Summary of Study ST002209
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 PR001410. The data can be accessed directly via it's Project DOI: 10.21228/M8G42C 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 | ST002209 |
| Study Title | Metabolomic analysis to assess response to immunotherapy for malignant brain tumors: Part 3 |
| Study Type | Cancer surrogate biomarker discovery |
| Study Summary | An effective immune response in patients with cancer treated with immunotherapy includes dendritic cell (DC) activation and migration followed by stimulation of CD8 and CD4 T cells. This then leads to the activation, proliferation and further activation of other immune cell populations including NK cells or immunosuppressive populations such as Tregs and myeloid derived suppressor cells (MDSCs). These studies were carried out utilizing murine brain tumor models treated with an RNA DC vaccine platform. We hypothesized that metabolomic analyses of urines would be sensitive to the action of this diverse set of immune cells. The objective of this study was to evaluate the feasibility of using metabolomics to follow immune responses after immunotherapy. We chose NMR as our analytical technique of choice, as it has many favorable qualities that make it ideal for analyses of urine. |
| Institute | University of Florida |
| Department | Applied Physiology and Kinesiology |
| Laboratory | Rm 042 |
| Last Name | Khattri |
| First Name | Ram |
| Address | 1200 Newell Dr., ARB 240, Gainesville, FL, 32611, USA |
| rbk11@ufl.edu | |
| Phone | 3307856045 |
| Submit Date | 2022-06-09 |
| Num Groups | 5 |
| Total Subjects | 24 |
| Num Males | NA |
| Num Females | NA |
| Study Comments | Metabolomic profiling of urine samples |
| Publications | Metabolomics journal (submitted) |
| Raw Data Available | Yes |
| Raw Data File Type(s) | fid |
| Analysis Type Detail | NMR |
| Release Date | 2023-06-16 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR001410 |
| Project DOI: | doi: 10.21228/M8G42C |
| Project Title: | Metabolomic analysis to assess response to immunotherapy for malignant brain tumors: Part 1 |
| Project Type: | Study of the urine and serum in mice treated with DC vaccine via 1H NMR |
| Project Summary: | The objective of this project was to identify a peripheral metabolomic profile to serve as a biomarker of response to immunotherapy for the treatment of malignant brain tumors. |
| Institute: | University of Florida |
| Department: | Neurosurgery Department, Medical college, University of Florida |
| Last Name: | Khattri |
| First Name: | Ram |
| Address: | 1200 Newell Dr., ARB 240, Gainesville, FL, 32611, USA |
| Email: | rbk11@ufl.edu |
| Phone: | 3307856045 |
| Funding Source: | This work was partially funded by the Florida Center for Brain Tumor Research (FCBTR), and by generous benefactors to the University of Florida, Peter and Angela Dziegielewski, who established the Eilzabeth Dziegielewski Glioblastoma Research Fund, and Rosalinde Wolfe, who established the Greg Wolfe Brain Tumor Research Fund. RK and MM were supported by funding from National Institutes of Health (U24-DK097209 and 5U2C-DK119889). All NMR portion of this study was performed in McKnight Brain Institute at National High Magnetic Field Laboratory’s Advanced Magnetic Resonance Imaging and Spectroscopy (AMRIS) Facility, which is funded by National Science Foundation Cooperative Agreement No. DMR-1644779 and the State of Florida. |
| Project Comments: | Study of the urine and serum in mice treated with DC vaccine via 1H NMR |
| Publications: | Metabolomics journal (submitted) |
| Contributors: | Farhad Dastmalchi, Ram B. Khattri, Marc A. McLeod, Kaitlyn Melnick, Loic P. Deleyrolle, Yusuf Mehkri, Aida Karachi, Paul Kubilis, Shu Wang, Duane A. Mitchell, Matthew E. Merritt, Maryam Rahman |
Subject:
| Subject ID: | SU002295 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
| Genotype Strain: | C57BL/6J |
| Gender: | Not applicable |
| Animal Animal Supplier: | Jackson Labs (Bar Harbor, ME) |
| Animal Housing: | Housed in a temperature of 22 oC |
| Animal Light Cycle: | 12-hour light/12-hour dark |
| Animal Water: | free access to food and water (3-5 animals per cage). |
Factors:
Subject type: Mammal; Subject species: Mus musculus (Factor headings shown in green)
| mb_sample_id | local_sample_id | Group |
|---|---|---|
| SA211487 | Maryam_serum_Control-1 | Control |
| SA211488 | Maryam_serum_Control-5 | Control |
| SA211489 | Maryam_serum_Control-4 | Control |
| SA211490 | Maryam_serum_Control-2 | Control |
| SA211491 | Maryam_serum_Control-3 | Control |
| SA211492 | Maryam_serum_PostV_24hrs-5 | PostV_24hrs |
| SA211493 | Maryam_serum_PostV_24hrs-4 | PostV_24hrs |
| SA211494 | Maryam_serum_PostV_24hrs-1 | PostV_24hrs |
| SA211495 | Maryam_serum_PostV_24hrs-3 | PostV_24hrs |
| SA211496 | Maryam_serum_PostV_24hrs-2 | PostV_24hrs |
| SA211497 | Maryam_serum_PostV_48hrs-4 | PostV_48hrs |
| SA211498 | Maryam_serum_PostV_48hrs-5 | PostV_48hrs |
| SA211499 | Maryam_serum_PostV_48hrs-1 | PostV_48hrs |
| SA211500 | Maryam_serum_PostV_48hrs-3 | PostV_48hrs |
| SA211501 | Maryam_serum_PostV_48hrs-2 | PostV_48hrs |
| SA211502 | Maryam_serum_PostV_7days-5 | PostV_7 days |
| SA211503 | Maryam_serum_PostV_7days-4 | PostV_7 days |
| SA211504 | Maryam_serum_PostV_7days-3 | PostV_7 days |
| SA211505 | Maryam_serum_PostV_7days-1 | PostV_7 days |
| SA211506 | Maryam_serum_PostV_7days-2 | PostV_7 days |
| SA211507 | Maryam_serum_PostV_Day0-4 | PostV_Day0 |
| SA211508 | Maryam_serum_PostV_Day0-2 | PostV_Day0 |
| SA211509 | Maryam_serum_PostV_Day0-3 | PostV_Day0 |
| SA211510 | Maryam_serum_PostV_Day0-1 | PostV_Day0 |
| SA211511 | Maryam_serum_PreV_Day0-4 | PreV_Day0 |
| SA211512 | Maryam_serum_PreV_Day0-5 | PreV_Day0 |
| SA211513 | Maryam_serum_PreV_Day0-3 | PreV_Day0 |
| SA211514 | Maryam_serum_PreV_Day0-1 | PreV_Day0 |
| SA211515 | Maryam_serum_PreV_Day0-2 | PreV_Day0 |
| Showing results 1 to 29 of 29 |
Collection:
| Collection ID: | CO002288 |
| Collection Summary: | Each animal was in a separate chamber individually. The chamber bottom was covered with clean parafilm. Animals were kept in the chamber until urination. Urine drops were collected using sterile syringes. Samples were stored at -80 °C until further analysis. Serum samples were collected from peripheral blood. Blood was drawn from the facial vein and was kept in the room temperature for 30 minutes to coagulate. Coagulated blood was centrifuged at 1500 rpm for 15 minutes at 4°C. Then serum was collected from the top and was stored at -20 °C until further analysis. |
| Sample Type: | Blood (serum) |
| Collection Method: | Each animal was in a separate chamber individually. The chamber bottom was covered with clean parafilm. Animals were kept in the chamber until urination. Urine drops were collected using sterile syringes. Samples were stored at -80 °C until further analysis. Serum samples were collected from peripheral blood. Blood was drawn from the facial vein and was kept in the room temperature for 30 minutes to coagulate. Coagulated blood was centrifuged at 1500 rpm for 15 minutes at 4°C. Then serum was collected from the top and was stored at -20 °C until further analysis. |
| Collection Location: | University of Florida, Neurosurgery Department, Medical college, University of Florida |
| Collection Frequency: | Pre-vaccination, post vaccination day0, 24 hrs, 48 hrs of post DC vaccination and after 7 days. |
| Storage Conditions: | -80℃ |
| Collection Vials: | cryovials |
| Storage Vials: | cryovials |
Treatment:
| Treatment ID: | TR002307 |
| Treatment Summary: | DC vaccine treatment Naïve C57/BL6 mice were used for tumor implantation. B16F10-OVA (2 × 104 cells/brain) tumor cell lines were injected intracranially. Antigen specific T cells (3 × 107 cells/50 µL of PBS) were infused into the animals intravenous (IV) five days after tumor implantation and the DC vaccine (1x106 cells/50 µL of PBS per animal) was injected intradermally on the same day. Both antigen-specific T cells and DC vaccine were administered once. Urine samples were collected after DC vaccination at several timepoints. Finally, animals were euthanized when they reached the endpoints. Sample collection Each animal was in a separate chamber individually. The chamber bottom was covered with clean parafilm. Animals were kept in the chamber until urination. Urine drops were collected using sterile syringes. Samples were stored at -80 °C until further analysis. Serum samples were collected from peripheral blood. |
| Animal Anesthesia: | isoflurane |
| Animal Fasting: | non-fasted |
| Animal Endp Euthanasia: | Euthanasia was carried out by thoracotomy followed by cervical dislocation. |
Sample Preparation:
| Sampleprep ID: | SP002301 |
| Sampleprep Summary: | To isolate metabolites from serum samples, perchloric acid (PCA) extraction was performed. Fifty microliters of serum sample was taken in a vial and 1 mL of ice cold 6 % (v/v) PCA was added to it. The mixture was vortexed for 2 minutes and centrifuged at 4 oC and 13.2 K rpm speed. Supernatant from the centrifuged sample was taken in a new vial and neutralized with 5 M potassium hydroxide making its pH near to 6.5. Centrifugation (4 oC temperature, 13.2 K rpm speed) of the neutralized solution was performed to remove the potassium perchlorate salt. The supernatant was dried using a room temperature lyophilizer (Thermo-Scentific, Dallas, USA). The dried powder was suspended in 200 µL of ultra-pure water. The pH of the mixture was adjusted to 7.0 using 1M sodium hydroxide and 1M hydrochloride. Centrifugation (4 oC temperature, 13.2 K rpm speed) of the mixture was performed to remove excess insoluble salt. The supernatant solution was again lyophilized. The resulted dried powder was resuspended in deuterated phosphate buffer system to prepare the NMR samples. In case of serum samples, the solid was dissolved in a 90/10 mixture of 50 mM sodium phosphate buffer (pH 7.0) with 2 mM of ethylene diamine tetra acetic acid (EDTA) and a Chenomx internal standard (Chenomx Inc., Alberta, Canada) consisting of 5 mM 2,2-Dimethyl-2-silapentane-5-sulfonate sodium salt (DSS-D6) and 0.2% NaN3 in D2O. |
| Sampleprep Protocol Filename: | DC_Vaccine_treated_NMR_serum_Procedures.docx |
| Processing Method: | Lyophilization |
| Processing Storage Conditions: | -80℃ |
| Extraction Method: | Perchloric acid extraction method |
| Extract Storage: | -80℃ |
| Sample Resuspension: | In 500 microliter of 150 mM phosphate buffer (pH 7.2) with 2 mM EDTA, 0.5 mM DSS and 0.2% sodium azide for serum samples. |
| Sample Spiking: | 0.5 mM DSS for urine samples |
Analysis:
| Analysis ID: | AN003612 |
| Laboratory Name: | McKnight Brain Institute |
| Analysis Type: | NMR |
| Acquisition Date: | 4/18/2017 |
| Acquisition Parameters File: | DC_Vaccine_treated_NMR_serum_Procedures.docx |
| Software Version: | Topspin |
| Operator Name: | Ram Khattri |
| Processing Parameters File: | DC_Vaccine_treated_NMR_serum_Procedures.docx |
| Detector Type: | Bruker 600 MHz |
| Data Format: | fid, 1r |
| Results File: | ST002209_AN003612_Results.txt |
| Units: | Peak intensity |
NMR:
| NMR ID: | NM000245 |
| Analysis ID: | AN003612 |
| Instrument Name: | Bruker 600 MHz |
| Instrument Type: | FT-NMR |
| NMR Experiment Type: | 1D-1H |
| Field Frequency Lock: | Deuterium |
| Standard Concentration: | 0.5mM DSS |
| Spectrometer Frequency: | 600 MHz |
| NMR Probe: | 5mm CPTXI 1H/D -13C/15N Z-GRD Z44866/0026 |
| NMR Solvent: | Phosphate buffer (pH 7.2) + 2 mM EDTA + 0.5 mM DSS + 0.2% of sodium azide in deuterated environment |
| NMR Tube Size: | 5 mm O.D. |
| Shimming Method: | Topshim |
| Pulse Sequence: | PROJECT |
| Water Suppression: | presat |
| Pulse Width: | 90-degree |
| Receiver Gain: | 512 |
| Offset Frequency: | 4.78 ppm |
| Chemical Shift Ref Cpd: | DSS |
| Temperature: | 298.2 oK |
| Number Of Scans: | 64 |
| Dummy Scans: | 8 |
| Acquisition Time: | 4.54s |
| Relaxation Delay: | 3s |
| Spectral Width: | 7211.5 Hz |
| Num Data Points Acquired: | 32768 |
| Real Data Points: | 65536 |
| Line Broadening: | 0.22 Hz |
| Zero Filling: | 65,536 points |
| Apodization: | Exponential |
| Baseline Correction Method: | Spline |
| Chemical Shift Ref Std: | 0 ppm for DSS |
| Binned Increment: | 0.001 ppm |
| Binned Data Excluded Range: | >9.5 ppm and < 0.5 ppm regions |
| NMR Results File: | NMR_data_3303.txt UNITS:Peak intensity |
