Dwarfism Gene Linked to Protection from Cancer and Diabetes

ScienceDaily (Feb. 16, 2011) — A 22-year study of abnormally short individuals suggests that growth-stunting mutations also may stunt two of humanity's worst diseases.

 long-term study finds extremely low incidence of cancer and diabetes among individuals with a growth-stunting genetic defect. The authors ask whether controlling growth hormone in healthy adults might provide similar protection.

Published in Science Translational Medicine, the study raises the prospect of achieving similar protection in full-grown adults by other means, such as pharmaceuticals or controlled diets.

The international study team, led by cell biologist Valter Longo of the University of Southern California and Ecuadorian endocrinologist Jaime Guevara-Aguirre, followed a remote community on the slopes of the Andes mountains.

The community includes many members with Laron syndrome, a deficiency in a gene that prevents the body from using growth hormone. The study team followed about 100 such individuals and 1,600 relatives of normal stature.

Over 22 years, the team documented no cases of diabetes and one non-lethal case of cancer in Laron's subjects.

Among relatives living in the same towns during the same time period, 5 percent were diagnosed with diabetes and 17 percent with cancer.

Because other environmental and genetic risk factors are assumed to be the same for both groups, Longo and his team concluded that -- at least for adults past their growing years -- growth hormone activity has many downsides.

"The growth hormone receptor-deficient people don't get two of the major diseases of aging. They also have a very low incidence of stroke, but the number of deaths from stroke is too small to determine whether it's significant," Longo said.

Overall lifespan for both groups was about the same, with the abnormally short subjects dying more often from substance abuse and accidents. The study did not include psychological assessments that could have helped explain the difference.

"Although all the growth hormone deficient subjects we met appear to be relatively happy and normal and are known to have normal cognitive function, there are a lot of strange causes of death, including many that are alcohol-related," Longo said.

Longo noted that any treatment for preventive reduction of growth hormone would have to show fewer and milder side effects than drugs used against a confirmed disease.

But he added that any preventive treatment would target adults with high growth hormone activity in order to bring it down to average, and not to the extremely low and potentially riskier state observed in Laron's subjects.

If high growth factor levels "become a risk factor for cancer as cholesterol is a risk factor for cardiovascular diseases," drugs that reduce the growth factor could become the new statins, Longo said.

Such drugs would be used at first only for families with a very high incidence of cancer or diabetes.

And because growth hormone activity decreases naturally with age, any preventive treatment would be appropriate only until the effects of advanced age took over, Longo explained.

Animal studies provide evidence for the health benefits of blocking growth hormone. Groups led by John Kopchick of Ohio University and Andrzej Bartke of Southern Illinois University achieved a record 40 percent lifespan extension with growth factor deficient mice in studies published in 2000 and 1996, respectively.

Later, the researchers linked growth factor deficiency to reduced tumor risk.

The Food and Drug Administration has already approved drugs that block growth hormone activity in humans. These are used to treat acromegaly, a condition related to gigantism.

Because studies have shown that growth hormone deficiency protects mouse and human cells against some chemical damage, Longo said his team would initially seek approval for a clinical trial to test such drugs for the protection of patients undergoing chemotherapy.

Growth hormone-blocking drugs such as pegvisomant appear to be well tolerated, Longo said. But even if chronic growth hormone blocking should come with a minor side effect, Longo predicted that societies and governments would make the trade in exchange for less chronic disease.

He called it the "square survival curve," where most of one's life is lived without major illness.

"It's the dream of every administration, anywhere in the world. You live a long healthy life, and then you drop dead," Longo said.

Exactly how growth hormone deficiency might protect a person is not fully understood.

In test tube studies, Longo's team found that serum from Laron's subjects had a double protective effect: it protected DNA against oxidative damage and mutations but it promoted the suicide of cells that became highly damaged.

Laron's subjects tend to have very low insulin levels and low insulin resistance, which may explain the absence of diabetes.

In joint experiments with a group led by Rafael de Cabo at the National Institute on Aging, human cells exposed to the Laron's serum also showed surprising changes in the activity of genes linked to life extension in yeast and other model organisms. Although Longo and colleagues had identified such genes 15 years ago, they had not been shown to be important for disease prevention in humans.

Artificial hormone blocking is not the only way to reduce these hormones in humans.

A natural method appears to achieve the same effect: restriction of calories or of specific components of the diet such as proteins.

Several studies are underway to assess the effect of dietary restriction in humans and other primates. The results are not yet known, but a recent study by Longo's group showed that fasting induces rapid changes in growth factors similar to those caused by the Laron mutation.

However, because fasting or restriction in particular nutrients for long periods can lead to dangerous conditions including anorexia, reduced blood pressure and immunosuppression -- and because individuals with rare genetic mutations can suffer life-threatening effects from even short periods of fasting -- Longo emphasized that additional studies are needed and that any changes in diet must be approved and monitored by a physician.

The study in Science Translational Medicine began as an attempt by Longo to test evidence from animal studies that longevity mutations prevent progressive DNA damage and/or cancer.

Co-author Guevara-Aguirre wanted to understand the reasons for the stunted growth of children in the remote community, centered in the Loja province of southern Ecuador.

Initially, Longo said, the children "were more looked at in search of problems than solutions."

But as the study wore on, Guevara-Aguirre began to notice that the adults in the community were not dying of the usual chronic diseases.

That was the clue Longo had been seeking. After hearing of the Ecuador study, he invited Guevara-Aguirre to present at a symposium on aging and cancer in 2006 at USC's Leonard Davis School of Gerontology, where Longo is associate professor.

Together, they obtained funding from the Center of Excellence in Genomic Science in the USC College of Letters, Arts and Sciences, which sponsored part of the initial field research in Ecuador, and from the National Institute on Aging, which sponsored the cellular studies.

Longo and Guevara-Aguirre's collaborators were co-lead author Priya Balasubramaniam, postdoctoral researcher in Longo's laboratory in the USC Leonard Davis School of Gerontology; Sue Ingles, associate professor in the Keck School of Medicine of USC; Min Wei, research assistant professor, Federica Madia, research associate, and Chia-Wei Cheng, graduate student, all in Longo's lab; Marco Guevara-Aguirre and Jannette Saavedra of the Institute of Endocrinology, Metabolism and Reproduction, in Quito, Ecuador; David Hwang and Pinchas Cohen of the David Geffen School of Medicine at UCLA; Rafael de Cabo of the National Institute on Aging; and Alejandro Martin-Montalvo of the National Institute on Aging and the Centre for Biomedical Research on Rare Diseases in Sevilla, Spain.

Balasubramaniam and Marco Guevara-Aguirre were responsible for major parts of the study in the laboratory and in the field, respectively.

New Pneumococcal Vaccine Approach Successful in Early Tests; Vaccine Inhibits Bacteria by Mimicking Naturally-Acquired Immunity

ScienceDaily (Feb. 16, 2011) — Pneumococcus (Streptococcus pneumoniae) accounts for as much as 11 percent of mortality in young children worldwide. While successful vaccines like Prevnar® exist, they are expensive and only work against specific pneumococcal strains, with the risk of becoming less effective as new strains emerge. Through a novel discovery approach, researchers at Children's Hospital Boston and Genocea Biosciences, Inc., in collaboration with the international nonprofit organization PATH, developed a new vaccine candidate that is potentially cheaper and able to protect against any pneumococcal strain.

Tested in mice, the protein-based vaccine successfully inhibited S. pneumoniae from establishing a foothold in the body, the researchers report in the February 17 issue ofCell Host & Microbe.

The current multivalent conjugate pneumococcal vaccines work by inducing people to make antibodies against the sugars on the bacterium's outer capsule. The antibodies then help fight off development of disease after the bacteria have colonized the body. But these vaccines are complex to manufacture, requiring separate individual components for sugars produced by multiple pneumococcal strains. Since pneumococci can make more than 90 different types of sugars, the vaccines may become less effective over time.

The new protein-based vaccine takes a different approach. Based on close to a decade of research at Children's Hospital Boston and utilizing Genocea's novel vaccine discovery technology developed at Harvard Medical School, it stimulates a group of cells in the body known as TH17 cells. These cells provide natural immunity to pneumococcal infection by clearing the bacteria from the surfaces of the upper respiratory tract where infection starts.

Six years ago, Children's Richard Malley, MD, and colleagues showed in mice that while antibodies against surface proteins can protect against pneumococcal disease, there is another mechanism of protection that doesn't require antibodies: the body has natural defenses that act as security guards, preventing the bacteria from becoming squatters in the upper respiratory tract. More recently, they showed that this protection is centered in TH17 cells and production of the chemical messenger IL-17A.

The current study, led by Malley and Kristin Moffit, MD of Children's and Todd Gierahn, PhD, and Jessica Flechtner, PhD, of Genocea Biosciences, began by evaluating a comprehensive library of S. pneumoniae proteins, seeking those that stimulated TH17 cells in mice. They identified specific pneumococcal proteins that activated TH17 cells and used them to make a new vaccine formulation.

When live mice were immunized with these antigens, they showed near-complete protection from S. pneumoniae upper respiratory tract colonization. These same antigenic proteins also potently stimulated human TH17 cells from healthy adult volunteers, causing them to secrete IL-17A.

"The next steps, already in motion, are to optimize the formulation of this vaccine, confirm its efficacy and safety in animals, and then proceed to human trials," says Malley.

In further collaboration with PATH, the researchers will refine and test the most promising formulation in preclinical studies. If the vaccine proves to be effective and safe, the group will prepare an Investigational New Drug (IND) application to the FDA to begin clinical trials.

Unlike existing conjugate vaccine components, the new pneumococcal protein-based vaccine antigens are common to all strains of S. pneumoniae. The researchers hope that a combination of 3 to 5 antigens will protect against pneumococcal colonization and disease from all strains, thereby providing comprehensive immunity with a universal vaccine that would be significantly less complex and expensive to manufacture.

Malley believes that an approach focusing on stimulating TH17 cells or IL-17A secretion may also be effective in providing protection against other pathogens such as Staphylococcus aureus, Mycobacterium tuberculosis, or Listeria monocytogenes.

"By combining advances in molecular biology, immunology and bioinformatics, the strategy we use at Genocea allows comprehensive, rapid, and unbiased screens of every protein produced by an infectious agent to identify the most effective T cell- stimulating antigens," says Flechtner. "We look forward to our continued collaboration and the development of an improved pneumococcal vaccine."

Overabundance of Protein Expands Breast Cancer Stem Cells; Two Drugs Block Cancer-Promoting Chain of Events

ScienceDaily (Feb. 16, 2011) — An essential protein for normal stem cell renewal also promotes the growth of breast cancer stem cells when it's overproduced in those cells, researchers at The University of Texas MD Anderson Cancer Center report in the February edition of Cancer Cell.

In mouse and lab experiments, the team also discovered that two drugs block the cascade of molecular events that they describe in the paper, thwarting formation of breast tumor-initiating cells.

"Overexpression of the EZH2 protein has been linked to breast cancer progression, but the molecular details of that connection were unknown," said senior author Mien-Chie Hung, Ph.D., professor and chair of MD Anderson's Department of Molecular and Cellular Oncology.

"Tumor-initiating cancer cells that arise from the primary cancer stem cells also are thought to drive cancer progression," Hung said. "This research connects EZH2 to the growth of breast tumor-initiating cells."

EZH2 blocks DNA damage repair

The molecular chain of events that improves self-renewal, survival and growth of these breast cancer stem cells can be initiated by oxygen-starved portions of a tumor, Hung said. This hypoxia stimulates a protein that in turn causes overexpression of EZH2.

Abundant EZH2, the team showed, dampens production of an important protein involved in DNA damage repair. Unrepaired chromosomal damage then amplifies production of RAF1, which unleashes a molecular cascade that promotes expansion of breast tumor-initiating cells and cancer progression.

Two drugs slash breast cancer stem cell population

The team tested five anti-cancer drugs against a culture of breast cancer cells and in tumor samples of human breast cancer in a mouse model. Sorafenib, a RAF inhibitor also known as Nexavar, eliminated more cancer stem cells and blocked tumor formation better than the other four.

Sorafenib inhibits multiple targets, so the researchers also tested an experimental drug called AZD6244, which specifically inhibits the MEK-ERK kinase cascade launched by RAF1. They found the drug eliminates EZH2-promoted breast cancer stem cells and blocks the formation of precancerous mammospheres.

"The drugs' inhibition of the breast tumor-initiating cells reveals a previously unidentified therapeutic effect for RAF1-ERK inhibitors to prevent breast cancer progression," Hung said. AZD6244 is being tested in multiple clinical trials, he noted, and it will be interesting to see whether the cancer-stem-cell-killing ability will be induced in those trials.

Hunting down a dangerous pathway

Breast cancer stem cells can be sorted from other primary cancer cells by the presence of the surface protein CD44 and low or absent CD24 in tumor cells.

The researchers found that high expression of EZH2 in breast cancer stem cells reduced the levels of the tumor suppressor RAD1 and correlated with high-grade tumors in a sample of 168 human breast cancer tumors. It also increased the number of tumor-initiating cells in culture.

EZH2 also is heavily expressed, and RAD1 diminished, by lack of oxygen in the tumor's environment, which causes activation of the HIF1a(alpha) protein. HIF1a(alpha), the researchers found, activates the EZH2 gene by binding to the gene's promoter region.

With RAD1 unable to repair damage, amplified RAF1 triggers the MEK-ERK-ß-Catenin pathway, a well-known cancer promoting molecular pathway. The team showed EZH1 enhances this signaling pathway, which they found correlates with breast cancer progression.

In December, Hung and colleagues published a paper in Nature Cell Biology showing that the enzyme CDK1 shuts down EZH2.

Research was funded by grants from the National Cancer Institute, including those for MD Anderson's Specialized Program in Research Excellence (SPORE) for breast cancer and MD Anderson's cancer center support grant; the U.S. Department of Defense, the National Science Council of Taiwan, the Cancer Center of Excellence in Taiwan, National Breast Cancer Foundation, Inc., Kadoorie Charitable Foundation, Beast Cancer Research Foundation and the MD Anderson-China Medical University and Hospital Sister Institution Fund.

Co-authors with Hun, who also is MD Anderson vice president of basic research, are lead author Chun-Ju Chang, Ph.D., Jer-Yen Yang, Ph.D., Weiya Xia, M.D., Chun-Te Chen, Xiaoming Xie, Ph.D., Chi-Hong Chao, Ph.D., and Jung-Mao Hsu, all of MD Anderson's Department of Molecular and Cellular Oncology; Wendy Woodward, M.D., Ph.D., MD Anderson's Department of Radiation Oncology; and Gabriel Hortobagyi, M.D., of MD Anderson's Department of Breast Medical Oncology. Hsu is a student in The University of Texas Graduate School of Biomedical Sciences, a joint program of MD Anderson and The University of Texas Health Science Center at Houston (UTHealth).

He also is affiliated with the Center for Molecular Medicine and the Graduate Institute of Cancer Biology, both of the China Medical University Hospital in Taichung, Taiwan and with Asia University in Taichung.

Designing New Molecular Tools to Study the Life and Death of a Cancer Cell

ScienceDaily (Feb. 16, 2011) — Basic and translational research on cancer, and development of new cancer therapeutics, has focused on different aspects of cancer cellular function. One area of focus is the life and death of a cancer cell. Apoptosis, also known as programmed cell death, is a fundamental process of cells including cancer cells. The signal transduction pathways of apoptosis involve many different proteins and their interactions with each other. Protein-protein interactions involved in these apoptotic signals, like those in many other biological processes, are often determined or influenced by a short fragment of protein sequence or even certain key amino acid residues with important functional or structural roles in the protein-protein interface. For biomedical and pharmaceutical scientists, developing new molecular tools to understand and control the functions of these small protein fragments or residues and the biological and pathological processes that they mediate is a task and challenge of both fundamental interest and practical value.

In the work published in the February issue of Experimental Biology and Medicine, Huang, Zhang, Reed, An and their coworkers have developed new synthetic molecules as models to study the structural and functional role of the proline residue and tetrapeptide sequence important for the regulation of cancer cell apoptosis by the XIAP protein. The work was carried out jointly by the laboratories of Ziwei Huang and Jing An, formerly at the Sanford-Burnham Medical Research Institute in La Jolla, California and now the Cancer Research Institute and Department of Pharmacology of the State University of New York (SUNY) Upstate Medical University in Syracuse, New York, Liangren Zhang at Peking University School of Pharmaceutical Sciences in Beijing, China, and John Reed at Sanford-Burnham Medical Research Institute.

Dr. Huang, who led this international research team, stated "research on protein-protein interactions and their synthetic modulators has become a new frontier for biomedical research and pharmaceutical development. We have chosen a proline containing tetrapeptide as the model to develop new peptidomimetic molecules to study the role of proline and tetrapeptide in the binding of XIAP protein and potential inhibition of XIAP mediated protein-protein interactions critical for apoptotic signaling in cancer cells. Our results suggest that these tetrapeptide analogs can be further developed into new molecular tools to analyze the mechanisms of protein-protein interactions and signal transduction pathways of XIAP in cancer and potential leads to develop anticancer drugs. This study combined the techniques in structure-based drug design, chemistry, and cancer biology and expertise and resources at institutions in America and China. It is an example of international collaboration to apply chemistry to biology and medicine with the long term goal of finding new anticancer therapeutics."

The research team used the crystal structure of a known tetrapeptide AVPI derived from Smac protein bound to XIAP protein as the guide to design a series of peptidomimetic analogs containing a conformationally constrained proline mimetic exo-2-azabicyclo[2.2.1]heptane-3-carboxylic acid. Structural analyses using nuclear magnetic resonance (NMR) and molecular modeling showed that some of these analogs can mimic the conformations of the parent tetrapeptide. Using a fluorescence polarization assay, one of these analogs was shown to be potent like the parent tetrapeptide in binding XIAP protein. This raises the possibility that such an analog may inhibit the antiapoptotic function of XIAP (a protein inhibitor of apoptosis), thus removing the roadblock of the death signal to kill a cancer cell.

Dr. John Reed, who led the binding and biological studies of these molecules in La Jolla, California, said, "The progress made through our collaborations with Dr. Huang and colleagues is a component of a substantial commitment we have made at Sanford-Burnham to discovery and design of small molecule chemical inhibitors of IAP family proteins as potential therapeutics for cancer. We are eager to advance the work towards drug-like leads that might provide renewed hope for those suffering from advanced malignancies."

The design and synthesis of the peptide analogs described in the study are the beginning steps in the long process of research and development of suitable pharmaceutical agents capable of penetrating a cancer cell membrane to reach the XIAP target and triggering the signaling pathway that causes the death of cancer cells in vivo. While further modifications and studies are needed on these peptide analogs in order to show their practical values as cell permeable anticancer agents, the present study of these analogs is of basic research interest for understanding the role of proline and conformation of prolyl peptide bond in mediating the biological function of a protein. It is known that the two different conformational isomers (cis and trans isomers) of the prolyl peptide bond can mediate distinct function of the protein. Many studies in the past of proline and proline mimics show either a mixture of cis and trans isomers or purely cis isomer. In this study, the proline mimic displayed strictly the trans conformation. The interesting conformational and functional effects of the synthetic unnatural mimic of proline discovered here suggest an alternative probe of prolyl isomerization in biology and that such a proline mimic can be applied to study the role of proline and proline containing sequence in other protein-protein interactions involved in a wide range of biological functions.

Steven R. Goodman, Editor-in-Chief of Experimental Biology and Medicine, said "This elegant study by Ziwei Huang and colleagues explores the role of proline containing peptides in inhibiting the anti-apoptotic function of XIAP. This will potentially lead the way to new designer anti-cancer drugs. The article is a wonderful example of the interdisciplinary and international research that is the focus of our journal."

Erg Gene Key to Blood Stem Cell 'Self-Renewal'

ScienceDaily (Feb. 16, 2011) — Scientists from the Walter and Eliza Hall Institute have begun to unravel how blood stem cells regenerate themselves, identifying a key gene required for the process.

Dr. Samir Taoudi from the Walter and Eliza Hall Institute in Melbourne, Australia, has identified a key gene involved in blood stem cells' unique ability to self-renew. 

The discovery that the Erg gene is vitally important to blood stem cells' unique ability to self-renew could give scientists new opportunities to use blood stem cells for tissue repair, transplantation and other therapeutic applications.

Professor Doug Hilton, Dr Samir Taoudi and colleagues from the institute's Molecular Medicine and Cancer and Haematology divisions led the study. Dr Taoudi said the research aimed to understand how blood stem cells are made.

"One of the key features of blood stem cells, one that could be exploited for therapeutic use, is their ability to regenerate or renew themselves," Dr Taoudi said. "However, relatively little is known about how this occurs, or the molecular pathways that specifically control regeneration."

Blood stem cells are required to produce and maintain the blood system throughout an organism's lifetime. They are multipotent cells, meaning they are able to form any cell of the blood system (but not other cells), and they self-renew, so they are a source of endless supply. However, one major barrier to their therapeutic use is that the cells can only be isolated in numbers too low for practical use and efforts to expand the number of cells often causes them to turn into more mature cells.

"At the moment, if you take stem cells from a person and try to expand them, many of the stem cells lose their ability to regenerate," Dr Taoudi said. "The practical aim of our research is to find ways in which you could take stem cells collected from bone marrow or cord blood and 'switch on' expression of particular sets of genes, encouraging the stem cells to expand, essentially creating your own endless supply of blood stem cells."

Institute researchers had previously discovered that ERG was vital for the proper function of adult blood stem cells. They decided to look at blood stem cells in a developing embryo, a time when the cells are particularly active, to determine ERG's role in stem cell production and maintenance.

"We found that during development, ERG was not needed for the original blood stem cells to be made, or to produce mature blood cells," Dr Taoudi said. "But without ERG, these new blood stem cells rapidly decreased as they divided to produce more blood, so that they were almost completely exhausted by the time the mouse was born."

Further testing revealed that two other genes important in embryonic development, GATA2 and RUNX1, were controlled by ERG at the blood producing stage of development.

"These genes are called transcription factors, they are the 'switches' that turn on and off other genes," Dr Taoudi said. "Individually, these genes are not essential for regeneration, but if you lose both, the stem cells are quickly exhausted. This is a key part of the puzzle, but we will continue to work to find out how these genes directly control self-renewal, and the signals that actually tell the stem cell to regenerate."

Dr Taoudi said that although the finding had promise for the future therapeutic use of blood stem cells, there was still a lot of work to be done.

"We have found part of the pathway required for the expansion of blood stem cells under normal conditions, but from a translation perspective, we still need to establish whether increasing expression of these genes will actually lead to expansion in a culture dish," he said.