Fumarylacetoacetate hydrolase (fah)-deficient and immunodeficient rats and uses thereof

Abstract

Described herein are rats with a hepatic deficiency comprising decreased function, activity, or expression of an enzyme in the tyrosine catabolic pathway (such as fumarylacetoacetate hydrolase), and methods of using the same for in vivo engraftment and expansion of heterologous hepatocytes, such as human hepatocytes, analysis of human liver disease, and analysis of xenobiotics. Also disclosed is the use of immunodeficient rats for the engraftment and expansion of heterologous hepatocytes.

1 . A genetically modified Fah-deficient rat whose genome is homozygous for a disruption in the Fah gene such that the disruption results in loss of expression of functional FAH protein and decreased liver function. 
     
     
         2 . (canceled) 
     
     
         3 . The Fah-deficient rat of  claim 1 , wherein the disruption is an insertion, wherein the insertion comprises a stop codon, a selectable marker, or both. 
     
     
         4 . The Fah-deficient rat of  claim 1 , wherein the disruption is in exon 3 of the Fah gene. 
     
     
         5 . The Fah-deficient rat of  claim 1 , wherein the rat is immunosuppressed. 
     
     
         6 . The Fah-deficient rat of  claim 5 , wherein the immunosuppression is the result of administration of one or more immunosuppressive agents. 
     
     
         7 . The Fah-deficient rat of  claim 6 , wherein the one or more immunosuppressive agents are selected from the group consisting of FK506, cyclosporin A, fludarabine, mycophenolate, prednisone, rapamycin, azathioprine, and combinations thereof. 
     
     
         8 . The Fah-deficient rat of  claim 5 , wherein the immunosuppression is the result of one or more genetic alterations that inhibit the development of functional immune cells. 
     
     
         9 .- 10 . (canceled) 
     
     
         11 . The Fah-deficient rat of  claim 1 , wherein the rat comprises transplanted heterologous hepatocytes. 
     
     
         12 . (canceled) 
     
     
         13 . The Fah-deficient rat of  claim 1 , wherein the rat comprises transplanted human hepatocytes. 
     
     
         14 . The Fah-deficient rat of  claim 1 , wherein the rat comprises expanded human hepatocytes, wherein liver function is restored in the rat. 
     
     
         15 . A method of expanding human hepatocytes in vivo, comprising:
 transplanting heterologous hepatocytes into a first rat, said first rat being a first Fah-deficient rat according to  claim 1 , or a first immunodeficient rat; and   allowing the heterologous hepatocytes to expand, thereby expanding heterologous hepatocytes in vivo.   
     
     
         16 . (canceled) 
     
     
         17 . The method of  claim 15 , wherein the heterologous hepatocytes transplanted into the first rat are isolated human hepatocytes. 
     
     
         18 . The method of  claim 17 , wherein the human hepatocytes are isolated from the liver of a humanized non-human mammal prior to transplanting into the first rat, wherein the non-human mammal is an FRG KO mouse. 
     
     
         19 .- 20 . (canceled) 
     
     
         21 . The method of  claim 15 , further comprising inducing acute liver damage in the first rat prior to transplanting the heterologous hepatocytes. 
     
     
         22 . The method of  claim 15 , further comprising administering a vector encoding urokinase to the first rat prior to transplanting the heterologous hepatocytes. 
     
     
         23 .- 29 . (canceled) 
     
     
         30 . The method of  claim 15 , wherein the first rat is an Fah-deficient rat, further comprising administering one or more immunosuppressive agents to the first Fah-deficient rat at least about 2 days prior to transplanting the heterologous hepatocytes. 
     
     
         31 . The method of  claim 15 , further comprising collecting the expanded heterologous hepatocytes from the first rat. 
     
     
         32 . (canceled) 
     
     
         33 . The method of claim  3132 , further comprising expanding the collected heterologous hepatocytes by serial transplantation, wherein said serial transplantation comprises:
 collecting expanded heterologous hepatocytes from the first rat;   transplanting the collected expanded heterologous hepatocytes from the first rat into a second rat, said second rat being a second Fah-deficient rat according to  claim 1 , or a second immunodeficient rat; and   allowing the heterologous hepatocytes to expand, thereby expanding heterologous hepatocytes in vivo.   
     
     
         34 .- 38 . (canceled) 
     
     
         39 . The method of  claim 15 , wherein the first rat is an immunodeficient rat, wherein the immunodeficiency of the rat is due to a genetic alteration, immunosuppression, or a combination thereof. 
     
     
         40 . The method of  claim 39 , wherein the immunodeficiency is due to one or more genetic alterations that inhibit the development of functional immune cells, wherein the one or more genetic alterations is selected from the group consisting of selected from the group consisting of Rag1 deficiency, Rag2 deficiency, Il2rg deficiency, SCID, Sirp-α hum/hum , nude, perforin −/− , and combinations thereof. 
     
     
         41 .- 54 . (canceled) 
     
     
         55 . An immunodeficient rat having a liver comprising transplanted and expanded human hepatocytes. 
     
     
         56 .- 77 . (canceled)