Gut Epithelial Cells as Targets for Gene Therapy of Hemophilia

Abstract

Gut epithelium is an attractive target for gene therapy of hemophilia due to the large number of rapidly dividing cells that should be readily accessible to a wide range of vectors by a noninvasive route of administration. We have performed in vitro tests to determine the suitability of gut epithelial cells for gene transfer, protein synthesis, and secretion of coagulation factors VIII and IX. The results with retroviral vectors indicate that transduced epithelial cells from human, rat, or porcine small or large intestine can synthesize significant amounts of factor VIII or factor IX and that two-thirds or more of the recombinant protein is secreted in a basolateral direction (i.e., away from the lumen and toward underlying capillaries and lymphatics). Furthermore, we have demonstrated that intestinal epithelial cells are susceptible to efficient gene transfer by lipofection and adenovirus vectors. In the case of factor IX, we have produced a high-titer adenovirus vector capable of transducing gut epithelial cells resulting in synthesis of factor IX. The results of our in vitro studies indicate that gene transfer targeting gut epithelium as a new approach to hemophilia gene therapy is rational and merits in vivo studies in hemophilia animal models.

Overview summary

Lozier et al. describe in vitro studies in which they investigate the feasibility of gene transfer to gut epithelial cells as a means of gene therapy of hemophilia A and B. In this report they demonstrate that small and large intestinal epithelial cell lines from various species are capable of synthesizing factor VIII and factor IX at rates comparable to other cell types previously tested. Using confluent epithelial cells in an in vitro model of gut epithelium, they have shown that the recombinant proteins are secreted by cells in a basolateral direction which is necessary for delivery of clotting factors into the circulation. Due to previous reports of poor efficiency of retroviral transduction of intestinal epithelium in vivo, transduction of gut epithelial cells with adenoviruses and transfection of gut epithelial cells with liposomes were investigated and shown to be capable of efficient gene transfer in vitro. On the basis of cell numbers, efficiency of gene transfer, rate of protein synthesis, and distribution of coagulant proteins in plasma, it can be seen that the gut epithelium is a rational target for gene transfer to treat hemophilia or other plasma protein deficiencies. Gene transfer by orally administered vectors might offer a noninvasive method for therapy that would be advantageous to patients with hemophilia.