Predictable and efficient retroviral gene transfer into murine bone marrow repopulating cells using a defined vector dose

Li, Zhixiong; Schwieger, Maike; Lange, Claudia; Kraunus, Janine; Sun, Han‐Dong; Akker, Eric van den; Modlich, Ute; Serinsöz, Ebru; Will, Elke; Laer, Dorotheé von; Stocking, Carol; Fehse, Boris; Schiedlmeier, Bernhard; Baum, Christopher

doi:10.1016/j.exphem.2003.08.008

Cited by 60 publications

(50 citation statements)

References 52 publications

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“…53 DNA samples for real-time PCR analysis (copy numbers) and flow cytometry (FACSCalibur; Becton-Dickinson, Franklin Lakes, NJ, USA) were obtained 4 days after the last transduction. Quantitative PCR was performed using the Applied Biosystems 7300 Real-Time PCR System (Foster City, CA, USA), using the Quantitect SYBR Green kit (Qiagen, Hilden, Germany), as described previously.…”

Section: In Vitro Immortalization Assaymentioning

confidence: 99%

CTF/NF1 transcription factors act as potent genetic insulators for integrating gene transfer vectors

Gaussin

Modlich

Bauche³

et al. 2011

Gene Ther

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Gene transfer-based therapeutic approaches have greatly benefited from the ability of some viral vectors to efficiently integrate within the cell genome and ensure persistent transmission of newly acquired transgenes to the target cell progeny. However, integration of provirus has been associated with epigenetic repercussions that may influence the expression of both the transgene and cellular genes close to vector integration loci. The exploitation of genetic insulator elements may overcome both issues through their ability to act as barriers that limit transgene silencing and/or as enhancer-blockers preventing the activation of endogenous genes by the vector enhancer. We established quantitative plasmid-based assay systems to screen enhancerblocker and barrier genetic elements. Short synthetic insulators that bind to nuclear factor-I protein family transcription factors were identified to exert both enhancer-blocker and barrier functions, and were compared to binding sites for the insulator protein CTCF (CCCTC-binding factor). Gamma-retroviral vectors enclosing these insulator elements were produced at titers similar to their non-insulated counterparts and proved to be less genotoxic in an in vitro immortalization assay, yielding lower activation of Evi1 oncogene expression and reduced clonal expansion of bone marrow cells.

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Section: In Vitro Immortalization Assaymentioning

confidence: 99%

CTF/NF1 transcription factors act as potent genetic insulators for integrating gene transfer vectors

Gaussin

Modlich

Bauche³

et al. 2011

Gene Ther

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“…23 The vector is referred to as SF91-IE. A PCR fragment containing the cDNA of DTrkA was generated and cloned into NotI site before the IRES-EGFP cassette of SF91-IE.…”

Section: Retroviral Vectors and Vector Productionmentioning

confidence: 99%

“…SF91 retroviral vector has been shown to mediate efficient transgene expression in hematopoietic cells. 23 Cell-free high-titer supernatants containing the ecotropic envelope protein were generated as previously described. 23 …”

Section: Retroviral Vectors and Vector Productionmentioning

confidence: 99%

Remarkable leukemogenic potency and quality of a constitutively active neurotrophin receptor, ΔTrkA

et al. 2007

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Neurotrophins and their receptors play a key role in neurogenesis and survival. However, we and others have recently obtained evidence for a potential involvement of this receptor system in leukemia. To investigate mechanisms underlying the leukemogenic potential of activated neurotrophin receptor signaling, we analyzed in vivo leukemogenesis mediated by DTrkA, a mutant of TRKA (tropomyosin-related kinase A) isolated from a patient with acute myeloid leukemia (AML). Retroviral expression of DTrkA in myeloid 32D cells induced AML in syngeneic C3H/Hej mice (n ¼ 11/11, latency B4 weeks). C57Bl/6J mice transplanted with DTrkA-transduced primary lineage negative (Lin À ) bone marrow cells died of a transient polyclonal AML (n ¼ 7/15, latency of o12 days). Serial transplantation of AML cells did not re-induce this disease but rather acute lymphoblastic leukemia (ALL, latency 478 days). All primary recipients surviving the early AML developed clonal ALL or myeloid leukemia (latency 472 days) that required additional genetic lesions. PI3K and mTOR-raptor were identified as the crucial mediators of leukemic transformation, whereas STAT and MAP kinase signaling pathways were not activated. Thus, our findings reveal potent and unique transforming properties of altered neurotrophin receptor signaling in leukemogenesis, and encourage further analyses of neurotrophin receptors and downstream signaling events in hematological malignancies.

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“…Before infection, BM cells from B6 wt mice were enriched for progenitors by depletion of Lin neg cells using a Mouse Lineage Panel (BD PharMingen, Hamburg, Germany). Infections and transplantations were carried out as previously described (Schwieger et al, 2002;Li et al, 2003). To determine incidence of LTR-HSC, BM cells from two transplanted mice were pooled and sorted on basis of GFP hi /cKit hi /Sca1 þ and injected i.p.…”

Section: Infection and Bm Transplantionmentioning

confidence: 99%

Defining the oncogenic function of the TEL/AML1 (ETV6/RUNX1) fusion protein in a mouse model

et al. 2005

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The t(12;21) translocation, generating the TEL/AML1 fusion protein, is the most common genetic lesion in childhood cancer. Using a bone marrow transplantation model, we demonstrate that TEL/AML1 expression impinges on normal hematopoietic differentiation, leading to the in vivo accumulation and persistence of an early progenitor compartment with a Sca1 þ /Kit hi /CD11b þ phenotype and an increased self-renewal capacity, as documented by replating assays in vitro. Differentiation of these cells is not blocked, but the frequency of mature blood cells arising from TEL/AML1-transduced progenitors is low. Impaired differentiation is prominently observed in the pro-B-cell compartment, resulting in an proportional increase in early progenitors in vivo, consistent with the t(12;21) ALL phenotype. Despite the accumulation of both multipotent and B-cell progenitors in vivo, no leukemia induction was observed during an observation period of over 1 year. These results are consistent with findings in twins with concordant ALL, showing that TEL/AML1 generates a preleukemic clone in utero that persists for several years in a clinically covert fashion. Furthermore, our studies showed that the pointed domain of TEL/AML1, which recruits transcriptional repressors and directs oligomerization with either TEL/ AML1 or wild-type TEL, was essential for the observed differentiation impairment and could not be replaced with another oligomerization domain.

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Predictable and efficient retroviral gene transfer into murine bone marrow repopulating cells using a defined vector dose

Cited by 60 publications

References 52 publications

CTF/NF1 transcription factors act as potent genetic insulators for integrating gene transfer vectors

CTF/NF1 transcription factors act as potent genetic insulators for integrating gene transfer vectors

Remarkable leukemogenic potency and quality of a constitutively active neurotrophin receptor, ΔTrkA

Defining the oncogenic function of the TEL/AML1 (ETV6/RUNX1) fusion protein in a mouse model

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