Ultrasound opens cell membranes for drug delivery
12 September 2006
Ultrasound can open holes in cell walls large enough to allow large drug
molecules to pass through, and the cells can repair the holes within
minutes, according to research carried out by the Georgia Institute of
Technology and Emory University.
Understanding this mechanism could advance the use of ultrasound for
delivering gene therapies, targeting chemotherapy and administering
large-molecule drugs that cannot readily move through cell membranes.
electron micrograph showing a prostate cancer cell immediately after
exposure to ultrasound. Image has been color enhanced to show to the
spot where the cell membrane has been removed. Image courtesy of
Robyn Schlicher, Robert Apkarian and Mark Baran.
Using five different microscopy techniques, the researchers showed that
the violent collapse of bubbles created by the ultrasound opens holes in the
membranes of cells suspended in a liquid medium. The holes, which are closed
by the cells in a matter of minutes, allow entry of therapeutic molecules as
large as 50 nanometers in diameter — larger than most proteins and similar
in size to the DNA used for gene therapy.
“The holes are made by mechanical interaction with the collapsing
bubbles,” said Mark Prausnitz, a professor in the School of Chemical and
Biomolecular Engineering at the Georgia Institute of Technology. “The
bubbles oscillate in the ultrasound field and collapse, causing a shock wave
to be released. Fluid movement associated with the resulting shock wave
opens holes in the cell membranes, which allow molecules from the outside to
enter. The cells then respond to the creation of the holes by mobilizing
intracellular vesicles to patch the holes within minutes.”
micrograph showing a prostate cancer cell immediately after exposure
to ultrasound. Image has been color enhanced to show to the spot
where the cell membrane has been removed. Image courtesy of Robyn
Schlicher, Robert Apkarian and Mark Baran.
The research was reported in the journal Ultrasound in Medicine and
(Vol. 32, No. 6) and was supported by the National Institutes of Health
(NIH) and the National Science Foundation (NSF).
Ultrasound is the same type of energy already widely used for diagnostic
imaging. Drug delivery employs higher power levels and different
frequencies, and bubbles may be introduced to enhance the effect.
Ultrasound drug delivery could be particularly attractive for gene
therapy, which has successfully used viruses to insert genetic material into
cells — but with side effects. It could also be used for more targeted
delivery of chemotherapy agents.
“One of the great benefits of ultrasound is that it is noninvasive,”
Prausnitz said. “You could give a chemotherapeutic drug locally or
throughout the body, then focus the ultrasound only on areas where tumours
exist. That would increase the cell permeability and drug uptake only in the
targeted cells and avoid affecting healthy cells elsewhere.”
Researchers have only recently found that the application of ultrasound
can help move drugs into cells by increasing the permeability of cell
membranes. It had been hypothesized, but not definitively shown, that the
ultrasound increased the permeability by opening holes in cell membranes.
Prausnitz and Robyn Schlicher use a confocal microscope to study
cells whose membranes have been opened by the application of
ultrasound. Photo: Georgia Tech, Gary Meek.
Prausnitz and collaborators Robyn Schlicher, Harish Radhakrisha, Timothy
Tolentino, Vladimir Zarnitsyn of Georgia Tech and Robert Apkarian (now
deceased) of Emory University set out to study the phenomenon in detail
using a line of prostate cancer cells. They used scanning and transmission
electron microscopy of fixed cells and two types of optical microscopy of
living cells to assess ultrasound effects and cell responses.
Beyond demonstrating that ultrasound punched holes in cell membranes, the
researchers also studied the mechanism by which cells repair the holes.
After the ultrasound exposure, they introduced into the cell medium a
chemical not normally taken up by the cells. By varying when the chemical
was introduced, they were able to determine that most of the cells had
repaired their membranes within minutes.
Though the researchers used prostate cancer cells in the study reported
in the journal, they have also studied other types of cells and believe
ultrasound offers a general way to briefly create openings in many classes
Researchers face a number of challenges, including FDA approval, before
ultrasound can be used to deliver drugs in humans. For example, the effects
of the ultrasound were not consistent across the entire volume of cells,
with only about a third affected. Researchers will also have to address
safety concerns and optimize the creation of collapsing bubbles – a
phenomenon known as cavitation – within bodily tissues.
“Before we can use ultrasound for therapy in the body, we will have to
learn how to control the exposure,” Prausnitz noted. “If we can properly
design the impact that ultrasound makes on a cell, we can generate an impact
that the cell can deal with. We want just enough impact to allow transport
into the cell, but not so much of an impact that the cell would be stressed
beyond its ability to repair the injury.”
Researchers don’t yet know if the membrane holes cause long-term harm to
the affected cells. General assays show that cells survive after resealing
the membrane holes, but detailed studies of cell behavior are still needed.
Evidence from other researchers suggests that cell membranes are frequently
damaged and repaired inside the body – without long-term ill effects. That
suggests cells may similarly tolerate ultrasound’s effects.
“One of the real challenges is going to be translating the successes that
have occurred in the laboratory and in small animals into clinical success
in people,” said Prausnitz. “Now that we better understand the mechanism of
ultrasound’s effects, we can more effectively take advantage of it for