Characterizing the quick-killing mechanism of action of azithromycin analogs against malaria parasites

Mao, EY; Nguyen, W; Jana, GP; Maity, BC; Pazicky, S; Giannangelo, C; Reader, J; Famodimu, MT; Birkholtz, LM; Delves, MJ; Creek, DJ; Bozdech, Z; Laleu, B; Burrows, JN; Sleebs, BE; Gancheva, MR; Wilson, DW

Author(s): Mao, EY; Nguyen, W; Jana, GP; Maity, BC; Pazicky, S; Giannangelo, C; Reader, J; Famodimu, MT; Birkholtz, LM; Delves, MJ; Creek, DJ; Bozdech, Z; Laleu, B; Burrows, JN; Sleebs, BE; Gancheva, MR; Wilson, DW;
Details: Publication Year 2025-09-03,Volume 69,Issue #9,Page e0178324
Journal Title: Antimicrobial Agents and Chemotherapy
Abstract: Drug resistance is steadily undermining the efficacy of frontline anti-malarials, highlighting the urgent need for novel therapies with alternative mechanisms of action. The chemical addition of different moieties to azithromycin yields compounds with improved quick-killing potency against malaria parasites, with the most active analogs typically containing a chloroquinoline group. Here, we investigated the quick-killing mechanism of five azithromycin analogs, two of which contain differentially oriented chloroquinoline moieties. The improvement in quick-killing activity over azithromycin for non-chloroquinoline analogs was around 10 -to 42-fold, with chloroquinoline-containing analogs showing a further 2- to 17-fold improvement over non-chloroquinoline compounds. Chemical inhibition of hemoglobin digestion and chloroquine's inhibitory effect against heme polymerization linked analogs with both chloroquinoline and non-chloroquinoline modifications to a chloroquine-like mechanism of action. However, none of the analogs showed a significant reduction in efficacy against chloroquine-resistant asexual blood-stage parasites. Multiple attempts at selecting for azithromycin analog-resistant parasites to elucidate the mechanism of quick-killing were unsuccessful. Application of cellular thermal shift proteomics revealed that azithromycin analogs significantly stabilized 34-155 different proteins in trophozoites, a high number that showed minimal overlap with chloroquine. Additionally, our most potent chloroquinoline-containing analog demonstrated a significant improvement in gametocytocidal activity over azithromycin and further maintained moderate inhibition of chloroquine-insensitive late-stage gametocytes. These findings support that this class of azithromycin analogs kills malaria parasites through a broad range of potential mechanisms, making them promising candidates for optimization as fast and broad-acting anti-malarials.
Publisher: ASM
Keywords: *Azithromycin/pharmacology/analogs & derivatives; *Antimalarials/pharmacology/chemistry; Chloroquine/pharmacology; *Plasmodium falciparum/drug effects; Drug Resistance; Humans; Parasitic Sensitivity Tests; Plasmodium; antimalarial agents; azithromycin; cellular thermal shift assay; malaria
Research Division(s): New Medicines and Diagnostics
PubMed ID: 40711456
Publisher's Version: https://doi.org/10.1128/aac.01783-24
Open Access at Publisher's Site: https://doi.org/10.1128/aac.01783-24
Terms of Use/Rights Notice: Refer to copyright notice on published article.

Creation Date: 2025-07-28 03:09:55

Last Modified: 2025-09-23 11:13:54