Gene Identified That May Provide Potential Therapy for Cerebral Cavernous Malformations

Researchers at University of California San Diego School of Medicine, with national collaborators, have identified a series of molecular clues to understanding the formation of cerebral cavernous malformations (CCMs). The study offers the first genome-wide analysis of the transcriptome of brain microvascular endothelial cells after KRIT1 inactivation. Findings were published September 28 in the Journal of Experimental Medicine.

“These mouse studies reveal a critical mechanism in the pathogenesis of cerebral cavernous malformations and point to the possibility of using angiogenesis inhibitors, such as TSP1 for potential therapy,” said Mark H. Ginsberg, MD, professor of medicine, UC San Diego School of Medicine.

CCMs are collections of enlarged and irregular blood vessels in the central nervous system (CNS), for which there is no drug therapy. The vessels are prone to leakage causing headaches, seizures, paralysis, hearing or vision loss, or bleeding in the brain. There are two forms of the condition: familial and sporadic, affecting 1 in 200 patients in the U.S. The current treatment for CCMs involves invasive surgery, however, surgery is not possible for all patients due to location of vascular lesions within the CNS.

The most common cause of familial cavernous malformations is mutations of KRIT1. The protein produced from this gene is found in the junctions connecting neighboring blood vessel cells. Loss of function mutations in KRIT1 result in weakened contacts between blood vessel cells and CNS vascular abnormalities as seen in CCMs.

“Inactivation of KRIT1 in endothelial cells causes a cascade of changes in the expression of genes that regulate cardiovascular development,” said Ginsberg. “What we learned is that reduced expression of a protein encoded by one of these genes, TSP1, contributes to the growth of CCMs. Loss of one or two copies of THBS1, the gene that encodes TSP1, makes a mouse model of the disease much worse. Conversely, administration of 3TSR, a fragment of TSP1, reduces lesions in this mouse model. This means that replacement of TSP1 by 3TSR or other angiogenesis inhibitors may be a preventative for CCMs or treatment of the disease.”

Penn’s Glowing Cancer Tool Illuminates Benign, but Dangerous, Brain Tumors during Pituitary Surgery

Fluorescent, targeted dye illuminates molecular signature of tumor tissue in personalized surgery.

An experimental imaging tool that uses a targeted fluorescent dye successfully lit up the benign brain tumors of patients during removal surgery, allowing surgeons to identify tumor tissue, a new study from researchers at the Perelman School of Medicine at the University of Pennsylvania shows. The tumors, known as pituitary adenomas, are the third most common brain tumor, and very rarely turn cancerous, but can cause blindness, hormonal disorders, and in some cases, gigantism.

Findings from the pilot study of 15 patients, published this week in the Journal of Neurosurgery, build upon previous clinical studies showing intraoperative molecular imaging developed by researchers at Penn’s Center for Precision Surgery can improve tumor surgeries. According to first author John Y.K. Lee, MD, MSCE, an associate professor of Neurosurgery in the Perelman School of Medicine at the University of Pennsylvania and co-director of the Center for Precision Surgery, this study describes the first targeted, near infrared dye to be employed in brain tumor surgery. Other dyes are limited either by their fluorescent range being in the busy visible spectrum or by lack of specificity.

“This study heralds a new era in personalized tumor surgery. Surgeons are now able to see molecular characteristics of patient’s tumors; not just light absorption or reflectance,” Lee said. “In real time in the operating room, we are seeing the unique cell surface properties of the tumor and not just color. This is the start of a revolution.”

Non-specific dyes have been used to visualize and precisely cut out brain tumors during resection surgery, but this dye is believed to be the first targeted, near infrared dye to be used in neurosurgery. The fluorescent dye, known as OTL38, consists of two parts: vitamin B9 (a necessary ingredient for cell growth), and a near infrared glowing dye. As tumors try to grow and proliferate, they overexpress folate receptors. Pituitary tumors can overexpress folate receptors more than 20 times above the level of the normal pituitary gland in some cases. This dye binds to these receptors and thus allows us to identify tumors.

“Pituitary adenomas are rarely cancerous, but they can cause other serious problems for patients by pushing up against parts of their brain, which can lead to Cushing’s disease, gigantism, blindness and death,” Lee explained. “The study shows that this novel, targeted, near infrared fluorescent dye technique is a safe, and we believe this technique will improve surgery.”

Lee says larger studies are warranted to further demonstrate its clinical effectiveness, especially in nonfunctioning pituitary adenomas.

A big challenge with this type of brain surgery is ensuring the entire tumor is removed. Parts of the tumor issue are often missed by conventional endoscopy approaches during removal, leading to a recurrence in 20 percent of patients. The researchers showed that the technique was safe and effective at illuminating the molecular features of the tumors in the subset of patients with nonfunctioning pituitary adenomas.

The technique uses near-infrared, or NIR, imaging and OTL38 fluoresces brightly when excited by NIR light. The VisionSense IridiumTM 4mm endoscope is a unique camera system which can be employed in the narrow confines of the nasal cavity to illuminate the pituitary adenoma. Both the dye and the camera system are needed in order to perform the surgery successfully.

The rate of gross-total resection (GTR) for the 15 patients, based on postoperative MRI, was 73 percent. The GTR with conventional approaches ranges from 50 to 70 percent. Residual tumor was identified on MRI only in patients with more severe tumors, including cavernous sinus invasion or a significant extrasellar tumor.

In addition, for the three patients with the highest overexpression of folate, the technique predicted post-operative MRI results with perfect concordance.

Some centers have resorted to implementing MRI in the operating room to maximize the extent of resection. However, bringing a massive MRI into the operating room theater remains expensive and has been shown to produce a high number of false-positives in pituitary adenoma surgery. The fluorescent dye imaging tool, Lee said, may serve as a replacement for MRIs in the operating room.

Co-authors on the study include M. Sean Grady, MD, chair of Neurosurgery at Penn, and Sunil Singhal, MD, an associate professor of Surgery, and co-director the Center for Precision Surgery.

Over the past four years, Singhal, Lee, and their colleagues have performed more than 400 surgeries using both nonspecific and targeted near infrared dyes. The breadth of tumor types include lung, brain, bladder and breast.

Most recently, in July, Penn researchers reported results from a lung cancer trial using the OTL38 dye. Surgeons were able to identify and remove a greater number of cancerous nodules from lung cancer patients with the dye using preoperative positron emission tomography, or PET, scans. Penn’s imaging tool identified 60 of the 66 previously known lung nodules, or 91 percent. In addition, doctors used the tool to identify nine additional nodules that were undetected by the PET scan or by traditional intraoperative monitoring.

Researchers at Penn are also exploring the effectiveness of additional contrast agents, some of which they expect to be available in the clinic within a few months.

“This is the beginning of a whole wave of new dyes coming out that may improve surgeries using the fluorescent dye technique,” Lee said. “And we’re leading the charge here at Penn.”

Attractive Drug Candidate Identified to Target Glioma Brain Tumors

This rapidly fatal brain cancer has seen only two improvements in therapy in 30 years.

In a paper published today in Cancer Research, researchers: 1) identify a biomarker enzyme associated with aggressive glioma brain tumors, 2) reveal the regulatory mechanism for that enzyme, and 3) demonstrate potent efficacy, using a mouse model of glioma, for a small molecule inhibitor they have developed.

The inhibitor, GA11, retains a core structure that resembles natural inhibitors of the biomarker enzyme; but the inhibitor has been modified to help it pass through the blood-brain barrier.

“In principle, both these features make GA11 an attractive drug candidate to target glioma stem-like cells in glioblastoma multiforme tumors,” said Ichiro Nakano, M.D., Ph.D., and colleagues in the paper.

Nakano, a professor of neurosurgery and academic neurosurgeon at the University of Alabama at Birmingham, and Vito Coviello and Concettina La Motta, University of Pisa, Italy, are doing further preclinical evaluation of the GA11 and its analogs.

Glioblastoma multiforme, or GBM, is a formidable cancer foe. Only two therapeutic improvements have appeared in the past 30 years, increasing the average survival of patients from five months to 15 or 16 months, Nakano says.

A GBM tumor is a mix of different cells that respond differently to therapies. Small numbers of glioma stem-like cells, or GSCs, drive the tumorigenicity of GBM and thus are prime targets for possible treatments. One GSC subtype called the mesenchymal GSC is more malignant and the most therapy-resistant, so Nakano and fellow researchers reasoned that identifying the regulatory molecules active in mesenchymal GSCs might lead to novel and effective therapeutics.

Study details
Nakano and colleagues found that one form of the enzyme aldehyde dehydrogenase — ALDH1A3 — is a specific marker for mesenchymal GSCs, and his group is the first to show that, among the heterogeneous mix of cells in a GBM tumor, cells with high levels of ALDH1A3 expression were more tumorigenic in vivo than are cells that are low in ALDH1A3.

The researchers also found that the FOXD1 transcription factor regulates the production of ALDH1A3 in mesenchymal GSCs. In clinical samples of high-grade gliomas from patients, the expression levels of both FOXD1 and ALDH1A3 were inversely correlated with disease progression — gliomas with high levels were more rapidly fatal than were gliomas with low levels.

Astonishingly, the same mechanism that drives the mesenchymal GSC tumorigenicity in humans acts in an evolutionarily distant organism, the fruit fly. Knocking down the expression of either the fruit fly version of the FOXD1 gene or the fruit fly version of ALDH1A3 blocks the formation of brain tumors in a brain cancer model of the fruit fly species Drosophila melanogaster, the researchers found. Thus, this signaling has been highly conserved in evolution.

The FOXD1 transcription factor is normally active during development from a fertilized egg and embryo to a fetus, and it is silent after birth. The role of FOXD1 in GBM, Nakano and colleagues say, suggests that the mesenchymal GSCs have hijacked the molecular mechanism of normal embryonic development to promote tumor growth.

In preclinical testing, GA11 was validated several ways. The researchers showed that it inhibited ALDH in yeast, reduced ALDH1 activity in cell-culture spheres of mesenchymal GSCs, inhibited proliferation of glioma spheres in cell culture, and inhibited xenograft growth of GBM in mouse brains.

“In conclusion,” Nakano and fellow researchers wrote, “the FOXD1-ALDH1A3 axis is critical for tumor initiation in mesenchymal GSCs, therefore providing possible new molecular targets for the treatment of GBM and other ALDH1-activated cancers.”

Nakano says his study of the role of GSCs in GBM is just one approach to treat glioma tumors. Other labs are pursuing immunotherapy, the use of check-point inhibitors, vaccination and efforts to increase sensitivity to radiotherapy.

It will take combined therapies to treat glioblastoma, Nakano says. “We don’t believe that one therapy will be effective.”

Nakano expects to launch a new clinical trial for glioblastoma in 2017, in conjunction with Burt Nabors, M.D., professor of neurology at UAB. Nakano says UAB will be the only site in the Deep South for this unique trial aimed at a molecular target in glioma stem cells, a target that is different from the ones described in the Cancer Research paper. The referral contact to Nakano’s service will be Lydia P. Harrell.

The Nakano lab is also working on brain metastases, tumors that spread into the brain from other parts of the body. Similar to high-grade gliomas, which originate in the brain, these metastatic brain tumors are lethal, and there are very few therapeutic options. Nakano believes the core stem cell genes and signaling pathways are shared between gliomas and brain metastases.

“If so,” he said, “the molecular targets identified for gliomas are most likely essential in brain metastases. Studies are underway, and similar to the glioma therapy development, I am working to develop clinical trials for brain metastasis, together with medical oncologists Mansoor Saleh, M.D., Andres Forero, M.D., and others at UAB.”

Tumor Paint Brings Light To Toddler’s Brain Tumor

In December of last year, Laura Coffman began to notice that something wasn’t quite right with her 2-year-old son, Hunter. He was leaning to one side and seemed to lose his balance easily. When he became lethargic and started vomiting a few days later on Dec. 28, she knew it was time to see the pediatrician.

After all standard tests came back normal, they were sent to Seattle Children’s for further testing and to find an answer. Unfortunately, it was far worse than anything Coffman could have imagined.

“What I thought was probably just Hunter being a wobbly toddler with a virus turned out to be a brain tumor,” said Coffman. “I will never forget that day. It was the most traumatic six hours of our lives.”

Tumor Paint sheds some light

Hunter was immediately scheduled for surgery to remove the brain tumor that was the size of a golf ball. In preparing for the operation, Coffman and her husband, Atom, were also presented with the opportunity to enroll Hunter in Seattle Children’s Phase 1 trial of BLZ-100 Tumor Paint. Since tumor cells can be difficult to distinguish from healthy cells, the drug aims to improve surgical outcomes by acting as a molecular flashlight that allows surgeons to visibly distinguish a tumor from normal brain tissue.

“We didn’t see how it could hurt and we wanted them to use every tool at their disposal so we enrolled him in the trial,” said Coffman.

Dec. 30 was the day of Hunter’s surgery. Another day the Coffman’s will never forget.

“As a parent you know that your child may never be the same after brain surgery – they may not be able to see, walk or speak,” Coffman said. “It’s the risk you have to take to save their life. But we trusted our surgeon, Dr. Amy Lee, with everything we had.”

Prior to surgery, BLZ-100 Tumor Paint was administered by intravenous injection. In the operating room, the tumor glows green when viewed under a laser light and imaged with a near-infrared camera system.

BLZ-100 Tumor Paint was invented by a team led by Dr. Jim Olson, pediatric neuro-oncologist at Seattle Children’s and Fred Hutch and co-founder of Blaze Bioscience: The Tumor Paint Company. While this Phase 1 trial is focused on examining the safety of the drug and how well it targets tumor tissue, the hope is that it will eventually help guide skilled surgeons to prevent the removal of healthy tissue that can lead to serious long-term side effects.

“Cure is not just about successful treatment of a tumor, but successful treatment of a child,” said Dr. Sarah Leary, principal investigator for the trial and Hunter’s oncologist at Seattle Children’s. “Much of cancer treatment for children is a trade-off where curative therapy comes with serious long-term side effects. By lighting the way for expert surgeons, we’re hopeful that BLZ-100 Tumor Paint could help improve the quality of life for children by reducing treatment-related damage to the healthy brain.”

Hunter’s surgery went well and the majority of his tumor was removed. One small piece on his brain stem was intentionally left because removal may have led to serious neurological injury.

After surgery, it was not long until Hunter was back to his young self.

“We were incredibly lucky to have such an amazing surgeon and we were thrilled that he was able to bounce back so quickly,” said Coffman. “He was able to walk, run and put words together only weeks after the operation.”

A terrifying diagnosis, the journey to remission

Once the tumor was removed, the Coffman’s faced their next hurdle as pathology determined that Hunter had a form of aggressive cancer called medulloblastoma. His next phase of treatment quickly began as he underwent seven months of chemotherapy and radiation to target the remaining tumor, as well smaller lesions in his brain and spine.

“I remember how terrifying that diagnosis was to hear, but Dr. Leary was so optimistic and immediately reassured us when she said ‘this is a cancer that we’ve cured many times,’” said Coffman. “And that’s exactly what they did.”

In August, four weeks after Hunter ended treatment, MRI scans confirmed he was in remission.

“The news was phenomenal – all of his cancer was gone,” she said. “You can’t ask for anything more.”

Today, Hunter is thriving.

“You’d never even know he had cancer or brain surgery,” she said. “His hair is growing back, he’s swimming, impressing us with his speaking skills and having playdates with friends.”

When reflecting on their decision to participate in the trial, Coffman is glad they had the opportunity to experience something that could be a game changer for future kids like Hunter.

“Brain surgery is not something you ever want to think about your child going through, but if that dreadful day ever comes, you definitely want a tool like Tumor Paint that could help guide the surgeon in making potentially life-altering decisions,” said Coffman.

What’s next for BLZ-100 Tumor Paint in children

Since the trial began in June 2015, Seattle Children’s has performed 15 brain surgeries with BLZ-100 Tumor Paint. To date, none of the patients have had any negative side effects and Leary and her team are working to determine the best dose. The drug also appears to be doing what it was designed to do – make tumor tissue glow.

“We have been excited to see that BLZ-100 Tumor Paint is binding to many different types of brain tumors in children and so far has not resulted in any side effects,” said Leary. “We are optimistic that in the future it could be an incredible tool when placed in the expert hands of a neurosurgeon that could lead to improved patient outcomes.”

Dr. Amy Lee, the lead neurosurgeon in the trial and Hunter’s surgeon, adds, “We believe BLZ-100 Tumor Paint holds tremendous potential and eventually could be a valuable aid for surgeons in differentiating tumor from healthy tissue, particularly when there are areas of question.”

Leary said their goal is that this Phase 1 trial at Seattle Children’s, which has the largest pediatric Brain Tumor Program and the most pediatric neurosurgeons in the Northwest, will be followed by other studies that lead BLZ-100 Tumor Paint to become a part of standard care for brain tumor surgery. The next step will be to determine the effectiveness of the drug, which will involve a larger collaborative study that will involve 15 of the leading pediatric brain tumor centers across the country.

“In the future, I hope we’ll look back and wonder how these surgeries were ever done without the lights on,” said Leary.