Two experimental antimalarial compounds, TM174 and TM240, are stuck in limbo because they appear to fail a lab test—but the failure may be a mirage caused by the test itself. These compounds belong to the 4-aminoquinoline drug class, a cornerstone of malaria treatment that has been undermined by drug-resistant parasites. TM174 and TM240 are potent against resistant strains, have long half-lives in the body, and show no heart safety issues—a major improvement over older drugs in the class. But in one difficult-to-culture parasite strain, the compounds show a shallow kill curve with about 10% of parasites appearing to survive at high drug concentrations. This "Eagle effect" has halted the compounds' path to becoming official drug candidates. The researchers suspect the effect is an artifact. The parasite strain grows slowly, and standard 72-hour assays may simply not give the drug enough time to finish killing. They propose to settle the question by testing the compounds in mice with human malaria infections—a more realistic model than a dish. If the Eagle effect disappears in mice, the compounds can proceed toward human trials. If not, the team will need to understand the mechanism behind the phenomenon.
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In partnership with MMV, we were challenged to deliver a new generation of 4-aminoquinoline (4-AQs), one of the most strategically important drug classes in MMV’s antimalarial portfolio, engineered to address current limitations of susceptibility to PfCRT-mediated resistance mechanisms and limiting safety profiles based on unacceptable hERG margins characteristic of the class. Such compounds are considered to have the potential to address the global need for new chemoprophylactic combinations with slow elimination characteristics, the highest TCP priority within MMV. Lead compounds TM174 (MMV1919471) and TM240 (MMV1960679) exhibit selective low-nanomolar potency across diverse P. falciparum strains, including chloroquine- and piperaquine-resistant (CQ-R, PPQ-R) clinical isolates Dd2 and RF12, without loss of efficacy. Both compounds demonstrate high oral bioavailability and exceptional metabolic stability with long predicted human half-lives. Importantly, they show no cardiac safety liabilities, yielding therapeutic indices >600- to 1500-fold above their antiplasmodial activity, an especially notable feature given that traditional 4-AQs typically have minimal hERG safety margins. MMV’s SOLA model predicts that TM174 and TM240 could deliver TCP-1 efficacy criteria in humans with a single 250 mg dose. Despite this compelling profile, progression toward candidate nomination is currently paused due to a reproducible in vitro “Eagle effect” observed in one strain, RF12, seen at high drug concentrations. This manifests as a shallow kill curve with an apparent ~10% residual parasite “survival”. Crucially, this phenotype is non-genetic: surviving parasites re-exposed to TM174/TM240 show no shift in IC50, indicating a reversible, phenotypic basis rather than an acquired resistance. RF12 is a challenging parasite to culture and has a parasite multiplication rate (PMR) ~33% lower than wild-type strains. We hypothesise this reduced PMR reflects impaired digestive vacuole (DV) function due to the H97Y PfCRT mutation on the K13 C580Y background. This may prolong the intra-erythrocytic life cycle, and because standard in vitro assays measure growth over a fixed window (48–72 h), such delay could produce premature “survival” readouts. Thus, the Eagle effect may be partly explained by a temporal artefact of the assay. In addition, we further hypothesise that RF12, due to impaired DV H? transport, is particularly vulnerable to DV pH disruption by weak bases like TM174 and TM240. Prior studies show that high concentrations of weak bases can alkalinise the DV, an effect that is more pronounced in CQ-R parasites. This enhanced susceptibility to DV alkalisation in RF12 may contribute to the Eagle Effect observed in vitro at elevated drug levels. Uncertainty as to the clinical relevance of these in vitro phenomena has triggered a pause in candidate selection until a credible explanation can be supported with experimental data confirming that the effect is indeed an in vitro artefact with limited if any relevance under physiologically relevant in vivo conditions of human malaria infection. To address these unresolved questions, we propose establishing an RF12-infected SCID mouse model (currently SCID is validated only for 3D7 CQ-sensitive isolate) to evaluate TM174/TM240 efficacy under physiologically-relevant conditions. This model will provide the definitive go/no-go data required for re-initiation of candidate nomination. Complementary in vitro studies will also be undertaken to further define the mechanisms unrepining this in vitro phenomenon. We seek MRC GAP support to resolve this final, decision-critical uncertainty and enable progression of a leading antimalarial candidate. MMV and their ESAC confirm that this is the only barrier to candidate selection at this time.
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