DNAzymes & Nanoparticles

The Gold Standard: A Promising New DNAzyme Technology for Targeted Therapeutics

Let's say you have asthma. What if, instead of taking a powerful medicine that affects your whole body and could have serious side effects, you could deliver a small amount of that drug straight to your lungs?

This is one of many uses Khalid Salaita, PhD, associate professor of chemistry at Emory University, envisions for his breakthrough technology: catalytic DNA molecules (DNAzymes) that are attached to gold nanoparticles for use as therapeutics. DNAzymes are short DNA sequences that cleave mRNA at a single, programmable site. More than 20 years ago, scientists discovered the ability of DNAzymes to act like catalysts or enzymes, and to cleave to the backbone of specific RNA when that piece is complementary, says Salaita. A DNAzyme can be designed to break any specific RNA target in half, deactivating it.

Khalid Salaita, PhD

Soon afterward, many groups became interested in using these DNAzymes as an improved approach for gene regulation, to knock down and inhibit the expression of a protein of interest. "It worked beautifully in a test tube," he says. But there were problems in translation that have resulted in very few DNAzyme approved drugs over the following two decades.

First was the challenge of delivering a charged piece of DNA across the lipid membrane of a cell. The second was the inherent instability of foreign DNA within cells: the nucleases in the cell "chewed up" the DNAzymes, processing them quickly, so they were not as potent as in the lab.

Salaita attached the DNAzyme to a gold nanoparticle, which increased its stability, making it more difficult for nucleases to eliminate these catalytic pieces of DNA and therefore increasing their half-life. Also, the DNAzyme-coated gold nanoparticle was able to move very efficiently into the cell, crossing the cell membrane.


"Our solution is a platform that addresses the main problems of using DNAzymes as a therapeutic drug, by focusing on stability and delivery," says Catherine Murari-Kanti, licensing associate at Emory Office of Technology Transfer.

This technology makes Salaita hopeful that the DNAzyme can be used therapeutically to block the production of disease-causing proteins that are being overly expressed—in other words, to "get rid of the RNA middle man" so the protein is never made.

Many conditions are caused by the overexpression of genes inflammation, cancer, cardiovascular disease, autoimmune diseases, etc. So to be able to disrupt this process instead of treating the results with pharmaceuticals would be a real advantage, saving lives and diminishing suffering. In fact, recent clinical successes with diseases like chronic myelogenous leukemia and Parkinson's have renewed interest in gene therapies.

Other advantages of Salaita's technique are that it can be customized to target any mRNA, and gold nanoparticles are non-toxic and are already being used to treat rheumatoid arthritis, which speeds up clinical testing.

After Salaita verified that this technique worked in vitro, he moved toward testing it in animals with colleague Michael Davis, PhD, associate professor of cardiology at Emory, who studies different treatments for heart disease. They tested the particles by injecting them into the hearts of rats in which they had induced heart attacks. Those with the injected nanoparticles showed less inflammation and improved heart function than those injected solely with non-active DNAzyme nanoparticles.

Michael Davis, PhD

This year, Salaita began working with pulmonologist Cherry Wongtrakool, MD, assistant professor of medicine at Emory, an expert on asthma and lung disease who herself has asthma. "A colleague and I were discussing DNAzymes as a potential intervention in my animal model of allergic asthma, and he suggested I speak with Khalid. After discussing our mutual projects, we realized that targeting these DNAzymes to the lung was going to be important and that inhaled delivery of the DNAzymes would be the best approach," she says. "Khalid developed a DNAzyme nanoparticle that could do that." If small amounts of the nanoparticles could be targeted to the lungs, the systemic effects could be "markedly reduced," Wongtrakool says.

"Many years ago people took the bronchodilator albuterol as a tablet and there were many side effects including palpitations and tremors. Now we inhale albuterol, which minimizes the frequency of side effects," she says. "We wanted to test whether the DNAzyme gold nanoparticles could be nebulized and be effective as a drug."

They now have proof of concept in an animal model that DNAzyme gold nanoparticles can be nebulized and reach the distal airways of the lung. "We also have tested nebulized DNAzyme gold nanoparticles as an intervention in a model of allergic asthma and the results look promising," Wongtrakool says. "Our collaboration has been fortuitous."

To model allergic asthma, they exposed mice to house dust mite antigens, a common allergen triggering symptoms in many asthmatics. Over a few weeks, these mice developed more airway hyper-responsiveness, a key feature of asthma that results in a greater propensity for narrowing airways and increasing resistance. Then the mice were given an active DNAzyme nanoparticle against a specific protein known to drive allergic asthma. "We found that in the animals that received the active agent, their airway response matched those who had never been exposed to the dust mites," Wongtrakool says. "The nanoparticle conjugate allows for organ- specific delivery, and can specifically target genes that drive inflammation."

A common treatment for asthma (corticoeroids) can be administered via the inhalation or oral route. Oral corticosteroid use is reserved for patients with severe asthma. However, taking corticosteroids orally is "like using a sledge hammer, it impacts cytokine production in the whole body and has a myriad of side effects," Wongtrakool says. Long-term use of oral corticosteroids can lead to weight gain, and elevated blood glucose levels, which can become diabetes.

Because these nanoparticles can be localized to the organ of interest, such as the lungs, the possibility exists to modify the disease with a dose that is "a thousand times lower than what's in the literature." The technology is promising not just for heart disease and asthma, but for a host of other diseases as well, says Salaita: "The possibilities are exciting."

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