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The highly repeated structure of the ribsosomal DNA (rDNA) locus and its high rates of transcription make it particularly vulnerable to genome instability and damage. Multiple studies have reported a link between rDNA stability and cellular aging, as
well as the association of proteins involved in genome integrity transiting the nucleolus. Aging in yeast is accompanied by nucleolar enlargement and fragmentation, suggesting a mechanism of cellular aging that may be related to nucleolar structure.
Concordantly a recent study reported that the premature aging disorder Hutchinson-Gilford progeria syndrome leads to nucleolar expansion and increased ribosome biogenesis. Furthermore, there is evidence suggesting an association of replication stress on
rDNA loci with the aging of hematopoietic stem cells, adding more evidence to the general function of the nucleolus in genome integrity and aging.

The nucleolus also impacts other vital cellular processes like the cell cycle and the response to cellular stress. One of the major tumor suppressor proteins central to regulating cell cycle is p53. The nucleolus acts as a platform connecting a cellular
stress response with cell cycle through the central tumor suppressor p53. Interestingly multiple studies have implicated p53 in aging in different organisms. The nucleolus has also been associated with regulation of cell senescence. Alterations in
nucleolar morphology have been reported in aging cells. In particular, presenescent cells exhibit multiple small-sized nucleoli compared to senescent cells which possess a single enlarged nucleolus.

The perception that the nucleolus is simply the place where ribogenesis takes place has clearly evolved. We now know that it is a highly dynamic organelle that coordinates signals from growth, energy, and stress to the balanced production and assembly of
multiple ribonucleoprotein particles and the maintenance of genome integrity. This has ramifications for essentially all levels of molecular organization from genome architecture, RNA metabolism, protein synthesis and quality control to metabolism.

Unexpectedly Better Results Cause Phase III Trial Failure for Gensight https://www.fightaging.org/archives/2018/05/unexpectedly-better-results-cause-phase-iii-trial-failure-for-gensight/

Clinical trials must produce exactly the expected result or they are declared a failure. A clinical trial can fail by producing unexpected benefit, and this has happened to Gensight Biologics' work on allotopic expression of the mitochondrial NH4 gene,
aimed at the treatment of inherited retinal degeneration caused by mutation in NH4. Allotopic expression is a process in which a copy of the correctly formed gene is placed into the cell nucleus, suitably altered to enable transport of the protein
produced back to the mitochondria where it is need.

In a sane world, this therapy would long ago have been available to anyone willing to accept the risk, based on positive earlier results and lack of serious side-effects. In that same sane world, the therapy would now be available to anyone willing
undergo the risk of gaining a greater benefit than was hoped for. Unfortunately we don't live in that world, and Gensight will now have to run further very expensive trials before regulators will permit the treatment to reach the clinic.

We watch progress at Gensight because allotopic expression of NH4 is one-thirteenth of a rejuvenation therapy - Gensight is the result of research supported and encouraged by the Methuselah Foundation some ten years ago. If all thirteen important
mitochondrial genes can be copied into the cell nucleus, then that would make an individual largely invulnerable to stochastic mitochondrial DNA damage and resultant dysfunction. It is thought that this is an important root cause of degenerative aging.

GenSight Biologics, a biopharma company focused on discovering and developing innovative gene therapies for retinal neurodegenerative diseases and central nervous system disorders, today announced topline results from the REVERSE Phase III clinical trial
evaluating the safety and efficacy of a single intravitreal injection of GS010 (rAAV2/2-ND4) in 37 subjects whose visual loss due to 11778-ND4 Leber Hereditary Optic Neuropathy (LHON) commenced between 6 and 12 months prior to study treatment.

Topline results further highlight the favorable safety and tolerability profile of GS010, and demonstrate a clinically meaningful improvement of +11 ETDRS letters in treated eyes at 48 weeks as compared to baseline in all 37 patients. Unexpectedly,
untreated contralateral eyes (treated with a sham injection) show a similar improvement of +11 ETDRS letters. Due to this improvement in untreated eyes, the trial did not meet its primary endpoint, defined as a difference of improvement in visual acuity
in GS010-treated eyes compared to sham-treated eyes at 48 weeks.

The improvement of visual acuity in sham-treated eyes was unexpected based on the natural history of LHON, for which partial spontaneous recovery is reported in only 8-22% of patients with the G11778 ND4 mutation. "This meaningful improvement of
untreated eyes observed at week 48 was totally unexpected given what is known and has been published about the natural history of this devastating disease. We will continue to analyze the data to better understand our results, but they suggest that GS010
benefits both eyes in a way that is still to be understood. The fact that structural measures of the retina showed such a large statistical difference with treatment is compelling and objective evidence that this gene therapy protects the integrity of
many retinal ganglion cells from the damage of LHON."

Based on preliminary analysis of the safety data, GS010 was well tolerated after 48 weeks. The ocular adverse events most frequently reported in the therapy group were mainly related to the injection procedure, except for the occurrence of intraocular
inflammation (accompanied by elevation of intraocular pressure in some patients) that is likely related to GS010, and which was responsive to conventional treatment and without sequelae. There were no withdrawals from the trial. GS010 is currently being
investigated in two additional ongoing Phase III trials, RESCUE and REFLECT, while patients in REVERSE continue to be followed for another 4 years.

Increased Mitochondrial DNA Copy Number Slows Vascular Aging in Mice https://www.fightaging.org/archives/2018/05/increased-mitochondrial-dna-copy-number-slows-vascular-aging-in-mice/

The open access paper here presents an interesting result in mitochondrial biology. Mitochondria are the power plants of the cell, a herd of bacteria-like structures responsible for packaging chemical energy store molecules. They have their own small
genome of a few mitochondrial genes. A mitochondrion may have one or several copies of this genome, and mitochondria promiscuously fuse together, divide, and swap around their component parts from one to another. This makes it quite hard to understand
how their age-related dysfunction and damage progresses in detail.

Nonetheless, it is well demonstrated that mitochondria become progressively less functional with advancing age, and this is particularly relevant in energy-hungry tissues such as muscles and the brain. Some of this decline may be reaction to forms of
cell and tissue damage, and some of this is due to stochastic mutational damage occurring to mitochondrial DNA. In this context, the researchers here show that forcing an increase in the number of copies of mitochondrial DNA can maintain mitochondrial
function in old age, and thereby slow vascular aging. It remains unclear, however, as to the exact chain of mechanisms that make this the case: the causes and immediate consequences of an age-related reduction in the number of copies of mitochondrial DNA
are not well understood at this point in time.

Mitochondria contain multiple copies of mitochondrial DNA (mtDNA) that encode ribosomal and transfer RNAs and many essential proteins required for oxidative phosphorylation. Loss of mtDNA integrity by both altered mitochondrial DNA copy number (mtCN) and
increased mutations is implicated in cellular dysfunction with aging. Reactive oxygen species (ROS), many of which are generated by mitochondria, also increase with age. However, the role of mitochondria in aging may extend beyond ROS, and it is unclear
whether decreased mitochondrial function promotes vascular aging directly or is just a consequence of aging.

Aging of the large conduit arteries is a major cause of morbidity and mortality, contributing to hypertension (high blood pressure) and stroke. Currently, it is unclear what the earliest time points that constitute vascular aging in laboratory mice are,
which physiological measures of large artery stiffness correspond most closely to humans, and whether similar processes underlie changes in mechanical properties in mouse and human arteries. Aging research is time-consuming and expensive because of the
long time courses needed. Therefore, identifying the earliest time points that show the most sensitive and reproducible changes and parameters is crucial in obtaining scientific consensus for mouse models of vascular aging.

We examined multiple parameters of vascular function, histological markers, and markers of mitochondrial damage and function during normal vascular aging, and the effects of reducing or augmenting mitochondrial function on the onset and progression of
vascular aging. We identify early, standardized time points and reproducible physiological parameters for vascular aging studies in mice. Vascular aging begins at far earlier time points than previously described in mice, with compliance, distensibility,
stiffness, and pulse wave velocity (PWV) being the best discriminators for normal aging and manipulations. Mitochondrial DNA copy number and mitochondrial respiratory function are reduced when functional and structural manifestations of vascular aging
begin. Rescue of the copy number deficit observed in normal aging improves mitochondrial respiration and delays all parameters of vascular aging, while reduced mtDNA integrity accelerates vascular aging. Together these data highlight the direct role of
mtDNA-mediated mitochondrial dysfunction in the progression of vascular aging.

Improved Approaches to Messing with Metabolism Will Use Gene Therapies https://www.fightaging.org/archives/2018/05/improved-approaches-to-messing-with-metabolism-will-use-gene-therapies/

I see that noted geneticist George Church has been discussing his new company Rejuvenate Bio in the media. The projects undertaken there are the logical progression of attempts to slow aging with pharmaceuticals, moving them into the era of gene therapy.
This is still guided by the a philosophy of what Aubrey de Grey would call "messing with metabolism." This means that researchers are attempting to alter the amounts of specific proteins in ways that adjust the operation of metabolism into what is
hopefully a more optimal state, one in which cell and tissue damage, or the consequences of that damage, accrue more slowly. Gene therapies are far more effective tools than pharmaceuticals when it comes to achieving this outcome with minimal side-
effects, and there are many candidate genes to explore.

This is not, however, likely to be as effective as repairing the underlying damage that causes aging. It is tinkering with the broken state of metabolism that arises due to damage, trying to make it more functional without addressing the root cause of
its problems. Clearly it is possible to do useful things via this approach, as demonstrated by the existence of statins, first generation stem cell therapies, and the like, but all of these technologies are in principle very limited in comparison to what
might be achieved by reversing the root causes of aging.

Professor George Church of Harvard Medical School has co-founded a new startup company, Rejuvenate Bio, which has plans to reverse aging in dogs as a way to market anti-aging therapies for our furry friends before bringing them to us. The company has
already carried some initial tests on beagles and plans to reverse aging by using gene therapy to add new instructions to their DNA. If it works, the goal is ultimately to try the same approach in people. "Dogs are a market in and of themselves. It's not
just a big organism close to humans. It's something that people will pay for, and the FDA process is much faster. We'll do dog trials, and that'll be a product, and that'll pay for scaling up in human trials."

Church and the team also understand that developing therapies that address aging in humans and getting them approved would not be so easy. It would take too long to prove something worked. "You don't want to go to the FDA and say we extend life by 20
years. They'd say, 'Great, come back in 20 years with the data.'" So, the team has taken a different tack; rather than aiming to increase human lifespan as its main focus, it is instead focusing on the typical age-related diseases common to dogs. The
hope is that by targeting the aging processes directly, these diseases could be entirely prevented from developing. If successful, this would lend additional supporting evidence that directly treating aging to prevent age-related diseases could also work
in humans.

The lab has been working on a collection of over 60 different gene therapies and has been testing their effects both individually and in combinations. The team intends to publish a report on an approach that extends mouse lifespan by modifying two genes
that protect against heart and kidney failure, obesity, and diabetes. Professor Church has commented that the results of this study are "pretty eye-popping". The new startup has been contacting dog breeders, veterinarians, and ethicists to discuss its
plans for restoring youth and increasing the lifespan of dogs. Its plan is to gain a foothold in the pet market and then use that as the basis for moving therapies to people.

An Interview with Reason at the Life Extension Advocacy Foundation https://www.fightaging.org/archives/2018/05/an-interview-with-reason-at-the-life-extension-advocacy-foundation/

Following the launch of Repair Biotechnologies, and since I'll be at the Life Extension Advocacy Foundation's one-day conference, Ending Age-Related Disease, this coming July in New York, I recently answered a few questions and offered a few opinions for
the LEAF volunteers. That interview was published yesterday. Regular readers here at Fight Aging! are no doubt all too familiar with many of those opinions already, since I'm not exactly what one might call reticent about putting them forward, but it
never hurts to check.

Thanks to the efforts of many advocates, yours included, public perception of rejuvenation is also shifting. How close do you think we are to widespread acceptance?

I don't think acceptance matters - that might be the wrong term to focus on here. Acceptance will occur when the therapies are in the clinic. People will use them, and everyone will conveniently forget all the objections voiced. The most important thing
is not acceptance but rather material support for development of therapies. The help of only a tiny fraction of the population is needed to fund the necessary research to a point of self-sustained development, and that is the important thing. Create
beneficial change, and people will accept it. Yet you cannot just go and ask a few people. Persuading many people is necessary because that is the path to obtaining the material support of the necessary few: people do not donate their time and funds to
unpopular or unknown causes; rather, they tend to follow their social groups.

Presently, rejuvenation is a relatively unknown topic; people who say they're against this technology probably don't think it's a concrete possibility anyway. However, as more important milestones will be reached - for example, robust mouse rejuvenation -
this might change. Do you think that these milestones will result in opponents changing their attitudes or becoming more entrenched?

Opposition to human rejuvenation therapies is almost entirely irrational; either (a) it's a dismissal of an unfamiliar topic based on the heuristic that 95% of unfamiliar topics turn out to be not worth the effort when investigated further, or (b) it's a
rejection of anything that might result in sizable change in personal opinion, life, and plans, such as the acceptance of aging and death that people have struggled to attain. This sort of opposition isn't based on an engagement with facts, so I think a
sizable proportion of these folk will keep on being irrational in the face of just about any scientific advance or other new factual presentation short of their physicians prescribing rejuvenation therapies to treat one or more of their current symptoms
of aging.

On the other hand, there will be steady progress in winning people over in the sense of supporting rejuvenation in the same sense as supporting cancer research: they know nothing much about the details, but they know that near everyone supports cancer
research, and cancer is generally agreed to be a bad thing, so they go along. Achieving this change is a bootstrapping progress of persuading opinion makers and broadcasters, people who are nodes in the network of society. Here, milestones and facts are
much more helpful.

After years of financially supporting other rejuvenation startups, you're now launching your own company focused on gene therapies relevant to rejuvenation. Your company's first objective is thymic regeneration. Why do you think the thymus is the ideal
initial target for your work?

It is a very straightforward goal, with a lot of supporting evidence from the past few decades of research. It think it is important to set forth at the outset with something simple, direct, and focused, insofar as any biotechnology project can be said
to have those attributes. This is a part of the SENS rejuvenation research agenda in the sense of cell atrophy: the core problem is loss of active thymic tissue, which leads to loss of T cell production and, consequently, immunodeficiency. However, the
immune system is so core to the health of the individual that any form of restoration can beneficially affect a great many other systems. The many facets of the immune system don't just kill off invading pathogens; they are also responsible for
destroying problem cells (cancers, senescent cells), and they participate in tissue maintenance and function in many ways.

You are using gene therapy; why have you chosen this delivery method specifically and not, for example, a small-molecule approach?

If your aim is to raise or lower expression of a specific protein, and you don't already have a small molecule that does pretty much what you want it to do without horrible side-effects, then you can pay 1-2M for a shot at finding a starting point in the
standard drug discovery databases. That frequently doesn't work, the odds of success are essentially unknown for any specific case, and the starting point then needs to be refined at further cost and odds of failure. This is, for example, the major
sticking point for anyone wanting to build a small-molecule glucosepane breaker - the price of even starting to roll the dice is high, much larger than the funding any usual startup crew can obtain.

On the other hand, assuming you are working with a cell population that can be transduced by a gene therapy to a large enough degree to produce material effects, then 1-2M will fairly reliably get you all the way from the stage of two people in a room
with an idea to the stage of having animal data sufficient enough to start the FDA approval process.

A View of Aging Centered Around Mutation and Senescence https://www.fightaging.org/archives/2018/05/a-view-of-aging-centered-around-mutation-and-senescence/

Many researchers see stochastic mutational damage to nuclear DNA as an important mechanism in aging, above and beyond its contribution to cancer risk. The challenge has always been that there don't seem to be enough mutations to explain significant harm,
if the harm remains restricted to only the cell in which the mutation occurs. One way to explain how DNA damage causes more general issues is through clonal expansion of detrimental mutations that occur in stem and progenitor cells. Another possible
explanation, presently being energetically explored by the research community, is that DNA damage can cause cellular senescence. In this case, just a few senescent cells can cause outsized amounts of harm in surrounding tissue through the potent mix of
signals they secrete: generating inflammation, remodeling the extracellular matrix, changing the behavior of other cells for the worse, and so on. We'll be seeing a great many papers like this one in the years ahead, I think.

During an organism's lifetime, cells are constantly exposed to exogenous and endogenous stressful agents. Cells can cope with these stressors by various response mechanisms, or in case of irreversible damage, programmed cell death (apoptosis), or
permanent cell-cycle arrest (cellular senescence). Cellular senescence is characterized by a halt in cellular replication, accompanied by a specific molecular phenotype. This phenotype can be the result of a few factors, such as accumulation of DNA
damage, telomere attrition, and various epigenetic alterations.

Cellular senescence is one of the cellular pathways contributing to organismal aging. Senescent cells can accumulate in tissues and organs and can ultimately result in tissue lesions that will cause organ dysfunction, such as through the senescence-
associated secretory phenotype (SASP). Age-related accumulation of DNA damage has been studied thoroughly, showing correlation between age and damage levels or mutation frequency. In the presence of DNA lesions or abnormalities, the DNA damage response (
DDR) is activated and can eventually lead to cell cycle arrest. In older organisms, accumulation of DNA damage and loss of regenerative potential consequently increase the number of senescent cells, leading to aging cells, tissues, organs, and inevitable
death.

The accumulation of genomic abnormalities is influenced by the quality of the repair pathways, which may also decline with age. Researchers studied age-related DNA damage in peripheral blood cells using single nucleotide polymorphism (SNP) microarray
data from over 50,000 individuals. The frequency of detectable genomic abnormalities was low (less than 0.5%) at birth and rose to 2-3% in 50-year-old donors. Peripheral blood cells were also studied using whole-exome sequencing data from DNA of 17,182
individuals lacking hematologic phenotypes. Somatic mutations were rare in young donors (~40 years old) but became more frequent with age. Furthermore, while studying subjects at 70-79 years, compared with 90-108 years, mutation frequency rose from 9.5
to 18.4%, respectively.

In conclusion, the connection between DNA damage and aging is emphasized by the secretion of senescence-associated proteins during cellular senescence, a phenotype which is activated by DNA damage and is common for both human and mice. Though much
progress has been achieved, full understanding of these mechanisms has still a long way to go.

XPO1 as a Novel Target for Therapies to Enhance Autophagy https://www.fightaging.org/archives/2018/05/xpo1-as-a-novel-target-for-therapies-to-enhance-autophagy/

Autophagy is the name given to a collection of cellular housekeeping processes that recycle damaged and unwanted proteins and structures inside a cell. Most of the means of slowing aging demonstrated in laboratory species involve increased autophagy: it
is an important response to any form of stress likely to result in more damage inside the cells. The less damage there is, the better off the cells. This in turn can leads to a longer, healthier life span to some degree. It is also worthy of note that
autophagy declines with age, and this is though important in a range of age-related conditions.

Autophagy enhancement therapies have been on the research community agenda for a long time now. There have been scores of papers published on this topic in the last decade alone, even putting to one side the point that all calorie restriction mimetic
development is likely based on increased levels of autophagy somewhere under the hood. Unfortunately, means of directly enhancing autophagy have not as yet made it out of the lab; there has been very little progress towards the clinic. This is worth
bearing in mind when reading publicity materials of the sort presented here. It is little different in tone from similar items published many years ago, and which subsequently went nowhere.

The process of autophagy involves the rounding up of misfolded proteins and obsolete organelles within a cell into vesicles called autophagosomes. The autophagosomes then fuse with a lysosome, an enzyme-containing organelle that breaks down those
cellular macromolecules and converts it into components the cell can re-use. Researchers wanted to see if they could increase autophagy by manipulating a transcription factor (a protein that turns gene expression on and off) that regulates autophagic
activity. In order for the transcription factor to switch autophagic activity on, it needs to be localized in the nucleus of a cell. So the team screened for genes that enhance the level of the autophagy transcription factor, known as TFEB, within nuclei.

Using the nematode C. elegans, the screen found that reducing the expression of a protein called XPO1, which transports proteins out of the nucleus, leads to nuclear accumulation of the nematode version of TFEB. That accumulation was associated with an
increase in markers of autophagy, including increased autophagosome, autolysosomes as well as increased lysosome biogenesis. There was also a marked increase in lifespan among the treated nematodes of between about 15 and 45 percent.

The next step was to see if there were drugs that could mimic the effect of the gene inhibition used in the screening experiment. The researchers found that selective inhibitors of nuclear export (SINE), originally developed to inhibit XPO1 to treat
cancers, had a similar effect - increasing markers of autophagy and significantly increasing lifespan in nematodes. The researchers then tested SINE on a genetically modified fruit fly that serves as a model organism for the neurodegenerative disease ALS.
Those experiments showed a small but significant increase in the lifespans of the treated flies.

As a final step, the researchers set out to see if XPO1 inhibition had similar effects on autophagy in human cells as it had in the nematodes. After treating a culture of human HeLa cells with SINE, the researchers found that, indeed, TFEB concentrations
in nuclei increased, as did markers of autophagic activity and lysosomal biogenesis. "Our study tells us that the regulation of the intracellular partitioning of TFEB is conserved from nematodes to humans and that SINE could stimulate autophagy in humans.
SINE have been recently shown in clinical trials for cancer to be tolerated, so the potential for using SINE to treat other age-related diseases is there."

Is the Architecture of the Nuclear Envelope Fundamental to the Evolution of Aging?
https://www.fightaging.org/archives/2018/05/is-the-architecture-of-the-nuclear-envelope-fundamental-to-the-evolution-of-aging/

Hydra are functionally immortal, given a suitably static environment. They exhibit continual proficient regeneration, and their mortality risk is low and constant over time. As a species they appear near unique in this. Why is aging and imperfect
regeneration almost universal among species? One explanation is that environmental change gives aging species an advantage: non-aging species can certain emerge in eras of comparative environmental stability, but will be out-competed when the environment
shifts. Other explanations involve the more complex structure in higher species, particularly in the central nervous system, where data must be stored as lasting molecular and cellular structures. Long-term persistence of fine cellular structure and
proficient, continual regeneration don't go well together.

This study looks at the complexity and structure of the nuclear envelope inside cells as a possible dividing line between the few immortal species such as hydra and all of the others. The authors propose that increased complexity of the nuclear structure,
and thus its greater vulnerability to certain kinds of molecular damage known to be associated with aging, limits the degree to which longevity and highly proficient regeneration can evolve - though I think that this is certainly something that could be
argued either way, and at length.

The freshwater polyp Hydra represents a rare case of an animal with extreme longevity. It demonstrates unlimited clonal growth with no detectable signs of senescence, such as age-dependent increase in mortality or decrease in fertility, and thus is
considered as non-senescent. Hydra body is made of cells of three lineages, originating from unipotent ectodermal and endodermal epithelial stem cells, and from multipotent interstitial stem cells. In contrast to most other animals, stem cells in Hydra
indefinitely maintain their self-renewal capacity, thus sustaining non-senescence and everlasting asexual growth.

While unlimited self-renewal capacity of the stem cells is long recognized fundamental for Hydra's non-senescence, the underlying molecular mechanisms remain poorly understood. So far, the transcriptional factor FoxO was found as critical regulator of
Hydra stem cell homeostasis and longevity, supporting the view that components of the insulin/insulin-like growth factor signaling pathways govern lifespan throughout the animal kingdom. Several other transcriptional factors are supposed to contribute to
the non-aging of Hydra. However, the putative effector molecules downstream from these transcriptional factors that might contribute to the sustained stem-cell activity and non-senescence in Hydra remain unclear.

Studies in bilaterian animals propose proteins of the Lamin family to be the major effector molecules involved in the age-related cellular senescence and, hence, in the genetic control of ageing and lifespan. These highly conserved intermediate filament
proteins form a complex network at the inner nuclear membrane, arrange the nuclear architecture and orchestrate multiple nuclear processes, such as DNA replication and repair, chromatin condensation, and transcription. Importantly, bilaterian cells are
highly sensitive to the nuclear lamina disturbances. Decline in the expression level of Lamin B1 and increase of an aberrant Prelamin A isoform are associated with the age-dependent alterations in the nuclear lamina morphology and chromatin organization
observed upon physiological ageing in mammals and invertebrates.


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