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Mono, Multiple Sclerosis, and Cells That Live Forever: Wistar’s Dr. Paul Lieberman on Progress in EBV Research

Written by The Wistar Institute on . Posted in Featured News.

Paul M. Lieberman, Ph.D., is leader of the Genome Regulation and Cell Signaling Program at the Ellen and Ronald Caplan Cancer Center. He studies how certain viruses, such as Epstein-Barr virus, establish a long-term latent infection that can lead to cancer or autoimmune disorders.

Why did you choose to focus your research on EBV?

I became interested in EBV during my graduate student introduction to virology. Someone explained that this virus is very good at immortalizing cells, and I became intrigued by the concept of immortalization.

EBV is very good at immortalizing otherwise-mortal B lymphocytes, or B-cells, which are a type of white blood cell. If you take a person’s blood and put it in culture, those B-cells will die in a few days. But if you infect the B-cells with EBV, they can live forever. It’s basically what an oncogene can do to a cell — it can take a cell from being normal and short lived to growing forever. I wanted to know: What can make a cell live forever?

Your research works on understanding how cancer-associated viruses persist in a latent state and increase the risk of cancer and autoimmune disorders. What does it mean for a virus to persist in a latent state? Do all viruses do this?

The viruses we study, like EBV, are unique in how they establish a long-term latent infection in cells that will both be long-lived and also can divide. Most viruses do not do this. The majority of viruses, like SARS CoV 2 and flu, don’t stick around. But with all herpesviruses — and EBV is a herpesvirus — the virus finds a place to live in the body for the life of the host. Chickenpox is a good example. You get infected as a child, but the virus lingers in a long-lived neuron and then can reactivate much later and cause shingles.

The viruses we study persist in blood cells, in particular B lymphocytes. The lymphocytes have what are called memory cells, which can live for decades, and the virus finds a place to survive in those long-lived cells. EBV, like other herpesviruses, goes through periodic reactivations. In most healthy individuals, EBV reactivation is asymptomatic because other immune cells like T-cells and natural killer cells do a great job at getting rid of those virally infected cells. Unfortunately, the virus is very good at playing cat and mouse so that the immune system manages to eliminate most of the virus, but not all of it. Then, the virus waits until the immune system is compromised to reemerge or in some cases, cause cancer.

How did scientists connect EBV to multiple sclerosis? And is your lab working on anything regarding this connection?

EBV was originally identified as a “tumor virus” because it was discovered in tumor samples of a rare childhood lymphoma called Burkitt lymphoma that occurred in Central Africa. Sir Tony Epstein, who recently passed away, was responsible for that discovery.

EBV’s ability to immortalize and induce tumors associated with a number of lymphomas has been fairly easy to prove because you can isolate the cancer cells and show that the virus is in those cancer cells. However, the plot thickened when scientists found that EBV also causes mono and many people who are infected with EBV never get cancer.

EBV is now suspected to be associated with autoimmune disease — multiple sclerosis (MS) being one of those. In 2022, Alberto Ascherio, an epidemiologist at Harvard, published a longitudinal study in Science that looked at 10 million blood samples of EBV-positive and EBV-negative individuals over 20 years and found that EBV infection preceded symptoms of MS. Thanks to his work, most scientists are convinced that EBV infection along with complications of mono create something like a 32-fold increase in risk of developing MS. That’s very significant — more than smoking cigarettes increases risk of getting cancer.

The trouble is that, unlike with EBV-infected tumors, sometimes we can find the virus in a patient with MS, but in a lot of cases we can’t. One hypothesis my lab is pursuing is that very few cells are infected with EBV, but it’s enough to cause sufficient inflammation to trigger the disease. However, right now, it’s difficult to definitively show exactly where EBV is in autoimmune disease and what it’s doing. This research is important because it will determine what the best therapeutic will be.

You’re also leading an NCI-funded team of scientists to explore the role of EBV in epithelial cancers. Why study epithelial cancers?

We understand a lot about how EBV infects lymphocytes, but EBV causes on the order of about 150,000 cases of epithelial cancer — meaning nasopharyngeal carcinomas and stomach cancer — every year. And we have very little understanding of why that happens, what the virus is doing in those cancers, or the best ways to treat EBV in that context.

For this program we built a multidisciplinary team that is well-situated to invent new small-molecule drugs and identify the right combination of drugs to treat EBV-epithelial cancers. EBV-infected cancer has allowed us to be very precise because we can target the virus. With this sort of precision medicine, you can reduce a lot of the off-target side effects and resistance issues that you see with other treatments like some of the more severe chemotherapeutics.

What excites you most about where EBV research is headed?

I think we are in the chase for a cure. This opportunity to target EBV across a number of different diseases is what keeps me wanting to get back to work, day after day. If we can eliminate the virus or eliminate the activity of the virus, we can cure a lot of disease.

To support the research of viruses like EBV, donate to Wistar’s Giving Tuesday campaign.

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The Wistar Institute and University of Pennsylvania Scientists Identify Potential Target Gene within Certain HIV Reservoir Cells

Written by Steven Schneible on . Posted in Press Releases.

PRESS RELEASE
Drs. Lieberman, Collman, and Co-Authors Link RSAD2/Viperin Gene with Certain
Chronically HIV-Infected Cells

PHILADELPHIA — (Oct. 3, 2024) —New findings could lead to different research tactics for scientists investigating a cure for HIV. Results from The Wistar Institute’s Lieberman lab, led by Hilary Koprowski, M.D., Endowed Professor Paul M. Lieberman, Ph.D., and researchers at the Perelman School of Medicine’s Center for AIDS Research and center director Ronald G. Collman, M.D. — have identified the RSAD2/Viperin gene as a potential HIV treatment target within certain HIV reservoir cells. Their results were published in the paper, “HIV-induced RSAD2/Viperin supports sustained infection of monocyte-derived macrophages,” in the Journal of Virology.

HIV does not have a cure because there is no known method — yet — for eliminating the virus from the body once infected. Although HIV can be managed with antiretroviral therapy (ART), the virus persists in infected cells throughout the body, called “HIV reservoirs.” Reservoirs not only serve as the main barrier in HIV cure research, which focuses in large part on strategies to destroy these HIV reservoirs, but also contribute to chronic inflammation and comorbidities in people living with HIV.

A certain type of immune cell — myeloid cells, including macrophages and microglia — often serves as an HIV reservoir because, unlike other cells infected with HIV, these cells tend not to be killed by HIV’s viral replication. Due to their comparative longevity as reservoirs and prevalence within the nervous system, HIV-infected macrophages often cause neurocognitive complications in people with HIV that develop even despite antiretroviral treatment (ART).

Lieberman and Collman, joined forces to research the genetics of macrophages that might play a role in maintaining the HIV reservoir status quo. Their investigation revealed a surprising candidate in the gene RSAD2/Viperin — which usually fights viruses. In HIV-infected macrophages, RSAD2/Viperin expression was quite high compared both with controls and HIV-infected CD4+ T cells (the other major type of cell that can become a viral reservoir of HIV).

RSAD2/Viperin is a gene associated with interferon response, and typically, both the gene itself and the interferons that trigger its activation have antiviral effects. However, certain interferons have been found to play paradoxical roles in chronic HIV by enabling the virus’ persistence, and upon finding RSAD2/Viperin’s elevated expression in reservoir macrophages, the researchers hypothesized that the gene must be abetting HIV’s continued presence within these cells.

To test this, the research team used the siRNA method to target and reduce RSAD2/Viperin’s expression in macrophages infected with HIV. Once they reduced the gene’s expression, several measures of HIV’s presence and activity fell, including viral transcripts, p24 protein production, and multinucleated giant cells (another indicator of active viral activity). Reduced RSAD2/Viperin also altered histone modification of HIV genomes — that is, the control of HIV’s latency by chromatin and epigenetic factors. These findings suggest a novel role for RSAD2/Viperin in regulating chromatin that otherwise might suppress HIV replication during latency.

“Looking closely at RSAD2/Viperin in these HIV-infected MDMs, we’ve identified yet another paradox of HIV infection,” said Lieberman, program leader, Genome Regulation and Cell Signaling Program, Ellen and Ronald Caplan Cancer. “Our data show that while, yes, this is an antiviral gene that can come to the body’s defense against the virus at first, it also seems to maintain HIV’s ability to persist as a chronic infection. That makes RSAD2/Viperin a compelling candidate for further research and possible targeting of HIV reservoirs — which is critical to future cure research.”

“We’ve come to understand yet another facet of chronic HIV infection’s complexity,” agreed Collman. “We’re hopeful that these findings will be helpful as the field continues to pursue possible therapeutic interventions that would eliminate the viral reservoir in the search for an HIV cure, or reduce negative consequences of infection that can persist even despite effective therapy, such as neurocognitive decline.”

Co-authors: Urvi Zankharia, Fang Lu, Olga Vladimirova, Bhanu Chandra Karisetty, Jayamanna Wikramasinghe, Andrew Kossenkov, and Paul M. Lieberman of The Wistar Institute; and Yanjie Yi and Ronald G. Collman of The Perelman School of Medicine at The University of Pennsylvania.

Work supported by: NIH grants R6133-133696, P30AI045008, and P30CA010815.

Publication information: “HIV-induced RSAD2/Viperin supports sustained infection of monocyte-derived macrophages,” from Journal of Virology.

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ABOUT THE WISTAR INSTITUTE:

The Wistar Institute is the nation’s first independent nonprofit institution devoted exclusively to foundational biomedical research and training. Since 1972, the Institute has held National Cancer Institute (NCI)-designated Cancer Center status. Through a culture and commitment to biomedical collaboration and innovation, Wistar science leads to breakthrough early-stage discoveries and life science sector start-ups. Wistar scientists are dedicated to solving some of the world’s most challenging problems in the field of cancer and immunology, advancing human health through early-stage discovery and training the next generation of biomedical researchers. wistar.org

ABOUT PENN MEDICINE:

Penn Medicine is one of the world’s leading academic medical centers, dedicated to the related missions of medical education, biomedical research, excellence in patient care, and community service. The organization consists of the University of Pennsylvania Health System and Penn’s Raymond and Ruth Perelman School of Medicine, founded in 1765 as the nation’s first medical school.

The Perelman School of Medicine is consistently among the nation’s top recipients of funding from the National Institutes of Health, with $550 million awarded in the 2022 fiscal year. Home to a proud history of “firsts” in medicine, Penn Medicine teams have pioneered discoveries and innovations that have shaped modern medicine, including recent breakthroughs such as CAR T cell therapy for cancer and the mRNA technology used in COVID-19 vaccines.

The University of Pennsylvania Health System’s patient care facilities stretch from the Susquehanna River in Pennsylvania to the New Jersey shore. These include the Hospital of the University of Pennsylvania, Penn Presbyterian Medical Center, Chester County Hospital, Lancaster General Health, Penn Medicine Princeton Health, and Pennsylvania Hospital—the nation’s first hospital, founded in 1751. Additional facilities and enterprises include Good Shepherd Penn Partners, Penn Medicine at Home, Lancaster Behavioral Health Hospital, and Princeton House Behavioral Health, among others.

Penn Medicine is an $11.1 billion enterprise powered by more than 49,000 talented faculty and staff

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Breaking Down the Science: Wistar’s New Genome Regulation and Cell Signaling Program in the Ellen and Ronald Caplan Cancer Center

Written by The Wistar Institute on . Posted in Featured News.

Upon the launch of The Wistar Institute’s new Genome Regulation and Cell Signaling Program, we sat down with the Program’s leader — Hilary Koprowski, M.D., Endowed Professor Paul M. Lieberman, Ph.D. — and co-leader, professor Bin Tian, Ph.D., to learn more about their vision for the Program and what it means for the future of cancer research at Wistar.

Tell us about the importance and excitement in this new Caplan Cancer Center Program.

PL: Wistar has been at the forefront of cancer research and discovery for decades. Recent advances in genomic technology and computational biology have inspired us to establish the new Genome Regulation and Cell Signaling (GRCS) Program. This new Program brings together a diverse group of investigators to solve complicated problems underlying cancer causation, including persistent viral infection, immune response to cancer, and cell signaling changes in cancer.

The GRCS Program combines multi-disciplinary expertise to solve these complicated problems from many different angles: from specialists in genome architecture and integrity like Drs. Tempera, Gardini, and Sarma, who focus on the physical structure of our genome, which has very critical ramifications for genetic diseases & cancers; to investigators interested in virus’ infection and cancer causation, like myself, Dr. Tempera, and Dr. Price; to researchers of inflammatory signals in cancer cells and metastasis with Drs. Chen and Altieri; to specialists in computational and informatic techniques like Drs. Madzo, Kossenkov, and Srivastava, who are essential for effective analysis and interpretation of the vast datasets our Program generates.

How does the combination of genome regulation and cell signaling synergize in this new Program?

BT: We want to better understand cancer, so we study how genes are regulated or dysregulated at the genomic level; cell signaling provides the biological context for understanding how genome regulation plays out in response to internal & external cues. Because of recent technological advances, gene regulation can now be studied across the entire genome — not just individual genes or small groups of genes with similar functions, but all the genes throughout the genome. The synergy between genome regulation and cell signaling is holistic.

PL: Genome regulation does not occur in a vacuum. Metabolic and environmental changes trigger signaling between cells, which, in turn, affect how the genome is regulated. How the genome responds to these types of signals is central to the problems in cancer biology and part of the new focus areas of the GCSR Program.

Why is genomics so critical to cancer research?

BT: All aspects of a cancer cell’s growth — from tumor formation, to tumor survival, to interactions with other cells in the tumor microenvironment — involve rewiring of our cells’ gene expression programs. And that is a fundamentally genome-based process — whether it is a physical change of some genome sequences; alternation of how the genome is structured in 3-dimentional space; transcription of DNA into RNA; or post-transcriptional regulations.

PL: Cancer is a disease of the genome: tumors start when the genome is changed in ways that give cancer a foot in the door. Genomics and informatics analyses allow us to understand the specific genetic changes — which, in cancer, are more like genetic injuries — that drive a particular individual’s cancer. Ideally, once you understand the underlying genetic nature of an individual cancer, you can design precision medicines targeted more accurately to a specific disease diagnosis.

What advances has sequencing technology unlocked in this area of cancer research?

BT: In essence, cancer is a genetic disease. And advanced sequencing technologies have enabled us to examine the genome with the resolution of a single nucleotide — the fundamental building block of DNA. Sequencing technologies have evolved to a point where we can even use these tools to understand the dynamics of genome regulation within individual cells or tiny regions in the body. Essentially, we see cancer’s real-time changes far more clearly, which is key to understanding and combatting the disease.

PL: Advances in genomics and sequencing technologies allow us to understand cancer as a personal disease. Each tumor is different, but we can use precision sequencing as a springboard for researching precision medicine. Armed with the latest advances like next-generation and ultra-high-throughput sequencing — methods that allow scientists to accurately assess entire genomic samples and in minute detail — the new Program’s scientists have the tools they need to move the field even further.

In state-of-the-science Wistar labs, our researchers can easily sample an entire genomic state with tools to improve and expand into new areas of application and translation. Our Program members combine these advances with technologies like CRISPR to identify, target, modify, and correct the genetic aberrations that drive cancer and other genetic diseases.

How do cancer researchers deal with the complexity of the different variables at play in cancer? And how will your Program’s approach account for that interconnectivity?

PL: Due to the complexity of biological systems — and cancer being among the most complicated biological problem because of the rapid, chaotic evolution of tumors and their surroundings — it’s quite unlikely that any single person or brain will solve this challenge. New artificial intelligence applications are welcome tools for investigators; by leveraging AI, we can sort through the massive amounts of biological information and identify potential vulnerabilities within cancer’s framework.

We do work in a reductionist mindset — where the entire complex network of information is reduced to one simple example — to identify new targets and pharmacological agents that can impact the whole system. While that might seem at odds with cancer’s enormous complexity, we still need to simplify the complex science of cancer. It’s a give and take: we zoom in to find a specific mechanism at play in cancer, and then we zoom out to see whether targeting that mechanism can work its way through the vast, interconnected complexity of the disease system to produce a therapeutic effect. We cut through the jungle one molecule at a time.

What is your plan for translating your Program’s discoveries into testable therapy strategies?

BT: We have several promising thematic areas for therapeutic intervention, including the emerging areas of mRNA vaccines and gene therapies, as well as continued progress in small molecules as drug candidates. So as we make progress on potential therapeutics, we seek to take full advantage of several technologies and investigate how they work together — similar to the multi-pronged approaches the HIV folks are using for disease containment and cure.

We believe in basic science, which pays off in the long run: any discovery and innovation moves the needle in cancer research and future therapeutics.

Five years from now, what do you hope to have achieved through the Program?

BT: We hope to achieve breakthroughs in both basic science research and cancer therapeutics; we can reach these goals because the GCRS Program has faculty with expertise in many cutting-edge and interdisciplinary technologies and is highly collaborative.

PL: The GRCS program has two main goals: advance our knowledge and understanding of the complex mechanisms of genome regulation and cell signaling in cancer; and second, identify new therapeutic targets and strategies to treat cancer and other complex diseases.

We anticipate publications in high-impact journals to highlight breakthroughs in genome regulation and cell signaling, and we also expect to see some of our findings advanced into new therapeutics — small molecules, gene therapies, and vaccines to treat cancer and other diseases — that will reach clinical trials thanks to our continued collaboration between the public and private sectors.

These are broad and ambitious goals, but they are achievable. With an excellent diversity of scientific expertise and supported by the most advanced technologies available from Wistar’s Shared Resources facilities, the GCRS Program is positioned to find answers to some of the most pressing questions in cancer biology.

Wistar Research Identifies Mechanisms for Selective Multiple Sclerosis Treatment Strategy

Written by The Wistar Institute on . Posted in Press Releases.

PRESS RELEASE
Wistar’s Lieberman lab stopped inflammatory signaling and immune response in lab samples

PHILADELPHIA — (May 28, 2024) — The Wistar Institute’s Paul M. Lieberman, Ph.D., and lab team led by senior staff scientist and first author, Samantha Soldan, Ph.D., have demonstrated how B cells infected with the Epstein-Barr virus (EBV) can contribute to a pathogenic, inflammatory phenotype that contributes to multiple sclerosis (MS); the group has also shown how these problematic B cells can be selectively targeted in a way that reduces the damaging autoimmune response of multiple sclerosis. The lab’s findings were published in Nature Microbiology in the paper, “Multiple sclerosis patient derived spontaneous B cells have distinct EBV and host gene expression profiles in active disease.”

EBV — a usually inactive, or latent, herpesvirus — affects most of the human population; more than 90% of people carry the virus as a passive, typically symptomless infection. However, EBV infection has been linked to several diseases, including MS: an incurable, chronic autoimmune disease that causes the body’s immune system to attack the myelin sheath of neurons in the brain and nervous system. Because myelin sheathing facilitates fast nervous system signaling (the fatty insulation of myelin along a neuron’s axon allows electrical impulses to travel through neuronal networks faster), its degradation can cause a wide variety of symptoms in both type and severity that may include motor control disruption, sensory issues, and speech difficulties.

Though researchers know that EBV can contribute to the development of MS, the exact mechanisms by which it does so aren’t completely understood. The Lieberman lab, in seeking to understand how EBV contributes to the development of MS, collaborated with Steven Jacobson, Ph.D., of the Neuroimmunology Branch at the National Institute of Neurological Disorders and Stroke, who contributed cell line samples from patients. The research team analyzed spontaneous lymphoblastoid cell line (SLCL) cell samples from a healthy control group; a group of patients with active MS (as opposed to so-called stable MS; the disease is characterized by unpredictable periods of flare-ups and eased symptoms); and a group of patients with stable MS.

B cells are crucial cells of the immune system that help regulate the body’s immune responses; they have also been implicated in autoimmune conditions due to their role as mediators of which biological signals warrant immune response. And B cells, when infected with EBV, become immortalized — that is, the cells are no longer constrained by senescence, so they can continue to divide an indefinite number of times — as “lymphoblastoid cell lines,” or LCLs. This immortalized B cell state can occur spontaneously within the body as a result of EBV infection, which is how the Lieberman lab was able to extract immortalized SLCL samples for study from the different patient groups.

Having obtained the matched samples, Dr. Lieberman and his team conducted genetic analyses of the SLCLs and confirmed that the MS-positive sample groups showed greater expression of genes associated with lytic EBV (“lytic” describes when latent viruses like EBV become active); they also saw increased inflammatory signaling and expression of the FOXP1 protein, the latter of which was shown to promote lytic EBV gene expression. As a whole, the group’s findings suggested a mechanism of lytic EBV in MS that promoted inflammation and disease.

Diving further, Lieberman’s group tested several antiviral compounds on all SLCL groups and found that one, TAF, reduced lytic EBV gene expression without killing the cells. TAF also significantly reduced the expression of inflammatory cytokines like IL-6 in the SLCLs from the patients with active MS. Finally, when cultured SLCLs from active MS, stable MS, and controls were administered TAF in the presence of antiviral T cells, the T cell response (a major factor in the autoimmune dysfunction of MS) was reduced in SLCLs from patients with MS but not reduced in the control SLCLs — an indication that TAF treatment has potential as a selectively cytotoxic anti-lytic treatment for MS.

“Our work with these SLCLs shows that the problematic inflammation signaling from lytic EBV can be selectively targeted in a way that demonstrably reduces damaging immune responses,” said Dr. Lieberman. “We’re excited about expanding this concept further; we have the potential to see whether TAF or other inhibitors of EBV might be a viable treatment for multiple sclerosis that can stop the autoimmune damage without causing wide-ranging and dangerous cell death.”

Co-authors: Samantha S. Soldan, Chenhe Su, Leena Yoon, Toshitha Kannan, Urvi Zankharia, Rishi J. Patel, Jayaraju Dheekollu, Olga Vladimirova, Jack Dowling, Natalie Brown, Andrew Kossenkov, Daniel E. Schäffer, Noam Auslander, and Paul M. Lieberman of The Wistar Institute; Maria Chiara Monaco and Steven Jacobson of the Neuroimmunology Branch at the National Institute of Neurological Disorders and Stroke; Jack Dowling and Simon Thebault of the Perelman School of Medicine; Annaliese Clauze, Frances Andrada, and Joan Ohayon of the Neuroimmunology Clinic at the National Institute of Neurological Disorders and Stroke; and Andries Feder and Paul J. Planet of the Children’s Hospital of Philadelphia.

Work supported by: This work was supported by grants from the National Institutes of Health (R01 CA093606, R01 AI153508, R01DE017336 to PML, the Wistar Cancer Center Core Grant P30 CA010815), and the Department of Defense (HT9425-23-1-1049 Log#MS220073). The funders had no role in study design; data collection and analysis; decision to publish; or preparation of the manuscript.

Publication information: “Multiple sclerosis patient derived spontaneous B cells have distinct EBV and host gene expression profiles in active disease,” from Nature Microbiology

ABOUT THE WISTAR INSTITUTE:

The Wistar Institute is the nation’s first independent nonprofit institution devoted exclusively to foundational biomedical research and training. Since 1972, the Institute has held National Cancer Institute (NCI)-designated Cancer Center status. Through a culture and commitment to biomedical collaboration and innovation, Wistar science leads to breakthrough early-stage discoveries and life science sector start-ups. Wistar scientists are dedicated to solving some of the world’s most challenging problems in the field of cancer and immunology, advancing human health through early-stage discovery and training the next generation of biomedical researchers. wistar.org

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Dr. Paul Lieberman talks with two scientists in the lab

Wistar-Led Team Awarded More Than $12 Million Grant from the NCI to Investigate Link Between Epstein-Barr Virus and Carcinomas

Written by The Wistar Institute on . Posted in Press Releases.

PHILADELPHIA — (JULY 26, 2023) — It’s been known since the 1960s that Epstein-Barr Virus (EBV) causes a variety of cancers, but research has overwhelmingly focused on its connection to lymphomas. Now, a multidisciplinary team of scientists led by The Wistar Institute has been awarded a more than $12 million National Cancer Institute (NCI) Program Project Grant (P01), a highly competitive five-year grant that includes a cross section of researchers from various disciplines and institutions throughout the country. The multidisciplinary team led by Wistar scientists is exploring the role of Epstein-Barr Virus in epithelial cancers. Epithelial cells form functional structures in organ tissue throughout the human body; they are often the site for solid organ cancers, including the most common cancers, which are known as carcinomas.

The new research will focus on basic questions about how EBV infection of normal epithelial cells transforms them into cancer-cells. Scientists also intend to build on this research to identify better and more selective therapeutic targets.

“We are investigating unexplored aspects of EBV and malignancies, potentially uncovering unique characteristics or pathways that can be targeted for therapeutic intervention,” said Italo Tempera, Ph.D., associate professor of the Gene Expression & Regulation Program of the Ellen and Ronald Caplan Cancer Center at The Wistar Institute. “This fresh perspective could lead to groundbreaking discoveries and innovative treatment strategies for EBV and epithelial malignancies.”

The project brings together scientists from The Wistar Institute and Harvard University, including experts in epigenetics, metabolomics and drug discovery. It’s the first time researchers from this variety of disciplines have combined their efforts to focus entirely on the EBV-epithelial cancer link.

“We’ve put together a new strategy, a new way of attacking the problem,” said Paul Lieberman, Ph.D., Hilary Koprowski, M.D., Endowed Professor and director of the Center for Chemical Biology and Translational Medicine at Wistar. “By working together across different modalities, there’s an opportunity for each of us to learn from the synergy and expertise of the other investigators.”

EBV is one of the most common human viruses, infecting an estimated 95% of people by the time they reach adulthood. Symptoms are usually mild, and most people recover within a few weeks. However, the virus can remain latent in the human body for years or even decades, and it causes some people to develop cancer later in life.

While research has historically focused on lymphomas, EBV-linked epithelial cancers are both more common and more deadly. Epithelial cancers represent 75% of the 200,000 EBV-related cancer cases diagnosed each year, and these cancers also have higher mortality rates and treatment failures.

“This grant put together a team that is now focused on this type of cancer that has been neglected, even though it’s the most common form of EBV cancers,” Lieberman said. The grant will fund three main research projects. The first will look at how EBV establishes a long-term infection within epithelial cells. The second will study how it causes genetic and metabolic changes to trigger cancer growth. Finally, researchers will use these findings to investigate new therapeutic strategies.

The research builds on past work by Lieberman’s lab, which has focused on developing small molecule inhibitors targeting EBV. He said the new project would focus on studying drugs that are already in development, and looking for ways to make them more targeted or use them in combination with other therapies.

Tempera said the group’s integrated approach sets it apart.“Our project will study both metabolic and epigenetic vulnerabilities simultaneously,” he said. “Combining these two aspects can provide a comprehensive understanding of the role of EBV infection in cancer and its underlying mechanisms, leading to unique insights and therapeutic opportunities.”

Co-authors: Ben Gewurz of Harvard; Joseph Salvino, Samantha Soldan, Andrew Kossenkov, Louise Showe, and Qin Liu of Wistar.

Unraveling the Ties That Bind: Epstein-Barr Virus and Multiple Sclerosis

Written by The Wistar Institute on . Posted in Featured News.

A conversation with the Lieberman Lab delves into how EBV can trigger MS and potential therapeutic solutions that can be developed with this knowledge.

Epstein-Barr virus (EBV) is ubiquitous, establishing lifelong infection found throughout the world. It targets the immune system’s B cells and typically remains silent in immune system memory cells. Though infection with the virus is largely asymptomatic, specific biological and environmental conditions can enable the virus to cause more serious diseases. For instance, the virus can cause rare cancers that occur at much higher rates in immunosuppressed individuals. More recent research has found a connection between EBV and the neurodegenerative disease, multiple sclerosis (MS).

In a recent article published in Nature Reviews Microbiology, Paul Lieberman, Ph.D., Hilary Koprowski, M.D., Endowed Professor; program leader, Gene Expression & Regulation Program, Ellen and Ronald Caplan Cancer Center; and director, Center for Chemical Biology & Translational Medicine; and Samantha Soldan, Ph.D., staff scientist in the Lieberman laboratory, review evidence of EBV as a cause of MS and the implications of this knowledge in research and clinical spheres.

“Eliminating EBV latent infection should be a safe and effective way to treat EBV cancers and autoimmune disease, especially multiple sclerosis,” Lieberman says.

We spoke with Soldan about the link between EBV and MS as well as how this knowledge can be harnessed into potential therapeutics for the disease.

Q: What is some of the background linking EBV to cancer?

A: EBV was the first virus to be implicated as a causative agent of human cancer. EBV has been linked to nasopharyngeal cancers, stomach cancers, primary CNS lymphomas, Hodgkin’s and non-Hodgkin’s lymphomas, NK/T-cell lymphomas, and leiomyosarcomas, as well as several autoimmune disorders, including MS. It is currently estimated that 1.5-2% of all human cancers are attributable to EBV infection. All EBV-related cancers are associated with latent infection, where no infectious virus is produced by the tumor cells. Different types of EBV-associated cancers express different EBV latency genes. Notably, all EBV cancers express the viral protein EBNA1—required to maintain the EBV genome in latently infected cells and a primary interest of the Lieberman lab.

Q: What inspired this review to investigate evidence behind EBV as a cause of MS?

A: The case for EBV as a causative agent in MS has been mounting over the last 40 years. This year, two landmark studies were published: one providing a strong epidemiologic link to MS and the other suggesting a mechanism by which EBV may drive disease pathogenesis. These two studies have intensified the MS community’s interest in the link between EBV and MS, both as a disease trigger and as a potential driver of disease pathogenesis.

Dr. Paul Lieberman is a leading expert in EBV and in the study of EBV EBNA1 specifically, and he was invited to write a review article focused on the role of EBV in MS for Nature Reviews Microbiology. Paul asked me to be a co-author for this manuscript and I was delighted to have this opportunity. My background is in neurovirology, and I have been involved in research investigating the relationship between viruses and neuroinflammatory diseases, including MS, since graduate school. I am deeply invested in trying to better understand the role that EBV plays in MS.

Q: Why is multiple sclerosis a complex disease to study?

A: Multiple sclerosis is the most common demyelinating disorder of young adults, effecting more than one million individuals in the U.S. alone. MS is a heterogenous and often disabling disease that develops because of the interplay between the immune system and the environment in genetically susceptible individuals. The clinical progression of MS is variable and unpredictable with several distinct disease courses. In addition, MS patients often transition from a relapsing-remitting to a progressive disease course over time and the mechanism of the disease and central nervous system damage also evolve, which are a hallmark of MS.

Further complicating matters, epidemiological studies have shown that environmental factors (including EBV infection) that contribute to one’s risk of developing MS often occur many years before clinical onset. Collectively, these complex interactions between genetic, immunologic, and environmental risk factors makes attributing disease-contributing agents and designing preventative measures and effective therapies for MS very challenging.

Q: How does EBV trigger MS? Can you explain these processes?

A: How a ubiquitous agent like EBV triggers disease in a small percentage of those who are infected is an enigma. We face this challenge in understanding the role of EBV in cancer as well as MS. For MS and many autoimmune diseases, we can identify inflammatory and autoreactive immune responses and characterize immune responses to infectious agents and antigens that trigger immune response in patients. However, understanding what set immune cells on an autoreactive and inflammatory path before the patient is symptomatic is a difficult task.

Nevertheless, there are several theories as to how EBV may be both a trigger and a driver of MS and we discuss these in the review. We believe that it is likely that EBV is involved in the pathogenesis of MS at many levels and in different anatomic compartments.

Q: What are some potential therapies that could arise out of understanding the role of EBV in MS?

A: In recent years, therapies depleting B-cells have proven to be tremendously beneficial in MS. While EBV primarily infects B-cells, these B-cell depletion therapies eliminate cells regardless of whether they are infected by EBV, making it difficult to determine if any of the clinical benefit derived from these drugs is related to their effects on the virus.

There are several EBV specific therapies in development that have the potential to present new, effective options for patients with MS. In addition, they may also help us better understand the role of EBV as a trigger and driver of disease pathogenesis. These include vaccines to prevent the development of infectious mononucleosis, MS, and EBV-associated cancers; cell-based immunotherapies, including EBV-specific cytotoxic T cell lines; and EBV specific antivirals.

Q: Why is it important to review existing literature and reveal new directions for research?

A: I find review articles to be incredibly helpful, both to the writer and the reader. When writing a review, you must commit time and energy to refamiliarizing yourself with the latest literature as well as the history of the field. The process helps you take the proverbial 30,000-foot view and see the whole picture, forcing you to get your head out of the specific aspect of research that you are generally focused on and enabling you to generate new ideas and consider new approaches to your work. Reviews are also very important for colleagues and especially trainees to get perspective on where the field is headed and where there are gaps in our knowledge.

Q: What do you currently work on with Dr. Lieberman regarding MS and EBV and where is your research headed?

A: We are working in several directions to better understand the role that that EBV plays in the pathogenesis of MS. Our current work focuses on characterizing virus-host interactions and maintenance of EBV latency in EBV infected B-cells from MS patients compared to healthy controls. Dr. Chenhe Su, a postdoctoral fellow in the Lieberman lab, is also working very hard on these studies.

I am especially interested in developing better animal models to understand how host factors like age of exposure, genetic background, metabolism, and sex influence host-virus interactions and EBV reprogramming of B-cells. We are also testing EBV-specific antiviral therapies, including the EBNA1 inhibitor developed in the Lieberman lab, to determine its potential as a therapeutic agent for use in MS.

Viruses and Cancer

Written by The Wistar Institute on . Posted in Featured News.

Viruses impact human health in many ways and can be the underlying agents that cause cancer.

The pandemic has reminded us how pervasive viruses are in human life. We know they can cause infection, making us very sick, but did you know that about 15 percent of all human cancers worldwide may be attributed to viruses?

Drs. Paul Lieberman, professor, and Italo Tempera, associate professor, both in the Gene Expression & Regulation Program of The Wistar Institute Cancer Center, investigate the link between the Epstein-Barr Virus (EBV) and malignant transformation to find ways to treat EBV-related cancers.

EBV is a very common virus. You know it as mononucleosis, and it is spread through saliva, but also through sexual contact, blood transfusions and organ transplants. It mostly infects B cells, a type of white blood cells that produces antibodies. The majority of the world population is infected and carries the virus in a silent state for life. However, in people with compromised immune systems, EBV increases the risk of B-cell lymphomas including Burkitt’s lymphoma and Hodgkin’s lymphoma.

Since it can also infect other types of cells, this virus is not only associated with blood tumors, but also with ten percent of all gastric cancers and most cancers of the nasopharynx, the region behind the nose and above the back of the throat.

How are viruses implicated in cancer?

They are particles made up of DNA or RNA surrounded by a protein coat. With such frugality comes the lack of the necessary machinery to replicate their genome or build proteins, so viruses hijack a host cell and use its machinery to make copies of themselves. In the case of oncogenic or cancer-causing viruses, this process can meddle with the cell’s genes and derail the mechanisms that keep cell proliferation in check, leading to uncontrolled growth.

Oncogenic viruses establish chronic but latent infections that don’t cause obvious symptoms in healthy individuals, because their immune system keeps infection at bay. In immunocompromised people, however, viruses are more likely to cause malignancies.

One approach to control EBV infection is by interfering with the complex and dynamic patterns of how viral genes are switched on or off — a process known as gene expression. The Tempera lab has explored the role of gene expression in the regulation of EBV latency, unveiling some viral and host cell factors that play key roles in regulating these patterns.

An EBV protein called latent membrane protein 1 (LMP1) is essential for the virus’s ability to make B cells cancerous.

The Tempera lab studies just how this protein affects the host cells. They discovered that LMP1 affects the function of an enzyme called poly(ADP-ribose) polymerase 1 (PARP1) and that inhibiting this protein suppresses malignant transformation, uncovering an important role of PARP1 in EBV-induced oncogenesis.

In a recent paper1, Tempera and colleagues demonstrated that LMP1 causes a switch in how infected cells produce the building blocks of fat that support tumorigenesis and cancer progression. Researchers also showed that targeting these metabolic changes could be an effective therapeutic strategy to treat EBV-associated cancer and that PARP1 inhibitors offset the metabolic changes caused by LMP1 that drive tumorigenesis.

“We have uncovered a potential new use for PARP inhibitors, which are currently used to treat recurrent ovarian cancers in women who have defects in their DNA repair system,” said Dr. Tempera. “Repurposing existing drugs saves time and money in the process of creating new therapies.”

Dr. Tempera joined Wistar as an associate professor in 2020, but that was not his first time working at the Institute. He had trained as a postdoctoral fellow in the Lieberman lab, which helped him launch his career in cutting-edge research on the epigenetic mechanisms underlying EBV infection. Returning to Wistar with his lines of research as an established investigator, he strengthened our program on virology and cancer.

“The outstanding scientific environment and technological support that benefited me during my training are a strong asset to further expand my research program.”

The lab of his mentor Dr. Lieberman is a reference point for EBV research as they have made seminal discoveries in the field and described several mechanisms that control replication and gene expression in latent EBV infection. One focus of their research is the Epstein-Barr nuclear antigen 1 (EBNA1) protein that is essential for efficient viral DNA replication. The team has been pursuing the development of small molecule inhibitors of EBNA1 as potential treatment against EBV-associated malignancies.

One of the lab’s latest studies2 led to an important basic discovery in the field, expanding the understanding of EBNA1’s function and providing new possibilities for inhibiting EBNA1 activity as an anticancer strategy.

The link between virus infection and human cancer is a complicated matter to unravel and has been studied for more than 100 years. The research underway in the Tempera and Lieberman labs is crucial to understanding the role viruses play in malignant transformation and hopefully finding future treatments to halt this process.

1 Epstein-Barr Virus-Encoded Latent Membrane Protein 1 and B-Cell Growth Transformation Induce Lipogenesis through Fatty Acid Synthase, J Virol, 2021
2 Cell-cycle-dependent EBNA1-DNA crosslinking promotes replication termination at oriP and viral episome maintenance, Cell, 2021

rendering of Epstein Barr Virus protein Genome Regulation and Cell Signaling (GCRS) Program

Wistar Scientists Make Pivotal Discovery on the Mechanism of Epstein-Barr Virus Latent Infection

Written by The Wistar Institute on . Posted in Press Releases.

PHILADELPHIA — (Jan. 21, 2021) — Researchers at The Wistar Institute have discovered a new enzymatic function of the Epstein-Barr Virus (EBV) protein EBNA1, a critical factor in EBV’s ability to transform human cells and cause cancer. Published in Cell, this study provides new indications for inhibiting EBNA1 function, opening up fresh avenues for development of therapies to treat EBV-associated cancers.

EBV establishes life-long, latent infection in B lymphocytes, which can contribute to development of different cancer types, including Burkitt’s lymphoma, nasopharyngeal carcinoma (NPC) and Hodgkin’s lymphoma.

The Epstein-Barr Nuclear Antigen 1 (EBNA1) serves as an attractive therapeutic target for these cancers because it is expressed in all EBV-associated tumors, performs essential activities for tumorigenesis and there are no similar proteins in the human body.

“We discovered an enzymatic activity of EBNA1 that was never described before, despite the intense research efforts to characterize this protein,” said Paul M. Lieberman, Ph.D., Hilary Koprowski, M.D., Endowed Professor, leader of the Gene Expression & Regulation Program at Wistar, and corresponding author of the study. “We found that EBNA1 has the cryptic ability to cross-link and nick a single strand of DNA at the terminal stage of DNA replication. This may have important implications for other viral and cellular DNA binding proteins that have similar cryptic enzyme-like activities.”

Lieberman and colleagues also found that one specific EBNA1 amino acid called Y518 is essential for this process to occur and, consequently, for viral DNA persistence in the infected cells.

They created a mutant EBNA1 protein in which the critical amino acid was substituted with another and showed that this mutant could not form covalent binding with DNA and perform the endonuclease activity responsible for generating single strand cuts.

In latently infected cells, the EBV genome is maintained as a circular DNA molecule, or episome, that is replicated by cellular enzymes along with the cell’s chromosomes. When the cell divides, the two episomal genomes segregate into the two daughter cells.

While it was known that EBNA1 mediates replication and partitioning of the episome during division of the host cell, the exact mechanism was not clear. The new study sheds light on the process and describes how the newly discovered enzymatic activity of EBNA1 is required to complete replication of the viral genome and maintenance of the episomal form.

“Our findings suggest that one could create small molecules to ‘trap’ the protein bound to DNA and potentially block replication termination and episome maintenance, similar to known inhibitors of topoisomerases,” said Jayaraju Dheekollu, Ph.D., first author on the study and staff scientist in the Lieberman Lab. “Such inhibitors may be used to inhibit EBV-induced transformation and treat EBV-associated malignancies.”

Co-authors: Andreas Wiedmer, Kasirajan Ayyanathan, Julianna S. Deakyne, and Troy E. Messick from The Wistar Institute. K.A. is currently employed at University of Pennsylvania and J.S.D. is currently employed at GlaxoSmithKline.

Work supported by: National Institutes of Health (NIH) grants RO1 CA093606, RO1 423 DE017336, P30 CA010815, and T32 CA09171.

Publication information: Cell Cycle-Dependent EBNA1-DNA Cross-Linking Promotes Replication Termination at oriP and Viral Episome Maintenance, Cell (2021). Online publication.

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The Wistar Institute is an international leader in biomedical research with special expertise in cancer, immunology, infectious disease research, and vaccine development. Founded in 1892 as the first independent nonprofit biomedical research institute in the United States, Wistar has held the prestigious Cancer Center designation from the National Cancer Institute since 1972. The Institute works actively to ensure that research advances move from the laboratory to the clinic as quickly as possible. wistar.org.

Spotlight on Wistar COVID-19 Researcher: Paul Lieberman, Ph.D.

Written by The Wistar Institute on . Posted in Featured News.

Creating a new drug from scratch takes many years and billions of dollars. While discovery of new molecules against SARS-CoV-2 and other emerging viruses is imperative, the world also needs solutions for COVID-19 now. This is why there has been wide interest in repurposing existing drugs, for which safety and pharmacologic profiles are already available, as a viable strategy to save critical time and resources and quickly discover potential candidate drugs to combat the pandemic. 

Dr. Paul Lieberman is a molecular virologist in Wistar’s Cancer Center, better known for his pioneering studies on how certain viruses persist in the body in a latent, long-term infection that can lead to cancer. He leads a talented  drug-discovery team that has created first-in-class molecules directed against  Epstein-Barr virus (EBV) as a novel therapeutic approach for potentially treating EBV-associated cancers including Burkitt’s lymphoma, nasopharyngeal carcinoma and Hodgkin’s lymphoma.

Applying their antiviral drug discovery expertise and innovative methods to “drug” the RNA component of viruses and cancer cells, the Lieberman lab is now working on a drug-repurposing project to quickly identify FDA-approved molecules that trap and inactivate SARS-CoV-2 RNA, the genetic material that carries the life information for the virus, just like DNA does in our cells. 

SARS-CoV-2 RNA forms unique three-dimensional structures known as pseudoknots that are essential for viral replication and for the ability of the SARS-CoV-2 virus to cause disease. RNA pseudoknots may be a promising new target for therapeutic intervention. 

Working closely with the Molecular Screening Facility, which provides state-of-the-art technologies and industry standard expertise, Dr. Lieberman and his team rapidly set up a high-throughput, novel screening assay based on identifying molecules that bind to the SARS-CoV-2 pseudoknot structures and disrupt their function. His team is in the process of evaluating thousands of FDA-approved small molecule drugs. 

The lead candidates identified in the screen will then be further tested in cells and in preclinical models for their ability to stop infection and disease progression, and eventually to advance the best molecules into the clinic.

“We are taking a new, very focused approach to evaluate existing molecules,” said Dr. Lieberman. “This allows us to look for a very specific activity to deploy against the virus in molecules that have already tested safe in humans.”

Because RNA pseudoknot structures are similar in other coronaviruses, the outcome of this project could also have broader applications against other respiratory diseases.