Dataset Information

Correction of murine ADAMTS13 deficiency by hematopoietic progenitor cell-mediated gene therapy.

ABSTRACT: ADAMTS13, a metalloprotease primarily synthesized in liver and endothelial cells, cleaves von Willebrand factor (VWF) at the central A2 domain, thereby reducing the sizes of circulating VWF multimers. Genetic or acquired deficiency of plasma ADAMTS13 activity leads to a potentially fatal syndrome, thrombotic thrombocytopenic purpura (TTP). To date, plasma infusion or exchange is the only proven effective therapy for TTP. In search for a better therapy, an autologous transplantation of hematopoietic progenitor cells transduced ex vivo with a self-inactivating lentiviral vector encoding a full-length murine Adamts13 and an enhanced green fluorescent protein (GFP) reporter gene was performed in Adamts13(-/-) mice after irradiation. All recipient mice showed detectable ADAMTS13 antigen and proteolytic activity in plasma despite only low levels of bone marrow chimerism. The levels of plasma ADAMTS13 were sufficient to eliminate the ultralarge VWF multimers and offered systemic protection against ferric chloride-induced arterial thrombosis. The data suggest that hematopoietic progenitor cells can be genetically modified ex vivo and transplanted in an autologous model to provide adequate levels of functional ADAMTS13 metalloprotease. This success may provide the basis for development of a novel therapeutic strategy to cure hereditary TTP in humans.

SUBMITTER: Laje P

PROVIDER: S-EPMC2652365 | biostudies-literature | 2009 Mar

REPOSITORIES: biostudies-literature

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Correction of murine ADAMTS13 deficiency by hematopoietic progenitor cell-mediated gene therapy.

Laje Pablo P Shang Dezhi D Cao Wenjing W Niiya Masami M Endo Masayuki M Radu Antoneta A DeRogatis Nicole N Scheiflinger Friedrich F Zoltick Philip W PW Flake Alan W AW Zheng X Long XL

Blood 20090113 10

ADAMTS13, a metalloprotease primarily synthesized in liver and endothelial cells, cleaves von Willebrand factor (VWF) at the central A2 domain, thereby reducing the sizes of circulating VWF multimers. Genetic or acquired deficiency of plasma ADAMTS13 activity leads to a potentially fatal syndrome, thrombotic thrombocytopenic purpura (TTP). To date, plasma infusion or exchange is the only proven effective therapy for TTP. In search for a better therapy, an autologous transplantation of hematopoie ...[more]

PMID: 19141866

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Project description:The chemokine stromal cell-derived factor (SDF)-1?/CXCL12 and its receptor CXC chemokine receptor 4 (CXCR4) play a crucial role in the homing/engraftment and retention of hematopoietic stem/progenitor cells (HSPCs) in the bone marrow. It has been shown using the viral gene transfer technique that CXCR4 overexpression on human CD34(+) HSPC significantly improves their engraftment in murine models. However, clinical trials with gene therapy have revealed safety concerns related to the immunogenicity of the viral carriers, due to the random integration of viral genes into the host genome. Therefore, a method for CXCR4 gene delivery into HSPC that is safe, nonviral, and highly efficient is needed to improve clinical transplantation and gene therapies. In this work, we investigated the nonviral CXCR4 gene delivery into HSPC using the cationic liposome agent IBAfect. We used CD34(+) cells from cord blood and the models of immature hematopoietic cells expressing CD34 antigen, namely, leukemic cell lines KG-1a and KG-1. Transfection efficiency was determined by flow cytometric analysis 12, 24, 48, and 72 h after transfection, and the viability of cells analyzed by trypan blue exclusion and MTS assays. The functional response of CXCR4-transfected HSPC toward an SDF-1? gradient was determined by chemotaxis assay. We found that ~25% transfection is achieved for KG-1a and KG-1 cells and 20% for HSPC, and that the viability of CXCR4-transfected HSPC is not significantly altered. More importantly, overexpression of CXCR4 using IBAfect significantly increased the chemotaxis of KG-1 cells and HSPC toward SDF-1?. However, we tested 2 other commercially available cationic liposomes (Lipofectamine 2000 and 1,2-dioleoyl-3-trimethylammonium-propane [DOTAP]) in parallel, and we found that they failed to deliver the CXCR4 gene into cells under the same conditions. These results suggest that IBAfect-mediated in vitro gene delivery to overexpress CXCR4 on HSPC is a safe and efficient technique with great potential for improving the efficacy of HSPC transplantation and gene therapy protocols.

Dataset Information

Correction of murine ADAMTS13 deficiency by hematopoietic progenitor cell-mediated gene therapy.

Publications

Correction of murine ADAMTS13 deficiency by hematopoietic progenitor cell-mediated gene therapy.

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