New ‘Litmus Test’ Could Aid Discovery of Anti-Cancer Drugs

March 28, 2006

New ‘Litmus Test’ Could Aid Discovery of Anti-Cancer Drugs

EVANSTON, Ill.—Using the unusual color properties of gold at the nanoscale, scientists at Northwestern University have developed a “litmus test” for DNA and small molecule binding that eventually could be used by pharmaceutical companies to rapidly identify promising candidates for new anti-cancer drugs.

The detection system, called colorimetric screening, can be used to detect a variety of targets, such as DNA and small molecules and proteins that bind to DNA. The strength of the bond is indicated by a simple color change.

In a paper reported online March 28 by the Journal of the American Chemical Society (JACS), the research team used the colorimetric method to screen for molecules that can facilitate the formation of a special form of DNA called a triple helix. Triple helix DNA involves three strands rather than the two associated with normal DNA. However, unlike double helix DNA, the triple helix is unstable alone and requires a small molecule triplex binder to increase its stability. This research builds on work reported March 6 in the German journal Angewandte Chemie in which the same method was used to screen small molecules for their binding affinity to duplex DNA.

“Pharmaceutical companies are targeting DNA for different therapies, and they need to identify DNA or small molecules that selectively bind to DNA to turn on or off the gene expression related to a particular disease,” said Chad A. Mirkin, PhD, George B. Rathmann Professor of Chemistry, professor of medicine, and professor of materials science and engineering, who led both studies. “Our method, which is simpler, faster, and more convenient than conventional methods, should help researchers zero in on potential anti-cancer agents from their large libraries of candidates more quickly.”

In the JACS paper, the researchers demonstrated that when a triplex binder binds to a given DNA triple helix in solution the strength of that binding event can be detected by the naked eye. The color of the solution changes from blue to red when heated, and the temperature at which this occurs indicates the strength of the triplex binder’s bond.

Much like tiny bits of gold produce the vibrant red in stained glass windows, the Northwestern team also takes advantage of gold’s intense color when the metal is measured on the scale of atoms. The researchers start with gold nanoparticles, each just 13 nanometers in diameter, held together by DNA in a triple helix conformation. Because they are held together within a certain critical distance, the gold nanoparticles—and the solution they are in—are blue. When the solution is heated, the DNA breaks apart, and the gold nanoparticles, no longer in close proximity to each other, are now bright red.

When a triplex binder binds to DNA it creates a stronger bond that requires a higher temperature to break apart the DNA strands. By adding different triplex binders to a solution with a specific DNA sequence between the gold nanoparticles, the colorimetric method can distinguish molecules that are strong binders, medium binders, and weak binders. (Current methods using fluorescence can only indicate whether or not a bond has formed.) When heated, the solution with a triplex binder that is a weak binder turns red at a lower temperature than the solution with a strong binder. This color change from blue to red allows the researchers by simple visual inspection to see which triplex binders bind to the DNA and how strongly they bind.

“It’s impossible to do a full-blown study on every triplex binder or small molecule,” said Dr. Mirkin, who is director of Northwestern’s Center for Cancer Nanotechnology Excellence. “You need to narrow down the possible candidates. This method allows researchers to identify the types of triplex binders or molecules that are effective for a given DNA sequence. Most diseases have a unique genetic code associated with them, and by manipulating the genes with the right triplex binders or small molecules you can develop new therapies.”

A certain DNA sequence might be linked to colon cancer, for example, and the proteins expressed by that DNA produce cancer cells. By identifying a triplex binder or small molecule that binds effectively to that particular DNA sequence, a drug can be developed that shuts down protein production and stops cancer cells from proliferating.

Their next step, said Dr. Mirkin, is to challenge the research community to provide libraries of triplex binders and small molecules for his research teams to test. Any interaction with DNA, whether it be with small molecules, proteins, or other DNA, can be identified by the colorimetric method. “We want to identify the real winners,” said Dr. Mirkin.

In addition to Dr. Mirkin, other authors on the JACS paper are postdoctoral fellow Min Su Han, PhD (first author), and graduate student Abigail K.R. Lytton-Jean, both from Northwestern University.

The research was supported by the National Cancer Institute, National Science Foundation, and Air Force Office of Scientific Research.