Sullivan Awarded RO1 Grant to Further RNA Research

By Bill Bruton

“This is an important opportunity for us to continue work to realize potential in RNA therapeutics with our recent ribozyme discoveries,” Sullivan says. “With our technology platform, we are now in a stronger position to develop nucleic acid therapeutics not only for our own particular disease targets, but also for others having such interests in ophthalmology and elsewhere. Biologics (protein and nucleic acid) therapeutics are an evolving aspect of human medicines.”

The project title is “Optimizing Enhanced Hammerhead Ribozymes for Retinal Nucleic Acid Therapeutics.” The grant will provide almost $470,000 a year in direct costs for research.

“What this particular award is about is to further develop hammerhead ribosomes as potential therapeutics, based upon our new knowledge that we can engineer them to go much faster than they did historically,” Sullivan says.

“We know the parts of the RNA that are involved in the interactions that support enhanced speed, and now we want to figure out if we can push the kinetics even further to make better therapeutics.  This will involve experiments that are highly multidisciplinary,” he adds.

The messenger RNA — after it is processed — leaves the nucleus and goes to the cytoplasm, where that information on the messenger RNA is read into a string of amino acids that become the folded protein. That protein has a specific role in the cell — for vision, for metabolism, or for something else entirely.

“We now know that we can make these hammerhead ribozymes cut target mRNAs much faster at cellular levels of free magnesium. Given that they can go so much faster (log-orders), we think that we may be able to get them to work in the initial stages of gene expression — while the target pre-messenger RNA is still in the nucleus, while it is being processed,” Sullivan adds.

Sullivan says a mature rhodopsin mRNA mostly lives in the cytoplasm of the cell on the time scale of 10 or more hours. The time frame in the nucleus is on the order of tens of minutes.

“Normally one would direct these therapeutics to the cytoplasm, but these agents are so fast now — in some cases over a hundred-fold faster — that we think we can get them to work in the nucleus, where it would probably be easier to target the messenger RNA, because the nucleus is smaller and the therapeutic can be more concentrated, rather than being diluted in the vast sea of the cytoplasm,” Sullivan says. 

“Historically, the nucleus was not a typical environment to consider ribozyme therapeutics, because the pre-mRNA processing in the nucleus happens on a 10s of minutes time scale, whereas the lifetime of the mature mRNA in the cytoplasm is typically on the scale of hours. Before, with very slow therapeutics, the cytoplasm appeared to be the sole choice. The kinetically enhanced ribozymes open the nucleus to a novel environment for ribozyme therapeutics.” 

“Something that has been historically slow is found to have potential to be very fast. You can use it to suppress target gene expression, presumably with lower levels of it, and with less toxicity,” Sullivan adds. “Given that the hammerhead ribozyme has been around quite a long time, the prior limitations got in the way of its clinical application.”

Sullivan says there were corporate and academic groups who tried to bring ribozymes through clinical trials in the past.

“I recall there were at least three clinical trials — they all flopped. They flopped in large part because ribozymes were so slow, and they needed oodles of magnesium, way beyond the free amounts that were in a cell.  Our measurements are done at the physiological levels of magnesium, and we’re getting activity rates a hundred-fold faster than previous agents that were measured at much higher levels of magnesium (10-40 fold),” Sullivan says.

“That tells us something: that we may have a new therapeutic modality. Given the history of hammerhead ribozymes and their prior failures, our discovery takes us to the point where we can think about revitalizing that technology toward therapeutics for arbitrary disease targets. It doesn’t have to be retina, it can be anything where you have a toxic protein that is made, or some protein that’s made in excess for what the cell needs.”

Jason Myers, MA, a research scientist in the Department of Ophthalmology, has played a key role in the research. “Jason has made major intellectual contributions to the project. He’s been a major driver on this development,” Sullivan says.

Also contributing to the research is Michael H. Farkas, PhD, associate professor of ophthalmology.

The research is also being done in collaboration with the RNA Institute at the University at Albany. There, Kenneth Halvorsen, PhD; Jia Sheng, PhD; and Sweta Vangaveti, PhD; bring biophysical, chemical and structural biological, and computational dynamics to the challenge of ribozyme optimization. 

“The Department of Ophthalmology is a relatively small department, but it’s been very successful in generating extramural grant dollars. Probably one of the most important sections of the department from a research point of view is the retina group. We have a number of investigators working on retina, diabetic retinopathy, retinal degeneration, etc. That’s a strong element of the group. This successful funding effort contributes to the overall success of that group. It is a major win for the department and for the Ross Eye Institute. This NIH/NEI grant appears to be the largest R01 grant that the Department of Ophthalmology has received,” Sullivan says.

Sullivan cares for patients with retinal degenerative diseases at both the Ross Eye Institute (UBMD), where he is director of the Retinal Degeneration Service, as well as the VA Western New York Healthcare System, where he is a staff physician-scientist.  

“The patients motivate and teach us about just how much more still needs to be learned and done to treat these diseases. Our findings help to bring the ‘landscape of hope,‘” Sullivan says.