Fight Aging! Newsletter, January 8th 2018

Sarcopenia is a geriatric syndrome with a high prevalence in older individuals; its presence is estimated in up to 35% of hospital wards. Elderly individuals generally experience a decline of nutrient and energy intake with increasing age. This phenomenon is generally due to age-related loss of appetite, the so-called "anorexia of aging", whose physiopathology is only partly understood. It may also depend on increased energy requirements due to acute or chronic inflammation, leading to "disease-related malnutrition". Malnutrition and sarcopenia often overlap in older patients, so that one of the mainstays of sarcopenia prevention and treatment is promoting adequate nutrition. The prescription of adequate intakes of proteins, vitamin D, antioxidant nutrients, and long-chain polyunsaturated fatty acids has been particularly emphasized in this field, since these nutrients are able to counteract anabolic resistance, promote protein synthesis, and modulate inflammation, thereby preventing its detrimental consequences on muscle cells.

The human gut microbiota is composed of as much as 1014 bacteria, viruses, fungi, protozoa, and Archaea, with a gene pool 150 times larger than that of the host. It establishes a symbiotic relationship with the host, whereby individual environmental and genetic factors can shape its composition, while the host physiology is influenced and gets adapted to its presence. In healthy individuals, the gut microbiome generally includes between 1100 and 2000 bacterial taxa, most of which cannot be cultivated with traditional microbiological techniques.

Geographical location and diet are the major environmental factors explaining the interindividual differences in healthy gut microbiota composition. After the age of 65, gut microbiota resilience is generally reduced, so that its overall composition is more vulnerable to lifestyle changes, drug treatments such as antibiotics, and disease. As a result, species richness (i.e., the number of taxa that metagenomic analyses are able to identify) is reduced, and interindividual variability is enhanced. A lower number of species, decrease in the representation of taxa with purported health-promoting activity, and expansion of Anaerotruncus, Desulfovibrio, Coprobacillus and Gram-negative opportunistic pathogens are the most important changes that have been demonstrated in different clinical settings. These distinctive features of older persons' gut microbiome allow hypothesizing its involvement in the aging process with multiple mechanisms.

In a pioneering study on the ELDERMET cohort, it was demonstrated that the species richness of the fecal microbiota of older subjects is inversely related to physical performance. A secondary analysis of the same cohort has recently revealed that, in community dwellers, the presence of frailty, as measured through the Barthel Index (BI), is associated with a gut microbiome profile similar to that typical of nursing-home residents, with an increased representation of Anaerotruncus, Desulfovibrio, and Coprobacillus. These results are not merely speculative; they have important clinical correlates. For example, gut microbiota dysbiosis can be associated with a reduced survival in older individuals with frailty or disability. Moreover, the over-representation of opportunistic pathogens in the gut microbiota of frail multimorbid older patients may also increase the risk of developing infections.

However, these studies do not establish any cause-effect relationship between gut microbiota dysbiosis and physical frailty, due to their cross-sectional design. Several compounds produced or modified by the gut microbiota can enter systemic circulation and ultimately influence skeletal muscle cells. For example, a healthy gut microbiota is able to produce significant amounts of folate and vitamin B12, which may improve muscle anabolism. The most studied putative mediators of the effect of gut microbiota on skeletal muscle function are short chain fatty acids (SFCAs). These substances are generally derived from the bacterial metabolism of nutrients, such as proteins, which are introduced with diet. Their main host targets are skeletal muscle mitochondria.

The only intervention study carried out on older patients and targeted at exploring the effects of gut microbiota modifications on skeletal muscle outcomes involved the administration of prebiotics, i.e., substances promoting the overexpression of beneficial bacteria. In a randomized controlled trial, researchers enrolled 60 older patients who received treatment with a prebiotic formulation including fructooligosaccharides and inulin versus placebo for 13 weeks. Surprisingly, the treatment group experienced improvement in two outcomes of muscle function: exhaustion and handgrip strength. Thus, these data support the hypothesis of a modulation of muscle function by gut microbiota. Unfortunately, no other studies have explored this field to date.

The current state-of-the-art literature supports the hypothesis that gut microbiota may be involved in the onset and clinical course of sarcopenia. Since nutrition is one of the main determinants of gut microbiota composition, and is also involved in the pathogenesis of sarcopenia, the gut microbiota may be at the physiopathological cross-road between these two elements. Some key microbial taxa may have a relevant role in determining muscle structure and function by producing metabolic mediators that influence the host physiology after intestinal mucosa absorption. Glycine betaine, tryptophan, biliary acids, and SCFA, namely butyrate, are the most promising of these putative mediators.

Donor age is one of the major concerns in Bone Marrow Transplantation (BMT). Studies on murine system have demonstrated that aged marrow harbors increased pool of hematopoietic stem cells (HSCs) exhibiting myeloid bias and having compromised competitive repopulating ability. Aged HSCs also exhibit multiple epigenome and transcriptome changes. DNA damage, replication stress, and ribosomal stress have been shown to cause aging of HSCs. Age-associated changes in human HSCs were similar to those observed in mouse HSCs, suggesting that hematopoietic aging is an evolutionarily conserved process. A retrospective study done in BMT patients showed age as the only donor trait associated with their overall and disease-free survival.

Since donor age is such an important concern in BMT, it might be argued that the upper limit of donor age may be lowered. However, patients having an older individual as the sole matched donor could be denied access to this potentially life-saving treatment. To overcome this impediment, efforts are being made to rejuvenate aged HSCs to improve their performance. Here we report a novel finding that a brief exposure of aged HSCs to young mesenchymal stromal cells (MSCs) rejuvenates them via intercellular transfer of microvesicles (MVs) containing "youth signals". We also demonstrate that intercellular transfer of aged exosomes carrying negative regulators of autophagy causes aging of HSCs. Our data are relevant in both allogenic as well as autologous transplantations involving older individuals as donors and recipients, respectively.

Rejuvenation of aged HSCs prior to transplantation could expand the donor cohort and also help older individuals undergoing autologous stem cell therapy. Application of the MSCs as well as MVs in clinical BMT/SCT might be logistically straightforward, since they can be cryopreserved as "ready-to-use" reagents. Use of pharmacological compounds to rejuvenate aged stem cells in general, and aged HSCs in particular, is being pursued to gain clinical advantage. However, most pharmacological compounds could show off-target effects and they also regulate diverse processes and pathways. Therefore, use of clinical grade "cellular products" in manipulating HSCs, albeit expensive, would be a safer approach than the direct application of pharmacological tools on them.

Reduced autophagy is associated with aging, whereas stimulation of autophagy is speculated to have anti-aging effects. Aged HSCs having high autophagy levels are known to preserve their regenerative capacity. Here we provide a direct evidence for this hypothesis. We demonstrate that young MSCs transfer autophagy initiating mRNAs to the aged HSCs via intercellular transfer of MVs, leading to their rejuvenation. ATG-7 is a critical component of the autophagy pathway and has been shown to be essential for the maintenance of human CD34+ HSCs. Here we show that young MSCs and their MVs transfer Atg7 to aged HSCs.

FOXO3a has been linked to longevity in multiple population studies. FOXO3a is known to stimulate autophagy in primary mouse renal cells. Similarly, FOXO3a inhibition or depletion prevents autophagy induction by starvation in vivo in mouse muscle, confirming a strong link between transcription factors of the FOXO family and autophagy. We found that aged HSCs treated with young MVs show high levels of FOXO3. In the light of these reports, our data clearly demonstrate that direct transfer of MVs containing autophagy-inducing mRNAs seems to be one of the important mechanisms involved in rejuvenation of aged HSCs by young MSCs.

Myeloid bias of HSCs has been considered as a hallmark of their aging. This has been attributed to an accumulation of myeloid-biased HSCs in the aged marrow. In transplantation between old and young individuals, microenvironment-mediated myeloid skewing has been demonstrated. Here, we report a novel finding that myeloid bias of aged HSCs could also be a non-cell-autonomous process involving intercellular communication mechanisms. We demonstrate that aged MVs contain higher levels of Itga2b, which is a myeloid commitment marker, whereas young MVs contain higher levels of IL7r, which is a lymphoid commitment marker. Importantly, we show that partitioning of these mRNAs depends upon the levels of activated AKT in the stromal cells.

Thus, a continuous transfer of aged MVs containing Itga2b to the HSCs could impose a myeloid bias in them, and this coupled with their low levels of apoptosis, could lead to the accumulation of aged HSCs in the marrow. Our data strongly suggest that the lineage bias of HSCs could be dictated by the mRNA profile of the MVs transferred to them, which in turn depends on the signaling mechanisms prevailing in the stromal cells. This aspect needs further investigation. Nonetheless, our findings have certainly added a new dimension to the existing academic debate.

Aubrey de Grey, Chief Science Officer, presents the SENS Research Foundation's current research into therapies that may add decades of healthy life for people who are adults today, as well as work that the Foundation has already spun out into successful startups. Dr. de Grey also explains how SRF's work fits within the context of the global anti-aging research effort and why it has gained broad expert support.

The fertility of women decreases with maternal aging, resulting from various kinds of reasons including decreased follicle number, altered reproductive endocrinology, increased reproductive tract defects, decreased embryo quality, and impaired oocyte quality. Among the possibilities, decreased oocyte quality with maternal aging is the main reason because oocyte donation from young women could rescue the low live birth rate in elder women. With maternal aging, both the nuclear maturation and cytoplasmic quality are affected, and oocyte aneuploidy arising from chromosome segregation error increases dramatically. The obvious change in ooplasm with maternal aging is mitochondrial dysfunction.

It is well known that mitochondria function in energy production and apoptosis in cells. As the most prominent cell organelles in oocytes, mitochondria play pivotal functions and determine the developmental competence of oocytes. With advanced maternal age in women, the most common aberrations in mitochondrial structure are mitochondrial swelling and cristae disruption. Mitochondria are the main source of ATP through oxidative phosphorylation in mammalian oocytes. It is reported that reduced ATP content and metabolic level could be detected in aged oocytes, which would affect oocyte quality and embyogenesis. Mitochondrial malfunction is highly related with defects in spindle organization, cell cycle progress and chromosome segregation in oocytes of aged women and mice. Mitochondrial dysfunction is a major contributing factor for negative outcomes in IVF in general, especially in women of advanced maternal age. The findings reminded the researchers that mitochondria supplement or replacement in oocytes might be a possible strategy for infertility treatment in elder women.

The mitochondria replacement by transfer of heterologous ooplasm, germinal vesicle, spindle, polar body, or pronuclei has been tested in animals and humans to improve developmental potential of aged defective oocytes or to prevent trans-generational mitochondrial disease transmission, but clinical translation of these techniques requires further validation for their efficacy and safety. Especially, the compatibility between donor and recipient mitochondrial DNA and mitochondrial heteroplasmy are still a concern. Transfer of autologous mitochondria from cumulus and granulosa cells were tested for oocyte quality rescue, but it is worth noting that cumulus and granulosa cells age similarly to oocytes. We supposed that autologous adipose tissue-derived stem cell (ADSCs) might be an ideal mitochondrial source for rescuing oocyte quality and fertility. In our study, we found that supplement of autologous ADSC mitochondria could improve oocyte quality, embryogenesis, and fertility in aged mice. We propose that autologous ADSC mitochondria supplement may be a promising strategy for fertility retrieval in women with advanced reproductive age.

Many people would tell you that death - or rather, aging - wasn't around until we started reproducing sexually. There's no reason sexual recombination in itself would demand our death, however. In fact it clearly doesn't: we know of two species of worm which reproduce by splitting themselves lengthwise and fusing together, who are nonetheless no likelier to die in old age than in youth. The famously immortal Hydra is also capable of reproducing sexually (although it usually chooses budding instead). On the flip side, there are multiple organisms who produce eggs asexually, but aren't any safer from senescence than we are.

The rather disturbing truth about life and death is that our bodies are just disposable vessels for the replicators cushioned safe and snug within our germ cells. While a few organisms like the Hydra kept things simple by remaining as one with their germline, others built free-standing bodies of somatic cells with ever more complex machinery to house and propagate a germline that was reincarnated each generation inside a new (and hopefully improved) body. The resulting collections of meat and bone eventually became complex enough to totally obscure the germline itself, and conducted lives with apparent independence - humans in particular enjoyed millennia of ignorance about our fundamental irrelevance. But despite its obscurity, the consequences of its influence could hardly escape notice, for one simple reason: once the germline had abandoned the body, we were all condemned to death.

Key to this fate was the fact that you would, at some point, probably die anyway. Maybe you'd starve in a famine, or be eaten by the resident apex predator, or just freeze to death. But whatever the cause, you were always less likely to live two years than to live one. In this way, each individual's reproductive potential was concentrated at the beginning of his life, and declined at some rate after sexual maturity. The most crucial task was getting you to reproductive age at all, and consequently a lot of selection power had to be spent on birth and development. By contrast, ages that organisms were rarely capable of reaching would experience extraordinarily little selective pressure, the bottom of a genetic slump that began very soon after reproductive maturity. The problem wasn't just that beneficial alleles in old age weren't selected for; it was that alleles with damaging effects later in life could curry favor by increasing fitness at the ages of highest reproductive potential - an effect known as "antagonistic pleiotropy".

Exactly how bad a deal you, the body, get is dependent on your niche. How dangerous is it? How long are you likely to survive? If you're a wild mouse, your chances are around 10% in the first year. There's no sense in spending a long time on growth and development, and you'd better have lots and lots of children, because most of them will die. On the opposite end of the spectrum are organisms that live in relative safety. They may live on an island with no predators, such as a particular strain of opossum with unusually long life, or have an unusually reliable food source. Humans, for our part, didn't come out as bad as we could have. We're among the longer-lived species, and there's a good chance that apparently unrelated medical advances in the last century have been pushing us in the direction of a slower intrinsic aging rate.

Confronting failure in trying to stem symptomatic Alzheimer's disease (AD), the field's main thrust has turned toward retooling its drug trials for ever-earlier disease stages. To sustain enthusiasm among participants and sites, scientists were advised to focus on learning from failure rather than conveying a sense of nihilism, both in internal discussions and in speaking with reporters.

They got to practice a positive attitude when another setback hit home. Merck announced an end to the EPOCH mild to moderate AD trial of the BACE inhibitor verubecestat for lack of efficacy, and data released later indicated that the drug had nudged down amyloid plaques without a hint of benefit, even in the more mildly symptomatic participants. Even so, BACE inhibitors are very much alive and being evaluated to a collective tune of billions in funding. Researchers believe EPOCH treated people too late, when they'd had brain amyloid for years and neuron loss was well underway. The hope now rests on Phase 3 trials in people with mild AD. In trials of anti-amyloid antibodies, the place to go in 2017 was up. The A4 trial joined the trend in Alzheimer's immunotherapy when it quadrupled the solanezumab dose and extended treatment time to five years. Solanezumab had shown hints of efficacy in its negative Phase 3 trial, suggesting a higher dose might work.

Two different α-synuclein antibodies advanced to Phase 2. Researchers desperately want biomarkers for the next round of α-synuclein trials, and the race is on for PET tracers that will detect it. This work currently plays out on the Parkinson's disease (PD) front, but tracers and therapeutics for this protein will come in handy in Alzheimer's and dementia with Lewy bodies as well, as they will help scientists dissect the significant overlap of pathology and symptoms across the AD-PD spectrum.

Biomarkers will continue to be a research priority until they are solidly in place as routine features of AD diagnoses and trials, and 2017 saw strides toward that end. Researchers improved the standardization of cerebrospinal fluid (CSF) amyloid and tau measurement with automated CSF assays that vary less between runs and can predict clinical progression in cognitively normal people. Ultimately, clinicians prefer to use blood over CSF, and this year saw the first signs that this may be possible. Trialists all over the world seek a blood-based indicator of brain amyloid deposition to help them cut down on the number of expensive amyloid PET scans currently needed to recruit for secondary prevention trials.

Whatever doubt might have lingered out there about microglia's role in Alzheimer's was put to rest when scientists fingered a protective polymorphism near the gene for a major microglial transcription factor. Called PU.1, it controls myriad responses, including expression of known AD genes. This protective variant reduces PU.1 expression, lowers amyloidosis, and delays onset of AD. In a bizarre twist, 2017 ended on news that microglia not only help clear amyloid plaques, they may also help seed them. Some activated microglia spew protein bundles that power inflammatory cascades and also latch onto amyloid, driving plaque assembly.

The gradual sickening and eventual death of neurons defines neurodegenerative disease, but how exactly do disease-related proteins do this to neurons? A single theme did not emerge from this line of research in 2017; rather, it seems toxic proteins have an arsenal of weapons at their disposal. Tau appears to mess with all manner of cellular functions. Researchers implicated toxic tau variants in mitochondrial dysfunction, bungling synaptic vesicle release, disrupting the nucleus, compromising the epigenome. No one mechanism rose to the fore, however.

Vascular dementia research used to be a slow backwater relative to the flow of data every year on AD, but 2017 was different. Researchers made inroads into the physiology underlying this disease, for example by toppling a long-held dogma with their demonstration that the human brain does have a lymph system. The finding comes two years after a dural lymph system was discovered in mice. Continuing this year, the rodent studies reported that the dural lymph vessels drain cerebrospinal fluid from the brain into the blood stream. Besides the excitement about lymphatics, there was buzz about the regulation of blood flow in the brain. Researchers found that microinfarcts shut down local clearance of amyloid from the brain, at least in mice. These tiny, "silent" strokes are known to occur in people with AD, and the findings suggest they could hasten amyloid buildup by blocking clearance.

Obesity continues to weigh on the blood-forming stem cell compartment, altering the balance of the cell types produced there, even after the body sheds excess weight. Under the stress of obesity, hematopoietic stem cells (HSCs) begin to overexpress a regulatory gene that tilts blood production toward myeloid cells, and may even promote preleukemic fates. This shift in gene expression, which worsens over time, results in lasting dysregulation, even if HSCs are transplanted into a normal environment.

Although these findings come from a study that relied on a mouse model of obesity, they raise questions about the use of HSCs isolated from obese people in therapeutic transplant procedures. "Little is known about how obesity in marrow donors could affect the quality of the hematopoietic stem cell compartment. We want to better understand the molecular alterations in obesity to predict potential risks associated with the therapeutic use of stem cells isolated from obese donors."

The research team traced the dysregulation of the HSC compartment to altered expression of Gfi1, a transcription factor. "Mechanistically, we establish that the oxidative stress induced by obesity dysregulates the expression of the transcription factor Gfi1 and that increased Gfi1 expression is required for the abnormal HSC function induced by obesity. These results demonstrate that obesity produces durable changes in HSC function and phenotype and that elevation of Gfi1 expression in response to the oxidative environment is a key driver of the altered HSC properties observed in obesity."

Although the effects of chronic organismal stresses are still poorly understood, research is showing that age and environmental stresses can lessen the healthy diversity of cells in our blood-making machinery. "There is now an understanding that the blood stem cell compartment is made up of numerous cell subsets, Keeping this compartment healthy is essential to human health. This includes maintaining the diverse pool of blood-making stem cells needed to produce blood cells the body needs to function properly."

As you get older, the chance that you will die goes up. As you get older, the chance that you will die from certain diseases also goes up. Why does this happen? A simple explanation would be that, like an old car, you accumulate damage in a random fashion. However, there are many simple things that we can do to make animals live longer. Why? We don't really know. Eating less makes mice live longer. Some genes, when mutated, make mice live longer. A few drugs, approved for human use, also make mice live longer. So what is the study of aging? I sum it up as the following: trying to figure out what kinds of damage accumulate with age, how to reverse that accumulation, and the search for switches that we could flip in human biology to increase lifespan.

In the 1930s, investigators wanted to do an experiment to see if stunted growth rates during the Great Depression might impact lifespan. They tested this in rats by feeding them less food than they would normally eat. To their surprise, this actually made the rats live longer! This was a seminal discovery. For the first time, we changed the environment of an animal to make it live longer than it normally would. Since then, investigators have tried to uncover how this works. While long-term human studies are sparse, investigators have run two caloric restriction experiments in monkeys, one of which showed promising results for an increase in survival.

In papers published in 1983-1993, investigators introduced the concept that a gene could control lifespan. Previously we'd known that caloric restriction could make animals live longer, but scientists found mutant genes that could make worms live longer. The first gene found encoded a protein that is similar to insulin-like growth factor and insulin receptors in humans. In mice, mutating members of both of those pathways can increase lifespan. One of the longest-lived mouse mutants we have today is a dwarf mouse. In one study, people with similar dwarf mutations seemed to suffer less age-related disease than their non-mutated relatives.

A paper published in the 70's showed that linking old and young female mice so that they share a bloodstream increased lifespan. Then, in 2011, a succession of papers came out showing that this procedure and others like it made mice better at remembering things, and improved heart and muscle function with age. These discoveries increased excitement and interest in the field, and lead to a wave of startups. Investigators in the field have proposed many possible causes for this phenomenon. Proteins, small vesicles, or cells in the young mouse cleaning the blood of the old mouse might all be part of the effect. Many companies are trying to figure out whether there is a special protein or molecule involved.

As you get old, so do your cells. But some of your cells get old in a way that is much worse than the others. If the cell refuses to die even when it stops working, and starts secreting signals to the immune system, we call that a 'senescent cell'. What happens when you get rid of these cells? Investigators found that getting rid of senescent cells in normal mice made them live a longer healthy lifespan. Knocking out senescent cells is tricky, because they don't have many unique identifiers. Companies are working to either find things empirically that kill senescent cells, or figure out specific mechanisms by which to try to destroy them.

Your body makes a lot of junk, on the molecular level, and cells need to clean this up. Just increasing the expression of one protein that helps to clean up this junk was enough to make mice live ~17% longer. Cells recycle old proteins and other molecules into a big vesicle, called a lysosome. It contains many proteins, and their job is to chop up old cell parts that it engulfs. Genes for proteins that do work in the lysosome are mutated in diseases such as Parkinson's. So improving this process has immediate relevance to neurodegenerative disease. As the lysosome gets older, more junk builds up in it that it cannot degrade. Finding ways to make more lysosomes, or help lysosomes degrade junk, may be interesting therapeutic avenues to pursue.

You may have heard mitochondria referred to as the 'powerhouses' of the cell. One concept that comes up when people talk about mitochondria is 'oxidative stress' - the idea that if molecules are very reactive, they are likely to interfere with a lot of other molecules in the cell that should be left to their own devices. Weirdly, the story has turned on its head over time. It's true that it is bad to pump an animal full of reactive oxygen species, and that you can make a mouse live longer by increasing the level of proteins that are supposed to clean up mitochondria. But you can also mutate things that should be helping the mitochondria, and end up increasing lifespan! It's counterintuitive, and one hypothesis is that a little bit of stress is good because it forces your cells to put up their defenses and ramp up production of molecules that neuter the reactive oxygen species. But we don't really know.

Researchers have cultured the first lab-grown skin tissue complete with hair follicles. This skin model, developed using stem cells from mice, more closely resembles natural hair than existing models. Although various methods of generating skin tissue in the lab have already been developed, their ability to imitate real skin falls short. While real skin consists of 20 or more cell types, these models only contain about five or six. Most notably, none of these existing skin tissues is capable of hair growth.

Researchers originally began using pluripotent stem cells from mice, which can develop into any type of cells in the body, to create organoids that model the inner ear. But the team discovered they were generating skin cells in addition to inner ear tissue, and their research shifted towards coaxing the cells into sprouting hair follicles. The team's recent research demonstrates that a single skin organoid unit developed in culture can give rise to both the epidermis (upper) and dermis (lower) layers of skin, which grow together in a process that allows hair follicles to form the same way as they would in a mouse's body.

While the researchers were unable to identify exactly which types of hairs developed on the surface of the organoid, they believe the skin grew a variety of hair follicle types similar to those present naturally on the coat of a mouse. The skin organoid itself consisted of three or four different types of dermal cells and four types of epidermal cells - a diverse combination that more closely mimics mouse skin than previously developed skin tissues. By observing the development of this more lifelike skin organoid, the researchers learned that the two layers of skin cells must grow together in a specific way in order for hair follicles to develop. As the epidermis grew in the culture medium, it began to take the rounded shape of a cyst. The dermal cells then wrapped themselves around these cysts. When this process was disrupted, hair follicles never appeared.

After discovering this recipe for lab-grown hair follicles, the researchers must now work to overcome a new roadblock in the study of in vitro hair development - physical limitations that prevent the hairs from shedding and regenerating. The shape of the tissue in culture causes the hair follicles to grow into the dermal cysts, leaving them with nowhere to shed. Nonetheless, the team thinks the mouse skin organoid technique could be used as a blueprint to generate human skin organoids.

Immunotherapy, which helps the body's immune system attack cancer, has revolutionized treatment for cancers such as melanoma and leukemia. However, many other kinds of cancer remain resistant. A new study suggests that a combination of two immunotherapies (oncolytic viruses and checkpoint inhibitors) could be much more successful in treating breast cancer and possibly other cancers. "It was absolutely amazing to see that we could cure cancer in most of our mice, even in models that are normally very resistant to immunotherapy. We believe that the same mechanisms are at work in human cancers, but further research is needed to test this kind of therapy in humans."

The researchers studied three mouse models of triple negative breast cancer, and found that all were resistant to a checkpoint inhibitor which is commonly used to treat other kinds of cancer. They also found that while an oncolytic virus called Maraba could replicate inside these cancers and help the mouse's immune system recognize and attack the cancer, the virus alone had minimal impact on overall survival.

The researchers then tested the virus and checkpoint inhibitor together in models that mimic the metastatic spread of breast cancer after surgery, which is very common in patients. They found that this combination cured 60 to 90 percent of the mice, compared to zero for the checkpoint inhibitor alone and 20 to 30 percent for the virus alone. In these models, the virus was given before the surgery and the checkpoint inhibitor was given after. "When you infect a cancer cell with a virus, it raises a big red flag, which helps the immune system recognize and attack the cancer. But in some kinds of cancer this still isn't enough. We found that when you add a checkpoint inhibitor after the virus, this releases all the alarms and the immune system sends in the full army against the cancer." Ongoing clinical trials are testing oncolytic viruses (including Maraba) in combination with checkpoint inhibitors in people with cancer.

Aging is a complex multifactorial process, meaning that multiple pathways need to be targeted to effectively prevent or slow aging. A number of molecular pathways are well known for influencing aging, but only a few have been successfully targeted with individual drugs, and these drugs do not individually target all aging pathways. However, combinations of these drugs might have the potential of effectively broadening the scope of aging targets. There are a number of drug combinations that could be combined based on different but overlapping pharmacological activities. Since the number one criterion for selecting drugs should be based on known anti-aging effects, for example, in preclinical mouse studies, the number of drugs available to consider is markedly reduced. Three drugs with well-validated anti-aging effects in laboratory animals, rapamycin, acarbose, and SS31, are well suited to therapeutic multiplexing as a way to enhance healthy aging and stop the development of lesions associated with aging and physiological dysfunction based on interactive cellular mechanisms of each drug.

The concept of drug multiplexing to slow aging looks good on paper, but drug combinations have yet to be tested in any meaningful way. Historically, testing single drugs in mouse lifespan studies has provided useful information, but it is costly and time consuming. More importantly, lifespan studies are difficult to recapitulate in humans, making translation of the preclinical information challenging. And especially relevant is the fact that lifespan studies in mice are not well-suited to testing drug combinations that could more effectively target multiple factors involved in aging. Thus, new paradigms for testing therapeutics aimed at slowing aging are needed.

While the future for expanded use of drug combinations in treating various diseases and conditions, including aging, is highly promising the path toward eventual regulatory approval can be challenging and should be considered in any preclinical studies undertaken. The potential beneficial functional synergy gained from the logical and judicious use of rational drug combinations, such as rapamycin, acarbose, and SS31, is obviously complicated by the fact that different drugs with different metabolic, pharmacokinetic, and toxicity profiles are being superimposed on top of one another. Focusing not just on the benefits of combination products but also the potential liabilities early on can speed the development process.

In summary, the concept of drug multiplexing as a powerful platform to slow aging is promising but has not yet entered the mainstream of aging research. The combination of rapamycin, acarbose, and SS31, three drugs with individually documented anti-aging effects, is a logical approach designed to complement mechanisms of action of their molecular targets and robustly enhance a delay of aging and age-related disease not seen with mono-therapeutic approaches. Support for the preclinical investigation of this drug combination as well as other drug combinations is urgently needed to determine dosages, frequency of administration, and criteria for when to start administering the drugs, i.e. focus on treatment at older ages, or prevention at younger ages.

Senescence is a cellular response to damage characterized by a stable cell cycle arrest and by the secretion of cytokines and other soluble factors with pleiotropic functions, collectively known as senescence-associated secretory phenotype or SASP. The primary role of senescence is thought to be the orchestration of tissue remodeling and repair. This has been demonstrated in a variety of settings, including tissue repair in the skin and liver. In general, senescent cells are efficiently cleared as part of a successful tissue repair process. However, upon severe or chronic damage, senescence-orchestrated tissue repair may fail and senescent cells may accumulate, contributing to disease and aging.

The power of cellular senescence in inducing tissue remodelling has been further extended to processes of cellular reprogramming in vivo. The transgenic expression of the four transcription factors abbreviated as OSKM (Oct4, Sox2, Klf4, and c-Myc) in adult mice induces dedifferentiation and cellular reprogramming within multiple tissues. However, in addition to reprogramming, the activation of OSKM also results in cellular damage and senescence. Therefore, OSKM induces two opposite cellular fates, namely senescence and reprogramming, that coexist in vivo in separate, but proximal, subsets of cells. Importantly, it has been demonstrated that senescence plays an active role in facilitating in vivo reprogramming through the paracrine action of the SASP, with interleukin-6 (IL6) as a critical mediator. Of note, IL6 plays an important role also during in vitro reprogramming. Moreover, the concept that senescence promotes cellular plasticity has been further extended to the activation of somatic stem/progenitor cells. In particular, the SASP can confer somatic stem/progenitor features onto proximal epithelial cells in several tissues.

The tumor suppressor genes p53, p21, Ink4a, and Arf act as cell-autonomous barriers for cellular reprogramming. These barriers are conceivably activated by cellular damages associated to reprogramming, most notably replication stress, which result in proliferation arrest and, consequently, inhibit reprogramming. At the same time, p53 and the genetic locus Ink4a/Arf also affect reprogramming, although in opposite directions, through cell extrinsic mechanisms. In the absence of p53, the induction of OSKM leads to exacerbated damage and senescence in tissues, which results in high levels of IL6 that further enhance reprogramming. The Ink4a/Arf locus plays a complex role in reprogramming: it promotes reprogramming through the paracrine influence of senescence, and, at the same time, it is a cell-autonomous barrier for reprogramming. In vivo, the absence of Ink4a/Arf severely impairs OSKM-senescence, IL6 levels are modestly increased, and reprogramming is very inefficient. Therefore, the positive cell-autonomous impact of Ink4a/Arf deficiency is completely obscured in vivo by the absence of senescence and IL6 secretion. The emerging picture is that tissue damage and senescence provide a tissue microenvironment that is critical for OSKM reprogramming in vivo.

Aging is the major risk factor for numerous chronic diseases and is responsible for the bulk of healthcare costs. To address this healthcare crisis, there is a growing interest in identifying ways to therapeutically target aging in order to prevent, delay, or attenuate multiple age-related diseases simultaneously. A number of therapeutic strategies have emerged. However, a major barrier to clinical trials targeting aging is the prolonged time between intervention and clinical outcomes. For these studies, surrogate endpoints will dramatically improve the economy and timescale in which we can measure the effects of interventions on biological age.

Biological age is defined by the health or fitness of an individual, and lack of age-related diseases, irrespective of their chronological age. Biological age can be quite distinct from chronological age. For example, cancer survivors are biologically older than their chronological age due to exposure to genotoxic agents, while centenarians are frequently biologically younger than their chronological age. A biomarker of biological age in accessible bodily fluids or tissues would be extremely valuable for clinical trials testing antigeronic factors, but also potentially for triaging patients facing onerous therapeutic procedures. Hundreds of studies have aimed to discover age-related changes in circulating factors including metabolites, advanced glycation end-products, exosome content, miRNA, and inflammatory molecules, with varying success.

Here, we identified MCP-1/CCL2, a chemokine responsible for recruiting monocytes, as a potential biomarker of biological age. Circulating MCP-1 levels increased in an age-dependent manner in wild-type (WT) mice. That age-dependent increase was accelerated in Ercc1-/Δ and Bubr1H/H mouse models of progeria. Genetic and pharmacologic interventions that slow aging of Ercc1-/Δ and WT mice lowered serum MCP-1 levels significantly. Finally, in elderly humans with aortic stenosis, MCP-1 levels were significantly higher in frail individuals compared to nonfrail. These data support the conclusion that MCP-1 can be used as a measure of mammalian biological age that is responsive to interventions that extend healthy aging.