Summary of Study ST004194

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR002643. The data can be accessed directly via it's Project DOI: 10.21228/M8154F This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

Study ID	ST004194
Study Title	PfK13-associated artemisinin resistance slows drug activation and enhances antioxidant defence, which can be overcome with sulforaphane
Study Summary	Goals - Identify mode of action of Sulforaphane using untargeted metabolomics. Results - Untargeted metabolomics of sulforaphane treated infected cells identified sulforaphane mode of action is via alkylation of parasite thiols especially glutathione. Conclusion - Sulforaphane acts on the parasite via alkylation of parasites's major antioxidants such as glutathione.
Institute	Monash University
Last Name	Siddiqui
First Name	Ghizal
Address	381 Royal Parade, Parkville, Melbourne, Victoria, 3052, Australia
Email	ghizal.siddiqui@monash.edu
Phone	99039282
Submit Date	2025-09-07
Raw Data Available	Yes
Raw Data File Type(s)	mzML, raw(Thermo)
Analysis Type Detail	LC-MS
Release Date	2025-10-06
Release Version	1

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Project:

Project ID:	PR002643
Project DOI:	doi: 10.21228/M8154F
Project Title:	PfK13-associated artemisinin resistance slows drug activation and enhances antioxidant defence, which can be overcome with sulforaphane
Project Summary:	Artemisinin resistance is increasingly prevalent in Africa, raising concerns and highlighting the need to better understand the cellular mechanisms behind this resistance. In Plasmodium falciparum, artemisinin resistance is primarily attributed to mutations in the PfKelch13 (PfK13) gene. In this study, we performed proteomics analysis on a range of artemisinin-resistant (both laboratory-generated and field isolates) and sensitive P. falciparum parasites at 3-6 (hours post invasion) hpi ring- and 22-24 hpi trophozoite-stage, revealing dysregulation of only PfK13 protein abundance. Reduced PfK13 levels were linked to impaired hemoglobin digestion, decreased free heme levels, and consequently, decreased artemisinin activation. Resistant parasites also exhibited elevated thiol levels, indicating a more reduced state. Targeting the parasite redox capacity with sulforaphane potentiated artemisinin activity in vitro and in vivo, offering a potential strategy to overcome resistance. Our findings provide critical insights into the molecular mechanisms of artemisinin resistance and suggest novel therapeutic interventions to restore drug sensitivity.
Institute:	Monash University
Last Name:	Siddiqui
First Name:	Ghizal
Address:	381 Royal Parade, Parkville, Melbourne, Victoria, 3052, Australia
Email:	ghizal.siddiqui@monash.edu
Phone:	99039282

Subject:

Subject ID:	SU004346
Subject Type:	Cultured cells
Subject Species:	Plasmodium falciparum
Taxonomy ID:	5833

Factors:

Subject type: Cultured cells; Subject species: Plasmodium falciparum (Factor headings shown in green)

mb_sample_id	local_sample_id	Treatment	Sample source
SA483649	DMSO_1	DMSO	iRBC
SA483650	DMSO_2	DMSO	iRBC
SA483651	DMSO_3	DMSO	iRBC
SA483652	DMSO_4	DMSO	iRBC
SA483653	DMSO_5	DMSO	iRBC
SA483654	DMSO_6	DMSO	iRBC
SA483655	SFN_1	Sulforaphane-30uM-3h	iRBC
SA483656	SFN_2	Sulforaphane-30uM-3h	iRBC
SA483657	SFN_3	Sulforaphane-30uM-3h	iRBC
SA483658	SFN_4	Sulforaphane-30uM-3h	iRBC
SA483659	SFN_5	Sulforaphane-30uM-3h	iRBC
SA483660	SFN_6	Sulforaphane-30uM-3h	iRBC

Showing results 1 to 12 of 12

Showing results 1 to 12 of 12

Collection:

Collection ID:	CO004339
Collection Summary:	Cultured malaria infected cells (Pf3D7 wildtype parasites) were harvested using magnet to achieve >80% parasitaemia. Infection was achieved using Pf3D7 parasite strains and infected red blood cells were then harvested using magnet enrichment, see DOI 10.1186/1475-2875-13-112.
Sample Type:	Blood (whole)

Treatment:

Treatment ID:	TR004355
Treatment Summary:	>80% parasitaemia parasites were treated with 30 µM of sulforaphane (SFN) for 3 h or DMSO control. High infection is required for the identification of parasite metabolites perturbed as a result of treatment.

Sample Preparation:

Sampleprep ID:	SP004352
Sampleprep Summary:	After drug treatments metabolites of magnet harvested parasites were collected. Briefly parasite metabolism was quenched by cooling samples to between 3-5°C. All samples were centrifuged at 650 g for 3 min, the supernatant was removed, and the pellet washed in 500 µL of ice-cold PBS. Samples were again centrifuged at 650 g for 3 min and pellets were resuspended in 150 µL of ice-cold extraction buffer (100% methanol) and quickly resuspended. The samples were then incubated on a vortex mixer for 1 h at 4°C before being centrifuged at 17,000 g for 10 min; from this 100 µL of supernatant was collected and stored at -80°C until analysis. For each sample, another 10 µL was collected and pooled, to serve as a quality control (QC) sample.

Sampleprep ID:

SP004352

Sampleprep Summary:

After drug treatments metabolites of magnet harvested parasites were collected. Briefly parasite metabolism was quenched by cooling samples to between 3-5°C. All samples were centrifuged at 650 g for 3 min, the supernatant was removed, and the pellet washed in 500 µL of ice-cold PBS. Samples were again centrifuged at 650 g for 3 min and pellets were resuspended in 150 µL of ice-cold extraction buffer (100% methanol) and quickly resuspended. The samples were then incubated on a vortex mixer for 1 h at 4°C before being centrifuged at 17,000 g for 10 min; from this 100 µL of supernatant was collected and stored at -80°C until analysis. For each sample, another 10 µL was collected and pooled, to serve as a quality control (QC) sample.

Chromatography:

Chromatography ID:	CH005290
Chromatography Summary:	Metabolite analysis was performed by liquid chromatography-mass spectrometry LC-MS using hydrophilic interaction liquid chromatography (HILIC) and high-resolution (Q-Exactive Orbitrap, Thermo Fisher) MS. Briefly, samples (10 μL) were injected onto a Dionex RSLC U3000 LC system (Thermo) fitted with a ZIC-pHILIC column (5 μm particle size, 4.6 by 150 mm; Merck) and 20 mM ammonium carbonate (A) and acetonitrile (B) were used as the mobile phases. A 30 min gradient starting from 80% B to 50% B over 15 mins, followed by washing at 5% B for 3 mins and re-equilibration at 80% B, was used.
Instrument Name:	Thermo Vanquish
Column Name:	Merck SeQuant ZIC-HILIC (150 x 4.6mm,5um)
Column Temperature:	25
Flow Gradient:	A 22 min gradient starting from 80% B to 50% B over 15 mins, followed by washing at 5% B for 3 mins and re-equilibration at 80% B, was used.
Flow Rate:	0.35 ml/min
Solvent A:	100% water; 20 mM ammonium carbonate
Solvent B:	100% Acetonitrile
Chromatography Type:	HILIC

Analysis:

Analysis ID:	AN006966
Analysis Type:	MS
Chromatography ID:	CH005290
Num Factors:	2
Num Metabolites:	244
Units:	normalised ion counts

Analysis ID:	AN006967
Analysis Type:	MS
Chromatography ID:	CH005290
Num Factors:	2
Num Metabolites:	227
Units:	normalised ion counts