title: Lentiviral gene transfer of TCIRG1 into peripheral blood CD34(+) cells restores osteoclast function in infantile malignant osteopetrosis.
abstract: Infantile malignant osteopetrosis (IMO) is a rare, lethal, autosomal recessive disorder characterized by non-functional osteoclasts. More than 50% of the patients have mutations in the TCIRG1 gene, encoding for a subunit of the osteoclast proton pump. The aim of this study was to restore the resorptive function of IMO osteoclasts by lentiviral mediated gene transfer of the TCIRG1 cDNA. CD34(+) cells from peripheral blood of five IMO patients and from normal cord blood were transduced with lentiviral vectors expressing TCIRG1 and GFP under a SFFV promoter, expanded in culture and differentiated on bone slices to mature osteoclasts. qPCR analysis and western blot revealed increased mRNA and protein levels of TCIRG1, comparable to controls. Vector corrected IMO osteoclasts generated increased release of Ca(2+) and bone degradation product CTX-I into the media as well as increased formation of resorption pits in the bone slices, while non-corrected IMO osteoclasts failed to resorb bone. Resorption was approximately 70-80% of that of osteoclasts generated from cord blood. Furthermore, transduced CD34(+) cells successfully engrafted in NSG-mice. In conclusion we provide the first evidence of lentiviral-mediated correction of a human genetic disease affecting the osteoclastic lineage.
authors.affiliation: Department of Molecular Medicine and Gene Therapy, Lund Strategic Center for Stem Cell Biology, Lund, Sweden.
authors.forename: Ilana
authors.initials: I
authors.lastname: Moscatelli
authors.forename: Christian Schneider
authors.initials: CS
authors.lastname: Thudium
authors.forename: Carmen
authors.initials: C
authors.lastname: Flores
authors.forename: Ansgar
authors.initials: A
authors.lastname: Schulz
authors.forename: Maria
authors.initials: M
authors.lastname: Askmyr
authors.forename: Natasja Stæhr
authors.initials: NS
authors.lastname: Gudmann
authors.forename: Nanna Merete
authors.initials: NM
authors.lastname: Andersen
authors.forename: Oscar
authors.initials: O
authors.lastname: Porras
authors.forename: Morten Asser
authors.initials: MA
authors.lastname: Karsdal
authors.forename: Anna
authors.initials: A
authors.lastname: Villa
authors.forename: Anders
authors.initials: A
authors.lastname: Fasth
authors.forename: Kim
authors.initials: K
authors.lastname: Henriksen
authors.forename: Johan
authors.initials: J
authors.lastname: Richter
mesh_terms: D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism*; D002478:Cells, Cultured; D002648:Child; D002675:Child, Preschool; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D015151:Immunoblotting; D007223:Infant; D007231:Infant, Newborn; D016086:Lentivirus / Q000235:genetics*; D008297:Male; D051379:Mice; D016513:Mice, SCID; D010010:Osteoclasts / Q000166:cytology / Q000378:metabolism; D010022:Osteopetrosis / Q000235:genetics* / Q000628:therapy*; D016133:Polymerase Chain Reaction; D025262:Vacuolar Proton-Translocating ATPases / Q000235:genetics* / Q000378:metabolism*
chemical_list: D018952:Antigens, CD34; C472421:TCIRG1 protein, human; D025262:Vacuolar Proton-Translocating ATPases
keywords: Hematopoietic stem cells; Infantile malignant osteopetrosis; Lentiviral gene transfer; Osteoclasts; TCIRG1