Marking of peripheral T-lymphocytes by retroviral transduction and transplantation of CD34 + cells in a canine X-linked severe combined immunodeficiency model

• Published in: Veterinary immunology and immunopathology Vol. 117; no. 3; pp. 183 - 196
• Main Authors: Suter, Steven E., Gouthro, Terry A., O’Malley, Thomas, Hartnett, Brian J., McSweeney, Peter A., Moore, Peter F., Felsburg, Peter J., Haskins, Mark E., Henthorn, Paula S.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 15-06-2007
• Subjects: animal models; Animals; Antigens, CD34 - analysis; bone marrow transplant; Bone Marrow Transplantation; canine X-linked severe combined immunodeficiency; CD34; cell transplantation; complementary DNA; dog diseases; Dogs; Ex vivo gene therapy; Flow Cytometry; fluorescent labeling; gene therapy; gene transfer; genetic disorders; genetic transduction; Genetic Vectors; green fluorescent protein; Green Fluorescent Proteins - genetics; Hematopoietic progenitor cells; Polymerase Chain Reaction; protein synthesis; Retroviral gene transfer; retroviral vectors; severe combined immunodeficiency; T-lymphocytes; T-Lymphocytes - immunology; tissue transplantation; Transduction, Genetic; X-Linked Combined Immunodeficiency Diseases - immunology; X-Linked Combined Immunodeficiency Diseases - therapy; Hematopoietic progenitor cells; Retroviral gene transfer; Ex vivo gene therapy; CD34
• ISSN: 0165-2427; 1873-2534
• DOI: 10.1016/j.vetimm.2007.03.004

A retrovirus vector containing an enhanced green fluorescent protein complimentary DNA (EGFP cDNA) was used to mark and dynamically follow vector-expressing cells in the peripheral blood of bone marrow transplanted X-linked severe combined immunodeficient dogs. CD34
+ cells isolated from young normal dogs were transduced, using a 2 day protocol, with an amphotropic retroviral vector that expressed enhanced green fluorescent protein (EGFP) and the canine common gamma chain (γc) cDNAs. Following transplantation of the transduced cells, normal donor peripheral blood lymphocytes (PBL) appeared by 1 month post-bone marrow transplant (BMT) and rescued three of five treated dogs from their lethal immunodeficiency. PCR and flow cytometric analysis of post-BMT PBL documented the peripheral EGFP expressing cells as CD3
+ T cells, which varied from 0% to 28%. Sorting of EGFP
+ and EGFP
− peripheral blood T cells from two dogs, followed by vector PCR analysis, showed no evidence of vector shutdown. EGFP expression in B cells or monocytes was not detected. These marking experiments demonstrate that the transduction protocol did not abolish the lymphoid engraftment capability of
ex vivo transduced canine CD34
+ cells and supports the potential utility of the MSCV retroviral vector for gene transfer to XSCID affected canine hematopoietic progenitor cells (HPC).