Scientists at The Scripps Research Institute Design Gene-Tipped Tumor
By Jason Socrates Bardi
A group of researchers from The Scripps Research Institute (TSRI) have
demonstrated what, in principle, could be a new way of treating cancer
and several other diseases where angiogenesis occurs. Angiogenesis, the
formation and differentiation of new blood vessels, is a crucial process
in cancer, and, when blocked, improves a patient's prognosis.
In cancer-related angiogenesis, tumors develop their own blood supplies
by causing cells that line blood vessels to proliferate, forming new vessels
and bringing more blood to the tumor. The increased oxygen and nutrients
the tumors receive allows them to grow and enables certain "metastatic"
cells to leave the tumor, enter the bloodstream, migrate to other tissues
of the body, and establish more tumors.
In an article appearing in the latest issue of the journal Science,
the TSRI investigators combined a gene that shuts off angiogenesis with
a 50 to100 nanometer-sized particle that selectively targets the cells
that form new blood vessels in cancer tumors. This approach combines gene
delivery with specific vascular targeting thereby effectively disrupting
the blood supply of tumors without influencing the normal blood vessels
or any other tissue.
This anti-cancer nanoparticle is like a smart bomb that delivers its
multiple warhead genetic payload into endothelial cells that proliferate
during angiogenesiswhich is the medical equivalent of cutting off
all the supply routes to destroy the tumor. Once angiogenesis is stopped,
the tumor cells starve, and the tumor is ultimately destroyed.
Anti-angiogenics have been known of and studied for many years, but
this anti-cancer nanoparticle is a new type of anti-angiogenic. Unlike
other, "systemic" angiogenesis blockers, which become diffused throughout
the blood steam upon injection, the nanoparticle-targeting vehicle directs
itself to areas of the body where the tumors exist and where local vascular
cells are expanding to form new blood vessels. The nanoparticle homes
in on these cells and drops off multiple copies of a gene that effectively
blocks angiogenesis and kills tumors.
"We saw strong regression of large tumors in every system we looked
at," says TSRI Immunology Professor David Cheresh, who led the study.
How Cancer Hijacks Blood Vessels
Angiogenesis, the process where blood vessels are formed and differentiated,
is the tumor's way of responding to hypoxia, a deficiency of oxygen-rich
blood reaching its proliferating cells.
As the cells of a tumor grow too fast to be fed by their existing blood
supply, they are starved of oxygen. They respond by exhibiting a hypoxic
responseturning on genes that produce growth factor proteins, which
tell the body to make new local blood vessels and bring more oxygen-rich
The body responds when endothelial cellsthe major cell type lining
the vasculaturereceive the growth factors and begin to proliferate,
producing more blood vessels. Once these new blood vessels are made, they
bring vital oxygen-rich blood to the tumor. This increased blood supply
allows the cells to survive oxygen deprivation, allows the tumor to grow,
and even allows cancerous cells to escape into the bloodstream and migrate
to other tissues (the process known as metastasis).
If blocking angiogenesis can kill tumors, the only question that remains
Stopping the Hijacking
The problem with tumor angiogenesis is that tumors typically produce
over 20 different growth factors that are capable of inducing angiogenesis.
Blocking only a few of these growth factors has a minimal effect on tumor
angiogenesis and tumor growth. Therefore, to produce an effective antiangiogenic
drug, one must block a general component of the angiogenic cascadesome
crucial mediator of angiogenesis that gets turned on by multiple growth
factors. Block this general component and you will block angiogenesis.
This general component proved to be the protein Raf-1 kinase.
"If you could block Raf-1 kinase, you could block the growth factors'
signaling," says Cheresh. "That should block all the initiators of angiogenesis."
The problem was how to target Raf-1 only in angiogenic endothelial cells.
But Cheresh already had the solution to this in hand. Several years ago,
he discovered a specific "integrin" receptor protein that is displayed
on only newly sprouting endothelial cells such as those on tumor-associated
"This receptor is called avb3,"
says Cheresh. "It goes from essentially zero to high levels very quickly."
By specifically targeting avb3,
it is possible to target only those endothelial cells involved in angiogenesis-like
those that line vessels bringing blood to tumors.
Cheresh and his colleagues had spent several years studying the role
played in angiogenesis. It turned out that avb3
was capable of regulating a number of intercellular enzymes, called kinases,
that appeared to play crucial roles in angiogenesis. One of these kinases
Cheresh and his colleagues designed a dominant negative mutant form
of the Raf gene that was defective and when delivered into endothelial
cells would shut down the normal Raf-1 kinase activityeffectively
shutting off angiogenesis.
The only question that remained was how to combine the therapeutic effect
of the mutant Raf gene with the angiogenic cell-specific targeting
of the avb3.
A Gene Warhead and its Delivery Vehicle
Interestingly, the solution was provided by viruses, certain types of
which had evolved the ability to use avb3
integrins to gain entry into cells.
Following the lead of the viruses, Cheresh and his collaborators designed
a "cationic nanoparticle" that contained polymerized, positively charged
fat molecules containing an organic ligand that mimicked the binding site
for the viral protein receptor for avb3.
The positive charges on the nanoparticles allowed them to stick genes
to their surfaces, since DNA is negatively charged.
These gene-loaded nanoparticles then direct themselves to endothelial
cells displaying the avb3
receptor, where they gain entry and deliver their genetic payload.
In the current study, the TSRI investigators first report how they successfully
delivered nanoparticles with "reporter" genessuch as those encoding
for luciferase or green fluorescent protein, proteins that glow like the
tail of a firefly. These reporter genes allowed dramatic demonstrations
of the specific targeting of the nanoparticles to tumors. (The tumors
glowed green under a microscope).
Cheresh and his colleagues then combined the nanoparticle with the mutant
Raf gene and tested whether they could regress tumors in vivo,
and they found the technique worked. Everywhere there were metastatic
lesions in the lung or liver, the Raf gene eliminated them.
The next step, says Cheresh, is to refine these particles, identify
other gene targets, and utilize them in diseases characterized by abnormal
neovascularizationlike heart disease, stroke, rheumatoid arthritis,
and certain types of blindness in elderly patients (age-related macular
degeneration) and in patients with diabetes (diabetic retinopathy).
The research article "Tumor Regression by Targeted Gene Delivery to
the Neovasculature" is authored by John D. Hood, Mark Bednarski, Ricardo
Frausto, Samira Guccione, Ralph A. Reisfeld, Rong Xiang, and David A.
Cheresh and appears in the June 28, 2002 issue of the journal Science.
The research was supported by the National Institutes of Health and
by a grant from Merck KGAa.
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