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Blood, 1 May 2004, Vol. 103, No. 9, pp. 3412-3419. Prepublished online as a Blood First Edition Paper on January 15, 2004; DOI 10.1182/blood-2003-10-3591. This Article Full Text Full Text (PDF) All Versions of this Article: 2003-10-3591v1 103/9/3412    most recent Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Rights and Permissions Citing Articles Citing Articles via HighWire Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Miao, H. Z. Articles by Pipe, S. W. Search for Related Content PubMed PubMed Citation Articles by Miao, H. Z. Articles by Pipe, S. W. Related Collections Hemostasis, Thrombosis, and Vascular Biology Gene Expression Social Bookmarking                   What's this?  Previous Article  |  Table of Contents  |  Next Article  HEMOSTASIS, THROMBOSIS, AND VASCULAR BIOLOGY Bioengineering of coagulation factor VIII for improved secretion Hongzhi Z. Miao, Nongnuch Sirachainan, Lisa Palmer, Phillip Kucab, Michael A. Cunningham, Randal J. Kaufman, and Steven W. Pipe From the Departments of Pediatrics and Biological Chemistry, Howard Hughes Medical Institute, University of Michigan Medical Center, Ann Arbor, MI; and the Department of Pediatrics, Faculty of Medicine, Ramathibodi Hospital, Bangkok, Thailand. Factor VIII (FVIII) functions as a cofactor within the intrinsic pathway of blood coagulation. Quantitative or qualitative deficiencies of FVIII result in the inherited bleeding disorder hemophilia A. Expression of FVIII (domain structure A1-A2-B-A3-C1-C2) in heterologous mammalian systems is 2 to 3 orders of magnitude less efficient compared with other proteins of similar size compromising recombinant FVIII production and gene therapy strategies. FVIII expression is limited by unstable mRNA, interaction with endoplasmic reticulum (ER) chaperones, and a requirement for facilitated ER to Golgi transport through interaction with the mannose-binding lectin LMAN1. Bioengineering strategies can overcome each of these limitations. B-domain-deleted (BDD)-FVIII yields higher mRNA levels, and targeted point mutations within the A1 domain reduce interaction with the ER chaperone immunoglobulin-binding protein. In order to increase ER to Golgi transport we engineered several asparagine-linked oligosaccharides within a short B-domain spacer within BDD-FVIII. A bioengineered FVIII incorporating all of these elements was secreted 15- to 25-fold more efficiently than full-length FVIII both in vitro and in vivo. FVIII bioengineered for improved secretion will significantly increase potential for success in gene therapy strategies for hemophilia A as well as improve recombinant FVIII production in cell culture manufacturing or transgenic animals. (Blood. 2004;103: 3412-3419) CiteULike    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this? This article has been cited by other articles: C. Vecchi, G. Montosi, K. Zhang, I. Lamberti, S. A. Duncan, R. J. Kaufman, and A. Pietrangelo ER Stress Controls Iron Metabolism Through Induction of Hepcidin Science, August 14, 2009; 325(5942): 877 - 880. [Abstract] [Full Text] [PDF] A. Ramezani and R. G. Hawley Correction of murine hemophilia A following nonmyeloablative transplantation of hematopoietic stem cells engineered to encode an enhanced human factor VIII variant using a safety-augmented retroviral vector Blood, July 16, 2009; 114(3): 526 - 534. [Abstract] [Full Text] [PDF] J. D. Malhotra, H. Miao, K. Zhang, A. Wolfson, S. Pennathur, S. W. Pipe, and R. J. Kaufman Antioxidants reduce endoplasmic reticulum stress and improve protein secretion PNAS, November 25, 2008; 105(47): 18525 - 18530. [Abstract] [Full Text] [PDF] M. Neyman, J. Gewirtz, and M. Poncz Analysis of the spatial and temporal characteristics of platelet-delivered factor VIII-based clots Blood, August 15, 2008; 112(4): 1101 - 1108. [Abstract] [Full Text] [PDF] B. Gangadharan, E. T. Parker, L. M. Ide, H. T. Spencer, and C. B. Doering High-level expression of porcine factor VIII from genetically modified bone marrow-derived stem cells Blood, May 15, 2006; 107(10): 3859 - 3864. [Abstract] [Full Text] [PDF] Y. Kang, L. Xie, D. T. Tran, C. S. Stein, M. Hickey, B. L. Davidson, and P. B. McCray Jr Persistent expression of factor VIII in vivo following nonprimate lentiviral gene transfer Blood, September 1, 2005; 106(5): 1552 - 1558. [Abstract] [Full Text] [PDF] B. Zhang, R. J. Kaufman, and D. Ginsburg LMAN1 and MCFD2 Form a Cargo Receptor Complex and Interact with Coagulation Factor VIII in the Early Secretory Pathway J. Biol. Chem., July 8, 2005; 280(27): 25881 - 25886. [Abstract] [Full Text] [PDF]

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