Screening and hit evaluation of a microbial metabolite library against the pathogenic Plasmodium falciparum and Toxoplasma gondii parasites

Abstract

Frontline drug treatments for malaria are at risk of failing due to emerging resistance, meanwhile drugs used to treat toxoplasmosis have suboptimal efficacy and safety. As demonstrated by the success of clinically used antiparasitic drugs, the diverse structural complexity and biological activity of natural products holds great potential for drug discovery and development, to address the need for new compounds with novel mechanisms. Here we screened the BioAustralis Discovery Plates Series I library, a collection of 812 microbial natural product compounds including rare microbial metabolites, against Plasmodium falciparum erythrocytic stage and Toxoplasma gondii tachyzoite parasites. We identified 219 compounds that inhibited P. falciparum growth by at least 80 % at a concentration of 2 μg/mL (1-10 μM for >90 % of compounds), whilst 149 compounds demonstrated equivalent activity against T. gondii. The active compounds were assigned based on chemical structure to more than 50 compound classes. After triaging active compounds for those with low mammalian cytotoxicity, we defined the in vitro half maximal inhibitory concentration (IC₅₀) of a selection of compounds against the parasites, identifying four compound groups with activity in the low nanomolar range. The macrocyclic lactone pladienolide B and cryptopleurine were found to be very potent against the parasites but also mammalian cells, warranting further structure-activity relationship investigation. Two groups, the monocyclic thiazole peptides, including micrococcin P1 and the thiocillins, and the pleuromutilins, exhibited both low antiparasitic IC₅₀ and low cytotoxicity, highlighting their potential for further analysis. This study defines the activity of the BioAustralis Discovery Plates Series I against two apicomplexan parasites of significant global importance, providing potential new tools to study parasite biology and possible starting points for novel antiparasitic development.

Keywords: Drug discovery; Malaria; Natural products; Toxoplasmosis.

Graphical abstract

Fig. 1

The antimalarial activity of the BioAustralis Discovery Plates Series I library of microbial metabolites. (a) Correlation between percentage survival of P. falciparum between biological duplicate screens of the microbial metabolite library. Each data point represents a single compound and dashed lines indicate 20 % survival of each replicate, used as the threshold for activity (i.e. active ≤20 % survival, inactive >20 % survival). Compound classes are assigned a colour if at least 2 compounds from the structural class demonstrate activity based on the average survival. Single active compounds from a class have been grouped as ‘others’, whilst non-active compounds (NAs) appear in grey. (b) Compound classes with at least 2 active compounds against P. falciparum, showing the proportion of active compounds in their classes and the total number of compounds in the respective classes within the library. Compound classes outlined in green showed the same activity in T. gondii. (c) UpSet plot depicting the overlap between antimalarial activity of microbial metabolites with anti-tumour (NS-1 cell line) and cytotoxic (fibroblast cells) activity. The filled circles on the x-axis specify the overlapping activities and the height of the bars indicate the number of overlapping active compounds. The horizontal bars represent the total number of active compounds in each screen.

Fig. 2

The anti-toxoplasma activity of the BioAustralis Discovery Plates Series I library of microbial metabolites. (a) Correlation between percentage survival of T. gondii between biological duplicate screens of the microbial metabolite library. Each data point represents a single compound and dashed lines indicate 20 % survival of each replicate, used as the threshold for activity (i.e. active ≤20 % survival, inactive >20 % survival). Compound classes are assigned a colour if at least 2 compounds from the structural class demonstrate activity based on the average survival. Single active compounds from a class have been grouped as ‘others’, whilst non-active compounds (NAs) appear in grey. (b) Compound classes with at least 2 active compounds against T. gondii, showing the proportion of active compounds in their classes and the total number of compounds in the respective classes within the library. Compound classes outlined in green showed the same activity in P. falciparum. (c) UpSet plot depicting the overlap between anti-toxoplasma activity of microbial metabolites with anti-tumour (NS-1 cell line) and cytotoxic (fibroblast cells) activity. The filled circles on the x-axis specify the overlapping activities and the height of the bars indicate the number of overlapping active compounds. The horizontal bars represent the total number of active compounds in each screen.

Fig. 3

Active compounds against P. falciparum and T. gondii, and their anti-tumour and cytotoxic activity. Venn diagram depicting the overlap between antimalarial (P. falciparum) and anti-toxoplasma (T. gondii) activity of the active microbial metabolites with anti-tumour (NS-1 cell line) and cytotoxic (foreskin fibroblast cells) activity. The total number of active compounds against P. falciparum and T. gondii are indicated in parentheses.