BUFFALO,
N.Y. -- A gene therapy method that doesn't rely on
potentially toxic viruses as vectors may be growing
closer as the result of in vitro research results
reported by University at Buffalo scientists in the
current online issue of the Proceedings of the National
Academy of Sciences.
The paper, which describes the successful uptake of
a fluorescent gene by cells using novel nanoparticles
developed as DNA carriers at UB, demonstrates that
the nanoparticles ultimately may prove an efficient
and desirable alternative vector to viruses.
Using confocal microscopy
and fluorescent spectroscopy, the UB scientists tracked
optically in real-time the process known as transfection,
including the delivery of genes into cells, the uptake
of genes by the nucleus and their expression.
"We have shown
that using photonics, the gene-therapy transfer can
be monitored, tracking how the nanoparticle penetrates
the cell and releases its DNA in the nucleus,"
explained Paras N. Prasad, Ph.D., executive director
of the UB Institute for Lasers, Photonics and Biophotonics,
SUNY Distinguished Professor in the Department of
Chemistry in the University at Buffalo's College of
Arts and Sciences, and a co-author of the paper.
"When the fluorescent
protein was produced in the cell, we knew transfection
had occurred," he said.
The work is important
in light of the difficulties that have plagued gene-therapy
human trials in recent years, including some fatalities
that may have resulted from the use of viral vectors.
"Efficient delivery
of the desired gene and substantial release inside
the cell is the major hurdle in gene therapy,"
explained Dhruba J. Bharali, Ph.D., a co-author and
postdoctoral researcher in the UB Department of Chemistry
and UB's Institute for Lasers, Photonics and Biophotonics,
where the work was done.
"Viruses have
been used as efficient delivery vectors due to their
ability to penetrate cells, but there is the chance
they can revert back to 'wild' type," he said.
While non-viral vectors
are safer, he noted that it is much more difficult
to get them into cells and then to achieve the release
of DNA once they do penetrate cells.
The advantage of the
UB team's approach, he explained, is that unlike most
other nonviral vectors, the DNA-nanoparticle complex
releases its DNA before it can be destroyed by the
cell's defense system, boosting transfection significantly.
The UB scientists
also were able to use photonic methods to provide
an unprecedented look at how transfection occurs,
from the efficient uptake of nanoparticles in the
cytoplasm to their delivery of DNA to the nucleus.
"No gene-delivery
vehicle -- either viral or non-viral -- has never
been tracked in the cell before," explained Tymish
Y. Ohulchanskyy, Ph.D., the third co-author and post-doctoral
research scholar at the institute. "By using
our photonics approach, we can track gene delivery
step by step to optimize efficiency," he said.
The research team
makes its nanoparticles from a new class of materials:
hybrid, organically modified silicas (ORMOSIL).
"The structure
and composition of these hybrid ORMOSILs yield the
flexibility to build an extensive library of tailored
nanoparticles for efficiently targeting gene therapy
into different tissues and cell types," said
Prasad.
The UB researchers
now are collaborating on in vivo studies with colleagues
from the UB School of Medicine and Biomedical Sciences
to use their novel nanoparticles to transfect neuronal
cells in the brains of mice.
This research was
supported by the U.S. Air Force through its Defense
University Research Initiative on Nanotechnology (DURINT)
grant.
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