Treatment for restoring ureagenesis in carbamoyl phosphate synthetase 1 deficiency

A pharmaceutical composition comprising:

• a first adeno-associated viral vector comprising:
• (a) a polynucleotide sequence comprising a terminal repeat sequence;
• (b) a polynucleotide sequence comprising a promoter;
• (c) a codon optimized polynucleotide sequence (SEQ ID NO:
• 2) comprising a segment of a gene encoding a carbamoyl phosphate synthetase 1 protein;
• and
• (d) a polynucleotide sequence comprising a polyA tail signal;
• and
• a pharmaceutical excipient selected from the group consisting of:
• a preservative, a tonicity adjusting agent, a detergent, a viscosity adjusting agent, a sugar or a pH adjusting agent;
• and
• a second adeno-associated viral vector comprising:
• (a) a polynucleotide sequence comprising an inverted terminal repeat;
• (b) a polynucleotide sequence comprising a promoter;
• and
• (c) a codon optimized polynucleotide sequence comprising a segment of a gene encoding a carbamoyl phosphate synthetase 1 protein;
• wherein:
• the first and second segments of the carbamoyl phosphate synthetase 1 polynucleotides overlap so that, following adeno-associated viral vector infection of a human liver cell, the carbamoyl phosphate synthetase 1 polynucleotides concatemerize via homologous recombination so as to reconstitute a carbamoyl phosphate synthetase 1 polynucleotide that expresses the carbamoyl phosphate synthetase 1 protein in the liver cell;
• and
• the first segment of the carbamoyl phosphate synthetase 1 polynucleotide is selected to comprise the sequence ATGCCTCAGA and the second segment of the carbamoyl phosphate synthetase 1 polynucleotide is selected to comprise the sequence TGCCGTGGAG.

A method of making a pharmaceutical composition comprising combining together in an aqueous formulation:

• at least one adeno-associated viral vector comprising a carbamoyl phosphate synthetase 1 polynucleotide sequence (SEQ ID NO:
• 2);
• and
• a pharmaceutical excipient selected from the group consisting of:
• a preservative, a tonicity adjusting agent, a detergent, a viscosity adjusting agent, a sugar or a pH adjusting agent, wherein:
• the vector and the carbamoyl phosphate synthetase 1 polynucleotide sequence are selected such that when the adeno-associated viral vector infects a human liver cell, carbamoyl phosphate synthetase 1 protein is expressed;
• the method combines two adeno-associated viral vectors comprising:
• a first adeno-associated viral vector comprising a first segment of a codon optimized carbamoyl phosphate synthetase 1 polynucleotide (SEQ ID NO:
• 2);
• and
• a second adeno-associated viral vector comprising a second segment of a codon optimized carbamoyl phosphate synthetase 1 polynucleotide (SEQ ID NO:
• 2);
• wherein:
• the first adeno-associated viral vector comprising the first segment of a codon optimized carbamoyl phosphate synthetase 1 polynucleotide and the second adeno-associated viral vector comprising the second segment of a codon optimized carbamoyl phosphate synthetase 1 polynucleotide are selected so that:
• the first and second segments of the carbamoyl phosphate synthetase 1 polynucleotides overlap such that, following first and second adeno-associated viral vector infection of the human liver cell, the carbamoyl phosphate synthetase 1 polynucleotides concatemerize via homologous recombination so as to reconstitute a carbamoyl phosphate synthetase 1 gene that expresses the carbamoyl phosphate synthetase 1 protein in the liver cell;
• and
• wherein the second segment of the carbamoyl phosphate synthetase 1 polynucleotide is selected to comprise the sequence TGCCGTGGAG.