Anti-Cancer Nanoparticles:
Scientists at The Scripps Research Institute Design Gene-Tipped Tumor Regressor "Smartbombs"

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 angiogenesis—which 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 response—turning on genes that produce growth factor proteins, which tell the body to make new local blood vessels and bring more oxygen-rich blood.

The body responds when endothelial cells—the major cell type lining the vasculature—receive 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 is: How?

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 cascade—some 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 blood vessels.

"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 avb3 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 was Raf-1.

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 activity—effectively 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" genes—such 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 neovascularization—like 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.

 

 


The Nanoparticles at work. Click to Enlarge
Reprinted with permission from Science. Copyright 2002 American Association for the Advancement of Science.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 


If blocking angiogenesis can kill tumors, the only question that remains is: How?


 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 



For more information:

The Cheresh Lab