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GEO help: Mouse over screen elements for information. |
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Status |
Public on Jul 21, 2021 |
Title |
Single-cell RNA sequencing of murine Colon26 tumors after combination treatment with HMGN1 immunostimulatory peptide (minP1) with anti-PD-L1 antibody |
Organism |
Mus musculus |
Experiment type |
Expression profiling by high throughput sequencing
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Summary |
In this study, we explore a new combination therapy of anti-PD-L1 antibody with an immunostimulatory peptide derived from an alarmin high mobility group nucleosome binding protein 1 (HMGN1). Combination treatment of HMGN1 immunostimulatory peptide (minP1) with anti-PD-L1 antibody exerted robust anti-tumor effects in the Colon26 subcutaneous murine model. To understand transcriptomic differences of the immune cell populations in the tumor-bearing mice after treatment with single-agent or combination therapy of minP1 with anti-PD-L1 antibody, we performed single-cell RNA sequencing (scRNA-seq) analysis on CD45+ immune cell populations using the BD Rapsody system, NEBNext UltraII FS library prep kit, and a Illumina Novaseq 6000 S4 flowcell.
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Overall design |
Single-cell RNA sequencing (scRNA-seq) was performed on CD45+ cells from the Colon26 tumors of BALB/c mice after treatments. There are untreated (n=3, biological replicates), minP1-treated (n=4, biological replicates), anti-PD-L1 antibody-treated (n=3, biological replicates), and minP1/anti-PD-L1 antibody-treated (n=4, biological replicates) groups. Intratumoral CD45+ cells from different treatment groups were stained with Hashtag anti-CD45 antibody to separate each sample first, were processed in separate experimental runs of the BD Rapsody system, were performed single-cell cDNA library by using NEBNext UltraII FS library prep kit, and were run by an Illumina Novaseq 6000 S4 flowcell (67 bp read1 and 140 bp read2) (Illumina, USA) to a depth of approximately 100,000 reads/cell. , which finally results in 14 samples across which 7,613 cells were passed through quality check and were analyzed by Seurat. For mouse data, after adapter removal by using Cutadapt-2.10 (ref), gene-expression libraries were aligned to the mm10 reference transcriptome by using Bowtie2-2.3.4.1, and count matrices were generated using home-built shell scripts and the modified python script of BD Rhapsody workflow. Valid cell barcodes were identified as cell barcodes above inflection threshold of knee-plot of total read counts of each cell barcode identified by DropletUtils package (Griffiths JA et al. Nat. Commun. 2018). Each sample origin and doublets were identified based on fold-change of the normalized read counts of Hashtag antibodies. Data were log-transformed [log(TPM + 1)] for all downstream analyses, most of which were performed using the R software package Seurat v2.3.4 (http://satijalab.org/seurat) to analyze immune cell populations. This submission represents 14 unique/individual tumor-bearing mice
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Contributor(s) |
Chen C, Shichino S, Satoshi U, Ishiwata Y, Yokochi S, Yang D, Oppenheim JJ, Ogiwara H, Deshimaru S, Kanno Y, Ogawa T, Shibayama S, Matsushima K |
Citation(s) |
34344641 |
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Submission date |
Feb 22, 2021 |
Last update date |
Oct 20, 2021 |
Contact name |
Alex Chang-Yu Chen |
E-mail(s) |
cachen@mgh.harvard.edu
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Organization name |
Massachusetts General Hospital
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Department |
Krantz Family Center for Cancer Research
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Lab |
Sen Lab
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Street address |
149 13th Street, 7th floor, Room 7103
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City |
Boston |
State/province |
MA |
ZIP/Postal code |
02129 |
Country |
USA |
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Platforms (1) |
GPL24247 |
Illumina NovaSeq 6000 (Mus musculus) |
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Samples (2) |
GSM5098817 |
Colon26 tumors, CD45+ cells, treated (cDNA library) |
GSM5098818 |
Colon26 tumors, CD45+ cells, treated (hashtag library) |
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Relations |
BioProject |
PRJNA703835 |
SRA |
SRP307464 |
Supplementary file |
Size |
Download |
File type/resource |
GSE167192_BDRhapsody_CellLabelSequences_Sept2017_SSED.xlsx |
12.8 Kb |
(ftp)(http) |
XLSX |
GSE167192_ProcessedData_Final_Matrix_Hashtag.txt.gz |
32.9 Mb |
(ftp)(http) |
TXT |
GSE167192_metadata_14individualmice.xlsx |
21.0 Kb |
(ftp)(http) |
XLSX |
SRA Run Selector |
Raw data are available in SRA |
Processed data are available on Series record |
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