/Cancer and Aging Overview: Would Better Nutrition Help Us to Age More Slowly?

Cancer and Aging Overview: Would Better Nutrition Help Us to Age More Slowly?

Video: Cancer and Aging Overview: Would Better Nutrition Help Us to Age More Slowly?

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>>Coordinator: Welcome and thank you for standing by. At this time all participants are in a listen-only mode for today's call. Today's conference is being recorded. If you have any objections you may disconnect at this time. And now I'll turn today's meeting over to Melissa Glim. Thank you. You may begin. >>Melissa Glim: Hi. My name is Melissa Glim and I am a contractor in the National Cancer Institute's Office of Communications and Education. I want to welcome you to the Frontier for Nutrition and Cancer Prevention online CME Webinar series brought to you by the National Cancer Institute's Division of Cancer Prevention.

Please note that closed captioning is available. You just have to click the word More at the top of your screen and a box will open up. To ensure uninterrupted streaming of the online portion of this presentation please close any additional programs and windows on your desktop. If you do have any technical difficulties during the presentation please press Star Zero for operator assistance for audio issues or call the technical support number shown on the screen. It's 800-857-8777 and choose option 3 if you are having Internet issues. As a reminder this Webinar is being recorded and will be archived on the Nutrition Science Researches group's Web site. We will post the address at the end. Before you get started. Before you get started please take a moment to familiarize yourself with the live meeting desktop. The question and answer box labeled Q and A is located on the top navigation bar. And you can click – keep it open by clicking on it and dragging the box to the right side of your screen.

You may type your questions here at any time during the presentation. And the speakers will answer them during the Q and A period after the presentation as time permits. Now I would like to introduce you to our moderator for today's workshop. That's Dr. Gabriela Riscuta, Program and Activity Director in the Nutritional Science Research Group Division of Cancer Prevention at the National Cancer Institute. >>Dr. Gabriela Riscuta: Good afternoon. I would like to welcome you all to the first presentation Can Nutrition Simultaneously Effect Cancer and Aging? Before we get started I want to thank my colleges, all the Steering Committee and Planning Committee in the Nutritional Science Research Group in the Division of Cancer Prevention and especially to our Acting Chief Doctor Harold Seifried for his report.

We are very excited that over 1000 people have joined the call today from every single state in the United States. We have physicians, nurses, registered dieticians and nutritionists, clinicians and public health officials, researchers, academics, again from United States and over 20 countries around the world. We expect to have a wide variety of interest and perspectives. And we will get questions and answer as much as we will be able to. And if we will not be able to answer every single question please follow-up with us on the emails and phone numbers that is provide at the end of the presentation. While aging is inevitable and increasing our chances for serious diseases including cancer we would like to know the mechanisms that are behind these two processes. And even more we would like to know if nutrition can play a role in slowing the aging processing and reducing the cancer risk at the same time.

We know this is a very ambitious topic but we have – we are very fortunate to have a distinguished panel of speakers to present this subject to you today. Now I will – I would like to turn over the program to our first speaker Dr. Joao Pedro de Magalhaes. And he's a Senior Lecturer in the Functional and Comparative Genomics Division at the Institute of Integrative Biology University of Liverpool England. He is going to speak today on common mechanists of cancer and aging. When Dr. Magalhaes is finished we will have a time for a few questions Thank you very much and thank for the invitation and for setting up this Webinar. What I'll do is I'll give an overview of the overlap between cancer and aging touching with but without going into a lot of detail about the major potential processes apply and also touching on some of the issues and then the other speakers will cover in more detail. So as I'm sure you're aware cancer incidence increases dramatically with age. Aging- related diseases affect typically other individuals.

And as you can see so for example on the right here you can see that's a log scale. So it's So it's the same data on the left and right. But on the right it's on the log scale. And so as you can see so cancer incidence increases exponentially with age. And this is true for the vast majority of tumor types although the pace of increase with age can defer between them. There are a few exceptions however the most notable of which is testicular cancer which quite strikingly has an incident peak relatively early in life around age 30 or which is quite surprising and the mechanisms or reasons for which are not well understood yet. Having said that aging clearly effects or is a major risk – age is a major risk factor for cancer. And when I think about aging I mean people are familiar with the aging process. But I'll show you in this (check) centenarians the interesting thing about aging is that it effects virtually every organ in the body and it entails a variety of changes from functional, physiological, molecular cellular changes.

And so I like the definition of aging as a progressive deterioration of physiological function accompanied by an increase in vulnerability and mortality with age. Now while aging is a complex process that affects multiple systems we don't yet know what drives the process of aging, I mean what are the causes of aging. There are a number of theories like DNA damage which I'll talk in a few minutes, the telomeres which we'll here, we'll talk about later, tissue damage. So there are a number of theories but none of which have been proven yet. So I would say that precise cellular molecular mechanisms that drive the aging process are still under debate. Having said that, we have made some dramatic progress in our understanding of aging. think arguably the most impressive development has been at the level of understanding that aging is a surprisingly plastic process in model systems and can be manipulated by gene and by environmental manipulations and diet in particular. So at the genetic level I guess the first gene shown to modulate aging was age-1 which is identifying the round worm C.

elegans by Tom Johnson and colleagues over 20 years ago. And it is quite dramatic and at the time even surprising how a real – a single gene can have a dramatic impact on lifespan and on aging. And we now know of hundreds of genes that have a significant impact on the aging process in model systems. So we know of hundreds in invertebrates and in worms for example. But even in mice there's about 100 genes that we know have to have a significant impact on aging phenotypes. So we have a number of genes that if you disrupt them they result in a significant increase in lifespan. So I believe the current record is a growth hormone receptor which if you knock out in mice you're saying – or animals that lives about 50percent longer than well-tapped controls.

And what is interesting as well is that these animals not only live longer but they live healthier. And I'll mention this in more detail in a little bit but cancer is one of the disease that is effected or that is retarded when lifespan is increased in this model. Conversely there's a number of models also of accelerated aging and is disruption of particular genes result in what is called segmental progeroid syndromes in mice that resemble accelerated aging. A number of these genes are actually related to DNA repair and DNA damage response. And so not surprisingly one potential areas of overlap between cancer and aging is exactly in terms of DNA damage because we know that mutations are one of the hallmarks of cancer. And we also know that DNA damage and DNA mutations accumulate with age. And so one of the theories of aging, the DNA theory of aging argues that with age there is an accumulation of DNA damage which impairs cell function which results in cell loss possibly at the level of stem cells as well and that this is one of the mains causes of aging.

There is some evidence for it as I mentioned before in terms of this segmental progeroid syndromes in mice. There's also one very good example from patients which is Werner's syndrome which is arguably the most striking progeroid syndrome in people which is what you see here. As you can see these patients look older at younger ages that they normally would. And these individual develop a variety of disease at early ages including cancer. Although the exact type of cancers that they developed is not the same or is not exactly the same as in normal individuals. But what is remarkable is that the disease which results from mutation in a single gene called the Werner's syndrome gene which is involved mainly is helicase exonuclease involved in DNA metabolism. These individuals develop this complex phenotype that appears to resemble accelerated aging. There's also been a number of studies at the level of quantifying DNA mutation, DNA damage with age. I do think this has not been as clear cut as probably once thought an issue. So for example what you see in this slide is the work of Jan Vijg and Colleagues.

And what they found in mice is that you do see an accumulation of DNA mutations in some tissues but not in others. So there seems at least for this particular assay that they use there tends to be tissue specific effects. And the other thing that is important to emphasize is that while we talk about an overlap between cancer and aging at the level of DNA damage which is certainly important for cancer and it appears to be important for aging as well there is also different types of DNA damage and there's also different type of DNA pathways. And without going into great detail we know from a variety of mouse models for example such as the work of (Gen Rachmacher)'s in the Netherlands that different types of damage can have different types of impact on cancer and aging. So for example DNA damage that appears to increase the mutational load that increases in chromosomal aberrations tend to impact more on cancer while on the other hand DNA damage that result in cell cycle or rest or cell death appears to be more important in cancer.

So while there is an overlap between the mechanism, the conceptual level of DNA damage impacting on aging and cancer when you go into the details there may be some differences between the specific types of DNA damage and specific types of DNA repair. Now I've mentioned this progeroid mice and accelerated aging. On the other hand of this (bench) you have this (unintelligible) and mutations in mice for which there's quite a few. I think arguably the most important pathway involved is the growth hormone IGF-1 pathway. IGF-1 stands for Insulin Growth Factor 1. And we now know that there's a variety of mutations in this pathway in which decreased growth hormone IGF-1 signaling extends lifespan and appears to retard aging. So it's not that just the animals are living longer, they are living longer healthier. So their health then is also extended.

So these animals tend to be smaller than controls. And they live longer. And they also appear to be protected from cancer. I mean that's a broad generalization when you look at the different mutants. I mean some of them have a lower cancer incidence. Others have different types of tumors. Others have kind of retarded onset of particular cancer types. But by and large as manipulations or decreased growth hormone IGF-1 signal does have not just extends lifespan but also protects against cancer but which not too surprising because growth hormone and IGF-1 essentially mitogens that makes cell proliferate. So it makes sense that if you decrease their levels animals are going to be protected from cancer. And then conversely if you over express bovine growth hormone mice appear to age faster. So they're bigger and they appear to age faster. And this has been the work of amongst others Andrzej Bartke. Now there's also been a few mutations in individuals that in the same pathway.

I'll show you one example which is in these individuals that have deficiency in the growth hormone receptor. As I mentioned earlier if you not got growth hormone receptor in mice the mice live up to 50 percent longer and they're also protected from cancer. But so there was a study conducted in Central America amongst a cohort of such individuals. And what's interesting or the most interesting finding in my opinion was that individuals at least in the cohort they do not die of cancer. So sample size was not too big. There were 30 individuals. But when compared to their relatives that don't have the growth hormone receptor deficiency for which about 20percent die of cancer it's quite remarkable that none of these individuals dies of cancer. I should say however that these individuals are not long lived per se. They appear to have a high incidence of some other diseases like heart disease.

So unlike the mice the growth hormone receptor knockout mice who live longer (unintelligible) protected these individuals that live appear to be cancer protected but do not live longer. And last this shows that the same pathways affect aging in mice and protect against cancer at least at levels one specific age related diseases in this case cancer they could be relevant for people. Now when you talk about environmental manipulation of aging and dietary manipulations in particular there's been quite a lot of work on this as well for the past fifty years. And by far the most widely studied manipulation is caloric restriction. And caloric restriction was discovered by Clive McCay and colleagues at Cornell University some earlier findings. And what the calorie affects so normally mice in the lab they fed at liberty. That means they can eat as much as they want.

But if you restrict the amount of calories that they eat without malnutrition this has a significant impact on longevity. It increases lifespan up to 50percent is – which is this plot you see here which I believe is from Edward Masoro's lab in San Antonio. So there is a very significant impact of restricting the amount of calories. And again just like in the mutated mouse it's not that the animals are living longer, they're also living longer healthier. And a variety of a age-related diseases are also ameliorated by caloric restriction and cancer in particular. So if I can summarize fifty years of research on caloric restriction in a single slide I would say we know now that it expands lifespan in most species but not all of them. There are a couple of exceptions. We know it does have also some negative side effects. So for example it appears to be detrimental in fighting infections.

There's been more recently done – work done on Rhesus monkeys under caloric restriction. The work is still ongoing because Rhesus monkeys live over forty years. So as you may imagine this is a very long project. But the results so far suggest that caloric restriction protects against cancer in Rhesus monkeys. Whether it will extend lifespan I think is still an open question. There are some conflicting results between two different groups in Wisconsin NIA. I would say that if caloric restriction extends lifespan in Rhesus monkeys it will be far less than what it does in rodents. So if you extrapolate to primates it appears that caloric restriction does have cancer protecting effects. But in terms of its impact on longevity per se it's probably less than what we observe in rodents. And so my own view is that caloric restriction isn't likely to be as effective in humans. And we certainly don't think it's going to be as effective in humans as it is in rodents.

And the results from Rhesus monkeys support that. But particularly give it its impact on age-related conditions and cancer in particular it could be helpful in trying to identify pathways involved and try to identify drug targets that could be used in the clinic in particular for treatment of age related diseases like cancer and Type 2 diabetes. And there's a number of work focusing on this. And I mean my own view of caloric restriction is that it involves endocrine mechanisms like growth hormone insulin IGF-1. But also there's a complex signaling (unintelligible) involving I think it was PI3K (Foxmontur) then – and many other players. That is also important. If you can understand this then it may provide applications in the clinic for age-related diseases. And so what we do know is that there are longevity pathways that can modulate aging.

And they do so to a very dramatic extent. And even surprisingly I think people didn't expect this impacts 30 or 40 years ago. And we do know that these pathways also tend to impact on cancer. So the same mechanisms of caloric restriction for example and these pathways that retard aging in rodents also have a significant impact on cancer. And conversely you have DNA damage and DNA repair systems that also probably are involved in aging and are certainly involved in cancer. Although as I said mentioned early they could like a different types of DNA damage and different types of DNA repair that are more important for cancer and aging. And so now the last thing I will mention is so in – is whether – so we know for example that mutations accumulate with age. And we know that that is important for cancer. But there's a number of changes in all of our organs during age. So I think a crucial question as well in terms of relationship between aging and cancer is whether tissues of – old tissues whether they are more fertile ground for cancer development.

And actually contrary and contrary to popular perception or sort of an urban myth is that so cancer is not less aggressive in the age. I mean there are some types of cancer that are less aggressive in the age and most notably breast cancer probably due to genotype differences. But by and large cancer is not less aggressive in the age. On the contrary it's actually more aggressive in the age. I mean there's some very clear examples like ovarian cancer which is clearly more aggressive in the age. And that is the rule, not the exception. So cancer does tend to be more aggressive in the age. So one question is whether there are process related to aging or aging duration that contributes to this. There's been some work which I will briefly summarize on this. One potential area of interest is related to senescent cells. These are cells that are from irreversible growth arrests either due to short telomeres or stress. And this is mostly the work of Judy Campisi's lab.

And what they've shown is that senescent cells which tend to accumulate with age at least in some tissues these contribu4e to create a tissue microenvironment that facilitates cancer development. And they've actually done some elegant experiments in injecting mice with malignant cells with senescent cells and with non-senescent cells. And what they showed was that if you inject mice with malignant cells together with senescent cells this forms more and larger tumors. And having said that there's probably several types of processes of aging that contribute to both help cancer development and also hinder cancer development. So on one hand we know the time or exposure. Time exposure is a crucial aspect in cancer. I mean there's mutations that accumulate. So what you see here in blue are cell accumulating mutations. But on the other hand you have a variety of aging processes that could also both help and hinder cancer development. So for example in terms of helping cancer development I've mentioned senescent cells. But another very important process in aging is inflammation.

So we know that inflammation is important in age-related disease. We know inflammatory levels increase with age. And we know inflammation's also important in at least some types of tumors. So it is certainly conceivable to hypothesize that age-related changes increases in inflammatory levels contribute to cancer development in the age while conversely you could have even mechanisms that hinder cancer development. And one obvious example would be hormonal levels. So I mention there earlier how growth hormone and IGF-1 if you reduce growth hormone IGF-1 (unintelligible) in mice it extends lifespan and retards cancer development. So what we also know is that these levels of these hormones decline with age normally. So it's conceivable to hypothesize again that this actually hinders cancer development in the elderly. So there's a variety of processes and it will also depend on cancer type and tissue type.

So to summarize we do know that some aging changes may render tissues more prone for cancer development like senescent cells and inflammation. On the other hand if you have normal processes that maybe degenerate with age but are important for tumor development then these changes with age may contribute to cancer resistance. There's been a variety of studies using venographs in animals of different ages where you can inject malignant cells of animals of different ages. But these have been providing conflicting results. And sometimes it forms these experiments yields there are more tumors in young, other times in elderly, old animals. But I – what I do think they show is that there's more to how age is a factor in cancer development than merely mutation accumulation. And I think there's a lot of work still to be done in this. The one thing I would emphasize is that there's also a lot of tissue in tumor specific effects. So it's difficult to generalize how this process, the aging processes effect cancer.

And to finalize I mean there's a lot of work still to be done in this area in understanding the overlap between aging and cancer. There's still a lot of open questions. I mean we still don't fully understand why testicular cancer incident peaks so early in life. As I just mentioned I still think there's a lot of work to be done in understanding which tissues are effected in which way to increase or to be more or less fertile for tumor development, the issue of how hormones and how the immune system changes with age, how this influence cancer development I think there's been very little study in so far. And lastly because we know that longevity pathways and dietary manipulations like caloric restriction impact on aging and cancer certainly one exciting prospect is whether we can employ this knowledge of these pathways and the players involved to ultimately translate these findings for to improve people's life and to develop anti-cancer treatments. So thank you very much. And so this is just some information about our lab if you wish to learn more about what we do.

Thank you. [Dr. Gabriela Riscuta:] Thank you Dr. Magalhaes for the great presentation. We have a question from the audience for you. And read a question is has the recent (unintelligible) calorie restriction accounted for the fact that eating is more amount of calories would be likely mean this is a lower consumption of carcinogens through the food supply? In short is it the calorie restriction that is protecting against cancer or is it – does it decrease amount of carcinogens that someone would ingest that would decrease the risk of cancer? [Dr. Joao Pedro de Magalhaes:] So I think there's two issues that you touched there. So the first one is what – I guess what constitutes a normal diet in the sense that it's okay so you're doing caloric restriction but on the other hand well maybe the problem is with the control animal that maybe they're eating too much of something that is bad for them.

And I think that's been an issue in the field for a very long time. And it's actually an issue with the studies of Rhesus monkeys because some of the differences between studies that have been published so far relate to how the controls are treated. I mean what is a normal diet. I think the take-home message in general is that eating a lot of calories and – will be detrimental both in terms of lifespan and in terms of cancer. As for the other issue which is whether in the food intake whether there's carcinogens that is something I'm a little bit outside of my expertise. But I'm not – I wouldn't necessarily think that there are carcinogens in the normal diet of animals in the lab depending on how you define a carcinogen or a mutagen. So I normally would not think that to be a problem. So my guess is not that it's the food that is causing the cancer. It's rather that there are pathways involved. You know, there's a lot of work on obesity and cancer.

And I think Steve will talk a little bit about this as well in his talk that so in terms of the metabolism and that this in turn impacts on cancer but not the carcinogens in the food themselves. [Dr. Gabriela Riscuta:] Okay thank you. And we have time for one more question. What percentage of caloric restriction was associated with decreased aging in animal models? [Dr. Joao Pedro de Magalhaes:] I think there's a variety. If my memory serves me correct there's a variety of diets that can result in a lifespan increase. I do think that the most striking effects are done with the most severe diets without malnutrition. So I think you can do like something like 40percent reduction of calories. I have seen studies along those ways. But there are other variants that can also result in an increasing lifespan so that there's different types of restriction of diets that can have an impact. Though I think that the ones that have the most impact are the strictest ones so to speak.

[Dr. Gabriela Riscuta:] Thank you so much Dr. Magalhaes for great presentation and for answering the questions. Again for additional questions as we are unable to ask you at this point we will follow-up with the participants and forward to your email address the questions to answer later. And now we need to move on. And I'm pleased to introduce our next speaker Dr. Maria Blasco. She is the Director and the head of the Telomeres and Telomerase Group at the Spanish National Cancer Research in Madrid, Spain. Dr. Blasco will present the role of the telomeres in health span and longevity. When Dr. Blasco is finished we'll have time for a few questions as well. Dr. Blasco? [Dr. Maria Blasco:] Hello. It's a pleasure for me to give this Webinar. So my talk is going to – to also to start by saying that one of the highest risk factors for developing cancer is aging.

I think is true for all the diseases that we just (unintelligible). So per incident increases with age and are 40 to 50 years old onward it increases very dramatically. So this means that it is one of the causes of diseases. This is our view of the drug then. While we are young and there is little degeneration also there is less disease. And this aging what causes the disease. So aging would be the cause and the disease would be the consequence of this aging process. It's very important to understand which is the cause of the aging, the genetic factors and environmental factors. Because if we are able to find this common cause of disease then the therapeutic interventions which are aimed to treat disease would be able to delay aging associated diseases. So in other words when there is a (unintelligible) health span and delay the presence of diseases. So this is a change in paradigm.

Now you can think that you can develop several different interventions which are different for every particular disease. And this has been the situation until now. Or you can think that there is a common cause for disease which is the aging person. And you would be able to develop therapeutic interventions which are directed to slowdown the aging process and in this manner being able to prevent or treat many different diseases. So which are these causes of aging? So during the last I would say 20 years there have been strong genetic evidence identified a number of molecular pathways. We heard some of them which are important to explain why cellular aging happens. And the work has been working in one of them, telomere shortening. And I will describe about the – our work as regarding in the role of telomeres in aging and disease. So the telomeres are the ends of the chromatic chromosomes. They are very important to protect chromosomes from fusion and rearrangements. And we also know that telomeres however they do shorten with every cell division. This is the effect which is multiplied with increasing cell divisions.

Life would not be actually possible if there would not be a way to compensate for this telomere loss. And in fact, there is an enzyme which is a – it is known as telomerase, telomerase is a pretty potent and essential (unintelligible) development and is able to recognize the chromosome end and by copying an RNA molecule which is part of enzyme is able to compensate for this telomere shortening. So telomerase resets telomere length at the blastocyst stage. When we are born telomerase is silent in the majority of cell types and tissues including also stem cell compartments. And this causes telomere shortening. When telomeres become critically short this triggers a DNA avalanche response leading to cell death or cell (unintelligible). And the – we think that this is one of the causes why aging happens. In the case of cancer however cancer cells need to reactivate telomerase. And this happens in the majority of human cancers in order to be able to maintain the telomeres always (young) and to be able to be (unintelligible) mortally. So this is the interest of the science telomerase in cancer and aging.

This is another way to look at the problem. So telomere length increases with increasing age. This includes (substantial) compartments. When telomeres reach a critical tissue length these would be causative of disease. Cancer is an aging associated disease as well. But in the case of cancer telomerase has to be reactive in order to allow the mortal growth of cancer cells. So we know that there are some human diseases which are known as telomere syndromes which are humans that have mutations in some of the telomerase or telomere components. And they have a premature loss of their regenerative capacity of tissues and premature death by different diseases like a plastic anemia pulmonary fibrosis among others. So this indicates that the amount of the telomerase that we have are very limiting and affecting telomerase will lead to a premature loss of the ability for our tissues to regenerate. We also know that not all individuals of the same age have the same telomere length.

Actually telomere length is very heterogeneous. And we know that there is a correlation between having shorter telomere and normal, meaning in a lower person (unintelligible) of telomere length with a higher risk of developing different diseases. And this brought the possibility that telomere length could be used as a biomarker of aging with prognostic value for many different diseases. In actually in my group we have been exploring this possibility and we have developed a technology which is a high throughput technology that is able to measure telomere in blood sample so you can determine both a mean telomere length and what is more importance the abundance of short telomere. These are the ones that trigger the DNA damage response. So we have evaluated this – these technology in a recent collaboration with a group of Michael Schneider's in Stamford, University of Stamford. Michael Schneider has a mutation in telomerase. And when we measure average telomere length with our technology, can be measured by our technologies.

We see that Michael Schneider has a normal telomere length when compared with individuals of the same age. However when we look at abundance of short telomere which as I said are the ones that cause disease we see that Michael Schneider has a very high percentage of critically short telomere, in this case telomere below 3 kilobases. And therefore we can correlate the phenotype which is a mutation in telomerase with – sorry the phenotype which is having shorter telomerase than normal with a genotype which is a mutation in telomerase. And we think this could be value for disease prevention. So currently we are exploring many different diseases which have been related to presence of short telomere. And I will give you an example with familiar breast cancer and which brings me to the following reasoning is that we know that genetic testings are important to determine the risk for developing diseases but they do not reflect on their way of life or the life habits.

And telomere length however, they are both an indication of the inheritance. So you can have a mutation in telomerase and also the environment that there are many different publications showing that the rate of these telomere shortening with aging depends on life habits and this is relevant- and we think that the combination of both genetic information and this case telomere length will give us a more powerful way to predict risk and time of onset of disease. And of course this could also aid in the development of new treatments to prevent disease. In the case of breast cancer, familiar breast cancer you know that mutations in BRCA1 and BRCA2 give a higher risk of development breast cancer (unintelligible). And in – well the – here this in (IO) they were both (unintelligible) demonstrated that when you combine genetic information with telomere length this is a more powerful way to predict risk.

So the women that have the mutations in BRCA1 and BRCA2 on top shorter telomere develop breast cancer early in life. So it's a factor of health assessing risk in the case of breast cancers. So one of the questions of course is can telomere length also predict lifespan of a given individual? And I would like to mention a couple of studies in humans. We don't know yet. But in birds there was a study which showed that – a longitudinal study that showed that telomere length at day 25 after risk was predictive of the longevity of this bird species. And we have done a similar study with mice. So we perform a longitudinal study. So following telomere length in individual mice with time and see whether any parameter associated to telomeres was able to predict longevity in mice. And the first thing that we all saw is that mice shorten the telomeres 100 times faster than humans. And this is interesting because this may explain why even though mice are born with longer telomeres than humans they live for much shorter time than humans. So their telomeres also shorten much faster than human telomeres. And what is important, you know, we found that the rate of increase of short telomeres was predictive of individual mouse longevity into different strains, a wild type strain and also in a strain which had increased telomerase discretion that we knew that had the way long – longer lived, that would delay aging.

So of course knowing what the short telomeres can be causative of aging you can think about possible therapeutic strategies to delay this process. In this review that we publish we propose that in the case of telomere shortening activation of telomerase in (other) tissues where we know that telomerase is silent we also proposed a strategy. And I would like to give you some data on that regard that we have published. So in the one – on the one hand inactivation of telomerase by making a deletion of telomerase gene in mice causes premature aging similar, very similar phenotypes to these telomere syndromes even though the studies were done before in mice than in humans. And also they were useful to validate that telomerase could be a good anti-cancer target because these mice also had less cancer because they didn't have telomerase. When we increased telomerase in (other) tissues using first trigenesis just making trigenic mice we saw that only over-expression telomerase was resulting in less aging. But however we saw that there was a slightly more cancer and the longevity was not increased due to this increase in cancer.

Because as I said telomerase is important also for cancer cells to evolve. So the way we had to solve this was combining telomerase over expression with increased suppression of tumor suppressed routine. And this resulted in a delay aging and an increase in lifespan which was very significant. And actually this was the first demonstration that telomerase can extend lifespan in any organism. And I'm sure you can see that the telomerase over-expression in this cancer resistant context can increase median longevity by 40percent. Not only that but if we look at the health span so which in a wild type mouse is around one year. After one year mice start developing different diseases including cancer.

In this strain of mice we were increasing lifespan by threefold more or less of the first mice that died much later such as in the case of the control mice. To prove that we are actually increasing – we are delaying aging or in other words we are increasing youth in these mice is that when we do different biomarkers of aging this is a neuromuscular coordination test. You can see that when mice age they fail their ability to hold onto a bar which is something that young mice can do very well. And these mice, the triple mice, the mice with the genetic modifications including telomerase overexpression, when they are chronologically old they are able to do the test as well as young mice. So we – this proves that with a single intervention in this case telomerase are especially able to increase – to delay aging.

So because we cannot do human trigenics we have been also exploring the possibility of developing (unintelligible) strategies to increase telomerase in our (unintelligible) tissues in humans. And this is a project that we did in mice. So here we are assuming that mice are aging like humans because they have a deficiency in telomerase among other things, among other mechanism. And if we are able to put back telomerase, you know, this is – we will be able to delay aging and delay disease. We chose that unassociated viruses because by the state of the art vectors that are being used right now in many different clinical trials for human genetic diseases deficiencies. And this (unintelligible) treat a single time middle aged mice or all mice with telomerase or with GFP as a control vector.

And you can see here how in the lung when there is no telomerase activity of course in a two year old lung that you can see here. But in the lungs of mice treated to single time with telomerase we were able to reactive telomerase. And this was also interesting because I mentioned that there are telomerase syndromes. One of the diseases factor pulmonary fibrosis. And this could be a way to reactivate telomerase in the lungs of these patients. So we see the same as we saw with the trigenic mice that when we treated with telomerase we could see an improvement in many different biomarkers of aging. This is the – again the tight hold success test that I showed you before. The mice with telomerase they performed better in this test. And what was important for us is the longevity. So with a single treatment of telomerase in the middle age or in the old group we could see an increase both in the median and in the maximum lifespan both way increase even in very old mice, in mice that were 2 year old.

And this was in the actions or more cancer. So the incidence of tumors was the same in the control group or in the telomerase treated. So to finish I would like to mention something – well and this is I forgot, this is the summary of the experiment. So we are injecting mice with telomerase. This is sufficient. I didn't show you but we show that. This is sufficient to delay the rate of telomere shortening and again to delay aging and increase longevity. So I wanted to finish by mentioning the impact of life habits, in this case calorie restriction on telomere maintenance. And for that what we did was a longitudinal study in mice where we (unintelligible) the same mice over time during the whole lifespan into group, mice that are – four groups, mice that are calorie restricted or they are having a normal diet. The normal diet is also controlled in terms of calories. So the mice are not eating unlimited.

They are in a controlled number of calories. And we are using two different strains, wild type mice or mice that over-expressed telomerase which have a slightly different longevity. This is just to show that the protocol is working and the mice that are on a calorie restriction which are in light blue, they have a lower weight as you would not expect). They have lower fat content. Metabolic parameters are also improved like glucose tolerance, et cetera. And we see a tendency to hear – to see bigger effects of calorie restriction when we use the calorie restriction of the telomerase of expressing mice. And I will mention that also later. We see also lower decrease of bone density in the mice that are calorie restricted as well as an improvement in this neuromuscular combination set that I show you before, mice that are calorie restricted from both genotypes. They show a better performance at one year – year after diet. So what happens with telomeres. So what we see is that telomeres are also better maintained in the mice that are under calorie restriction.

You can see here on the top telomere length decreasing with age in wild type mice like in humans. Mice also shorten the telomeres as I said before and a corresponding increase in the percentage of short telomeres mice. And how this strength is similar in the strains in wild type and telomere trigenics. However in the mice that are calorie restricted we see that after initial decrease in telomere length then telomeres are maintained and also there is a – not an increase of short telomeres. So this results in having longer telomeres with time in the mice that are calorie restricted compared to the control mice. We can see individually every mouse and see how it behaves. And we observe that in the case of wild type mice 100 percent of the mice that were under the control diet they show telomere shortening so like in the green lines here in the graph. However 20 percent of the mice, wild type mice that were calorie restricted they were actually able to elongate telomeres over time which is something that never happens in wild type mice on their controlled diet.

This ability to elongate telomeres was also seen in some of the telomerase trigenic mice because they are over-expression telomerase. Also, when we look at the percentage of short telomeres we saw that the wild tail mice under calorie restriction but not the ones that were on the control diet were able to decrease their abundance of short telomeres over time as here as shown in red lines, which is again indicating that calorie restriction has a beneficial effect on telomere maintenance. We have looked also in different tissues. These are not longitudinal studies, but at the endpoint we measure telomeres in the kidney, the lung, the muscles and in all cases we see an improved telomere length or better telomere maintainers in the calorie restricted mice. We have also looked in the bone morrow where we can actually look at metaphases and look not only at telomere length but also chromosomal instability as associated with telomeres function.

And we see that telomeres are better maintained in the calorie restricted mice and also there is less chromosomes which lack telomeric signals or less chromosomes, which have a type of aberration that we call multi-telomerasic, which is an indication of replicative expression of the telomere. So they're suggesting that calorie restriction may be decreasing replicative problems of telomere. And that's why also they have beneficial effect on telomere length. On just mentioned finally that we were able to see also a synergistic effect of dietary restriction with TERT overexpression, with telomerase overexpression. So as I mentioned before, when we overexpressed telomerase along there is more cancer.

After we combine these with calorie restriction we are canceling the pro-tumorigenic effect of telomeres like we did before with combining telomeres with chemo suppressor. So in this sense, calorie restriction is acting as a tumor suppressor mechanism. When combined with telomeres we can sell the effect of telomerase in increasing cancer. And as a consequence of this, we think, and also as a consequence of having longer telomeres we see a significant synergy in also in increasing longevity or in the combination between telomeres or expression on calorie restriction. I'm finished – the conclusion of my talk will be that we think that there are common causes for many different diseases. One of these common causes is telomere loss which can be influenced both by genetic and environmental factors and that if we are able to delayed this telomere loss by in this case by an intervention which is telomerase activation in tissues then we're able to extend the time of life free of disease and also increase longevity.

And my last slide is the people of the lab that have done the work that I mentioned to you here. And finally the acknowledgments of the funding agencies. Thank you very much. [Dr. Gabriela Riscuta:] Thank you so much, Dr. Blasco. I have a few questions for you from the audience. And the first question is, are there now a variants in the human population of individual who lose slower or who do not lose telomerase activity? And if so what is the phenotype? [Dr. Maria Blasco:] Yes. Well, this is a very interesting question. What we know so far is that there telomere length is very heterogeneous and not all individuals shorten telomeres at the same pace. We know some of the genetic causes of that like being mutant for telomerase or some of the telomere binding proteins. But we don't know whether individuals that have longer telomeres they have some – we don't know which genetic alterations may have individuals that have much longer telomeres than normal. We know what causes shorter telomeres than normal but not the other way around. But I think it's a very interesting question. [Dr.

Gabriela Riscuta:] Yes I agree it would be nice to know… [Dr. Maria Blasco:] Yes. [Dr. Gabriela Riscuta:] …(unintelligible) as well. We have one other question is related to the stem cell dysfunction and you said that telomerase deficiency would result in stem cell dysfunction. But how would the percentage, I understand that the stem cells differ – is different in young people or animals or in old animals or old people. [Dr. Maria Blasco:] Yes we have done the study in mice. We have studied in particular the telomerase deficient mice which is an artificial model where we induce dramatic telomere shortening. And we show that when mice have particularly short telomeres actually, we saw that they are more label retaining cells which is that there's techniques to identify stem cells. In particular, we did (we did a study in the skin. So that there were more mutated stem cells in the skin when telomeres were short. But the stem cells were not able to react to stimuli like when we treated with agents that are known to (model like) the stem cell then the stem cells that have too short telomeres were not responding to the mitogens in this case.

And they were not mobilizing and they were not regenerating the skin or the hair. So what happens we think with when the stem cells that have critically short telomeres is that they are there but they are not able to actually mobilize and regenerate tissues. [Dr. Gabriela Riscuta:] And this will be the last question. How is telomere length and cancer rates in young people with the aging syndrome compares with normal people of the same age? [Dr. Maria Blasco:] You mean with the telomere syndromes because there are other aging syndromes which are not related to telomere shortening? But in the case of the telomere shortening syndromes in the telomere syndromes, they have increased cancer as well. So, because as I mentioned with the breast cancer study so aging is also and telomere shortening – skin is also to be increasing the risk of cancer.

And individuals that have faster rates of telomere shortening because they are mutants for some of the telomere binding proteins so at the long rate they also seem to have an increased risk of cancer. [Dr. Gabriela Riscuta:] Thank you so much, Dr. Blasco for a great presentation and for answering the questions. And we will follow-up with you with additional questions that we have. At the time, we have to move on with the meeting now. So if there are a lot of them. And we're going to move on to our third and final speaker Dr. Stephen Hursting. Dr. Hursting is a Professor for the McKean-Love endowed a Chair of Nutritional, Molecular and Cellular Sciences in the Department of Nutritional Sciences at University of Texas. And Dr. Hursting will discuss the role of nutrition at the intersection of aging and cancer. And at the end we'll have time for a few more questions.

Dr. Hursting? [Dr. Steve Hursting:] Thank you very much Gabriela and hello everyone, from beautiful Austin, Texas. It's an honor to join you today and talk about nutrition and cancer and aging. What I'd like to cover is some of the links between diet and aging and the cancer process both in humans and animals. And I'll focus on obesity, which as we know, has emerged as a major risk factor for cancer, many types of cancer. And it's also really contributing to some accelerated aging. And we'll talk about that. We'll also particularly focus on lessons from aging research in terms of molecular targets and strategies to break some of the links between obesity and cancer. We've heard about some of this already in today's Webinar. We'll focus on IGF-1 and so micro factor 1 and its downstream signals, adipokines and inflammatory signals.

There are some emerging targets that we've heard mentioned already, certainly telomerase we've just heard here. But Sirtuins and their pathway also seem to be coming into the fore. And we'll wrap up with a discussion about possible diet and pharmacologic strategy to break these obesity-cancer links. So that's what we'll cover. Now if you pick up a newspaper or magazine almost always there's a reference to this obesity epidemic that's occurring in the United States. And certainly we're not alone. There are an estimated 700 million obese adults in the world, many countries affected by this. In the US over the last 30 years we've seen our obesity prevalence more than double. And about seven out of ten adults are currently obese as determined by a BMI greater than 25. About half of those, about 36percent currently of our US adults are in the obese category BMI greater than 30. And there is an age affect here that there is even a higher rate, possibly 42percent of our US adults in the 65 to 74 year category are obese. This certainly has a major impact on number of diseases related to aging.

So this obese phenotype we know is associated with alterations in metabolism. In fact, in more than half of the individuals with this phenotype we actually refer to them as a metabolic syndrome. And they're characterized by insulin resistance and hyperglycemia, dyslipidemia, the classic factors. There are also alterations and inflammation obesity related to pro-inflammatory state. And we see increased cytokines and chemokines in circulation in these individuals. There are vascular perturbations linked to increased factor such as vascular endothelial growth factor and plasma antigen inhibitor one. But these can impact vascular health and neovascularization that occurs with cancer. There are altered adipokines particularly we see increases of leptin and decreases of adiponectin, these factors produce by a fat cell that have a number of biological factors.

And as we've heard already, certainly IGF-1, Insulin Growth Factor 1 is elevated, particularly the bioavailable form of IGF-1 in its relationship with the number of growth, binding proteins that impact bioavailability. So there's a whole spectrum of metabolic perturbations associated with this phenotype. And we know, you know, a number of chronic diseases for many years, we've known, for example, that diabetes and heart disease and hypertension have been linked to this phenotype. Really it's just been the past decade, maybe a little longer than that now what – we've had the appreciation of how major a risk factor obesity is for number of types of cancer. In recent estimates particularly I'll alert you to a paper by Graham Colditz last year did an analysis and an estimate of roughly 25 percent or nearly 150,000 cancer deaths per year are caused by this overweight, obesity state. And so this is a major risk factor, something we've got to get a handle on in terms of breaking this link between obesity and cancer.

Now energy balance is certainly sort of the driver here. We've got a nation increasingly in a positive energy balance state. And I think it's worth looking at this with energy balance equation and how it relates to aging and cancer. On the energy inside we're really referring to amount of calories consumed. The type of calorie can also have an impact whether it's a carbohydrate or a fat calorie or even within carbohydrates the type of calories for example. Pattern, you know, if there's – if the calories are spread over the course of the day, versus, you know, maybe three large, you know, sort of the traditional American three square meal a day kind of approach, that can have an impact as well. On the energy-out side to really the only modulatable aspect of this side of the equation is physical activity. It's very difficult to modulate metabolic rate, thermal regulatory mechanisms, growth or storage mechanisms.

So really, physical activity, increased exercise is certainly a target here and is really the only modulatable aspect. Now in terms of the connections between energy balance aging and cancer says as you've heard already the best studied aspect really is the energy-in side, particularly calorie intake, calorie restriction. There's a large literature on it. And to define what we're talking about with calorie restriction it's really a reduced calorie intake. And typically a 24 to 20 percent to 40 percent reduction relative to an ad libitum-fed control without malnutrition. So pains are taken to ensure adequate nutrition, adequate vitamin, mineral, amino acid, fatty acid nutriger is there. It's really a reduction in energy is what we're talking about. And the best study of this in terms of its connection about the aging. As already has been mentioned calorie restriction extends lifespan in multiple species.

I've just taken samples where a 20 percent reduction was reported in these studies. And you see across organism to yeast, the worms, C. elegans, the fruit fly, the Bowl and Doily spiders, even dogs and cows have been looked at in this regard. And a 20 percent reduction in calories results in a at least a 20 percent perhaps even 40 percent reduction in longevity in some of these cases so just consistent strong affective calorie restriction in terms of both medium and maximum lifespan has been seen in a number of organisms. The best study certainly has been in rodents. And I am – I was struck by this paper. I had started my graduate work in 1987. And this paper by Rick Weindruch about general nutrition had just come out in �86, showing that compared to a non-calorie restricted group a 25, 55, or 65 percent calorie restriction leads to an extension in lifespan. And Rick Weindruch kind of equated that to human lifespan equivalence in years.

And so, you see a median lifespan in mice of about 29 months in the non-calorie restricted with a 25 percent reduction in calories see extensions to a median to, something like 100 or so years in terms of human equivalent so and in about 39 months or so in terms of the mouse lifespan, and increasing with 55 percent even a more extended lifespan. In fact this does show that this can be sort of maximized and in fact you can run into malnutrition much above 40 percent so most of the studies are done in that 20 to 40 percent range. But a clear effect– and this is been consistently shown — a life extension from calorie restriction in a number of other species. Now how about cancer? And certainly part of this life extension is a reduction in tumor development. And, you know, Pedro even mentioned earlier that this seems to be occurring in monkeys as well. But in the rodent systems a number of – I'll just show you spontaneous tumor development in a number of experimental model systems — mammary, liver, leukemia, so on, a number of mouse and rat strains used for this and a variety of degrees of calorie restriction limited, you know, 20 to 40 percent. But what it sees as a very consistent and strong effect in terms of the ratio of incidents all systems comparing the ad lib group to the calorie restricted group.

So that ratio think of it as something of is relative risk. And it's in the order of anywhere between, you know, two, five, all the way up to six-fold increase. And what you see is always positive and it's always in many cases is quite strong this effect of calorie restriction in reducing the development of cancer in these models. This is spontaneous tumors. It – there's a huge literature on chemically induced tumor model systems, same story, very strong, very consistent, effect on tumor development in response to calorie restriction. Now a number of mechanisms have been proposed for how this might work. I've taken this from an article by Longo and Fontana that shows that a decreased calorie intake, this is really sort of a checklist of some of the known and adaptations that occur with calorie restriction.

You see at the metabolic level we've already mentioned some of these insulin-like growth factor 1, insulin, steroid and hormones and cortisol, a number of oxidative stress markers, inflammation, adiponectin we've mentioned and thyroid hormones. So they are clear metabolic adaptations. At the molecular level what's striking is particularly we've already mentioned the mTOR pathway, a number of components of this IGF insulin signaling pathway through PF3 kinases down through mTOR is involved number of those components. Also oxidative stress components mediated by the anti-oxidant regulator NRF2. Sirtuin pathway we'll mention in a minute. But that also is emerging. So a number of molecular alterations, and in terms of cellular adaptations we see generally a decrease in cell proliferation, apoptosis has generally increased.

The process of autophagy has increased. We see DNA repair in carcinogen detoxification processes improved in this low-calorie environment. Oxidative damage has decreased, genomic stability, possibly involving certain 1 regulatory mechanisms there, and immuno surveillance generally improved in terms of anti-cancer immuno surveillance. So the net result here as we said is a consistent story of an increase in median and maximal lifespan. For many years this was really the only experimental modality to increase median and maximal lifespan and also we know a decrease in cancer risk and progression. So these two linked processes are occurring and responding to calorie restriction. Now we can gain some clues from some several long-lived mutant organisms.

We've already seen the calorie restriction effects of the lifespan in what – higher organisms cancer in these organisms. And one clue we've gotten from some of these long-lived mutant organisms again relates to the insulin growth factor one TOR pathway. So we see in C. elegans there's a mutant called daf 2 that is really an alteration in this highly conserved pathway, IGF-1 pathway all the way down to C. elegans. The mutations in that pathway can lead to life extension. And we also in the fruit fly there's a chico mutant that is an IGF-1 receptor mutation once again a life extension in lifespan in occurrence with IGF-1 mutation. And in mice we can actually manipulate genetically the components of this pathway. For example, here's an IGF receptor of heterozygote that has an extended lifespan relative to control. So some evidence that the indeed this pathway is important in life span and in the aging process is really quite clear. And again in rodents we can manipulate these pathways.

And one approach that we've taken is to knock out IGF-1 in the liver. Most of our insulin growth factor one circulation comes from the liver, about 80 percent or so. And so we use the pre-lock approach to actually knock it out and try to mimic what we kind of are seeing with calorie restriction which is a reduction in IGF-1 level in response to the diet. So it's a genetic approach to try to mimic that. And indeed the liver IGF deficients or LID mice have, you know, about a 55 to 60 percent reduction in circulating IGF-1. And as already been mentioned there are examples in humans of individuals with alterations in these pathways. So here's an example from the Laron Syndrome group, of Ecuadorians are sort of a hot cluster in Ecuador of these Laron Syndrome individuals. And as I think has already been mentioned some of these dwarf syndrome individuals have very low IGF-1. They have very low inflammatory cytokines. They tend to have a trend for the longevity and virtually no cancer or diabetes in these individuals.

And so there seems to be some clear evidence that this pathway is important. In terms of cancer we've looked at this in a number of model systems. And one where we see particularly response to calorie restriction is the use MMTV, Mouth Mammary Tumor Virus- induced-Wnt-1 transgenic mouse model. And it's a – in terms of a nice mammary cancer model. It's a basal like model of mammary cancer. And we can actually transplant these tumors into the fat pad of a mouse in this case a wild type mouse. And we see tumor volume as a function of their time after tumor cell injection. In about six weeks into the study the tumors would start to form in the wild type mice and really take off by about the second month and grow rapidly and kill the animal I think before 75 days. In contrast the low IGF-1 group, this LID, liver IGF-deficient mouse never really gets going in terms of tumor growth. We see just barely detectable tumors out at the time when the wild type tumors are really at a point where we have to terminate the study. So reduction in tumor development in response to low IGF-1 is clear and it correlates with what we see in the calorie restricted state where about a 50 percent reduction in IGF-1 is associated with reduced tumor development in a number of models.

We've seen this effect as well in multiple models. This is mammary multiple mammary models, multiple pancreatic cancer models, prostate models told a very similar story, reduced tumor growth and tumor development in the IGF deficient mice. Now one thing we were a bit surprised about is this connection between the IGF-1 and inflammation. And we mentioned information as a part of this metabolic perturbation that occurs with obesity and is also associated with the aging process. Every time we manipulated IGF-1 either through diet, through pharmacologic approaches we see a reduction in circulating cytokines. And here's a list of a panel of cytokines. And in every case, the LID mice, these IGF deficient mice are lower than their wild type counterparts. And this is true also in calorie restriction. Every cytokine we look at is lower in the restricted mouse than in their ad libitum fed counterparts.

So there seems to be a connection between these pathways at least we didn't appreciate. And we'll talk more about that as we go. But these interacting pathways I think are quite important. Now we know that IGF-1 insulin are examples of receptors and kinases. And so they signal through a pathway. And this TOR pathway, target of Rapamycin that's been mentioned in the case of a mouth it's – or in other mammals. It's a mammalian tour, mTOR that is, it seems to be a critical kind of target, critical collection point for a number of these signals that perhaps gives us an opportunity to intervene. And in this case we were curious whether IGF-1 and its downstream signal through TOR may be a critical piece to this. And so we've Leticia Nogueira, a grad student in the lab did a nice experiment where she compared again using this wnt 1 model control mice, a 30 percent calorie restricted group of mice and replicated what we saw before, which is a reduction in tumor growth in response to calorie restriction.

And then Leticia took another step. She added IGF-1 back to these animals because there — we know there's about a 50 percent reduction in IGF-1 in those CR mice. So these mice are still restricted but she exogenously added IGF-1 back in. And she rescued much of the tumor growth in that restricted group simply by adding IGF-1. She also repeated the experiment, but in this time she basically locked mTOR on in the cells that she injected. So she compared a wild type mTOR construct that she introduced into the cells. So they have normal mTOR and again responded to calorie restriction as we saw here with a reduced tumor development in response to the CR and then mTOR was normal and indeed she saw calorie restriction reducing the the activity levels of mTOR. In a group what she locked it on so she did a conceptually activated, basically a myristoylated mTOR construct. She lost to the calorie restriction effect.

There was no longer an anticancer effect with – even though the animals were restricted all the hormones changed, et cetera. They couldn't signal through this mTOR to reduce the growth of the tumor. So further evidence that this pathway is important. And indeed, we've seen this in a number of models as I mentioned — insulin receptor, IGF-1 receptor for example, the receptor tyrosine kinase is a signal through a cascade of enzymes, kinases and what mTOR is sort of the collection point regulator of this path. And it's a key And it's a key regulator of a cell proliferation, as well as protein translation machinery. So it makes sense that this is what we refer to as an energy sensing or nutrient sensing pathway because it – these processes are tied end. You don't want to turn on a lot of protein, you know, machinery, if there's not adequate substrate. So this senses how much energy is in the system and allows this process of protein translation to proceed.

We haven't seen in both in normal and tumor tissue and in multiple tumor tissue types — skin, liver, prostate, colon, pancreas and mammary – that in the obese state when we have animals feeding, you know, fed with a diet induced obese diet that every step of this pathway is activated. All the – all these kinases are phosphorylated and with increased phosphorylation. So it's an activated mTOR signaling. In contrast, every step of the pathway is decreased with the decreased phosphorylation in all these kinases is calorie restriction and interestingly, when we compare the liver IGF deficient, these LID mice which are not restricted and they're not particularly lean, they're short because IGF-1 is low in these animals but they're not particularly lean, but their IGF-1 levels are low and they mimic the calorie restricted animal each step of the way.

So it shows that maybe it's not so much the diet, the body size, but the metabolic state that's driving some of these signals and really we think mTOR becoming a major target. So it kind of further proves that principal that mTOR is involved. And certainly there's evidence in terms of the aging world that it is. We – I'll cite this paper from Harrison in Nature from a couple years ago were they fed in an aging study they started Rapamycin, which is a direct mTOR inhibitor at 600 days of age. So the animals, you know, were sort of eating and living normally. And then at 600 days they were – Rapamycin was introduced into their diet. And what they saw was indeed an extension of lifespan. This -here is a male and here is a female, so a separation and extension of lifespan in response to the mTOR inhibitor so further evidence that this was indeed – in fact, in the aging world they claim now that, in addition to calorie restriction they now have a second experimental mode to increase medium and maximum lifespan. Although really, I think most of the effect is primarily on median lifespan. But still some life extending effects with (unintelligible).

And so we were curious if mTOR inhibitors a pharmacologic approach would have an effect on cancer development as well, and particularly could we offset the effects of obesity? And so we – so Rebecca DeAngel, a grad student in the lab, did this comparing tumor weight in the lean calorie restricted animal, the control, really overweight control animals and obese animals that were on a diet induced obesity diet. So the blue bars are just kind of what we've been seeing before. This is the placebo where the lean and restricted animals had minimal tumor growth, the intermediate controls had intermediate tumor growth, the obese rapid tumor growth. So if you add RAD001 or Afinitor, which is an FDA approved mTOR inhibitor we – even in the lean group there's room for improvement. They basically took the restricted animals and further decreased the tumor development really to a non-detectable level that most of the tumors were barely palpable.

In the controls it took the kind of intermediate (path) tumor growth down to a both calorie restricted level even though the animals were still eating a controlled diet by simply blocking mTOR because you were able to disconnect some of that effect. And I think most importantly in the obese group the addition of the mTOR inhibitor really reversed many of the effects of obesity. So 10 mg per kg dose the tumor growth was comparable to the control even though the animals are still obese many of the metabolic perturbations still intact the only thing was blocked was the mTOR signal in a little higher dose. And this gets to an issue that we're working hard on right now as well is this this obesity effect on drug resistance. It often takes more drug to have the same effect in the obese state.

And that's true even with the mTOR. Of the 15 mg per kg we were able to take the obese mice and really turn them into – in terms of tumor development the level of a calorie restricted animal simply by blocking a single target here, mTOR. And so I think that's encouraging that we can indeed develop and identify targets, develop strategies to disconnect from this obese cancer connection. So here's kind of where we are in our thinking. The calorie restriction to these metabolic alterations really kind of tuning down the insulin IGF-1, improving leptin – adiponectin ratio, decrease in cytokines affecting the vascular factors in a positive way through their growth factor signaling an inflammatory problem that are related to these factors and in terms of growth factor we think that this pathway particularly is promising, the mTOR pathway. In terms of information it really appears that NF-kappa B, the master regulator of a number of cytokines seems to be quite energy responsive and COX-2, cyclooxgenase 2 which is the – a key enzyme prostaglandin metabolism, these two seem to be particular targets in this realm.

that's certainly these are contributing to the anticancer and anti-aging effects of calorie restriction. Now there's a big wide box in the middle. And that's where I think the future directions go. We're seeing preclinical evidence emerging. We already heard about telomerase from Maria and so there's a number of factors that I think. Sirtuins have been a hot topic the last few years and I'll talk more about that. Certainly there's a decrease in reactive oxygen species, oxidative stress with calorie restriction. That can certainly contribute. I think genomic stability we heard about epigenetics, telomerase contribute here. And really, I think understudied still is the role of immune function and how that can play a role in this connection. So these are some of the emerging topics. I'll mention just really kind of Sirtuin and then wrap up. But it in a way it's kind of a second clue from long-lived organisms.

We know in C. elegans, Surtuin is the human analog or the human analog (cert one) that really we're talking about is sort of a whole lot of sir-2 in lower organisms. And what we – what has been shown by particularly Leonard Guarente's group and others is that genetic alteration in the sir-2 gene can impact lifespan. So when they introduce an extra sir-2 and they see extended lifespan when they knockout, mutate the sir-2 they see a trend toward decreased lifespan. And indeed in mice the sir-1 is engineered mice. But the sir-1 overexpression they're smaller. They have lower body fat, lower insulin, glucose, insulin sensitivity and parametabolic rates. There's really been a calorie restriction in that regard. And so the story of cancer is just emerging. They're just beginning to really characterize that. But I think this is a promising additional target besides TOR that bears further work. There is a link between CR and longevity that's been – beginning to be well-established. So lifespan is not extended by calorie restriction in mice that lack the sir-2.

Uh, not mice, these are yeast actually in these studies that lack sir-2. There are sir-2 inactivating compounds, one which is resveratrol, the compound, the phytolectin in red wine that's gotten some claim from the French paradox. They can cut the survival (unintelligible) mice (unintelligible) currently, some data emerging possibly in mice as well. And it appears to do it by mimicking calorie restrictions and how this again from the Guarente group how they think this is working. It in calorie excess like in the obese state NAD is really probably driving glycolysis. Most of it is shown in that path space. And with calorie restriction, you know, a reduction in calories the (unintelligible) NAD really has pushed in towards the sir-2 pathway which is it's sort of a chromosome silencing pathway through NAD acetylation processes and that's been linked to increased longevity. So this I think is an emerging target and an interesting pathway.

And as I've said there are bioactive food components that are known to hit this pathway including resveratrol the bears further study and so that's indicated here. And mentioning bioactive food components – I'll close here – but that there are a number of bioactive food components in fruits and vegetables and other food stuff that can indeed in preclinical studies, you know, have been shown to influence aging and cancer processes. So we mentioned the ñ we heard about, the telomerase story. Sir-1 we just talked about, micro-RNAs also appear to be responsive to a number of bioactive components. And this can certainly impact chromosome stability. There are a number of growth signals and survival signals mTOR which we've talked about in that kinase. There are bioactive food components. Acid we found to be quite a nice influencer.

That's curcumin, the component of curry can also impact mTOR signaling to an extent. There are bioactive food components known to affect cell cycle machinery p53 and Bcl2 for example in terms of cell cycle and apoptotic mechanism. Interrupt 2 has been mentioned and that clearly is a diet response and oxidative stress regulator. Inflammation again the curry compound, curcumin, and a number of compounds, control, can also hit COX-2. So there's I think emerging evidence that these are candidates there may be ways to target some of these aging and cancer processes through an approach (unintelligible). Some even can mimic its own signals, you know, including insulin IGF-1 and steroid (cytogenscene) that can actually play a role in terms of phytoestrogen activity. So I think there's some real possibilities here to manipulate the system. And in fact a number of bioactive food components have been established with anti-aging and anticancer effects. Again these are really limited (pre-clinical) studies or not (unintelligible) bringing this (unintelligible).

And that's really where I think the focus needs to be. If we can translate not only (unintelligible) but, you know, as Pedro mentioned, I'd say the biggest challenge of calorie restriction research is no one wants to do it. It's, you know, it's difficult to maintain a low-calorie diet. And, you know, I think our evidence would save you don't have to go very low to see positive effects. But still if we could complement by some targeting of these key mechanisms with bioactive food components in addition to low-calorie diets and increased physical activity I think we could really have an impact on these processes. So I'll stop there and happy to answer any questions here to my collaborators and (unintelligible). And thank you very much for joining us today and again I look forward to your questions.

I'm working on hang as close to that as I can and I think that's probably the key. It probably has less to do with the extent of restriction and more to do with sort of maintaining a healthy body size and probably more important I mean that's a good marker I think of metabolic state. And we all know that there are individuals that are there probably obese individuals with a more healthy metabolic state and their risk seems to be for cancer anyway seems to be lower. [Dr. Gabriela Riscuta:] So the first question is any age (ready) to start calorie restrictions so would any age be better than other when we should be starting calorie restriction? [Dr. Steve Hursting:] Yes, you know, it's a bit of an experimental phenomena isn't it that we've, you know, we've kind of packaged this thing. I would say the key thing in my mind is really preventing, you know, the – what's so characteristic. I mean the average American is gaining more – almost 2 pounds a year is our rate of increase here. So preventing that and really maintaining sort of a good healthy weight and I would say the key thing within that is a good healthy metabolic state in preventing the sort of the insulin resistance and inflammation and other components that we've talked about is the key thing.

And so whatever it takes to do that. You know, I've certainly am not the same weight I was when I was, you know, 22 years old. I'm working on hang as close to that as I can and I think that's probably the key. It seems like we cannot make recommendations for the general population other than recommending and maintaining an optimal weight, active lifestyle and healthy diet which we keep saying all the time. And I would like also to thank you again to Dr. Joao Pedro de Magalhaes, Dr. Maria Blasco, Dr. Steve Hursting for their great presentation. And And thank you the – to our audience for participating today. I would like to thank you Dr. Harold Seifried is our Acting Chief and the rest of the Planning and Steering Committee and Office of Communications and Education for helping putting this Webinar together.

And if you have additional questions please send it to us. My name is Gabriela Riscuta and you see my email Gabriela dot Riscuta at nih dot gov. Again to remind you this Webinar is being archived and will be available through our Web site in about two weeks. Also you will receive an email with information to – how to request your credit hours. And again thank you all for joining us today. Goodbye. [Dr. Steve Hursting:] Bye-bye. [END OF TRANSCRIPT] [Coordinator:] Thank you for your participation. That does conclude today's conference. You may disconnect at this time.