High-level protein production in erythroid cells derived from in vivo transduced hematopoietic stem cells

• Authors: Hongjie Wang, University of Washington; Zhinan Liu, University of Washington; Chang Li, University of Washington; Sucheol Gil, University of Washington; Thalia Papayannopoulou, University of Washington; Christopher Doering, Emory University; Andre Lieber, University of Washington
• Journal Title: Blood Advancer
• Volume: Volume 3, Number 19
• Publisher: American Society of Hematology | 2019-10-08, Pages 2883-2894
• Type of Work: Article | Final Publisher PDF
• Publisher DOI: http://doi.org/10.1182/bloodadvances.2019000706
• Subjects: Biology, Genetics; Health Sciences, Pharmacology
• Keywords: Science & Technology; Life Sciences & Biomedicine; Hematology; GENE-THERAPY; EXPRESSION; VECTORS; DEFICIENCY; MICE

Abstract

We developed an in vivo hematopoietic stem cell (HSC) transduction approach that involves HSC mobilization from the bone marrow into the peripheral bloodstream and the IV injection of an integrating, helper-dependent adenovirus (HDAd5/3511) vector system. HDAd5/3511 vectors target human CD46, a receptor that is abundantly expressed on primitive HSCs. Transgene integration is achieved by a hyperactive Sleeping Beauty transposase (SB100x) and transgene marking in peripheral blood cells can be increased by in vivo selection. Here we directed transgene expression to HSC-derived erythroid cells using b-globin regulatory elements. We hypothesized that the abundance and systemic distribution of erythroid cells can be harnessed for high-level production of therapeutic proteins. We first demonstrated that our approach allowed for sustained, erythroid-lineage specific GFP expression and accumulation of GFP protein in erythrocytes. Furthermore, after in vivo HSC transduction/selection in hCD46-transgenic mice, we demonstrated stable supraphysiological plasma concentrations of a bioengineered human factor VIII, termed ET3. High-level ET3 production in erythroid cells did not affect erythropoiesis. A phenotypic correction of bleeding was observed after in vivo HSC transduction of hCD461/1/F82/2 hemophilia A mice despite high plasma anti-ET3 antibody titers. This suggests that ET3 levels were high enough to provide sufficient noninhibited ET3 systemically and/or locally (in blood clots) to control bleeding. In addition to its relevance for hemophilia A gene therapy, our approach has implications for the therapy of other inherited or acquired diseases that require high levels of therapeutic proteins in the blood circulation.