Restores age-related vision loss through epigenetic reprogramming

• The proof-of-concept study represents the first successful attempt to reverse the aging clock in animals through epigenetic reprogramming.
• Scientists activated embryonic genes to reprogram the cells of the mouse retinas.
• Approach to Inverted Glaucoma-Induced Eye Damage in Animals.
• The approach also restored age-related vision loss in aged mice.
• Work spells promise to use the same approach in other tissues, organs besides the eyes.
• Success sets the stage for the treatment of various age-related diseases in humans.

Harvard Medical School scientists successfully restored vision in mice by turning back the clock on aged eye cells in the retina to regain juvenile gene function.

The team’s work, described today (December 2, 2020) in Nature, represents the first demonstration that it may be possible to safely reprogram complex tissues, such as nerve cells in the eye, at an earlier age.

In addition to restoring the cell’s aging clock, the researchers successfully reversed vision loss in animals with a condition that mimicked human glaucoma, a leading cause of blindness worldwide.

The result represents the first successful attempt to reverse glaucoma-induced vision loss, rather than simply curbing its progression, the team said. If replicated through further studies, the approach could pave the way for therapies aimed at promoting tissue repair in various organs and reversing aging and age-related diseases in humans.

“Our study shows that it is possible to safely reverse the age of complex tissues such as the retina and restore its juvenile biological function,” said senior author David Sinclair, professor of genetics at the Blavatnik Institute at Harvard Medical. School, co-director of the Paul F. Glenn Center for Biology of Aging Research at HMS and an aging expert.

Sinclair and colleagues caution that the results need to be replicated in further studies, including in different animal models, before any human experiments. However, they add, the findings offer a proof of concept and a path to designing treatments for a range of age-related human diseases.

“If affirmed through further studies, these results could be transformative for the treatment of age-related vision diseases such as glaucoma and for the fields of biology and medical therapies for the disease in general,” Sinclair said.

For their work, the team used an adeno-associated virus (AAV) as a vehicle to deliver three youth-restoring genes – Oct4, Sox2 and Klf4 – into the retinas of mice, which are normally activated during embryonic development. The three genes, along with a fourth, which was not used in this work, are collectively known as Yamanaka factors.

The treatment had multiple beneficial effects on the eye. First, it promoted nerve regeneration following optic nerve injury in mice with damaged optic nerves. Second, it reversed vision loss in animals with a condition that mimicked human glaucoma. And third, it reversed vision loss in older animals without glaucoma.

The team’s approach is based on a new theory of why we age. Most of the cells in the body contain the same DNA molecules but they have very different functions. To achieve this degree of specialization, these cells only need to read genes specific to their type. This regulatory function is the responsibility of the epigenome, a system of activating and deactivating genes in specific patterns without altering the underlying DNA base sequence of the gene.

This theory postulates that changes to the epigenome over time cause cells to read the wrong genes and malfunction, giving rise to diseases of aging. One of the most important modifications to the epigenome is DNA methylation, a process by which methyl groups are attached to DNA. DNA methylation patterns are established during embryonic development to produce the various types of cells. Over time, juvenile DNA methylation patterns are lost and genes within cells that should be activated are turned off and vice versa, resulting in impaired cell function. Some of these DNA methylation changes are predictable and have been used to determine the biological age of a cell or tissue.

However, it is still unclear whether DNA methylation results in age-related changes within cells. In this study, the researchers hypothesized that if DNA methylation actually controls aging, erasing some of its fingerprints could reverse the age of cells within living organisms and restore them to their earlier, younger state.

Previous work had achieved this feat in cells grown in laboratory dishes, but failed to demonstrate the effect on living organisms.

The new findings show that the approach could also be used in animals.

Overcoming an important obstacle

The lead author of the study, Yuancheng Lu, a researcher in genetics at HMS and a former doctoral student in Sinclair’s lab, developed a gene therapy that could safely reverse the age of cells in a living animal.

Lu’s work is based on the discovery of the Nobel laureate Shinya Yamanaka, who identified the four transcription factors, Oct4, Sox2, Klf4, c-Myc, which could erase epigenetic markers on cells and return these cells to their embryonic state. primitive from which they can develop into any other type of cell.

Subsequent studies, however, have shown two major setbacks. First, when used in adult mice, the four Yamanaka factors could also induce tumor growth, making the approach unsafe. Second, the factors could restore the cellular state to the most primitive cellular state, thereby completely erasing a cell’s identity.

Lu and colleagues worked around these obstacles by slightly modifying the approach. They dropped the c-Myc gene and provided only the remaining three genes Yamanaka, Oct4, Sox2, and Klf4. The modified approach successfully reversed cellular aging without fueling tumor growth or losing its identity.

Gene therapy applied to optic nerve regeneration

In this study, the researchers targeted cells of the central nervous system because it is the first part of the body affected by aging. After birth, the central nervous system’s ability to regenerate rapidly decreases.

To test whether the regenerative capacity of juvenile animals could be imparted to adult mice, the researchers delivered the modified combination of three genes via an AAV into retinal ganglion cells of adult mice with optic nerve lesions.

For the work, Lu and Sinclair collaborated with Zhigang He, HMS professor of neurology and ophthalmology at Boston Children’s Hospital, who studies neuro-regeneration of the optic nerve and spinal cord.

Treatment resulted in a two-fold increase in the number of surviving retinal ganglion cells after injury and a five-fold increase in nerve regrowth.

“At the start of this project, many of our colleagues said our approach would either fail or be too dangerous to ever use,” Lu said. “Our results suggest that this method is safe and could potentially revolutionize the world. treatment of the eye and many other organs affected by aging “.

Reversal of glaucoma and age-related loss of vision

Following encouraging results in mice with optic nerve injuries, the team collaborated with colleagues at the Schepens Eye Research Institute at Massachusetts Eye and Ear Bruce Ksander, HMS associate professor of ophthalmology, and Meredith Gregory-Ksander, HMS assistant professor of ophthalmology. They planned two sets of experiments: one to test if the cocktail of three genes could restore vision loss due to glaucoma and another to see if the approach could reverse vision loss resulting from normal aging.

In a mouse model of glaucoma, the treatment resulted in increased electrical activity of nerve cells and a noticeable increase in visual acuity, as measured by the animals’ ability to see moving vertical lines on a screen. Surprisingly, it did so after glaucoma-induced vision loss had already occurred.

“Recovery of visual function after injury has rarely been demonstrated by scientists,” said Ksander. “This new approach, which successfully reverses multiple causes of vision loss in mice without the need for a retinal transplant, represents a new treatment modality in regenerative medicine.”

The treatment worked equally well in 12-month-old mice with decreased vision due to normal aging. After treatment of the elderly mice, the gene expression patterns and electrical signals of the optic nerve cells were similar to the young mice, and vision was restored. When the researchers analyzed the molecular changes in the treated cells, they found inverted patterns of DNA methylation, an observation that suggests that DNA methylation is not a mere marker or spectator in the aging process, but rather an active agent that drives him.

“What this tells us is that the clock doesn’t just represent time, it is time,” Sinclair said. “If you rewind the hands of the clock, time goes back too.”

The researchers said that if their findings are confirmed in further animal work, they could start clinical trials within two years to test the approach’s effectiveness in people with glaucoma. So far, the results are encouraging, the researchers said. In the current study, a one-year full-body treatment of mice with the three-gene approach showed no negative side effects.

Reference: 2 December 2020, Nature.
DOI: 10.1038 / s41586-020-2975-4

Other authors on the article include Benedikt Brommer, Xiao Tian, ​​Anitha Krishnan, Margarita Meer, Chen Wang, Daniel Vera, Qiurui Zeng, Doudou Yu, Michael Bonkowski, Jae-Hyun Yang, Songlin Zhou, Emma Hoffmann, Margarete Karg, Michael Schultz, Alice Kane, Noah Davidsohn, Ekaterina Korobkina, Karolina Chwalek, Luis Rajman, George Church, Konrad Hochedlinger, Vadim Gladyshev, Steve Horvath and Morgan Levine.

This work was supported in part by a grant and epigenetics seed grant from Harvard Medical School, The Glenn Foundation for Medical Research, Edward Schulak, National Institutes of Health (grants R01AG019719, R37AG028730, R01EY026939, R01EY021526, R01AG067765, R01GM0 , R01, R24EY028767 and R21EY030276) and the St. Vincent de Paul Foundation.

Relevant Disclosures: David Sinclair is a consultant, licensed patent inventor, board member and shareholder of Iduna Therapeutics, a Life Biosciences company that develops epigenetic reprogramming therapies and a free consultant to Zymo Research, a tools company epigenetics. Yuancheng Lu, Luis Rajman and Steve Horvath are shareholders of Iduna Therapeutics. George Church and Noah Davidsohn are co-founders of Rejuvenate Bio.