First Large-Scale Genomic Analysis of Key Acute Leukemia Will Likely Yield New Therapies

A consortium including St. Jude Children’s Research Hospital and the Children’s Oncology Group has performed an unprecedented genomic sequencing analysis of hundreds of patients with T-lineage acute lymphoblastic leukemia (T-ALL). The results provide a detailed genomic landscape that will inform treatment strategies and aid efforts to develop drugs to target newly discovered mutations.

The data will also enable researchers to engineer better mouse models to probe the leukemia’s aberrant biological machinery.

The project’s 39 researchers were led by Charles Mullighan, M.D., MBBS, a member of the St. Jude Department of Pathology, with co-corresponding authors Jinghui Zhang, Ph.D., chair of the St. Jude Department of Computational Biology and Stephen Hunger, M.D., of the Children’s Hospital of Philadelphia. The research was selected for advance online publication today in the journal Nature Genetics.

“This first comprehensive and systematic analysis in a large group of patients revealed many new mutations that are biologically significant as well as new drug targets that could be clinically important,” Mullighan said. “Leukemias typically arise from multiple genetic changes that work together. Most previous studies have not had the breadth of genomic data in enough patients to identify the constellations of mutations and recognize their associations.”

T-ALL is a form of leukemia in which the immune system’s T cells acquire multiple mutations that freeze the cells in an immature stage, causing them to accumulate in the body. ALL is the most common type of childhood cancer, affecting about 3,000 children nationwide each year. T-ALL constitutes about 15 percent of those cases. While about 90 percent of children with ALL can be cured, many still relapse and require additional treatment.

The multi-institutional effort involved sequencing the genomes of 264 children and young adults with T-ALL—the largest such group ever analyzed. The study involved sophisticated analysis of multiple types of genomic data, led by Yu Liu, Ph.D., a postdoctoral fellow in Zhang’s Computational Biology laboratory and first author of the study. Their analyses identified 106 driver genes—those whose mutations trigger the malfunctions that block normal T cell development and give rise to cancer. Half of those mutated genes had not been previously identified in childhood T-ALL.

The study enabled the researchers to compare the frequencies of mutations among patients whose cancerous cells were sequenced at the same detailed level, Mullighan said. Also important, he said, was that all the patients had uniform treatment, which enabled the researchers to draw meaningful associations between the genetics of their cancer and the response to different treatments. Such associations will enable better diagnosis and treatment of T-ALL with existing drugs.

Researchers analyzed the cancerous T cells as well as those that treatments had rendered non-cancerous. Comparing the two populations of cells could reveal valuable clues about why specific treatments were successful in thwarting particular cancer-causing mutations.

The findings revealed significant unexpected findings. “We went into this study knowing that we didn’t know the full genomic landscape of T-ALL,” Hunger said. “But we were surprised that over half of the new targets and mutations were previously unrecognized. It was particularly unexpected and very striking that some mutations were exclusively found in some subtypes of T-ALL, but not others.”

Cancers are driven by mutations in genes that are the blueprint for protein enzymes in signaling pathways in cells—the biological equivalent of circuits in a computer. While a cancer may arise from an initial founding mutation, that mutation triggers a cascade of other mutations that help drive the cancer.

The new genomic analysis confirmed that T-ALL was driven by mutations in known signaling pathways, including JAK–STAT, Ras and PTEN–PI3K.

However, the new analysis identified many more genetic mutations in those known pathways. The findings offered more targets for drugs to shut down the aberrant cells. “So the frequency of the patients that are potentially amenable to these targeted approaches is higher than we appreciated before,” Mullighan said.

The researchers also found cases in which the same T-ALL subtype had mutations in different pathways triggered by the same cancer-causing founding mutation. “We believe this finding suggests we can target such subtypes with an inhibitor drug for one of the pathways, and it’s likely to be effective,” Mullighan said.

The multitude of new mutations uncovered in the new study will also enable researchers to use genetic engineering to create mouse models that more accurately reflect human cancer, he said. Such models are invaluable for understanding the biological machinery of T-ALL, as well as testing new drug strategies. “We now have a launching pad, if you will, to design mouse models that include multiple genetic mutations to more faithfully reflect the leukemias we see in humans,” Mullighan said.

The research also offers a broader lesson for genomic studies of cancers, Zhang said. “Our study is further evidence that if you systematically study a large enough population with careful, detailed genomic analysis, you will discover new mutational patterns of collaboration or exclusion across multiple genes unique to each T-ALL subtype,” she said.

The study was a collaboration between the St. Jude Children’s Research Hospital – Washington University Pediatric Cancer Genome Project, the Children’s Oncology Group (COG) and the National Cancer Institute’s Therapeutically Applicable Research to Generate Effective Treatments (TARGET) initiative. COG is a federally supported clinical trials group focused exclusively on childhood cancer. TARGET uses genomic analysis of COG samples to identify therapeutic targets and spur development of more effective treatment for childhood cancer.

First-In-Human Clinical Trial Aims to Extend Remission for Children and Young Adults With Leukemia Treated With T-Cell Immunotherapy

Phase 1 pilot study utilizes T-cell antigen presenting cells to prolong the persistence of cancer-fighting chimeric antigen receptor (CAR) T cells, reduce the relapse rate

After phase 1 results of Seattle Children’s Pediatric Leukemia Adoptive Therapy (PLAT-02) trial have shown T-cell immunotherapy to be effective in getting  93 percent of patients with relapsed or refractory acute lymphoblastic leukemia (ALL) into complete initial remission, researchers have now opened a first-in-human clinical trial aimed at reducing the rate of relapse after the therapy, which is about 50 percent. The new phase 1 pilot study, PLAT-03, will examine the feasibility and safety of administering a second T-cell product intended to increase the long-term persistence of the patient’s chimeric antigen receptor (CAR) T cells that were reprogrammed to detect and destroy cancer.

The research team, led by Dr. Mike Jensen at the Ben Towne Center for Childhood Cancer Research at Seattle Children’s Research Institute, is exploring this strategy after discovering that of the patients who relapse in the PLAT-02 trial, about half of them have lost their CAR T cells. Lasting persistence of the CAR T cells is critical in combating a recurrence of cancer cells.

“While it’s promising that we’re able to get these patients who are very sick back into remission, we’re also seeing that the loss of the CAR T cells in some patients may be opening the door for the cancer to return,” said Dr. Colleen Annesley, an oncologist at Seattle Children’s and the lead investigator of the PLAT-03 trial. “We’re pleased to now be able to offer patients who have lost or are at risk of losing their cancer-fighting T cells an option that will hopefully lead to them achieving long-term remission.”

In the PLAT-03 trial, patients will receive “booster” infusions of a second T-cell product, called T antigen-presenting cells (T-APCs). The T-APCs have been genetically modified to express the CD19 target for the cancer-fighting CAR T cells to recognize. Patients will receive a full dose of T-APCs every 28 days for at least one and up to six doses. By stimulating the CAR T cells with a steady stream of target cells to attack, researchers hope the CAR T cells will re-activate, helping to ensure their persistence long enough to put patients into long-term remission.

PLAT-03 is now open to patients who first enroll in phase 2 of Seattle Children’s PLAT-02 trial and who are also identified as being at risk for early loss of their reprogrammed CAR T cells, or those who lose their reprogrammed CAR T cells within six months of receiving them.

The PLAT-03 trial is one of several trials that Seattle Children’s researchers are planning to open within the next year aimed at further improving the long-term efficacy of T-cell immunotherapy. In addition to the current T-cell immunotherapy trial that is open for children with neuroblastoma, researchers also hope to expand this promising therapy to other solid tumor cancers.

“We are pleased to be at a pivotal point where we are now looking at several new strategies to further improve CAR T-cell immunotherapy so it remains a long-term defense for all of our patients,” said Dr. Rebecca Gardner, Seattle Children’s oncologist and the lead investigator of the PLAT-02 trial. “We’re also excited to be working to apply this therapy to several other forms of pediatric cancer beyond ALL, with the hope that T-cell immunotherapy becomes a first line of defense, reducing the need for toxic therapies and minimizing the length of treatment to only weeks.”

To read about the experience of one of the patients in the PLAT-02 trial, please visit Seattle Children’s On the Pulse blog.

The T-cell immunotherapy trials at Seattle Children’s are funded in part by Strong Against Cancer, a national philanthropic initiative with worldwide implications for potentially curing childhood cancers. If you are interested in supporting the advancement of immunotherapy and cancer research, please visit Strong Against Cancer’s donation page.

For more information on immunotherapy research trials at Seattle Children’s, please call (206) 987-2106 or email immunotherapy@seattlechildrens.org.

Monoclonal Antibody Drug Superior to Chemotherapy for Advanced Acute Lymphoblastic Leukemia

More than 100 centers participate in Phase III randomized trial revealing longer overall survival

A Phase III clinical trial involving 101 centers in 21 countries revealed the monoclonal antibody blinatumomab to be more effective than standard chemotherapy for treatment of advanced acute lymphoblastic leukemia (ALL). Study findings were published in the March 1 online issue of the New England Journal of Medicine.

The study, led by The University of Texas MD Anderson Cancer Center, randomly assigned 405 patients 18 years or older to groups receiving either blinatumomab or chemotherapy. Overall survival was significantly longer in the blinatumomab group with median survival of 7.7 months versus four months for those on chemotherapy. Remission rates within 12 weeks after treatment began were higher in the blinatumomab group with complete remission rates of 34 percent reported in this group versus 16 percent for those on chemotherapy. The study also showed that patients treated with blinatumomab had a lower rate of adverse effects.

While the prognosis for newly diagnosed ALL has improved over the last three decades with intensive chemotherapy regimens resulting in complete remission rates of 85 to 90 percent and long-term survival rates of 30 to 50 percent, most adult patients with the B-cell precursor ALL, the most common form, ultimately relapse and die from disease complications. The accepted standard of care is to help the patient maintain remission long enough to receive allogeneic or donor stem-cell transplantation, considered the most effective therapy.

“Among adults with relapsed ALL, remission rates are18 to 44 percent with standard chemotherapy but the duration of remission is typically short. A major goal for these patients is to induce remission with sufficient duration to prepare for stem-cell transplantation,” said Hagop Kantarjian, M.D., chair of the Department of Leukemia, and lead author for the New England Journal of Medicine paper. “In this study, 24 percent of patients in each treatment group underwent allogeneic stem cell transplantation.”

Blinatumomab, developed by Amgen, works by binding simultaneously to specific cytotoxic T-cells and B-cells, which allows the patient’s healthy T-cells to recognize and eliminate cancer stem cells called blasts.

“The activity of an immune-based therapy such as blinatumomab, which depends on functioning T-cells for its activity, provides encouragement that responses may be further enhanced and made durable with additional immune activation strategies,” said Kantarjian.

The study, designed and funded by Amgen, did not include patients who had other active cancers, relevant central nervous system conditions, autoimmune disease, acute or chronic graft-versus-host disease, chemotherapy or radiotherapy within two weeks before the study, donor stem cell transplantation within 12 weeks before the trial or autologous stem cell transplantation within six week preceding the study. Patients who received immunotherapy in the month before the trial or who were undertaking other investigational treatments were also excluded.

Long-Sought Genetic Model Of Common Infant Leukemia Described

After nearly two decades of unsuccessful attempts, researchers from the University of Chicago Medicine and the Cincinnati Children’s Hospital Medical Center have created the first mouse model for the most common form of infant leukemia. Their discovery, published in the Nov. 14, 2016, issue of Cancer Cell, could hasten development and testing of new drug therapies.

Pro-B acute lymphoblastic leukemia (ALL) with the (4;11) translocation is responsible for about 70 percent of infant and 10 percent of both childhood and adult acute lymphoblastic leukemias. The new mouse model replicates the human genetic flaw that causes this disease, making it much easier to study.

This subtype of leukemia results from a genetic fusion t(4;11), known as a translocation. This combines parts of two separate genes. One of those genes, MLL (short for mixed-lineage leukemia), comes from chromosome 11. The other fragment, AF4 (short for ALL fused gene) from chromosome 4. The hybrid MLL-AF4 gene results in leukemia.

Children and adults with this disease produce vast numbers of dysfunctional blood cells, which eventually crowd out functional cells. MLL-AF4 leukemia has a dismal prognosis, among the worst of any subset of acute leukemia.

“For 20 years, scientists have repeatedly tried and consistently failed to make a model of MLL-AF4 Pro-B acute lymphoblastic leukemia,” said Michael Thirman, MD, Associate Professor of Medicine at the University of Chicago. “Even though we understood the basic genetic flaw, no one had been able create a mouse model that mimicked the human disease, which is crucial for evaluating potential therapies.”

That frustrated many researchers, who shifted their focus to test alternative hypotheses on the causes of this leukemia or refocused their laboratories to study different aspects of this disease.

Thirman’s team, including longtime colleague Roger Luo, PhD, began working on this problem “years ago,” he said, and stayed with it. They quickly identified two hurdles.

The first was a problem with the retrovirus that scientists used to insert the leukemia-causing gene into mouse cells. That gene, acquired from leukemia patients, consisted of a human gene fragment from MLL linked to the human fragment from AF4.

“We soon discovered that the virus wasn’t working,” Thirman explained. “We knew that certain parts of human DNA can decrease viral titers. So we switched from the human version of AF4 to the mouse version, Af4, which is slightly different. This increased viral titers 30 fold.”

That worked, but it led to hurdle two. The mice injected with virus transporting MLL-Af4 developed leukemia, but it was the wrong kind. They developed acute myeloid instead of acute lymphoblastic leukemia. “Despite the use of lymphoid conditions,” the study authors wrote, “no lymphoid leukemia was observed.”

Next, they collaborated with James Mulloy, PhD, at Cincinnati Children’s Hospital Medical Center, whose graduate student Shan Lin inserted the fused MLL-Af4 gene into human CD34 cells, derived from cord or peripheral blood from volunteer donors. They transferred those cells to mice with immune systems that permit the growth of human cells. This time, the mice developed Pro-B ALL, identical to the leukemia found in humans.

“The model worked perfectly,” Thirman said. Within 22 weeks, all of the mice developed exactly the same type of leukemia as observed in patients.

Expression of MLL-Af4 in human cells “recapitulates the pro-B ALL observed in patient with t(4:11) as shown by immunophenotype, chromatin targeting of the fusion, nuclear complex formation, and gene expression signatures,” the authors wrote. “It mimics the disease found in humans both phenotypically and molecularly.”

“The differences in the type of leukemia that developed using mouse versus human cells were striking,” said Mulloy. “Researchers need to consider these differences carefully when choosing which model to use to mimic human disease. The available evidence now indicates that the approaches are not equivalent.”

They conclude that “our MLL-Af4 model will be a valuable tool to study this most prevalent MLL-fusion leukemia with such a poor prognosis.”

However, there is more work to be done. “MLL fusion disease is not a single genetic entity,” the authors note. “Each has its own genetic and biological features associated with particular fusion partners.” This highlights the need for “more models specific to each fusion. Our MLL-Af4 model will be a valuable tool.”

Hybrid Treatment Hunts Down and Kills Leukemia Cells

Researchers at UC Davis and Ionis Pharmaceuticals have developed a hybrid treatment that harnesses a monoclonal antibody to deliver antisense DNA to acute lymphoblastic leukemia (ALL) cells and that may lead to less toxic treatments for the disease.

The study, published in the journal Molecular Medicine, demonstrated that once delivered, the therapeutic DNA reduced levels of MXD3, a protein that helps cancer cells survive. This novel conjugate therapy showed great promise in animal models, destroying ALL cells while limiting other damage.

“We’ve shown, for the first time, that anti-CD22 antibody-antisense conjugates are a potential therapeutic agent for ALL,” said Noriko Satake, associate professor in the Department of Pediatrics at UC Davis. “This could be a new type of treatment that kills leukemia cells with few side effects.”

ALL is the most common type of childhood cancer. It is a disease in which the bone marrow makes too many immature lymphocytes, a type of white blood cell. While most children survive ALL, many patients suffer late or long-term side effects from treatment, which may include heart problems, growth and development delays, secondary cancers and infertility.

Antisense oligonucleotides are single strands of DNA that can bind to messenger RNA, preventing it from making a protein. While antisense technology has long shown therapeutic potential, getting the genetic material inside target cells has been a problem.

In the study, researchers attached antisense DNA that inhibits the MXD3 protein to an antibody that binds to CD22, a protein receptor expressed almost exclusively in ALL cells and normal B cells.

Once the antibody binds to CD22, the conjugate is drawn inside the leukemia cell, allowing the antisense molecule to prevent MXD3 production. Without this anti-apoptotic protein, ALL cells are more prone to cell death.

The hybrid treatment was effective against ALL cell lines in vitro and primary (patient-derived) ALL cells in a xenograft mouse model. Animals that received the hybrid therapy survived significantly longer than those in the control group.

Designed to be selective, the treatment only targets cells that express CD22. While it does attack healthy B cells, the therapy is expected to leave blood stem cells and other tissues unscathed.

“You really don’t want to destroy hematopoietic stem cells because then you have to do a stem cell transplant, which is an extremely intensive therapy,” noted Satake. “Our novel conjugate is designed so that it does not harm hair, eyes, heart, kidneys or other types of cells.”

While the study shows the conjugate knocked down MXD3, researchers still have to figure out how this was accomplished. In addition, they will investigate combining this treatment with other therapies. Because it hastens cell death, the conjugate could make traditional chemotherapy drugs more effective. In addition, the approach might work against other cancers.

“You can see this as proof of principle,” Satake said. “You could switch the target and substitute the antibody, which could be used to treat other cancers or even other diseases.”

Cancer Trial for Leukemia Halted after Death of Two Patients

The U.S. Food and Drug Administration placed a hold on a Juno Therapeutics clinical trial of a treatment for a form of leukemia following the death of two trial patients last week.

The Company said in a press announcement that both deaths occurred last week after the patients, who had relapsed or refractory B cell acute lymphoblastic leukemia, took the drug fludarabine before receiving the chimeric antigen receptor (CAR) T cells that Juno had taken from their bodies and re-engineered to better attack cancer cells. Another patient had died earlier in the trial, but “confounding factors” spurred Juno to continue with the study at that time

Juno announced that it has received notice from the U.S. Food and Drug Administration (FDA) that a clinical hold has been placed on the Phase II clinical trial of JCAR015 in adult patients with relapsed or refractory B cell acute lymphoblastic leukemia (r/r ALL), known as the “ROCKET” trial.

Juno has proposed to the FDA to continue the ROCKET trial using JCAR015 with cyclophosphamide pre-conditioning alone. In response, the FDA has requested that Juno submit, as a Complete Response to the Clinical Hold several items including a revised patient informed consent form, a revised investigator brochure, a revised trial protocol, and a copy of the presentation made to the agency yesterday. Juno stated they will submit the requested information to the FDA this week.

Juno’s trials and plans for its other CD19-directed CAR-T cell product candidates, including JCAR017, are not affected.