Could a cocktail of transcription factors reverse aging of the eye?

Turning cells back to a more youthful state

In the 19th century, the German biologist August Weismann stated that there are two types of cells, one that ages and the other seems immortal. In the human body, the immortal cells are the cells in the germline. Furthermore, because inheritance is only mediated by germ cells (that is, eggs and sperm), unnecessary genetic codes must be deleted or terminally inactivated in somatic cells that are committed to a specific state. We now know that at least part of this process occurs via DNA methylation. What is intriguing is that the ovary and the testis undergo aging and methylaton as one gets older, but the sperms and eggs where an embryo comes from revert back to a youthful state when an embryo is formed. As a result, when a baby is born, the cells in the baby are at a youthful state, rather than at the same aged state as the parents.

Cellular aging can be assessed by DNA methylation, and is commonly known as epigenetic clock. In a study published in 2021, the authors showed that epigenetic clock decreases significantly during embryogenesis, i.e. rejuvenation, followed by an increase in later stages. The author then stated that there is a natural rejuvenation event during embryogenesis, and this marks the beginning of aging as the point of minimal biological age.

The natural question is how this mechanism of rejuvenation occurs naturally. Yamanaka quoted this observation as central to the answer this question. When the nucleus of a somatic cell is transferred into an egg with its nucleus removed, this hybrid egg develops into an embryo that is genetically identical to the donor of the somatic cell. This implies that the somatic cell, although aged and well-differentiated, still contains the necessary genetic material for an embryo to form and that there is something in the egg that is able to revert the DNA expression back to embryogenic stage. Initially, this observation was made in frogs by John Gurdon, and subsequently the same technique was used in cloning of other animals, such as Dolly the Sheep in 1997.

Based on this observation, Yamanaka discovered that a combination of four transcription factors, namely Oct3/4, Sox2, c-Myc, and Klf4, can revert mature cells into cells that resemble embryonic stem cell in 2006. These cells are termed induced pluripotent stem cells. The immediate application of this discovery is to provide a source of stem cells for further biological research, bearing in mind that prior to this discovery, the source of stem cells is from embryos only, and this posts an obvious ethical problem in human stem cell research. This discovery also opens the door to the formation of organoid, which is an in vitro model of multiple cell types and exhibiting a tissue architecture and function seen in the actual organ.

A natural question is that if the Yamanaka factors can revert mature cells back to a more youthful state, could these factors also reverse aging in mature animals? Prolonged expression of the Yamanaka factors will revert mature cells back to stem cell like state, losing its cellular identity. These cells can then lose their cellular function and could even form teratomas. These initial concerns were addressed by timed or cyclic expression of the Yamanaka in mouse models, which demonstrated that such a treatment could lead to increased lifespan and improved regeneration.

Another approach is to use only three transcription factors Oct3/4, Sox2, and Klf4, and without c-Myc (as high c-Myc level can lead to cancer) for partial reprogramming. Expression of these three transcription factors in old mice led to an extension of lifespan and a number of health measures, with no formation of cancer.

The connection to optic neuropathy

I had the opportunity to attend a talk by Dr Joe Rizzo from Harvard Medical School on non-arteritic ischaemic optic neuropathy (NAION) and normal tension glaucoma (NTG). He stated that it is feasible that a younger optic nerve is more able to withstand insults than an older optic nerve. Given the above technique in partial reprogramming of the epigenome to return a cell to a more youthful state, would it be possible to improve outcomes in patients with NAION or NTG by rejuvenating the old optic nerve, even after the development of these conditions?

Animal studies support this approach. In an article in Nature in 2020, it was shown that expression of the genes for Oct3/4, Sox2, and Klf4 in the retinal ganglion cells in the eye in mice using a gene therapy like technique (adeno-associated virus platform) led to a more youthful epigenome, promotes axon regeneration after injury and restore vision in a mouse glaucoma model.

Life Biosciences has now commenced a phase one human trial on using the same technique for the treatment of NAION and NTG. The primary completion date is mid 2027, and hopefully there will be a positive result.

How about age-related macular degeneration

The treatment of dry (non-neovascular) age-related macular degeneration (AMD) remains an ongoing clinical problem with our current approaches being limited in efficacy in stopping the progression of the disease or to improve vision. It is feasible that expression of the same transcription factors in the photoreceptors and retinal pigment epithelium (RPE) could help in this condition, although there is no evidence to back this up currently.

Initial results on such an approach showed that expression of Oct3/4, Sox2, and Klf4 in the retinal pigment epithelium of a mouse model of AMD showed protection against this condition, prolonged expression of these factors beyond 8 weeks were shown to induce photoreceptor toxicity as measured by electroretinogram (ERG). Furthermore, the same study showed that the beneficial effect in oxidative stress from Oct3/4, Sox2, and Klf4 are mediated by GSTA4, and upregulation of GSTA4 leads to mitochondrial resilience and rejuvenates RPE cells.

Based on this animal study, direct application of Oct3/4, Sox2, and Klf4 for AMD could risk photoreceptor damage so a modified approach, potentially with GSTA4, is required to pursue this further in human studies. I am also concerned the possibility of Oct3/4, Sox2, and Klf4 activation in the photoreceptors when treating NAION and NTG as discussed above leading to photoreceptor toxicity, and will be looking out for any signs of this in future publications.

Conclusion

Partial epigenetic reprogramming using three Yamanaka factors is an exciting new development and may prove to be a novel way of treating age-related eye diseases. Its greatest promise, based on animal models, is in the treatment of optic neuropathy. However, with any novel treatment, there can be risks that are yet unknown. In this case, photoreceptor toxicity can limit the use of these factors in the human eye. I eagerly await future research in this field.