An investigation into the effects of allosteric modulators of the human 5-Ht3_3A_A receptor

The 5-Ht$_3$$_A$ receptor is a cys-loop ligand gated ion channel re-emerging as an attractive target in irritable bowel syndrome (IBS), which currently affects 5-10% of the global population. At present, IBS therapies involve the complete blockade of the 5-Ht$_3$$_A$ receptor by orthosteric antagonists, which leads to complications such as severe constipation and ischaemic colitis. Allosteric modulation could bypass such side effects, as receptor function relies upon the endogenous neurotransmitter to retain physiological control. Here, we investigate the structure and function of the 5-Ht$_3$$_A$ receptor allosteric binding site by the identification of novel allosteric compounds and the generation of an α7/5-HT$_3$$_A$ chimeric receptor. Intracellular calcium assays and competitive radioligand binding experiments indicated that the halogenated indole derivatives, 5-chloroindole (5-Cl) (Newman et al., 2013) and 5- (trifluoromethyl)indole (5-TFMI) are positive allosteric modulators (PAM) of the 5-Ht$_3$$_A$ receptor; however 5-TFMI also displayed a degree of orthosteric binding. The structural analogues, 5-bromoindazole (5-BI) and 5-bromo-benzimidazole (5-BBI) exhibited contrasting effects, by potentiating and decreasing 5-HT-evoked responses, respectively. Further studies suggested some orthosteric binding by 5-BI and 5-BBI at high concentrations. The allosteric binding site for 5-Cl was previously located in the N-terminus of the mouse 5-Ht$_3$$_A$ receptor. To identify the site in the human receptor, we attempted to construct a human chimeric α7V$_2$$_0$$_1$5-HT$_3$$_A$ receptor to allow further investigation into this question. Our data suggest these halogenated indoles are allosteric compounds and, when studied in combination with the α7V$_2$$_0$$_1$5-HT$_3$$_A$ chimera, could identify the required core structure of high affinity compounds needed for negative allosteric modulation in the treatment of IBS

Lactam Truncation Yields a Dihydroquinazolinone Scaffold with Potent Antimalarial Activity that Targets PfATP4

The emergence of resistance against current antimalarial treatments has necessitated the need for the development of novel antimalarial chemotypes. Toward this goal, we recently optimised the antimalarial activity of the dihydroquinazolinone scaffold and showed it targeted PfATP4. Here, we deconstruct the lactam moiety of the tricyclic dihydroquinazolinone scaffold and investigate the structure-activity relationship of the truncated scaffold. It was shown that SAR between scaffolds was largely transferrable and generated analogues with potent asexual stage activity. Evaluation of the truncated analogues against PfATP4 mutant drug resistant parasite strains and in assays measuring PfATP4-associated ATPase activity demonstrated retention of PfATP4 as the molecular target. Analogues exhibited activity against both male and female gametes and multidrug resistant parasites. Limited efficacy of analogues in a P. berghei asexual stage mouse model was attributed to their moderate metabolic stability and low aqueous stability. Further development is required to address these attributes toward the potential use of the dihydroquinazolinone class in a curative and transmission blocking combination antimalarial therapy

7-N-Substituted-3-oxadiazole Quinolones with Potent Antimalarial Activity Target the Cytochrome bc1 Complex.

The development of new antimalarials is required because of the threat of resistance to current antimalarial therapies. To discover new antimalarial chemotypes, we screened the Janssen Jumpstarter library against the P. falciparum asexual parasite and identified the 7-N-substituted-3-oxadiazole quinolone hit class. We established the structure-activity relationship and optimized the antimalarial potency. The optimized analog WJM228 (17) showed robust metabolic stability in vitro, although the aqueous solubility was limited. Forward genetic resistance studies uncovered that WJM228 targets the Qo site of cytochrome b (cyt b), an important component of the mitochondrial electron transport chain (ETC) that is essential for pyrimidine biosynthesis and an established antimalarial target. Profiling against drug-resistant parasites confirmed that WJM228 confers resistance to the Qo site but not Qi site mutations, and in a biosensor assay, it was shown to impact the ETC via inhibition of cyt b. Consistent with other cyt b targeted antimalarials, WJM228 prevented pre-erythrocytic parasite and male gamete development and reduced asexual parasitemia in a P. berghei mouse model of malaria. Correcting the limited aqueous solubility and the high susceptibility to cyt b Qo site resistant parasites found in the clinic will be major obstacles in the future development of the 3-oxadiazole quinolone antimalarial class