Tests with topical treatment strategy for fighting skin cancer yield positive results

Researchers at the University of São Paulo (USP), in Brazil, are testing a technique in mice that combines low-intensity electric current with a formulation containing nanoencapsulated chemotherapy to treat skin cancer.

Applying a low-intensity unidirectional current is one of the ways to ensure that chemical substances penetrate the skin, pushed into the bloodstream through the electric field using a technique known as iontophoresis.

According to preliminary results of the study, cancer-induced mice which received the formulation combined with iontophoresis presented a significantly greater reduction in the size of the tumor than those that received it through injection.

“One of the challenges involved in this type of topical treatment is ensuring that the drug penetrates the stratum corneum – the outermost layer of the epidermis, composed mainly of dead cells. It is an important tissue barrier against the entry of microorganisms, but it also makes it more difficult for medicines to penetrate,” explained Renata Fonseca Vianna Lopez, who supervises the Thematic Project supported by the São Paulo Research Foundation – FAPESP and is also a at the School of Pharmaceutical Sciences of Ribeirão Preto (FCFRP-USP).

In the case of skin cancer, however, the intent is not that the drug penetrates the tissue to get into the bloodstream, but rather that it becomes concentrated in the area below the stratum corneum that requires treatment. This is the reason why, in the study led by Lopez, she chose to place the chemotherapeutic agent inside nanoparticles.

In vivo tests

Using mice, the researchers induced the formation of a tumor associated with one of the most common types of skin cancer – squamous cell carcinoma – through a subcutaneous injection of human tumor cells that overexpress the epidermal growth factor receptor (EGFR). Lopez explained that the presence of this protein causes the tumor to become more aggressive.

The treatment was conducted using a formulation containing chemotherapy agent 5-fluorouracil encapsulated in a nanoparticle (liposome) that functions as an anti-EGFR antibody. The malignant cells are able to capture a larger quantity of the drug encapsulated in these liposomes.

One group of rodents received the tumor formulation through subcutaneous injections and another group received it through topical application combined with iontophoresis. Lopez compared both methods and thus assessed:

“In addition to reducing the size of the tumor, the topical treatment left the tumor less aggressive. We believe that this method combined with iontophoresis allows the drug to be dispersed over the entire area of the tumor, whereas the subcutaneous application causes it to be concentrated in a single location,” Lopez noted.

Versatile technique

In another study, Lopez’ group used a stiffer type of polymeric nanoparticle, one containing the anti-inflammatory dexamethasone associated with iontophoresis for the treatment of uveitis – an inflammation of the eye tissue. The results, published in 2015 in the Journal of Controlled Release, is the outcome of the doctoral thesis of Joel Gonçalves Souza, winner of the 2015 Capes Thesis Award in Pharmacy.

“When we apply the medicine directly to the eye, it is quickly eliminated through the defense mechanisms, such as tears. Increased penetration and better results are obtained by using the application method combined with iontophoresis,” Lopez said.

Currently, in dissertation research by Camila Lemos, the group plans to test a method that uses iontophoresis in the treatment of chronic wounds such as those that develop in patients with diabetes.

“In this case, we are not dealing with the stratum corneum barrier. We use iontophoresis to assess its influence on release of the substance of interest in a formulation, and to investigate its effect on the growth of microorganisms,” Lopez explained.

The strategy consists of placing a peptide having anti-inflammatory properties on a film made of fibers extracted from the cocoon of a silkworm (fibroin). The film is placed on the wound as a dressing, to which an electric current is then applied.

“When we placed the peptide directly on the wound, it degraded very quickly. When placed on the film, however, release occurs in a slower and more sustained way. Iontophoresis allows a larger amount of the peptide to be released from the film at the start of treatment to accelerate healing,” the researcher explained.

Lopez went on to say that preliminary results suggest that iontophoresis also stops the proliferation of some types of microorganisms (particularly gram-positive bacteria) that could aggravate wounds.

Stabilizing TREM2 — a potential strategy to combat Alzheimer’s disease

A gene called triggering receptor expressed on myeloid cells 2, or TREM2, has been associated with numerous neurodegenerative diseases, such as Alzheimer’s disease, Frontotemporal lobar degeneration, Parkinson’s disease, and Nasu-Hakola disease. Recently, a rare mutation in the gene has been shown to increase the risk for developing Alzheimer’s disease.

Independently from each other, two research groups have now revealed the molecular mechanism behind this mutation. Their research, published today in EMBO Molecular Medicine, sheds light on the role of TREM2 in normal brain function and suggests a new therapeutic target in Alzheimer’s disease treatment.

Alzheimer’s disease, just like other neurodegenerative diseases, is characterized by the accumulation of specific protein aggregates in the brain. Specialized brain immune cells called microglia strive to counter this process by engulfing the toxic buildup. But as the brain ages, microglia eventually lose out and fail to rid all the damaging material.

TREM2 is active on microglia and enables them to carry out their protective function. The protein spans the microglia cell membrane and uses its external region to detect dying cells or lipids associated with toxic protein aggregates. Subsequently, TREM2 is cut in two. The external part is shed from the protein and released, while the remaining part still present in the cell membrane is degraded. To better understand TREM2 function, the two research groups took a closer look at its cleavage. They were led by Christian Haass at the German Center for Neurodegenerative Diseases at the Ludwig-Maximilians-University in Munich, Germany, and Damian Crowther of AstraZeneca’s IMED Neuroscience group in Cambridge, UK together with colleagues at the Tanz Centre for Research in Neurodegenerative Diseases, University of Toronto and the Cambridge Institute for Medical Research, University of Cambridge, UK.

Using different technological approaches, both groups first determined the exact site of protein shedding and found it to be at amino acid 157. Amino acid 157 was no unknown. Only recently, researchers from China had uncovered that a mutation at this exact position, referred to as p.H157Y, increased the risk of Alzheimer’s disease. Together, these observations indicate that protein cleavage is perturbed in the p.H157 mutant and that this alteration promotes disease development.

As a next step, Haass and Crowther’s groups investigated the biochemical properties of the p.H157Y mutant protein more closely. They found that the mutant was cleaved more rapidly than a healthy version of the protein. “Our results provide a detailed molecular mechanism for how this rare mutation alters the function of TREM2 and hence facilitates the progression of Alzheimer’s disease,” said Crowther.

While most TREM2 mutations affect protein production, the mechanism behind p.H157Y is somewhat different. The p.H157Y mutation allows the protein to be correctly manufactured and transported to the microglia cell surface, but then it is cleaved too quickly. “The end result is the same. In both cases, there is too little full-length TREM protein on microglia,” said Haass. “This suggests that stabilizing TREM2, by making it less susceptible to cleavage, may be a viable therapeutic strategy.”

Repairing damaged hearts with self-healing heart cells

New research has discovered a potential means to trigger damaged heart cells to self-heal. The discovery could lead to groundbreaking forms of treatment for heart diseases. For the first time, researchers have identified a long non-coding ribonucleic acid (ncRNA) that regulates genes controlling the ability of heart cells to undergo repair or regeneration. This novel RNA, which researchers have named “Singheart”, may be targeted for treating heart failure in the future. The discovery was made jointly by A*STAR’s Genome Institute of Singapore (GIS) and the National University Health System (NUHS), and is now published in Nature Communications.

Unlike most other cells in the human body, heart cells do not have the ability to self-repair or regenerate effectively, making heart attack and heart failure severe and debilitating. Cardiovascular disease (CVD) is the leading cause of death worldwide, with an estimated 17.7 million people dying from CVD in 2015 (1). CVD also accounted for close to 30% of all deaths in Singapore in 2015 (2).

In this project, the researchers used single cell technology to explore gene expression patterns in healthy and diseased hearts. The team discovered that a unique subpopulation of heart cells in diseased hearts activate gene programmes related to heart cell division, uncovering the gene expression heterogeneity of diseased heart cells for the first time. In addition, they also found the “brakes” that prevent heart cells from dividing and thus self-healing. Targeting these “brakes” could help trigger the repair and regeneration of heart cells.

“There has always been a suspicion that the heart holds the key to its own healing, regenerative and repair capability. But that ability seems to become blocked as soon as the heart is past its developmental stage. Our findings point to this potential block that when lifted, may allow the heart to heal itself,” explained A/Prof Roger Foo, the study’s lead author, who is Principal Investigator at both GIS and NUHS’ Cardiovascular Research Institute (CVRI) and Senior Consultant at the National University Heart Centre, Singapore (NUHCS).

“In contrast to a skin wound where the scab falls off and new skin grows over, the heart lacks such a capability to self-heal, and suffers a permanent scar instead. If the heart can be motivated to heal like the skin, consequences of a heart attack would be banished forever,” added A/Prof Foo.

The study was driven by first author and former Senior Research Fellow at the GIS, Dr Kelvin See, who is currently a Postdoctoral Researcher and Mack Technology Fellow at University of Pennsylvania.

“This new research is a significant step towards unlocking the heart’s full regenerative potential, and may eventually translate to more effective treatment for heart diseases. Heart disease is the top disease burden in Singapore and strong funding remains urgently needed to enable similar groundbreaking discoveries,” said Prof Mark Richards, Director of CVRI.

Executive Director of GIS, Prof Ng Huck Hui added, “This cross-institutional research effort serves as a strong foundation for future heart studies. More importantly, uncovering barriers that stand in the way of heart cells’ self-healing process brings us another step closer to finding a cure for one of the world’s biggest killers.”

UC research examines lung cell turnover as risk factor & target for treatment of influenza pneumonia

Influenza is a recurring global health threat that, according to the World Health Organization, is responsible for as many as 500,000 deaths every year, most due to influenza pneumonia, or viral pneumonia. Infection with influenza most typically results in lung manifestations limited to dry cough and fever, and understanding how the transition to pneumonia occurs could shed light on interventions that reduce mortality. Research led by University of Cincinnati (UC) scientists takes a different approach to investigating how influenza spreads through the lungs by focusing on how resistant or susceptible cells lining the airway are to viral infection.

The work published today in the Proceedings of the National Academy of Sciences (PNAS) shows how stimuli that induce cell division in the lung promote spread of influenza from the airway to the gas exchanging units of the lung, known as the alveoli. The UC study also demonstrates that interventions that prevent alveolar cells from dividing reduce influenza mortality in animal models, suggesting a potential prophylactic and/or therapeutic strategy for influenza pneumonia.

“Almost all research into susceptibility or resistance to influenza focuses on host immune responses,” says Nikolaos Nikolaidis, PhD, research scientist in the Division of Pulmonary, Critical Care and Sleep Medicine in the Department of Internal Medicine at the UC College of Medicine and lead author on the paper. “Our approach was to examine factors that influence the vulnerability of alveolar cells to influenza infection, separate from how the immune system is dealing with the virus.”

“Less than 1 percent of alveolar cells are actively dividing at any given time in the healthy lung, rendering it naturally resistant to influenza infection,” says Frank McCormack, MD, Gordon and Helen Hughes Taylor Professor of Internal Medicine and director of the Division of Pulmonary, Critical Care and Sleep Medicine and senior author on the paper. “Recovery from lung injury due to supplemental oxygen therapy, cigarette smoke or scarring lung diseases is associated with expression of growth factors that result in multiplication of lung cells. Our work demonstrated that these mitogenically stimulated cells are rich targets for influenza infection while they are dividing.”

The researchers found that when sirolimus, which is FDA-approved for use as an anti-growth agent for the rare lung disease, lymphangioleiomyomatosis (LAM), was given to influenza-infected animal models, it prevented alveolar cells from dividing, and as a result, protected the mice from viral pneumonia and death.

“Although sirolimus also has off target immunosuppressive properties that could potentially pose added risks of side effects in virus-infected patients, trials of inhaled sirolimus could lead to approaches that do not entail systemic exposure,” says McCormack.

The McCormack lab expressed optimism that this observation has the potential to ultimately inform understanding of other unexplained risk factors for influenza, including very young age and pregnancy, and perhaps even to change medical management, such as more judicious use of supplemental oxygen in patients admitted with suspected viral pneumonia. Further, the team has hopes that the research could lead to a paradigm shift in the approach to therapy.

Nikolaidis says the next step in this research is to further explore why the multiplying alveolar epithelial cell is a better target for influenza. “Is it because the virus gets into the dividing cell more easily, because multiplying stimuli expand the pool of cellular machinery used by the virus to replicate, or because proliferation is associated with a reduction in innate cellular defenses? We are anxious to explore these and other potential mechanisms of viral susceptibility,” he adds.

Altered virus may expand patient recruitment in human gene therapy trials

For many patients, participating in gene therapy clinical trials isn’t an option because their immune system recognizes and fights the helpful virus used for treatment. Now, University of Florida Health and University of North Carolina researchers have found a solution that may allow it to evade the body’s normal immune response.

The discovery, published May 29 in the Proceedings of the National Academy of Sciences, is a crucial step in averting the immune response that prevents many people from taking part in clinical trials for various disorders, said Mavis Agbandje-McKenna, Ph.D., a professor in the University of Florida College of Medicine department of biochemistry and molecular biology and director of the Center for Structural Biology.

During gene therapy, engineered viruses are used to deliver new genes to a patient’s cells. While the recombinant adeno-associated virus, or AAV, is effective at delivering its genetic cargo, prior natural exposure to AAV results in antibodies in some people. As many as 70 percent of patients have pre-existing immunity that makes them ineligible for gene therapy clinical trials, Agbandje-McKenna said.

The findings provide a road map for designing virus strains that can evade neutralizing antibodies, said Aravind Asokan, Ph.D., an associate professor in the department of genetics at the University of North Carolina, who led the study. At UF Health, the structural “footprints” where pre-existing antibodies interact with the virus were identified using cryo-electron microscope resources provided by the UF College of Medicine and the UF Office of Research’s Division of Sponsored Programs. The UNC researchers then evolved new viral protein shells. Using serum from mice, rhesus monkeys and humans, the researchers showed that the redesigned virus can slip past the immune system.

“This is the blueprint for producing AAV strains that could help more patients become eligible for human gene therapy. Now we know how to do it,” Agbandje-McKenna said.

While the findings prove that one variation of AAV can be evolved, further study in preclinical models is needed before the approach can be tested in humans. Next, the immune profile of one particularly promising virus variant will need to be evaluated in a larger number of human serum samples, and dose-finding studies are needed in certain animal models. Researchers may also need to study whether the same virus-manipulating technique can be used in a broader range of gene therapy viruses, Agbandje-McKenna said.

Although human gene therapy remains an emerging field and has yet to reach patients on a wide scale, researchers elsewhere have used AAV therapy to successfully treat hemophilia, a blood-clotting disorder, in a small trial. It has also been or is now being studied as a way to treat hereditary blindness, certain immune deficiencies, neurological and metabolic disorders, and certain cancers.

The latest findings are the result of more than 10 years of studying the interactions between viruses and antibodies and a long-standing collaboration with Asokan, who heads the synthetic virology group at the UNC Gene Therapy Center, according to Agbandje-McKenna.

URI Scientist: Rare Childhood Disease Linked to Major Cancer Gene

Researcher discovers novel connection between Fanconi anemia, PTEN

A team of researchers led by a University of Rhode Island scientist has discovered an important molecular link between a rare  childhood genetic disease, Fanconi anemia, and a major cancer gene called PTEN. The discovery improves the understanding of the molecular basis of Fanconi anemia and could lead to improved treatment outcomes for some cancer patients.

According to Niall Howlett, URI associate professor of cell and molecular biology and Rhode Island’s leading expert on Fanconi anemia, the disease is characterized by birth defects, bone marrow failure and increased cancer risk. He said the genes that play a role in the development of the disease are also important in the development of hereditary breast and ovarian cancer.

Howlett’s new study now establishes a molecular link between Fanconi anemia and a gene strongly associated with uterine, prostate and brain cancer. This research was published this month in the journal Scientific Reports, with URI graduate student Elizabeth Vuono as lead author.

About 1 in 150,000 children in the United States is born with Fanconi anemia.

“People often ask why we study such a rare disease,” said Howlett, who has been studying Fanconi anemia for nearly 20 years. “First and foremost, there is no cure or effective treatments for it. So a greater understanding of the molecular basis of Fanconi anemia is critical to address this need.”

In addition, Howlett said there are countless examples of how the study of Fanconi anemia has greatly benefited the general population. The first umbilical cord blood transplant, for example, was performed with a Fanconi anemia patient. Bone marrow transplants have become much safer and more effective because of studies with Fanconi anemia patients. And new breast and ovarian cancer genes have been discovered as a result of studies on the molecular biology of Fanconi anemia.

Howlett’s current research is another example of the broader impact of Fanconi anemia studies.

The URI researcher speculated about the existence of a biochemical link between Fanconi anemia and PTEN. Mutations in PTEN occur frequently in uterine, prostate and brain cancer.

“The PTEN gene codes for a phosphatase – an enzyme that removes phosphate groups from proteins,” explained Howlett. “Many Fanconi anemia proteins have phosphate groups attached to them when they become activated. However, how these phosphate groups are removed is poorly understood.”

Howlett said that cells from Fanconi anemia patients are characteristically sensitive to a class of drugs widely used in cancer chemotherapy called DNA crosslinking agents.

“So we performed an experiment to determine if Fanconi anemia and PTEN were biochemically linked,” he said. “By testing if cells with mutations in the PTEN gene were also sensitive to DNA crosslinking agents, we discovered that Fanconi anemia patient cells and PTEN-deficient cells were practically indistinguishable in terms of sensitivity to these drugs. This strongly suggested that the Fanconi anemia proteins and PTEN might work together to repair the DNA damage caused by DNA crosslinking agents.”

By using epistasis analysis, a genetic method that determines if genes work together, Howlett and his research group found that the Fanconi anemia proteins and PTEN do indeed function together in this repair pathway.

“Before this work, Fanconi anemia and PTEN weren’t even on the same radar,” said Howlett. “This is really important to understanding how this disease arises and what its molecular underpinnings are. The more we can find out about its molecular basis, the more likely we are to come up with strategies to treat the disease.”

Howlett’s research is equally important to cancer patients who do not have Fanconi anemia. He said that since his study found that cells missing PTEN are highly sensitive to DNA crosslinking agents, it should be possible to predict whether a particular cancer patient will respond to this class of chemotherapy drug by conducting a simple DNA test.

“We can now predict that if a patient has cancer associated with mutations in PTEN, then it is likely that the cancer will be sensitive to DNA crosslinking agents,” he said. “This could lead to improved outcomes for patients with certain types of PTEN mutations.”

Heart disease, leukemia linked to dysfunction in nucleus

We put things into a container to keep them organized and safe. In cells, the nucleus has a similar role: keeping DNA protected and intact within an enveloping membrane. But a new study by Salk Institute scientists, detailed in the November 2 issue of Genes & Development, reveals that this cellular container acts on its contents to influence gene expression.

“Our research shows that, far from being a passive enclosure as many biologists have thought, the nuclear membrane is an active regulatory structure,” says Salk Professor Martin Hetzer, who is also holder of the Jesse and Caryl Philips Foundation chair. “Not only does it interact with portions of the genome to drive gene expression, but it can also contribute to disease processes when components are faulty.”

Using a suite of molecular biology technologies, the Salk team discovered that two proteins, which sit in the nuclear envelope, together with the membrane-spanning complexes they form, actively associate with stretches of DNA to trigger expression of key genes. Better understanding these higher-level functions could provide insight into diseases that appear to be related to dysfunctional nuclear membrane components, such as leukemia, heart disease and aging disorders.

Historically, the nuclear membrane’s main purpose was thought to be keeping the contents of the nucleus physically separated from the rest of the cell. Complexes of at least thirty different proteins, called nucleoporins, form gateways (pores) in the membrane, controlling what goes in or out. But as the Hetzer lab’s work on nucleoporins shows, these nuclear pore complexes (NPCs), beyond being mere gateways into the nucleus, have surprising regulatory effects on the DNA inside.

“Discovering that key regulatory regions of the genome are actually positioned at nuclear pores was very unexpected,” says Arkaitz Ibarra, a Salk staff scientist and first author of the paper. “And even more importantly, nuclear pore proteins are critical for the function of those genomic sites.”

Curious about all the regions of DNA with which nucleoporins potentially interact, the team turned to a human bone cancer cell line. The scientists used a molecular biology technique called DamID to pinpoint where two nucleoporins, Nup153 and Nup93, came into contact with the genome. Then they used several other sequencing techniques to understand which genes were being affected in those regions, and how.

The Salk team discovered that Nup153 and Nup93 interacted with stretches of the genome called super-enhancers, which are known to help determine cell identity. Since every cell in our body has the same DNA, what makes a muscle cell different from a liver cell or a nerve cell is which particular genes are turned on, or expressed, within that cell. In the Salk study, the presence of Nup153 and Nup93 was found to regulate expression of super-enhancer driven genes and experiments that silenced either protein resulted in abnormal gene expression from these regions. Further experiments in a lung cancer cell line validated the bone cancer line results: Nucleoporins in the NPC were found to interact with multiple super-enhancer regions to drive gene expression, while experiments that altered the NPC proteins made related gene expression faulty, even though the proteins still performed their primary role as gatekeepers in the cell membrane.

“It was incredible to find that we could perturb the proteins without affecting their gateway role, but still have nearby gene expression go awry,” says Ibarra.

The results bolster other work indicating that problems with the nuclear membrane play a role in heart disease, leukemia and progeria, a rare premature aging syndrome.

“People have thought the nuclear membrane is just a protective barrier, which is maybe the reason why it evolved in the first place. But there are many more regulatory levels that we don’t understand. And it’s such an important area because so far, every membrane protein that has been studied and found to be mutated or mis-localized, seems to cause a human disease,” says Hetzer.

Antibody Breaks Leukemia’s Hold, Providing New Therapeutic Approach

Acute myeloid leukemia (AML) is an aggressive cancer known for drug resistance and relapse. In an effort to uncover new treatment strategies, researchers at University of California San Diego School of Medicine and Moores Cancer Center discovered that a cell surface molecule known as CD98 promotes AML. The study, published October 27 by Cancer Cell, also shows that inhibiting CD98 with the therapeutic antibody IGN523 blocks AML growth in patient-derived cells and mouse models.

“To improve therapeutic strategies for this disease, we need to look not just at the cancer cells themselves, but also at their interactions with surrounding cells, tissues, molecules and blood vessels in the body,” said co-senior author Tannishtha Reya, PhD, professor of pharmacology at UC San Diego School of Medicine and Moores Cancer Center. “In this study, we identified CD98 as a critical molecule driving AML growth. We showed that blocking CD98 can effectively reduce leukemia burden and improve survival by preventing cancer cells from receiving support from the surrounding environment.”

Reya led the study together with Mark Ginsberg, MD, professor of medicine at UC San Diego School of Medicine and Moores Cancer Center. Co-author Edward van der Horst, PhD, senior director at Igenica Biotherapeutics Inc., provided the anti-CD98 antibody IGN523.

AML is a type of cancer in which the bone marrow makes abnormal white blood cells, red blood cells or platelets. Reya’s team and others have previously shown that leukemia cells interact with their surroundings in the body via molecules on their cell surfaces, and that these interactions can help the cancer cells divide, replicate and metastasize.

CD98 is a molecule found on the surface of cells, where it controls how cells stick to one another. CD98 is known to play a role in the proliferation and activation of certain immune cells. CD98 levels are also known to be elevated in some solid tumors, and linked to poor prognosis.

To determine CD98’s role in AML, in this latest study Reya’s team engineered mouse models that lack the molecule. They found that the loss of CD98 blocked AML growth and improved survival. CD98 loss largely spared normal blood cells, which the researchers said indicates a potential therapeutic window. Further experiments revealed that leukemia cells lacking CD98 had fewer stable interactions with the lining of blood vessels — interactions that were needed to fuel AML growth.

Next, the researchers wanted to see what would happen if they blocked CD98 in AML with a deliverable inhibitor. In 2015, Igenica Biotherapeutics Inc. tested IGN523, a humanized antibody that specifically binds and inhibits CD98, in a phase 1 clinical trial at Moores Cancer Center and elsewhere. The trial’s goal was to determine a safe dose for IGN523 administration in AML patients. In this study, Reya and team tested IGN523 in their own AML models.

The researchers found that IGN523 blocks CD98’s AML-promoting activity in both mouse models of AML and human cells in the laboratory. They also transplanted human patient-derived AML cells into mice and treated the recipients soon after with either IGN523, the anti-CD98 antibody, or with a control antibody. Anti-CD98-treatment effectively eliminated AML cells. In contrast, AML in control mice expanded more than 100-fold.

“This study suggests that human AML can’t get established without CD98, and that blocking the molecule with anti-CD98 antibodies could be beneficial for the treatment of AML in both adults and children,” Reya said.

Moving forward, Reya and team are working to further define whether CD98 could be targeted to treat pediatric AML.

“Many of the models we used in this work were based on mutations found in childhood AML,” she said. “While many childhood cancers have become very treatable, childhood AML continues to have a high rate of relapse and death. We plan to work with pediatric oncologists to test if anti-CD98 agents can be effective against pediatric AML, and whether it can improve responses to current treatments. I think this is particularly important to pursue since the anti-CD98 antibody has already been through phase I trials, and could be more easily positioned to test in drug-resistant pediatric AML.”

The American Cancer Society estimates that there will be about 19,950 new cases of AML and about 10,430 deaths from the disease in the United States in 2016, mostly adults. Approximately 500 children are diagnosed with AML in the U.S. each year, and it’s the most common second cancer among children treated for other cancers, according to St. Jude Children’s Research Hospital.

Cytomegalovirus Infection Relies On Human RNA-Binding Protein

Viruses hijack the molecular machinery in human cells to survive and replicate, often damaging those host cells in the process. Researchers at the University of California San Diego School of Medicine discovered that, for cytomegalovirus (CMV), this process relies on a human protein called CPEB1. The study, published October 24 inNature Structural and Molecular Biology, provides a potential new target for the development of CMV therapies.

“We found that CPEB1, one of a family of hundreds of RNA-binding proteins in the human genome, is important for establishing productive cytomegalovirus infections,” said senior author Gene Yeo, PhD, professor of cellular and molecular medicine at UC San Diego School of Medicine.

CMV is a virus that infects more than half of all adults by age 40, and stays for life. Most infected people are not aware that they have CMV because it rarely causes symptoms. However, CMV can cause serious health problems for people with compromised immune systems, or babies infected with the virus before birth. There are currently no treatments or vaccines for CMV.

In human cells, RNA is the genetic material that carries instructions from the DNA in a cell’s nucleus out to the cytoplasm, where molecular machinery uses those instructions to build proteins. CPEB1 is a human protein that normally binds RNAs that are destined to be translated into proteins.

Yeo’s team discovered that CPEB1 levels increase dramatically in human cells infected by CMV. Using genomics technologies, the researchers also found that increased CPEB1 levels in CMV-infected cells leads to abnormal processing of RNAs encoding thousands of human genes. In addition, they were surprised to find that CPEB1 was necessary for proper processing of viral RNAs. Without the host CPEB1 protein, viral RNA did not mature properly and the virus was weakened.

CMV-infected human cells undergo abnormal changes and produce more virus, which ultimately infects other cells. In collaboration with Deborah Spector, PhD, Distinguished Professor at UC San Diego School of Medicine and Skaggs School of Pharmacy and Pharmaceutical Sciences, the team went on to show that suppressing CPEB1 levels during CMV infection reversed these harmful cellular changes and reduced viral production tenfold.

“CPEB1 was previously shown to play a role in neuronal development and function, but this involvement in active viral infections is unexpected,” said first author Ranjan Batra, PhD, a postdoctoral fellow in Yeo’s lab. “This discovery has important implications for many viral infections.”

Yeo said the next steps are to determine the therapeutic value of inhibiting CPEB1 in CMV infections and identify other RNA-binding proteins that may be important in other viral infections.

Potential therapeutic target for Huntington’s disease

There is new hope in the fight against Huntington’s disease. Scientists at the Gladstone Institutes discovered that changing a specific part of the huntingtin protein prevented the loss of critical brain cells and protected against behavioral symptoms in a mouse model of the disease.

Huntington’s disease causes jerky, uncoordinated movements and a loss of control of motor function. It also results in deficits in learning and memory, as well as personality changes, such as dementia, depression, and aggression. Huntington’s is ultimately fatal, and there are no treatments to stop or slow its progression.

The disease is linked to a mutation in the Huntingtin gene, which causes a protein of the same name to fold up incorrectly like misshapen origami. Neurons cannot get rid of the misfolded protein, so it builds up in the brain, wreaking havoc in the cells.

In the new study, published in the Journal of Clinical Investigation, scientists in the laboratory of Steve Finkbeiner, MD, PhD, showed that modifying the huntingtin protein through a process called phosphorylation can actually make the protein less toxic and allows cells to eliminate it more easily. In fact, phosphorylating a specific spot on the protein called S421 protected a mouse model of Huntington’s from developing symptoms of the disease.

“I was shocked at the profound effect phosphorylation had on the Huntington’s model mice,” said first author Ian Kratter, MD, PhD, a former graduate student at Gladstone and the University of California, San Francisco (UCSF). “They showed few signs of the motor dysfunction, depression, or anxiety that are characteristic of the disease. In most of our tests, they were virtually indistinguishable from healthy mice.”

The mice were also protected against neuron death, particularly in the striatum, the movement and reward center of the brain that is first affected in Huntington’s disease. The scientists think that phosphorylation enables neurons to remove more of the harmful protein so it does not accumulate and damage the cell.

“Phosphorylation helps control how proteins fold and the systems in cells that clear proteins,” explained Finkbeiner, who is a senior investigator at Gladstone. “This is exciting, because a lot of the work we’ve done points to these protein removal pathways as being important not only for Huntington’s disease, but also for other neurodegenerative disorders. Understanding how phosphorylation links to these pathways could help treat several different brain diseases.”

The researchers are now exploring ways to mimic the effects of phosphorylation with a drug.

Some genetic causes of ALS may need an epigenetic trigger to activate the disease

A new research report appearing online in The FASEB Journal shows why, for some people, having a genetic predisposition to amyotrophic lateral sclerosis (ALS) may not be enough to actually guarantee having the disease. In short, researchers examined identical twins–one afflicted with familial ALS and one not–and found that environmental factors were likely necessary to alter the expression of some immune genes (epigenetic changes) before the disease could take hold. This discovery may pave the way toward developing preventive strategies for those who are at risk for ALS.

“Inflammation is a cause of damage in the central nervous system in other neurodegenerative diseases, including mild cognitive impairment,” said Milan Fiala, M.D., a senior researcher involved in the work from the David Geffen School of Medicine at the University of California, Los Angeles. “Further studies will show how inflammation can be regulated in different diseases.”

To make their discovery, Fiala and colleagues studied a set of identical twins, one with an inherited form of ALS and the other healthy. Both twins had mutations in their genomes related to ALS, but the ALS twin had certain epigenetic changes in the genome suspected to be related to the disease. In particular, the team looked at the production of IL-6 because in previous observations they saw that tocilizumab (an IL-6 receptor antibody) seemed to benefit another ALS patient. In the study they also demonstrated the presence of neurotoxic cytokines in the afflicted twin. Further research is necessary to understand how these epigenetic changes relate to actual immune function, and ultimately disease progression.

“We are now beginning to see a number of new clues on both the familial and sporadic ALS pathogenesis landscapes, some quite unanticipated as in this important study,” said Thoru Pederson, Ph.D., Editor-in-Chief of The FASEB Journal. “Considering the dearth of effective treatments for this devastating disease, all new findings such as these are welcome indeed.”

Body’s Own Gene Editing System Generates Leukemia Stem Cells

Cancer stem cells are like zombies — even after a tumor is destroyed, they can keep coming back. These cells have an unlimited capacity to regenerate themselves, making more cancer stem cells and more tumors. Researchers at University of California San Diego School of Medicine have now unraveled how pre-leukemic white blood cell precursors become leukemia stem cells. The study, published June 9 in Cell Stem Cell, used human cells to define the RNA editing enzyme ADAR1’s role in leukemia, and find a way to stop it.

While DNA is like the architect’s blueprint for a cell, RNA is the like the engineer’s interpretation of the blueprint. That RNA version is frequently flawed in cancer. While many studies have uncovered pivotal DNA mutations in cancer, few have addressed the roles of RNA and mechanisms that regulate RNA.

“In this study, we showed that cancer stem cells co-opt a RNA editing system to clone themselves. What’s more, we found a method to dial it down,” said senior author Catriona Jamieson, MD, PhD, associate professor of medicine and chief of the Division of Regenerative Medicine at UC San Diego School of Medicine.

The enzyme at the center of this study, ADAR1, can edit the sequence of microRNAs, small pieces of genetic material. By swapping out just one microRNA building block for another, ADAR1 alters the carefully orchestrated system cells use to control which genes are turned on or off at which times.

ADAR1 is known to promote cancer progression and resistance to therapy. In this study, Jamieson’s team used human blast crisis chronic myeloid leukemia cells in the lab, and mice transplanted with these cells, to determine ADAR1’s role in governing leukemia stem cells.

The researchers uncovered a series of molecular events. First, white blood cells with a leukemia-promoting gene mutation become more sensitive to signs of inflammation. That inflammatory response activates ADAR1. Then, hyper-ADAR1 editing slows down the microRNAs known as let-7. Ultimately, this activity increases cellular regeneration, or self-renewal, turning white blood cell precursors into leukemia stem cells. Leukemia stem cells promote an aggressive, therapy-resistant form of the disease called blast crisis.

“This is the first mechanistic link between pro-cancer inflammatory signals and RNA editing-driven reprogramming of precursor cells into leukemia stem cells,” said Jamieson, who is also deputy director of the Sanford Stem Cell Clinical Center at UC San Diego Health, director of the CIRM Alpha Stem Cell Clinic at UC San Diego School of Medicine and director of stem cell research at UC San Diego Moores Cancer Center.

After learning how the ADAR1 system works, Jamieson’s team looked for a way to stop it. By inhibiting sensitivity to inflammation or inhibiting ADAR1 with a small molecule tool compound, the researchers were able to counter ADAR1’s effect on leukemia stem cell self-renewal and restore let-7. Self-renewal of blast crisis chronic myeloid leukemia cells was reduced by approximately 40 percent when treated with the small molecule, called 8-Aza, as compared to untreated cells.

“Based on this research, we believe that detecting ADAR1 activity will be important for predicting cancer progression. In addition, inhibiting this enzyme represents a unique therapeutic vulnerability in cancer stem cells with active inflammatory signaling that may respond to pharmacologic inhibitors of inflammation sensitivity or selective ADAR1 inhibitors that are currently being developed,” Jamieson said.

Botox’s Sweet Tooth Underlies Its Key Neuron-Targeting Mechanism UCI

The Botox toxin has a sweet tooth, and it’s this craving for sugars – glycans, to be exact – that underlies its extreme ability target neuron cells in the body … while giving researchers an approach to neutralize it.

A study co-led by Rongsheng Jin, professor of physiology & biophysics at the University of California, Irvine; Min Dong with Boston Children’s Hospital-Harvard Medical School; and Andreas Rummel with the Hannover Medical School in Germany, reveals an important general mechanism by which the pathogen is attracted to, adapts to and takes advantage of glycan modifications in surface receptors to invade motor neurons. Glycans are chains of sugars synthesized by cells for their development, growth, functioning or survival. Results appear June 13 in Nature Structural and Molecular Biology.

“Our findings reveal a new paradigm of the everlasting host-pathogen arms race, where a pathogen develops a smart strategy to achieve highly specific binding to a host receptor while also tolerating genetic changes on the receptor,” Jin said. “And to some extent, this mechanism by which the toxin attacks human is similar to the one that is utilized by some important broad-neutralizing human antibodies to fight viruses, such as dengue viruses and HIV.”

Botulinum neurotoxin A (BoNT/A), commonly known as the Botox toxin, is widely used in a weakened form for treating various medical conditions as well as for cosmetics. The clinical product Botox contains extremely low doses of the toxin and is safe to use. But at higher doses, it can be lethal, and it’s also classified as a potential bioterrorism agent.

Its potency and therapeutic effects rely on its extraordinary ability to target motor neurons and subsequently block release neurotransmitters that control the movement of muscle fiber, which can result in paralysis. This specific targeting is achieved by highly selective interactions between the toxin and its receptors – host proteins that bind to the toxin in a lock and key manner.

The intricate detail of how the Botox toxin recognizes its receptors also reveals novel ways to neutralizing these deadly toxins. “With this new structural information,” Rummel said, “we were able to pinpoint key amino acids in the toxin that are required for binding to sugars, and we found that even mutating a single amino acid is sufficient to abolish the toxicity by more than a million fold.”

Remarkably, the newly mapped glycan-binding site turns out to be also the site recognized by a potent human neutralizing antibody called CR2/CR1, which is currently in clinical trial for the treatment and prevention of BoNT/A poisoning. “This antibody fights the toxin exactly by preventing it from binding to sugars on the receptor. Our study thus provides a new strategy for anti-toxin inhibitor design,” Dong said.

Guorui Yao and Kwok-ho Lam at UCI, Sicai Zhang at Harvard, Stefan Mahrhold at the Hannover Medical School, Daniel Stern at Berlin’s Center for Biological Threats & Special Pathogens, Kay Perry at the Argonne National Laboratory in Illinois, and Karine Bagramyan and Markus Kalkum the Beckman Research Institute at the City of Hope in Duarte, Calif., contributed to the study, which was primarily supported by the National Institutes of Health.

No symptoms, but could there be cancer? Our chemosensor will detect it!

Many cancers could be successfully treated if the patient consulted the doctor sufficiently early. But how can a developing cancer be detected if it doesn’t give rise to any symptoms? In the near future, suitably early diagnosis could be provided by simple and cheap chemical sensors – thanks to special recognizing polymer films developed at the Institute of Physical Chemistry of the Polish Academy of Sciences in Warsaw.

These days, cancer is no longer a death sentence for the patient. However, the best chances of recovery are when the correct treatment is undertaken at an early stage of the disease. This is where the trouble starts, because many tumours develop over a long period without any symptoms. One solution to this problem could be diagnostic tests available to everyone that could be performed by people themselves and on a relatively regular basis. A step bringing us closer to this sort of personalized medical diagnosis and cancer prophylaxis is the chemical sensor devised and fabricated by Prof. Wlodzimierz Kutner’s group from the Institute of Physical Chemistry of the Polish Academy of Sciences (IPC PAS) in Warsaw using a grant from the National Science Centre, in collaboration with the team of Prof. Francis D’Souza of the University of North Texas in Denton TX, USA.

The most important element of the chemosensor devised at the IPC PAS is a thin film of the polymer that detects molecules of neopterin. Neopterin – in chemical terminology known as 2-amino-6-(1,2,3-trihydroxypropyl)-1H-pteridin-4-one) – is an aromatic compound present in human body fluids, such as serum, urine, and cerebrospinal fluid. Produced by the immune system, it is regarded as a universal marker in medical diagnosis. The concentration of this biomarker rises significantly particularly in the case of certain neoplastic diseases, e.g., malignant lymphoma, although elevated levels of neopterin are also seen in some viral and bacterial infections, as well as in diseases of parasitic aetiology. In turn, in transplant patients, increased levels of neopterin signal probable rejection.

“How can we detect the presence of neopterin? A reasonable approach is to use special recognizing materials for this purpose, prepared by molecular imprinting. This technique involves ‘stamping out’ molecules of the desired compound – their shape, but also at least some of the chemical characteristics – in a carefully designed polymer,” explains Dr. Piyush Sindhu Sharma (IPC PAS), the lead author of an article published in the Biosensors and Bioelectronics journal.

During the preparation of the polymer film, molecules of the substance being detected – in this case neopterin – are in a working solution in which their binding sites have to link with recognizing sites of so-called functional monomers. In turn, these monomers should be able to form connections with another monomer, a cross-linking agent which together, after polymerization, form a rigid support structure of the polymer. Next, the molecules of the compound used as a template are washed out from the structure. The result is a durable polymer with molecular cavities of a shape and chemical properties ensuring the capture of molecules of the desired compound from its surroundings.

The basic difficulty in molecular imprinting is the selection of the appropriate functional and cross-linking monomers as well as solvents, their proportions and reaction conditions. PhD student Agnieszka Wojnarowicz (IPC PAS) explains: “With the aid of quantum-chemical calculations, we first check whether there is bonding between our template molecule and selected functional monomers, and whether they will be stable in the solvent used. We also check whether the molecular cavities formed are sufficiently selective, i.e., whether they will primarily capture the molecules we are detecting, and not any that are similar to them. When the calculation results confirm our expectations, that is when we proceed to their experimental confirmation.”

At the IPC PAS a recognizing polymer film with molecular cavities from neopterin has been produced on the surface of an electrode. After immersion in artificial blood serum spiked with neopterin, the film on the electrode captured molecules of the latter, thus leading to a decrease in electrical potential in the connected measuring system. The tests showed that the molecular cavities of the polymer were almost entirely filled with molecules of neopterin despite the presence of molecules of similar structure and properties. This result means that the probability of false positive detection (detecting the presence of neopterin in body fluids not containing it) is negligibly small. The new chemical sensor therefore mainly reacts to what it should react to – and nothing else.

“At present, our chemosensor is a piece of laboratory equipment. However, the production of its key element, that is, the recognizing polymer film, does not pose major problems, and the electronics responsible for electrical measurements can easily be miniaturized. There is nothing standing in the way of building simple and reliable diagnostic equipment, based on our development, in just a few years’ time, which would be affordable not only for medical institutions and doctors’ surgeries, but also for the public in general,” predicts Prof. Kutner (IPC PAS).

Genetic Variations that Boost PKC Enzyme Contribute to Alzheimer’s Disease

In Alzheimer’s disease, plaques of amyloid beta protein accumulate in the brain, damaging connections between neurons. Now, researchers at University of California San Diego School of Medicine and Harvard Medical School have found that the enzyme Protein Kinase C (PKC) alpha is necessary for amyloid beta to damage neuronal connections. They also identified genetic variations that enhance PKC alpha activity in patients with Alzheimer’s disease.

The study, published May 10 in Science Signaling, may present a new therapeutic target for the disease.

“Until recently, it was thought that PKC helped cells survive, and that too much PKC activity led to cancer. Based on that assumption, many companies tested PKC inhibitors as drugs to treat cancer, but they didn’t work,” said co-senior author Alexandra Newton, PhD, professor of pharmacology at UC San Diego School of Medicine.

“Instead, we recently found that the opposite is true. PKC serves as the brakes to cell growth and survival, so cancer cells benefit when PKC is inactivated. Now, our latest study reveals that too much PKC activity is also bad, driving neurodegeneration. This means that drugs that failed in clinical trials for cancer may provide a new therapeutic opportunity for Alzheimer’s disease.”

The study was a three-way collaboration between experts in PKC (Newton), neuroscience (Roberto Malinow, MD, PhD, Distinguished Professor of Neurosciences and Neurobiology and holder of the Shiley-Marcos Endowed Chair in Alzheimer’s Disease in Honor of Dr. Leon Thal at UC San Diego School of Medicine) and genomics (Rudolph Tanzi, PhD, professor of neurology at Harvard Medical School).

Malinow’s team found that when mice are missing the PKC alpha gene, neurons functioned normally, even when amyloid beta was present. Then, when they restored PKC alpha, amyloid beta once again impaired neuronal function. In other words, amyloid beta doesn’t inhibit brain function unless PKC alpha is active.

Enter the Tanzi team, which has a database of genetic information for 1,345 people in 410 families with late-onset Alzheimer’s disease. Tanzi and team use this database to look for rare variants — genetic mutations found only in family members with the disease. Here, the team found three variants in one form of the PKC enzyme, PKC alpha that were associated with the disease in five families.

The researchers replicated these three PKC alpha gene variants in laboratory cell lines. In each instance, PKC alpha activity was increased.

While this study surfaced only five families with these rare mutations in the PKC alpha gene, there are many ways to influence PKC alpha’s activity, Newton said. She believes there could be many other inherited genetic variations that indirectly boost or inhibit PKC activity, and therefore also influence a person’s likelihood of developing Alzheimer’s disease.

“Next we want to identify more molecules participating in the pathophysiology,” said Malinow. “The more steps in the mechanism we can understand, the more therapeutic targets we’ll find for Alzheimer’s disease.”