About five years ago, George Church and his new postdoctoral fellow, Ying Kai Chan, sat leaning over a laptop in the Harvard genetics pioneer’s office, staring in bewilderment at an old newspaper.
The paper documented early trials for Glybera, the first and at the time only gene therapy to be approved around the world. Less than three dozen patients ever got it, but in the years before Luxturna and Z Answerma, it gave researchers an example to point to that gene therapy really worked.
Chan and Church, however, were shocked to see that researchers conducting clinical trials had given patients a battery of high-dose immunosuppressants often reserved for organ transplants, names like mycophenolic acid and cyclosporine. And when they biopsied patients’ muscles, they were filled with T cells, indicating an active immune response.
The results were especially surprising because Glybera used the viral vector AAV, a delivery system that had led to a resurgence of the gene therapy precisely because it was widely believed to bypass the immune system overreaction that broke the field in the 1990s.
Ying Kai Chan“There are so many people working on gene therapy now and even if you tell them, oh, ‘did you know cyclosporine was being used? Did you know that all these things were used? People say, ‘Huh, what?’ Chan told Endpoints News. “Glybera was the poster child, but it seems people didn’t realize how much immunosuppression was needed.”
Trained as a viral immunologist, Chan entered the Church lab with the intuition that viral vectors, those hollowed-out and staged gene taxis, were still viruses and were still treated as such by the body. Gradually, the field has come into view. Multiple monkey studies showed that high doses of AAV can be toxic to certain neurons, results companies have been reluctantly accepting. And last year, three deaths in a high-dose study raised concerns about AAV safety, even if it is not yet definitively associated with an immune response.
Meanwhile, Chan has been working on new methods of disguising AAV to make it safer and reduce the need for immunosuppressants. This week he, the Church and a larger team from the Wyss Institute published the work in Science Translational Medicine, show how weaving specific strands of human DNA into the vector can neutralize one of the body’s main defense mechanisms against foreign invaders.
“It was very much inspired by nature,” said Chan.
One of the first ways gene therapy pioneer Jim Wilson showed that the body could respond to AAV was through a series of sentinels called toll-like receptors. These sentinels are one of the first layers of defense of the immune system, sounding the alarm when they detect anything that seems strange. However, that means that normal cells need a way to tell the receptors they are safe – an encryption key that only human cells know.
That encryption key is encoded in a pair of strands of DNA at the ends of telomeres, those paperclip-shaped strands at the ends of chromosomes that are sometimes involved in aging. Chan incorporated those strands into the DNA of an AAV2 vector, the serotype used in Luxturna. When the vector is injected, the strands must bind to the toll-like receptors throughout the body and tell the receptors not to sound an alarm.
When the team injected it into the muscle, liver, and eyes of pig and mouse models, it caused a markedly reduced immune response than a traditional vector, Chan reported in. STM.
The results add to a range of new technologies emerging in labs across the country to combat AAV immunogenicity. Wilson’s lab has provided a way to use microRNAs – short strands that minimize the expression of a particular gene in a particular cell – to mitigate its neural effects. And Dyno Therapeutics, a church lab spin-out, is using engineering and machine learning to invent entirely new vectors, hoping to find one that the immune system can avoid.
Chan has now helped launch a new company, in collaboration with ARCH and a few other VCs to form Ally Therapeutics, a still-hidden biotech trying to minimize the immunogenicity of viral vectors.
Still, he admits that he had hoped for more dramatic results than he ultimately had. Although his technology successfully suppressed the immune response in pigs and mice, the results were less profound in monkeys.
Chan’s team injected the vector into the eyes of non-human primates, an area of the body that much of the immune system cannot penetrate and as a result, toll-like receptors are acutely important. They saw improved safety when administered under the retina, but by injecting it directly into the glassy jelly in the center of the eye, it still caused significant inflammation. Intravitreal injection is important for addressing a variety of conditions and for safer and easier delivery of ocular gene therapies in a broader sense, as only ophthalmic surgeons can deliver subretinally.
However, the new article is only version 1.0 of the approach, Chan said, and they’ve made significant improvements since then.
More generally, the field still has a long way to go. Animal models, for example, are still poor predictors of the immune response in humans, making translation difficult and safety testing large gaps. A vector that appears immune-silent in monkeys can still trigger responses in humans and vice versa. Although their role in animals is well-documented, it is still unclear how great a role toll-like receptors play in the human response to AAV.
Still, Chan says they accomplished what they intended: they improved the vector and helped wake the field up to a problem that had been overlooked for years.
“There are still challenges,” said Chan. “What we really wanted to achieve was increase awareness and come up with a promising solution. I would say we have made progress on both fronts. “