Alteplase stability and bioactivity after low-power ultrasonic energy delivery with the OmniSonics resolution system.
OBJECTIVE
Low-power ultrasonic (US) energy is capable of clot dissolution in vivo. The combination of US energy plus alteplase may further accelerate clot lysis; however, the effects of cavitation could potentially denature and inactivate the lytic protein. The purpose of this study was to determine the bioactivity and stability of alteplase when exposed to US energy with use of a novel intravascular US wire in an in vitro model.


METHODS
The model consisted of a 6.4-mm-diameter silicone tube closed at one end and filled with alteplase (1 mg/mL) in a water bath (37 degrees C). A 95-cm US wire (0.025-inch diameter, 20 kHz) was inserted into the tube and connected to a variable power generator. The wire delivers low-power acoustic energy 360 degrees around its 20-cm active length and was irrigated by a continuous infusion of purified water. Fresh 6-mL alteplase aliquots were exposed to US energy and tested in duplicates. Zero (control), 1 W, or 2 W of energy was delivered to individual test samples for zero (control), 0.5, 3, or 6 minutes. Alteplase samples were assayed for optical clarity and protein concentration with use of UV spectrophotometry, for percent protein monomer with use of high-performance size-exclusion chromatography, and for in vitro clot lysis activity.


RESULTS
In the control samples, optical clarity was clear or colorless in all samples; protein concentration was 1.02 mg/mL +/- 0; protein monomer was 98%; and clot lysis activity was 108% per mg +/- 1. In the test samples, optical clarity was clear or colorless in all samples; protein concentrations at 0.5, 3, and 6 minutes were 0.98 mg/mL +/- 0.02, 0.93 mg/mL +/- 0.01, and 0.86 mg/mL +/- 0.02, respectively, at 1 W, and 1.00 mg/mL +/- 0.03, 0.94 mg/mL +/- 0.10, and 0.84 mg/mL +/- 0.17, respectively, at 2 W. Protein monomer was 98% for all samples. Clot lysis activity levels at 0.5, 3, and 6 minutes were 111% per mg +/- 1, 110% per mg +/- 1, and 115% per mg +/- 1, respectively, at 1 W, and 110% per mg +/- 0, 111% per mg +/- 1, and 116% per mg +/- 2, respectively, at 2 W.


CONCLUSIONS
Alteplase solutions exposed to low-power US energy for as long as 6 minutes remained fully active and stable as determined by protein assays. Further investigation is warranted with use of combinations of US energy and alteplase.
Smikahl
John
J
Pharmaceutical Research and Development, Genetech, South San Francisco, CA 94080, USA.
Yeung
Doug
D
Wang
Sharon
S
Semba
Charles P
CP
D001681:Biological Assay; D002404:Catheterization; D016503:Drug Delivery Systems; D004355:Drug Stability; D004867:Equipment Design; D066298:In Vitro Techniques; D012996:Solutions; D010959:Tissue Plasminogen Activator / Q000008:administration & dosage / Q000737:chemistry*; D014464:Ultrasonic Therapy / Q000295:instrumentation*
D012996:Solutions; D010959:Tissue Plasminogen Activator