Abstract
Fanconi anemia (FA) is an autosomal-recessive disorder associated with hematopoietic failure and it is a candidate for hematopoietic stem cell (HSC)-directed gene therapy. However, the characteristically reduced HSC numbers found in FA patients, their ineffective mobilization from the marrow, and re-oxygenation damage during ex vivo manipulation have precluded clinical success using conventional in vitro approaches. We previously demonstrated that lentiviral vector (LV) particles reversibly attach to the cell surface where they gain protection from serum complement neutralization. We reasoned that cellular delivery of LV to the bone marrow niche could avoid detrimental losses during FA HSC mobilization and in vitro modification. Here, we demonstrate that a VSV-G pseudotyped lentivector, carrying the FANCC transgene, can be transmitted from carrier to bystander cells. In cell culture and transplantation models of FA, we further demonstrate that LV carrier cells migrate along SDF-1α gradients and transfer vector particles that stably integrate and phenotypically correct the characteristic DNA alkylator sensitivity in murine and human FA-deficient target bystander cells. Altogether, we demonstrate that cellular homing mechanisms can be harnessed for the functional phenotype correction in murine FA hematopoietic cells.Electronic supplementary materialThe online version of this article (doi:10.1186/s13287-016-0431-z) contains supplementary material, which is available to authorized users.
Highlights
Fanconi anemia (FA) is a multisystem disorder resulting from mutations in one of at least 22 genes that encode the FANC family of DNA repair proteins [1, 2]
Considerable evidence supports the involvement of CXCR4SDF-1α as the critical axis for homing to the bone marrow compartment [31,32,33]
In proof of principle studies, we used low-dose radiation to elicit SDF-1αinduced homing of carrier hematopoietic stem and progenitor cell (HSPC) to the bone marrow to mediate in situ delivery of the therapeutic lentiviral vector (LV)-GFPFANCC vector to endogenous Fancc–/– HSPCs
Summary
Fanconi anemia (FA) is a multisystem disorder resulting from mutations in one of at least 22 genes that encode the FANC family of DNA repair proteins [1, 2]. We previously reported that cell surface-bound HIV-1-derived LV particles gain protection from serum neutralization and transfer between cells with relatively greater efficiency compared with several cell-free virions [18, 19]. We exploited those observations in a strategy that combines in vitro exposure of LV to “carrier” cells that traffic to the bone marrow, thereby minimizing in vitro manipulation of FA HSCs while achieving stable transduction through cellular in situ delivery. Results showed that carrier cells can migrate along a chemotactic gradient of stromal derived factor-1α (SDF-1α) and traffic FANCC expressing LV to Fancc–/– murine hematopoietic stem and progenitor cell (HSPC) target cells, with subsequent transduction (TD) and expansion under selection pressure
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