Can the patterns in tree branches or the meandering bends in a river provide clues that could lead to better cancer therapies? According to a new study from Virginia Commonwealth University Massey Cancer Center, these self-similar, repeating patterns in nature known as fractals help scientists better understand how the immune system is organized and may one day be used to help improve stem cell transplant outcomes in leukemia patients by predicting the probability of transplant complications.

Recently published in the journal Biology of Blood and Marrow Transplantation, the study led by Amir Toor, M.D., found a fractal pattern in the T cell repertoire of 10 unrelated stem cell transplant donors and recipients. T cells are a family of immune system cells that keep the body healthy by identifying and launching attacks against pathogens such as bacteria, viruses or cancer. T cells have small receptors that recognize antigens, which are proteins on the surface of foreign cells. Once T cells encounter a foreign cell, the antigen fits into the T cell’s receptor like a key in a lock and the T cell’s deadly arsenal is unleashed on the threat. Once activated, T cells divide into many clones with receptors designed to recognize and guard against that specific pathogen. Over the course of a person’s life, he will develop millions of these clonal families, which make up his T cell repertoire and protect him against the many threats that exist in his unique environment.

“The technological advancements of high throughput sequencing have only recently allowed scientists to sequence the genetic material responsible for T cell repertoire. At first glance, the data looks like a chaotic jumble of information,” says Toor, a hematologist in the Bone Marrow Transplant Program and researcher in the Developmental Therapeutics program at VCU Massey Cancer Center. “However, if you study a person’s T cell repertoire by analyzing the DNA segments responsible for the various types of T cell receptors, you begin to notice a fractal pattern based on segment usage.” Toor and his team are hopeful that this information will give them clues that will help them better understand the recovery of immune function following stem cell transplantation and possibly predict complications such as graft-versus-host disease in transplant recipients.

Much like a child can assemble Lego blocks to create a range of different models, humans have evolved a highly efficient process by which a short span of the genome called the T cell receptor locus rearranges gene fragments to create a multitude of different T cell receptor families. In this process, DNA segments known as variable (V), diversity (D) and joining (J) segments are rearranged to create the millions of T cell receptor families, or clones, that the body uses to combat disease. Similar to how the branching pattern of a tree is faithfully replicated from the trunk all the way to its farthest branches, T cells have families that are created from DNA segments branching out from one another to form a shield that provides protection from diseases.

Toor’s team looked at the frequency of T cell clones bearing different V, D and J segments in stem cell transplant donors and recipients following stem cell transplantation. Using a circular diagram designed by researcher Jeremy Meier, B.S., to better visualize the arrangement of the different DNA segments, the team observed a similar fractal order in the T cell receptor families of the donors. This order was even apparent in donors of different ethnicities living on different continents. In patients who had received a stem cell transplant, Toor found that this pattern was disrupted and the patients displayed a lower level of complexity in their T cell receptor repertoire at three months after transplant, followed by a modest improvement when a full year had elapsed after transplantation.

“Attempting to restore the fractal order of a patient’s T cell receptor repertoire by optimizing the stem cell transplant process could serve as a valuable therapeutic target,” says Toor. “Additionally, our findings lend an insight into nature, such that even in complex biological systems bereft of physical form, mathematically determined organization is observed.”

Toor and his colleagues plan to continue using high throughput sequencing of patients’ T cell receptors to learn more about how the immune system recovers following stem cell transplantation. The team hopes this will give them valuable information about the effectiveness of future stem cell transplant and immunotherapy clinical trials developed in their clinic.

In addition to Meier, Toor collaborated on this research with Catherine Roberts, Ph.D., and Allison Hazlett, M.S., both from the Bone Marrow Transplant Program at VCU Massey Cancer Center; Masoud Manjili, D.V.M., Ph.D., VCU Massey researcher in the Cancer Cell Signaling program, and Kassi Avent, Jennifer Berrie and Kyle Payne, all from the Department of Microbiology and Immunology at VCU School of Medicine; Kevin Hogan, Ph.D., senior scientific writer at VCU Massey; Kellie Archer, Ph.D., director of the Biostatistics Shared Research Core at VCU Massey; and Catherine Sanders, Ph.D., Cindy Desmarais, Ph.D., and David Hamm, M.Sc., from Adaptive Technologies.

The full manuscript of this study is available at: http://www.sciencedirect.com/science/article/pii/S1083879112011408.

Preclinical, laboratory studies suggest a novel immunotherapy could potentially work like a vaccine against metastatic cancers, according to scientists at Virginia Commonwealth University Massey Cancer Center. Results from a recent study show the therapy could treat metastatic cancers and be used in combination with current cancer therapies while helping to prevent the development of new metastatic tumors and train specialized immune system cells to guard against cancer relapse.

Recently published in the journal Cancer Research, the study detailed the effects of a molecule engineered by lead author Xiang-Yang Wang, Ph.D., on animal and cell models of melanoma, prostate and colon tumors. The molecule called Flagrp-170 consists of two distinct proteins, glucose-regulated protein 170 (Grp170), known as a “molecular chaperone,” and a “danger signal” derived from flagellin, a protein commonly found in bacteria. The researchers used modified viruses, or adenoviruses, that can no longer replicate to transport Flagrp-170 directly to the tumor site to achieve localized vaccination. The novel therapy caused a profound immune response that significantly prolonged survival in animal models.

“Successfully promoting antitumor immunity will help eradicate tumor cells, control cancer progression and help prevent tumor relapse,” says Wang, Harrison Scholar, member of the Cancer Molecular Genetics research program at VCU Massey Cancer Center and associate professor of Human and Molecular Genetics at VCU School of Medicine. “This immunotherapy has the potential to be used alone or in combination with conventional cancer treatments to develop and establish immune protection against cancer and its metastases.”

Grp170 is currently being explored for its potential as a “cancer vaccine” because it has been shown to help the immune system recognize cancer antigens. Antigens are molecules from foreign objects such as bacteria, viruses or cancer that, when detected, provoke an immune response aimed at attacking them. However, because cancer cells can alter the microenvironment surrounding a tumor, they are able to suppress immune responses and continue replicating without being attacked by the body’s natural defenses.

The chimeric chaperone Flagrp-170, created by strategically fusing a fragment of flagellin to Grp170, not only enhances antigen presentation, it also stimulates additional immune signals essential for functional activation of specialized immune cells, including dendritic cells, CD8+ T lymphocytes and natural killer (NK) cells. Dendritic cells act as messengers between the innate and adaptive immune systems. Once activated in response to a stimulus such as Flagrp-170, dendritic cells migrate to lymph nodes where they interact with other immune cells such as T lymphocytes to shape the body’s immune response. CD8+ T lymphocytes and NK cells are known to respond to tumor formation and kill cancer cells by triggering apoptosis, a form of cell suicide.

“Overcoming cancer’s ability to suppress the body’s natural immune responses and restore or develop immunity for tumor eradication is the goal of cancer immunotherapy,” says Wang. “More experiments are needed, but we are hoping Flagrp-170 may one day be used in formulating more effective therapeutic cancer vaccines.”

Moving forward, Wang and his team are working to better understand the molecular mechanisms responsible for Flagrp-170’s therapeutic effects. Additional studies are underway to more efficiently target and deliver Flagrp-170 to tumor sites in order to provoke a more robust and durable immune response.

Wang collaborated on this research with Paul Fisher, M.Ph., Ph.D., Thelma Newmeyer Corman Endowed Chair in Cancer Research and program co-leader of Cancer Molecular Genetics at VCU Massey Cancer Center, chairman of VCU’s Department of Human and Molecular Genetics and director of the VCU Institute of Molecular Medicine; Xiaofei Yu; Chunquing Guo, Ph.D.; Huanfa Yi; and Jie Qian, Ph.D., all from VCU’s Department of Human and Molecular Genetics and the VCU Institute of Molecular Medicine; and John R. Subjeck from the Department of Cellular Stress Biology at Roswell Park Cancer Institute. This research was supported by NIH grants CA129111 and CA154708; the American Cancer Society, the Department of Defense and, in part, by funding from VCU Massey Cancer Center’s NIH-NCI Cancer Center Support Grant P30 CA016059.

The full manuscript of this study is available online at: http://cancerres.aacrjournals.org/content/early/2013/01/18/0008-5472.CAN-12-1740.long.

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Laboratory experiments conducted by scientists at Virginia Commonwealth University Massey Cancer Center suggest that a novel combination of the drugs ibrutinib and bortezomib could potentially be an effective new therapy for several forms of blood cancer, including diffuse large B-cell lymphoma (DLBCL) and mantle cell lymphoma (MCL).

The study, published in the British Journal of Hematology, showed that the experimental drug combination killed cancer cells through a form of cell suicide known as apoptosis, but was relatively non-toxic to normal, healthy cells. Ibrutinib is a new agent that inhibits the B-cell receptor (BCR) signaling complex, which plays an important role in the survival of malignant B-cells. It has shown very promising initial results in the treatment of patients with B-cell malignancies, including chronic lymphocytic leukemia (CLL), DLBCL and MCL. The synergistic interaction of the two drugs proved lethal even to lymphoma cells that had become resistant to bortezomib, when used alone.

“Bortezomib is currently used to treat MCL and multiple myeloma, but, unfortunately, many patients develop resistance to the drug,” says the study’s principle investigator Steven Grant, M.D., Shirley Carter Olsson and Sture Gordon Olsson Chair in Oncology Research, associate director for translational research, program co-leader of Developmental Therapeutics and Cancer Cell Signaling research member at VCU Massey Cancer Center. “We are hopeful that this combination therapy may circumvent such resistance and eventually help fill an urgent need for more effective therapies for patients with these uncommon blood disorders.”

With cultured DLBCL and MCL cells in laboratory experiments spearheaded by Girija Dasmahapatra, Ph.D., lead author of the study’s manuscript and instructor in the Department of Internal Medicine at VCU School of Medicine, the scientists found that ibrutinib blocked several molecular pathways that the cancer cells use for growth and survival. When ibrutinib was combined with bortezomib, the scientists observed a high level of synergism between the two drugs that resulted in profound cell death due to DNA damage, culminating in apoptosis. The research findings suggest that the effectiveness of the combination therapy against bortezomib-resistant lymphoma cells may stem from ibrutinib’s ability to block signaling pathways used by the cancer cells to survive bortezomib exposure.

Specifically, exposure of DLBCL and MCL cells to ibrutinib blocked the cancer-promoting NF-κB, AKT and ERK1/2 signaling pathways. These signaling pathways provide cells with the ability to adapt to otherwise harmful environmental stimuli by transmitting messages from receptors located at the cell’s surface to proteins within the cell that trigger a variety of biological processes. In particular, NF-κB, AKT and ERK1/2 have been shown to carry out many functions that allow cancer cells to survive and proliferate. Significantly, each of these pathways has been implicated in the development of resistance to proteasome inhibitors such as bortezomib.

“We have provided a framework for understanding how an agent like ibrutinib might be employed to enhance the activity of an established anti-cancer agent like bortezomib,” says Grant. “We are currently working with representatives from the pharmaceutical industry and the National Cancer Institute to develop a new treatment strategy in which ibrutinib will be combined with proteasome inhibitors like bortezomib for the treatment of patients with lymphomas and potentially other blood cancers.”

Grant and Dasmahapatra collaborated on this study with Hiral Patel and Tri Nguyen, Ph.D., from the Department of Internal Medicine at VCU School of Medicine; Paul Dent, Ph.D., Universal Corporation Distinguished Professor for Cancer Cell Signaling, vice chair of the department of neurosurgery and member of the Developmental Therapeutics research program at VCU Massey; and Richard I. Fisher, M.D., and Jonathan Friedberg, M.D., from the James T. Wilmot Cancer Center at the University of Rochester.

This research was supported by National Institutes of Health grants CA63753, CA93738 and CA100866; Lymphoma SPORE award 1P50 CA130805; award R6059-06 from the Leukemia and Lymphoma Society of America; the Multiple Myeloma Research Foundation; Myeloma Spore grant P50CA142509; the V Foundation; and, in part, by funding from VCU Massey Cancer Center’s NIH-NCI Cancer Center Support Grant P30 CA016059.

Follow this link to view the full manuscript of this study online.

After uncovering a mechanism that promotes chronic intestinal inflammation and the development of colorectal cancer, scientists from Virginia Commonwealth University Massey Cancer Center have found that fingolimod, a drug currently approved for the treatment of multiple sclerosis, could potentially eliminate or reduce the progression of colitis-associated cancer (CAC).

The study, published online in the journal Cancer Cell, was led by Sarah Spiegel, Ph.D., Mann T. and Sara D. Lowry Chair in Oncology, co-leader of the Cancer Cell Signaling program at VCU Massey Cancer Center and chair of the Biochemistry and Molecular Biology Department at the VCU School of Medicine. Spiegel’s team discovered that increased production of an enzyme known as sphingosine kinase 1 (SphK1) causes cells lining the intestine to produce more of a signaling molecule known as sphingosine-1-phosphate (S1P), which activates a variety of biological mechanisms that lead to chronic intestinal inflammation and the development and progression of CAC. The researchers then used animal models to demonstrate that the drug fingolimod decreased expression of SphK1 and S1P’s receptor, S1PR1, which subsequently interfered with the development and progression of CAC, even after tumors were established.

“Perhaps the most significant aspect of this study is the therapeutic potential of fingolimod in the treatment of colitis-associated cancer,” says Spiegel. “Since this drug is already approved for clinical use, we’re hoping to initiate a clinical trial to study its efficacy in patients with CAC in combination with approved therapies.”

Essentially, the researchers discovered a self-feeding loop that results in chronic intestinal inflammation and increases the progression of CAC. The team showed that increased production of SphK1 and S1P lead to sustained activation of NF-kB and Stat3, which are both proteins called transcription factors that control the way DNA is transcribed in a cell’s nucleus in order to respond to environmental stimuli. This increased activation of NF-kB and Stat3 led to an increased production of TNF-a and IL-6, which are small pro-inflammatory molecules secreted by immune system cells. The increased inflammation, in turn, led to increased production of SphK1 and S1P, which continued the malicious cycle.

This is the first time that SphK1 and S1P have been linked to NF-kB, Stat3, chronic inflammation and CAC.

“Because one of the consequences of inflammatory bowel diseases is an increased risk of developing colorectal cancer, the next step in our research is to examine blood samples from patients with irritable bowel syndrome and colitis-associated cancer to measure levels of S1P,” says Spiegel. “Colorectal cancer is one of the leading causes of cancer-related deaths, and we’re hopeful that this research will lead to more effective treatments.”

Spiegel collaborated on this study with Kazuaki Takabe, M.D., Ph.D., and Tomasz Kordula, Ph.D., both members of the Cancer Cell Signaling program at VCU Massey; and Jie Liang, Masayuki Nagahashi, M.D., Ph.D., Eugene Y. Kim, Ph.D., Kuzhuvelil B. Harikumar, Ph.D., Akimitsu Yamada, M.D., Wei-Ching Huang, Nitai C. Hait, Ph.D., Jeremy C. Allegood, Ph.D., Megan M. Price, Dorit Avni, Ph.D., and Sheldon Milstien, Ph.D.,  all from VCU Massey Cancer Center  and the Department of Biochemistry and Molecular Biology at VCU School of Medicine.

This study was supported by NIH grants R37GM043880, RO1CA61774, U19AIO77435, T32HL094290, P30CA16059 and K12HD055881, a Susan G. Komen for the Cure Research Foundation grant and National Institute of Neurological Disorders and Stroke core grant 5P30NS047463.

The full manuscript of this study is available online at: http://www.sciencedirect.com/science/article/pii/S1535610812004928

After recently announcing success in eliminating melanoma metastasis in laboratory experiments, scientists at Virginia Commonwealth University Massey Cancer Center have made another important discovery in understanding the process by which the gene mda-9/syntenin contributes to metastasis in melanoma (the spread of skin cancer) and possibly a variety of other cancers.

Published in the journal Cancer Research, the study demonstrated that mda-9/syntenin is a key regulator of angiogenesis, the process responsible for the formation of new blood vessels in tumors. Mda-9/syntenin was originally cloned in the laboratory of the study’s lead author Paul B. Fisher, M.Ph., Ph.D., Thelma Newmeyer Corman Endowed Chair in Cancer Research and program co-leader of Cancer Molecular Genetics at Virginia Commonwealth University Massey Cancer Center, chairman of VCU’s Department of Human and Molecular Genetics and director of the VCU Institute of Molecular Medicine.

“Our research brings us one step closer to understanding precisely how metastatic melanoma, a highly aggressive and therapy-resistant cancer, spreads throughout the body,” says Fisher. “Additionally, analysis of the human genome has indicated that mda-9/syntenin is elevated in the majority of cancers, which means novel drugs that target this gene could potentially be applicable to a broad spectrum of other deadly cancers.”

Fisher’s team discovered that mda-9/syntenin regulates the expression of several proteins responsible for promoting angiogenesis, including insulin growth factor binding protein-2 (IGFBP-2) and interleukin-8 (IL-8). The study is the first to provide proof of the pro-angiogenic functions of IGFBP-2 in human melanoma.

In in vivo and in vitro experiments, the scientists confirmed that mda-9/syntenin binds with the extracellular matrix (ECM) to start a series of biological processes that eventually cause endothelial cells to secrete IGFBP-2. The ECM is the substance that cells secrete and in which they are embedded. Endothelial cells are the cells that line the interior surface of blood vessels throughout the entire circulatory system. The secretion of  IGFBP-2, in turn, caused the endothelial cells to produce and secrete vascular endothelial growth factor-A (VEGF-A), a protein that mediates the development of and formation of new blood vessels.

The researchers also noted that IGFBP-2 could potentially serve as a novel biomarker to monitor for disease progression in melanoma patients.

“This is a major breakthrough in understanding angiogenesis and its impact in melanoma metastasis,” says Fisher. “We are now focusing on developing novel small molecules that specifically target mda-9/syntenin and IGFBP-2, which could be used as drugs to treat melanoma and potentially many other cancers.”

Fisher collaborated on this study with Devanand Sarkar, M.B.B.S., Ph.D., Harrison Scholar and research member of the Cancer Molecular Genetics program at VCU Massey, Blick Scholar and assistant professor in the Department of Human and Molecular Genetics and member of the VCU Institute of Molecular Medicine at VCU School of Medicine; Swadesh K. Das, Ph.D., Santanu Dasgupta, Ph.D., and Luni Emdad, M.B.B.S., Ph.D., from the Department of Human and Molecular Genetics at VCU School of Medicine, the VCU Institute of Molecular Medicine and the Cancer Molecular Genetics research program at VCU Massey;  Sujit K. Bhutia, Ph.D., Belal Azab, Ph.D., Upneet K. Sokhi, Timothy P. Kegelman, Leyla Peachy, Prasanna K. Santhekadur, Ph.D., and Rupesh Dash, Ph.D., all from the Department of Human and Molecular Genetics; Paul Dent, Ph.D., Universal Corporation Distinguished Professor for Cancer Cell Signaling and Developmental Therapeutics program member at VCU Massey, and professor and vice chair of research in the Department of Neurosurgery at VCU School of Medicine; Steven Grant, M.D., Shirley Carter Olsson and Sture Gordon Olsson Chair in Oncology Research, associate director for translational research, program co-leader of the Developmental Therapeutics program and Cancer Cell Signaling program member at VCU Massey; and Maurizio Pellacchia, Ph.D., from Sanford-Burnham Medical Research Institute.

This research was supported by National Institutes of Health grant CA097318, the Thelma Newmeyer Corman Endowment, the National Foundation for Cancer Research, the Goldhirsh Foundation for Brain Tumor Research, the Dana Foundation and, in part, by funding from VCU Massey Cancer Center’s NIH-NCI Cancer Center Support Grant P30 CA016059.

The full manuscript of this study is available online at: http://cancerres.aacrjournals.org/content/early/2012/12/08/0008-5472.CAN-12-1681.long

Approximately 68 percent of U.S. adults are overweight or obese, according to the National Cancer Institute, which puts them at greater risk for developing cancer, cardiovascular disease, diabetes and a host of other chronic illnesses.  But an international team of scientists led by Virginia Commonwealth University Massey Cancer Center researcher Andrew Larner, M.D., Ph.D., has successfully reversed obesity in mice by manipulating the production of an enzyme known as tyrosine-protein kinase-2 (Tyk2). In their experiments, the scientists discovered that Tyk2 helps regulate obesity in mice and humans through the differentiation of a type of fat tissue known as brown adipose tissue (BAT).

Published today in the online edition of the journal Cell Metabolism, the study is the first to provide evidence of the relationship between Tyk2 and BAT. Previous studies by Larner and his team discovered that Tyk2 helps suppress the growth and metastasis of breast cancer, and now the current study suggests this same enzyme could help protect against and even reverse obesity.

The scientists were able to reverse obesity in mice that do not express Tyk2 by expressing a protein known as signal transducer and activator of transcription-3 (Stat3). Stat3 mediates the expression of a variety of genes that regulate a host of cellular processes. The researchers found that Stat3 formed a complex with a protein known as PR domain containing 16 (PRDM16) to restore the development of BAT and decrease obesity.

“We discovered that Tyk2 levels in mice are regulated by diet. We then tested tissue samples from humans and found that levels of Tyk2 were more than 50 percent lower in obese humans,” said Larner, Martha Anne Hatcher Distinguished Professor in Oncology and co-leader of the Cancer Cell Signaling program at VCU Massey Cancer Center. “Our findings open new potential avenues for research and development of new pharmacological and nutritional treatments for obesity.”

There are two different types of fat, white adipose tissue (WAT) and BAT. WAT is the primary site of energy storage. BAT is responsible for energy expenditure in order to maintain body temperature. BAT deposits are present in all mammals, but until recently, scientists thought BAT was only active in infants and not in adult humans. Only in the last four years have scientists realized that BAT is present in adults and helps to regulate energy expenditure. Additionally, research has shown that diminished BAT activity is associated with metabolic syndrome, a combination of medical disorders that increase the risk of developing cardiovascular disease and diabetes. Researchers estimate metabolic syndrome could affect as much as 25 percent of the U.S. population.

“We have made some very interesting observations in this study, but there are many questions left unanswered,” said Larner. “We plan to further investigate the actions of Tyk2 and Stat3 in order to better understand the mechanisms involved in the development of brown adipose tissue. We’re hopeful this research will help lead to new targets to treat a variety of obesity-related diseases such as cancer, cardiovascular disease and diabetes.”

Larner collaborated on this study with Marta Derecka, Magdalena Morgan, Vidisha Raje, Jennifer Sisler and Quifang Zhang, all from the Department of Biochemistry and Molecular Biology at VCU School of Medicine; Karol Szczepanek from the Department of Internal Medicine at VCU School of Medicine; Tomasz Kordula, Ph.D., Cancer Cell Signaling program member at VCU Massey; Agnieszka Gornicka, from the Cleveland Clinic Foundation; Sergei B. Koralov, Ph.D., from New York University Medical School; Dennis Otero, Ph.D., from the University of California; Joanna Cichy, Ph.D., from Jagiellonian University in Krakow, Poland; Klaus Rajewsky, Ph.D., from Harvard Medical School; Kazuya Shimoda, M.D., Ph.D., from Miyazaki University in Japan; Valeria Poli, Ph.D., from the University of Turin in Torino, Italy; Brigit Strobl, Ph.D., from the University of Veterinary Medicine in Vienna, Austria; Sandra Pellegrini, Ph.D., from Institut Pasteur in Paris, France; Thurl E. Harris, Ph.D., and Susanna R. Keller, M.D., from University of Virginia School of Medicine; Patrick Seale, Ph.D., from the University of Pennsylvania School of Medicine; Aaron P. Russell, Ph.D., from Deakin University in Burwood, Australia; Andrew J. McAinch, Ph.D., from Victoria University in St. Albans, Australia; Paul E. O’Brien, M.D., from Monash University in Melbourne, Australia; and Colleen M. Croniger, Ph.D., from Case Western University School of Medicine.

This study was supported by National Institutes of Health grants R01 AI059710-01 and R21 AI088487, the Austrian Science Fund and, in part, by funding from VCU Massey Cancer Center’s NIH-NCI Cancer Center Support Grant P30 CA016059.

The full manuscript is available online: http://www.cell.com/cell-metabolism/abstract/S1550-4131(12)00458-5