Expression of active human factor IX in mammary tissue of transgenic animals

Abstract

Recombinant Factor IX characterized by a high percentage of active protein can be obtained in the milk of transgenic animals that incorporate chimeric DNA molecules according to the present invention. Transgenic animals of the present invention are produced by introducing into developing embryos DNA that encodes Factor IX, such that the foreign DNA is stably incorporated in the DNA of germ line cells of the mature animal. Particularly efficient expression was accomplished using a chimeric construct comprising a mammary gland specific promoter, Factor IX cDNA that lacked the complete or any portion of the 5′-untranslated and 3′-untranslated region, which is substituted with a 5-′ and 3′- end of the mouse whey acidic protein gene. In vitro cell cultures of cells explanted from the transgenic mammal of the invention and methods of producing Factor IX from such said culture and methods of treating hemophilia B are also described.

What is claimed is:  
     
         1 . A method of treating a patient having hemophilia B comprising administering to said patients a hemophilia B symptom preventing or ameliorating amount of Factor IX produced by the non-human transgenic mammal and a pharmaceutically acceptable carrier.  
     
     
         2 . A method of  claim 1 , wherein said Factor IX is biologically active human Factor IX.  
     
     
         3 . A method of  claim 1 , wherein said non-human transgenic mammal is selected from the group consisting of mice, rats, rabbits, pigs, sheep, goats and cows.  
     
     
         4 . A method of  claim 3 , wherein said non-human transgenic mammal is a pig and secretes biologically active human Factor IX per milliliter of milk.  
     
     
         5 . A method of  claim 2 , wherein said biologically active human Factor IX has a specific activity that is at least about 5-200% greater than the specific activity of human Factor IX isolated from human plasma.  
     
     
         6 . A method of  claim 5 , wherein said non-human transgenic mammal is a pig.  
     
     
         7 . A method of  claim 2 , wherein said amount of said human Factor IX that is administered comprises about 18-30 IU/kg administered once a week.  
     
     
         8 . A method of  claim 2 , wherein said administration of said human Factor IX raises Factor IX levels 20% or greater in the blood.  
     
     
         9 . A biologically active human Factor IX produced in a non-human transgenic mammal, said Factor IX comprising a specific activity that is at least about 5-200% greater than the specific activity of human Factor IX isolated from human plasma.  
     
     
         10 . A biologically active human Factor IX of  claim 9 , wherein said non-human transgenic mammal is a pig.