Defining the Optimal FVIII Transgene for Placental Cell-Based Gene Therapy to Treat Hemophilia A

• Authors: Nadia El-Akabawy, Wake Forest University; Martin Rodriguez, Wake Forest University; Ritu Ramamurthy, Wake Forest University; Andrew Rabah, Wake Forest University; Brady Trevisan, Wake Forest University; Alshaimaa Morsi, Wake Forest University; Sunil George, Wake Forest University; Jordan Shields, Childrens Healthcare Atlanta; Diane Meares, Wake Forest University; Andrew Farland, Wake Forest University; Anthony Atala, Wake Forest University; Christopher Doering, Emory University; H Trent Spencer, Emory University; Christopher B. Porada, Wake Forest University; Graca Almeida-Porada, Wake Forest University
• Journal Title: Molecular Therapy: Methods & Clinical Development
• Volume: Volume 17
• Publisher: Cell Press & Elsevier | 2020-06-12, Pages 465-477
• Type of Work: Article | Final Publisher PDF
• Publisher DOI: http://doi.org/10.1016/j.omtm.2020.03.001
• Subjects: Biology, Cell; Health Sciences, Medicine and Surgery; Health Sciences, Rehabilitation and Therapy; Health Sciences, Oncology
• Keywords: Science & Technology; Life Sciences & Biomedicine; Medicine, Research & Experimental; Research & Experimental Medicine; Mesenchymal stem cells; High level expression; Porcine factor VIII; Hematopoietic stem; Amniotic fluid; HLA-E; Transplantation; Integration; Generation; Sequence

Abstract

The delivery of factor VIII (FVIII) through gene and/or cellular platforms has emerged as a promising hemophilia A treatment. Herein, we investigated the suitability of human placental cells (PLCs) as delivery vehicles for FVIII and determined an optimal FVIII transgene to produce/secrete therapeutic FVIII levels from these cells. Using three PLC cell banks we demonstrated that PLCs constitutively secreted low levels of FVIII, suggesting their suitability as a transgenic FVIII production platform. Furthermore, PLCs significantly increased FVIII secretion after transduction with a lentiviral vector (LV) encoding a myeloid codon-optimized bioengineered FVIII containing high-expression elements from porcine FVIII. Importantly, transduced PLCs did not upregulate cellular stress or innate immunity molecules, demonstrating that after transduction and FVIII production/secretion, PLCs retained low immunogenicity and cell stress. When LV encoding five different bioengineered FVIII transgenes were compared for transduction efficiency, FVIII production, and secretion, data showed that PLCs transduced with LV encoding hybrid human/porcine FVIII transgenes secreted substantially higher levels of FVIII than did LV encoding B domain-deleted human FVIII. In addition, data showed that in PLCs, myeloid codon optimization is needed to increase FVIII secretion to therapeutic levels. These studies have identified an optimal combination of FVIII transgene and cell source to achieve clinically meaningful levels of secreted FVIII.