title: Imaging stem cell-derived persistent foci after in vivo selection of lentiviral MGMT-P140K transduced murine bone marrow cells.
abstract: The engraftment of hematopoietic stem cells (HSCs) after drug resistance gene transfer and drug selection may recapitulate stress response hematopoiesis, but the processes remain elusive. Homing, trafficking, and localization of transduced cells and the impact of insertion site on focal expansion have not been well characterized. With the goal of optimizing and understanding these processes under conditions of low multiplicity of infection (MOI) lentiviral gene transfer, we used drug resistance gene O(6)-methylguanine-DNA methyltransferase (MGMT)-P140K and in vivo selection to enrich for transduced and transgene-expressing HSCs. To systemically monitor homing, trafficking, and expansion after transplantation and drug selection over time, we linked MGMT-P140K to the firefly luciferase gene in lentiviral self-inactivating vectors. Periodic bioluminescence imaging (BLI) of transplanted recipients was followed for up to 9 months after both primary and secondary transplantation. Initial dispersion and widespread early homing and engraftment were transient, followed by detection of persistent and discrete foci at stable tissue sites after in vivo drug selection. From these studies, we concluded that drug resistance gene transfer followed by early or late drug selection can result in stable gene expression and cell expansion in persistent foci of transduced bone marrow cells that often remain in fixed sites for extended periods of time.
authors.affiliation: Division of Hematology/Oncology, Case Comprehensive Cancer Center, Case Western Reserve University, Cleveland, Ohio 44106, USA.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Lin
authors.forename: Perrin
authors.initials: P
authors.lastname: Cheung
authors.forename: Justin C
authors.initials: JC
authors.lastname: Roth
authors.forename: David L
authors.initials: DL
authors.lastname: Wilson
authors.forename: Stanton L
authors.initials: SL
authors.lastname: Gerson
mesh_terms: D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology* / Q000378:metabolism*; D016026:Bone Marrow Transplantation; D002460:Cell Line; D002478:Cells, Cultured; D015254:DNA Modification Methylases / Q000235:genetics*; D045643:DNA Repair Enzymes / Q000235:genetics*; D005260:Female; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D051379:Mice; D008807:Mice, Inbred BALB C; D014161:Transduction, Genetic / Q000379:methods*; D025521:Tumor Suppressor Proteins / Q000235:genetics*
chemical_list: D025521:Tumor Suppressor Proteins; D015254:DNA Modification Methylases; C528331:MGMT protein, mouse; D045643:DNA Repair Enzymes