H U M A N G E N E THERAPY 2:113 (1991) Mary Ann Liebert, Inc., Publishers
M E E T I N G PRESENTATION
Correction o f M u r i n e M u c o p o l y s a c c h a r i d o s i s T y p e V I I ( M P S V I I ) b y B o n e M a r r o w Transplantation a n d G e n e Transfer T h e r a p y
EDWARD H. BIRKENMEIER
T H E gusmpsIgusmps mouse has no P-glucuronidase activity and develops murine mucopolysaccharidosis type VII ( M P S VII). The clinical and pathologic abnormalities are similar to those found in humans with severe M P S VII. Mutant mice are dysmorphic, dwarfed, and have a shortened life span. Pathologic findings include widespread lysosomal storage. Genetic analysis showed that the mutation is inherited as an autosomal recessive that maps to the (^-glucuronidase gene complex (Gus) on the distal end of chromosome 5. Molecular studies comparing the mutant and normal P-glucuronidase genes revealed a 1-bp deletion in exon 10 ofthe mutant gene. This deletion changes the reading frame and introduces a stop codon eight codons downstream. Since M P S VII mice have a well-defined genetic background, they provide an attractive animal model to test potential therapies for lysosomal storage diseases. Our initial studies described the consequences of introducing the human P-glucoronidase gene, G U S B , into gusmpsIgusmps mice. Transgenic mice homozygous for the mucopolysaccharidosis VII mutation expressed high levels of human p-glucuronidase activity in all tissues examined and were phenotypically normal. Biochemically, both the intralysosomal storage of glycosaminoglycans and the secondary elevation of other acid hydrolases were corrected. These findings demonstrated that the G U S B transgene is mucopolysaccharidosis storage disease. T o determine whether bone marrow transplantation ( B M T ) also corrects these abnormalities, genetically identical mutant animals were given syngeneic bone marrow transplants using cells from + / + mice. Initial experiments showed that levels of P-glucuronidase activity in recipient tissues correlated with the amount of radiation given before B M T . T w o groups of mice given B M T therapy were followed for periods of 1 and 2 years, respectively. These mice were evaluated using a combination of
clinical, biochemical, histochemical, and pathologic analyses. Spleen, liver, cornea, and glomerular mesangial cells showed essentially complete correction at all radiation doses. Storage was partially corrected in meninges and perivascular cells in brain, and in renal tubular epithelial cells at the higher radiation doses. Life span in B M T treated animals was increased approximately threefold, approaching that seen in normal mice after B M T . These results support the position that B M T has a place in the therapeutic regimen for mucopolysaccharidoses. Our future plans are to use recombinant retroviruses carrying P-glucuronidase c D N A s to infect bone marrow cells from M P S VII mice. Preliminary experiments used an N 2 retroviral vector containing a rat p-glucuronidase c D N A driven by a thymidine kinase promoter ( N T K - B G E O - 1 ) to infect normal bone marrow enriched for hematopoietic stem cells. Over 8 0 % ofthe C F U - S in irradiated hosts showed presence ofthe virus as determined by P C R analysis. Parallel experiments using infected marrow cells from M P S VII mice showed low levels of P-glucuronidase activity in the C F U - S of irradiated mutant hosts. Secondary recipients showed low levels of P-glucuronidase in peripheral W B C for at least 6 months following bone marrow transplantation and at 10 months of age were killed for detailed pathologic and biochemical analyses. Our long-term plan is to continue the retroviral-mediated gene transfer experiments in M P S VII mice using direct copy (DC) recombinant viruses carrying the human P-glucuronidase gene driven by its o w n promoter.
The Jackson Laboratory, Bar Harbor, M E 04609.
113
Address reprint requests to: Dr. Edward H . Birkenmeier The Jackson Laboratory 600 Main Street Bar Harbor, M E 04609
M E E T I N G PRESENTATION
Correction o f M u r i n e M u c o p o l y s a c c h a r i d o s i s T y p e V I I ( M P S V I I ) b y B o n e M a r r o w Transplantation a n d G e n e Transfer T h e r a p y
EDWARD H. BIRKENMEIER
T H E gusmpsIgusmps mouse has no P-glucuronidase activity and develops murine mucopolysaccharidosis type VII ( M P S VII). The clinical and pathologic abnormalities are similar to those found in humans with severe M P S VII. Mutant mice are dysmorphic, dwarfed, and have a shortened life span. Pathologic findings include widespread lysosomal storage. Genetic analysis showed that the mutation is inherited as an autosomal recessive that maps to the (^-glucuronidase gene complex (Gus) on the distal end of chromosome 5. Molecular studies comparing the mutant and normal P-glucuronidase genes revealed a 1-bp deletion in exon 10 ofthe mutant gene. This deletion changes the reading frame and introduces a stop codon eight codons downstream. Since M P S VII mice have a well-defined genetic background, they provide an attractive animal model to test potential therapies for lysosomal storage diseases. Our initial studies described the consequences of introducing the human P-glucoronidase gene, G U S B , into gusmpsIgusmps mice. Transgenic mice homozygous for the mucopolysaccharidosis VII mutation expressed high levels of human p-glucuronidase activity in all tissues examined and were phenotypically normal. Biochemically, both the intralysosomal storage of glycosaminoglycans and the secondary elevation of other acid hydrolases were corrected. These findings demonstrated that the G U S B transgene is mucopolysaccharidosis storage disease. T o determine whether bone marrow transplantation ( B M T ) also corrects these abnormalities, genetically identical mutant animals were given syngeneic bone marrow transplants using cells from + / + mice. Initial experiments showed that levels of P-glucuronidase activity in recipient tissues correlated with the amount of radiation given before B M T . T w o groups of mice given B M T therapy were followed for periods of 1 and 2 years, respectively. These mice were evaluated using a combination of
clinical, biochemical, histochemical, and pathologic analyses. Spleen, liver, cornea, and glomerular mesangial cells showed essentially complete correction at all radiation doses. Storage was partially corrected in meninges and perivascular cells in brain, and in renal tubular epithelial cells at the higher radiation doses. Life span in B M T treated animals was increased approximately threefold, approaching that seen in normal mice after B M T . These results support the position that B M T has a place in the therapeutic regimen for mucopolysaccharidoses. Our future plans are to use recombinant retroviruses carrying P-glucuronidase c D N A s to infect bone marrow cells from M P S VII mice. Preliminary experiments used an N 2 retroviral vector containing a rat p-glucuronidase c D N A driven by a thymidine kinase promoter ( N T K - B G E O - 1 ) to infect normal bone marrow enriched for hematopoietic stem cells. Over 8 0 % ofthe C F U - S in irradiated hosts showed presence ofthe virus as determined by P C R analysis. Parallel experiments using infected marrow cells from M P S VII mice showed low levels of P-glucuronidase activity in the C F U - S of irradiated mutant hosts. Secondary recipients showed low levels of P-glucuronidase in peripheral W B C for at least 6 months following bone marrow transplantation and at 10 months of age were killed for detailed pathologic and biochemical analyses. Our long-term plan is to continue the retroviral-mediated gene transfer experiments in M P S VII mice using direct copy (DC) recombinant viruses carrying the human P-glucuronidase gene driven by its o w n promoter.
The Jackson Laboratory, Bar Harbor, M E 04609.
113
Address reprint requests to: Dr. Edward H . Birkenmeier The Jackson Laboratory 600 Main Street Bar Harbor, M E 04609
