Biopharmconsortium Blog

Expert commentary from Haberman Associates biotechnology and pharmaceutical consulting.

Posts filed under: Anti-Aging

Obesity, sarcopenia, aging, and health

Fatmouse_1

In our  June 25, 2010 article on the Biopharmconsortium Blog, we discussed the “contrarian” views of Dr. Katherine M. Flegal and her colleagues at the National Center for Health Statistics of the Centers for Disease Control and Prevention (CDC) on the epidemiology of obesity.

According to Dr. Flegal, based on epidemiological data from the National Health and Nutrition Examination Survey (NHANES), people in the overweight class have a lower risk of death than those in either the normal weight or the obese class. These weight classes are determined on the basis of the body mass index (BMI), with underweight at <18.5, normal weight at 18.5-24.9,  overweight at 25-29.9, and obesity at >30.

Dr. Flegal’s conclusions–as summarized in our 2010 article–were mainly based on work published in the 2005-207 period, as well as other analyses of her results published between 2005 and 2010. In January 2013, Dr. Flegal and her colleagues published a report in the Journal of the American Medical Association. This report was based on an analysis of a wide variety of published reports indexed in PubMed and EMBASE that reported all-cause mortality for weight categories based on standard BMI categories.

In this study, the researchers compared all-cause mortality in the normal weight class (BMI 18.5-24.9) with that in the overweight (BMI 25-<30), grade 1 obese (BMI of 30-<35) and grade 2 and 3 obese (BMI of ≥35) classes. They came to similar conclusions as in their earlier studies. Specifically, both obesity (all grades) and grades 2 and 3 obesity were associated with significantly higher all-cause mortality as compare to normal weight. Grade 1 obesity was not associated with higher all-cause mortality, and overweight was associated with significantly lower all-cause mortality.

Reactions to Dr. Flegal’s 2013 study

As usually happens when one of Dr. Flegal’s “contrarian” studies is published, other leaders of the obesity epidemiology and nutrition community who hold the “majority” view react strongly against it. This was detailed, for example, in a feature article  in the 23 May issue of Nature written by science writer Virginia Hughes. On 20 February 2013, a meeting was held at the Harvard School of Public Health “to explain why [Dr. Flegal’s new study] was absolutely wrong”. The organizer of the meeting, Dr. Walter Willett, said in an earlier radio interview, “This study is really a pile of rubbish, and no one should waste their time reading it.” At the meeting, speaker after speaker got up to criticize the Flegal study.

The major concern of Dr. Willett and the other speakers was that Dr. Flegal’s study (and the commentary on that study in the popular press) would serve as a license for the general public and for doctors to let up on weight loss programs, and to undermine public policies aimed at curbing the rate of obesity. Dr. Willett was also concerned that the Flegal studies might be “hijacked by powerful special-interest groups, such as the soft-drink and food lobbies, to influence policy-makers”.

Nevertheless, as also detailed in Ms. Hughes’ article, other researchers accept Dr. Flegal’s results, and see them as part of the evidence for what they call “the obesity paradox”. Although for the general population overweight increases one’s risk of type 2 diabetes, cardiovascular disease, and cancer, overweight in some populations may not be harmful and may even lower the risk of death. These populations especially include people over 50 and especially those over 60 or 70, as well as patients with cardiovascular disease or cancer. We also discussed the decreased association of mortality with weight in older people in our June 25, 2010 article.

Explaining the “obesity paradox”, and the need for better metrics than BMI

In the 23 August issue of Science, Rexford S. Ahima, M.D., Ph.D. and Mitchell A. Lazar, MD., Ph.D. (both metabolic disease researchers at the Perelman School of Medicine, University of Pennsylvania, Philadelphia PA) published a Perspective entitled “The Health Risk of Obesity—Better Metrics Imperative”. The goal of this essay was to enable researchers to find better means to study and to explain the “obesity paradox”, and to use the results of their studies to improve the health of patents with metabolic diseases and their complications (e.g., cardiovascular disease).

These researchers noted that although it is easily measured and widely used, BMI does not adequately measure body composition (especially the proportion of muscle and fat) and the distribution of fat in the body. These factors may be especially important for such health outcomes as development of insulin resistance and type 2 diabetes, and cardiovascular risk. Other researchers, notably Dr. José Viña and his colleagues at the University of Valencia in Spain, who wrote a critical response to Dr. Flegal’s 2013 article, came to similar conclusions. The Spanish researchers criticized Flegal’s studies because they were based on BMI. However, unlike Dr. Willett, they accept the validity of the “obesity paradox”.

Notably, the Ahima and Lazar article includes a figure that shows metabolically healthy people with  normal and obese BMIs, and contrasts them with metabolically unhealthy people with normal and obese BMIs. The main difference between metabolically healthy versus unhealthy people (whatever their BMI) is muscle mass and fitness. The unhealthy subjects exhibit muscle loss, or sarcopenia, and reduced fitness.

The authors note that skeletal muscle accounts for the majority of glucose disposal. Thus loss of muscle mass, or sarcopenia, due to aging and/or physical inactivity, can result in reduced insulin sensitivity, development of diabetes, and poor cardiovascular health. This applies people with poor metabolic health, whether they have apparently normal BMIs or are obese. Metabolically unhealthy individuals–whether of normal BMI or obese–also have excess visceral fat. Excess visceral fat is associated with the metabolic syndrome and development of type 2 diabetes and cardiovascular disease.

Drs. Ahima and Lazar call for better metrics than BMI, in order to assess a patient’s risk of metabolic disease. They cite the “body shape index”, which quantifies abdominal adiposity (and thus visceral adiposity) relative to BMI and height as potentially a better predictor of mortality than BMI. The body shape index is based on measuring waist circumference, and adjusting it for height and weight. They further call for the development of “accurate, practical, and affordable tools to assess  body composition, adipose hormones, myokines, cytokines, and other biomarkers” to use in assessing obesity and other metabolic disorders in order to determine the risk of developing diabetes and cardiovascular disease, and the risk of mortality.

Appreciating the role of muscle mass in health and disease

The analysis of Ahimsa and Lazar also suggests the hypothesis that loss of muscle mass–sarcopenia–due to aging and/or lack of exercise may be a key factor in the development of obesity-related diseases.

There are at least two other recent reports that focus on sarcopenic obesity. The first, a 2012 paper in Nutrition Reviews entitled “Sarcopenic obesity in the elderly and strategies for weight management” is authored by Zhaoping Li, M.D., Ph.D. and David Heber, M.D., Ph.D. of the Center for Human Nutrition, David Geffen School of Medicine, University of California at Los Angeles. The second paper, entitled “Sarcopenic obesity: strategies for management”, by Melissa J. Benton, PhD, MSN and her colleagues (Valdosta State University College of Nursing, Valdosta, GA) was published in 2011 in the American Journal of Nursing. The first of these reports is a scientific review article, while the second is a practically-oriented report for nurses (carrying continuing education credits); the lead author is a nurse with advanced training in education, sports medicine, and gerontology.

The Li and Heber paper covers much of the same ground as the Ahimsa and Lazar Science Perspective, with respect to the inadequacy of BMI as a metric for obesity, and the need to have better measures of body composition (especially with respect to fat versus skeletal muscle). However, it goes beyond this concern for metrics, by focusing on “sarcopenic obesity”, its relationship with a sedentary lifestyle and with aging, and how sarcopenic obesity might be treated.

Loss of muscle mass as a function of aging in sedentary individuals results in age-associated decreases in resting metabolic rate and muscle strength, and is also a major factor in decreases in activity levels.  These factors result in the decreased energy requirement in aging individuals. If (as is usual) calorie intake does not decrease to match the decreased energy requirements, obesity (i.e., accumulation of excess body fat) results. Sarcopenic obesity in aging individuals is associated not only with type 2 diabetes and other metabolic and cardiovascular diseases, but also with loss of independence and increased risk of mortality. It is a major public health challenge in the over-65 population.

Li and Heber discuss various means to measure body composition, and thus to diagnose sarcopenia and sarcopenic obesity. They then go on to discuss ways to treat this condition, via emphasizing resistance training and increased intake of protein, in order to increase muscle mass and the resting metabolic rate. The authors cite resistance training as “the most effective intervention for reversing sarcopenia in the elderly”. Based on evidence in the field, the authors also hypothesize that increased dietary protein (especially the use of protein supplements or meal replacements) is also important in building muscle mass and as a result reducing fat mass.

It is known that increased dietary protein results in maintenance of muscle mass during calorie-restricted diets, as compared to diets with “normal” or inadequate intakes of protein. However, the authors see the need for more research to determine whether a high-protein diet (up to 35% of caloric intake) will be beneficial in improving muscle anabolic responses to resistance exercise in older adults.

The Benton et al. paper also emphasizes the role of resistance training and a high-protein diet in treatment of sarcopenic obesity. However, being a practically-oriented nursing article, it gives specific recommendations for exercise, as well as sources of high-quality protein in the diet. (This article focuses on high-protein foods, not protein supplements.)

This article also states that nurses should be knowledgeable about sarcopenic obesity and its management. They should also educate older patients on utilizing resistance training and dietary protein to prevent or reverse sarcopenia and sarcopenic obesity. This education should also apply to educating now-healthy aging adults on the need to prevent these conditions, since prevention is easier than reversing sarcopenic obesity once it has developed.

It would seem that not only nurses, but also primary care physicians and other doctors need to be aware of these issues as well.

The Benton et al. paper also wisely counsels that patients contemplating diet and exercise programs such as recommended in their article should first consult with their primary care physician. We agree with this recommendation. We also once again emphasize that this blog does not exist to give diet or exercise advice, or to receive comments or guest posts that purport to give such advice.

However, you are welcome to use this article, or better yet the publications we have cited herein, to help your primarily care provider to be aware of issues involving sarcopenic obesity. Some medical facilities also include physical therapists and/or access to gyms with trainers who can help patients with exercise programs, once one’s primary care physician has been consulted.

Conclusions

1. Currently marketed drugs for obesity–and for such conditions as type 2 diabetes, dyslipidemia, and other metabolic diseases that are usually found in obese individuals and metabolically unhealthy individuals with normal BMI–are generally prescribed as adjuncts to diet and exercise. “Diet and exercise” generally means the types of hypocaloric diets and aerobic exercise conventionally prescribed for weight loss. Researchers and physicians may need to take sarcopenic obesity into account when prescribing these drugs for patients with this condition, and in designing and conducting clinical trials. Diet and especially exercise recommendations may be different for patients with sarcopenic obesity than the current recommendations.

2. We have discussed “alternative” (i.e., non-CNS  or gut targeting) antiobesity therapies now in development in several articles on this blog. Unlike CNS-targeting drugs [e.g., lorcaserin (Arena’s Belviq) and phentermine/topiramate (Vivus’ Qsymia)], which are aimed at curbing appetite, these novel therapeutics are designed to increase energy expenditure or to inhibit the biosynthesis of fat. These drugs, if and when they are approved, will be indicated for patients with extreme obesity, such as those who may currently be candidates for bariatric surgery.

Similarly, we have discussed Novartis’ bimagrumab, an anti-muscle wasting drug now entering Phase 3 clinical trials in patients with the rare muscle wasting disease sporadic inclusion body myositis (sIBM). Bimagrumab is also in Phase 2 clinical trials in sarcopenic older adults with mobility limitations. If and when this drug is approved, it will be at least initially indicated for patients with sIBM, and perhaps eventually for older adults with severe sarcopenia (with or without obesity) that has resulted in mobility limitations.

It will be an extremely long time–if ever–before such drugs are approved for the broader obese and obese-sarcopenic population (or those at risk for these conditions). The diet and resistance exercise approaches discussed in this article may be appropriate for many in this broader group of individuals, and are free of drug-related adverse effects. They may also prevent the development of extreme obesity and its complications, as well as loss of independence due to sarcopenia or obese sarcopenia.

3. We have also discussed the development of anti-aging therapies in various articles in this blog. This field has generated a lot of interest in the news lately, because of Google’s launch of the anti-aging company Calico. As we discussed, for example, in our August 15, 2013 aging article, no pharmaceutical company can run a clinical trial with longevity as an endpoint. Companies must test their drugs against a particular aging-related disease. Many such companies test their agents (e.g., drugs that target sirtuins) against type 2 diabetes.

Why develop an “anti-aging” drug for type 2 diabetes rather than a specific antidiabetic drug? The hope is that an “anti-aging” drug approved for treatment of, for example, type 2 diabetes, will have pleiotropic effects on multiple diseases of aging, and will ultimately be found to increase lifespan or “healthspan” (the length of a person’s life in which he/she is generally healthy and not debilitated by chronic diseases).

Given the major role of sarcopenia and sarcopenic obesity in aging-related disability and mortality, those involved in research and development of anti-aging therapeutics need to take preservation and restoration of muscle mass into account, as they study and/or target pathways involved in aging and longevity.

__________________________________________

As the producers of this blog, and as consultants to the biotechnology and pharmaceutical industry, Haberman Associates would like to hear from you. If you are in a biotech or pharmaceutical company, and would like a 15-20-minute, no-obligation telephone discussion of issues raised by this or other blog articles, or of other issues that are important to  your company,  please contact us by phone or e-mail. We also welcome your comments on this or any other article on this blog.

Novartis’ breakthrough therapy for a rare muscle-wasting disease

skeletal muscle

Skeletal muscle. http://bit.ly/15BgVYY

On August 20, 2013, Novartis announced in a press release that the FDA had granted breakthrough therapy designation to its experimental agent BYM338 (bimagrumab) for treatment of the rare muscle wasting disease sporadic inclusion body myositis (sIBM).

sIBM is a rare–but increasingly prevalent–disease. It is the most common cause of inflammatory myopathy in people over 50. sIBM has a yearly incidence of 2 to 5 per million adults with a peak at ages 50 to 70, with male predominance. Muscle wasting caused by sIBM is superimposed upon the sarcopenia (degenerative loss of muscle mass) that typically occurs with aging.

sIBM is characterized by a slowly progressive asymmetric atrophy and weakness of muscles. Typically, patients become wheelchair bound within 10 to 15 years of onset. Death may occur due to falls, respiratory infection, or malnutrition.

The causes of sIBM are not well-understood. In sIBM, an autoimmune process and a degenerative process appear to occur in muscle cells in parallel. In the autoimmune process, T cells that appear to be driven by specific antigens invade muscle fibers. In the degenerative process, holes appear in muscle cell vacuoles, and inclusion bodies containing abnormal proteins are deposited in muscle cells.

Despite the lack of understanding of the causes of sIBM, in recent years researchers have developed potential therapeutic approaches to this disease. These therapeutic strategies are based on the hypothesis that enhancing muscle regeneration can be beneficial in treating muscle-wasting diseases regardless of their cause. Researchers have thus been working on several approaches, principally 1. developing drugs that stimulate myofiber regeneration, and 2. cell-mediated therapies to replace damaged myofibers (e.g., autologous stem cell therapy). It is the first approach that led to the discovery of Novartis’ bimagrumab.

The myostatin pathway

Myostatin is a regulator of muscle growth in mammals and other vertebrates. It is a secreted protein that is a member of the transforming growth factor beta (TGF-β) family. It is secreted in an inactive form, and must be activated via cleavage by a metalloproteinase. The activated myostatin then binds to its receptor, activin receptor type IIB (ActRIIB). The binding of myostatin to ActRIIB on myoblasts initiates an intracellular signaling cascade, which (as with other members of the TGF-β family), includes activation of transcription factors of the SMAD family. The SMADs (SMAD2 and SMAD3) in the myostatin pathway go on to induce myostatin-specific gene regulation, which inhibits the proliferation of myoblasts and their differentiation into mature muscle fibers.

Bimagrumab, the myostatin pathway, and muscle-wasting diseases

Bimagrumab is a novel, fully human monoclonal antibody (MAb), which was developed by the Novartis Institutes for Biomedical Research (NIBR), in collaboration with the human MAb specialist company MorphoSys (Martinsried, Germany). MorphoSys’ HuCAL (Human Combinatorial Antibody Library) technology was used to identify bimagrumab.

Bimagrumab binds with high affinity to the ActRIIB receptor, thus blocking myostatin binding. The researchers working on development of bimagrumab hypothesized that treatment with the drug would have the same physiological result as myostatin deficiency. For example, myostatin knockout mice have a two-fold to three-fold increase in muscle mass, without other abnormalities. Humans with a loss-of-function mutation in myostatin exhibit marked increase in muscle mass, as well as increased strength. These findings suggest that a myostatin receptor antagonist such as bimagrumab would be a potent stimulator of muscle growth.

According to Novartis’ press release, this hypothesis has been borne out in human studies. The FDA granted breakthrough status for bimagrumab based on the results of a Phase 2 proof-of-concept (POC) study that showed that the drug substantially benefited patients with sIBM compared to placebo. The results of this study will be presented at the American Neurological Association meeting on October 14, 2013. Novartis also expects to published the results of the study in a major medical journal later in 2013.

In addition to sIBM, Novartis is developing bimagrumab for the common muscle-wasting disease of aging sarcopenia, as well as for cachexia in cancer and in chronic obstructive pulmonary disease (COPD) patients, and for muscle wasting in mechanically ventilated patients. In particular, the company is sponsoring a Phase 2 POC study (Clinical Trial Number NCT01601600) of bimagrumab in older adults with sarcopenia and mobility limitations. The study is designed to determine the effects of the drug on skeletal muscle volume, mass, and strength and patient function (gait speed). It will also generate data on the safety, tolerability, and pharmacokinetics of bimagrumab in older adults, as well as (via an extended study duration) the stability of drug-induced changes in skeletal muscle and patient function.

As we discussed in the Conclusions section of our August 15, 2013 blog article on aging, aging-related sarcopenia is a major causes of disability and death. We also said in that section that sarcopenia is not normally a target for drug development. At that time, we did not know about Novartis’ development of bimagrumab. We are happy to be proven wrong about drug development for sarcopenia.

Another approach to myostatin pathway-based drug development

A fully-human anti-myostatin MAb, Regeneron/Sanofi’s REGN1033 (SAR391786), is in Phase 1 clinical development for treatment of sarcopenia. Unlike bimagrumab, which binds to the myostatin receptor ActRIIB, REGN1033 binds directly to myostatin. REGN1033 thus represents an alternative approach to treatment of sarcopenia and other muscle-wasting conditions via the myostatin pathway.

Attempts to address the causes of muscle degeneration in sIBM directly

Despite the evidence from early clinical trials that therapies that enhance muscle regeneration may be effective in treating sIBM, some researchers believe that it will be necessary to identify the causes of muscle degeneration in sIBM and to address them. For example, there is evidence that in some patients, autoantibodies may recognize antigens that are enriched in regenerating muscle fibers. Some researchers therefore hypothesize that treating such patients with therapies that enhance muscle regeneration without addressing the autoimmune pathology may be counterproductive. Therefore, continuing research on the causes of muscle degeneration in sIBM and on potential therapies to slow this degeneration may still be important, despite the apparent progress of clinical trials of such drugs as bimagrumab.

For example, some researchers hypothesize that sIBM is a primary degenerative disease, like Alzheimer’s and Parkinson’s disease. As with these neurodegenerative diseases, some researchers have found evidence that misfolded proteins may be involved in the pathogenesis of sIBM. This avenue of research has led to the hypothesis that agents that enhance correct protein folding may slow muscle degeneration in sIBM patients. One such agent, CytRx’ arimoclomol has been in clinical trials in sIBM patients. [Arimoclomol is also in clinical trials in patients with amyotrophic lateral sclerosis (ALS)].  Arimoclomol appears to act as a coinducer of chaperone proteins such as heat shock protein 70 (Hsp70). Chaperone proteins promote the correct folding of intercellular proteins.

In a small POC Phase 2a clinical trial in Europe, arimoclomol showed early signs of efficacy, in addition to being well tolerated. There was a trend toward slower degeneration in physical function, muscle strength, and right-hand grip muscle strength in arimoclomol-treated patients as compared to placebo over an 8-month period.

Other researchers are attempting to address the inflammatory aspects of sIBM. For example, there are early clinical trials in progress of  the-anti-lymphocyte agent alemtuzumab (Genzyme’s Campath/Lemtrada) and the anti-tumor necrosis factor agent etanercept (Amgen/Pfizer’s Enbrel).

Meanwhile, additional basic research on the causation of sIBM continues. Some of these approaches may eventually lead to additional drug discovery strategies for this disease.

However, whether or not muscle-enhancing therapies such as bimagrumab might provide adequate treatment for at least some classes of sIBM patients (without addressing the autoimmune and/or degenerative aspects of the causation of the disease) will depend on the results of late-stage clinical trials now in the planning stage.

Conclusions

The development of bimagrumab represents an example of Novartis’ pathway-based rare disease strategy. We discussed this strategy in our July 20, 2009 Biopharmconsortium Blog article. Novartis researchers note that in many cases rare diseases are caused by disruptions of pathways that are also involved in other rare diseases and/or in more common diseases. The researchers therefore develop drugs that target these pathways, and obtain POC for these drugs by first testing them in small populations of patients with a specific rare disease. Drugs that have achieved POC in this rare disease may later be tested in other indications (especially including more common diseases) that involve the same pathway.

As we discussed in our July 20, 2009 article, the first drug that Novartis developed by using this strategy is the interleukin-1β inhibitory MAb drug Ilaris (canakinumab). The company conducted its first clinical trials in patients with cryopyrin-associated periodic syndromes, (CAPS), a group of rare inherited auto-inflammatory conditions that are characterized by overproduction of IL-1β. In 2009, the FDA and the European Medicines Agency approved Ilaris for treatment of CAPS. Since that time, Novartis has been conducing clinical trials of canakinumab in such conditions as systemic juvenile idiopathic arthritis (SJIA), gout, acute gouty arthritis, type 2 diabetes, and several others. Canakinumab had also been tested in rheumatoid arthritis, but these trials have been discontinued.

In the case of bimagrumab, Novartis researchers are targeting the myostatin pathway. The strategy is to first target the rare disease sIBM, and to obtain POC in human studies in that disease. Novartis claims (and the FDA concurs with them) that they have obtained POC in sIBM, and the company plans to present the results of its POC clinical trial later this year, both in a scientific meeting and in a publication. Novartis then plans to complete development of bimagrumab for sIBM, while also developing the drug for other muscle-wasting conditions, especially the more common aging-related condition sarcopenia, which is becoming a major public health problem.

________________________________

As the producers of this blog, and as consultants to the biotechnology and pharmaceutical industry, Haberman Associates would like to hear from you. If you are in a biotech or pharmaceutical company, and would like a 15-20-minute, no-obligation telephone discussion of issues raised by this or other blog articles, or an initial one-to-one consultation on an issue that is key to your company’s success, please contact us by phone or e-mail. We also welcome your comments on this or any other article on this blog.

Does inflammation in the brain cause aging?

Hypothalamic nuclei on one side of the hypothalamus, in 3-D. Source: Was a bee. http://bit.ly/13o91HU

Hypothalamic nuclei on one side of the hypothalamus, in 3-D. Source: Was a bee. http://bit.ly/13o91HU

The Biopharmconsortium Blog has been following novel developments in anti-aging medicine and biology for several years. Much of the interest in this field has centered around sirtuins and potential drugs that modulate these protein deacetylase enzymes. Recently–on May 29, 2013–we published our latest blog article on sirtuins.

However, we have long been aware that studies in the biology of aging reveal that lifespan is controlled by sets of complex, interacting pathways. Sirtuins represent only one control point in these pathways, which might not be the most important one.

Now comes a research article in the 9 May 2013 issue of Nature, on the role of inflammatory pathways in the hypothalamus of the brain in the control of systemic aging. This article (Zhang et al.) was authored by Dongsheng Cai, M.D., Ph.D. of the Albert Einstein College of Medicine (Bronx, NY) and his colleagues. The same issue of Nature contains a News and Views mini-review of Zhang et al., authored by Dana Gabuzda, M.D. and Bruce A. Yankner, M.D., Ph.D. (Harvard Medical School).

The role of the neuroendocrine system–and other tissues–in the regulation of lifespan

The News and Views review begins with the statement that classic studies of aging-related pathways in Caenorhabditis elegans by such pioneering researchers as Cynthia Kenyon and Leonard Guarente, as well as later studies in Drosophila suggested that genetic changes that affect the function of nutrient-sensing and environmental-stress-sensing neurons can regulate aging of the entire organism.

In our May 11, 2010 Biopharmconsortium Blog article that reviewed the aging field, we focused on biochemical pathways that may affect lifespan, not the specific tissues in which they act. That article included a link to a 25 March 2010 review in Nature by Dr. Kenyon, entitled “The genetics of ageing”. As we said in our article, that review “discussed the panoply of aging-related pathways in worms, flies, and mice, especially the insulin/insulin-like growth factor-1 (IGF-1) and TOR pathways, as well as several other biomolecules and biological processes”. However, Dr. Kenyon’s article, and several of the references she cites, also deal with the tissues in which these pathways act.

Numerous leading researchers have found evidence that IGF signaling in the C. elegans nervous system regulates longevity. A 2008 article by Laurent Kappeler (INSERM U893, Hopital Saint-Antoine, Paris, France) and his colleagues referenced in the Kenyon review also found evidence that IGF-1 receptors in the brain control lifespan and growth in mice via a neuroendocrine mechanism.

Nevertheless, there is evidence that the insulin pathway in such tissues as the intestine of C. elegans can also regulate lifespan in a non-cell autonomous manner. These studies indicate that changes in insulin pathway gene expression in one tissue–perhaps with certain tissues (the neuroendocrine system, endoderm, adipose tissue) being especially important–results in coordinated changes in insulin pathway activity among all the relevant tissues of the organism. These studies complicate the determination that the neuroendocrine system is the locus of longevity regulation by the insulin pathway and other aging-related pathways.

The potential role of the hypothalamus in the regulation of mammalian aging

Based on the results of studies of neural regulation of lifespan in C. elegans and Drosophila, Zhang et al. asked whether the hypothalamus may have a fundamental role in the process of aging and in regulation of lifespan. The hypothalamus is a region of the brain that is critically involved in regulating such functions as growth, reproduction and metabolism. An important function of the hypothalamus is to link the nervous system to the endocrine system via the pituitary gland, an endocrine gland that is intimately associated with the hypothalamus, and is the master regulator of the endocrine system. According to the Gabuzda and Yankner News and Views article, the mammalian hypothalamus has similar functions to the nutrient-sensing and environmental-stress-sensing neurons of C. elegans and Drosophila that have been implicated in regulation of aging in those organisms.

Zhang et al. studied the increase in NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) in the hypothalamus of mice as a function of aging. NF-κB is a transcriptional regulator that is involved in cellular responses to stress, and in particular mediates inflammatory responses; it has also been implicated as a driver of aging-related gene expression in mice. The researchers found that the numbers of microglia (central nervous system cells that functionally resemble macrophages) in the hypothalamus increased as the mice aged. These microglia exhibited inflammatory function, expressing activated NF-κB and overproducing tumor necrosis factor alpha (TNFα). In turn, secretion of TNF-α (an inflammatory cell-signaling molecule) stimulated NF-κB-mediated signaling in hypothalamic neurons.

Zhang et al. showed–via using genetic models such as specific gene knockouts–that activation of the NF-κB pathway in hypothalamic neurons accelerated the aging process and shortened lifespan. Conversely, inhibiting the NF-κB pathway resulting in delayed aging and increased lifespan. They further showed that activation of neural NF-κB signaling resulted in declines in gonadotropin-releasing hormone (GnRH) levels. Since GnRH stimulates adult neurogenesis in the hypothalamus and hippocampus, decline in GnRH secretion suppressed neurogenesis. Conversely, hypothalamic administration of GnRH reversed aging-associated declines in neurogenesis.

The researchers also treated old mice with GnRH peripherally (i.e., via subcutaneous injection, rather than via hypothalamic administration). Such treatment with GnRH resulted in amelioration of aging-related changes in muscle, skin, and the brain. Notably, GnRH treatment resulted in amelioration of  aging-related cognitive decline. The researchers hypothesize that peripherally-administered GnRH (a neurohormone that is secreted by specific neurons in the hypothalamus) exerts its anti-aging effects via its action on one or more of the GnRH-responsive brain regions that lack a blood–brain barrier, such as the median eminence, subfornical organ and area postrema.

As pointed out by Gabuzda and Yankner in their News and Views article, a 2011 study showed that dendrites of hypothalamic GnRH-producing neurons extend through the blood–brain barrier. These dendrites are able to sense inflammatory and metabolic signals in the blood. Gabuzda and Yankner hypothesize that inflammatory signals in the periphery (which are known to be associated with aging, and such aging-related conditions as insulin resistance, obesity, and cardiovascular disease) may feed back via these dendrites to downregulate GnRH production in the hypothalamus. Such a feedback loop might be analogous to the coordination between peripheral and neural tissues of aging-related pathways seen in C. elegans by Dr. Kenyon and other researchers.

Despite these hypotheses (as pointed out by Zhang et al.), the mechanisms by which GnRH (especially peripherally-administered GnRH) exerts its anti-aging effects are not well-understood, and need further investigation. The researchers conclude, however, that the hypothalamus can integrate NF-κB-directed immunity and the GnRH-driven neuroendocrine system to program development of aging.

With respect to anti-aging therapy, the study of Zhang et al. suggests two potential therapeutic strategies–inhibition of inflammatory microglia in the hypothalamus, and restoration of levels of GnRH. Given the difficulties of specific targeting of microglia in the hypothalamus (across the blood-brain barrier), the second of these alternatives seems to be the more feasible of these strategies.

The GnRH receptor agonist leuprolide as a potential therapy for Alzheimer’s disease

GnRH has a short half-life in the human body, and thus cannot be used as a medication unless it is delivered via infusion pumps. However, the GnRH receptor agonist leuprolide acetate (AbbVie’s Lupron, Sanofi’s Eligard) has been approved by the FDA since 1985. It is available as a slow-release implant (AbbVie’s Lupron Depot) or a formulation delivered via subcutaneous/intramuscular injection. Leuprolide is approved for treatment of prostate cancer, endometriosis, fibroids, and several other conditions. Although leuprolide is the largest-selling GnRH agonist, there are other approved nanopeptide GnRH agonists, such as goserelin (AstraZeneca’s Zoladex) and histrelin (Endo’s Supprelin and Vantas).

As discussed in a 2007 review by Wilson et al., leuprolide acetate has also been under investigation as a therapeutic for Alzheimer’s disease (AD). Studies in mice indicated that leuprolide modulated such markers of AD as amyloid-β (Aβ) and tau phosphorylation, and prevented AD-related cognitive decline. For example, in a 2006 study in a classic mouse model of AD (Tg2576 amyloid precursor protein transgenic mice carrying the Swedish mutation) by Casadesus et al., the researchers demonstrated that leuprolide acetate halted Aβ deposition and improved cognitive performance.

Although Casadesus et al. attributed the efficacy of leuprolide to its suppression of the production of luteinizing hormone, Wilson et al. speculated that it might be possible that leuprolide works directly via GnRH receptors in the brain. Those receptors had only been identified in 2006 by the first author of the review, Andrea Wilson, and her colleagues at the University of Wisconsin. The direct action of leuprolide on GnRH receptors in the brain to ameliorate aging-related cognitive decline–and perhaps AD itself–is consistent with the 2013 findings of Zhang et al.

Development of a leuprolide-based Alzheimer’s disease treatment by Voyager Pharmaceuticals

As discussed in the review of Wilson et al., a Phase 2 clinical trial in women with mild-to-moderate AD receiving acetylcholinesterase inhibitors and implanted subcutaneously with leuprolide acetate showed a stabilization in cognitive decline at 48 weeks. A subsequent study in men (clinical trial number NCT00076440) was documented on ClinicalTrials.gov between 2004 and 2007. However, no results of this trial were ever posted.

The clinical development of a formulation of leuprolide (known as Memryte) as a treatment for AD had been carried our by a small Durham, NC biotech company called Voyager Pharmaceutical Corporation. Memryte was a biodegradable implant filled with leuprolide acetate, designed to treat mild to moderate AD.

After struggling to develop their treatment for nearly a decade, Voyager ran out of money, and stopped its R&D operations in 2007. In 2009, Voyager acquired a new set of investors, and changed its name to Curaxis. In 2010, Curaxis did a reverse stock merger, which enabled it to become a publicly traded company. Also in 2010, Curaxis attracted $25 million from a Connecticut investment firm. Nevertheless, Curaxis noted in its SEC filing that it would need “at least $48 million through 2014” to complete development of Memryte and file a New Drug Application (NDA) with the FDA.

However, Curaxis failed to find the needed funding and/or a partner to complete its development plans. In July 2012, the company filed for bankruptcy.

The failure of Voyager/Curaxis as a company does not necessarily mean that leuprolide may not be a viable treatment for AD. However, as we noted in earlier Biopharmconsortium Blog articles, development of an AD therapy is an enormously long, expensive, and risky proposition, which is beyond the capacity of a small biotech (such as Voyager/Curaxis) unless it attracts a Big Pharma partner. Moreover, treating AD once it reaches the “mild to moderate” stage is unlikely to work.

As we discussed in our April 5, 2013 article, the FDA has been working with industry and academia to develop guidelines for clinical trials of agents to treat the very earliest stages of AD, before the development of extensive irreversible brain damage. If another company endeavors to develop a formulation of leuprolide (or another GnRH pathway activator) for AD, it would probably do best to aim to treat very early-stage disease using the new proposed FDA guidelines.

Conclusions

Why should drug discovery and development researchers and executives be interested in anti-aging research? No pharmaceutical company will be able to run a clinical trial with longevity as an endpoint. However, the hope is that an “anti-aging” drug approved for treatment of one disease of aging will have pleiotropic effects on multiple diseases of aging, and will ultimately be found to increase lifespan or “healthspan” (the length of a person’s life in which he/she is generally healthy and not debilitated by chronic diseases). Numerous pharmaceutical and biotechnology companies have been working on discovery and development of treatments for major aging-related diseases, such as type 2 diabetes and AD. It would be truly spectacular if a new drug for (for example) type 2 diabetes would also be effective against AD.

Moreover, their are also major aging-related conditions, such as sarcopenia (aging-related loss of muscle mass, quality, and strength) that are not normally targets for drug development, but are major causes of disability and death. It would also be amazing if an anti-aging drug aimed at (for example) AD would be effective against sarcopenia as well. Note that Zhang et al. showed that GnRH treatment ameliorated aging-related changes in muscle in their mouse models. (Update: The statement that sarcopenia is “not normally a target for drug development” is no longer true. See our September 4, 2013 article on this blog, “Novartis’ Breakthrough Therapy For A Rare Muscle-Wasting Disease”.)

The study of Zhang et al. constitutes a new approach to anti-aging biology and target and drug discovery. However, aging biology is complex and not well-understood. Researchers will need to study many aspects of aging-related pathways for anyone to be able to discover and develop successful anti-aging drugs. This includes, of course, the sirtuin field, as well as the role of mitochondria in aging related metabolic imbalance, as exemplified by a recent paper in Cell.  As discussed in our other Biopharmconsortium Blog articles on anti-aging biology and medicine, there are many other avenues for investigation as well.

__________________________________________

As the producers of this blog, and as consultants to the biotechnology and pharmaceutical industry, Haberman Associates would like to hear from you. If you are in a biotech or pharmaceutical company, and would like a 15-20-minute, no-obligation telephone discussion of issues raised by this or other blog articles, or of other issues that are important to  your company,  please contact us by phone or e-mail. We also welcome your comments on this or any other article on this blog.

New findings on mechanism of activation of sirtuins may vindicate Sirtris founders

Sir2, the yeast homologue of SIRT1

Sir2, the yeast homologue of SIRT1

The Biopharmconsortium Blog has from time to time been following novel developments in anti-aging medicine, including attempts to develop activators of sirtuins. However, we have not had an article on sirtuins since December 1, 2010. At that time, we reported on the discontinuation by GlaxoSmithKline (GSK) of its lead sirtuin activator, SRT501, a proprietary formulation of the natural product resveratrol (which is found in red wine).

Sirtuins are nicotinamide adenine dinucleotide (NAD+)–dependent protein deacetylases, which have been implicated in control of lifespan in yeast, the nematode Caenorhabditis elegans, and the fruit fly Drosophila. Mammalian sirtuins have been implicated (via animal model studies) in protection against aging-related diseases such as metabolic and cardiovascular diseases, neurodegeneration, and cancer.

As we discussed in our December 1, 2010 article, GSK acquired the sirtuin-pathway specialty company Sirtris (Cambridge, MA) for $720 million in June 2008. This gave GSK ownership of Sirtris’ sirtuin modulator drugs. As stated in that article, although GSK discontinued development of SRT501, it was continuing  development of Sirtris’ non-resveratrol synthetic selective sirtuin 1 (SIRT1) activators, which in addition to their greater potency, had more favorably drug-like properties.

Recently, resveratrol and synthetic sirtuin activators such as those developed by Sirtris have come to be known as  “sirtuin-activating compounds” (STACs).

Sirtuin-activating compounds (STACs) under a cloud

As we discussed in our February 10, 2010 blog article, researchers at Amgen found evidence that the apparent in vitro activation of SIRT1 by resveratrol depended on the substrate used in the assay. The Amgen group found that the fluorescent SIRT1 peptide substrate used in the Sirtris assay is a substrate for SIRT1, but in the absence of the covalently linked fluorophore, the peptide is not a SIRT1 substrate. Resveratrol did not activate SIRT1 in vitro as determined by assays using two other non-fluorescently-labeled substrates.

Researchers at Pfizer also found that resveratrol and three of Sirtris’ second-generation STACs activated SIRT1 when a fluorophore-bearing peptide substrate was used, but were not SIRT1 activators in in vitro assays using native peptide or protein substrates.The Pfizer researchers also found that the Sirtris compounds interact directly with the fluorophore-conjugated peptide, but not with native peptide substrates.

Moreover, the Pfizer researchers were not able to replicate Sirtris’ in vivo studies of its compounds. Specifically, when the Pfizer researchers tested SRT1720 in a mouse model of obese diabetes, a 30 mg/kg dose of the compound failed to improve blood glucose levels, and the treated mice showed increased food intake and weight gain. A 100 mg/kg dose of SRT1720 was toxic, and resulted in the death of 3 out of 8 mice tested.

The Pfizer researchers also found that the Sirtris compounds interacted with an even greater number of cellular targets (including an assortment of receptors, enzymes, transporters, and ion channels) than resveratrol. For example, SRT1720 showed over 50% inhibition of 38 out of 100 targets tested, while resveratrol only inhibited 7 targets. Only one target, norepinephrine transporter, was inhibited by greater than 50% by all three Sirtris compounds and by resveratrol. Thus the Sirtris compounds have a different target selectivity profile than resveratrol, and all of these compounds exhibit promiscuous targeting.

Finally, as we reported in our December 1, 2010 blog article, NIH researcher Jay H. Chung and his colleagues found evidence that resveratrol works indirectly, via the energy sensor AMP-activated protein kinase (AMPK), to activate sirtuins. Since activation of AMPK increases fatty acid oxidation and upregulates mitochondrial biogenesis, this study suggested that the effect of resveratrol on AMPK may be more important than its more indirect activation of sirtuins in the regulation of insulin sensitivity.

All of these studies left Sirtris/GSK’s STACs under a cloud.

On March 13, 2013, GSK reported that it was shutting down Sirtris and its Cambridge MA facilities, just five years after its $720 million acquisition. GSK also said that it was offering transfers to the Philadelphia area for some of the 60 remaining Sirtris employees. Although GSK was closing Sirtris, it said that it remained confident in Sirtris’ drug candidates. The pharma company said that following Sirtris’ “highly successful” research on the biology of sirtuins, further development of Sirtris’ drug candidates “requires the resource and expertise available from our broader drug discovery organization.” GSK will be “exami[ing] [its] research against a variety of therapeutic conditions, with the aim of moving potential assets into the clinic within the next three to four years.”

New evidence that STACs activate SIRT1 in vitro under certain conditions

On 8 March 2013, the journal Science published a report by Sirtris founder David A. Sinclair, Ph.D. (Harvard Medical School, Boston MA) and his colleagues [from academia and from Sirtris, GSK, and from Biomol (Plymouth Meeting, PA)] that identified conditions under which STACs activate SIRT1 in vitro. This research report was accompanied by a Perspective in the same issue of Science authored by Hua Yuan, Ph.D. and Ronen Marmorstein, Ph.D. (Wistar Institute, Philadelphia, PA).

Dr. Sinclair and his colleagues hypothesized that the fluorophore tags on peptide substrates that were used in the original, successful SIRT1 activation assays might mimic hydrophobic amino acid residues of natural substrates at the same position as the fluorophore (i.e, +1 relative to the acetylated lysine that is engaged by SIRT1). Consistent with this hypothesis, the researchers found that non-fluorophore-tagged natural SIRT1 substrates with a large hydrophobic amino acid residue [i..e, tryotophan (Trp), tyrosine (Tyr), or phenylalanine (Phe)] at positions +1 and +6 or +1 were selectively activated by STACs. Examples of such substrates are peroxisome proliferator-activated receptor γ coactivator 1α acetylated on lysine at position 778 (PGC-1α–K778), and forkhead box protein O3a acetylated on lysine at position 290 (FOXO3a-K290). The PGC-1α–K778 peptide contains Tyr at the +1 position and Phe at the +6 position, and FOXO3a contains Trp at the +1 position. Substitution of these aromatic amino acids on either acetylated peptide with alanine (Ala) resulted in complete abolition of SIRT1 activity.

The researchers identified over 400 nuclear acetylated proteins that are potential SIRT1 targets that support STAC-mediated activation of SIRT1, on the basis of their structure. They tested five of these native sequences and found that three of them supported SIRT1 activation.

Kinetic analysis of SIRT1 activation by STACs in the presence of the above peptide substrates showed that the enhancement in the rate of SIRT1 deacetylation was mediated primarily through an improvement in peptide binding. This is consistent with an allosteric mechanism of activation. In allosteric regulation, an allosteric activator (in this case, a STAC) binds to a regulatory site (also known as an allosteric site) that is distinct from the catalytic site of an enzyme (in this case, SIRT1). Binding of the activator to the allosteric site results in the enhancement of the activity of the enzyme, for example by causing a conformational change in the protein that results in improved biding of the catalytic site to the substrate.

In order to investigate the nature of the hypothesized SIRT1 allosteric site, the researchers screened  for SIRT1 mutant proteins that could not be activated by STACs in the presence of an appropriate peptide substrate. As a result of these studies, the researchers identified a critical glutamate (Glu) residue at position 230 of SIRT1, which is immediately N-terminal to the catalytic core of SIRT1.  Glu230 of SIRT1 is conserved from flies to humans. Replacement of Glu230 with another amino acid, such as lysine or alanine, resulted in attenuation of SIRT1 activation by STACs, independent of the substrate used.  Structural studies identified a rigid N-terminal domain that contains Glu230, and is critical for activation by STACs.

The researchers then studied the effects of STACs on cultured cells (murine myoblasts), expressing either wild-type SIRT1 or mutant SIRT1 in which Glu230 is replaced with lysine (SIRT1-E222K, which is the murine equivalent of human SIRT1-E230K). Cells expressing the mutant SIRT1 did not respond to STACs, but cells expressing wild-type SIRT1 did. Specifically, cells expressing wild-type SIRT1 exhibited STAC-stimulated increases in ATP levels, mitochondrial mass, and mitochondrial DNA copy number, but cells expressing mutant SIRT1 did not. In STAC-treated cells, the researchers found no evidence of SIRT1-independent AMPK phosphorylation. This goes against studies discussed earlier in this article, that indicate that resveratrol works via activating AMPK. They also found no evidence for inhibition of phosphodiesterase isoforms in the STAC-treated cells. This goes against a study, published in Cell in 2012, that indicates that resveratrol ameliorates aging-related metabolic conditions by inhibiting cAMP phosphodiesterases, thus engaging a pathway that activates AMPK.

The researchers conclude that STACs act via a mechanism of direct “assisted allosteric activation” mediated by the Glu230-containing N-terminal activation domain of SIRT1. They further conclude that their findings support the hypothesis that allosteric activation of SIRT1 by STACs constitutes a viable therapeutic intervention strategy for many aging-related diseases. thus apparently vindicating the Sirtris/GSK development program.

However, the authors of the companion Perspective hypothesize that the reason that existing STACs only work with SIRT1 substrates that contain hydrophobic residues at position +1 to the acetylated lysine is because they were identified via screening with a substrate that contained a hydrophobic residue mimetic–i.e., a fluorophore tag. A new screen that is not biased in this way might possibly identify STACs that exhibit selectivity for SIRT1 substrates that contain other sequence signatures. It is possible that such STACs might be better therapeutics for certain aging-related diseases than the current STACs being investigated by Sirtris/GSK. There also remain many unknowns in the biology of SIRT1, and in the biochemistry of STACs –i.e., mechanisms by with certain STACs modulate the activity of biomolecules other than SIRT1 (e.g,, cAMP phosphodiesterases). Such issues might affect the success or failure of any program to develop STACs as therapeutic compounds.

________________________________

As the producers of this blog, and as consultants to the biotechnology and pharmaceutical industry, Haberman Associates would like to hear from you. If you are in a biotech or pharmaceutical company, and would like a 15-20-minute, no-obligation telephone discussion of issues raised by this or other blog articles, or an initial one-to-one consultation on an issue that is key to your company’s success, please contact us by phone or e-mail. We also welcome your comments on this or any other article on this blog.

Identification of a novel Alzheimer’s disease pathway provides potential new avenues for drug discovery

 

Neurofibrillary tangle.

Neurofibrillary tangle.

In August and September of 2012, we published three articles on Alzheimer’s disease on the Biopharmconsortium Blog:

Subsequent to the publication of our articles–on 21 November, 2012–the Wellcome Trust announced the identification of a novel pathway involved in the pathogenesis of Alzheimer’s disease (AD). This research was led by Professor Simon Lovestone and Dr Richard Killick (Kings College, London U.K.), and was published in the online edition of Molecular Psychiatry on 20 November 2012. The Wellcome Trust helped to fund the research.

As we have discussed in earlier articles on this blog, the dominant paradigm among AD researchers and drug developers is that the disease is caused by aberrant metabolism of amyloid-β (Aβ) peptide, resulting in accumulation of neurotoxic Aβ plaques. This paradigm is known as the “amyloid hypothesis”. AD is also associated with neurofibrillary tangles (NFTs) which are intracellular aggregates of hyperphosphorylated tau protein. In contrast to the amyloid hypothesis, some AD researchers have postulated that NFT formation is the true cause of AD. The new research links amyloid toxicity to the formation of NFTs, and identifies potential new drug targets.

The new study is based on the discovery of the role of clusterin–an extracellular chaperone protein–in sporadic (i.e., late-onset, non-familial) AD. The gene for clusterin, CLU, has been identified as a genetic risk factor for sporadic AD via a genome-wide association study published in 2009. Clusterin protein levels are also increased in the brains of transgenic mouse models of AD that express mutant forms of amyloid precursor protein (APP), as well as in the serum of humans with early stage AD.

The researchers first studied the relationship between Aβ and clusterin in mouse neuronal cells in culture. Aβ rapidly increases intracellular concentrations of clusterin in these cells. Aβ-induced increases in clusterin drives transcription of a set of genes that are involved in the induction of tau phosphorylation and of Aβ-mediated neurotoxicity. This pathway is dependent on the action of a protein known as Dickkopf-1 (Dkk1), which is an antagonist of the cell-surface signaling protein wnt. The transcriptional effects of Aβ, clusterin, and Dkk1 are mediated by activation of the wnt-planar cell polarity (PCP) pathway. Among the target genes in the clusterin-induced DKK1-WNT pathway that were identified by the researchers are EGR1 (early growth response-1), KLF10 (Krüppel-like factor-10) and NAB2 (Ngfi-A-binding protein-2)–all of these are transcriptional regulators. These genes are necessary mediators of Aβ-driven neurotoxicity and tau phosphorylation.

The researchers went on to show that transgenic mice that express mutant amyloid display the transcriptional signature of the DKK1-WNT pathway, in an age-dependent manner, as do postmortem human AD and Down syndrome hippocampus. (Most people with Down syndrome who survive into their 40s or 50s suffer from AD.) However, animal models of non-AD tauopathies (non-AD neurodegenerative diseases associated with pathological aggregation of tau, and formation of NFTs, but no amyloid plaques) do not display upregulation of transcription of genes involved in the DKK1-WNT pathway, nor does postmortem brain tissue of humans with these diseases.

The Kings College London researchers concluded that the clusterin-induced DKK1-WNT pathway may be involved in the pathogenesis of AD in humans. They also hypothesize that such strategies as blocking the effect of Aβ on clusterin or blocking the ability of Dkk1 to drive Wnt–PCP signaling might be fruitful avenues for AD drug discovery. According to the Wellcome Trust’s 21 November 2012 press release, Professor Lovestone and his colleagues have shown that they can block the toxic effects of amyloid by inhibiting DKK1-WNT signaling in cultured neuronal cells. Based on these studies, the researchers have begun a drug discovery program, and are at a stage where potential compounds are coming back to them for further testing.

________________________________

As the producers of this blog, and as consultants to the biotechnology and pharmaceutical industry, Haberman Associates would like to hear from you. If you are in a biotech or pharmaceutical company, and would like a 15-20-minute, no-obligation telephone discussion of issues raised by this or other blog articles, or an initial one-to-one consultation on an issue that is key to your company’s success, please contact us by phone or e-mail. We also welcome your comments on this or any other article on this blog.

The great metformin mystery–genomics, diabetes, and cancer

 

Galega officinalis (Goat’s Rue) From JoJan http://bit.ly/l5Ybco

Metformin (Bristol-Myers Squibb’s Glucophage, generics), an oral biguanide antidiabetic drug, is the most widely prescribed agent for treatment of type 2 diabetes. The drug mainly works by lowering glucose production by the liver, and thus lowering fasting blood glucose.

Although metformin–approved in the United States in 1994, and in Europe prior to that–has been used for many years, its mechanism of action is not well understood. In 2005, signal-transduction pioneer Lewis Cantley (Beth Israel Deaconess Cancer Center/Harvard Medical School, Boston MA), and his colleagues–including Reuben J. Shaw (now at the Howard Hughes Medical Institute, The Salk Institute for Biological Studies, La Jolla, CA)–published a report showing that metformin targets the adenosine monophosphate (AMP)-activated kinase (AMPK) pathway in the liver. We discussed this report and its implications in our 2007 Cambridge Healthtech Institute Insight Pharma Report, Diabetes and Its Complications.

AMPK is found in all eukaryotic organisms, and serves as a sensor of intracellular energy status. In mammals, it also is involved in maintaining whole-body energy balance, and helps regulate food intake and body weight. We  have discussed the potential role of AMPK in regulation of lifespan, and as a target in anti-aging medicine and in metabolic disease, in earlier articles on this blog. (See here and here.)

AMPK is activated by increases in the ratio of AMP to ATP, caused by energy stress. Under conditions of energy stress, AMP levels go up, and AMP binds to a specific site on the AMPK γ subunit. This induces a conformational change that exposes the activation loop of the α subunit. This allows an upstream serine/threonine kinase to phosphorylate this activation loop. In several mammalian cell types, including liver and skeletal muscle, that kinase is LKB1. Drs. Cantley and Shaw in 2005 showed that metformin targets the LKB1-AMPK pathway in the liver, and that metformin requires LKB1 to lower glucose production by the liver. However, neither LKB1 nor AMPK is the direct target of metformin, and as of 2005, that direct target was unknown.

A new genetic study that suggests that ATM kinase may affect the ability of patients to respond to metformin

Now–as of February 2011–comes a Nature Genetics paper that indicates that the serine/threonine kinase ATM (ataxia telangiectasia mutated) acts upstream of AMPK to mediate the therapeutic effects of metformin. ATM is a DNA repair protein that is recruited and activated by double-strand breaks in DNA. It initiates activation of the DNA damage checkpoint, leading to cell cycle arrest, followed by DNA repair or apoptosis. Thus the role of ATM in the AMPK pathway and in the therapeutic effects of metformin is surprising indeed.

In the study reported in the Nature Genetics paper, researchers of The GoDARTS and UKPDS Diabetes Pharmacogenetics Study Group and The Wellcome Trust Case Control Consortium 2 performed a genome-wide association study (GWAS) for glycemic response to metformin in type 2 diabetes patients in the U.K. In a population of nearly 4,000 patients, they identified a single-nucleotide polymorphism (SNP) designated rs11212617, which was associated with treatment success. This SNP occurs in a genetic locus that also contains the gene that encodes ATM. In a rat hepatoma cell line, inhibition of ATM by the specific inhibitor KU-55933 (KuDOS Pharmaceuticals, Cambridge, U.K., which was acquired by AstraZeneca in 2005) attenuated metformin-mediated phosphorylation and activation of AMPK.

The analysis by Morris Birnbaum and Reuben Shaw in the 17 February 2011 issue of Nature

The 17 February 2011 issue of Nature contained a Forum entitled “Genomics: Drugs, diabetes and cancer.” This consisted of two analyses of the implications of the Nature Genetics paper for metformin’s mechanism of action, and for understanding diabetes and the connections of the metformin-activated ATM/AMPK pathway with cancer. The first analysis was by Morris J. Birnbaum, M.D., Ph.D. (University of Pennsylvania Medical School, Philadelphia, PA), who does research on the role of AMPK and insulin in energy metabolism and in diabetes. The second analysis is by Dr. Reuben Shaw, mentioned earlier. Dr. Shaw’s research centers around LKB1 [also known as serine/threonine kinase 11 (STK11)]. LKB1, a serine/threonine kinase, is not only a regulator of hepatic glucose production via AMPK, but is also a tumor suppressor. Germline mutations in LKB1 are associated with the familial cancer Peutz-Jegher syndrome, and somatic mutations in LKB1 are present in various other cancers. In particular, the Lkb1 gene is one of the most frequently muted genes in human lung adenocarcinomas.

Dr. Birnbaum’s analysis

Dr. Birnbaum notes that the finding of a role for ATM in metformin responsiveness may be an important clue to the mechanism of action of this drug. However, it may also be a false lead, with ATM having only an indirect effect on metformin’s action. He cites recent evidence that metformin acts independently from LKB1 and AMPK and of transcriptional regulation in general. In these studies, genetic ablation of LKB1 and AMPK was used to show that these mediators are dispensable for metformin’s glucose-lowering activity. Instead, metformin appears to work by inhibiting mitochondrial production of ATP in the liver, thus reducing the level of liver glucose production via gluconeogenesis (which uses ATP). This is in apparent contradiction to the 2005 results of Dr Shaw and his colleagues. Nevertheless, metformin’s inhibition of mitochondrial ATP production increases the ratio of AMP to ATP, and thus activates AMPK. There are also other pathways by which inhibition of mitochondrial ATP production may inhibit gluconeogenesis. Thus the mechanisms by which metformin causes a decrease in glucose production by the liver appear to be very complex, and are not well understood.

Dr. Birnbaum therefore speculates that ATM may affect blood glucose levels via pathways that are parallel to, but not the same as, those modulated by metformin. However, the effects of these other pathways may be synergistic with those modulated by metformin when patients are treated with the drug. Dr. Birnbaum notes that 40 years ago, it was found that patients with ataxia telangiectasia often display a type 2-diabetes-like condition, including insulin resistance. Ataxia telangiectasia is a familial disease caused by germline mutations in ATM. This suggests that  ATM may act to counteract hyperglycemia and insulin resistance.

Dr. Birnbaum concluded that biochemical and cell biology studies should be conducted to determine the nature of the interaction of ATM and the antidiabetic effects of metformin. Key to these endeavors is to determine whether there are any biomolecules other than AMPK that both are influenced by ATM and control metabolism.

Dr. Shaw’s analysis

Dr. Shaw first discusses several animal studies that help elucidate the role in glucose regulation of the biomolecules involved in the putative ATM-LKB1-AMPK pathway. He notes notes that deletion of the Lkb1 gene in mouse liver results in loss of AMPK activity in that organ, and to the development of hyperglycemia and hepatic steatosis–two conditions that are seen in type 2 diabetes. Dr. Shaw also cites the 40-year-old finding about the connection between  ataxia telangiectasia and insulin resistance and diabetes. But as he also mentions the more recent (2006) finding that mice with defective ATM activity show increased insulin resistance and abnormal glucose regulation.

Dr. Shaw then speculates as to how ATM might work to modulate patients’ antidiabetic responses to metformin. He notes that ATM is known to phosphorylate LKB1, which is the key activator of AMPK in the liver. Alternatively, ATM might also regulate AMPK independently of LKB1, and might affect responsiveness of patients to metformin by regulating other relevant targets, independently of AMPK. In this context, ATM is known to phosphorylate other, LKB1 and AMPK-independent components of the insulin signaling pathway.

In the light of these considerations, Dr. Shaw says that it is important to determine whether the rs11212617 genetic variant results in modulation of ATM activity toward AMPK activation or toward other targets relevant to glucose regulation, or indeed whether this SNP affects ATM activity at all.

Dr. Shaw then focuses on the potential relevance of metformin to cancer therapy. Researchers have found, in retrospective studies, that diabetes patients who take metformin have a lower risk of developing cancer than those treated with other antidiabetic medications. Animal studies confirm the anticancer effects of metformin, but–as discussed in a 2010 review by Dr. Michael Pollak (McGill University, Montreal, Quebec, Canada)–they indicate that the anticancer effects of this drug are mechanistically complex. Dr. Shaw asks whether metformin is a general activator of ATM (and/or its targets) in the DNA damage-response pathway, or whether its specific effects on LKB1 and/or AMPK might be responsible for the apparent beneficial effects of metformin on cancer risk.

Dr. Shaw concludes with the statement that future studies of the relationship between metformin action, ATM, LKB1, and AMPK should shed light on the relationship between metformin’s antidiabetic effects and its apparent anticancer effects.

Our conclusions

The finding, based on a genome-wide association study, which suggests that ATM, a kinase best known for its involvement in DNA repair pathways, may also be involved in diabetics’ response to metformin is surprising and intriguing. It may eventually be important in unraveling metformin’s mechanism of action in inhibition of liver gluconeogenesis, and in other antidiabetic activities. This finding indicates a connection between pathways by which metformin exerts its antidiabetic activities, and pathways that are involved in cancer.

Nevertheless, the elucidation of metformin’s mechanism(s) of action in diabetes remains a work in progress. This situation is an example of how science works in the real world (as opposed to textbooks or much of science journalism)–generating more questions than answers.

A drug like metformin, with its complex and still poorly understood mechanism of action, could not have been discovered by modern, post-genomics drug discovery strategies. Metformin was discovered via research on natural products derived from the plant Galega officinalis (known as the French lilac, goat’s rue, and by various other names), which had been known by herbalists for centuries. It is fortunate that researchers were able to study the effects of extracts of this plant, and ultimately to develop metformin, well in advance of the modern era of drug discovery. Diabetics and their physicians now have access to metformin as an inexpensive generic drug.

The continued study of the antidiabetic mechanism(s) of action of metformin may yield additional insights into control of gluconeogenesis and other metabolic pathways. Some of the findings of these studies might be relevant to drug discovery and development, for example the development and use of AMPK activators in metabolic disease and in anti-aging medicine.

Continued study of the mechanism(s) of action of metformin may also be relevant to developing new therapies for cancer. As suggested by Dr. Pollak, although metformin is off-patent and is thus not an attractive agent for development as an oncology drug by pharmaceutical or biotechnology companies, other biguanides or related compounds might be better anticancer compounds, and would be patentable. In addition to identifying such compounds, it will be important to determine and define which groups of cancer patients could best benefit from them (perhaps via biomarkers). It will then be important to conduct personalized medicine hypothesis-testing clinical trials (as discussed in an earlier blog post) designed to obtain proof-of-concept that such compounds can indeed benefit specific groups of patients.

___________________________________

As the producers of this blog, and as consultants to the biotechnology and pharmaceutical industry, Haberman Associates would like to hear from you. If you are in a biotech or pharmaceutical company, and would like a 15-20-minute, no-obligation telephone discussion of issues raised by this or other blog articles, or of other issues that are important to  your company, please click here. We also welcome your comments on this or any other article on this blog.

GlaxoSmithKline stops development of resveratrol drug SRT501

Resveratrol

In statements to Fierce Biotech and to the Myeloma Beacon, GlaxoSmihtKline (GSK) said that it has stopped all development of its proprietary resveratrol formulation SRT501. Thanks also to the “In the Pipeline” blog for the information on the Myeloma Beacon statement.

As you all may recall, GSK acquired the sirtuin-pathway specialty company Sirtris (Cambridge, MA) for $720 million in June 2008. This gave GSK ownership of Sirtris’ sirtuin modulator drugs, including SRT501. GSK also appointed Christoph Westphal, then CEO of Sirtris, as the Senior Vice President of GSK’s Centre of Excellence in External Drug Discovery (CEEDD), and Michelle Dipp, then vice president of business development at Sirtris, as Vice President and the head of the US CEEDD at GSK.

According to the Fierce Biotech article, the precipitating factor in GSK’s decision to halt development of SRT501 was the result of a Phase 2a study of the drug in advanced multiple myeloma. The company suspended the study after several patients developed kidney failure. GSK said that in its analysis, the company concluded that SRT501 “may only offer minimal efficacy while having a potential to indirectly exacerbate a renal complication common in this patient population.” It then said that the company has “no further plans to develop SRT501.”

Instead, GSK intends to focus on development of Sirtris’ non-resveratrol synthetic selective sirtuin 1 (SIRT1) activators, which in addition to their greater potency, have more favorably drug-like properties. In its statement to the Myeloma Beacon, GSK in particular mentioned SRT2104 and SRT2379 as the focus of its continuing activity. According to the Sirtris website, SRT2104 is in Phase 2 studies in metabolic and cardiovascular disease, and SRT2379 is in Phase 1 studies in healthy volunteers. Neither compound is currently being tested in cancer.

We discussed Sirtris’ SIRT1 activators in the context of the anti-aging biology field, in a February 10, 2010 blog post. In summary, the mechanism of action of reseveratrol and of Sirtris/GSK’s sirtuin activators is unclear. They apparently activate multiple targets, and they may not be direct SIRT1 activators at all. Nevertheless, Sirtris’ studies of these compounds in mice indicate that they have efficacy in treatment of metabolic diseases. The Phase 2 clinical trials in humans are still ongoing.

To complicate matters further, a study published in the journal Diabetes in March 2010 by NIH researcher Jay H. Chung and his colleagues indicates that resveratrol works indirectly, via the energy sensor AMP-activated protein kinase (AMPK), to activate sirtuins. Since activation of AMPK increases fatty acid oxidation and upregulates mitochondrial biogenesis, the effect of resveratrol on AMPK may be more important than its more indirect activation of sirtuins, at least in the case of metabolic diseases.

Thus Sirtris/GSK’s “sirtuin activators” are under a cloud.

However, as we discussed in our blog posts of November 8, 2009 and February 10, 2010, basic research on anti-aging biology has yielded ample material for drug discovery which may eventually lead to novel treatments for metabolic diseases, and perhaps for other conditions such as various cancers. For example, several companies are developing AMPK activator drugs. Thus there are other avenues for harnessing basic research on anti-aging pathways to discover and develop novel drugs for multiple conditions, even if the Sirtris compounds prove to be a dead end.

_______________________________________________________

As the producers of this blog, and as consultants to the biotechnology and pharmaceutical industry, Haberman Associates would like to hear from you. If you are in a biotech or pharmaceutical company, and would like a 15-20-minute, no-obligation telephone discussion of issues raised by this or other blog articles, or of other issues that are important to  your company, please click here. We also welcome your comments on this or any other article on this blog.

More on anti-aging research: Continuing controversy, opportunity, and good news

1. Continuing Controversy

In our blog post on 10 February 2010, we discussed the controversy over Sirtris/GlaxoSmithKline’s reseveratrol formulation, and its second-generation sirtuin-1 (SIRT1) activators. Researchers at Amgen and Pfizer found that the apparent in vitro activation of SIRT1 by these compounds was an artifact of the experimental method used by Sirtris researchers. The Amgen group found that the fluorescent SIRT1 peptide substrate used in the Sirtris assay is a substrate for SIRT1, but in the absence of the covalently linked fluorophore, the peptide is not a SIRT1 substrate. Although resveratrol appears to be an activator of SIRT1 if the artificial fluorophore-conjugated substrate is used, resveratrol does not activate SIRT1 in vitro as determined by assays using two other non-fluorescently-labeled substrates.

Last month, I attended two meetings at which this controversy was discussed. One was the Bio-IT World Conference & Expo in Boston. At that conference, Christoph Westphal (then CEO of Sirtris) gave a keynote address. In that presentation, Mr. Wesphal stuck with the story that Sirtis’ compounds and its assays are valid. The day after his presentation, Mr. Westphal resigned as Sirtris’ CEO, and now is the head of GSK’s SR One venture fund. He and other Sirtris and Vertex founders also started the Longwood Founders Fund in February of this year.

At the other meeting (which was Harvard-related), one of the most respected leaders of the longevity-related pathway field (whose name I am withholding) stated that the whole resveratrol/sirtuin-activator story is nonsense. He did, however, concur with our views on anti-aging pathways as expressed in our November 8, 2009 article on this blog. We do not go as far as calling the resveratrol story nonsense, but remain unconvinced of the mechanistic basis for resveratrol action pending further evidence.

Meanwhile, Derek Lowe’s “In the Pipeline” blog has a discussion of Mr. Wesphal’s talk at the Bio-IT conference.

In its 25 March 2010 issue, Nature also has a News Feature centered upon the controversy. This article (written by Cambridge MA-based Nature reporter Heidi Ledford) basically says that the controversy remains unsettled, but that several laboratories are working to resolve the assay issue. These include corporate researchers at Sirtris, Leonard Guarente of MIT (another leader in the longevity-related pathway field, who is co-chair of Sirtris’ scientific advisory board), and Anthony Sauve of Weill Cornell Medical School (also a member of Sirtris’ scientific advisory board).

2. Opportunity

There was a review of longevity-related pathways in the 16 April 2010 issue of Science. It covers all the bases of anti-aging research in yeast, worms, flies, and mammals, with an emphasis on the TOR and insulin-like growth factor-1 (IGF-1) pathways. Sirtuins and resveratrol rate a minimal mention in the review.

Cynthia Kenyon, another leader in the longevity pathway field, published a review on the genetics of aging in a special Nature Insight section on aging in the 25 March 2010 issue. In this review, Dr. Kenyon discussed the panoply of aging-related pathways in worms, flies, and mice, especially the insulin/IGF-1 and TOR pathways, as well as several other biomolecules and biological processes. Dr. Kenyon discusses sirtuins, but notes the unknowns in aging-related mechanisms involving sirtuins, especially in mammals. She also notes the difficulties in interpreting results with resveratrol. In addition to the issue with the assays involving the fluorescent substrate, she notes that although (in studies conducted by Sirtris researchers and their academic colleagues) resveratrol has been found to extend the lifespan of mice fed a high-fat diet, it did not extend the lifespan of mice fed a normal diet. Dr. Kenyon also cited the results of studies with resveratrol in yeast, worms, and flies that are not consistent with the hypothesis that resveratrol extends lifespan by acting as a sirtuin activator.

The bottom line of the discussion in the two reviews in Science and Nature is that lifespan is controlled by sets of complex, interacting pathways. Sirtuins represent only one control point in these pathways, which might not be the most important one. Thus no one company “owns” the anti-aging field in terms of drug discovery and development, and there is a lot of opportunity out there. Even Mr. Westphal stated as much in his Bio-IT World presentation.

Interestingly, Dr Kenyon notes that different closely related animals can have large differences in lifespan. For example, rats live for three years, but squirrels for 25. She speculates that differences in longevity might be easily evolvable, and mechanisms by which lifespan changes during evolution (perhaps involving mutations in regulatory genes or that affect rates of respiration) might constitute novel intervention points.

3. Good News

Now for some good news about aging. In an article in the 25 March 2010 Nature Insight section by James W Vaupel (Max Planck Institute for Demographic Research, Rostock, Germany, University of Southern Denmark, Odense, and Duke University), the author presents evidence that human senescence (i.e., deterioration with age)—at least in advanced countries—has been postponed by a decade. This process, first noted in 1994, is continuing. The factors that are making this possible are prosperity (which promotes good health) and medicine (including medical and surgical interventions to prevent or treat disability, and public health efforts). These two factors enable people to reach old age in better health, as well as promoting better health in older people.

This ongoing postponement of senescence and mortality provides a foundation for ongoing anti-aging research and eventual treatments based on that research. (One must remember, however, that regulatory agencies as well as the practical considerations of drug development will not permit researchers and companies to utilize mortality as an endpoint in clinical trials. Companies must therefore develop putative “anti-aging drugs” for specific diseases associated with aging, such as diabetes, cancer, various cardiovascular indications, and dementia.) The postponement of senescence also has profound implications for how one lives one’s life, as well as for social policy and the practice of medicine.

Update on anti-aging biology, sirtuins, and Sirtris/GlaxoSmithKline

On November 8, 2009, we posted an article entitled “Anti-aging biology: new basic research, drug development, and organizational strategy” on this blog. This article focused on new findings in anti-aging biology, their applications to drug discovery and development, and on how this field has affected the organizational strategy of GlaxoSmithKline (GSK).

GSK acquired Sirtris for $720 million in 2008. Later that year, GSK appointed Christoph Westphal, the CEO and co-founder of Sirtris, as the Senior Vice President of GSK’s Centre of Excellence in External Drug Discovery (CEEDD). The CEEDD works to develop external alliances with biotech companies, with the goal of acquiring promising new drug candidates for GSK’s pipeline. Michelle Dipp, who was the vice president of business development at Sirtris at the time of GSK’s appointment of Dr. Wesphal, became Vice President and the head of the US CEEDD at GSK. Thus GSK has been using its relationship with Sirtris to restructure its organizational strategy, attempting to become more “biotech-like” in order to improve its R&D performance.

Now we learn that several research groups and companies have been questioning whether resveratrol (a natural product derived from red wine which has been the basis of Sirtris’ sirtuin-activator platform), as well as Sirtris’ second-generation compounds, may not modulate the sirtuin SIRT1 at all. Thanks to Derek Lowe’s “In the Pipeline” blog for the information. This issue was also covered in a second post on the same blog. It was also covered by articles in the 15 January 2010 issue of New Scientist and in the January 26, 2010 issue of Forbes. Nature also covered this story in an online news article.

In a report published in December 2009, researchers at Amgen found evidence that the apparent in vitro activation of SIRT1 was an artifact of the experimental method used by Sirtris researchers. The Amgen group found that the fluorescent SIRT1 peptide substrate used in the Sirtris assay is a substrate for SIRT1, but in the absence of the covalently linked fluorophore, the peptide is not a SIRT1 substrate. Although resveratrol appears to be an activator of SIRT1 if the artificial fluorophore-conjugted substrate is used, resveratrol does not activate SIRT1 in vitro as determined by assays using two other non-fluorescently-labeled substrates.

More recently, researchers at Pfizer published a study of SIRT1 activation by resveratrol and three of Sirtris’ second-generation sirtuin activators (which the Pfizer researchers synthesized themselves, using published protocols). These researchers also found that although these compounds activated SIRT1 when a fluorophore-bearing peptide substrate was used, they were not SIRT1 activators in in vitro assays using native peptide or protein substrates. The Pfizer researchers also found that the Sirtris compounds interact directly with the fluorophore-conjugated peptide, but not with native peptide substrates.

Moreover, the Pfizer researchers were not able to replicate Sirtris’ in vivo studies of its compounds. Specifically, when the Pfizer researchers tested SRT1720 in a mouse model of obese diabetes, a 30 mg/kg dose of the compound failed to improve blood glucose levels, and the treated mice showed increased food intake and weight gain. A 100 mg/kg dose of SRT1720 was toxic, and resulted in the death of 3 out of 8 mice tested.

The Pfizer researchers also found that the Sirtris compounds interacted with an even greater number of cellular targets (including an assortment of receptors, enzymes, transporters, and ion channels) than resveratrol. For example, SRT1720 showed over 50% inhibition of 38 out of 100 targets tested, while resveratrol only inhibited 7 targets. Only one target, norepinephrine transporter, was inhibited by greater than 50% by all three Sirtris compounds and by resveratrol. Thus the Sirtris compounds have a different target selectivity profile than resveratrol, and all of these compounds exhibit promiscuous targeting.

Sirtris and GSK dispute the findings of the Amgen and Pfizer researchers. One issue raised by Sirtris is that the Sirtris compounds synthesized by Pfizer may have contained impurities, resulting in the toxicity and lack of specificity of the compounds in vivo. Researchers associated with Sirtris and GSK also contend that although the Sirtris compounds only work with fluorophore-conjugated peptides in vitro, they appear to increase the activity of SIRT1 in cells. However, other researchers assert that since resveratrol interacts with many targets in cells, the results of the cellular assays are difficult to interpret. In the Forbes article, GSK’s CEO Andrew Witty is quoted as calling the dispute over the activity of the Sirtris compounds “a bit of a storm in a teacup”. He says that the compounds that Pfizer tested in mice are not the same ones that Sirtris and GSK are currently testing in clinical trials for treatment of diabetes and cancer. (Sirtris’ compounds in clinical trials, discussed in the next paragraph, are in fact different from the ones tested by the Pfizer researchers.)

Currently, Sirtris is testing its proprietary formulation of resveratrol, SRT501, in a Phase IIa clinical trial in cancer. The company reports that SRT501 lowered blood glucose and improved insulin sensitivity in patients with type 2 diabetes in a Phase IIa trial. Sirtris is also testing a second-generation SIRT1 activator, SRT2104, in Phase IIa trials in patients with metabolic, inflammatory and cardiovascular diseases. SRT2104 was found to be safe and well tolerated in Phase I trials in healthy volunteers. Sirtris is also testing another second-generation SIRT1 activator, SRT2379, In Phase I trials. SRT2379 is structurally distinct from resveratrol and from SRT2104.

As we discussed in our original blog post, Elixir Pharmaceuticals is also developing various sirtuin inhibitors and activators for metabolic and neurodegenerative diseases and for cancer. One of Elixir’s products, the SIRT1 inhibitor EX-527, was in-licensed by Siena Biotech (Siena, Italy) in 2009, and was entered into Phase I clinical trials in January 2010. Siena Biotech is developing this compound for treatment of Huntington’s disease.

Despite the dispute over whether Sirtris’ compounds are real SIRT1 activators, the numerous studies on the biology of sirtuins are still valid. Companies with assays that use native peptide substrates and are amenable to high-throughput screening could use these assays to discover novel sirtuin activators. For example, Amgen published a report in 2008 describing such assays. The ability of companies such as Amgen and Pfizer to commercialize such novel sirtuin activators would depend on whether they could overcome the intellectual property position of Sirtris (and Elixir). Since Amgen and Pfizer are working in this area, this indicates that they believe that they can do so.

The efficacy of high doses of resveratrol in improving metabolic parameters of mice fed a high-calorie diet is also not invalidated by the Amgen and Pfizer studies. However these studies cast doubt on the mechanisms by which resveratrol exerts these effects. The apparent efficacy of SRT501 in improving metabolic parameters in patients with type 2 diabetes in a Sirtris Phase IIa trial is consistent with the mouse studies.

Finally, as we discussed in our November 8, 2009 blog post, longevity is controlled by numerous interacting pathways, which may provide at least several targets for drug discovery. Researchers are hard at work to gain additional understanding of these pathways, and some companies are working to discover and develop compounds that modulate these targets. For example, several companies are developing AMPK activators, as discussed in our original blog post. And numerous research groups are reportedly attempting to find drugs that act similarly to rapamycin in increasing lifespan in mice (the main focus of our November blog post), without rapamycin’s immunosuppressive effects.

Anti-aging biology: new basic research, drug development, and organizational strategy

In the 2 October issue of Science (the “Ardipithecus ramidus issue”), there was a Perspective (authored by Matt Kaeberlein and Pankaj Kapahi) and a Report (authored by Colin Selman and his colleagues) on recent findings in anti-aging biology.

Since the late 1980s, researchers have found that caloric restriction (CR) (reduction in caloric intake while maintaining essential nutrients) slows aging in a variety of organisms—yeasts, nematodes, fruit flies, mice, and most recently rhesus macaques. In the recently published 20-year study in rhesus macaques, CR not only increased lifespan, but also delayed the onset of a suite of aging-related disease conditions—diabetes, cancer, cardiovascular disease, and brain atrophy. This parallels the studies with other organisms.

Researchers who have been studying the CR model have been attempting to elucidate the mechanisms by which CR works to slow the aging process and to retard aging-related disease. They hope to find targets for drugs to mimic the effects of CR in humans, since long-term CR is not practical for most people. Over the years, researchers have discovered several pathways by which CR appears to exert its effects. The Report describes new research results on one such pathway, the mammalian target of rapamycin (mTOR) pathway. The Perspective reviews this research in the context of related recent studies.

In a report published in Nature earlier this year (16 July 2009), researchers found that rapamycin administered in food increased the median and maximal lifespan of genetically heterogeneous laboratory mice, whether it was fed to middle-aged (600 days old) or young adult (270 days old) mice. Rapamycin feeding beginning at 600 days of age led to an increase in lifespan of 14% for females and 9% for males, on the basis of age at 90% mortality.

Rapamycin targets mTOR (mammalian target of rapamycin), a kinase that regulates signaling pathways that affect many cellular processes. mTOR forms two protein complexes that are active in intracellular signaling—mTORC1 and mTORC2. It is mTORC1 that is most sensitive to rapamycin. mTORC1 works to coordinate cellular growth and survival responses induced by changes in the availability of nutrients, and also responses to cellular stresses (e.g., hypoxia, DNA damage and osmotic stress). Genetic inhibition of TORC1 in yeast and invertebrates has been found to extend their lifespan. In particular, in the nematode Caenorhabditis elegans, TORC1 interacts with the insulin pathway (via raptor, a component of TORC1) to control lifespan. The role of the insulin pathway in the enhancement of lifespan by CR in C. elegans has been known for many years. The role of mTORC1 at the junction of nutrient and stress sensing pathways, together with these results in invertebrates and now mice, has led researchers to hypothesize that the mTORC1 pathway may be involved in CR-mediated enhancement of lifespan, and that drugs that modulate this pathway may substitute for CR in lifespan extension.

In other studies, inhibition of the mTOR pathway in mice was found to retard development of such aging-related conditions as cancer, metabolic disease, and cardiovascular disease. This effect has also been seen in studies of CR in mice and in nonhuman primates, as stated above.

Rapamycin is an immunosuppressant that is marketed as Wyeth’s (now Pfizer’s, since the October 2009 merger) Rapimmune, to prevent organ transplant rejection. More recently, a derivative of rapamycin, temsirolimus (Wyeth/Pfizer’s Toricel) has been approved for treatment of renal cell carcinoma. The authors of the Nature paper therefore hypothesized that rapamycin may have extended lifespan in the mice either by working via CR-related pathways that control lifespan, by postponing death from cancer, or both.

The finding that oral rapamycin can retard aging in mice, even when fed to 600-day-old mice (the equivalent of 60 years old in humans) raises hope for the development of anti-aging drugs for human use. However, rapamycin itself cannot be used for this purpose because of its immunosuppressant effects. (In the mouse rapamycin feeding studies, the mice were kept under specific pathogen-free conditions.) If researchers were to attempt to modulate the mTORC1 pathway to extend lifespan, they would therefore need to discover other drugs that modulate that pathway without rapamycin’s side effects. Learning more about specific pathway components that may be targeted to increase lifespan may help researchers discover such drugs.

In the new Selman et al. report, researchers endeavored to learn more about how the mTORC1 pathway might extend lifespan in mice. They constructed knockout mice that lacked S6 protein kinase 1 (S6K1). S6K1 is a downstream target of mTORC1, which upregulates mRNA translation and protein synthesis in response to mTORC1 signaling. The researchers found that deletion of the gene for S6K1 resulted in a 19% increase in median lifespan in female mice (as compared to wild-type females), and also increased maximum lifespan. S6K1 deletion had no effect on the lifespan of male mice. This was in contrast to the study with rapamycin feeding, which showed lifespan extension in both sexes, even though the effect in female mice was greater. However, the results of the two studies are not strictly comparable, since mice of different genetic background were used in the two studies.

Female S6K1 knockout mice also showed improvement in several biomarkers of aging (e.g., motor and neurological function, level of physical activity, insulin sensitivity, glucose tolerance, fat mass, immunological parameters). Hepatic gene expression in 600-day-old female S6K1 knockout mice resembled that of wild type mice subjected to CR. Female S6K1 knockout mice showed increased hepatic, muscle, and adipose tissue expression (as compared to wild-type mice) of genes associated with other pathways associated with longevity, including genes for sirtuin-1 (SIRT1) and adenosine monophosphate-activated protein kinase (AMPK).

Selman et al. went on to obtain evidence that the effect of S6K1 knockout on lifespan in female mice is due to activation of AMPK. The gene expression profile of muscle tissue of long-lived female S6K1 knockout mice resembled the profile of wild-type mice treated with the AMPK activator aminoimidazole carboxamide ribonucleotide (AICAR). Hepatocytes from S6K1 knockout mice also showed enhanced AICAR activation of AMPK as compared to hepatocytes from wild type mice. A parallel study in C. elegans showed that deletion of the aak-2 gene, which encodes a subunit of AMPK, suppresses lifespan extension in mutants that lack rsks-1, the nematode homolog of S6K1. These results suggest that S6K1 knockout may exert its pro-longevity effects via activation of AMPK.

AMPK is found in all eukaryotic organisms, and serves as a sensor of intracellular energy status. In mammals, it also is involved in maintaining whole-body energy balance, and helps regulate food intake and body weight. AMPK has been implicated in metabolic response to CR in eukaryotic organisms from yeasts to humans, and it mediates the effects on lifespan of at least one type of CR regimen in C. elegans. Thus the hypothesis that lifespan extension via the mTORC1-S6K1 pathway works via AMPK activation is an attractive one.

However, it is not known how deletion of S6K1 (or its inhibition via mTORC1 in rapamycin-treated mice) might activate AMPK. Moreover, as pointed out by Kaeberlein and Kapahi, there are other downstream targets of S6K1 that might play a role in anti-aging effects of SK61 deletion or inhibition. Among these is hypoxia-inducible factor-1α (HIF-1α). Moreover, there are other biomolecules and pathways that have been implicated in the effects of CR on retarding aging. These especially include the sirtuins, an evolutionarily conserved family of nicotinamide adenine dinucleotide (NAD+)-dependent protein deacetylases.

As shown by the Perspective and Report in the 2 October issue of Science, anti-aging research is an exciting area of basic biological research, and researchers still have much to learn about pathways that mediate the effects of CR on longevity. However, this field is already being applied to drug discovery and development. A basic issue in applying anti-aging research to the development of drugs is that one clearly cannot use increased lifespan as an endpoint in clinical trials. Companies must test putative anti-aging drugs against one or more diseases of aging. The hope is that any “anti-aging” drugs approved for treatment of one disease of aging will have pleiotropic effects on multiple diseases of aging, and will ultimately be found to increase lifespan or “healthspan” (the length of a person’s life in which he/she is generally healthy and not debilitated by chronic diseases).

The two principal types of “anti-aging” drugs currently in company pipelines are sirtuin modulators and AMPK activators. Sirtris Pharmaceuticals (Cambridge, MA, a wholly-owned subsidiary of GlaxoSmithKline [GSK]) is developing the SIRT1 activators SRT501 (a proprietary formulation of the natural product resveratrol) and SRT2104 (a novel synthetic small-molecule SIRT1 activator that is structurally unrelated to resveratrol and is up to 1000-fold more potent). SRT501 is in Phase II clinical trials in type 2 diabetes. SRT2104 has been tested in Phase I trials in healthy volunteers, and was found to be safe and well tolerated. Elixir Pharmaceuticals (Cambridge, MA) is developing a preclinical-stage SIRT1 inhibitor for treatment of Huntington’s disease and certain cancers, and a preclinical-stage SIRT1 activator for treatment of type 2 diabetes and obesity. Elixir also has a research-stage SIRT2 inhibitor under development for treatment of type 2 diabetes and obesity.

Companies developing AMPK activators include a collaboration between Metabasis Therapeutics (La Jolla, CA; about to be acquired by Ligand Pharmaceuticals, San Diego, CA) and Merck–preclinical oral AMPK activators, for treatment of type 2 diabetes and hyperlipidemia), Mercury Therapeutics (Woburn, MA)–research and preclinical-stage oral AMPK activators for treatment of type 2 diabetes, and Betagenon (Umea, Sweden)–the preclinical-stage oral AMPK activator BG8702, for treatment of type 2 diabetes.

The relationship between sirtuin-modulator developer Sirtris and GSK represents a prime example of the attempt of large pharmaceutical companies to become more “biotech-like” in order to improve their R&D performance. We discussed this strategy in our recent report, Approaches to Reducing Phase II Attrition. GSK acquired Sirtris for $720 million in June 2008. In December 2008, GSK announced that it had appointed Christoph Westphal, the CEO and co-founder of Sirtris, as the Senior Vice President of GSK’s Centre of Excellence in External Drug Discovery (CEEDD). The CEEDD works to develop external alliances with biotech companies, with the goal of acquiring promising new drug candidates for GSK’s pipeline. Michelle Dipp, who was the vice president of business development at Sirtris at the time of GSK’s appointment of Dr. Wesphal, is now Vice President and the head of the US CEEDD at GSK. Dr. Westphal, who is also a former venture capitalist, remains as CEO of Sirtris, and is based at Sirtris’ Cambridge location.

Thus anti-aging research, despite the fact that it is mainly in the basic research stage, is not only beginning to produce drug candidates, but has also been having an impact on the organizational strategy of one of the major pharmaceutical companies, GSK.