Abstract:
:Insertional mutagenesis has been associated with malignant cell transformation in gene therapy protocols, leading to discussions about vector security. Therefore, clonal analysis is important for the assessment of vector safety and its impact on patient health. Here, we report a unique approach to assess dynamic changes in clonality of lentivirus transduced cells upon Sanger sequence analysis of a specially designed genetic barcode. In our approach, changes in the electropherogram peaks are measured and compared between successive time points, revealing alteration in the cell population. After in vitro validation, barcoded lentiviral libraries carrying IL2RG or LMO2 transgenes, or empty vector were used to transduce mouse hematopoietic (ckit+) stem cells, which were subsequently transplanted in recipient mice. We found that neither the empty nor IL2RG encoding vector had an effect on cell dynamics. In sharp contrast, the LMO2 oncogene was associated with altered cell dynamics even though hematologic counts remained unchanged, suggesting that the barcode could reveal changes in cell populations not observed by the frontline clinical assay. We describe a simple and sensitive method for the analysis of clonality, which could be easily used by any laboratory for the assessment of cellular behavior upon lentiviral transduction.
journal_name
Mol Ther Methods Clin Devjournal_title
Molecular therapy. Methods & clinical developmentauthors
Zanatta DB,Tsujita M,Borelli P,Aguiar RB,Ferrari DG,Strauss BEdoi
10.1038/mtm.2014.52subject
Has Abstractpub_date
2014-11-19 00:00:00pages
14052issn
2329-0501pii
S2329-0501(16)30121-8journal_volume
1pub_type
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journal_title:Molecular therapy. Methods & clinical development
pub_type: 杂志文章
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abstract::[This corrects the article DOI: 10.1038/mtm.2015.16.]. ...
journal_title:Molecular therapy. Methods & clinical development
pub_type: 已发布勘误
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pub_type: 杂志文章
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journal_title:Molecular therapy. Methods & clinical development
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journal_title:Molecular therapy. Methods & clinical development
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