title: Lentiviral vector induced insertional haploinsufficiency of Ebf1 causes murine leukemia.
abstract: Integrating vectors developed on the basis of various retroviruses have demonstrated therapeutic potential following genetic modification of long-lived hematopoietic stem and progenitor cells. Lentiviral vectors (LV) are assumed to circumvent genotoxic events previously observed with γ-retroviral vectors, due to their integration bias to transcription units in comparison to the γ-retroviral preference for promoter regions and CpG islands. However, recently several studies have revealed the potential for gene activation by LV insertions. Here, we report a murine acute B-lymphoblastic leukemia (B-ALL) triggered by insertional gene inactivation. LV integration occurred into the 8th intron of Ebf1, a major regulator of B-lymphopoiesis. Various aberrant splice variants could be detected that involved splice donor and acceptor sites of the lentiviral construct, inducing downregulation of Ebf1 full-length message. The transcriptome signature was compatible with loss of this major determinant of B-cell differentiation, with partial acquisition of myeloid markers, including Csf1r (macrophage colony-stimulating factor (M-CSF) receptor). This was accompanied by receptor phosphorylation and STAT5 activation, both most likely contributing to leukemic progression. Our results highlight the risk of intragenic vector integration to initiate leukemia by inducing haploinsufficiency of a tumor suppressor gene. We propose to address this risk in future vector design.
authors.affiliation: Institute of Experimental Hematology, Hannover Medical School, Hannover, Germany.
authors.forename: Dirk
authors.initials: D
authors.lastname: Heckl
authors.forename: Adrian
authors.initials: A
authors.lastname: Schwarzer
authors.forename: Reinhard
authors.initials: R
authors.lastname: Haemmerle
authors.forename: Doris
authors.initials: D
authors.lastname: Steinemann
authors.forename: Cornelia
authors.initials: C
authors.lastname: Rudolph
authors.forename: Britta
authors.initials: B
authors.lastname: Skawran
authors.forename: Sabine
authors.initials: S
authors.lastname: Knoess
authors.forename: Johanna
authors.initials: J
authors.lastname: Krause
authors.forename: Zhixiong
authors.initials: Z
authors.lastname: Li
authors.forename: Brigitte
authors.initials: B
authors.lastname: Schlegelberger
authors.forename: Christopher
authors.initials: C
authors.lastname: Baum
authors.forename: Ute
authors.initials: U
authors.lastname: Modlich
mesh_terms: D000818:Animals; D016000:Cluster Analysis; D015536:Down-Regulation / Q000235:genetics; D005260:Female; D020869:Gene Expression Profiling; D015973:Gene Expression Regulation, Leukemic; D023061:Gene Order; D005822:Genetic Vectors; D042822:Genomic Instability; D057895:Haploinsufficiency; D006412:Hematopoietic Stem Cells / Q000378:metabolism; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D007938:Leukemia / Q000235:genetics*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D016254:Mutagenesis, Insertional; D015452:Precursor B-Cell Lymphoblastic Leukemia-Lymphoma / Q000235:genetics / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional; D050799:STAT5 Transcription Factor / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic; D014161:Transduction, Genetic; D016662:Virus Integration
chemical_list: C489921:Ebf1 protein, mouse; D050799:STAT5 Transcription Factor; D015534:Trans-Activators