Summary of Study ST003160
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 PR001965. The data can be accessed directly via it's Project DOI: 10.21228/M8QQ8P 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 | ST003160 |
| Study Title | New class of heterospirocyclic compounds present strong and rapid activity against artemisinin- and multidrug-resistant P. falciparum parasites |
| Study Summary | Malaria remains a significant health burden and a leading contributor to global mortality rates. Increasing drug resistance creates an urgent demand for novel treatment options. We have synthesised a new class of heterospirocyclic compounds with novel chemical connectivities. Compounds 25 and 26 display antimalarial activity within 24 h and have similar potency against a panel of drug-resistant strains of Plasmodium falciparum, the most virulent of human malaria parasites, including parasites resistant to the frontline artemisinin antimalarials. C25 and C26 do not induce major toxicity in kidney- and hepatic-derived human cell lines, highlighting their specificity. Untargeted metabolomics analysis of P. falciparum infected red blood cells revealed that the mechanism of action of C25 involves disruption of the pyrimidine biosynthesis pathway and haemoglobin catabolism. These heterospirocyclic compounds represent a promising opportunity for antimalarial drug development and could prove relevant against drug resistant malaria. |
| Institute | Monash University |
| Last Name | Giannangelo |
| First Name | Carlo |
| Address | 381 Royal Parade, Parkville, Victoria, 3052, Australia |
| carlo.giannangelo@monash.edu | |
| Phone | 99039282 |
| Submit Date | 2024-04-06 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | raw(Thermo) |
| Analysis Type Detail | LC-MS |
| Release Date | 2024-04-29 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR001965 |
| Project DOI: | doi: 10.21228/M8QQ8P |
| Project Title: | New class of heterospirocyclic compounds present strong and rapid activity against artemisinin- and multidrug-resistant P. falciparum parasites |
| Project Summary: | Malaria remains a significant health burden and a leading contributor to global mortality rates. Increasing drug resistance creates an urgent demand for novel treatment options. We have synthesised a new class of heterospirocyclic compounds with novel chemical connectivities. Compounds 25 and 26 display antimalarial activity within 24 h and have similar potency against a panel of drug-resistant strains of Plasmodium falciparum, the most virulent of human malaria parasites, including parasites resistant to the frontline artemisinin antimalarials. C25 and C26 do not induce major toxicity in kidney- and hepatic-derived human cell lines, highlighting their specificity. Untargeted metabolomics analysis of P. falciparum infected red blood cells revealed that the mechanism of action of C25 involves disruption of the pyrimidine biosynthesis pathway and haemoglobin catabolism. These heterospirocyclic compounds represent a promising opportunity for antimalarial drug development and could prove relevant against drug resistant malaria. |
| Institute: | Monash University |
| Last Name: | Giannangelo |
| First Name: | Carlo |
| Address: | 381 Royal Parade, Parkville, Victoria, 3052, Australia |
| Email: | carlo.giannangelo@monash.edu |
| Phone: | 99039282 |
Subject:
| Subject ID: | SU003279 |
| Subject Type: | Cultured cells |
| Subject Species: | Plasmodium falciparum |
| Taxonomy ID: | 5833 |
| Species Group: | 3D7 strain |
Factors:
Subject type: Cultured cells; Subject species: Plasmodium falciparum (Factor headings shown in green)
| mb_sample_id | local_sample_id | Sample source | Treatment |
|---|---|---|---|
| SA342802 | ATV_1 | iRBC | Atovaquone |
| SA342803 | ATV_4 | iRBC | Atovaquone |
| SA342804 | ATV_3 | iRBC | Atovaquone |
| SA342805 | ATV_2 | iRBC | Atovaquone |
| SA342806 | C25_4 | iRBC | C25 |
| SA342807 | C25_3 | iRBC | C25 |
| SA342808 | C25_1 | iRBC | C25 |
| SA342809 | C25_2 | iRBC | C25 |
| SA342810 | C25_0HR_3 | iRBC | C25_0HR_control |
| SA342811 | C25_0HR_2 | iRBC | C25_0HR_control |
| SA342812 | C25_0HR_4 | iRBC | C25_0HR_control |
| SA342813 | C25_0HR_1 | iRBC | C25_0HR_control |
| SA342814 | DMSO_3 | iRBC | DMSO |
| SA342815 | DMSO_2 | iRBC | DMSO |
| SA342816 | DMSO_1 | iRBC | DMSO |
| SA342817 | DMSO_4 | iRBC | DMSO |
| SA342818 | PS027_1 | iRBC | PS027 |
| SA342819 | PS027_4 | iRBC | PS027 |
| SA342820 | PS027_2 | iRBC | PS027 |
| SA342821 | PS027_3 | iRBC | PS027 |
| Showing results 1 to 20 of 20 |
Collection:
| Collection ID: | CO003272 |
| Collection Summary: | Plasmodium falciparum cultures (3D7 strain) synchronised to the trophozoite stage were magnet purified to achieve a parasitaemia of >90% and haematocrit of 0.5%. Four different experiment conditions were used: (i) the active heterospirocyclic compound, compound 25 (C25); (ii) the inactive precursor of compound 25, PS027; (iii) atovaquone; and (iv) DMSO alone as the negative (no drug) control. Following compound incubation (5 h), cultures were centrifuged at 2,000 RPM for 5 minutes, the media was removed and the cell pellets were washed in ice cold PBS. All samples were centrifuged at 1,200g for 3 minutes, after which metabolites were extracted from 5e7 cells using 100 µL of ice-cold methanol extraction solvent. Samples were placed on a shaker at 4°C for 1 h and spun at 13,000 RPM for 10 minutes at 4 °C. Supernatants were transferred into high-performance liquid chromatography (HPLC) vials and stored at -80°C until liquid chromatography-mass spectrometry (LC-MS) analysis. A pooled biological quality control sample consisting of a 15 µL aliquot from each sample was also generated for analytical quality control and to aid metabolite identification. |
| Sample Type: | Plasmodium cells |
Treatment:
| Treatment ID: | TR003288 |
| Treatment Summary: | Plasmodium falciparum cultures (3D7 strain) synchronised to the trophozoite stage were magnet purified to achieve a parasitaemia of >90% and haematocrit of 0.5%. Four different experiment conditions were used: (i) the active heterospirocyclic compound, compound 25 (C25); (ii) the inactive precursor of compound 25, PS027; (iii) atovaquone; and (iv) DMSO alone as the negative (no drug) control. Following compound incubation (5 h), cultures were centrifuged at 2,000 RPM for 5 minutes, the media was removed and the cell pellets were washed in ice cold PBS. All samples were centrifuged at 1,200g for 3 minutes, after which metabolites were extracted from 5e7 cells using 100 µL of ice-cold methanol extraction solvent. Samples were placed on a shaker at 4°C for 1 h and spun at 13,000 RPM for 10 minutes at 4 °C. Supernatants were transferred into high-performance liquid chromatography (HPLC) vials and stored at -80°C until liquid chromatography-mass spectrometry (LC-MS) analysis. A pooled biological quality control sample consisting of a 15 µL aliquot from each sample was also generated for analytical quality control and to aid metabolite identification. |
Sample Preparation:
| Sampleprep ID: | SP003286 |
| Sampleprep Summary: | Plasmodium falciparum cultures (3D7 strain) synchronised to the trophozoite stage were magnet purified to achieve a parasitaemia of >90% and haematocrit of 0.5%. Four different experiment conditions were used: (i) the active heterospirocyclic compound, compound 25 (C25); (ii) the inactive precursor of compound 25, PS027; (iii) atovaquone; and (iv) DMSO alone as the negative (no drug) control. Following compound incubation (5 h), cultures were centrifuged at 2,000 RPM for 5 minutes, the media was removed and the cell pellets were washed in ice cold PBS. All samples were centrifuged at 1,200g for 3 minutes, after which metabolites were extracted from 5e7 cells using 100 µL of ice-cold methanol extraction solvent. Samples were placed on a shaker at 4°C for 1 h and spun at 13,000 RPM for 10 minutes at 4 °C. Supernatants were transferred into high-performance liquid chromatography (HPLC) vials and stored at -80°C until liquid chromatography-mass spectrometry (LC-MS) analysis. A pooled biological quality control sample consisting of a 15 µL aliquot from each sample was also generated for analytical quality control and to aid metabolite identification. |
Combined analysis:
| Analysis ID | AN005184 | AN005185 |
|---|---|---|
| Analysis type | MS | MS |
| Chromatography type | HILIC | HILIC |
| Chromatography system | Thermo Dionex Ultimate 3000 RS | Thermo Dionex Ultimate 3000 RS |
| Column | Merck SeQuant ZIC-HILIC (150 x 4.6mm,5um) | Merck SeQuant ZIC-HILIC (150 x 4.6mm,5um) |
| MS Type | ESI | ESI |
| MS instrument type | Orbitrap | Orbitrap |
| MS instrument name | Thermo Q Exactive Orbitrap | Thermo Q Exactive Orbitrap |
| Ion Mode | POSITIVE | NEGATIVE |
| Units | Peak height | Peak height |
Chromatography:
| Chromatography ID: | CH003922 |
| Instrument Name: | Thermo Dionex Ultimate 3000 RS |
| Column Name: | Merck SeQuant ZIC-HILIC (150 x 4.6mm,5um) |
| Column Temperature: | 25 |
| Flow Gradient: | 0–15 min, 80–50% B; 15–18 min, 50–5% B; 18–21 min, 5% B; 21–24 min, 5–80% B and 24–32 min, 80% B |
| Flow Rate: | 0.3 mL/min |
| Solvent A: | 100% Water; 20 mM ammonium carbonate |
| Solvent B: | 100% Acetonitrile |
| Chromatography Type: | HILIC |
MS:
| MS ID: | MS004917 |
| Analysis ID: | AN005184 |
| Instrument Name: | Thermo Q Exactive Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | Liquid chromatography-mass spectrometry (LC-MS) data was acquired on a Q-Exactive Orbitrap mass spectrometer (Thermo Scientific) coupled with high-performance liquid chromatography system (HPLC, Dionex Ultimate 3000 RS, Thermo Scientific) as described previously (Creek et al. 2016). Samples within the LC-MS batch were sorted according to blocks of replicates and randomized. To facilitate metabolite identification, approximately 350 authentic metabolite standards were analysed prior to the LC-MS batch and their peaks and retention time manually checked using the MZmine software. Pooled biological quality control samples and extraction solvent blanks were analysed periodically throughout the batch to monitor LC-MS signal reproducibility and aid downstream metabolite identification procedures. Raw LC-MS metabolomics data were analysed using the open source software, IDEOM (http://mzmatch.sourceforge.net/ideom.php). Briefly, the IDEOM workflow uses msconvert to convert raw files to mzXML format, XCMS (Centwave) to pick LC-MS peak signals and MZmatch for alignment and annotation of related metabolite peaks. Default IDEOM parameters were used to eliminate unwanted noise and artefact peaks. Confident metabolite identification was made by matching accurate masses to retention time of the ~350 authentic standards. When these authentic standards were unavailable, putative metabolite identification used accurate mass and predicted retention times. Metabolite abundance was represented by LC-MS peak height. |
| Ion Mode: | POSITIVE |
| MS ID: | MS004918 |
| Analysis ID: | AN005185 |
| Instrument Name: | Thermo Q Exactive Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | Liquid chromatography-mass spectrometry (LC-MS) data was acquired on a Q-Exactive Orbitrap mass spectrometer (Thermo Scientific) coupled with high-performance liquid chromatography system (HPLC, Dionex Ultimate 3000 RS, Thermo Scientific) as described previously (Creek et al. 2016). Samples within the LC-MS batch were sorted according to blocks of replicates and randomized. To facilitate metabolite identification, approximately 350 authentic metabolite standards were analysed prior to the LC-MS batch and their peaks and retention time manually checked using the MZmine software. Pooled biological quality control samples and extraction solvent blanks were analysed periodically throughout the batch to monitor LC-MS signal reproducibility and aid downstream metabolite identification procedures. Raw LC-MS metabolomics data were analysed using the open source software, IDEOM (http://mzmatch.sourceforge.net/ideom.php). Briefly, the IDEOM workflow uses msconvert to convert raw files to mzXML format, XCMS (Centwave) to pick LC-MS peak signals and MZmatch for alignment and annotation of related metabolite peaks. Default IDEOM parameters were used to eliminate unwanted noise and artefact peaks. Confident metabolite identification was made by matching accurate masses to retention time of the ~350 authentic standards. When these authentic standards were unavailable, putative metabolite identification used accurate mass and predicted retention times. Metabolite abundance was represented by LC-MS peak height. |
| Ion Mode: | NEGATIVE |
