Biologic agents and methods of use
Disclosed herein are nucleic acid compositions and methods of use. The nucleic acid compositions may have a therapeutic nucleic acid sequence operably linked to a nuclear targeting sequence that increases expression of the therapeutic nucleic acid in a cell by at least 1.25 fold; at least 80% sequence identity to SEQ ID NO: 6; or a first regulatory element comprising a promoter sequence operably linked to a hemoglobin subunit gamma intron (hBGi) sequence, and a second regulatory element comprising a woodchuck hepatitis posttranscriptional regulatory element (WPRE) sequence. The nucleic acid compositions with these features may enhance therapeutic nucleic acid transfection and expression. Also disclosed herein are transgenes optimized for gene therapy applications, including novel FVIII transgene sequences, in which the B domain may be non-naturally occurring the A1 and/or A3 domain may include at least one amino acid substitution.
1 . A nucleic acid composition comprising a nucleic acid sequence of SEQ ID NO: 6.
2 . The nucleic acid composition of claim 1 , further comprising a therapeutic nucleic acid sequence, wherein the nucleic acid sequence of SEQ ID NO: 6 increases expression of the therapeutic nucleic acid sequence in a cell by at least 1.25-fold as compared to an otherwise identical control composition lacking the nucleic acid sequence of SEQ ID NO: 6.
3 . The nucleic acid composition of claim 1 , wherein the nucleic acid sequence of SEQ ID NO: 6 is a nuclear targeting sequence.
4 . The nucleic acid composition of claim 1 , wherein the nucleic acid composition is an isolated nucleic acid molecule.
5 . The nucleic acid composition of claim 1 , wherein the nucleic acid composition further comprises one or more Inverted Terminal Repeat (ITR) sequences.
6 . The nucleic acid composition of claim 1 , wherein the nucleic acid composition is a non-viral vector.
7 . A kit comprising a sonoactive agent and a nucleic acid composition, wherein the nucleic acid composition comprises a nucleic acid sequence of SEQ ID NO: 6.
8 . The kit of claim 7 , wherein the nucleic acid composition further comprises a therapeutic nucleic acid sequence, wherein the nucleic acid sequence of SEQ ID NO: 6 increases expression of the therapeutic nucleic acid sequence in a cell by at least 1.25-fold as compared to an otherwise identical control composition lacking the nucleic acid sequence of SEQ ID NO: 6.
9 . The kit of claim 7 , wherein the nucleic acid sequence of SEQ ID NO: 6 is a nuclear targeting sequence.
10 . The kit of claim 7 , wherein the nucleic acid composition is an isolated nucleic acid molecule.
11 . The kit of claim 7 , wherein the nucleic acid composition further comprises one or more Inverted Terminal Repeat (ITR) sequences.
12 . The kit of claim 7 , wherein the nucleic acid composition is a non-viral vector.
13 . The kit of claim 7 , wherein the sonoactive agent comprises a shell filled with a perfluorinated gas.
14 . The kit of claim 7 , wherein the sonoactive agent comprises protein-stabilized microstructures.
15 . The kit of claim 7 , wherein the sonoactive agent comprises lipid-stabilized microstructures.
16 . The kit of claim 15 , wherein the lipid-stabilized microstructures comprise a lipid stabilized shell surrounding a perfluorinated gas core.
17 . The kit of claim 16 , wherein the lipid stabilized shell comprises a monomolecular membrane of hydrogenated egg yolk phosphatidyl serine, wherein the perfluorinated gas core comprises perfluorobutane gas.
18 . The kit of claim 7 , further comprising instructions for administering ultrasound acoustic energy to a subject to facilitate delivery of the nucleic acid composition to the subject.