Thank you very much for the kind introduction and for the invitation to speak to all of you at this event. It was a great event last year and I'm very excited that we extended it to a two day event this year. I am thrilled to be back. We brought a whole army of our staff, up there in the back, so I'm very excited. Before I dive in, a couple of housekeeping things. Everything I do is for-profit, so financial disclosures: I have a financial interest in everything I am about to talk about. I also need to acknowledge the wonderful team that assisted with building out the research program that I'm going to describe, in particular one of our grad students is in the audience, and a lot of the figures in the work I'm going to be presenting were the result of his extraordinary efforts, so thank you. Also this program is a spin out of Aubrey de Grey's SENS Research Foundation, and received founding investment from Kizoo Technology Ventures - I think Michael Greve is around here somewhere - but thank you guys for getting this off the ground.

Just to give a very brief overview of Ichor Therapeutics in general: Ichor is a vertically integrated biopharmaceutical company, and we, since 2013, have focused exclusively on diseases and mechanisms of aging. Within the Ichor umbrella we have a variety of portfolio companies. Some are platform technologies, some are single asset plays, all of which are focused on different indications and contributing to the field of anti-aging research. Today we're going to focus on our longest-standing program, LysoClear, which is an enzyme therapy that we're developing for age-related macular degeneration, and the subject of my thesis work as well. Age-related macular degeneration is the leading cause of vision loss in individuals over the age of 50. About 200 million people worldwide have this disease. One of the major problems with this disease is that it robs patients of their high acuity central vision that gives them the ability to interact with the world in a meaningful way. If we think about our own lives, vision is really how we interact. We can hear, we can smell, we move around, and are involved in our everyday lives, but vision is a primary means for that. When you look at a geriatric population, that isn't as mobile, can't hear as well, can't smell as well, the devastating effects of macular degeneration are very real. We hope to put an end to that.

Now when we look at the origins of where macular degeneration originates and manifests, anatomically we're talking about the posterior segment of the eye, the retina, which is responsible for all of our vision. At the very back of the eye there is this little indented part of the retina that is called the macula. The retina lets us see, and the little indented part, the macula, is what gives us that high acuity central vision. If we dive in just a little bit closer, we can appreciate that there is a great microanatomy, a lot of crosstalk between a variety of different cellular layers that are responsible for taking light and allowing us to see.

Starting in the vitreous and moving posteriorly, we have a variety of different neural layers that are responsible for the electrochemical signal transduction that gets sent to the brain and we interpret as vision. Further on, we have a variety of different photoreceptors. Those include the rods and the cones. Those are the cells that get hit by light and can kick off that entire cascade. Now a general rule in biology is that if you have a cell type or a system that is repeatedly stressed, in this case the photoreceptors with light, you need to turnover those systems in order to eliminate the accumulation of damage. So the photoreceptors are supported by very critical phagocytic cell called the retinal pigmented epithelium or RPE. These cells are essential for photoreceptor function and survival. The photoreceptor outer segments are constantly growing towards the upper RPE and the RPE are constantly gobbling off little bits of the photoreceptors in an effort to turn them over.

Like any cells in the body, the RPE cells' phagocytic potential is really based on the lysosome, the organelle that is responsible for degrading things. LIke any other cell in the body, like any other lysosome in the body, a normal healthy lysosome has a plethora of different enzymes that are able to break down all of the little bits of stuff that make up us. However, there are cases where this doesn't function properly. Canonically we're thinking about lysosomal storage diseases. These are congenital diseases where patients are born missing one of those essential lysosomal enzymes. A lot of the time these diseases are lethal in utero, but for the patients who are born with these diseases, the clinical manifestation can be very, very severe. The patients accumulate molecular junk uncontrollably and can't do anything about it. Our central hypothesis for how macular degeneration works is that age-related macular degeneration is an evolutionarily silent lysosomal storage disease, driven by a very specific sort of junk accumulation called lipofuscin. In the context of retinal lipofuscin, it is a combination of retinoid derivatives and lipids. When I say "evolutionarily silent" I mean that in our evolutionary history we haven't lived long enough to accumulate enough of this junk to evolve enzymes able to break it down. But in our view, the onset and progression of macular degeneration is very akin, mechanistically, to what we see in conventional lysosomal storage diseases, and this is going to be a central theme for how we approach treating the disease, and our translational pathway.

Lipofuscin accumulates in the lysosomes of these essential support cells, the RPE. In the very earliest stages of macular degeneration progress, we don't see anything clinically at all, because it is all happening inside of the cell. But at some point in time, lipofuscin accumulates to a level sufficient to promote pathology. It crosses a threshold and problems start to emerge. I've got two types of figures here. The ones on the left are fundus images, which is when you take an ophthalmoscope and look right into the back of the eye and snap a picture. We can see the optic nerve coming through and this little darkened area here is the macula. If we were to take a cut cross-section through the back of the eye and then flip it and look at that cross-section, then that's what these images on the right are. It is called optical coherence tomography, OCT.

As lipofuscin accumulates in the RPE cells, eventually the RPE cells start to become dysfunctional and choke on the accumulating lipofuscin. To cope with the stress, they do what cells need to do - they dump the junk. So the brightest white line here is the RPE layer, and we can see these bumps underneath it. Those are extracellular junk deposits that the RPE are laying down. We call those drusen. So lipofuscin: junk in the cells; drusen: the junk outside the cells that the cells are depositing. We can also see this very easily clinical via fundus imaging, we just look and we see all these little white specks. All of those are drusen in the back of the eye.

Now what is important to note about this, in this early-stage mild form of age-related macular degeneration, we maintain a nice concave morphology of the macula. We also have integrity in the RPE layers, and in the photoreceptors above them. Even though we have this junk being deposited, even though we see these morphological changes happening, usually there is no clinical presentation of this because the photoreceptors that are required to see are not yet disrupted. As the disease progresses, however, eventually the RPE cannot handle the stress burden and the cells start to die, and with them the photoreceptors that rely upon them. So we see by OCT the thinning of the RPE and photoreceptor layers, we see a collapse of the nice morphology of the macula. Because of this thinning, when you look by fundus imaging you can see the choroid, the blood vessel layers at the back of the eye. That presents as geographic atrophy.

This is the intermediate form of age-related macular degeneration, and, collectively, mild and intermediate are termed dry or atropic macular degeneration. Which suggests that there is a wet form. As these cells are dying and having all kinds of problems, it creates a pro-inflammatory environment in which you have complement activation, an immune response, and, importantly, hypoxia. Part of the way that the body attempts to cope with this mess is to create new blood vessels. New blood vessel formation occurs at the choroid, and protruding into the eye. When you have new blood vessels in the body, frequently they are very leaky vessels, so you can have exudate and outright hemorrhage, either into the subretinal space, which can lead to retinal detachment, or into the vitreous itself, which can lead to total blindness. This process of neovascularization is called neovascular or wet age-related macular degeneration, and this is the most severe and advanced form.

So our disease model hypothesis is that lipofuscin, which accumulates inside of the RPE cells, so intracellular stress, drives the accumulation of extracellular drusen. This in turn causes ROS, inflammation, complement activation, hypoxia, and then these stressors eventually lead to the disease state. Our goal at LysoClear is to develop an enzyme therapy that targets lipofuscin at the earliest stages of disease onset and progression. How are we going to go about doing that?

It is first of all important to note that there is no good standard of care for this disease, and, by the way, the dry form is about 90% of patients. Only 10% have the advanced wet form. So for 90% of patients there are no FDA approved drugs. They do have a vitamin formulation that will mildly reduce your risk of progessing, and if you are a smoker you should not be smoking, for this among many other reasons, but it is really not until you get to the really severe form that there is any sort of really efficacious treatment regimen. These are mostly monoclonal antibodies that target the VEGF pathway to inhibit further neovascularization. So there is a huge overwhelming need for more effective treatments for this condition.

I mentioned previously that we're viewing age-related macular degeneration in a manner akin to conventional lysosomal storage disease. So we are wondering if we can borrow some aspects of how we will treat a normal lysosomal storage disease, and with that motif put together a strategy for going after AMD. What is that strategy? Well if your lysosomal storage disease is based on the idea of missing an enzyme in the lysosome, then at the high level it is a simple process: you make the missing enzyme, and you introduce it in such a way that it goes to the lysosome of the target cells. This has been developed and has been approved clinically, and used very successfully by Genzyme. It has been a conventional way to treat lysosomal storage diseases since the 1990s.

The pathways that are used: you make your enzyme, you decorate it in such a way that it gets recognized by a receptor on your target cell, mannose 6-phosphate glycosylation is the canonical delivery path for lysosomal enzymes to reach the lysosome. Mannose receptor is another pathway that is used, and then through an endocytosis pathway your enzyme is able to be selectively delivered to the lysosome. So this is how lysosomal storage diseases are treated. The problem is that we don't have any good human enzymes that are able to break down RPE lipofuscin. We aren't supposed to. We've never evolved them. So we can't rely on human enzymes to facilitate this junk removal. Instead we have to look elsewhere in the world.

That is where SENS Research Foundation (SRF) came in. Some years ago SRF said that this is a problem, and this is the canonical LysoSENS paradigm: here is a junk accumulation, let's look for enzymes that can break it down and remove the junk. By way of wonderful research by John Schloendorn and his team, SRF identified a variety of fungal peroxidases, among other enzymes, that are capable of breaking down certain lipofuscin components. The specific component that they looked at was a molecule called A2E - again, lipofuscin is comprised of a variety of retinoids and lipid derivatives. A2E is one of the best studied and most resistant to degradation, and so that is what SRF used for their assays. They identified a bunch of enzymes that can break down A2E from wild-type sources.

This is where we came in. Our goal was to take this very encouraging proof of concept research and build a program to establish a proof of concept that we could actually remove lipofuscin both within cells and within animals to pave the way for moving into patients for the first time. The first thing we needed to do is to get out of a wild-type system - there is just a lot of variability when dealing with wild-type enzymes. So we moved into a recombinant system to make the best performing enzyme, manganese peroxidase. We expressed that enzyme in the yeast strain Pichia pastoris. The reason for that is that Pichia preferentially mannosylates unlinked glyosylation sites, so Pichia automatically adds those sugars to the enzyme that we need for it to be delivered into the cell.

When we make our enzyme and run a gel, we see a beautiful big smear, which is characteristic of hyperglycosylation, and when we treat our enzyme with PNGase F, an enzyme that selectively cleaves off all of your sugars, we see a nice tightening of the band at the expected molecular weight of about 54 kDa, suggesting that we have an enzyme that is indeed glycosylated. Of course we wanted to know how many sugars we have, so we sent the sugar trees out for glycoanalysis and identified 26 mannose residues on the sugar chain. For context, for those of you who aren't glycobiologists, and neither am I, you probably want two to five mannose sugars minimum to achieve uptake optimally - though depending on the enzyme you can go more or less than that.

So, ok. We have an enzyme, it has the sugars that are required for delivery. The next step is that we need to figure out its selectivity against lipofuscin targets. I mentioned to you that lipofuscin is a hodgepodge of different things, and the hardest part of it to break down are these retinoids, vitamin A derivatives. Unfortunately, it is not just one thing but many things. So we scoured the literature and synthesized all of the major lipofuscin components that we could find. We tested our recombinant enzyme using our local HPLC, and we showed that our enzyme is actually able to break down every single lipofuscin component that we tested. Now this isn't entirely surprising, because these are all chemically related species, so we would have expected the enzyme to work similarly on these different species. We have EC50 variants of 1 μmol all the way up to 20 μmol; interestingly, A2E, the molecule that SRF did their screen with, was the most resistant to degradation by the enzyme. As I move forward and present both the cell based work we did and some of the in vivo work, we are going to use A2E as the readout, quantitatively, and the reason for that it is the easiest to analyze, it is the most resistant to degradation, so we believe we are under-reporting the effectiveness of the enzyme by using the hardest target to degrade.

One of the problems that we ran into very very early with this program is that, go figure, cells outside the body behave a little bit differently than they do when they are in the body. So although RPE cells have mannose receptors in vivo, and this is very well established, the second you take those cells out and grow them in culture, the receptor disappears, which makes it really difficult to use that as a model for update and delivery of your drug. So we had to split the efficacy and the uptake into separate experiments. For efficacy, we used a canonical line of RPE cells, ARPE-19. The graph on the left shows them to be negative by flow cytometry for mannose receptor, that is what CD206 is. So what we did is that we had a layer of these cells, untreated they are high viability. When we added excessive levels of A2E as a stressor, we see that the viability drops, and then we saw that our manganese peroxidase is able to rescue some of that viability when we had A2E loaded cells and treated them with the enzyme.

The way that we delivered the enzyme for these studies is with a lipid reagent called BioPORTER, a way of forcing the enzyme artificially into the cell. So this isn't our proposed delivery system, this is just asking the question of if we could get the enzyme into the cells, would it be able to protect against some of this retinoid toxicity, and indeed it seems that it can. Then of course we did manganese peroxidase by itself just to make sure it wasn't toxic, and we have a bunch of other studies along these lines as well, and we see that it doesn't different in any meaningful way from the untreated cells.

We then had to ask the separate question, which is whether the uptake is able to occur through a mannose receptor dependant manner. The cell line that we used for this is a mouse monocyte line called RAW 264.7, which is known to have an intact mannose receptor and mannose receptor endocytosis pathway. For this study we started off with mannosylated bovine serum albumin that was flourescently conjugated and we added that to the cells and looked by flow cytometry for an increase in fluorescence. That is the solid pink line here. So we see over time an increase in fluorescence, which we interpret as internalization of the enzyme by our target cell. When we introduce the competitive inhibitor mannan, which also binds to mannose receptor, we see an attenuation of that effect size, suggesting that we're impairing the uptake, and that this is in fact a mechanism by which the enzyme is entering cells. We then did the exact same thing for our recombinant manganese peroxidase, again we think it has mannose because we checked that, and we also fluorescently labelled it. In much the same way we see an increase in the fluorescence of the enzyme, and we can reduce that signal by competing with mannan, suggesting that the increase in fluorescence is mediated by mannose receptor endocytosis.

Next we really wanted to get to this proof of concept point: are we able to remove or degrade existing lipofuscin components like A2E in vivo. There has been a lot of work showing that you can kind of shovel it around or maybe reduce the rate of its accumulation, but no-one to my knowledge has ever shown that you can actually get rid of it and break it down. So that is really what we wanted to get to as a primary inflection point. We ran an intravitreal pharmacokinetics study, and this is just where we injected the vitreous with our enzyme, did sampling measured by ELISA, and identified a half-life of about 9.6 hours. That is great because this is supposed to be a highly targeted enzyme that just goes into these highly phagocytic RPE cells, so we would expect a pretty short half-life. Indeed, we don't want an enzyme that is hanging out there for no particular reason. We want it to be internalized rapidly - versus monoclonal antibody therapies, which are acting on extracellular targets, and where you'd be looking for a half-life measured in days. So we're very excited by that.

We then asked the big question: can we actually have efficacy? To do this, we used a mouse model of age-related macular degeneration, and the juvenile onset form Stargardt's disease. This is an ABCA4-null mouse, and these mice accumulate accelerated levels of A2E. So what we did, we had our mice and we injected them with six doses of either phosphate buffered saline (PBS), low dose enzyme, or high dose enzyme. We see a little bit of a trend, maybe, on the low dose, but nothing significant, and then at the high dose we saw a significant reduction in A2E burden as measured by analytical HPLC.

We wanted to do an intermediate dose, so we redid a separate experiment, where we treated with PBS and an intermediate dose of the enzyme. We did see a statistically significant drop there. When we pull this data together, we see a nice dose-response in our efficacy model in terms of reduction of our molecular target when treated with increasing doses of our enzyme. So collectively we're really excited about this. We've shown that manganese peroxidase is able to break down all lipofuscin fluorophores that we've tested, we've shown that it can be taken up into cells by way of mannose receptor endocytosis, and, most importantly, for the first time to my knowledge, we're actually able to eliminate existing lipofuscin in an in vivo system.

These results were published in December 2018, and later on that same month we successfully closed a financing round for our LysoClear program, to take these very promising lead series, and engineer them into clinical candidates. All of that work is ongoing at Ichor Therapeutics, I'm very excited about it. Our rate-limiting step right now is actually that we need to move into large animal models for safety, toxicity, and so on, to inform a pre-IND meeting with the FDA. I'm super-excited that my large animal vivarium is scheduled to complete construction in the next three weeks, so hopefully we'll have some very exciting data coming out for our next presentations on this topic.

Despite it long having been the consensus that DNA damage and the resulting epigenetic changes are drivers of aging, some recent studies have questioned the importance of mutations in aging. For example, the number of mutations present in aged yeast cells is fairly low, and some genetically engineered strains of mice with high levels of free radicals or mutation rates do not appear to age prematurely, nor do they have shorter lifespans than their wild-type counterparts.

This appears to suggest that mutational load may not have such a strong influence on aging as was once thought, and the researchers of this new study consider further evidence suggesting the same. They also suggest that epigenetic alterations are perhaps the most important driver of aging and that, far from being random in nature, these changes are predictable and reproducible.

Researchers suggest that DNA double-strand breaks (DSBs) are a possible reason for epigenetic changes and show that there are clues to be found in yeast. In yeast cells, DSBs trigger a DNA damage signal that summons epigenetic regulators and takes them away from gene promoters to the site of the DSB on the DNA, where they then facilitate the repair of the break. The researchers suggest that after these repairs, the regulators responsible for repairing the DSBs return to their original locations on the genome, thus turning off the DNA damage signal, but this does not always happen.

The researchers suggest that with each successive cycle of DNA damage response and repair, the epigenetic landscape begins to change and regulators gradually become displaced, reaching a point where the DNA damage response remains active, leaving cells in a chronic state of stress. This stressed state then causes them to become dysfunctional and ultimately alters their cellular identity.

There are numerous hallmarks of aging in mammals, but no unifying cause has been identified. In budding yeast, aging is associated with a loss of epigenetic information that occurs in response to genome instability, particularly DNA double-strand breaks (DSBs). Mammals also undergo predictable epigenetic changes with age, including alterations to DNA methylation patterns that serve as epigenetic "age" clocks, but what drives these changes is not known. Using a transgenic mouse system called "ICE" (for inducible changes to the epigenome), we show that a tissue's response to non-mutagenic DSBs reorganizes the epigenome and accelerates physiological, cognitive, and molecular changes normally seen in older mice, including advancement of the epigenetic clock. These findings implicate DSB-induced epigenetic drift as a conserved cause of aging from yeast to mammals.

Calorie restriction (CR), a dietary regimen in which calories are reduced without causing malnutrition, extends the lifespan of many diverse species and is the gold standard for interventions that promote the health and longevity of mammals. Importantly, CR extends not only longevity but also healthspan. There has therefore been great interest in identifying the physiological and molecular mechanisms by which CR promotes health and longevity.

In mammals fed a CR diet, one of the most striking and broadly conserved effects is improved sensitivity to insulin. Many dietary and pharmaceutical interventions that extend mammalian lifespan and healthspan likewise promote insulin sensitivity, while conversely, there is a well-known association of insulin resistance with diabetes and poor health. Given the central role of the insulin signaling pathway in the lifespan of worms, flies, and mammals, improved insulin sensitivity has been proposed as an essential mechanism by which a CR diet extends mammalian lifespan. While the effects of CR are systemic, some of its most prominent effects are on adipose tissue; CR reduces adiposity in mammals, mobilizing fat stores in white adipose tissue (WAT) while also activating WAT lipogenesis, which is associated with improved systemic insulin sensitivity and metabolic health.

Despite the strong correlative evidence that CR promotes health and longevity through improved insulin sensitivity, there is clear evidence that insulin sensitivity may not necessarily be essential for healthy aging. Several genetically modified mouse models in which insulin resistance has been induced in one or more tissues have extended lifespan, while mice treated with rapamycin, an inhibitor of the mTOR (mechanistic target of rapamycin) protein kinase that extends lifespan, develop insulin resistance in multiple tissues.

Over the last decade, a critical role for mTOR complex 2 (mTORC2) in the control of organismal metabolism has become apparent. In contrast to the well-known mTOR complex 1 (mTORC1), which functions as a key integrator of many different environmental and hormonal cues, mTORC2 functions primarily as an effector of phosphatidylinositol 3-kinase (PI3K) signaling, contributing to the downstream activation of many kinases, including AKT, by insulin. Deletion of Rictor, which encodes an essential protein component of mTORC2, results in insulin resistance in tissues, including liver, adipose tissue, and skeletal muscle. The organismal consequences of inactivating adipose mTORC2 have been unclear.

While an important role for CR-induced insulin sensitivity in the health and survival benefits of CR has long been assumed, the contribution of improved insulin sensitivity to the benefits of CR has not been directly examined. Here, we have tested the role of CR-induced insulin sensitivity on the metabolic health, frailty, and longevity of mice by placing mice lacking adipose mTORC2 signaling (AQ-RKO) and their wild-type littermates on either ad libitum or CR diets. Critically, the insulin sensitivity of AQ-RKO mice does not improve on a CR diet, enabling us to discern the role of CR-induced insulin sensitivity in CR-induced phenotypes. Although the WAT of AQ-RKO mice has a blunted metabolic response to CR and female AQ-RKO mice fed an ad libitum diet have a slightly reduced lifespan, we find that AQ-RKO mice of both sexes fed a CR diet have increased fitness and extended lifespan. We conclude that the CR-induced increase in insulin sensitivity is dispensable for the effects of CR on fitness and longevity.

As a key regulator of vascular physiology, the renin-angiotensin system (RAS) has been implicated in the development and progression of vascular aging. Interruption of the RAS pathway, either by preventing the formation of angiotensin II (Ang II) or by blocking the Ang II type 1 (AT1) receptor, has been proven to be highly successful in retarding vascular aging phenotypes. Meanwhile, inappropriate activation of the RAS, independent of the classic bioactive molecule Ang II, may cause excessive activation of the AT1 receptor and induce chronic inflammation, but how this occurs is not fully understood.

At the end of the twentieth century, a specific autoantibody against AT1 receptor (AT1-AA) was discovered and found to exist in patients with preeclampsia, malignant hypertension, refractory hypertension, and renal-allograft rejection. AT1-AAs could specifically bind to the AT1 receptor and were found to have a receptor agonist-like effect. AT1-AAs were proven to be pro-inflammatory via the transcription factor nuclear factor-kappa B (NF-κB) pathway, thus enhancing the expression of inflammatory factors in endothelial cells (ECs). Moreover, we have previously demonstrated that AT1-AAs induced endothelial damage and contributed to endothelial dysfunction in vivo. Most importantly, AT1-AAs have been reported to accelerate aortic atherosclerosis in mice. A recent study demonstrated that higher AT1-AAs level was associated with inflammation, hypertension, and adverse outcomes. All the above evidence suggests a close relationship between AT1-AAs and vascular aging. Nevertheless, whether AT1-AAs can induce vascular aging or EC senescence has never been explored.

In this study, AT1-AAs were detected in the sera of patients with peripheral arterial disease (PAD) and the positive rate was 44.44% vs. 17.46% in non-PAD volunteers. In addition, analysis showed that AT1-AAs level was positively correlated with PAD. To reveal the causal relationship between AT1-AAs and vascular aging, an AT1-AAs-positive rat model was established by active immunization. The carotid pulse wave velocity was higher, and the aortic endothelium-dependent vasodilatation was attenuated significantly in the immunized rats. Morphological staining showed thickening of the aortic wall. Histological examination showed that levels of the senescent markers were increased in the aortic tissue, mostly located at the endothelium. In addition, purified AT1-AAs-IgGs from both the immunized rats and PAD patients induced premature senescence in cultured human umbilical vein endothelial cells. These effects were significantly blocked by the AT1 receptor blocker. Taken together, our study demonstrates that AT1-AAs contribute to the progression of vascular aging and induce EC senescence through AT1 receptor.

The alterations of the B-cell compartment in aging have been evaluated by contrast to B-cell physiology in young adults. Overall, B-cell generation and function demonstrate large similarities between young mice and humans. In the more detailed mouse context, B cells arise from uncommitted progenitors nested in the bone marrow. Overall, aging disturbs B-cell development in the mouse bone marrow. Strikingly, aging seems to introduce a high mouse-to-mouse variability in early progenitor B cellularity compared to young mice. Impaired B-cell development occurs as a result of affected RAG and SLC expression, as well as decreased sensitivity to IL-7 signals. The in-depth situation in humans remains to be established. Nevertheless, available studies suggest that the amount of B cells decreases, although proportions of progenitor and mature B subpopulations may not be substantially changed in the aged bone marrow.

Various clues point at a role for inflammation in the altered B-cell development in aging, albeit the data is generally based on similarities with acute inflammatory responses. Indeed, pro-inflammatory senescent cells as well as terminally differentiated CD8+ effector T cells accumulate in the bones of old mice and humans respectively. Correspondingly, concentrations of pro-inflammatory molecules or their production by cells are increased in bone marrow during aging. The balance between the negating effects of anti-inflammatory cytokines and the intensity, as well as variety, of pro-inflammatory molecules expressed could contribute to the observed variability in the deterioration of early B-cell development in susceptible aged organisms.

Inflammation can affect the differentiation of multi-lineage hematopoietic progenitors. Thus, aged mouse and human hematopoietic stem cell (HSC) physiology is altered and the output of this compartment reveals a bias against the production of lymphocytes due to the accumulation of stem cells with a propensity to differentiate into myeloid cells. This limits the production of B cells. Persistent inflammation in aging could also compromise the differentiation of B lineage-restricted progenitor cells. A major effect of aging is the repression in B cells of the expression and/or activity of the transcription factors E2A and EBF1, which control the RAG and SLC genes.

Despite the alteration of the B-cell compartment in the bone marrow, the cellularity of mature B cells in the spleen is comparable between aged and young mice. However, this apparent stability masks underlying disparities in the distribution of mature B-cell subsets. The situation in humans appears more difficult to appreciate, since most studies performed analyses based on blood samples, which have an inherent variability and may not reflect mature B-cell representation within tissues. Altogether, the B-cell fraction in the blood of elderly people appears decreased and the proportions of naïve and memory subsets altered. Similar to the bone marrow, various inflammatory molecules could influence the distribution of mature B-cell populations.

In the past two decades, the molecular signatures of aging have been started to be uncovered. A remarkable conservation of these cell signaling pathways has been shown across various invertebrate and vertebrate species. Autophagy is a cellular process that has emerged as a nexus at which these various pathways have been shown to converge. Autophagy is the catabolic process by which the cell eliminates unnecessary cellular components to maintain energy homeostasis and prevent the build-up of toxic material.

Autophagic activity has been shown to decline with age in various animal models. For example, body-wide quantification of autophagic flux in Caenorhabditis elegans revealed a general decline in activity in various tissues, including the intestine and neurons. A similar decline in function has been observed in mammals. For example, electron microscopy analysis of aged mouse livers revealed a depression in the rate of autophagic vesicle formation.

Various groups have identified a necessary role of autophagy in mediating the effects of longevity-enhancing mutations. Inhibiting autophagy in a long-lived mutant model nullifies the longevity-promoting effects of the mutation. C. elegans worms that carry a loss-of-function mutation in their daf-2 gene, which encodes for a common single insulin/Insulin-like Growth Factor (IGF)-1 Receptor in this organism, live significantly longer than their wild-type counterparts. RNAi-mediated knockdown of the autophagy gene bec-1 significantly reduced the lifespan of the daf-2 mutants, clearly identifying autophagy as a process that is required for the increased longevity of this mutant.

To demonstrate a causal relationship between autophagy and longevity, some groups have evaluated the effects of overexpressing autophagy genes. A positive relationship between autophagic activity and lifespan was first demonstrated in Drosophila. Neuron-specific overexpression of the Atg8a gene resulted both in an increase in lifespan and a reduction in the accumulation of toxic protein aggregates in neurons. Similarly, body-wide overexpression of Atg5 resulted in a significant increase in lifespan in mice. Increase in autophagy via disruption of the beclin1-BCL2 complex has been shown to promote both healthspan and lifespan in mice. In summary, autophagy has convincingly been shown to play a pivotal role in healthspan and lifespan extension.

In view of the inability of drugs to slow or reverse the cognitive impairment of Alzheimer's disease (AD) thus far, investigating non-pharmaceutic interventions against AD as a possible alternative is now clearly warranted. Neuromodulatory approaches have consequently emerged and are currently being clinically tested in AD subjects. These approaches include transcranial magnetic stimulation (tMS), transcranial direct current stimulation, and deep brain stimulation. All three approaches provide a generalized stimulatory/inhibitory effect on neuronal activity. The most recent and largest clinical studies involving long-term tMS (Phase III clinical trial) or deep brain stimulation (Phase II clinical trial) in AD subjects have reported minimal or no cognitive benefits.

As the newest neuromodulatory approach against AD, Transcranial Electromagnetic Treatment (TEMT) is very different from tMS because TEMT involves perpendicular magnetic and electric waves emanating away from an antenna/emitter source (rather than magnetic waves radiating from and returning to a conductor in tMS). For our studies, these "electromagnetic waves" are actually within the radiofrequency range (around 1 GHz), which can easily penetrate the human cranium and underlying brain areas.

In a number of pre-clinical studies involving AD transgenic mice, we have administered TEMT daily for up to 8 months. We have demonstrated the ability of TEMT to prevent/reverse both oligomeric and insoluble amyloid-β aggregation - both inside and outside neurons. These TEMT-induced reductions in brain Aβ aggregation are accompanied by brain mitochondrial enhancement and prevention or reversal of cognitive impairment in AD transgenic mice at multiple age. In view of our extensive pre-clinical platform and the aforementioned wide spectrum of human safety studies, clinical trials of TEMT technology in AD were clearly warranted. Therefore, we designed and built a first-of-its-kind head device for administration of TEMT to human subjects in their homes and by their caregivers. The present study reports on safety and efficacy endpoints in an open-label clinical trial to provide daily TEMT to AD subjects over a 2-month period, as well as evaluation at two weeks following completion of treatment.

No deleterious behavioral effects, discomfort, or physiologic changes resulted from 2 months of TEMT. TEMT induced clinically important and statistically significant improvements in the Alzheimer's Disease Assessment Scale-Cognitive, as well as in the Rey Auditory Verbal Learning Test. TEMT also produced increases in cerebrospinal fluid (CSF) levels of soluble amyloid-β, cognition-related changes in CSF oligomeric amyloid-β, a decreased CSF phosphorylated-tau/amyloid-β ratio, and reduced levels of oligomeric amyloid-β in plasma. TEMT administration to AD subjects appears to be safe, while providing cognitive enhancement, changes to CSF/blood AD markers, and evidence of stable/enhanced brain connectivity.

A recently published study reports findings of a study in which researchers compared how much muscle strength older people could muster voluntarily with how much force their muscles put out when stimulated electrically. The results of this research suggest that physical weakness in aging may be due, at least in part, to impairments in brain and nerve function, rather than changes in the muscles themselves.

The study looked at a group of 66 older adults (average age in their 70s), who were first categorized as severely weak, modestly weak, or strong based on their measured performance on a standardized physical test. In the study, the subjects were asked to push against resistance with their leg extensor muscles, using as much strength as they could generate. When they reached their self-perceived limit, the muscle they were using was then stimulated electrically. If this caused the muscle to put out more force, it was a sign that the strength limitation the person experienced came from somewhere other than the muscle itself.

When the added force that came from electrical stimulation was expressed as a percentage increment, it showed that the weaker the test subjects, the larger a boost their muscles got. The subjects in the "severely weak" group (who were on average older) got an increase of 14.2 percent - twice the 7.1 percent increase shown by those in the "strong" group. When the conventional scientific wisdom linked such weakness mainly to loss of muscle mass, many drug companies looked for medications that acted directly on the muscles, but few proved effective. The new study provides further evidence that the nervous system plays a significant role in the problem.

Contrary to popular belief, cartilage in human joints can repair itself through a process similar to that used by creatures such as salamanders and zebrafish to regenerate limbs. The mechanism for cartilage repair appears to be more robust in ankle joints and less so in hips. The finding could potentially lead to treatments for osteoarthritis, the most common joint disorder in the world.

Researchers devised a way to determine the age of proteins using internal molecular clocks integral to amino acids, which convert one form to another with predictable regularity. Newly created proteins in tissue have few or no amino acid conversions; older proteins have many. Understanding this process enabled the researchers to use sensitive mass spectrometry to identify when key proteins in human cartilage, including collagens, were young, middle-aged or old. They found that the age of cartilage largely depended on where it resided in the body. Cartilage in ankles is young, it's middle-aged in the knee and old in the hips. This correlation between the age of human cartilage and its location in the body aligns with how limb repair occurs in certain animals, which more readily regenerate at the furthest tips, including the ends of legs or tails.

The researchers further learned that molecules called microRNA regulate this process. Not surprisingly, these microRNAs are more active in animals that are known for limb, fin or tail repair, including salamanders and zebrafish. These microRNAs are also found in humans - an evolutionary artifact that provides the capability in humans for joint tissue repair. As in animals, microRNA activity varies significantly by its location: it was highest in ankles compared to knees and hips and higher in the top layer of cartilage compared to deeper layers of cartilage.

"We were excited to learn that the regulators of regeneration in the salamander limb appear to also be the controllers of joint tissue repair in the human limb. We believe we could boost these regulators to fully regenerate degenerated cartilage of an arthritic joint. If we can figure out what regulators we are missing compared with salamanders, we might even be able to add the missing components back and develop a way someday to regenerate part or all of an injured human limb. We believe this is a fundamental mechanism of repair that could be applied to many tissues, not just cartilage."

Obesity is an established risk factor for several cancers. Adult weight gain has been associated with increased cancer risk, but studies on timing and duration of adult weight gain are relatively scarce. We examined the impact of body mass index (BMI) and weight changes over time, as well as the timing and duration of excess weight, on obesity- and non-obesity-related cancers. We pooled health data from six European cohorts and included 221,274 individuals with two or more height and weight measurements during 1972-2014. Several BMI and weight measures were constructed. Cancer cases were identified through linkage with national cancer registries. Hazard ratios (HRs) of cancer were derived from time-dependent Cox-regression models.

During follow-up, 27,881 cancer cases were diagnosed; 9,761 were obesity-related. The HR of all obesity-related cancers increased with increasing BMI at first and last measurement, maximum BMI and longer duration of overweight (men only) and obesity. Participants who were overweight before age 40 years had an HR of obesity-related cancers of 1.16 and 1.15 in men and women, respectively, compared with those who were not overweight. The risk increase was particularly high for endometrial cancer (70%), male renal-cell cancer (58%) and male colon cancer (29%). No positive associations were seen for cancers not regarded as obesity-related. In conclusion, adult weight gain was associated with increased risk of several major cancers. The degree, timing, and duration of overweight and obesity also seemed to be important. Preventing weight gain may reduce the cancer risk.

The global longevity and anti-senescence market will witness rapid growth over the forecast period (2018-2023) owing to an increasing emphasis on stem cell research and increasing demand for cell-based assays in research and development. An increasing geriatric population across the globe and rising awareness of antiaging products among generation Y and later generations are the major factors expected to promote the growth of a global longevity and anti-senescence market. Factors such as a surging level of disposable income and increasing advancements in anti-senescence technologies are also providing traction to the global longevity and anti-senescence market growth over the forecast period (2018-2023).

Senolytics, placenta stem cells, and blood transfusions are some of the hot technologies picking up pace in the longevity and anti-anti-senescence market. Companies and start-ups across the globe such as Unity Biotechnology, Human Longevity Inc., Calico Life Sciences, Acorda Therapeutics, etc. are working extensively in this field for the extension of human longevity by focusing on the study of genomics, microbiome, bioinformatics, and stem cell therapies, etc. Senolytic drug therapy held the largest market revenue share in 2017. The fastest growth of the gene therapy segment is due to the large investments in genomics.

The scope of this report is broad and covers various therapies currently under trials in the global longevity and anti-senescence therapy market. The market estimation has been performed with consideration for revenue generation in the forecast years 2018-2023 after the expected availability of products in the market by 2023. The global longevity and anti-senescence therapy market has been segmented by the following therapies: senolytic drug therapy, gene therapy, immunotherapy, and other therapies which includes stem cell-based therapies, etc. Forecasts from 2028 to 2023 are given for each therapy and application, with estimated values derived from the expected revenue generation in the first year of launch.

The term "sarcopenia" was first introduced to describe the progressive age-related loss of muscle mass and is correlated with poor health-related quality of life. In this context, the handgrip dynamometer (HGD) is a useful tool to evaluate muscle strength because it provides simple, fast, reliable, and standardized measurements of total muscle strength. In addition, handgrip strength (HGS) is considered an important measure to diagnose dynapenia because low HGS is a robust predictor of low muscle mass and a clinical marker of poor physical performance.

In the respiratory system, the incidence of chronic lung diseases (CLDs) is comparatively higher in individuals aged 65 and older. HGS is an indicator of overall physical capacity. It is not limited to assessing the upper limbs and is a good predictor of morbidity and mortality, indicating that the HGD is a potentially useful instrument for evaluating different populations with different respiratory conditions. Despite these advantages, HGS is rarely used as a functional measure in patients with respiratory diseases, perhaps because it is erroneously considered a part of a complex battery of functional tests.

Current evidence indicates the presence of different phenomena linking lower muscle mass and function with the occurrence of CLDs in this population. Chronic systemic inflammation is related to nontransmissible CLDs in the elderly, and this inflammatory status may be one of the main links to reduced HGS. In addition to systemic inflammation, other contributors that appear to be important are the chronic effects of hypoxemia due to CLDs, physical inactivity, respiratory and peripheral myopathy, malnutrition, and the use of corticosteroids, which is common in many CLDs. Sarcopenic obesity is increasingly diagnosed in different clinical conditions and may be an important link between decreased HGS and adiposity in CLDs. Reduced HGS in CLDs should be considered a systemic phenomenon requiring a holistic approach to restore physical reconditioning and nutritional status. Therefore, early targeted interventions should be developed in patients with CLDs to delay muscle strength decline and prevent functional limitations and disabilities.

Ischemic stroke is a devastating and major cause of morbidity and mortality worldwide. However, due to the narrow time window of thrombolytic therapy, new pharmacological therapeutic approaches are still necessary. Cyclin-dependent kinase 5 (CDK5) is a proline-directed serine/threonine kinase that interacts with NR2B and phosphorylates NR2B to promote ischemic neuronal death. Targeting aberrant CDK5 is neuroprotective for the neuronal loss, tauopathy, and microglial hyperreactivity induced by stroke.

Previously, a membrane-permeant targeting peptide-based method that rapidly and reversibly knocks down endogenous proteins through chaperone-mediated autophagy (CMA) had been validated. In this study, we synthesized a membrane-permeable peptide (Tat-CDK5-CTM) that specifically disrupts the binding of CDK5 and NR2B and then leads to the degradation of CDK5 by a lysosome-mediated pathway.

We found that the administration of Tat-CDK5-CTM not only retards calcium overload and neuronal death in oxygen and glucose deprivation (OGD)-treated neurons but also reduced the infarction area and neuronal loss and improved the neurological functions in MCAO (middle cerebral artery occlusion) mice. The peptide-directed lysosomal degradation of CDK5 is a promising therapeutic intervention for stroke.

Matthew Scholz is co-founder and CEO of Oisín Biotechnologies. When asked what led him to focus on aging, Scholz responds, "Aging has been on my mind for a long time. Even at Immusoft, my long-term goal for the platform was to recreate the biochemical environment of youth in old age. I reasoned that it would never be possible to take enough drugs to accomplish this but, if you can program the body, you can do anything."

It wasn't until a chance meeting at a Health Extension Salon sponsored by Joe Betts-LaCroix in 2012, however, that Oisín Biotechnologies was first conceived. Judy Campisi took the stage to present her work at the Buck Institute for Research on Aging and the results of a recent mouse study. She explained how researchers had created transgenic mice in which senescent cells could be easily cleared with an otherwise innocuous drug. Scholz thought the results were amazing but didn't think it was a feasible approach from a clinical perspective. He leaned over to the guy sitting next to him - Gary Hudson - and said, "That's amazing, but I would do it totally differently."

That comment led to drinks at the bar, where Scholz explained to Hudson his strategy. And that conversation led to a collaboration that would become Oisín Biotechnologies. Gary Hudson not only liked what Scholz had to say, he also knew Dave Gobel at the Methuselah Foundation. And Gobel and the Methuselah Foundation were eager to fund Scholz's proof of principle.