Ultrasound enhances gene delivery of human factor IX plasmid.
Delivery of plasmid DNA can be enhanced by treatment with ultrasound (US); acoustic cavitation appears to play an important role in the process. Ultrasound contrast agents (UCAs; stabilized microbubbles) nucleate acoustic cavitation, and lower the acoustic pressure threshold for inertial cavitation occurrence. Fifty micrograms of a liver-specific, high-expressing human factor IX plasmid, pBS-HCRHP-FIXIA, mixed with UCA or phosphate-buffered saline was delivered to mouse livers by intrahepatic injection, with simultaneous exposure to 1 MHz-pulsed US using various acoustic protocols. Variable pulse duration (PD) at constant treatment time, pulse repetition frequency, and an acoustic peak negative pressure amplitude of 1.8 MPa produced 2- to 13-fold enhancements in hFIX gene expression, but PD was not a strong determinant. In contrast, a dose-response relationship was demonstrated for the peak negative pressure (P-), with significant enhancement of gene transduction at P- >/= 2 MPa. Up to 63 ng/ml (approaching the therapeutic range for treating hemophilia patients) could be achieved by transducing one liver lobe at 4-MPa P-, corresponding to a 66- fold increment relative to treatment with naked DNA alone. Under the same conditions, mouse livers could also be transduced with a GFP plasmid. Histology showed transient liver damage caused by intrahepatic injection and US exposure at 4-MPa P-; however, the damage was repaired in a few days. We conclude that therapeutic US in combination with UCA has the potential to promote safe and efficient nonviral gene transfer of hFIX for the treatment of hemophilia.
Miao
Carol H
CH
Department of Pediatrics, University of Washington and Children's Hospital and Regional Medical Center, Seattle, WA 98195, USA. miao@u.washington.edu
Brayman
Andrew A
AA
Loeb
Keith R
KR
Ye
Peiqing
P
Zhou
Ling
L
Mourad
Pierre
P
Crum
Lawrence A
LA
D000410:Alanine Transaminase / Q000378:metabolism; D000818:Animals; D001219:Aspartate Aminotransferases / Q000378:metabolism; D005164:Factor IX / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics; D006467:Hemophilia A / Q000235:genetics / Q000628:therapy; D008099:Liver / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D045423:Microbubbles; D010957:Plasmids; D014465:Ultrasonics
D049452:Green Fluorescent Proteins; D005164:Factor IX; D001219:Aspartate Aminotransferases; D000410:Alanine Transaminase