Hepatocyte Based Insulin Gene Therapy For Diabetes

Abstract

A method and vectors for controlling blood glucose levels in a mammal are disclosed. In one embodiment, the method comprises the steps of: treating the hepatocyte cells of a patient with a first, second or third vector, wherein the first vector comprises a promoter enhancer, glucose inducible regulatory elements, a liver-specific promoter, a gene encoding human insulin with modified peptidase and an albumin 3′UTR and lacks an HGH intron, wherein the second vector comprises an HGH intron, glucose inducible regulatory elements, a liver-specific promoter, a gene encoding human insulin with modified peptidase site and an albumin 3′UTR and lacks a promoter enhancer, wherein the third vector comprises an HGH intron, glucose inducible regulatory elements, a liver-specific promoter, a gene encoding human insulin with modified peptidase site, an albumin 3′UTR and a promoter enhancer and observing the patient's insulin levels, wherein the patient's insulin levels are controlled.

1 . A method for obtaining glucose-regulated expression of insulin ex vivo in hepatocyte cells, wherein the method comprises delivering a first, second or third genetic vector for glucose-regulated synthesis of insulin into an isolated hepatocyte cell,
 wherein the first vector comprises a promoter enhancer, 1-5 glucose inducible regulatory elements, a liver-specific promoter, a gene encoding insulin with modified peptidase sites and an albumin 3′UTR and lacks an HGH intron,   wherein the second vector comprises an HGH intron, glucose inducible regulatory elements, a liver-specific promoter, a gene encoding insulin with modified peptidase sites and an albumin 3′UTR and lacks a promoter enhancer,   wherein the third vector comprises an HGH intron, glucose inducible regulatory elements, a liver-specific promoter, a gene encoding insulin with modified peptidase sites, an albumin 3′UTR and a promoter enhancer, and   wherein glucose-regulated expression of insulin occurs.   
     
     
         2 . The method of  claim 1  additionally comprising the step of transplanting the hepatocytes back into a mammal. 
     
     
         3 . The method of  claim 1 , wherein the genetic vector is delivered by exposing the cells to a virus infective for the cells, wherein the virus comprises the genetic construct, and whereby at least a portion of the cells are infected by the virus under suitable conditions and at a sufficient multiplicity. 
     
     
         4 . The method of  claim 1  wherein the mammal is human. 
     
     
         5 . The method of  claim 1  wherein the insulin is human insulin. 
     
     
         6 . A vector suitable for controlling blood glucose levels in a mammal,
 wherein the vector comprises a promoter enhancer, glucose inducible regulatory elements, a liver-specific promoter, a gene encoding insulin with modified peptidase sites and an albumin 3′UTR and lacks an HGH intron or   wherein the vector comprises an HGH intron, glucose inducible regulatory elements, a liver-specific promoter, a gene encoding insulin with modified peptidase sites and an albumin 3′UTR and lacks a promoter enhancer or   wherein the vector comprises an HGH intron, glucose inducible regulatory elements, a liver-specific promoter, a gene encoding human insulin with modified peptidase sites, an albumin 3′UTR and a promoter enhancer.   
     
     
         7 . The vector of  claim 6  wherein additionally comprising a VEGF-Enhancer. 
     
     
         8 . The vector of  claim 6  wherein the promoter is an albumin promoter. 
     
     
         9 . The vector of  claim 6  wherein the insulin is human insulin. 
     
     
         10 . A method of controlling blood glucose levels in a mammal, comprising the steps of:
 treating a mammal with a first, second or third vector,   wherein the first vector comprises a promoter enhancer, 1-5 glucose inducible regulatory elements, a liver-specific promoter, a gene encoding insulin with modified peptidase sites and an albumin 3′UTR and lacks an HGH intron and   wherein the second vector comprises an HGH intron, glucose inducible regulatory elements, a liver-specific promoter, a gene encoding insulin with modified peptidase sites and an albumin 3′UTR and lacks a promoter enhancer,   wherein the third vector comprises an HGH intron, glucose inducible regulatory elements, a liver-specific promoter, a gene encoding insulin with modified peptidase sites and an albumin 3′UTR and a promoter enhancer, and   observing the mammal's insulin levels, wherein the mammal's insulin levels are controlled.   
     
     
         11 . The method of  claim 1  wherein the vector is in a minicircle format. 
     
     
         12 . The method of  claim 10  wherein the vector is in a minicircle format. 
     
     
         13 . The method of  claim 10  wherein the mammal is human. 
     
     
         14 . The method of  claim 13  wherein the insulin is human insulin. 
     
     
         15 . The method of  claim 10  wherein the mammal's cholesterol level decreases after treatment. 
     
     
         16 . The method of  claim 10  wherein the mammal's triglyceride level decreases after treatment. 
     
     
         17 . The method of  claim 10  wherein the mammal is a cat. 
     
     
         18 . The method of  claim 10  wherein the mammal is a dog. 
     
     
         19 . The method of  claim 10  wherein the mammal is selected for the group consisting of hamsters, gerbils, rats, mice, rabbits, guinea pigs, chinchillas and ferrets. 
     
     
         20 . The method of  claim 10  wherein the mammal is a non-human mammal. 
     
     
         21 . The method of  claim 10  wherein the patient has a decrease in the plasma level of a compound selected from the group of AST, ALT, and alkaline phosphatase after treatment.