Targeting EGFL6 Protein Halts Growth and Spread of Ovarian Cancer


Dr. Ronald J. Buckanovich, professor of hematology/oncology and gynecologic oncology at the University of Michigan Medical School, and his colleagues have identified a protein that help ovarian cancer cells multiply and spread to other organs.  When he and his coworkers inhibited this protein with an antibody they were able to stop the spread of ovarian cancer cold.

The EGFL6 or epidermal growth factor like 6 precursor protein, which is also known as MAEG, maps to human Xp22 chromosome.  The EGFL6 protein is expressed primarily in fetal tissues and during early development (see Yeung G., et al., (1999) Genomics 62, 304307; and Buchner G., et al., (2000) Genomics 65, 1623).  The expression of MAEG has also been detected in several tissues, including the dermis of the trunk, hair follicles, and the mesenchyme of the cranio-facial region (see Buchner G., and others, (2000) Mech. Dev. 98, 179182).  EGFL6 protein has been proposed as a possible biomarker in ovarian cancer (Buckanovich R. J., and others, (2007) J. Clin. Oncol. 25, 852861).

In this paper, which appeared in Cancer Research, Buckanovich and others amplified the expression of EGFL6 in ovarian cancer cells.  Increased EGFL6 expression stimulated cancer growth some two-three times.  This effect was observed in cultured ovarian cancer cells and in a mouse model of ovarian cancer.  Conversely, elimination of EGFL6 greatly reduced ovarian cancer growth, decreasing the rate of growth some four-fold.

EGFL6 specifically acts in cancer stem cells.  To review, in tumors, not all cancer cells are the same.  Inside malignant tumors or even among circulating cancerous cells (as in the case of leukemia) there are usually a variety of different types of cancer cells.  The stem cell theory of cancer proposes that among cancerous cells, a small population among them act as stem cells that reproduce themselves and sustain the cancer.  Cancer stem cells, therefore, are like normal stem cells that renew and sustain our organs and tissues.  Therefore, cancer cells that are not stem cells can certainly adversely affect health, but they cannot sustain the cancer long-term.  Therefore, cancer stem cells fuel the growth and spread of cancers and also are often resistant to chemotherapy and radiation treatments.

Further experiments by Buckanovich and his colleagues showed that EGFL6 cause cancer stem cells to divide asymmetrically so that the one of the daughter cells remains a cancer stem cell while the other daughter cell is a cancer cell that can affect the patient but cannot sustain the cancer. This asymmetric cell division also generates a good deal of diversity among cancer cells.

Buckanovich noted: “What this means is that the stem cell population remains stable.  But the daughter cells, which can have a burst of growth, multiply, and allow the cancer to grow.”.

EGFL6 does more than just promote cancer cell proliferation.  EGFL6 is also a promoter of cancer stem cell migration.  When blood vessels were engineered to express EGFL6, tumor metastasis (spread) was even more robust.

Treatment of tumor-afflicted mice with an antibody that specifically binds to EGFL6 and inactivates it caused a 35% reduction in cancer stem cells and significantly reduced overall tumor growth.  Additionally, the antibody also prevented tumor metastasis.

Buckanovich thinks that targeting EGFL6 might be a potential therapy for women with stage 3 ovarian cancer.  Such a treatment might control the growth and spread of ovarian cancers.

Dr. Buckanovich added: “The bigger implication is for women at high risk of ovarian cancer.  These patients could be treated before cancer develops, potentially blocking cancer from developing or preventing it from spreading.  If cancer did develop, it could be diagnosed at an early stage, which would improve patient outcomes.”.

The next step for Buckanovich and his colleagues is developing an antibody that can properly work in human cancer patients.

Vitamin A as a Treatment For Colon Cancer


A new study that was published in the journal Cancer Cell, has introduced a way to treat colon, which is not only a leading cause of cancer deaths worldwide, but is notoriously resistant to treatment.

This collaboration between Swiss and Japanese scientists has identified an anticancer mechanism that includes vitamin A that can be tapped to inhibit colon cancer.

Colon cancer patients are typically treated with chemotherapy, which kills off most of the cancer cells, but leaves a few resistant cells that then aggressively grow back to form another deadlier tumor that can readily spread throughout the body. Chemotherapy ultimately fails because there are a core of rouge stem cells that divide uncontrollably called cancer stem cells that drives the growth of the cancer. These cancer stem cells are what is driving the growth of the tumor and if these cancer stem cells are not eliminated, the tumor will simply come back after chemotherapy. When a colon cancer patient receives treatment such as chemotherapy, most of the cancer cells die off.

Joerg Huelsken, Ph.D., of Ecole Polytechnique Federale de Lausanne (EPFL) led his research team to understand how stem cells populations in the colon give rise to new colon cells to replace dead, dying or cells that have been sloughed off. In mice and in tissue samples from human patients, a protein called HOXA5 kept asserting itself. HOXA5 turns out to be an integral part of the machinery of the cell that ensures that the cells of the colon properly differentiate after they are born from colon stem cells.

Huelsken’s team showed that in the colon, HOXA5 helps restrict the number of stem cells. Cancerous stem cells, however, block the expression of HOXA5 and prevent it from restricting stem cell numbers. HOXA5 is part of a signaling pathway that activates aspects of the cellular machinery that negatively controls cell growth. By blocking expression of the HOXA5 gene, these cancerous stem cells in the colon can grow uncontrollably and spread, thereby causing relapses and metastasis or spread of the colon cancer.

Huelsken and his team, in collaboration with Japanese researchers from Kyoto University investigated ways to unblock the expression of HOXA5 in colon cancer stem cells. The answer came from an unexpected corner – vitamin A. Vitamin A is a member of the retinoid family of molecules and has been known for some time to be able to induce differentiation of skin-based stem cells. Huelsken’s group showed that retinoids like vitamin A can upregulate HOXA5 and antagonize the mechanism in colon cancer stem cells that staunch its expression.

In a mouse colon cancer model system, treatment with retinoids not only blocked progression of the tumors, but normalized the tissue. The activation of the expression of the HOXA5 gene eliminated cancer stem cells and prevented metastasis in live animals. These results were then faithfully recapitulated in samples from actual patients.

From this study, it seems that screening tumors for the absence of HOXA5 expression is a relatively easy way to determine if a patient’s colon cancer will respond to treatment with vitamin A. Treatment with vitamin A or other retinoids might not only prove effective against colon cancer, but also as a preventive measure in high-risk patients.

Pancreatic Cancer Stem Cells Could Be “Suffocated” by an Anti-Diabetic Drug


A new study by researchers from Queen Mary University of London’s Barts Cancer Institute and the Spanish National Cancer Research Centre (CNIO) in Madrid shows that pancreatic cancer stem cells (PancSCs) are very dependent on oxygen-based metabolism, and can be “suffocated” with a drug that is already in use to treat diabetes.

Cancer cells typically rely on glycolysis; a metabolic pathway that degrades glucose without using oxygen. However, it turns out that not all cancer cells are alike when it comes to their metabolism.

PancSCs use another metabolic process called oxidative phosphorylation or OXPHOS to completely oxidize glucose all the way to carbon dioxide and water. OXPHOS uses large quantities of oxygen and occurs in the mitochondria. However, this is precisely the process that is inhibited by the anti-diabetic drug, metformin.

Ever crafty, some PancSCs manage to adapt to such a treatment by varying their metabolism, and this leads to a recurrence of the cancer. However, this English/Spanish team thinks that they have discovered a way to prevent such resistance and compel all PancSCs to keep using OXPHOS. This new discovery might open the door to new treatments that stop cancer stem cells using oxygen and prevent cancers from returning after conventional treatments. A clinical trial is planned for later next year.

Dr Patricia Sancho, first author of the research paper, said: “We might be able to exploit this reliance on oxygen by targeting the stem cells with drugs that are already available, killing the cancer by cutting off its energy supply. In the long-term, this could mean that pancreatic cancer patients have more treatment options available to them, including a reduced risk of recurrence following surgery and other treatments.”

PancSCs become resistant to metformin by suppressing a protein called MYC and increasing the activity of a protein called PGC-1α. However, this resistance mechanism of PancSCs can be abolished if a drug called menadione is given. Menadione increases the amount of reactive oxygen species in mitochondria. Additionally, resistance to metformin can be prevented or even reversed if the MYC protein is inhibited by genetic or pharmacological means. Therefore, the specific metabolic features of pancreatic Cancer Stem Cells are amendable to therapeutic intervention and can provide the basis for developing more effective therapies to combat this lethal cancer.

Pancreatic cancer is still one of the most difficult cancer types to treat. It rarely causes symptoms early on and does not usually trigger diagnosis until its later and more advanced stages. Unfortunately, many patients do not live longer than a year after being diagnosed. These cancers are also becoming more common due to obesity, which increases the patient’s risk for metabolic syndrome and diabetes, which are pancreatic cancer risk factors. Limited treatment options and a failure to improve survival rates mean that finding new treatment strategies is a priority.

PancSCs could be an important but as yet overlooked piece of this puzzle, since they compose only a small proportion of the tumor. PancSCs also have the potential to make new tumors, even if all the other cells are killed, and are prone to spreading around the body (metastasis). Therefore killing these PancSCs is a better way to treat such dangerous cancers.

Two Genes Control Breast Cancer Stem Cell Proliferation and Tumor Properties


When mothers breastfeed their babies, they depend upon a unique interaction of genes and hormones to produce milk and deliver it to their hungry little tyke. Unfortunately, this same cocktail of genes and hormones can also lead to breast cancer, especially if the mother has her first pregnancy after age 30.

A medical research group at the Medical College of Georgia at Georgia Regents University has established that a gene called DNMT1 plays a central role in the maintenance of the breast (mammary gland) stem cells that enable normal rapid growth of the breasts during pregnancy. This same gene, however, can also maintain those cancer stem cells that enable breast cancer. According to their work, the DNMT1 gene is highly expressed in the most common types of breast cancer.

Also, another gene called ISL1, which encodes a protein that puts the brakes on the growth of breast stem cells, is nearly silent in the breasts during pregnancy and in breast cancer stem cells.

Dr. Muthusamy Thangaraju, a biochemist at the Medical College of Georgia, who is the corresponding author of this study, which was published in the journal Nature Communications, said, “DNMT1 directly regulates ISL1. If the DNMT1 expression is high, this ISL1 gene is low.” Thangaraju and his team first observed the connection between DNMT1 and ISL1 when they knocked out the DNMT1 gene mice and noted an increase in the expression of ISL1. These results inspired them to examine the relationship of these two genes in human breast cancer cells.

Thangaraju and his co-workers discovered that ISL1 is silent in most human breast cancers. Furthermore, they demonstrated that restoring higher levels of ISL1 to human breast cancer cells dramatically reduced cancer stem cell populations, the growth of these cells, and their ability to spread throughout the tissue; all of which are hallmarks of cancer.

Conversely, Thangaraju and his team knocked out the DNMT1 gene in a breast-cancer mouse model, the breast will not develop as well. However, according to Thangaraju, this same deletion will also prevent the formation of about 80 percent of breast tumors. In fact, DNMT1 also down-graded super-aggressive, triple-negative breast cancers, which are negative for the estrogen receptor (ER-), progesterone receptor (PR-), and HER2 (HER2-).

The findings from this work also point toward new therapeutic targets for breast cancer and new strategies to diagnose early breast cancer. For example, a blood test for ISL1 might provide a marker for the presence of early breast cancer. Additionally, the anti-seizure medication valproic acid is presently being used in combination with chemotherapy to treat breast cancer, and this drug increases the expression of ISL1. This might explain why valproic acid works for these patients, according to Thangaraju. Workers in Thangaraju’s laboratory are already screening other small molecules that might work as well or better than valproic acid.

Mammary stem cells maintain the breasts during puberty as well as pregnancy, which are both periods of dynamic breast cell growth. During pregnancy, breasts may generate 300 times more cells as they prepare for milk production. Unfortunately, these increased levels of cell growth might also include the production of tumor cells, and the mutations that lead to breast cancer increase in frequency with age. If the developing fetus dies before she comes to term, immature breast cells that were destined to become mature mammary gland cells can more easily become cancer, according to Rajneesh Pathania, a GRU graduate student who is the first author of this study.

DNMT1 is essential for maintaining a variety of stem cell types, such as hematopoietic stem cells, which produce all types of blood cells. However, the role of DNMT1 in the regulation of breast-specific stem cells that make mammary gland tissue and may enable breast cancer has not been studied to this point.

For reasons that unclear, there is an increased risk of breast cancer if the first pregnancy occurs after age 30 or if mothers lose their baby during pregnancy or have an abortion. Women who never have children also are at increased risk, but multiple term pregnancies further decrease the risk, according to data compiled and analyzed by the American Cancer Society.

Theories to explain these phenomena include the coupling of the hormone-induced maturation of breast cells that occurs during pregnancy with an increase in the potential to produce breast cancer stem cells. Most breast cancers thrive on estrogen and progesterone, which are both highly expressed during pregnancy and help fuel stem cell growth. During pregnancy, stem cells also divide extensively and as their population increases, DNMT1 levels also increase.

In five different types of human breast cancer, researchers found high levels of DNMT1 and ISL1 turned off. Even in a laboratory dish, when they reestablished normal expression levels of ISL1, human breast cancer cells and stem cell activity were much reduced, Thangaraju said.

Promising New Drug Attacks Cancer Stem Cells in Mesothelioma


The anticancer drug defactnib is currently the subject of clinical trials being conducted in multiple countries. Data from this trial in patients with a type of cancer called mesothelioma have shown that defactnib is potentially quite effective in the treatment of these cancers. Defactnib, which is being marketed by biopharmaceutical company Verastem, Inc.

Mesothelioma attacks the lining of the lung, abdomen and, in some cases, the heart. It has only one known cause; the ingestion or inhalation of asbestos fibers.

This trial is currently in its second phase, and it involves 180 patients in 13 countries. The goal of this trial is to evaluate the ability of Defactnib to kill cancer stem cells that are the main cause of the spread of the tumors and their recurrence. Even though the main focus of this trial has been on treating mesothelioma, positive results have also been achieved by using defactnib to treat other types of cancer as well.

In this trial, mesothelioma patients were treated with a combination of defactnib and pemetrexed for 12 days prior to undergoing surgery. Pemetrexed has been approved by the U.S. Food and Drug Administration to treat mesothelioma. Tumors shrunk in 70 percent of the treated mesothelioma patients. Defactnib is designed to inhibit the activity of a protein called the Focal Adhesion Kinase or FAK. FAK is very important for cancer stem cell function and without it, cancer stem cells cannot grow.

“These and other exciting developments continue to build belief that there may be an end to this horrible disease in sight,” said Russell Budd, president and managing shareholder of the mesothelioma law firm Baron and Budd. “It is extremely encouraging to see signs of progress occur on a regular basis.”

Stem Cells Lurk in Tumors and Can Resist Treatment


Regenerative medicine seeks to train stem cells to transform into nearly any kind of cell type. Unfortunately, this ability that makes stem cells so useful also is cause for concern in cancer treatments. Malignant tumors contain resident stem cells, which prompts worries among cancer experts that the cells’ transformative powers help cancers escape treatment.

Data from new research shows that the threat posed by cancer stem cells is more prevalent than previously thought. Until now, stem cells had been identified only in aggressive, fast-growing tumors. However, a mouse study at Washington University School of Medicine in St. Louis has revealed that slow-growing tumors also have treatment-resistant stem cells.

Brain tumor stem cells (orange) in mice express a stem cell marker (green). Researchers at Washington University School of Medicine in St. Louis are studying how cancer stem cells make tumors harder to kill and are looking for ways to eradicate these treatment-resistant cells. Credit: Yi-Hsien Chen
Brain tumor stem cells (orange) in mice express a stem cell marker (green). Researchers at Washington University School of Medicine in St. Louis are studying how cancer stem cells make tumors harder to kill and are looking for ways to eradicate these treatment-resistant cells. Credit: Yi-Hsien Chen

In mice, low-grade brain cancer stem cells were less sensitive to anticancer drugs. When compared to healthy stem cells, tumor-based stem cells from brain tumors, revealed the reasons behind their resistance to treatments, which points to new therapeutic strategies.

“At the very least, we’re going to have to use different drugs and different, likely higher dosages to make sure we kill these tumor stem cells,” said senior author David H. Gutmann, MD, PhD, the Donald O. Schnuck Family Professor of Neurology.  Their data were published in the March 12 edition of Cell Reports.

First author Yi-Hsien Chen, who is a senior postdoctoral research associate in Gutmann’s laboratory, used a mouse model of neurofibromatosis type 1 (NF1), which forms low-grade brain tumors, to identify cancer stem cells and demonstrate that they could form tumors when transplanted into normal, cancer-free mice.

Neurofibromatosis type I is caused by mutations in the NF1 genes, and such mutations affect about 1 in every 2,500 babies. Neurofibromatosis type I can cause an array of physical problems, including brain tumors, impaired vision, learning disabilities, behavioral problems, heart defects and bone deformities.

In children with NF1 mutations, the most common brain tumor is optic gliomas. Treatment for NF1-related optic gliomas usually includes drugs that inhibit a cell growth pathway originally identified by Gutmann. In laboratory tests conducted as part of the new research, it took 10 times the dosage of these drugs to kill the low-grade cancer stem cells.

Compared with healthy stem cells from the brain, cancer stem cells made multiple copies of a protein called Abcg1 that helps those cells survive stress.

“This protein blocks a signal from inside the cells that should make them more vulnerable to treatment,” Gutmann explained. “If we can identify a drug that disables this protein, it would make some cancer stem cells easier to kill.”

Even though these laboratory mice were bred to model NF1 optic gliomas, Gutmann and others said that their findings could be applied more broadly to other brain tumors.

“Because stem cells haven’t differentiated into specialized cells, they can easily activate genes to turn on new developmental programs that allow the cells to survive cancer treatments,” said Gutmann, who directs the Washington University Neurofibromatosis Center. “Based on these new findings, we will have to develop additional strategies to keep these tumors from evading our best treatments.”

Inhibiting Leukemic Stem Cells


Blood cancers, also known as leukemias, are, in many ways, a disease of stem cells. A core of cancerous stem cells divide and produce progeny that overpopulate, overwhelm, and in some cases invade and destroy other tissues. However many cancer treatments are designed to specifically attack the progeny of the cancer stem cells and not the leukemic stem cells. Therefore, the disease is destined to relapse, since the main driving entities of the leukemia are left to flourish while their progeny have been killed off.

Acute myeloid leukemia or AML is largely a disease of older adults, and even though the survival rates have increased, only about one out of every four adult patients survives for five years after the AML has been diagnosed. The mean survival time for this disease, which predominantly occurs in the elderly, is less than a year for patients over 65 years. Younger adults tend to do much better than older people. For example, more than half (50%) of the people under 45 diagnosed with AML will live for at least 5 years, and some will be cured. But in others, however, the AML will return and there is no way to tell in advance who has been cured and who will relapse. In people over 65 years of age the outlook tends not to be so good and around 12 out of 100 people (12%) are alive for more than 5 years.

Relapses and treatment failure in AML is almost certainly due to leukaemic stem cells, which cannot be completely eliminated during treatment. However, researcher Dr. Marin Ruthardt from the Hematology Department of the Medical Clinic II and Dr. Jessica Roos, Prof. Diester Steinhilber and Prof. Thorsten Jürgen Maier from the Institute for Pharmaceutical Chemistry has discovered a chink in the armor of leukemic stem cells. They report their data in the current edition of the journal “Cancer Research.”

AML stem cells require an enzyme called 5-lipoxygenase or 5-LO in order to survive. 5-LO plays a very important role in inflammatory diseases like asthma. Ruthardt, Roos, Steinhilber, and Maier and the members of their research teams showed that the leukemic stem cells in a subgroup of AML could be selectively and efficiently attacked by chemical inhibitors of 5-LO. These inhibitors killed the leukemic stem cells in cell culture model systems and in leukemia mouse models.

“These results provide the basis for the potential implementation of 5-LO-inhibitors as stem cell therapeutic agents for a sustained AML cure, although this must be investigated further in preclinical and clinical studies in humans,” explains Dr. Ruthardt.

Prof. Maier continued: “In addition, there are plans for further molecular biological studies with the objective of understanding exactly how the 5-LO inhibitors act on the leukemic cells.”

Hopefully these inhibitors can be fast-tracked for Phase I studies in human patients, and if they prove safe under clinical conditions, then maybe, if all seems well, they can be used to treat AML patients with aggressive cancers that do not respond well to more traditional cancer treatments.

Cancer Stem Cell Research Leads to Clinical Trials


Dennis Slamon and Zev Wainberg from the UCLA Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research have been awarded a Disease Team Therapy Development award to begin clinical trials in human patients early in 2014.

In this clinical trial, Slamon and Wainberg will test a new drug that targets cancer stem cells. This drug was developed by research and development over the last decade on the cancer stem cell hypothesis. The cancer stem cell hypothesis predicts that proliferating stem cells are the main drivers of tumor growth and are also resistant to standard cancer treatments.  This new drug, CFI-400945, has prevented cancer growth in an extensive series of laboratory animal tests.

An important extension of the cancer stem cell hypothesis is that cancer stem cells inhabit a particular niche that prevents anticancer drugs from reaching them. Alternatively, tumors become resistant to cancer drugs by a process called “cell fate decision,” in which some cancer stem cells are killed by chemotherapy, but other cells replace them and repopulate the tumor. This tumor repopulation is the main reason for cancer recurrence.

The new anticancer drug to be tested in this clinical trial targets the “polo-like kinase 4.” Inhibition of this enzyme effectively blocks cell fate decisions that cause cancer stem cell renewal and tumor cell growth. Thus inhibition of this enzyme effectively stops tumor growth.

This clinical trial will test this novel chemotherapeutic agent in patients to establish the safety of the drug. After these initial safety tests, the trial will quickly proceed to further clinical tests. “We are excited to continue to test this drug in humans for the first time,” said Wainberg. Slamon, Wainberg and others will also look for biological markers to determine how well their drug is working in each patient.

The US Food and Drug Administration approved the Investigational New Drug (IND) for this drug trial. Also, Health Canada, the Canadian government’s therapeutic regulatory agency, also approved this trial. These approvals are part of an international effort to bring leading-edge stem cell science to patients.

A Stem Cell-Based Therapy for Colon Cancer


Colorectal cancer is the third leading cause of death in the Western World. Like many other types of cancer, colorectal cancer spreads and is propagated by cancer stem cells. Therefore, understanding how to inhibit the growth of cancer stem cells provides a key to treating the cancer itself.

By inactivating a gene that drives stem cell renewal in cancer stem cells, scientists and surgeons at the Princess Margaret Cancer Centre in Toronto, Canada, have discovered a promising new approach to treating colorectal cancer.

John Dick, a senior scientist at the Princess Margaret Cancer Centre, said, “This is the first step toward clinically applying the principles of cancer stem cell biology to control cancer growth and advance the development of durable cures.”

In preclinical experiments with laboratory rodents, Dick and his team identified a gene called BMI-1 as a pivotal regulator of colon cancer stem cell proliferation. With this knowledge in hand, Dick’s laboratory dedicated many hours to finding small molecules that disarm BMI-1. Then Dick and his co-workers replicated human colorectal cancer in mice, and used their BMI-1-inhibiting small molecules to treat these cancer-stricken mice.

According to lead author of this work, Antonija Kreso: “Inhibiting a recognized regulator of self-renewal is an effective approach to control tumor growth, providing strong evidence for the clinical relevance of self-renewal as a biological process for therapeutic targeting.”

Dr. Dick explained: “When we blocked the BMI-1 pathway, the stem cells were unable to self-renew, which resulted in long-term and irreversible impairment of tumor growth. In other words, the cancer was permanently shut down.”

The clinical potential of this approach is significant, since it provides a viable treatment that specifically targets colon cancer. About 65% of all colorectal cancers have an activated BMI-1 pathway. Since physicians now have techniques for identifying the presence of BMI-1 and the tools to inhibit it, this strategy could translate into a clinical treatment that might radically transform the treatment of aggressive, advanced colorectal cancers. Such a treatment would be specific, personal, and specific. May the phase 1 trials begin soon!!!

Adult Stem Cells Suppress Cancerous Growth While Dormant


William Lowry and his postdoctoral fellow Andrew White at UCLA’s Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research have discovered the means by which particular adult stem cells suppress their ability to trigger skin cancer during their dormant phase. A better understanding of this mechanism could provide the foundation to better cancer-prevention strategies.

This study was published online Dec. 15 in the journal Nature Cell Biology. William Lowry, Ph.D. is an associate professor of molecular, cell and developmental biology in the UCLA College of Letters and Science.

Hair follicle stem cells are those tissue-specific adult stem cells that generate the hair follicles. Unfortunately, they also are the cell population from which cutaneous squamous cell carcinoma, a common skin cancer, begins. However, these stem cells cycle between active periods, when they grow, and dormant periods, when they do not grow.

Diagram of the hair follicle and cell lineages supplied by epidermal stem cells. A compartment of multipotent stem cells is located in the bulge, which lies in the outer root sheath (ORS) just below the sebaceous gland. Contiguous with the basal layer of the epidermis, the ORS forms the external sheath of the hair follicle. The interior or the inner root sheath (IRS) forms the channel for the hair; as the hair shaft nears the skin surface, the IRS degenerates, liberating its attachments to the hair. The hair shaft and IRS are derived from the matrix, the transiently amplifying cells of the hair follicle. The matrix surrounds the dermal papilla, a cluster of specialized mesenchymal cells in the hair bulb. The multipotent stem cells found in the bulge are thought to contribute to the lineages of the hair follicle, sebaceous gland, and the epidermis (see red dashed lines). Transiently amplifying progeny of bulge stem cells in each of these regions differentiates as shown (see green dashed lines).
Diagram of the hair follicle and cell lineages supplied by epidermal stem cells. A compartment of multipotent stem cells is located in the bulge, which lies in the outer root sheath (ORS) just below the sebaceous gland. Contiguous with the basal layer of the epidermis, the ORS forms the external sheath of the hair follicle. The interior or the inner root sheath (IRS) forms the channel for the hair; as the hair shaft nears the skin surface, the IRS degenerates, liberating its attachments to the hair. The hair shaft and IRS are derived from the matrix, the transiently amplifying cells of the hair follicle. The matrix surrounds the dermal papilla, a cluster of specialized mesenchymal cells in the hair bulb. The multipotent stem cells found in the bulge are thought to contribute to the lineages of the hair follicle, sebaceous gland, and the epidermis (see red dashed lines). Transiently amplifying progeny of bulge stem cells in each of these regions differentiates as shown (see green dashed lines).

White and Lowry used transgenic mouse models for their work, and they inserted cancer-causing genes into these mice that were only expressed in their hair follicle stem cells. During the dormant phase, the hair follicle stem cells were not able to initiate skin cancer, but once they transitioned into their active period, they began growing cancer.

Dr. White explained it this way: “We found that this tumor suppression via adult stem cell quiescence was mediated by PTEN (phosphatase and tensin homolog), a gene important in regulating the cell’s response to signaling pathways. Therefore, stem cell quiescence is a novel form of tumor suppression in hair follicle stem cells, and PTEN must be present for the suppression to work.”

Retinoids are used to treat certain types of leukemias because they drive the cancer cells to differentiate and cease dividing. Likewise, understanding cancer suppression by inducing quiescence could, potentially, better inform preventative strategies for certain patients who are at higher risk for cancers. For example, organ transplant recipients are particularly susceptible to squamous cell carcinoma, as are those patients who are taking the drug vemurafenib (Zelboraf) for melanoma (another type of skin cancer). This study also might reveal parallels between squamous cell carcinoma and other cancers in which stem cells have a quiescent phase.

New Treatment for Mesothelioma Attacks Cancer Stem Cells


The term mesothelium refers to a membrane that forms the lining of several body cavities such as the pleural cavity that contains the lungs or the peritoneum that contains the gastrointestinal tract. Inhalation of asbestos fibers repeatedly injures the mesothelium, and the cycle of injury and regeneration selects for cells that grow faster and faster. Such conditions predispose people to cancer and cancer of the mesothelium or mesothelioma is a consequence of repeated exposure to asbestos fibers.

Mesothelium

In the United Kingdom (UK), asbestos was banned in 1985, but the number of asbestos-related deaths has climbed from 153 in 1968 to 2,321 in 2009 and epidemiologists have estimated that the number of asbestos-related deaths will continue to rise over the next 20 years, peaking in 2020.

Mesothelioma treatments do not provide a terribly good prognosis. The drug cisplatin alone or in combination with pemetrexed (brand name Alimta) or cisplatin in combination with raltitrexed, but raltitrexed is no longer commercially available for this type of treatment regime. Cisplatin has also been used in combination with gemcitabine or vinorelbine. If cisplatin cannot be used then carboplatin can be substituted. In all cases, survival rates are rather underwhelming.

The importance of this clinical issue to stem cell biology is that mesothelioma is a type of cancer driven by rogue stem cells that grow uncontrollably. To that end, Dean Fennell and his team from Leicester University, UK have conducted a clinical trial to test a new treatment for mesothelioma.

The Meso2 study, conducted by Synta Pharmaceuticals, examined the efficacy of a drug called ganetspib as a treatment for mesothelioma. The trial is being led by Fennell and his research team, and will enroll about 140 patients.

Ganetespib is a unique drug in that it targets a protein called HSP90 (heat shock protein 90). Heat shock proteins help proteins fold and help unfolded proteins refold. They are called heat shock proteins because their expression increases when temperatures are raised. Since cancer cells grow quickly and make large quantities of protein, hamstringing those proteins that fold other proteins can gum up the internal workings of the cell and cause them to die.

HSP90

Fennell said, “We think this is a new way to being able to target mesothelioma. Laboratory tests show ganetespib is extremely active in mesothelioma, and combined with chemotherapy, this treatment could shrink cancers down and improve symptoms for patients.”

There is also a second clinical trial called COMMAND (Control of Mesothelioma with MAintenance Defactinib) is being sponsored by a Verastem, a Cambridge, Massachusetts-based pharmaceutical company and it will test a new drug called defactinib.

Defactinib inhibits a protein called FAK (focal adhesion kinase), which is also crucial for cancer stem cell function and for the conversion of cancer stem cells into tumors.  FAK acts as an adapter between the cell adhesion molecules on the surface of the cell, and the internal skeleton proteins of the cell.  Therefore when the cell attaches to another cell or a substratum, FAK and the proteins associated with it transmits a message to the rest of the cell that the cell has attached to another cell.  For a great website about FAK, see here.

FAK

Defactinib inhibits FAK and prevents the cell from adapting to its environment and since FAK is involved with spread of the cell over its substratum, proliferation of the cell and migration of the cell.  Inhibition of FAK prevents the cell from properly responding to surface stimuli, and the cell stops growing.

The COMMAND trial will enroll some 350-400 people and Fennell’s lab is involved with starting this trial.

Cancer stem cells can cause cancer to return to return after chemotherapy because most chermotherapeutic strategies attack the progeny of cancer stem cells and not the cancer stem cells themselves.  Inhibiting the FAK protein takes away something cancer stem cells crucially need and Fennell hopes that treatments like defactinib or ganetespib will positively help mesothelioma patients.

Tumor Suppressor Gene is Required For Neural Stem Cells to Differentiate into Mature Neurons


Cancer cells form when healthy cells accumulate mutations that either inactivate tumor suppressor genes or activate proto-oncogenes. Tumor suppressor genes work inside cells to put the brakes on cell proliferation. Proto-oncogenes work to drive cell proliferation. Loss-of-function mutations in tumor suppressor genes remove controls on cell proliferation, which causes cells to divide uncontrollably. Conversely activating mutations in proto-oncogenes removes the controls on the activity of proto-oncogenes, converting them into oncogenes and driving the cell to divide uncontrollably. If a cell accumulates enough of these mutations, they can grow in such an uncontrollable fashion that they start to gain extra chromosomes or pieces of chromosomes, which contributes to the genetic abnormality of the cell. Accumulation of more mutations allows the cell to break free from the original tumorous mass and spread to other tissues.

There are over 35 identified tumor suppressor genes and one of these, CHD5, has another role besides controlling cell proliferation. Researchers at Karolinska Institutet in Stockholm, Swede, in collaboration with other laboratories at Trinity College in Dublin and BRIC in Copenhagen has established a vital role for CHD5 in normal nervous development.

Once stem cells approach the final phase of differentiation into neurons, the CHD5 protein is made at high levels. CHD5 reshapes the chromatin structure into which DNA is packaged in cells, and in doing so, it facilitates or obstructs the expression of other genes.

Ulrika Nyman, postdoc researcher in Johan Holmberg’s laboratory, said that when they switched of CHD5 expression in stem cells from mouse embryos at the time when the brain develops, the CHD5-less stem cells were unable to turn off those genes that are usually expressed in other tissues, and equally unable to turn on those genes necessary for making mature neurons. Thus these CHD5-less stem cells were trapped in a nether-state between stem cells and neurons.

CHD5 function in stem cell differentiationretinoic

The gene that encodes the CHD5 protein is found on chromosome 1 (1p36) and it is lost in several different cancers, in particular neuroblastomas, a disease found mainly in children and is thought to arise during the development of the peripheral nervous system.

Neuroblastomas that lack this part of chromosome 1 that contains the CHD5 gene are usually more aggressive and more rapidly fatal.

Treatment with retinoic acid forces immature nerve cells and some neuroblastomas to mature into specialized nerve cells. However, when workers from Holmberg’s laboratory prevented neuroblastomas from turning up their expression of CHD5, they no longer responded to retinoic acid treatment.

Holmberg explained, “In the absence of CHD5, neural tumor cells cannot mature into harmless neurons, but continue to divide, making the tumor more malignant and much harder to treat. We now hope to be able to restore the ability to upregulate CHD5 in aggressive tumor cells and make them mature into harmless nerve cells.”

How Stem Cells Maintain Skin


Professor Kim Jensen from BRIC, University of Copenhagen and Cambridge University has used careful mapping studies to challenge current ideas of how the skin renews itself.

Skin is a rather complex organ system that consists of many cell types and structures. Skin includes proliferating cells in the stratum germanitivum, differentiating cells in the upper layers of the epidermis, hair cells, fat, sensory neurons, Langerhans cells, and sweat and sebaceous glands.

Jensen explained, “Until now, the belief was that the skin’s stem cells were organized in a strict hierarchy with a primitive stem cells type at the top of the hierarchy, and that this cell gave rise to all other cell types of the skin. However, our results show that there are differentiated levels of stem cells and that it is their close micro-environment that determines whether they make hair follicles, fat- or sweat glands.”

Jensen’s work completes what was a “stem cell puzzle.” As Jensen put it, “our data complete what is already known about the skin and its maintenance. Researchers have until now tried to fit their results into the old model for skin maintenance. However, the results give much more meaning when we relate them to the new model that our research purposes.”

To give an example of what Jensen is talking about, over-proliferation of skin cells can initiate skin cancer, but the stem cells of the skin that help maintain the integrity of the skin will lack any detectable genetic changes. According to Jensen, the reason these stem cells lack detectable genetic changes in that they do not take part in over-proliferation.

To demonstrate this, Jensen used a unique technique to label skin cells. They made a mouse strain that expresses a glowing protein from the control region of the Lrig1 gene. The Lrig1 gene is expressed in all proliferating skin stem cell populations. Therefore, making a mouse strain in which all cells expressing Lrig1 also express a glowing protein is a sure-fire way to label the skin stem cell populations.

Jensen and his cohorts used several experimental strategies. First, they simply mapped out the glowing cells in the skin. Jensen and his colleagues discovered that the skin contains several stem cell populations that reside in distinct compartments.  These different compartmentalized skin stem cells contributed to specific tissues and their domains did not over lap.

Basic RGB

 

When the mice were wounded, the proliferating stem cells freely crossed over into each other’s domains and helped heal and remake structures that they normally would not make.  This shows that upon wounding, the stem cells compartment boundaries break down as the stem cells proliferate to recreate the compartments that might have been lost as a result of wounding.  Therefore, Jensen’s work shows that Lrig1 marks stem cells in the epidermis, and that these stem cells have a unique lineage potential.  Secondly, the epidermis is maintained in discrete compartments by these multiple stem cell populations.  These stem cell populations largely keep to themselves and do not invade other compartments.  Therefore, stem cell compartmentalization underlies maintenance of the tissue complexity of the skin and not “hierarchy.”  This simply means that where the stem cells live is far more important to skin stem cell function than who their parents were.  Finally, wounding alters stem cell fate and break down the boundaries.

Wounding does more than that.  When Jensen and his colleagues made a mouse with an activated form of the ras gene that was expressed in skin, the skin showed no signs of tumor formation.  This is odd, since activating mutations in ras are extremely common in human and mouse tumors and cultured cells with activated ras mutations grow like cancer cells.  However, if the skin of these mice with the activated ras gene in their skin is wounded, then tumors form.  Therefore, wounding not only breaks down the compartments in which stem cells reside, it also potentiates cancer formation.

Jensen said of his results, “Our research will now take two directions.  We will establish mathematical models for organ maintenance in order to measure what stem cells are doing in the skin.  Also, we will expand our investigations in cancer initiation, hoping for results that can contribute to cancer diagnostics and improved treatment.”

Using Sleeping Stem Cells to Treat Aggressive Leukemias


British scientists have discovered that aggressive forms of leukemia (blood cancers) do not displace normal stem cells from the bone marrow, but instead, put them to sleep. If the normal stem cells are asleep, it implies that they can be awakened. This offers a new treatment strategy for acute myeloid leukemia or AML.

This work comes from researchers at Queen Mary, University of London with the support of Cancer Research UK’s London Research Institute.

In the United Kingdom, approximately 2,500 people are diagnosed with AML each year. The disease strikes young and old patients and the majority of patients die from AML.

In healthy patients, the bone marrow contains hematopoietic stem cells (HSCs) that divide to form either a common myeloid precursor (CMP) or a common lymphoid precursor (CLP) that differentiate into various kinds of white blood cells or red blood cells or lymphocytes. Individuals afflicted with AML, however, have bone marrow invaded by leukemic myeloid blood cells. Since red blood cells are derived from the myeloid lineage, AML causes red blood cell deficiencies (anemia), and the patient becomes tired, and is at risk for excessive bleeding. AML patients are also more vulnerable to infection those white blood cells that fight infections are not properly formed.

HSC differentiation2

David Taussig from the Barts Center Institute at Queen Mary, University of London said that the widely accepted explanation for these symptoms is that the cancerous stem cells displace or destroy the normal HSCs.

However, Taussig and his colleagues have found in bone marrow samples from mice and humans with AML contain plenty of normal HSCs. Thus, AML is not destroying or displacing the HSCs. Instead, the cancerous stem cells appear to be turning them off so that they cannot form HSCs. If Taussig and his coworkers and collaborators had determine how these leukemic myeloid blood cells are shutting off the normal HSCs, they might be able to design treatments to turn them back on.

Such a treatment strategy would increase the survival of AML patients. Only 40% of younger patients are cured of AML, and the cure rate for older patients in much lower. Current treatments that include chemotherapy and bone marrow transplants are not terribly successful with older patients.

Taussig’s group examined the levels of HSCs in the bone marrow of mice that had been transplanted with human leukemic myeloid cells from AML patients. They discovered that the numbers of HSCs stayed the same, but these same HSCs failed to transition through the developmental stages that result in the formation of new blood cells. When Taussig and his group examined bone marrow from 16 human AML patients, they discovered a very similar result.

Even though AML treatment has come a long way in the last ten years, there is still an urgent need for more effective treatments to improve long-term survival. This present study greatly advances our understanding of what’s going on in the bone marrow of AML patients. The future challenge is to turn this knowledge into treatments.

Under normal circumstances, stress on the body will boost HSC activity. For example, when the patient hemorrhages, the HSCs kick into action to produce more red blood cells that were lost during the bleed. However, the cancer cells in the bone marrow are somehow over-riding this compensatory mechanism and the next phase of this research will determine exactly how they do it.

Breast Cancer Clinical Trial Targets Cancer Stem Cells


Even though my previous posts about cancer stem cells have generated very little interest, understanding cancer as a stem cell-based disease has profound implications for how we treat cancer. If the vast majority of the cells in a tumor are slow-growing and not dangerous but only a small minority of the cells are rapidly growing and providing the growth the most of the tumor, then treatments that shave off large numbers of cells might shrink the tumor, but not solve the problem, because the cancer stem cells that are supplying the tumor are still there. However, if the treatment attacks the cancer stem cells specifically, then the tumor’s cell supply is cut off and the tumor will wither and die.

In the case of breast cancer, the tumors return after treatment and spread to other parts of the body because radiation and current chemotherapy treatments do not kill the cancer stem cells.

This premise constitutes the foundation of a clinical trial operating from the University of Michigan Comprehensive Cancer Center and two other sites. This clinical trial will examine a drug that specifically attacks breast cancer stem cells. The drug, reparixin, will be used in combination with standard chemotherapy.

Dr. Anne Schott, an associate professor of internal medicine at the University of Michigan and principal investigator of this clinical trial, said: “This is one of only a few trials testing stem cell directed therapies in combination with chemotherapy in breast cancer. Combining chemotherapy in breast cancer has the potential to lengthen remission for women with advanced breast cancer.”

Cancer stem cells are the small number of cells in a tumor that fuel its growth and are responsible for metastasis of the tumor. This phase 1b study will test reparixin, which is given orally, with a drug called paclitaxel in women who have HER2-negative metastatic breast cancer. This study is primarily designed to test how well patients tolerate this particular drug combination. However, researchers will also examine how well reparixin appears to affect various cancer stem cells indicators and signs of inflammation. The study will also examine how well this drug combination controls the cancer and affects patient survival.

This clinical trial emerged from laboratory work at the University of Michigan that showed that breast cancer stem cells expressed a receptor on their cell surfaces called CXCR1. CXCR1 triggers the growth of cancer stem cells in response to inflammation and tissue damage. Adding reparixin to cultured cancer stem cells killed them and reparixin works by blocking CXCR1.

Mice treated with reparixin or the combination of reparixin and paclitaxel had significantly fewer (dramatically actually) cancer stem cells that those treated with paclitaxel alone. Also, riparixin-treated mice developed significantly fewer metastases that mice treated with chemotherapy alone (see Ginestier C,, et al., J Clin Invest. 2010, 120(2):485-97).

Stem Cell-Promoting Gene Also Promotes the Growth of Head and Neck Cancer


Nanog is a very funny name for a gene, but the Nanog gene is an essential part of the cellular machinery that keeps embryonic stem cells from differentiating and maintains them in a pluripotent state. Unfortunately, Nanog also has other roles if it is mis-expressed and that includes in the genesis of cancers of the head and neck.

Nanog function during development
Nanog function during development

This study emerged from work done by researchers at the Ohio State University Comprehensive Cancer Center – Arthur G. James Cancer Hospital, and Richard J. Solove Research Institute or OSUCCC-James. Since Nanog has been studied in some depth, understanding Nanog activity might provide vital clues in the design of targeted drugs and reagents for treating particular cancers.

“This study defines a signaling axis that is essential for head and neck cancer progression, and our findings show that this axis may be disrupted at three key steps,” said Quintin Pan, associate professor of otolaryngology at OSUCCC-James and principal investigator in this research effort. “Targeted drugs that are designed to inhibit any or all of these three steps might greatly improve the treatment of head and neck cancer.”

What kind of signaling axis is Dr. Pan referring to? An enzyme called protein kinase C-epsilon or PKC-epsilon can place phosphate groups on the Nanog protein. Phosphate groups are negatively charged and are also quite bulky. Attaching such chemical groups to a protein can effectively change its structure and function. In the case of Nanog, phosphorylation of stabilizes it and activates it.

Phosphorylated Nanog proteins can bind together to form a dimer, which attracts a third protein to it; p300. This third protein, p300, in combination with the paired Nanog proteins acts as a potent activator of gene expression of particular genes, in particular a gene called Bmi1. When expressed at high levels, Bmi1 stimulates the proliferation of cells in an uncontrolled fashion.

Bmi1 - Nanog interaction

“Our work shows that the PKCepsilon/Nanog/Bmi1 signaling axis is essential to promote head and neck cancer,” Pan said. “And it provides initial evidence that the development of inhibitors that block critical points in this axis might yield a potent collection of targeted anti-cancer therapeutics that could be valuable for the treatment of head and neck cancer.”

Isolating Mammary Gland Stem Cells


Female mammary glands are home to a remarkable population of stem cells that grow in culture as small balls of cells called “mammospheres.” Clayton and others were able to identify these stem cells in 2004 (Clayton, Titley, and Vivanco, Exp Cell Res 297 (2004): 444-60), and Max Wicha’s laboratory at the University of Michigan showed that a signaling molecule called Sonic Hedgehog and a Polycomb nuclear factor called Bmi-1 are necessary for the self-renewal of normal and cancerous mammary gland stem cells (Lui, et al., Cancer Res June 15, 2006 66; 606). The biggest problem with mammary gland stem cells is isolating them from the rest of the mammary tissue.

Mammary gland stem cells or MaSCs are very important for mammary gland development and during the induction of breast cancer. Getting cultures of MsSCs is really tough because the MaSCs share cell surface markers with normal cells and they are also quite few in number.

Gregory Hannon and his co-workers at Cold Spring Harbor Laboratory used a mouse model to identify a novel cell surface protein specific to MaSCs. By exploiting this unusual marker, Hannon and his team were able to isolate MaSCs from mouse mammary glands of rather high purity.

Camila Do Santos, the paper’s first author, said that “We are describing a marker called Cd1d.” Cd1d is also found on the surfaces of cells of the immune system, but is specific to MaSCs in mammary tissue. Additionally, MaSCs divide slower than the surrounding cells. Do Santos and her colleagues used this feature to visually isolate MaSCs from cultured mammary cells.

They used a mouse strain that expresses a green glowing protein in its cells and then made primary mammary cultures from these green glowing mice. After shutting of the expression of the green glowing protein with doxycycline, the cultured cells divided, and diluted the quantity of green glow protein in the cells. This caused them to glow less intensely. However, the slow-growing MaSCs divided much more slowly and glowed much more intensely. Selecting out the most intensely glowing cells allowed Dos Santos and her colleagues to enrich the culture for MaSCs.

“The beauty of this is that by stopping GFP expression, you can directly measure the number of cell divisions that have happened since the GFP was turned off,” said Dos Santos. She continued: “The cells that divide the least will carry GFP the longest and are the ones we characterized.”

Using this strategy, Dos Santos and others selected stem cells from the mammary glands in order to examine their gene expression signature. They also confirmed that by exploiting Cd1d expression in the MaSCS, in combination with other techniques, they could enhance the purity of the cultures several fold.

Hannon added, “With this advancement, we are now able to profile normal and cancer stem cells at a very high degree of purity, and perhaps point out which genes should be investigated as the next breast cancer drug targets.”

Will we be able to use these cell for therapeutic purposes some day?  Possibly, but at this time, more must be known about them and MaSCs must be better characterized.

A Protein Responsible for Cancer Stem Formation Provides a Drug Target


Eighty-five percent of all tumors are carcinomas, which are tumors that form in layers of cells that line surfaces.  Such cell layers are known as an epithelium. When carcinomas form, they undergo an “epithelial-mesenchymal” transformation” or EMT.  EMT means that cells go from being closely aligned and tightly bound to each other in a an organized layer to cells that have little to do with each other and grow in unorganized clumps.  Is there a molecule that unites the carcinomas and if so is this molecule a potential drug target for cancer treatments?

Mammary Carcinoma
Mammary Carcinoma

Researchers at the University of Texas MD Anderson Cancer Center have identified a protein that seems to play a pivotal role in EMT.  This protein, FOXC2, may lay at the nexus of why some carcinomas resist chemotherapy and grow uncontrollably and spread.  FOXC2 could, conceivably represent a novel drug target for chemotherapy.

Sendurai Mani, assistant professor of Translational Molecular Pathology and co-director of the Metastasis Research Center at MD Anderson, said, “We found that FOXC2 lies at the crossroads of the cellular properties of cancer stem cells and cells that have undergone EMT, a process of cellular change associated with generating cancer stem cells.”

Cancer stem cells are fewer in number than other tumor cells, yet research has tied them to cancer progression and resistance to treatment.  Abnormal activation of EMT can actually create cancer stem cells, according to Mani.

Mani continued, “There are multiple molecular pathways that activate EMT.  We found many of these pathways also activate FOXC2 expression to launch this transition, making FOXC2 a potentially efficient check point to block EMT from occurring. ”  Mani’s research group used experiments with cultured cells and mice to discover these concepts, but the next step will require assessing the levels of FOXC2 expression in human tumors samples.

In the meantime, these new data from Mani’s research team may have profound implication for the treatment of particular types of carcinomas that have proven to be remarkably stubborn.  Breast cancers, for example, are typically carcinomas of the mammary gland ductal system.  A specific group of breasts cancers are very notoriously resistant to treatment, and FOXC2 seems to be at the center of such breast cancers.

The anti-cancer drug sunitinib, which is marketed under the trade name Sutent, has been approved by the US Food and Drug Administration (US FDA) for three different types of cancers.  In this study, sunitinib proved effective against these particularly stubborn types of breast cancer; the so-called “triple-negative, claudin-low” breast cancers.

Sunitinib
Sunitinib

Mani explained why such breast cancers are so resistant to treatment:  “FOXC2 is a transcription factor, a protein that binds to DNA in the promoter region of genes to activate them.  For a variety of reasons, transcription factors are hard to target with drugs.”

However, sunitinib seems to target these triple-negative breast cancers.  When mice with triple-negative breast cancer were treated with sunitinib, the treated mice had smaller primary tumors, longer survival, and fewer incidences of metastasis.  The cancer cells also showed a marked decreased in their ability to form “mammospheres,” or balls of cancer stem cells (this is an earmark of cancer stem cells).  Thus sunitinib seem to attack cancer stem cells.

As it turns out, FOXC2 activates the expression of the platelet-derived growth factor receptor-beta (PDGFRc-beta).  Activation of PDGFRc-beta drives cell proliferation in FOXC2-positive cells, and sunitinib inhibits PDGFRc-beta and inhibits cells that have active FOXC2 and undergoing EMT.

Triple-negative breast cancer cells lack receptors that are used by the most common anti-cancer drugs.  These deficiencies are responsible for the resistance of these cancers to treatment.  Such cancer cells also tend to under go EMT because they lack the protein claudin, which binds epithelial cells together.  Without claudin, these cancer cells become extremely aggressive.

Since cells undergoing EMT are heavily expressing FOXC2, Mani and his colleagues used a small RNA molecule that makes a short hairpin and inhibits FOXC2 synthesis.  Unfortunately, blocking FOXC2 had no effect on cell growth, but it did alter the physical appearance of the cells and reduced their expression of genes associated with EMT and increased the expression of E-cadherin, a protein necessary for epithelial cell organization.  Breast cancer cells also became less invasive when FOXC2 was inhibited, and they down-regulated CD44 and CD24, which are markers of cancer stem cells..  Additionally, triple-negative breast cancer cells that had FOXC2 inhibited had a reduced ability to make mammospheres.  Thus, FOXC2 expression is elevated in cancer stem cells, and inhibition of FOXC2 decreased the ability of the cancer stem cells to behave as cancer stem cells.

Mammospheres
Mammospheres

Mani’s group also approached these experiments from another approach by overexpressing FOXC2 in malignant mammary epithelial cells.  This forced FOXC2 expression drove cells to undergo EMT and become much more aggressive and metastatic (the cancer spread to the liver, hind leg, lungs, and brain).  Breast cancer cells without forced FOXC2 overexpression showed no tendency to metastasize.

Finally, Mani’s group examined metastatic mammary tumors that were highly aggressive when implanted into nude mice (mice that cannot reject transplants).  Two of the tumors were claudin-negative and both of these tumors showed elevated FOXC2 expression.  When FOXC2 expression was blocked by Mani’s hairpin RNA, the claudin-negative tumors became less aggressive and grew more as mesenchymal cells.  The cells that underwent EMT also showed high levels of PDGF-RC-beta expression.

Mani said of these data: “We thought PDGF-B might be a drugable target in these FOXC2-expressing cells.”  Mani’s group also showed that suppressing FOXC2 reduced the expression of PDGFRC-Beta.  Thus, this small molecule might be an effective therapeutic strategy for treating these hard-to-treat breast cancers.

MD Anderson has filed a patent application connected to this study.

See Hollier B.G., Tinnirello A.A., Werden S.J., Evans K.W., Taube J.H., Sarkar T.R., Sphyris N., Shariati M., Kumar S.V., Battula V.L., Herschkowitz J.I., Guerra R., Chang J.T., Miura N., Rosen J.M., and Mani S.A.,. FOXC2 expression links epithelial-mesenchymal transition and stem cell properties in breast cancer. Cancer Research. e-Pub 2/2013.

Isolation of Cancer Stem Cells from Childhood Tumor


Benjamin Dekel, the head of the Pediatric Stem Cell Research Institute in Tel Aviv, Israel, and his team have isolated cancer stem cells from tumors found in the kidneys of some children. Wilms’ tumor is an inherited for of cancer that is found in the kidneys of particular children at a young age. Fortunately, these tumors are easily removed, but these children are at risk for other cancers throughout their lives.

Wilms tumor

Accord to Professor Dekel, “In earlier studies, cancer stem cells were isolation from adult cancers of the breast, pancreas, and brain, but so far much less is known about stem cells in pediatric cancers.” Professor Dekel continued, “Cancer stem cells contain the complete genetic machinery necessary to start, sustain and propagate tumor growth and they are often referred to as cancer initiating cells. As such, they not only represent a useful system to study cancer development but they also serve as a way to study new drug targets and potential treatments designed to stop the growth and spread of different types of cancer. We have demonstrated for the first time the presence of cancer stem cells in a type of tumor that is often found in children.”

Wilm’s tumors represent the most frequent type of kidney tumor found in children, and while children do usually respond well if the tumors are removed early surgically and if the patients are given chemotherapy, recurrences are possible and they can spread to other tissues.

Conventional chemotherapy is toxic to all cells in the body and if given to children may lead to the development of secondary cancers when they become adults. Thus, scientists would like to target tumor cells in as specific a manner as possible.

Researchers were able to remove parts of the tumors of cancer patients and graft them into mice. This procedure allowed researchers to test for the presence of cancer stem cells, since only the cancer stem cells could propagate the tumor from one animal to another. In the case of Wilms’ tumor, it was clear that cancer stem cells were present and could even be isolated from the rest of the tumor cells.

Enzyme Promotes Cancer Stem Cell Growth in Chronic Myeloid Leukemia


An international research team led by researchers at the University of California, San Diego School of Medicine has identified an enzyme that plays a key role in cancer stem cell reprogramming in a blood-based cancer known as chronic myeloid leukemia (CML).

CML treatment has received a tremendous boost by the discovery and development of chemotherapuetic agents known as tyrosine kinase inhibitors. Tyrosine kinase inhibitors attack a very specific group of signaling molecules that go awry in CML, and because of their high degree of specificity, these drugs are well tolerated and rather effective.

tyrosine
Tyrosine

Tyrosine kinase inhibitors or TKIs block receptors called receptor tyrosine kinases that bind growth factors. Such receptors include molecules like Epidermal Growth Factor Receptor, which binds the growth factor EGF (Epidermal Growth Factor), Plate-Derived Growth Factor Receptor, which binds Platelet-Derived Growth Factor, and several others. Receptor tyrosine kinases are proteins that are embedded in the membrane of cells and when they are engaged by a specific growth factor, they pair up with another molecule and bind the growth factor tightly. Because this binding of their growth factor targets pairs two receptor tyrosine kinases together, the portion of the receptor protein that sticks toward the inside of the cell is activated. This internal portion of the receptor has “kinase” activity. Kinases are enzymes that stick phosphate groups on other molecules. Kinases place their phosphate groups on very specific targets. In the case of receptor tyrosine kinases, the target is the amino acid tyrosine. It just so happens that the internal piece of receptor tyrosine kinases contains several tyrosine residues and the paired receptors molecules tag each other with several phosphate groups on their tyrosines.

EGF activity

Phosphotyrosine acts as a signal to the inside of the cell, because specific protein contain pieces that can bind to phosphotyrosine. These phosphotyrosine-binding proteins (SH2-domain proteins for those who care about such things) drag powerful signaling molecules to the cell membrane. These signaling molecules are activated and the cell undergoes changes that cause it to move, growth, divide, or do other types of things.

EGFR signaling

In the case of blood cells, activation of particular receptor tyrosine kinases induces cells to grow and divide. Because there are exquisite controls on the signals set in motion by tyrosine, these signals cells divide and then stop. However, if the genes that encode these receptor tyrosine kinases undergo mutations that allow the receptors to pair up without binding growth factors, then the receptors will activate themselves at will without being dependent on the availability of growth factors. Cell will grow uncontrollably and fill up the bone marrow and blood.

At this point, we can see how TKIs work. These small molecules bind to the kinase part of receptor tyrosine kinases and gum them up. Because cells do not receive the signal to grow, they stop growing uncontrollably and this send the cancer into remission. TKIs include such famous drugs as Gleevec (imatinib), which was one of the first TKIs and has provent very successful against CML. However, after long periods of time on Gleevec, tumor cells can become resistant to it, and the physician must change drugs. Other TKIs include gefitinib (Iressa), and erlotinib (Tarceva), which inhibit Epidermal Growth Factor Receptor, Lapatinib (Tykerb), which is a dual inhibitor of EGFR and a subclass called Human EGFR type 2, and Sunitinib (Sutent) which is multi-targeted drug that inhibits Platelet-Derived Growth Factor Receptor and Vascular Endothelial Growth Factor Receptor.

Sunitinib
Sunitinib
Gefitinib
Gefitinib
Erlotinib
Erlotinib
imatinib
imatinib
Laptinib
Laptinib

 

 

 

 

 

 

 

 

 

There are also other most specialized TKIs such as sorafenib (Nexavar), which targets a complex pathway that leads to a kinase signaling cascade, and nilotinib (Tasinga) which inhibits the fusion protein bcr-abl and is typically prescribed when a patient has shown resistance to imatinib (Gleevex).

Well, with all these new drugs, what’s the problem? The problem is that blood cancers, leukemias, can find ways around these treatments. Therefore, we must learn more these cancers in order to improve treatment of them. Leukemias are definitely tumors that emerge from cancer stem cells. Therefore, if you kill the cancer stem cells, you kill the tumor.

Principle investigator of this research, Catriona Jamieson, associate professor of medicine at UC San Diego, in collaboration with colleagues from Canada and Italy reported that inflammation, a phenomenon long associated with the development of cancer, increases the activity of the enzyme ADAR1 or adenosine deamiinase 1.

ADAR1 is expressed during embryonic development and it is essential in blood cell development. After embryonic development, ADAR1 switches off, but is reactivated by viral infections. Its role during viral infections is to protect blood cell-making stem cells from viral attacks. In leukemia stem cells, however, ADAR1 enhances the abnormal processing of RNA molecules. This causes enhanced cell renewal and resistance of malignant stem cells to chemotherapy.

Jamieson has already studied the link between cancer stem cell instability and abnormal RNA processing. She said, “People normally think about DNA instability in cancer, but in this case, it’s how the RNA is edited by enzymes that really matters in terms of cancer stem cell generation and resistance to conventional therapy.”

Because this RNA processing process is basic to cells and occurs in closely related organisms as well, studying it should be possible in model systems. It also represents a novel target for new therapies. According to Jamieson, inflammation is “an essential driver of cancer relapse and therapeutic resistance.”

Jamieson continued, “ADAR1 is an enzyme that we may be able to specifically target with a small molecule inhibitor, an approach we have already used effectively with other inhibitors. If we can block the capacity of leukemia stem cells to use ADAR1, if we can knock down that pathway, maybe we can put stem cells back on the right track and stop malignant cloning.”

The initiation of CML requires a mutation that fuses two genes together. One of these genes, BCR, fuses to a protein tyrosine kinase called ABL to generate the BCR-ABL gene that makes a fusion protein with uninhibited activity. The white blood cells that contain the BCR-ABL fusion protein expand slowly. The slowness with which this leukemia expands makes it difficult to diagnose early, and diagnosis is only possible once there are large numbers of precursors and malignant cells throughout the bloodstream and bone marrow. The median age at which CML is diagnosed is 66, and despite the advances in chemotherapeutic treatments, the vast majority of patients relapse if therapy is discontinued, since the cancer stem cells are dormant and resistant to treatment. If ADAR1 is addressed as a target, then perhaps treatments that target ADAR1 will overcome cancer stem cell resistance and prevent relapse.

Formation of the Philadelphia chromosome that produces the bcr-abl oncogene.
Formation of the Philadelphia chromosome that produces the bcr-abl oncogene.

In the United States alone, there are an estimated 70,00 people with CML and as the population ages, the prevalence of this disease is projected to jump to approximately 181,000 by 2050.

See “ADAR1 promotes malignant progenitor reprogramming in chronic myeloid leukemia.” Qingfei Jianga et al., PNAS DOI: 10.1073/pnas.2123021110.