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Frontiers of Medicine

ISSN 2095-0217

ISSN 2095-0225(Online)

CN 11-5983/R

Postal Subscription Code 80-967

2018 Impact Factor: 1.847

Front Med    2011, Vol. 5 Issue (4) : 356-371     DOI: 10.1007/s11684-011-0159-1
Stem cell gene therapy: the risks of insertional mutagenesis and approaches to minimize genotoxicity
Chuanfeng Wu, Cynthia E. Dunbar()
Hematology Branch, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA
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Virus-based vectors are widely used in hematopoietic stem cell (HSC) gene therapy, and have the ability to integrate permanently into genomic DNA, thus driving long-term expression of corrective genes in all hematopoietic lineages. To date, HSC gene therapy has been successfully employed in the clinic for improving clinical outcomes in small numbers of patients with X-linked severe combined immunodeficiency (SCID-X1), adenosine deaminase deficiency (ADA-SCID), adrenoleukodystrophy (ALD), thalassemia, chronic granulomatous disease (CGD), and Wiskott-Aldrich syndrome (WAS). However, adverse events were observed during some of these HSC gene therapy clinical trials, linked to insertional activation of proto-oncogenes by integrated proviral vectors leading to clonal expansion and eventual development of leukemia. Numerous studies have been performed to understand the molecular basis of vector-mediated genotoxicity, with the aim of developing safer vectors and lower-risk gene therapy protocols. This review will summarize current information on the mechanisms of insertional mutagenesis in hematopoietic stem and progenitor cells due to integrating gene transfer vectors, discuss the available assays for predicting genotoxicity and mapping vector integration sites, and introduce newly-developed approaches for minimizing genotoxicity as a way to further move HSC gene therapy forward into broader clinical application.

Keywords gene therapy      hematopoietic stem cells      insertional mutagenesis      genotoxicity      induced pluripotent stem cell     
Corresponding Authors: E. Dunbar Cynthia,   
Issue Date: 05 December 2011
URL:     OR
DiseasesPatientsVector backboneTransgeneFollow-upSevere adverse effectsInsertion sites (clonal dominance)Clinical outcomeReferences
SCID-X19MFG γ-retroviral vectorγC9 years (range, 8 to 11)T-ALL in four patientsNo.1LMO2Died despite chemotherapy[1,2,10,15]
No.2LMO2Remission with chemotherapy
No.3CCND2Remission with chemotherapy
No.4LMO2, BMI1Remission with chemotherapy
Other 5No clonal dominanceAlive, improved immune function
SCID-X110MFG γ-retroviral vectorγC5 yearsT-ALL in one patientNo.1LMO2Remission with chemotherapy[14,16]
Other 9No clonal dominanceAlive with improved immune function
ADA-SCID10GIADAl MLV-based retroviral vectorADA4.0 years (range, 1.8 to 8.0)NoneIntegration hotspots near DYRK1A,BLM,LMO2, CCND2 and BCL2 no clonal selection )All alive with improved immune and metabolic parameters[3,4]
X-CGD2pSF7, SFFV-based retrovirus vectorgp91phox4 yearsMDSNo.1MDS1/Evi1, PRDM16, and SETBP1Died from sepsis 27 months[5,9]
MDSNo.2MDS1/Evi1, PRDM16, and SETBP1Underwent an allogeneic HSC transplantation at month 45
X-CGD3MFGS retroviral vectorgp91phox3 yearsNoneNo clonal dominanceClinical improvement[6]
X-ALD2HIV-1-derived lentiviral vectorABCD124 to 30 monthsNoneNo clonal dominanceAlive with decreased progression of neurologic phenotype[7]
β-thalassemia1HIV-1-derived lentiviral vectorβ-globin33 monthsNoneDominant HMGA2 expression cloneAlive and transfusion-independent[26]
WAS2CMMP retroviral vectorWASP3 yearsT-ALL in one patient(LMO2)LMO2, CCND2, and BMI1 in T-cells, and MDS1/EVI1, PRDM16, and SETBP1 in granulocytesT-ALL patient with ongoing chemotherapy, the other alive with improved immune function and decreased cytopenias[8]
Tab.1  Reported genotoxic events in HSC gene therapy clinical trials
Fig.1  Schematic diagrams for summarizing the different methods available to identify proviral insertion sites. (A)Methods requiring restriction enzyme digestion adjacent to insertion sites, including inverse PCR, LM-PCR, LAM-PCR and multi-arm optimized LAM-PCR. (B) Methods without restriction enzyme digestion, including FLEA-PCR, nrLAM-PCR and transposase MuA based-PCR. (Timeline shows the year when each method was first described.)
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