAB compounds are bacteriostatic against actively growing _M. tb_, as with other QcrB inhibitors such as Q203 and LPZS, they are bactericidal against _M. tb_ grown under conditions of nutrient starvation, in contrast to the lack of activity of other QcrB inhibitors in non-replicating models [129]. This discrepancy may be due to a potential involvement of a secondary target of PAB, for instance, rv1339, the second gene in which mutations were identified from resistant mutants. The screening of a library of small polar molecules in a distinctive chemical space conducted at the Novartis Institute for Tropical Diseases led to the identification of hit compound pyrrol[3,4-c]pyridine-1,3(2H)-dione, which was further optimised to increase its stability by replacing the ester moiety with a methyl oxadiazole biosostere [131]. Lead compound 5h has an MIC\({}_{90}\) below 0.156 uM in replicating _M. tb_, and is non-cytotoxic. The high clearance and poor plasma exposure of 5h in mouse PK studies could be improved by co-dosing with a pan-CYP inhibitor, such that a compound concentration above that of the MIC could be achieved in the blood for 6 h at a dose of 20 mg/kg. While attempts at isolating resistant mutants to this novel chemical class were unsuccessful, _M. tb_ with a Cyt-bd oxidase deletion was hypersusceptible to the analogues. Moreover, the presence of mutation A317T in QcrB in this strain rendered _M. tb_ resistant to the compounds, thus implicating QcrB in the mechanism of action of this class. Further studies on the pyrrol[3,4-c]pyridine-1,3(2H)-diones will be aimed at improving their plasma concentration in vivo. In 2013, GlaxoSmithKline published the results of a large phenotypic screening campaign against _M. bovis_ BCG and _M. tb_, in which 177 hits belonging to several structurally distinct groups were identified [125]. To date, these chemical starting points have led to the discovery of two new classes of QcrB inhibitors, the 2-quinolin-4-yloxyacetamides (QOA) and the arylvinylpiperazine amides. SeveralSAR studies of the QOAs have been reported and are in agreement regarding the potency of this chemical entity against _M. tb_, the lack of toxicity in a zebrafish model and the narrow-spectrum activity specific for _M. tb_[132; 154; 155]. Independent work through cross-resistance studies and isolation of resistance mutants harbouring the mutation T313A in QcrB confirmed cytochrome bcc as the target of QOAs [155; 156]. The solubility and stability of this series, as well as their in vivo potency, remain to be addressed [133; 154]. The arylvinylpiperazine amides were generated through lead optimisation and SAR studies conducted with GW861072X from the GSK screen, which was an attractive starting point for its structural simplicity and potent activity against _M. bovis_ and _M. tb_[76]. The most potent analogues have improved MICs compared to the parent scaffold, displaying low cytotoxicity and activity against intracellular _M. tb_. The lead compound, AX-35 and two other analogues, AX-37 and AX-39, were also active in an acute TB infection model in mice. While isolation of _M. tb_-resistant mutants on agar proved unsuccessful, continual and increasing exposure of _M. tb_ to AX-35 over several passages in liquid culture resulted in the selection of resistant clones harbouring the mutations S182P, M342V and M342I in QcrB. Cross-resistance studies provided insight on residues which were important for the binding of the compound to QcrB, indicating a slightly different interaction with the quinol binding pocket than Q203 and LPZS. Improving the metabolic stability of this series would likely improve the in vivo activity in mice, although these compounds appear to be more stable in human microsomes.