Long-term expression of human coagulation factor VIII and correction of hemophilia A after in vivo retroviral gene transfer in factor VIII-deficient mice

Abstract

Hemophilia A is caused by a deficiency in coagulation factor VIII (FVIII) and predisposes to spontaneous bleeding that can be life-threatening or lead to chronic disabilities. It is well suited for gene therapy because a moderate increase in plasma FVIII concentration has therapeutic effects. Improved retroviral vectors expressing high levels of human FVIII were pseudotyped with the vesicular stomatitis virus G glycoprotein, were concentrated to hightiters (10 9 -10 10 colony-forming units͞ml), and were injected intravenously into newborn, FVIII-deficient mice. High-levels (>200 milliunits͞ml) of functional human FVIII production could be detected in 6 of the 13 animals, 4 of which expressed physiologic or higher levels (500-12,500 milliunits͞ml). Five of the six expressers produced FVIII and survived an otherwise lethal tail-clipping, demonstrating phenotypic correction of the bleeding disorder. FVIII expression was sustained for >14 months. Gene transfer occurred into liver, spleen, and lungs with predominant FVIII mRNA expression in the liver. Six of the seven animals with transient or no detectable human FVIII developed FVIII inhibitors (7-350 Bethesda units͞ml). These findings indicate that a genetic disease can be corrected by in vivo gene therapy using retroviral vectors.

Dangerous liaisons: the role of “danger” signals in the immune response to gene therapy

“…Although many of these viruses have not been shown to cause human disease or significant toxicity, which may explain their partial success in mediating long-term transgene expression, their use has still been limited, in part, by host immune responses. 1,[61][62][63][64][65] Human immunodeficiency virus (HIV)-based lentiviral vectors are a class of retroviral vectors capable of infecting and integrating into dividing and nondividing tissues. 66 Experiments were performed in which portal vein injections of a lentiviral vector containing a human factor VIII (hFVIII) cDNA were administered to C57Bl/6 mice following a partial hepatectomy.…”

Transient blockade of the inducible costimulator pathway generates long-term tolerance to factor VIII after nonviral gene transfer into hemophilia A mice

“…In most preclinical experiments using immunocompetent hemophilia A murine and canine models, strong immune responses against FVIII after gene therapy have completely inhibited circulating FVIII activity and thus subverted the effect of gene therapy. [2][3][4][5]8,9,[14][15][16] Recent gene transfer studies 1,5,9,[17][18][19][20] indicate that the risk of transgene-specific immune responses depends on multiple factors, including the type and dose of the vector, the expression cassette and tissue specificity of the promoter, the type and level of transgene expression, route of administration, transduced cell type, and the age and the underlying mutation of the gene therapy model. Some of these factors have been extensively reviewed.…”

“…However, immune responses against exogenously introduced transgene products occur in many gene therapy model systems. Whereas transgene induced immune responses primarily comprise humoral responses, [1][2][3][4][5] cytotoxic lymphocytes (CTLs) may also be induced in the presence of other strong signals, such as viral vector components. [6][7][8][9][10] Transgene-specific antibodies and/or CTLs can significantly reduce or eliminate functional transgene products and/or transduced cells.…”

Cell Culture Processes for the Production of Viral Vectors for Gene Therapy Purposes

“…Therefore, the actual viral dose may be 10-1000 times greater (Braas et al 1996;Lyddiat and O'Sullivan 1998). It was subsequently shown empirically that titers >5 · 10 7 IP ml À1 of infusion product can efficiently obtain in vivo gene transfer in animal models of hemophilia, arthritis, cancer, chronic HBV infection, cystic fibrosis and other diseases (Ghivizzani et al 1997;Karavodin et al 1998;Sallberg et al 1998;Wang et al 1998;Nemunaitis et al 1999;Van den Driessche et al 1999). This still leaves a gap between the number of viral particles required and the number that can be produced.…”