Towards clinically relevant dose ratios for Cabamiquine and Pyronaridine combination using P. falciparum field isolate data

Abstract

The selection and combination of dose regimens for antimalarials involve complex considerations including pharmacokinetic and pharmacodynamic interactions. In this study, we use immediate ex vivo P. falciparum field isolates to evaluate the effect of cabamiquine and pyronaridine as standalone treatments and in combination therapy. We feed the data into a pharmacometrics model to generate an interaction map and simulate meaningful clinical dose ratios. We demonstrate that the pharmacometrics model of parasite growth and killing provides a detailed description of parasite kinetics against cabamiquine-susceptible and resistant parasites. Pyronaridine monotherapy provides suboptimal killing rates at doses as high as 720 mg. In contrast, the combination of a single dose of 330 mg cabamiquine and 360 mg pyronaridine provides over 90% parasite killing in most of the simulated patients. The described methodology that combines a rapid, 3R-compliant in vitro method and modelling to set meaningful doses for new antimalarials could contribute to clinical drug development.

Fig. 1. A pharmacodynamic drug–drug interaction approach for identifying relevant clinical trial combination doses using an in vitro viability assay with ex vivo P. falciparum field isolates (IEV).

A Patient P. falciparum parasites were collected and cultured ex vivo to determine (B) in vitro parasite-killing kinetics of an antimalarial drugs at different concentrations either monotherapy or combinations at different timepoints using a rapid viability assay (MitoTracker™ Deep Red FM staining). From there (C), an in silico drug–drug-PD interaction map modeled and coupled with (D) individual antimalarial PK simulation obtained from previous human clinical trial to simulate (E) individual parasite-killing rate profiles per dose in monotherapy and in combination.

Fig. 2. Concentration-time-dependent EC₅₀ of cabamiquine in IEV P. falciparum wild-type field isolates simulated at different concentrations over 72 h.

The orange dashed line represents the static EC₅₀ of cabamiquine.

Fig. 3. Individual model fits for cabamiquine–pyronaridine combination for the in vitro dug–drug–PD interaction assay data using IEV P. falciparum wild-type field isolates.

Model predictions from the final model (lines) and original data (points) for each parasite (colors); each facet displays a studied scenario for cabamiquine (A1) and pyronaridine (A2). The numbers in each facet present the concentration of either cabamiquine or pyronaridine in nM in each well. Readout: total SYBR Green and MitoTracker positive parasite count.

Fig. 4. Heat map of the simulated, apparent killing rate for cabamiquine and pyronaridine at 6, 12, 48, and 72 h for the typical IEV P. falciparum wild-type field isolates using the population parameters of the mixed effects model illustrating the time-dependent killing for cabamiquine and stable killing for pyronaridine.

The color gradient depicts the killing rate (green faster; pink slower), while the isoboles visualize lines where the killing rate is identical. The red isobole refers the EC₅₀.

Fig. 5. Clinical trial simulations and parasite-killing rates in IEV wild-type parasites.

Distribution of the apparent killing rate against susceptible field isolates of P. falciparum at 24 h (upper panel) and 96 h (lower panel) from clinical trial simulations using a single dose of cabamiquine (free base) and pyronaridine (tetraphosphate); percentage in label: virtual patients exceeding a killing rate of 90% (dashed line); box: 25th to 75th percentile (interquartile range) with median (black horizontal line); whiskers: 1.5 times interquartile range below 25th or above 75th percentile, respectively; dots: values outside whiskers’ range; n = 1000 simulations for each dosing scenario.

Fig. 6. Clinical trial simulations and parasite-killing rates in cabamiquine-resistant parasites.

Distribution of the apparent killing rate against a cabamiquine-resistant P. falciparum strain at 24 h (upper panel) and 96 h (lower panel) from clinical trial simulations using a single dose of cabamiquine and pyronaridine; percentage in label: virtual patients exceeding a killing rate of 90% (dashed line); box: 25th to 75th percentile (interquartile range) with median (black horizontal line); whiskers: 1.5 times interquartile range below 25th or above 75th percentile, respectively; dots: values outside whiskers’ range; n = 1000 simulations for each dosing scenario.

Fig. 7. Clinical trial heat map of parasite-killing rates against P. falciparum (wild-type) at 96 hours following a single dose of cabamiquine and pyronaridine.

The map shows the percentage of virtual patients exceeding a 90% killing rate.