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A call for global oversight of unproven stem cell therapies

Source: Cell Press
Date: 6/8/2021
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The promotion and marketing of unproven stem cell therapies is a global problem that needs a global solution, say experts in a perspective published June 8 in the journal Stem Cell Reports. The authors of the paper call for the World Health Organization (WHO) to establish an advisory committee on regenerative medicine to tackle this issue and provide guidance for countries around the world.

“The field of regenerative medicine, which entails the manipulation of cells and tissues to obtain therapeutic properties, has been hailed as the most promising research field in modern medicine,” says senior author Mohamed Abou-el-Enein, the executive director of the joint University of Southern California/Children’s Hospital of Los Angeles Cell Therapy Program. “Starting in the early 2000s, however, unregulated stem cell clinics offering untested and poorly characterized treatments with insufficient information on their safety and efficacy began emerging all over the world, taking advantage of the media hype around stem cells and patients’ hope and desperation.”

“Advancing regenerative medicine is key to addressing the chronic disease burden, which the WHO considers to be a top international priority,” says first author Zubin Master, an associate professor of biomedical ethics at the Mayo Clinic. “And even though we have had significant advancement in the field of regenerative medicine, more needs to be done to ensure the safe and timely delivery of evidence-based interventions to patients, many of whom have exhausted all available options.”

The three authors of the commentary are Lawrence Goldstein Science Policy Fellows for the International Society for Stem Cell Research (ISSCR) and ex-officio members of ISSCR’s Public Policy Committee. They decided to write it after realizing that the majority of efforts in this space have focused on the marketing practices of providers and clinics, with much less focus on the tightening of regulations and enforcement to deal with the issue from a global standpoint.

The commentary highlights several proposals, including the harmonization of regulatory definitions surrounding stem cell and regenerative medicine and the importance of balancing the scientific evidence demonstrating safety and efficacy of stem cell and regenerative products with the needs of the patient.

“Many patients with chronic diseases who desire a regenerative medicine option have exhausted conventional medicine treatments and have no other suitable option,” Master says. “We should aim to develop pathways to provide patients with evidenced-based experimental regenerative intervention as possible options where there is oversight, especially in circumstances where there is no suitable alternative left.”

The authors write that the WHO, which already has established an expert advisory committee to develop global standards for governance and oversight of human genome editing, could initiate a similar committee to address the spread of unproven stem cell treatments. They say that the committee could promote greater harmonization among regulatory standards, convince member nations to prioritize this issue on their public health agendas, and develop educational and outreach tools that could help increase awareness among physicians and patients on the danger of this practice.

“I believe that the global spread of unproven stem cell therapies reflects critical gaps in the international system for responding to health crises, which could put the life of thousands of patients in danger,” Abou-el-Enein says. “Urgent measures are needed to enhance the global regulatory capacity to detect and respond to this eminent crisis rapidly.”

The authors also propose the establishment of an active surveillance mechanism to collect and analyze information on dubious clinics performing these procedures and share it with the public to increase awareness, as well as taking the appropriate legal actions. They note the importance of educational programs for patients and physicians on the realistic potential of stem cells and the regulatory pathways that are in place for developing these promising therapies.

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Defects in mitochondria may explain many health problems observed during space travel

Source: Cell Press
Date: 11/25/2020
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For space exploration to be successful, it’s vital to understand–and find ways to address–underlying causes of the health issues that have been observed in astronauts who have spent extended periods of time off world. These problems include loss of bone and muscle mass, immune dysfunction, and heart and liver problems. Using data collected from a number of different resources, a multidisciplinary team is reporting discovery of a common thread that drives this damage: mitochondrial dysfunction. The researchers used a systems approach to look at widespread alterations affecting biological function. The findings are reported November 25 in the journal Cell.

“We started by asking whether there is some kind of universal mechanism happening in the body in space that could explain what we’ve observed,” says senior author Afshin Beheshti (@AfshinBeheshti), a principal investigator and bioinformatician at KBR in the Space Biosciences Division of the National Aeronautics and Space Administration (NASA) at Ames Research Center in California’s Silicon Valley and a visiting researcher at the Broad Institute. “What we found over and over was that something is happening with the mitochondria regulation that throws everything out of whack.”

The investigators analyzed data obtained from NASA’s GeneLab platform, a comprehensive database that includes data from animal studies, the NASA Twin Study, and samples collected from 59 astronauts over decades of space travel. Many of the scientists who participated in this study are involved with GeneLab’s Analysis Working Groups, which draw from institutions all over the world. The platform contains a range of “omics” data related to changes in tissues and cells that occur due to the combined effects of space radiation and microgravity, including proteomic, metabolomic, transcriptomic, and epigenomic data. 

The researchers used an unbiased approach to look for correlations that could explain the widespread changes observed. “We compared all these different tissues from mice that were flown in space on two different missions, and we saw that mitochondrial dysfunction kept popping up,” Beheshti says. “We looked at problems in the liver and saw they were caused by pathways related to the mitochondria. Then we looked at problems in the eyes and saw the same pathways. This is when we became interested in taking a deeper look.”

He explains that mitochondrial suppression, as well as overcompensation that can sometimes occur because of that suppression, can lead to many systemic organ responses. They can also explain many of the common changes seen in the immune system.

Using their discoveries from mice as a starting point, the researchers then looked at whether the same mechanisms could be involved with humans in space. Examining data from the NASA Twins Study, in which identical twins Scott and Mark Kelly were followed over time, the former on the International Space Station and the latter on the ground, they saw many changes in mitochondrial activity. Some of these changes could explain alterations in the distribution of immune cells that occurred in Scott during his year in space. They also used physiological data and blood and urine samples that had been collected from dozens of other astronauts to confirm that mitochondria activity in different cell types had been altered. 

“I was completely surprised to see that mitochondria are so important, because they weren’t on our radar,” Beheshti says. “We were focusing on all the downstream components but hadn’t made this connection.” He adds that mitochondrial dysfunction can also help explain another common problem with extended space travel: disrupted circadian rhythms. Since the team first reported their findings within NASA, other NASA scientists have begun making connections between mitochondrial changes and common space-related cardiovascular problems as well.

The hope is that now that mitochondrial issues have been identified as a cause of so many health risks related to space travel, countermeasures could be developed to address them. “There are already many approved drugs for various mitochondrial disorders, which would make it easier to move them toward this application,” Beheshti notes. “The low-hanging fruit now would be to test some of these drugs with animal and cell models in space.”

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Mitochondria may metabolize ADP differently in aging muscle, despite exercise resistance

Source: Cell Press
Date: 03/13/2018
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Most adults reach their peak levels of muscle mass in their late 30s or early 40s. Even for those who exercise regularly, strength and function start to decline after that point. For those who don’t exercise, the drops can be dramatic. Now, a study of twenty men published March 13 in the journal Cell Reports provides new clues about the cellular mechanisms of aging muscles, showing a key role for how mitochondria, the powerhouses of the cell, process ADP, which provides energy to cells.

ADP, or adenosine diphosphate, plays a role in how our cells release and store energy. But previous lab models that have looked at the mechanisms of aging in human cells have not included ADP. When ADP is metabolized in the mitochondria, it stimulates cellular respiration and decreases reactive oxidative species (ROS; also known as free radicals). Higher ROS levels are linked to damage in different components of the cell, a process also called oxidative stress.

In the study, the investigators developed an in vitro system employing individual muscle fibers taken from muscle biopsies. The fibers were put into a system in which mitochondrial function and respiration could be measured across a range of ADP concentrations that are relevant to those found in the human body. “The way people normally measure ROS is in a system that has ADP removed,” says senior author Graham Holloway, an associate professor at the University of Guelph in Ontario. “But biologically, we always have ADP in the system. We started to think that maybe how we get ADP into the mitochondria is important for aging.”

In the first part of the paper, the researchers compared muscle from ten healthy men in their 20s with muscle from ten healthy men in their early 70s. They found that there was an 8- to 10-fold decrease in ADP sensitivity, and therefore, when ADP was added to the system, there was a 2- to 3-fold higher rate of ROS emission in the muscle taken from the older men. ROS levels were determined by measuring emissions of hydrogen peroxide, a byproduct of activity in the cell.

The findings suggested that mitochondrial ADP sensitivity was somehow impaired in the muscles of the older men and that increased levels of ROS were contributing to sarcopenia, or the degenerative loss of muscle mass. “The magnitude of change was quite striking to us,” Holloway explains. “For humans, it’s remarkable to have such a big difference.”

In the second part, the older men undertook a program of supervised resistance training, which included leg presses and upper-body exercises. But, after 12 weeks, there were no changes in the levels of hydrogen peroxide emitted, suggesting no improvements in age-associated cellular stress.

“This doesn’t mean there’s no hope for building strength in aging muscle,” Holloway says. “I actually think that endurance training would be potentially beneficial, because we know with that kind of training you get increases in mitochondrial content.” Endurance training includes aerobic exercise like cycling and swimming. “Moving forward, we plan to look at other types of exercise, to see if it can improve the dynamic response of mitochondria to ADP,” he adds.

Other future work will use rodent models to delve into the cause-and-effect relationships of the molecular mechanisms of ADP metabolism. The investigators also plan to extend their studies to looking at different types of exercise in aging women. Early research in healthy young people has indicated that there are differences in sensitivity to ADP between men and women.

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Something New Under the Sun: Study in Leukemia Finds Role for Helios Protein

Source: Memorial Sloan Kettering - On Cancer
Date: 11/21/2018
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Proteins are one of the fundamental building blocks of life, controlling many of the vital functions carried out by cells. These activities include cell growth, division, and death. Sometimes the same protein can have more than one job, depending on how it interacts with other proteins in a cell.

The latest example of a protein with a dual role is one called Helios. When it’s missing, Helios can contribute to a type of pediatric blood cancer called B-acute lymphoblastic leukemia (B-ALL). But researchers at Memorial Sloan Kettering are now learning that when Helios is abundant, it can actually drive the formation of a different and a more common type of blood cancer called acute myeloid leukemia (AML). The findings were published in Cell Stem Cell.

“There are not a lot of examples of this type of situation,” says senior author Michael Kharas, a cancer biologist in the Molecular Pharmacology Program in the Sloan Kettering Institute. ”Occasionally, there are proteins that have completely different activities in different kinds of cancer. In this study, we figured out the ways in which the protein works and showed how it turns different genes on and off.”

A Protein That Regulates Genes

Helios is the name for the ancient Greek god of the sun. (Genes and proteins are sometimes given whimsical names, and Helios is a member of a family of proteins that are all named after characters from Greek mythology.) The Helios protein acts as an epigenetic regulator, which means it can control which genes get turned on and which genes get turned off. It exerts control by regulating chromatin, the part of the cell that packages DNA. If you imagine DNA as long strands of yarn, chromatin is the spool that the strands wrap around. When DNA is tightly wound, it’s hard for proteins to get made because the machinery that’s needed to start the process can’t make contact with the DNA. But when it’s unwound, the DNA becomes accessible.

In the new study, investigators found that Helios unwraps DNA from chromatin in areas that are important for the survival of leukemia cells. It also winds up the DNA in locations that are important for blood stem cells to turn into specific cell types. This process is called differentiation.

In someone with AML, the bone marrow produces immature white blood cells called myeloblasts rather than healthy, normal blood cells. Myeloblasts are unable to function like normal blood cells. They grow out of control and crowd out healthy cells. High levels of Helios are present in leukemia stem cells, which are essential for leukemia to grow. These cells are also thought to be the cause for relapse. 

“In the case of AML, Helios controls the program that leukemia stem cells use,” Dr. Kharas explains. “In some areas of the genome, it keeps the chromatin open, and in other places, it keeps the chromatin closed. The genes that are important for the cells’ ability to keep growing are left on, and those that would drive normal differentiation are turned off.”

By contrast, in the pediatric blood cancer B-ALL, it’s the absence of Helios that causes trouble. Earlier research found that Helios was lost in about half of people with a type of B-ALL called hypodiploid B-ALL.

In this study, however, investigators found that knocking down, or deleting, Helios can reduce the number of leukemia stem cells. Furthermore, Helios is required for leukemia cells to survive, and when it’s removed, leukemia cells stop growing and differentiating, and ultimately die.

To confirm that Helios contributes to AML development, the investigators also transplanted human leukemia cells into mice models. They found that when Helios was deleted, the mice had a greater reduction in leukemia cells and lived longer.

What’s Next? Targets for Drug Development

Based on the findings, the researchers hope to develop drugs that target Helios as a treatment for AML. Dr. Kharas doesn’t believe there is any danger in causing B-ALL by blocking Helios, since several other mutations are needed to drive the formation of that cancer. In addition, the mice that had Helios blocked in their blood cells didn’t show any signs of other cancers, including B-ALL, and had normal blood stem cell function. But until further research is conducted, investigators won’t know for sure.

Dr. Kharas notes that other researchers have discovered that Helios plays a role in the proper functioning of a type of white blood cell that affects immune response. This suggests that drugs that influence Helios could also be used to boost immune therapies for cancer.

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A Death Wish That Allows Worms to Thrive — and What It Tells Us About Cancer Biology

Source: Memorial Sloan Kettering - On Cancer
Date: 03/28/2019
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In biology, how cells die is as important as how they live. Cell death provides a counterbalance to cell division, maintaining the proper number of cells throughout an organism’s lifetime. Gaining a better understanding of how cells die is also important for cancer research. It can teach us how the body naturally fends off cancer — by preventing runaway cell growth — as well as point to new ways to target and kill tumor cells.

The lab of Sloan Kettering Institute cell biologist Michael Overholtzer explores the mechanisms of different kinds of cell death. More than a decade ago, while studying breast cancer cells, he was the first to observe a type of cell death called entosis, in which one cell engulfs and kills another.

In a study published in March 2019 in Cell Reports, Dr. Overholtzer described for the first time the role of entosis in the development of a tiny worm called C. elegans. The discovery is significant because until now very little has been known about the role that entosis plays in natural developmental processes.

“Understanding cell death in normal development can provide new clues about how it works and why it evolved,” says Dr. Overholtzer, who was recently named dean of the Gerstner Sloan Kettering Graduate School of Biomedical Sciences. “We hope this research will help us find ways to harness it for cancer treatment.”

Characterizing Different Kinds of Cell Death

The most well-studied form of cell death is called apoptosis. Apoptosis is a type of programmed cell death sometimes likened to cellular suicide, in which a cell breaks down in a regulated, systematic fashion. Apoptosis can occur in response to cell damage, but it’s also a normal part of development in embryos. For example, apoptosis in the hands and feet allows individual fingers and toes to form, by killing cells in the spaces in between them.

In the March 2019 study, the researchers focused on the development of the gastrointestinal and reproductive tracts of the C. elegans worm, a popular lab model for studying development. Research from a team at Rockefeller University had suggested that forms of cell death other than apoptosis were important in the formation of parts of the worm’s body. Dr. Overholtzer’s team decided to continue this line of inquiry.

In particular, the researchers looked at the role of cell death in the formation of the cloaca, the dual-purpose orifice at the hind end of worms, as well as many other organisms, from which excrement is discharged. In males, it is also where sperm are released. It turns out that entosis is vital to ensuring that the genital tract connects to the cloaca. Without it, male worms are sterile.

A Mysterious Process

Unlike apoptosis, entosis requires two cells. It might sound more like a murder than a suicide, but cells that undergo death by entosis still have a death wish. They actually burrow themselves into the other cell, where they are broken down and eaten.

“This is a really enigmatic process,” Dr. Overholtzer says. “Why would a cell decide to do this? It turns out that in the worm, it’s required for normal development.”

In the C. elegans embryo, a type of cell called a linker cell pulls the developing genital tract into the cloaca. The linker cell then burrows into another cell, where it is destroyed by entosis. Destruction of the linker cell creates an opening that allows sperm to enter the cloaca.

“There’s an ongoing question in biology, which is, ’Where do all these different cell death programs come from and why do they exist?’” he notes. “For entosis, this study demonstrates that a process observed initially in breast cancer cells also has a role in normal development.”

Further research will focus on figuring out whether entosis contributes to other developmental processes, as well as molecular changes in the cell that lead to death by entosis. The researchers also plan to study a piece of the linker cell that is left behind after the cell undergoes entosis, called the lobe. “These pieces stick around for a long time, so we think they may have some purpose,” Dr. Overholtzer concludes.

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An Old Protein Gets a New Look: Researchers Target TGF-ß to Make Immunotherapy More Effective

Source: Memorial Sloan Kettering - On Cancer
Date: 04/05/2019
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The immunotherapy drugs called checkpoint inhibitors have transformed treatment for some people with cancer. Currently, a major focus in cancer research is looking for ways to make these drugs effective for a greater number of people and more types of cancer.

One of the many approaches that scientists are now studying is combining immunotherapies with drugs that block the actions of a molecule called transforming growth factor-beta (TGF-ß). The importance of TGF-ß was uncovered more than 30 years ago by Sloan Kettering Institute Director Joan Massagué.

“There has been tremendous interest in this new avenue of treatment,” Dr. Massagué says. “Right now, we have a serious opportunity to leverage the robust knowledge that we’ve built about TGF-ß and use it to make immunotherapy more effective. TGF-ß is actually a hormone that is released by cells. This means it is quite accessible to drugs that can block its activity.”

A Fundamental Molecule with Many Roles

TGF-ß is critical for regulating how cells function. It works by sending signals from a cell’s membrane to its nucleus, telling it which genes to make into proteins. Among the most important jobs of TGF-ß are directing embryonic development and regulating the immune system. It also plays a role in whether cancer spreads.

It is the effect of TGF-ß on the immune system that many researchers are now most interested in studying.

A study published in February 2018 in Nature reported that, in people being treated with the immunotherapy drug atezolizumab (Tecentriq®), levels of TGF-ß in tumors correlated with how well people responded. In short, the more TGF-ß that tumors had, the less likely it was that atezolizumab worked. This is because TGF-ß can prevent the immune cells called T cells from infiltrating and attacking tumors. Checkpoint inhibitors boost the activity of T cells, but if the immune cells can’t get into the tumor, they will not be effective.

Another study in the same issue of Nature reported that combining a TGF-ß-blocker called galunisertib with immunotherapy allowed the immune system in mouse models of colon cancer to attack tumors that it had previously not been able to go after.

“This research showed that TGF-ß prevents the incoming T cells from penetrating and infiltrating the tumor,” Dr. Massagué says. “If you block TGF-ß, however, T cells infiltrate and kill the tumor cells.”

Combining Approaches in the Clinic for Many Cancer Types

Several clinical trials now under way are looking at how to combine immunotherapy with drugs that block TGF-ß. MSK medical oncologist Anna Varghese is the co-principal investigator of one such trial for pancreatic cancer.

“Immunotherapy alone is not effective against pancreatic cancer for most patients,” Dr. Varghese explains. “This is because pancreatic tumors have an immunosuppressive microenvironment.” That means the cells and tissue around the tumor prevent immune cells from getting in and attacking the cancer.

Dr. Varghese’s trial combines the immunotherapy drug durvalumab (Imfinzi®) with galunisertib. The study is still ongoing, and it’s too early to know how effective the combination will be or what side effects it will have, but Dr. Varghese hopes to report preliminary findings soon.

Research on TGF-ß blockers goes beyond immunotherapy to look at their effectiveness when combined with other kinds of drugs. Recently, MSK medical oncologist James Harding was an investigator in a trial that looked at galunisertib, either alone or in combination with other drugs, for the treatment of liver cancer.

Early analysis of the study showed that when galunisertib was combined with the targeted therapy sorafenib (Nexavar®), people had more favorable outcomes compared with other treatments. However, Dr. Harding comments that “the design of this early study makes it difficult to say how impactful this combination strategy will ultimately be for liver cancer.

“Furthermore,” he adds, “with the advent of effective immunotherapy, the focus of the investigation has shifted to pairing galunisertib and other TGF-ß blockers with immune checkpoint inhibitors.”

Clinical trials focusing on TGF-ß blockers are ongoing at other cancer centers as well as at MSK.

Taking a New Look at a Familiar Target

“For a long time, drug companies have been interested in harnessing the power of TGF-ß blockers, not only for cancer but for other diseases as well,” Dr. Massagué says. “Until now, there’s been concern about doing that because we know that TGF-ß has so many different functions in cells.” Long-term use of these drugs, he explains, would have too many harmful side effects.

“However, combining a TGF-ß blocker with checkpoint immunotherapy and using it as temporary way to boost the effectiveness of these other drugs may be possible,” he concludes.

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Ro Versus Musashi: How One Molecule Can Turn Cancer Cells Back to Normal

Source: Memorial Sloan Kettering - On Cancer
Date: 06/19/2019
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Since 2012, Memorial Sloan Kettering cancer biologist Michael Kharas has focused on studying a family of proteins called Musashi. These proteins play a role in acute myeloid leukemia (AML) as well as in many solid tumors, including colorectal, breast, lung, and pancreatic cancers. Musashi proteins function by binding to messenger RNAs. These molecules serve as a template for making proteins.

On June 19, 2019, in Nature Communications, Dr. Kharas’s team reported that they have identified a molecule that appears to block the function of Musashi-2. This protein plays a role in making cancer grow and spread. The compound appears to eliminate tumor cells in human cancer cell lines and in mice.

“This research provides a strategy for how to develop inhibitors for RNA-binding proteins,” says Dr. Kharas, who is in the Sloan Kettering Institute’s Molecular Pharmacology Program. “Historically, it’s been difficult to develop inhibitors to proteins that bind to RNA because of their challenging structural properties.

“We don’t think this particular compound will ultimately make it into clinical trials,” he adds, “but we now have a road map to guide us in future drug development.”

Turning Cancer Cells Back to Normal

This latest work builds on earlier research from Dr. Kharas’s lab, in which the investigators started with more than 150,000 molecules that could potentially block Musashi-2. They then developed a number of tests that could rapidly look for effective molecules in an automated way. Eventually, they settled on a molecule called Ro 08-2750, or just Ro for short.

In the current study, the team used structural biology to look at where Ro binds to Musashi-2. “Based on this research, we have an idea of where to start in designing additional molecules that could be used as drugs,” Dr. Kharas says. “We know the binding region and how the drug fits.”

Researchers know that Musashi-2 plays a role in how aggressive cancer is. The protein is present in more than 70% of people with AML. Solid tumors that contain a high level of the protein are more likely to grow, spread, and resist treatment. It appears that Musashi-2 allows cancer cells to continue growing and resist signals to die.

“Musashi-2 is required for cancer stem cells to survive,” Dr. Kharas explains. Cancer stem cells are cancer cells that have the ability to give rise to all types of cells within a tumor. “When Ro was added to AML cells in a dish, the cells became normal. They stopped growing and died.” The same effects were observed in mice that had AML

A Cooperative Effort among Several Labs

This research was possible due to collaboration among many different experts at MSK. The project was overseen by Gerard Minuesa, a postdoctoral researcher in Dr. Kharas’s lab.

SKI computational chemist John Chodera, SKI structural biologist Dinshaw Patel, and Yehuda Goldgur, Head of MSK’s X-Ray Crystallography Core Facility, helped determine the structure of the Musashi-2 protein and how Ro binds to it. SKI computational biologist Christina Leslie helped with the gene expression data generated from this research.

“Thanks to this study, we’ve shown that it’s possible to develop drugs for these difficult targets,” Dr. Kharas concludes. “It provides a path forward for future work, so we can eventually develop drugs that can be tested in clinical trials in people with cancer.”

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Research Clarifies How IDH Mutations Cause Cancer

Source: Memorial Sloan Kettering - On Cancer
Date: 07/01/2019
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A family of genes called IDH are associated with cancer. These genes make enzymes called isocitrate dehydrogenases. The enzymes help break down nutrients and generate energy for cells. Mutations in IDH genes prevent cells from differentiating, or specializing, into the kind of cells they are ultimately supposed to become.

When cells can’t differentiate properly, they may begin to grow out of control. Scientists are still learning about what controls this process.

Now a team of researchers working in the lab of Memorial Sloan Kettering President and CEO Craig Thompson have made discoveries about how this malfunction occurs, at least in test tubes. Although the work is still in an early stage, they hope their findings will eventually contribute to new approaches for developing cancer drugs.

“Although IDH mutations are not very common overall, there are some diseases where these genetic changes contribute to a significant portion of cases,” says Juan-Manuel Schvartzman, a postdoctoral fellow in the Thompson lab, an instructor in the Gastrointestinal Oncology Service, and the first author of a paper recently published in the Proceedings of the National Academy of Sciences (PNAS). “For these subtypes of cancer, better targeted therapies are needed.”

IDH mutations are found in about one-quarter of people with acute myeloid leukemia (AML), the most common type of leukemia in adults. They may also be found in a type of bile duct cancer called cholangiocarcinoma, a bone cancer called chondrosarcoma, low-grade glioma, and some kinds of lymphoma. The mutations occur much less frequently in more common cancers, such as colon cancer, breast cancer, and lung cancer.

Deciphering Underlying Changes

To learn more about how IDH mutations block differentiation, the investigators studied them in the context of a well-characterized model: cells called fibroblasts that can be coaxed to become muscle cells. By figuring out how the mutations prevent muscle cells from forming properly, the team aimed to get at the underlying defects in cells that these mutations cause.

Earlier research showed that IDH mutations influence cells through epigenetic changes. Epigenetics involves changes in gene expression that do not cause changes in the DNA sequence. Many of these have to do with the way DNA is packaged in the nucleus of a cell. The strands are wrapped around spool-like proteins called histones. Small chemical groups attached to DNA and histones — including fragments called methyl groups — can affect how DNA is spooled. Ultimately, this can influence how and when genes get made into proteins.

Specifically, IDH mutations lead to the formation of a molecule called 2-hydroxyglutarate (2HG). This molecule, in turn, can block the removal of methyl groups.

In the PNAS paper, the investigators dove deeper into the specific epigenetic changes caused by IDH mutations. “What we found was that they didn’t have much to do with DNA methylation, which is what we previously thought,” Dr. Schvartzman says. “Instead, they were related to methylation on histones.”

This change affects how the DNA strands are wrapped around histones. When they are tightly wrapped, it can prevent certain regions of DNA from being accessible. This can affect which genes get made into proteins.

Expanding the Development of Drugs

There already are drugs that are approved to work in AML caused by IDH mutations. Ivosidenib (Tibsovo®) targets IDH1, and enasidenib (Idhifa®) targets IDH2. Both of these drugs prod cancer cells into differentiating normally. But investigators say that there are many more avenues to be explored for new drugs that work against IDH-mutant cancers.

“I’m very interested in looking not just at tumors that are IDH mutant but more broadly at how these cellular changes affect the ability of those cells to differentiate,” Dr. Schvartzman says. “In addition to the buildup of 2HG, there are other changes in the cell that may prevent methyl groups from being removed from histones. We want to study those as well.

“It’s a little early to talk about how this could be applied to new drugs,” he concludes. “But one thing that’s exciting is the ability to understand more about how cells are wired and how different cellular changes affect levels of methylation. There are many enzymes we can start to explore that could be interesting for new cancer drugs.”

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Scientists Develop a Tool to Watch a Single Gene Being Transcribed in a Living Cell

Source: Memorial Sloan Kettering - On Cancer
Date: 07/05/2020
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A picture is worth a thousand words, or so the saying goes. But it can be quite a challenge to capture a picture of something that’s so tiny it’s on the scale of individual molecules.

The field of structural biology is dedicated to constructing images of very, very small things. But most of the techniques used by structural biologists to take these pictures require that the molecules are frozen in one position. This makes it difficult to watch the dynamic, shifting processes that are essential to life.

For the first time, researchers from the Sloan Kettering Institute have found a way to peer inside living cells and observe gene transcription. This is the process by which DNA is copied into messenger RNA (mRNA), which then specifies how a protein is made.

“Gene transcription is one of the most fundamental processes in all of biology,” says SKI structural biologist Alexandros Pertsinidis, senior author of the study, which was published in Cell. “We know that it’s highly regulated and uses complicated molecular machinery. Being able to watch this process as it happens is an important step forward in understanding what goes on inside cells.”

Following the Recipe

If you think of the genetic code as a universal cookbook, containing all of the instructions needed to make every part of a living organism, you can think of the various cell types as different restaurants, Dr. Pertsinidis explains. “French restaurants follow French recipes to make French dishes, and Italian restaurants follow Italian recipes to make Italian dishes,” he says. “In the same way, brain cells make the proteins that brain cells need to function, and liver cells make the proteins for liver function.”

A family of enzymes called RNA polymerases and a large set of factors that are associated with them regulate the transcription of individual genes and control the characteristics that cells exhibit. “The interplay between RNA polymerases and regulatory factors helps determine which genes are turned on and off in specific cells,” he says. “They also control how cells respond to outside signals, which can influence their activities.”

Until now, the function of RNA polymerases and associated regulatory factors has been studied indirectly, through biochemical reactions: Cells are broken open in a test tube and purified into individual parts. By adding or removing components and measuring the outcomes, scientists have been able to figure out certain molecular activities. How the machine as a whole works inside cells, however, has remained obscure.

“For 50 years, hundreds of researchers all over the world have studied these reactions,” Dr. Pertsinidis says. “But the problem has been that nobody has been able to directly observe how gene transcription happens inside a live cell.”

Zooming In on a Single Gene

In the study, the investigators used a highly specialized optical microscope to look at the activity of one RNA polymerase, called RNA polymerase II, as it interacted with genes and synthesized mRNA. The new method, developed by Dr. Pertsinidis’s lab, is called single-molecule nanoscopy.

To be able to look at the individual parts of cells, researchers label molecules with a fluorescent tag that makes them glow under the microscope. “But a cell is very crowded, and there are many reactions happening at the same time,” Dr. Pertsinidis says. “If you label all the polymerases in a cell, the whole nucleus is just a big glow.

“What’s new about this technology is the ultrasensitive, integrated system that lets us zoom in on a single tagged gene even when the cell nucleus is moving around and the specific chromosomal location is jiggling due to random microscopic motion,” he says. “At the same time, the system suppresses the signals from the other reactions that are happening, casting them into the background. This enables us to extract the signal for only the gene of interest and zoom in on it.”

The organization and dynamics of RNA polymerase II in the nucleus have been a topic of intense study over the past few decades. “Here, we directly observed the activity of this molecule and how it functions in the nucleus of live cells,” Dr. Pertsinidis says. “Being able to see how it interacts with other regulatory factors has unveiled the intricate hierarchies and interdependencies of these various factors. These insights enable us to reach a more detailed and comprehensive picture of transcription in live cells.”

Expanding to Other Cellular Processes

The researchers hope that their tool will be widely used to study complicated reactions inside living cells.

“There are enough details in our paper that other labs will be able to pick up and implement the technology,” Dr. Pertsinidis says. “We also have labs both inside and outside MSK that are interested in collaborating with us on specific projects.”

He adds that although he is focused on understanding gene transcription, the tools his team has developed could be used to study the details of other vital biological processes, such as DNA repair and protein synthesis.

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How Errors in Divvying Up Chromosomes Lead to Defects in Cells

Source: National Institutes of General Medical Sciences - Biomedical Beat Blog
Date: 03/25/2020
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Mitosis is fundamental among all organisms for reproduction, growth, and cell replacement. When a cell divides, it’s vital that the two new daughter cells maintain the same genes as the parent.

In one step of mitosis, chromosomes are segregated into two groups, which will go into the two new daughter cells. But if the chromosomes don’t divide properly, one daughter cell may have too many and the other too few. Having the wrong number of chromosomes, a condition called aneuploidy, can trigger cells to grow out of control.

How chromosome segregation errors disrupt cell division is an important area of research. Although it’s been studied for decades, new aspects are still being uncovered and much remains unknown. NIGMS-funded scientists are studying different aspects of mitosis and chromosome segregation. Understanding the details can provide vital insight into an essential biological process and may also be the key to developing better drugs for cancer and other diseases.

Decoding the Mechanics of Chromosome Segregation

Sophie Dumont, Ph.D.  of the University of California, San Francisco, originally trained in physics, but fell in love with biology during graduate school. Her background in physics continues to inform her work, which focuses largely on the mechanical forces that help cells organize themselves during mitosis.

In particular, she’s studying the mitotic spindle (the structure of microtubule filaments that pulls the chromosome pairs apart) and the kinetochore (the protein structure that attaches the chromosome to the spindle).The mitotic spindle and kinetochores are essential to cell organization during mitosis. Credit: Judith Stoffer and NIGMS.

“We’re trying to figure out how these structures, which are made of many small parts, robustly generate and respond to the force that’s needed to move the chromosomes over large distances in the cell,” she explains.

Dr. Dumont’s lab works with human cells, but she also uses a surprising model—kidney cells from rat kangaroos, a small marsupial. Rat kangaroo cells can be helpful because they have many fewer chromosomes than human cells, which makes it easier to follow a single chromosome’s trajectory during mitosis.

“Over millions of years, evolution has found a lot of clever solutions to the challenges of proper chromosome segregation,” she says. “What’s exciting to me about my research is that not only are we making discoveries that have implications for human health, but we are gaining an understanding of how nature can build a diversity of complex structures with very simple parts.”

Recreating Important Processes in the Lab

Chip Asbury, Ph.D. , a scientist at the University of Washington in Seattle, also studies the mechanics of mitosis and chromosome segregation.

“I think of the mitotic spindle as a kind of machine,” he says. “Its job is to move the chromosomes in the appropriate way, while at the same time detecting any errors that may have occurred.”

Dr. Asbury’s lab conducts experiments with the spindle and related components that have been isolated or reconstructed outside the cell. Because kinetochores are made of hundreds of proteins, isolating individual parts of them allows the lab to do experiments they couldn’t do with whole cells.Two Trypanosoma brucei seen through a microscope. Credit: Jeffrey DeGrasse, Rockefeller University.

One experiment introduces tiny artificial cargoes to the kinetochores, enabling Dr. Asbury to recreate and observe how the microtubules form attachments. In addition, Dr. Asbury has a project focusing on mitosis in trypanosomes, a unicellular organism that causes sleeping sickness. This work is a collaboration with Bungo Akiyoshi, Ph.D. , and it may lead to new drugs for targeting these parasites.

“I’m drawn to studying these kinds of fundamental processes because not only are they fascinating, but all life depends on them,” Dr. Asbury says. “Ultimately, our ability to fight complex diseases is limited by our fundamental lack of understanding about what’s happening inside the cell. Understanding how kinetochores work will eventually enable us to develop smarter drugs.”

Linking Chromosomal Errors with Cancer

Jennifer (Jake) DeLuca, Ph.D. , a researcher at Colorado State University in Fort Collins, is looking at how errors in chromosome segregation can lead to cancer. She aims to determine whether it’s possible to develop cancer drugs that target defective segregation.

“The strength of the attachment of the microtubules at the kinetochore must be very tightly controlled,” she explains. “We know that these attachments are misregulated in cancer cells, so we’re working backwards to find out why that is the case.”A human cell undergoing mitosis with the microtubules shown in green, chromosomes shown in blue, and kinetochores shown in red.

To study the chromosome malfunctions seen in cancer, Dr. DeLuca and her team recreate them in human cell models. They transform the cells with cancer-causing genes and then use a number of imaging techniques to watch mitosis in the cells. This allows them to get at the heart of how different proteins regulate microtubule connections in cancer cells.

One of the long-term goals of this research is to figure out how to target these proteins with drugs. “Although we are a basic research lab, not a cancer lab, our work has opened up a huge toolbox of targets to explore,” Dr. DeLuca concludes. “The integration of basic research with clinical research is critical for developing effective new therapies.”

Dr. Dumont’s research is supported by NIGMS grants DP2GM119177 and R01GM134132. Dr. Asbury’s research is supported by R01GM079373 and P01GM105537. Dr. DeLuca’s research is supported by R35GM130365.