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Series GSE184895 Query DataSets for GSE184895
Status Public on Oct 01, 2021
Title Monitoring CAR T cell generation with a CD8-targeted lentiviral vector by single cell transcriptomics
Organism Homo sapiens
Experiment type Expression profiling by high throughput sequencing
Summary Quantifying gene expression in individual cells can substantially improve our understanding about complex genetically engineered cell products such as chimeric antigen receptor (CAR) T cells. Here we designed a single-cell RNA sequencing (scRNA-seq) approach to monitor the delivery of a CD19-CAR gene via lentiviral vectors (LVs), i.e. the conventional VSV-LV and the CD8-targeted CD8-LV. LV exposed human donor PBMC were evaluated for a panel of 400 immune-response related genes including LV-specific probes. The resulting data revealed a trimodal expression for the CAR and CD8A demanding for a careful distribution-based identification of CAR T cells and CD8+ lymphocytes in scRNA-seq analysis. The fraction of T cells expressing high CAR levels was in concordance with flow cytometry results. More than 97% of the cells hit by CD8-LV expressed the CD8A gene. Remarkably, the majority of the potential off-target cells were in fact on-target cells resulting in a target cell selectivity of above 99%. Beyond that, the differential gene expression analysis revealed the upregulation of restriction factors in CAR-negative cells thus explaining their protection from CAR gene transfer. In summary, we provide a workflow and subsetting approach for scRNA-Seq enabling reliable distinction between transduced and untransduced cells during CAR T cell generation.
 
Overall design Analysis of lentiviral vector generated products
 
Contributor(s) Charitidis FT, Adabi E, Thalheimer FB, Clarke C, Buchholz CJ
Citation(s)
  • Charitidis FT, Adabi E, Thalheimer FB, Clarke C et al. Monitoring CAR T cell generation with a CD8-targeted lentiviral vector by single-cell transcriptomics. Mol Ther Methods Clin Dev 2021 Dec 10;23:359-369. PMID: 34729382
Submission date Sep 28, 2021
Last update date Dec 14, 2022
Contact name Christian J. Buchholz
E-mail(s) christian.buchholz@pei.de
Organization name Paul Ehrlich Institut
Street address Paul-Ehrlich-Strasse 51-59
City Langen
State/province Hessen
ZIP/Postal code 63225
Country Germany
 
Platforms (1)
GPL21697 NextSeq 550 (Homo sapiens)
Samples (3)
GSM5599953 PBMCs, Untransduced
GSM5599954 PBMCs, VSV-LV
GSM5599955 PBMCs, CD8-LV
Relations
BioProject PRJNA766855
SRA SRP339084

Download family Format
SOFT formatted family file(s) SOFTHelp
MINiML formatted family file(s) MINiMLHelp
Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE184895_1046_vial_gene_masked_amplicons.fasta.gz 47.5 Kb (ftp)(http) FASTA
GSE184895_RAW.tar 3.6 Mb (http)(custom) TAR (of CSV)
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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