RRC ID 53914
Author Hamada M, Nishio N, Okuno Y, Suzuki S, Kawashima N, Muramatsu H, Tsubota S, Wilson MH, Morita D, Kataoka S, Ichikawa D, Murakami N, Taniguchi R, Suzuki K, Kojima D, Sekiya Y, Nishikawa E, Narita A, Hama A, Kojima S, Nakazawa Y, Takahashi Y.
Title Integration Mapping of piggyBac-Mediated CD19 Chimeric Antigen Receptor T Cells Analyzed by Novel Tagmentation-Assisted PCR.
Journal EBioMedicine
Abstract Insertional mutagenesis is an important risk with all genetically modified cell therapies, including chimeric antigen receptor (CAR)-T cell therapy used for hematological malignancies. Here we describe a new tagmentation-assisted PCR (tag-PCR) system that can determine the integration sites of transgenes without using restriction enzyme digestion (which can potentially bias the detection) and allows library preparation in fewer steps than with other methods. Using this system, we compared the integration sites of CD19-specific CAR genes in final T cell products generated by retrovirus-based and lentivirus-based gene transfer and by the piggyBac transposon system. The piggyBac system demonstrated lower preference than the retroviral system for integration near transcriptional start sites and CpG islands and higher preference than the lentiviral system for integration into genomic safe harbors. Integration into or near proto-oncogenes was similar in all three systems. Tag-PCR mapping is a useful technique for assessing the risk of insertional mutagenesis.
Volume 34
Pages 18-26
Published 2018-8-1
DOI 10.1016/j.ebiom.2018.07.008
PII S2352-3964(18)30251-2
PMID 30082227
PMC PMC6116345
MeSH DNA Transposable Elements / genetics* High-Throughput Nucleotide Sequencing Humans Lentivirus / genetics Polymerase Chain Reaction / methods* Receptors, Antigen, T-Cell / genetics* Retroviridae / genetics T-Lymphocytes*
IF 6.68
Times Cited 3
Resource
DNA material CSII-CMV-MCS (RDB04377)