title: High Level of Perforin Expression Is Required for Effective Correction of Hemophagocytic Lymphohistiocytosis.
abstract: Perforin-1 mutations result in a potentially fatal hemophagocytic lymphohistiocytosis (HLH) with heightened immune activation, hypercytokinemia, pancytopenia, and end-organ damage. At present, hematopoietic stem cell (HSC) transplantation is curative, but limited by donor availability and associated mortality, making gene therapy an attractive alternative approach for HLH. We reported that perforin expression driven by cellular promoters in lentiviral (LV) vectors resulted in significant, albeit partial, correction of the inflammatory features in a murine model of HLH. We hypothesized that the level of perforin expression achieved per cell from ectopic moderate-strength cellular promoters (phosphoglycerate kinase gene/perforin-1 gene) is inadequate and thus engineered an LV vector using a viral promoter (MND; a modified Moloney murine leukemia virus long terminal repeat with myeloproliferative sarcoma virus enhancer) containing microRNA126 target sequences to restrict perforin expression in HSCs. We show here that the MND-LV vector restored perforin expression to normal levels in a perforin-deficient human natural killer cell line and perforin gene-corrected Perforin1-/-transplant recipients, whereas cellular promoters drove only partial correction. On lymphocytic choriomeningitis virus challenge, the clinical scores and survival improved only with the MND-LV vector, but inflammatory markers and cytotoxicity were improved with all LV vectors. Our studies suggest that although moderate levels of expression can result in partial amelioration of the HLH phenotype, high levels of perforin expression per cell are required for complete correction of HLH.
authors.affiliation: 1 Department of Molecular Genetics, Biochemistry, and Microbiology, University of Cincinnati , Cincinnati, Ohio.|2 Division of Experimental Hematology and Oncology, Cincinnati Children's Hospital Medical Center (CCHMC), Cincinnati, Ohio.
authors.forename: Swati
authors.initials: S
authors.lastname: Tiwari
authors.affiliation: 3 Division of Allergy/Immunology, Department of Pediatrics, CCHMC , Cincinnati, Ohio.
authors.forename: Adrianne
authors.initials: A
authors.lastname: Hontz
authors.affiliation: 2 Division of Experimental Hematology and Oncology, Cincinnati Children's Hospital Medical Center (CCHMC), Cincinnati, Ohio.
authors.forename: Catherine E
authors.initials: CE
authors.lastname: Terrell
authors.affiliation: 2 Division of Experimental Hematology and Oncology, Cincinnati Children's Hospital Medical Center (CCHMC), Cincinnati, Ohio.
authors.forename: Paritha
authors.initials: P
authors.lastname: Arumugam
authors.affiliation: 4 Infection, Immunity, Inflammation, and Physiological Medicine Programme, Molecular and Cellular Immunology Section, UCL Institute of Child Health , London, United Kingdom.
authors.forename: Marlene
authors.initials: M
authors.lastname: Carmo
authors.affiliation: 3 Division of Allergy/Immunology, Department of Pediatrics, CCHMC , Cincinnati, Ohio.
authors.forename: Kimberly
authors.initials: K
authors.lastname: Risma
authors.affiliation: 5 Division of Immunobiology, CCHMC , Cincinnati, Ohio.|6 Department of Bone Marrow Transplantation and Immune Deficiency, CCHMC , Cincinnati, Ohio.
authors.forename: Michael
authors.initials: M
authors.lastname: Jordan
authors.affiliation: 2 Division of Experimental Hematology and Oncology, Cincinnati Children's Hospital Medical Center (CCHMC), Cincinnati, Ohio.
authors.forename: Punam
authors.initials: P
authors.lastname: Malik
mesh_terms: D000818:Animals; D049109:Cell Proliferation / Q000235:genetics; D004195:Disease Models, Animal; D005786:Gene Expression Regulation; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000627:therapeutic use; D018380:Hematopoietic Stem Cell Transplantation; D006801:Humans; D007249:Inflammation / Q000235:genetics / Q000473:pathology / Q000628:therapy*; D007694:Killer Cells, Natural / Q000378:metabolism / Q000473:pathology; D016086:Lentivirus / Q000235:genetics; D051359:Lymphohistiocytosis, Hemophagocytic / Q000235:genetics / Q000473:pathology / Q000628:therapy*; D051379:Mice; D035683:MicroRNAs / Q000235:genetics / Q000627:therapeutic use; D009154:Mutation; D054353:Perforin / Q000235:genetics* / Q000627:therapeutic use
chemical_list: D035683:MicroRNAs; D054353:Perforin