Metabolomics reveals synergistic antimalarial drug pairing effects against Plasmodium falciparum in vitro
preprint
OA: closed
CC-BY-NC-4.0
Abstract
Background The development of new drugs against afflictions that disproportionately impact poorly resourced areas around the globe is an expensive endeavor. As cost-effective alternatives, strategic combinations of approved drugs can be used to enhance the efficacy against Plasmodium falciparum . Understanding the metabolic consequences of such combinations is essential for optimizing treatment strategies and delaying drug resistance. Methods An integrated metabolomic and pharmacological analysis was performed on P. falciparum exposed to chloroquine (CQ), pyrimethamine (PY), sulfadoxine (SD), and their combinations (SDPY and SDCQ). Dose□response assays were used to quantify drug potency, whereas untargeted metabolomic profiling was used to assess pathway-level perturbations associated with individual and combined treatments. Results Dose □ response assays confirmed the nanomolar potency of PY (IC₅₀ = 12.5 nM) and CQ (IC₅₀ = 11 nM) compared with the micromolar efficacy of SD (IC₅₀ = 9.1 µM), which is consistent with its role as a synergistic antifolate partner. Metabolomic profiling revealed that PY strongly disrupted folate-dependent pyrimidine biosynthesis, leading to deoxyuridine and dUMP accumulation, whereas SD caused milder perturbations, which was consistent with DHPS inhibition. CQ produced modest metabolic effects alone but markedly amplified antifolate-induced stress when combined with PY. Drug combinations generated metabolic responses that are distinct from those resulting from individual treatments. Across antifolate combinations, consistent trends included reduced amino acid pools, suppression of thiamine and glutathione metabolism, and enhanced PPP inhibition, leading to broad disruption of nucleotide, redox, and carbon metabolism. Conclusions Pyrimidine suppression has emerged as the central hallmark of antifolate-based therapy in P. falciparum . The distinct synergistic signatures observed with drug combinations support their potential to enhance efficacy and delay resistance. These findings provide a mechanistic foundation for guiding antimalarial combination policies, optimizing therapeutic regimens, and strengthening rational drug-design strategies.
Full text
4,425 characters
· extracted from
oa-doi-fallback
· 4 sections
· click to expand
Abstract
Background The development of new drugs against afflictions that disproportionately impact poorly resourced areas around the globe is an expensive endeavor. As cost-effective alternatives, strategic combinations of approved drugs can be used to enhance the efficacy against Plasmodium falciparum. Understanding the metabolic consequences of such combinations is essential for optimizing treatment strategies and delaying drug resistance.
Methods
An integrated metabolomic and pharmacological analysis was performed on P. falciparum exposed to chloroquine (CQ), pyrimethamine (PY), sulfadoxine (SD), and their combinations (SDPY and SDCQ). Dose□response assays were used to quantify drug potency, whereas untargeted metabolomic profiling was used to assess pathway-level perturbations associated with individual and combined treatments.
Results
Dose□response assays confirmed the nanomolar potency of PY (IC₅₀ = 12.5 nM) and CQ (IC₅₀ = 11 nM) compared with the micromolar efficacy of SD (IC₅₀ = 9.1 µM), which is consistent with its role as a synergistic antifolate partner. Metabolomic profiling revealed that PY strongly disrupted folate-dependent pyrimidine biosynthesis, leading to deoxyuridine and dUMP accumulation, whereas SD caused milder perturbations, which was consistent with DHPS inhibition. CQ produced modest metabolic effects alone but markedly amplified antifolate-induced stress when combined with PY.
Drug combinations generated metabolic responses that are distinct from those resulting from individual treatments. Across antifolate combinations, consistent trends included reduced amino acid pools, suppression of thiamine and glutathione metabolism, and enhanced PPP inhibition, leading to broad disruption of nucleotide, redox, and carbon metabolism.
Conclusions
Pyrimidine suppression has emerged as the central hallmark of antifolate-based therapy in P. falciparum. The distinct synergistic signatures observed with drug combinations support their potential to enhance efficacy and delay resistance. These findings provide a mechanistic foundation for guiding antimalarial combination policies, optimizing therapeutic regimens, and strengthening rational drug-design strategies.
Competing Interest Statement
The authors have declared no competing interest.
Data availability
Data available in the NIH data repository, Metabolomics Workbench Study project doiPR002764, and Study ID ST004358, available at https://dev.metabolomicsworkbench.org:22222/data/DRCCMetadata.php?Mode=Study&StudyID=ST004358&Access=WubV3794
List of Abbreviations
- ACT
- Artemisinin-based Combination Therapy
- ADP
- Adenosine Diphosphate
- ATP
- Adenosine Triphosphate
- CMP
- Cytidine Monophosphate
- CDP
- Cytidine Diphosphate
- CQ
- Chloroquine
- dCDP
- Deoxycytidine Diphosphate
- dGTP
- Deoxyguanosine Triphosphate
- DHFR
- Dihydrofolate Reductase
- DHFR-TS
- Dihydrofolate Reductase–Thymidylate Synthase
- DHPS
- Dihydropteroate Synthase
- DMSO
- Dimethyl Sulfoxide
- dTMP
- Deoxythymidine Monophosphate
- dUMP
- Deoxyuridine Monophosphate
- dTTP
- Deoxythymidine Triphosphate
- DV
- Digestive Vacuole
- FAD
- Flavin Adenine Dinucleotide
- FPIX
- Ferriprotoporphyrin IX
- G1P
- Glucose-1-Phosphate
- Glc6P
- Glucose-6-Phosphate
- GDP
- Guanosine Diphosphate
- GTP
- Guanosine Triphosphate
- HPLC
- High-Performance Liquid Chromatography
- IRS
- Indoor Residual Spraying
- ITN
- Insecticide-Treated Net
- LC-MS
- Liquid Chromatography-Mass Spectrometry
- MPAG
- Malaria Policy Advisory Group
- NADP⁺
- Nicotinamide Adenine Dinucleotide Phosphate ion
- NADPH
- Nicotinamide Adenine Dinucleotide Phosphate
- PABA
- para-Aminobenzoic Acid
- PBS
- Phosphate-Buffered Saline
- PfCRT
- Plasmodium falciparum Chloroquine Resistance Transporter
- PfATP4
- Plasmodium falciparum Na⁺/H⁺-Dependent ATPase
- PfA-M1
- Plasmodium falciparum Alanyl-Aminopeptidase M1
- PPP
- Pentose Phosphate Pathway
- PY
- Pyrimethamine
- RDT
- Rapid Diagnostic Tests
- RPMI
- Roswell Park Memorial Institute Medium
- SAGE
- Strategic Advisory Group of Experts
- SD
- Sulfadoxine
- SDCQ
- Combination of Sulfadoxine + Chloroquine
- SDPY
- Combination of Sulfadoxine + Pyrimethamine
- SEM
- Standard Error of the Mean
- SMC
- Seasonal Malaria Chemoprevention
- TCA
- Tricarboxylic Acid Cycle
- THF
- Tetrahydrofolate
- TPP
- Thiamine Pyrophosphate
- UDP
- Uridine Diphosphate
- UDP-Glc
- Uridine Diphosphate-Glucose
- UTP
- Uridine Triphosphate
Text is read by the "Ask this paper" AI Q&A widget below. Extraction quality varies by source — PMC NXML preserves structure cleanly, OA-HTML may include some navigation residue, and OA-PDF can have broken hyphenation. The publisher copy (via DOI) is the canonical version.
My notes (saved in your browser only)
Ask this paper
Answers must be backed by verbatim quotes from this paper's full text. Hallucinated quotes are dropped automatically; if no verbatim passage answers the question, we say so. How this works
Citation neighborhood (no data yet)
We don't have any in-corpus citations linked to this paper yet. This is a recent paper (2025) — citers typically take a year or two to land, and the OpenAlex reference graph may still be filling in.
Source provenance
- europepmc
- last seen: 2026-05-20T01:45:00.602351+00:00
- unpaywall
- last seen: 2026-05-26T02:00:01.498150+00:00
License: CC-BY-NC-4.0