Fight Aging!

Perhaps the most important early measure of the quality of a given approach to the treatment of aging is its effect on remaining life span in old mice. Prevention is a good approach, but it has the disadvantage of only working to its greatest effect in those who are not yet old. The best approaches to the treatment of aging will produce rejuvenation, and thus be applicable to both (a) prevention of degenerative aging in people who are entering later life and (b) reversal of degenerative aging in those already suffering its effects. Reversal of the cell and tissue damage that causes aging, when periodically applied prior to the worst pathology of aging, is prevention. Allowing mice to age into dysfunction followed by application of a therapy to restore health and extend life is a good indication that the therapy is producing rejuvenation.

In today's preprint paper, researchers outline an interesting approach to partial reprogramming, a way to restore a more youthful pattern of gene expression in the cells present in aged tissue. Evidence to date suggests that widespread partial reprogramming in most tissues is beneficial. The research employed an adeno-associated virus (AAV)-mediated gene therapy to introduce a conditional construct into the cells of aged mice, allowing the expression of Yamanaka factors in response to oral administration of the antibiotic doxycycline. Partial reprogramming could therefore be induced intermittently in all of the cells transduced by the AAV vector for the remainder of the mouse life span. The researchers used AAV9, an AAV variant that tends to give decent coverage of the major organs at the dose used in this study. The result was a doubling of remaining life span in the treated mice. This was a small study, 20 mice to a group, but the size of the outcome is large, a compelling result.

The intravenous AAV dose used here was at the high end of the practical and safe range for mice, and there have been deaths in human clinical trials at an equivalent dose. It puts stress on the liver, for example. AAV as it stands is a poor choice for uses that will require high dose systemic administration in large numbers of older patients, for a variety of logistical and regulatory reasons. There must be improvements to the AAV technology, or better gene therapy options must arise to replace it. Some lipid nanoparticles (LNPs) show promise, but they must be coupled with a payload able to reliably deliver genetic machinery into the cell nucleus. Many lines of work seem potentially able to solve one or other of these challenges, but none have as yet reached the goal to become both robustly functional and readily available for other projects.

One of the ways in which we will come to see improvement in gene therapy technologies is the continued demonstration in animal studies of ever more compelling outcomes that can be achieved via their use. Sizeable extension of remaining life span in old mice, achieved via use of a hot-topic technology currently backed by billions in funding for research and development, is likely to draw increased investment in ways to deliver the important advance of a gene therapy capable of cost-effective, safe-enough whole body introduction of long-lasting genetic additions.

Gene Therapy Mediated Partial Reprogramming Extends Lifespan and Reverses Age-Related Changes in Aged Mice

Using a cocktail of transcription factors, OCT4 (O), SOX2 (S), KLF4 (K), and c-MYC (M), collectively known as OSKM or Yamanaka factors, seminal studies showed that somatic cells can be reversed to a pluripotent state, thereby reversing a long-held paradigm of unidirectional differentiation. By short or cyclic induction of the Yamanaka factors in transgenic mice, investigators have demonstrated age extension in progeroid mice. These transgenic mouse models encoded a polycistronic OSKM cassette driven by a reverse tetracycline transactivator (rtTA) (4F mice); cyclic administration of doxycycline led to partial reprogramming without teratoma formation. This paradigm partially ameliorated aging phenotypes and extended the lifespan in the 4F-progeroid model. The study further showed that the epigenetic profile assessed by epigenetic methylation clocks of tissues, correlated with improved function.

The translation of these proof-of-concept genetic studies toward therapeutic interventions is to benefit the increasingly large aging population. In support of this endeavor, we have generated a systemically delivered two-part AAV9 system with doxycycline-inducible OSK, where one vector carried a constitutively expressed rtTa and the other vector contained a polycistronic OSK expression cassette driven by doxycycline responsive TRE promoter. We selected AAV9 capsid to ensure maximal distribution to most tissues. We injected 124-week-old wild type C57BL6/J mice retro-orbitally with 100 μl containing either PBS or 1E12 viral particles of each vector. We initiated the doxycycline induction for both the control and AAV administered groups the day after injections and alternated weekly on/off cycles for the remainder of the animals' lives.

Intriguingly, we observed a 109% extension in median remaining life in response to OSK expression (control mice had 8.86 weeks of life remaining vs. 18.5 weeks for TRE-OSK mice). Doxycycline-treated control mice had a median lifespan of ∼133 weeks, while the TRE-OSK mice had a median lifespan of 142.5 weeks. We observed a significant reduction in the frailty index from 7.5 points for doxycycline treated control mice to 6 points for TRE-OSK mice, suggesting that increased lifespan correlated to overall better health of the animals. We isolated DNA from heart and liver tissue from control and TRE-OSK treated mice at time of death and found that the Lifespan Uber Clock (LUC) for both liver and heart trended towards reduced epigenetic age.