Gene therapy in hemophilia

Trials of gene therapy in hemophilia A and B, to determine safety and efficacy, are under way (2). Three patients with hemophilia B were treated in a phase I trial with a recombinant adenovirus-associated vector expressing human blood-coagulation factor IX (3). There was no evidence of formation of inhibitory antibodies against factor IX. In a phase I trial with a recombinant adenovirus-associated vector expressing human blood-coagulation factor IX, there was no evidence of germ-line transmission of vector sequences (3). 

Wliite GC. Gene therapy in hemophilia clinicai trials update. Thromb Haemost 2001 86 172-7. 

Fig. 3.2. Schematic of recombinant AAV dual-vector strategies for gene therapy for hemophilia A. (A) depicts the individual expression of the 5 and 3 ends of the factor VIII protein followed by heterodimerization to generate a function protein. (B) shows concatamerized 5 and 3 vectors in the correct head-to-tail orientation. Splicing of vectors to remove the intron and inverted terminal repeats leads to expression of the entire factor VIII protein from a single mature mRNA transcript. ITR, AAV inverted terminal repeat SD, splice donor SA, splice acceptor An, poly A.

Dwarki, V. J., Belloni, P., Nijjar, T., Smith, J., Couto, L., Rabier, M., Clift, S., Berns, A. and Cohen, L. K. (1995). Gene therapy for hemophilia A Production of therapeutic levels of human factor VIII in vivo in mice. Proc. Natl. Acad. Sci. USA 92, 1023-1027. 

Yao, S. N., Wilson, J. M., Nabel, E. G. et al. (1991). Expression of human factor-Ix in rat capillary endothelial-cells—Toward somatic gene-therapy for hemophilia-B. Proc. Natl. Acad. Sci. USA 88(18), 8101-8105. 

R.W. Herzog and K.A. High. 1998. Problems and prospects in gene therapy for hemophilia Curr. Opin. Hematol 5 321-326. (PubMed)... 

Wang L, Nichols TC, Read MS, Bellinger DA, Verma IM. Sustained expression of therapeutic level of factor IX in hemophilia B dogs by AAV-mediated gene therapy in liver. Mol Ther 2000 1 154-158. 

Gene therapy of hemophilia B also seems promising. Here, AAV vectors encoding a smaller but functional version of the human coagulation factor IX-gene were administered by intramuscular injection. A detectable increase in factor IX plasma concentration was observed. Even repeated AAV injections were well tolerated. 

Hortelano G, Al-Hendy A, Ofosu FA, Chang PL. Delivery of human Factor DC in mice by encapsulated recombinant myoblasts a novel approach towards allogeneic gene therapy of hemophilia B. Blood 1996 87 5095-5103. 

Most gene therapy protocols currently under way involve the replacement of a missing gene product in a cell (e.g., the replacement of clotting fitctor Vin in a hemophilia A patient). Recombinant DNA techniques (see Section I, Chapter 6 Recombinant DNA and Gene Cloning) are used to insert a normal DNA sequence into a mJw, wUdi dm carries the DNA into the patient s cells, where it suppUes a template fiir the normal gme prodnct (KgU-6-2). 

The investigators evaluated the safety of their nonviral somatic-cell gene therapy system, which they call transkaryotic implantation, in six patients with severe hemophilia. The procedure involved isolation of dermal fibroblasts from the patients upper arms. The fibroblasts were then transfected with a factor VIII genebearing plasmid. Cells that expressed factor VIII were cloned, propagated, and implanted into the patients abdomens. This technique can be considered as a less invasive form of ex vivo gene therapy. 

Gene therapy holds great promise for the treatment of many diseases (e.g., cancer, AIDS, cystic fibrosis, adenosine deaminase deficiency, cardiovascular diseases, Gaucher disease, a 1-antitrypsin deficiency, rheumatoid arthritis, and several others) (1,2). Advances in genomics and molecular biology have revealed that almost all diseases have a genetic component. In some cases, such as cystic fibrosis or hemophilia,... 

Thee future prospects for naked DNA gene therapy include clinical trials for genetic diseases (e.g., Duchenne muscular dystrophy, ischemia, hemophilia), which would be initiated in the next few years, and tail vein injections in rodents, which will become a widely used technique for rapidly testing expression vectors/gene therapy approaches (101). 

Kay, M.A., Landen, C.N., Rothenberg, S.R., et al. (1994). In vivo hepatic gene therapy complete albeit transient correction of factor IX deficiency in hemophilia dogs. Proc. Natl. Acad. Sci. U.S.A., 91, 2353-2357. 

Several protocols are routinely used in assessing the success of gene therapy vector treatment in the animal models of hemophilia and are described below. 

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