Summary of Study ST002755
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 PR001716. The data can be accessed directly via it's Project DOI: 10.21228/M8WM6F 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 | ST002755 |
| Study Title | Metabolomics Profiling of the Antiproliferative, Anti-migratory and Anti-invasive Potential of Amlodipine in Lung Cancer Cells |
| Study Type | LC/MS/MS |
| Study Summary | Lung cancer is still among the most leading causes of cancer-related deaths across the world. Although chemotherapy is considered as a critical choice to manage/limit cancer growth in lung cancer patients with early-stage and advanced cancer stages, it has many limitations including, at least, the severe side effects and chemoresistance. The latter is one of the considerable challenges to lung cancer treatment. Therefore, identification of new alternative therapies with lesser cytotoxic effects when compared to the currently used chemotherapeutics is one of the current research approaches. Calcium channel blockers (CCBs) are emerging as anti-cancer agents in several cancer types. Our objective is to determine the cytotoxic effect of amlodipine on non-small cell lung cancer (NSCLC) cells. Colorimetric MTT cell proliferation assay was used to analyze cell viability following treatments with amlodipine in A549 and H1299 NSCLC cell lines. ANOVA and Tukey’s multiple comparison test were used to detect statistical significance. Half maximal (50%) inhibitory concentration (IC50) values were obtained by applying nonlinear regression curve fit analysis. To assess the effect of amlodipine on A549 and H1299 NSCLC cells migration and invasion scratch wound-healing assay and cell invasion assay were used. Our study revealed that amlodipine significantly reduced proliferation of cancer cells in a dose-dependent fashion with half maximal (50%) inhibitory concentration (IC50) values of 23 and 25.66 µM in A549 and H1299 cells, respectively. Furthermore, amlodipine was able to reduce the invasiveness and migration of cancer cells, both of which are hallmarks in the pathogenesis of cancer, in both cell lines in a dose-dependent manner. Accordingly, our study provides empirical evidence that amlodipine expresses anti-cancer effect to NSCLC cells. However, additional investigations are required to further confirm our results on a larger scale at the clinical level. |
| Institute | Sharjah Institute for Medical Research |
| Last Name | Facility |
| First Name | Core |
| Address | M32, SIMR, College of Pharmacy, Health Sciences, University of Sharjah |
| tims-tof@sharjah.ac.ae | |
| Phone | 065057656 |
| Submit Date | 2023-06-22 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | d |
| Analysis Type Detail | LC-MS |
| Release Date | 2023-07-18 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR001716 |
| Project DOI: | doi: 10.21228/M8WM6F |
| Project Title: | Metabolomics Profiling of the Antiproliferative, Anti-migratory and Anti-invasive Potential of Amlodipine in Lung Cancer Cells |
| Project Type: | LC-MS/MS |
| Project Summary: | Lung cancer is still among the most leading causes of cancer-related deaths across the world. Although chemotherapy is considered as a critical choice to manage/limit cancer growth in lung cancer patients with early-stage and advanced cancer stages, it has many limitations including, at least, the severe side effects and chemoresistance. The latter is one of the considerable challenges to lung cancer treatment. Therefore, identification of new alternative therapies with lesser cytotoxic effects when compared to the currently used chemotherapeutics is one of the current research approaches. Calcium channel blockers (CCBs) are emerging as anti-cancer agents in several cancer types. Our objective is to determine the cytotoxic effect of amlodipine on non-small cell lung cancer (NSCLC) cells. Colorimetric MTT cell proliferation assay was used to analyze cell viability following treatments with amlodipine in A549 and H1299 NSCLC cell lines. ANOVA and Tukey’s multiple comparison test were used to detect statistical significance. Half maximal (50%) inhibitory concentration (IC50) values were obtained by applying nonlinear regression curve fit analysis. To assess the effect of amlodipine on A549 and H1299 NSCLC cells migration and invasion scratch wound-healing assay and cell invasion assay were used. Our study revealed that amlodipine significantly reduced proliferation of cancer cells in a dose-dependent fashion with half maximal (50%) inhibitory concentration (IC50) values of 23 and 25.66 µM in A549 and H1299 cells, respectively. Furthermore, amlodipine was able to reduce the invasiveness and migration of cancer cells, both of which are hallmarks in the pathogenesis of cancer, in both cell lines in a dose-dependent manner. Accordingly, our study provides empirical evidence that amlodipine expresses anti-cancer effect to NSCLC cells. However, additional investigations are required to further confirm our results on a larger scale at the clinical level. |
| Institute: | Sharjah Institute for Medical Research |
| Last Name: | Facility |
| First Name: | Core |
| Address: | M32, SIMR, College of Pharmacy, Health Sciences, University of Sharjah |
| Email: | tims-tof@sharjah.ac.ae |
| Phone: | 065057656 |
Subject:
| Subject ID: | SU002862 |
| Subject Type: | Human |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
Factors:
Subject type: Human; Subject species: Homo sapiens (Factor headings shown in green)
| mb_sample_id | local_sample_id | Treatment |
|---|---|---|
| SA289539 | Amlodipine 05-02-10455 | Amlodipine |
| SA289540 | Amlodipine 06-01-10456 | Amlodipine |
| SA289541 | Amlodipine 01-01-10446 | Amlodipine |
| SA289542 | Amlodipine 04-02-10453 | Amlodipine |
| SA289543 | Amlodipine 06-02-10457 | Amlodipine |
| SA289544 | Amlodipine 05-01-10454 | Amlodipine |
| SA289545 | Amlodipine 04-01-10452 | Amlodipine |
| SA289546 | Amlodipine 01-02-10447 | Amlodipine |
| SA289547 | Amlodipine 02-02-10449 | Amlodipine |
| SA289548 | Amlodipine 02-01-10448 | Amlodipine |
| SA289549 | Amlodipine 03-01-10450 | Amlodipine |
| SA289550 | Amlodipine 03-02-10451 | Amlodipine |
| SA289551 | Control 05-02-10442 | Control |
| SA289552 | Control 05-01-10441 | Control |
| SA289553 | Control 06-02-10444 | Control |
| SA289554 | Control 04-02-10440 | Control |
| SA289555 | Control 06-01-10443 | Control |
| SA289556 | Control 01-01-10433 | Control |
| SA289557 | Control 02-01-10435 | Control |
| SA289558 | Control 01-02-10434 | Control |
| SA289559 | Control 02-02-10436 | Control |
| SA289560 | Control 03-01-10437 | Control |
| SA289561 | Control 03-02-10438 | Control |
| SA289562 | Control 04-01-10439 | Control |
| Showing results 1 to 24 of 24 |
Collection:
| Collection ID: | CO002855 |
| Collection Summary: | A549 human lung adenocarcinoma cell line was obtained from American Type Culture Collection (ATCC, Manassas, VA, USA). Cells were maintained in Roswell Park Memorial Institute (RPMI-1640) media supplemented with 10% fetal bovine serum (FBS), 100 U/mL penicillin, and 0.1 mg/mL streptomycin in a humidified atmosphere with 5% CO2 at 37°C. |
| Sample Type: | Lung |
Treatment:
| Treatment ID: | TR002871 |
| Treatment Summary: | Amlodipine was purchased from Tocris Bioscience (Bristol, UK). Amlodipine was dissolved in dimethyl sulfoxide (DMSO) to produce stock concentrations of 10, 50 and 100 mM DMSO. Of note, to minimize the cytotoxic effects of DMSO, the final concentration has never exceeded 0.1% in all treatment groups. Furthermore, the cells ere treated in a quadruplicate manner with DMSO as a positive control. |
Sample Preparation:
| Sampleprep ID: | SP002868 |
| Sampleprep Summary: | we used chloroform/methanol extraction protocol to increase the coverage of the extracted metabolites. At first, the samples (cells and buffer) were transferred into Eppendorf tubes then centrifuged at 14000 rpm for 5 minutes. Afterward, the buffer was discarded, and the cells were preserved. To each sample, 400 µL of the mixture containing one protease inhibitor tablet and 10 mL of lysis buffer was added. Following rest for 10 minutes, samples were transferred to 10 mL tubes, vortexed for 2–4 minutes, and sonicated with a COPLEY probe-sonicator (QSONICA SONICATOR, USA) for 30 seconds while utilizing a 30% amplifier in an ice bath. The samples were then transferred to Eppendorf tubes and centrifuged for 5 minutes at 14000 rpm. The supernatant was then transferred to another Eppendorf, and 400 µL of methanol and 300 µL of chloroform were added. Following that, the samples were vortexed for 30 seconds and centrifuged for 5 minutes at 14000 rpm. After that, two metabolite-containing layers are obtained, after transferring the upper layer of each sample to glass vials, 400 µl of methanol was added, followed by vertexing and centrifugation. The remaining supernatant was transferred to the same glass vials used before for the drying step, with the remaining protein pellets being air-dried for proteomics. A dried metabolomics sample was resuspended in 200 µL (0.1% formic acid in water) and injected into HPLC to be analysed by Q-TOF MS. |
| Extract Storage: | -80℃ |
Combined analysis:
| Analysis ID | AN004472 |
|---|---|
| Analysis type | MS |
| Chromatography type | Reversed phase |
| Chromatography system | Bruker Elute |
| Column | Hamilton Intensity Solo 2 C18(100 x 2.1 mm, 1.8 um) |
| MS Type | ESI |
| MS instrument type | QTOF |
| MS instrument name | Bruker timsTOF |
| Ion Mode | POSITIVE |
| Units | AU |
Chromatography:
| Chromatography ID: | CH003357 |
| Chromatography Summary: | amples were chromatographically separated by inline reversed-phase chromatography using the Elute HPG 1300 pumps and Elute Autosampler (Bruker, Darmstadt, Germany) with solvent A 0.1% FA in HPLC grade water and solvent B 0.1% FA in ACN. A Hamilton Intensity Solo 2 C18 column (100 mm x 2.1 mm, 1.8µm beads) was maintained at 35C. For metabolomics, 10 µL was injected twice for each sample and eluted using a 30-minute gradient as follows: 1% ACN was held for 2 minutes, ramping to 99% ACN over 15 minutes, held at 99% ACN for 3 minutes before re-equilibrating to 1% ACN for 10 minutes. Flow rates were 250 µL/min for elution and 350 µL/min for re-equilibration. |
| Instrument Name: | Bruker Elute |
| Column Name: | Hamilton Intensity Solo 2 C18(100 x 2.1 mm, 1.8 um) |
| Column Temperature: | 35 |
| Flow Gradient: | The gradient program was: 0–2 min, 99% A: 1% B; 2–17 min, 99–1% A: 1–99% B; 17–20 min, 99% B: 1% A. The flow rate was fixed at 0.25 mL/min. Subsequently, 20–20.1 min 99% B to 99% A; 20.1–28.5 min, 99% A: 1% B at 0.35 mL/min flow rate; 28.5–30 min; 99% A: 1% B at 0.25 mL/min. |
| Flow Rate: | 250 uL/min |
| Solvent A: | Water (0.1% Formic Acid) |
| Solvent B: | ACN (0.1% Formic Acid) |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS004219 |
| Analysis ID: | AN004472 |
| Instrument Name: | Bruker timsTOF |
| Instrument Type: | QTOF |
| MS Type: | ESI |
| MS Comments: | The MS analysis was performed using a TimsTOF (Bruker, Darmstadt, Germany) with Apollo II electrospray ionization (ESI) source. The drying gas was set to flow at 10 L/min and the drying temperature to 220C and the nebulizer pressure to 2.2 bar. The capillary voltage was 4500 V and the end plate offset 500V. For metabolomics the scan range was 20-1300 m/z. The collision energy was set to 20 eV, the cycle time to 0.5 seconds with a relative minimum intensity threshold of 400 counts per thousand and target intensity of 20,000. Sodium formate was injected as an external calibrant in the first 0.3 minutes of each LC-MS/MS run. MetaboScape 4.0 software was used for metabolite processing and statistical analysis (Bruker Daltonics). The following parameters for molecular feature identification and "bucketing" were set in MS Comments the T-ReX 2D/3D workflow: For peak detection, a minimum intensity threshold of 1,000 counts is required, as well as a minimum peak duration of 7 spectra, with feature quantification determine using peak area. The file masses were recalibrated based on the external calibrant injected between 0-0.3 min. |
| Ion Mode: | POSITIVE |
