The Academy: May 26, 2010

The Academy

Academic research is fueling tremendous strides in oncology and biotechnology. The Academy delivers the latest news on biotech and oncology research, providing a link between the clinical world of cancer care and the university researchers who are pushing the envelope of knowledge and discovery.

Emory University

Three-Dimensional Technology Applied to Breast Cancer Screening

Scientists at Emory University, Atlanta, are using new technology that is being compared with a View-Master (a classic children’s toy) to check for breast cancer in women with breasts too dense for today’s mammograms to give a clear picture.

The research is being watched closely because the need for better breast cancer detection is so great; roughly half of women younger than 50 and a third of women over 50 are estimated to have dense breasts—generally acknowledged to be one of the Achilles heel’s of successful mammography-based detection. In addition to making detection more challenging, women with dense breasts are also at higher risk of getting breast cancer.

The reason that dense breasts present such a challenge to traditional mammography is a matter of perspective. Mammograms are two-dimensional, flat pictures of a surface that is simply not flat. When technicians squeeze women’s breasts into the mammography unit, they are trying to spread the tissue out so less is hidden from the X-ray machine. The so-called ‘stereo mammograms’ that Emory researchers are currently experimenting with allow radiologists to see those X-ray images in three dimensions (3-D), so that a small spot on the bottom might not be hidden by normal tissue laying over it.

“We have depth perception because each eye gets a slightly different view, allowing your brain to construct a 3-D view when it overlays the two,” explained Mary S. Newell, MD, assistant breast-imaging chief, Emory University. That is the concept behind stereoscopes, gadgets that help people see pictures in 3-D like the old cartoons of a View-Master. Stereo mammograms, which are being developed by Cambridge, Massachusetts—based BBN Technologies, work essentially the same way. Separate X-rays are taken at slightly different angles. Then radiologists wear special glasses that make each eye see a separate image on special monitors. The brain ‘reads’ that as a single, 3-D view.

In a soon-to-be-published study, Emory radiologists gave nearly 1,500 women at increased risk of breast cancer both a mammogram and a stereo mammogram. Different radiologists analyzed each test. When researchers put together the results, the stereo mammograms increased detection of cancer by 23%. In addition to detecting tumors that mammograms missed, the stereo technology was able to more accurately map and better distinguish areas that traditional mammograms tend to classify as cancerous.

According to the Emory study, the stereo mammography technology decreased the incidence of false-positives by 46% when compared with traditional mammograms. The decrease in false alarms has the potential to eliminate a substantial amount of both patient stress and medical resource utilization.

According to Dr. Newell, the research demonstrates that “we can do better than we’re doing.”

M. D. Anderson Cancer Center

Potential Treatment-Resistant Ovarian Cancer Target Identified

Scientists from the University of Texas M. D. Anderson Cancer Center have discovered that overexpression of tissue-type transglutaminase (TG2) in ovarian cancer is associated with increased tumor cell growth and adhesion, resistance to chemotherapy and lower overall survival rates. When researchers targeted and silenced TG2 in animal models, cancer progression was reversed, suggesting the protein may also provide a novel therapeutic approach for late-stage ovarian cancer.

A significant need for new therapeutic approaches to late-stage ovarian cancer exists. The American Cancer Society estimates that more than 70% of women with ovarian cancer will suffer a recurrence and perish as a result of the disease.

Cancer Research

The M.D. Anderson research team, whose findings appear in the most recent issue of , was led by Anil K. Sood, MD, professor in the Departments of Gynecologic Oncology and Cancer Biology, and Kapil Mehta, PhD, professor in the Department of Experimental Therapeutics. According to the investigators, the effort represents one of the first explorations of TG2’s functionality in ovarian cancer.

The study, which examined 93 ovarian cancer samples of ranging stages, found that high levels of TG2 corresponded with significantly lower patient survival than those with low levels of TG2. Sixty-nine percent of high-stage ovarian cancers overexpressed TG2 compared with 30% of lowstage cancers. In-depth analysis demonstrated that tumors which overexpressed the protein tended to have an increased ability to invade healthy tissue and to survive or avoid the affects of chemotherapy.

According to the researchers, once it became clear that TG2 activated the survival pathway p13K/Akt in these tumors (explaining the resistant behavior of the tumors), the next step was to silence TG2. Researchers shut off TG2 with a small interfering RNA strand (TG2 siRNA) targeted to the protein, reducing the ability of the tumor cells to invade and killing them through programmed cell death (apoptosis). When exposed to the targeted intervention, ovarian cancer cells greatly reduced proliferation and blood vessel development, while increasing apoptosis.

Studies of chemotherapy-sensitive and chemotherapy-resistant models in mice showed considerable antitumor activity both with TG2 siRNA alone and in combination with docetaxel chemotherapy. The combination therapy of TG2 siRNA with docetaxel reduced tumor weight by 86%, proving to have the greatest efficacy compared with control groups or those without chemotherapy.

“TG2 appears to fuel different types of cancer through multiple molecular pathways, making it an important therapeutic target,” stated Dr. Mehta, whose lab also has connected TG2 overexpression to drugresistant and metastatic melanoma, breast cancer and pancreatic cancer.

“Drug resistance and metastasis are major impediments to the successful treatment of ovarian cancer and until now we had little information about the role TG2 played in ovarian cancer. We began to see its story unfold as we translated these data from tissue samples to cell lines to animal models,” added Dr. Sood.

In the near future, Dr. Sood and Dr. Mehta will be moving TG2 siRNA toward phase I clinical trials for ovarian and pancreatic cancers.

University of North Carolina

Study Provides Clues to Cancer Questions

How can cancer cells from one organ, such as the skin, travel and take root in a totally different organ, like the lung? Why do certain cancers tend to metastasize to certain places (i.e., prostate cancer usually moves to bone; colon cancer tends to migrate to the liver)?

In an effort to answer these questions, Hendrik van Deventer, MD, assistant professor of medicine, University of North Carolina (UNC) at Chapel Hill, and a member of the UNC Lineberger Comprehensive Cancer Center, and colleagues have dusted off an over 100- year-old idea of how cancer spreads: 19th Century English surgeon Stephen Paget’s so-called ‘seed and soil’ theory. Dr. Paget asserted that the spread of cancer is not just about the tumor itself (the seed), but also the environment where it grows (the soil). Dr. Van Deventer and his colleagues reaffirmed Dr. Paget’s teachings, then drilled down (in ‘the soil,’ as it were) to find further explanations.

The American Journal of Pathology

In short, the UNC team wanted to determine what mysterious non-tumor cell could change a normal organ so cancer cells would invade. The idea behind the research was that if scientists could discover the identity of that normal cell, maybe they could devise treatments to stop metastases. In a study published in the most recent issue of , UNC investigators showed for the first time that the aforementioned cell could also be a fibrocyte. Fibrocytes are cells that travel around the body, rushing to the site of an injury to aid in healing when needed. The study also suggests ways to develop treatments to prevent metastases using already available medications.

“This study shows it’s possible for fibrocytes to form the premetastatic niche,” Dr. van Deventer asserted. In healthy humans, fibrocytes travel through the bloodstream to areas of injury. Once there, they produce changes that are good for wounds. Unfortunately, these same changes can help cancers grow.

The UNC research team’s work with fibrocytes began when they wanted to figure out why ‘knockout mice’ that are missing the cell receptor CCR5 get fewer cancer metastases than normal mice. CCR5 helps control the migration of cells through the body. Dr. van Deventer and colleagues injected these knockout mice with all types of cells from normal mice to try to make the mice form more metastases caused by melanoma. The only cells that did so were those that appeared to be fibrocytes.

When Dr. van Deventer injected the mice with just 60,000 of these cells, the rate of metastases nearly doubled. “That’s a big effect for a relatively small number of cells,” stated Dr. van Deventer. The experiment also showed that injection of the cells induced MMP9, an enzyme that is known to promote cancer. The researchers considered this good news, since drugs are available that block MMP enzymes and have proven beneficial in treating cancer.

Stanford University

Molecule that Kills Kidney Cancer Cells Is Found

A new molecule found by Stanford University School of Medicine researchers kills kidney cancer cells. Ideally, the researchers said, a drug created from this molecule would help fight the life-threatening disease while leaving patients’ kidneys intact.

Cancer Cell

“You now have a potential means of going after a disease that’s been difficult to treat,” said Amato Giaccia, PhD, professor and director of radiation oncology and radiation biology at Stanford Medical School. His findings were recently published in the journal .

Dr. Giaccia said his lab focused on renal cell carcinoma, or kidney cancer, because there is no known cure for it except for removing a damaged kidney from a patient’s body. Dr. Giaccia’s work focused on the von Hippel- Lindau (VHL) tumor suppressor gene, which normally slows tumor growth in humans but does not work in 75% of kidney tumor cells.

Dr. Giaccia’s team searched for a small molecule that would kill cancer cells when this VHL gene is broken. They found their weapon in a molecule called STF-62247. While STF- 62247 is toxic to kidney cancer, it is generally harmless to most other cells in the human body because they carry a working VHL gene. As an added benefit, patients treated with STF-62247 should not suffer some of chemotherapy’s infamous side effects, like nausea and hair loss, because STF-62247 is not toxic to the entire body. “Clinical trials could begin in the next couple years,” Dr. Giaccia predicted.

Study co-author and postdoctoral fellow Denise A. Chan, PhD, said she believed the new findings could affect how all types of cancer are treated in the future. The study is one of the first to identify a trait unique to a certain form of cancer—in this case, kidney cancer’s deficient VHL gene—and exploit it to defeat the disease, Dr. Chan said. She predicted other scientists soon would follow suit, looking for characteristics in other cancers that could be manipulated.

Researchers’ motivation could be twofold, the study’s authors said: (1) to find cures for deadly cancers, and (2) to rein in the debilitating adverse effects caused by many current cancer treatments. “These results can be extended far beyond kidney cancer,” Dr. Chan said.

The findings also speak well for Stanford’s High-Throughput BioScience Center, which opened in 2004. The results of this study are some of the first to use the center’s equipment. The high-throughput equipment at Stanford can analyze thousands of molecules for their cytotoxicity at the same time, allowing researchers like those in Dr. Giaccia’s lab to search for hidden genes and molecules that previously would have been extremely labor intensive to find. In fact, Dr. Chan asserted that the study would not have been possible without the BioScience Center facilities.

Harvard Medical School

The Ins and Outs of Entosis

Cell Death Process Explains ‘Bird’s-Eye Cells’

Although pathologists have observed for over a century a peculiar cytological feature of human cancers (cells internalized in other cells) they remained puzzled as to how such so-called “bird’s-eye cells” formed in the first place. That is, until Harvard Medical School researchers in the laboratory of Joan Brugge, PhD, professor, department of cell biology, Harvard Medical School stumbled over the cell death process that explains the cellin-cell phenomenon. The discovery of this novel, nonapoptotic type of programmed cell death illustrates the serendipity of scientific research.

A major focus of Dr. Brugge’s laboratory is exploring mechanisms of cell survival relevant to breast cancer. In one research approach, investigators detach normal breast epithelial cells from their niches in the extracellular matrix (ECM), and then observe how the displaced cells die.

As postdoctoral researcher Michael Overholtzer, PhD, was watching these cells drift in suspension, he began to notice that some became completely encased in their neighbors. When he happened to mention this curious finding to pathologist Andrea Richardson, MD, PhD, she told him that pathologists have witnessed this phenomenon in cancer cells for decades.

Intrigued by the cancer link, Dr. Overholtzer conducted additional experiments using normal breast epithelial cells, MCF10As, and cancerous MCF7 cells. Through time-lapse microscopy, he found that about 25% of MCF10A cells and 30% of MCF7 cells were forming into bird’s-eye structures. Moreover, unlike apoptotic cells that become engulfed by adjacent cells, matrix-deprived cells appeared to be aggressively pushing their way into neighboring cells—like refugees seeking temporary shelter.

Not all these internalized cells are fated to die. Dr. Overholtzer observed that host cells formed capsules, or vacuoles, around the invaders, killing most of them through acidification and degradation by lysosomes; but a few escaped the host cell and slipped back into the culture. While internalized, these cells appeared to be very much alive—moving about and even dividing into daughter cells—clearly demonstrating their viability and distinguishing this type of cell death from apoptosis or necrosis. “Pathologists have speculated for years that some internalized cells are alive,” says Dr. Overholtzer, “and our data suggest they were right.”

Cell

Through this work, says Dr. Brugge, “we have elucidated the death program that underlies the formation of cell-in-cell structures and is provoked by the cell’s loss of attachment to the extracellular matrix, which occurs independent of apoptosis.” Researchers named the novel process entosis, from a Greek word meaning within or inside, and published their findings in the November 30 issue of . “What is incredibly exciting is that we have also found live cell-in-cell invasion in fluid exudates from metastatic tumors and in primary tumor tissue from breast cancer patients,” added Dr. Brugge.

However, the reasons why entosis occurs, what it reveals about cancer, and whether it happens in vivo remain unclear. “Our first instinct is that entosis inhibits tumor progression by killing ‘homeless’ cancer cells before they can colonize distant sites,” says Dr. Overholtzer. “One could also imagine that entosis promotes tumor progression by providing nutrients for some cells from their neighbors.” Clearly, asserted the investigators, more research is needed to more narrowly and accurately define the role of entosis in tumor growth.

Mayo Clinic Cancer Center

Investigators Discover Liver Cancer Treatment Target

Hepatology

The Mayo Clinic Cancer Center, Rochester, Minnesota, in collaboration with the National Cancer Institute, Bethesda, Maryland, reported in a recent issue of that the protein sulfatase 2 (SULF2) may provide one of the keys needed to begin the design of new therapies.

According to a Mayo Clinic Cancer Center spokesperson, the institution currently leads the field in researching the impact and effect of SULF1, a protein whose normal role is to degrade heparin sulfate proteoglycans—molecules that are part sugar and part protein. Mayo scientists have found that the protein also helps inhibit tumor growth. Now Mayo researchers are studying a related gene, SULF2. The role of the SULF2 gene and protein has not been fully defined, but in this study, researchers investigated the effect of SULF2 on liver tumor growth in the laboratory. They found that increased expression of SULF2 enhances cancer cell growth and migration, whereas decreased expression reduces both.

The researchers sought answers by examining a protein related to one they already knew had a role in suppressing liver tumors. SULF1 and SULF2 are similar proteins, but they cause opposing results. SULF1 removes sulfate groups that allow growth factors to bind to cells, thus inhibiting growth. The investigators found that SULF2 did the opposite. It increased binding of a specific growth factor—fibroblast growth factor 2 (FGF2)—to tumor cells, and also increased expression of the heparin sulfate proteoglycan glypican 3 (GPC3), which plays an important role in cell division and growth.

Findings indicated that expression of SULF2 was increased in 79 of 139 (57%) hepatocellular carcinomas (HCCs) and 8 of 11 (73%) HCC cell lines. Forced expression of SULF2 increased HCC cell growth and migration, whereas knockdown of SULF2 using short hairpin RNA targeting SULF2 abrogated HCC cell proliferation and migration in vitro. The effects of SULF2 on up-regulation of GPC3 and tumor growth were confirmed in nude mouse xenografts. Moreover, HCC patients with increased SULF2 expression in resected HCC tissues had a worse prognosis and a higher rate of recurrence after surgery.

Researchers concluded that, in contrast to the tumor suppressor effect of SULF1, SULF2 has an oncogenic effect in HCC mediated in part through up-regulation of FGF signaling and GPC3 expression.

This discovery indicates if scientists can decrease the levels or activity of SULF2 in a tumor, they might be able to stop its development. Mayo researchers are exploring use of an agent that mimics heparin and inhibits SULF2. They are also examining whether preventing heparin sulfate synthesis would inhibit tumor growth.

“The liver is designed to excrete toxins, and its tumors are no exception. Our problem is that the tumors tend to excrete chemotherapeutic agents rather than be affected by them. So we are looking for ways to get around that,” explained the study’s primary investigator, Mayo Clinic gastroenterologist Lewis Roberts, MD, PhD.

“If something has a very broad effect on signaling by growth factors, it may lead to an effective treatment. SULF2 has a number of characteristics that make it an attractive target, such as the fact that it is widely present in tumors. We are exploring a number of options with SULF2 as a focal point for treatment not only in liver cancer, but also in head and neck, pancreas, breast and other types of cancer,” stated Jinping Lai, MD, PhD, a Mayo oncology researcher and the lead author of the study.

The researchers hope to identify drugs that block SULF2, and they seek to thoroughly understand the mechanisms involved, including determining other growth signaling pathways affected by SULF2.

UCLA’s Jonsson Cancer Center

Common Cold Virus Used to Locate Cancer Cells

Using an engineered common cold virus, UCLA researchers delivered a genetic payload to prostate cancer cells that allowed them, using positron emission tomography (PET) scanning, to locate the diseased cells as they spread to the lymph nodes, the first place prostate cancer goes before invading other organs.

The tiny cancer metastases in the pelvic lymph nodes are very difficult to find using conventional imaging tools such as CT scanning. This discovery could aid oncologists in finding the cancer’s spread earlier, when it’s more treatable, and before it invades distant organs.

The next step for Lily Wu, MD, a researcher at UCLA’s Jonsson Cancer Center and the senior author of the study, and her colleagues is to link the noninvasive imaging advance with a treatment component, activating a toxic agent in the genetic payload to kill the spreading cancer cells. Dr. Wu hopes one day to be able to find tiny prostate cancer metastases in patients and kill them at the same time, watching it all on a PET scanner. She currently is refining this image-guided therapy in her lab in mouse models.

“I think this is very exciting for many reasons,” said Dr. Wu, who also is an associate professor of pharmacology and urology. “We now know we can reach these prostate cancer metastases at an earlier stage than before, and we know we can deliver genes to those cancer cells that produce proteins that can be imaged by PET. Now we will find out how effective this genetic toxic payload is in preventing further spread of the cancer to other vital organs.”

The spread of prostate cancer to the pelvic lymph nodes is the most reliable indicator that the patient will have a poor prognosis, with disease recurrence and progression likely. “Accurately assessing pelvic lymph node involvement in patients is critical in planning their treatment,” stated Dr. Wu.

Currently, physicians do not know if a treatment is attacking cancer cells until, using traditional imaging, they see a decrease in tumor size. This insensitive approach can take weeks and months. If the treatment is not working, the patient is exposed to a toxic therapy that will not help them. If Dr. Wu is successful, an oncologist would know within days if the cancer has spread and whether the treatment is killing the cancer.

Using mouse models, Dr. Wu and her team engineered a virus to travel to the lymph nodes, using a prostate cancer—specific vector that dictates its protein payload be expressed only in prostate cells. The payload in this case is a protein that can be imaged by PET scanning. The virus was introduced into the tumor in the mouse, and Dr. Wu and her team were able to detect PET signals only in the lymph nodes with cancer cell involvement, indicating the virus reached and infected the prostate cancer cells and produced the imaging protein.

As part of this study, Dr. Wu co-developed TSTA, a two-step transcriptional amplification method, which increased the expression of the genetic payload inside the cancer cells—in effect boosting the imaging signals and potential killing activity of the engineered virus.

Dr. Wu believes this type of image-guided therapy has the potential to improve the way advanced prostate cancer is treated.

“It would represent a treatment advance in patients for whom the outcome is not good. This would help improve the prognosis for these patients by letting us find and treat these metastases early. If we can catch the cancer before it invades other organs, we have a better chance to change the outcomes for these patients,” explained Dr. Wu.

Nature Medicine

The study appeared July 11, 2008, in the early, on-line edition of the peer-reviewed journal .