The study was led by Dr. Moran Rubinstein and graduate student Saja Fadila, along with Anat Mavashov, Marina Brusel and Karen Anderson, all from the Sackler Faculty of Medicine and the Sagol School of Neuroscience at Tel Aviv University, and Dr. Eric Kremer, from the University of Montpellier in France. Also participating in the study were Bertrand Beucher and Iria González-Dopeso Reyes from Montpellier and other researchers from France, the USA and Spain. The research was published in the Journal of Clinical Investigation.

Dr. Moran Rubinstein

Dravet syndrome affects approximately one in 16,000 births and is considered relatively common among rare genetic diseases. Currently, there are around 70 affected children in Israel. The syndrome presents as thermally-induced seizures around six months of age, with progress to frequent spontaneous epileptic seizures and motor and cognitive developmental delays after one year.

Dr. Rubinstein highlights that existing epilepsy drugs are insufficient for children with DS, who face a significant risk of early death. The syndrome results from a genetic mutation that occurs randomly during fetal development in a gene called SCN1A and is not inherited from the parents. Unfortunately, the disease cannot be predicted or discovered during pregnancy, making early diagnosis challenging.

According to the researchers, it is customary nowadays to perform a genetic analysis for children who suffer from complex thermally-induced seizures around the age of six months. However, even if the test detects that the problem is in the SCN1A gene, the final diagnosis is often given after the epilepsy worsens, with the appearance of severe spontaneous convulsions and developmental delays. Although it is important to have an early diagnosis, diagnosis is often delayed, and most children are diagnosed only at the age of one or two years and sometimes even later.

Promising Results in Lab Models

Although genetic therapies have shown promise in DS mice and some of them are undergoing clinical trials in humans, they have only been effective when administered at very early stages, prior to symptom onset. Given the complex and invasive nature of gene therapy, it cannot be administered without a confirmed diagnosis of DS. Hence, the researchers focused on developing a treatment that could be effective after seizure onset, even at a relatively late age. Additionally, since DS involves cognitive impairments, the team aimed to alleviate both epilepsy and cognitive symptoms.

Dr. Rubinstein explains that viruses are commony used as carriers in genetic therapies to introduce normal genetic material into patients, enabling normal cellular function. For this purpose, the virus is engineered: its original genetic material is removed so it cannot cause disease or replicate itself, and instead, the relevant normal gene is packed inside. In the case of Dravet syndrome, since the SCN1A gene is very large, it was not possible to use common viruses that are usually used for this purpose and a virus capable of carrying and transferring large genes was needed. The team solved this problem by using a virus called Canine adeno virus type 2, as a carrier of the normal gene.

The carrier virus was directly injected into the brains of DS mice since its properties prevent it from crossing the blood-brain barrier. The treatment was administered to 31 mice at three weeks of age,  after spontaneous convulsions had commenced (equivalent to one to two years of age in children), and to 13 mice at five weeks of age (equivalent to approximately six to eight years of age in children). The injection was performed in multiple brain areas, while an empty virus was injected into the brains of 48 control mice.

Potential for Rare Diseases

Promising results followed, with the highest efficacy observed when the treatment was administered at three weeks of age. In these mice, seizures ceased entirely within 60 hours of injection, life expectancy significantly increased, and cognitive impairment, assessed through spatial memory tests, was completely restored. Even in mice treated at five weeks of age, there was notable improvement, characterized by reduced epileptic activity and protection against thermally-induced seizures. In the control group that received the empty virus, no improvement was observed, and the mice experienced symptoms akin to untreated mice, with approximately 50% succumbing to early death due to severe epilepsy. The treatment was also applied to healthy mice without any adverse effects, demonstrating its safety.

The researchers clarify that their treatment restored normal function to damaged neurons in the brain by introducing a complete, normal gene. This approach is crucial in treating Dravet syndrome since the mutation can occur at different locations within the gene, and administering a complete gene provides a univform treatment suitable for all DS patients. Furthermore, the chosen virus infected numerous nerve cells and spread widely beyond the injection site, enhancing its effectiveness.  

Dr. Rubinstein concludes that their treatment is the first proven to be effective for Dravet syndrome after the onset of spontaneous convulsions, offering improvement in cognitive function for DS mice. The team has already registered a patent, and hopes to see the treatment reach clinical settings to benefit children affected by this debilitating disease. They are also exploring its potential applicability to other genetic neurodevelopmental diseases. The developed platform represents a plug-and-play system for genetic therapies, with the possibility of incorporating different types of normal genetic material into the carrier virus for treating additional diseases in the future.

The study was the doctoral dissertation of Dr. Pnina Stern, under the guidance of Prof. Lilach Shalev-Mevorach of The Jaime and Joan Constantiner School of Education at Tel Aviv University. The encouraging results of the study were recently accepted for publication in the Journal of Attention Disorders.

“We developed the CPAT system years ago, and it produced good results in previous studies that we conducted, mainly in children," explains Prof. Shalev-Mevorach. "Furthermore, in the only study that we conducted in adults with ADHD, positive findings were obtained, but without indications of ‘far transfer,’ meaning an improvement in functions for which participants were not directly trained in the treatment.”

According to Prof. Shalev-Mevorach, it is challenging for researchers to make scientific claims about the effectiveness of non-medication treatments because it is difficult to compare them to a "non-medication placebo." In other words, when studying non-medication treatments, it's hard to distinguish the effects of the treatment itself from other factors like the attention participants receive during training sessions or the effort they put into the research. This makes it complex to determine the true impact of non-medication interventions.

Prof. Lilach Shalev-Mevorach

Students with ADHD Enrolled

In the current study, the team of researchers tried to resolve this by employing a research design that included two control groups: a regular control group, which performed the various assessment tasks at two points in time without any intervention as part of the research (the passive control group) and a second control group that participated in mindfulness training sessions under the guidance of a professional instructor. This type of training has yielded positive results in previous studies in people with ADHD.

For the experiment 54 students, male and female, diagnosed with ADHD were recruited from Tel Aviv University and other academic institutions. The subjects were blindly divided into three groups: a zero-intervention control group, a mindfulness group and a CPAT group.

Participants in the CPAT and the Mindfulness groups attended two-hour long group meetings on the University campus once a week, where the CPAT group received Computerized Progressive Attention Training and the mindfulness group received training from a certified mindfulness instructor.

Before and after the intervention protocol, the participants of the three groups performed a comprehensive series of assessment tests: standard computerized tests to assess attention functions, behavioral assessment questionnaires (self-reported ADHD symptoms), and mindfulness questionnaires (self-reported feelings such as stress, anxiety and well-being). In addition, a novel measurement was used for this intervention study, whose participants were, as mentioned, higher-education students: they were asked to read a text from a scientific article while their eye movements were tracked by an eye-tracker. The indices produced using the eye-tracking system made it possible to identify a pattern of inattentive reading, which was used as a measure of reading efficiency in an academic context. Finally, the participants filled out a questionnaire regarding their academic difficulties.

Improvements Maintained Over Time

Prof. Shalev-Mevorach says the results were very positive: “We saw improvements in the attention functions themselves, that is, ‘near transfer,’ for example in sustained attention, the ability to remain attentive for a long period of time, and in attention control, the ability to delay a routine response. But the main thing, is that we saw significant improvements in the participants’ daily and academic functioning, such as reduced repeated reading while reading a scientific article. Furthermore, the improvements in these attention functions were connected to the reduction in behavioral symptoms of ADHD and in repetitive reading."

"In other words, the CPAT trained the attention mechanisms themselves, and their improvement was related to the improvement achieved in behavioral symptoms and reading patterns. 33% of the participants who received the CPAT protocol showed a significant improvement in ADHD symptoms, compared to only 11% of those who underwent the mindfulness protocol. The improvements obtained were preserved in the testing that was carried out about four months after the end of the intervention protocol.”

Prof. Shalev-Mevorach notes that the effects of stimulant drugs (psychostimulants) such as Ritalin and Concerta are ‘on-off’: patients who take Ritalin daily enjoy significant improvements, but when they stop the treatment, the improvements fade, and they return to the starting point. She says the researchers wish to bring about "a profound change in basic attention functions, a change that will be significant in the long term, as an additional option alongside medication, and of course as an alternative to drug treatment in cases in which it isn’t applicable.”

The unique study, which was published in the leading journal Nature Neuroscience, involved an international collaboration led by Dr. Maya Geva-Sagiv (today at UC Davis). The study was a collaboration between the laboratories of Prof. Yuval Nir from the Sackler Faculty of Medicine, Department of Biomedical Engineering at The Iby and Aladar Fleischman Faculty of Engineering, and Sagol School of Neuroscience at Tel Aviv University, and Prof. Itzhak Fried from the Department of Neurosurgery at UCLA and the Sackler Faculty of Medicine at Tel Aviv University.

"Intervention during sleep represents a unique approach that can be further developed in the future to provide hope for people with memory impairments such as dementia." - Prof. Yuval Nir

The researchers (from left to right): Dr. Maya Geva-Sagiv, Prof. Yuval Nir and Prof. Itzhak Fried

"This study was made possible by a rare group of 18 patients with epilepsy at the UCLA Medical Center," says Prof. Nir. "Prof. Fried implanted electrodes in these patients' brains to try and pinpoint the areas that cause their epileptic seizures, and they volunteered to take part in a study investigating the effects of deep-brain stimulation during sleep. Close work with expert neurologists led by Prof. Dawn Eliashiv at UCLA enabled our team to integrate advanced brain stimulation in the research. Thus, we were able to test, for the first time in humans, the long-held hypothesis - that coordinated activity of the hippocampus and cerebral cortex during sleep is a critical mechanism in consolidating memories."

"Moreover, we improved memory consolidation through a special stimulation protocol that enhanced synchronization between these two areas in the brain. Intervention during sleep represents a unique approach that can be further developed in the future to provide hope for people with memory impairments such as dementia."

"In this study we directly examined the role of neural activity and electrical brain waves during sleep. Our goal was to enhance the natural mechanisms at play, to discover exactly how sleep assists in stabilizing memories." – Dr. Maya Geva-Sagiv

Unraveling Mechanism

"We know that a good night's sleep is critical for the consolidation of long-lasting memories, but so far, we had little evidence regarding the precise processes that are at work during human sleep," explains Dr. Maya Geva-Sagiv. "In this study we directly examined the role of neural activity and electrical brain waves during sleep. Our goal was to enhance the natural mechanisms at play, to discover exactly how sleep assists in stabilizing memories."

The researchers developed a deep-brain stimulation system that improves electrical communication between the hippocampus – a deep-brain region involved in acquiring new memories, and the frontal cortex – where memories are stored for the long term. By monitoring activity in the hippocampus during sleep, the system enables precisely timed delivery of electrical stimulation to the frontal cortex.

The study's participants completed two memory tests, and their performance was compared after two different nights – one undisturbed and one with deep-brain stimulation. On both occasions, they were asked in the morning to recognize famous persons whose pictures they had been shown the previous evening. The study found that deep-brain stimulation significantly improved the accuracy of their memory.

"To our surprise, we also discovered that the intervention did not significantly increase the number of right answers given by participants, but rather reduced the number of wrong answers. This suggests that sleep sharpens the accuracy of our memory…"   - Prof. Yuval Nir

Sharpening Memory Accuracy

"We found that our method had a beneficial effect on both brain activity during sleep and memory performance," says Prof. Fried. "All patients who had received synchronized stimuli to the frontal cortex demonstrated better memory performance, compared to nights of undisturbed sleep. The control group, which received similar yet unsynchronized stimuli, showed no memory improvement. Our deep-brain stimulation method is unique because it is close-looped – stimuli are precisely synchronized with hippocampal activity. In addition, we monitored the stimuli's impact on brain activity at a resolution of individual neurons."

"Our findings support the hypothesis that precise coordination between sleep waves assists communication between the hippocampus that takes in new memories, and the frontal cortex that stores them for the long term," adds Prof. Nir.

"To our surprise, we also discovered that the intervention did not significantly increase the number of right answers given by participants, but rather reduced the number of wrong answers. This suggests that sleep sharpens the accuracy of our memory, or in other words, it removes various distractions from the relevant memory trace."   

The study was supported by grants from the US National Institutes of Health (NIH), the European Research Council (ERC), the US National Science Foundation (NSF), the US-Israel Bilateral Science Foundation (BSF), and the Human Frontier Science Program (HFSP). The paper’s other co-authors are: Prof. Dawn Eliashiv, Dr. Emily Mankin, Natalie Cherry, Guldamla Kalender, and Dr. Natalia Tchemondanov of UCLA, and Dr. Shdema Epstein from Tel Aviv University.

The studies were led by Dr. Omri Bronstein and PhD students Rotem Zirler, Lisa-Maria Schmidt, Gal Eviatar, and Lachan Roth from the School of Zoology, at The George S. Wise Faculty of Life Sciences, and The Steinhardt Museum of Natural History at Tel Aviv University. The papers were published in Frontiers in Marine science and Royal Society Open Science.

The researchers underscore the vital importance of sea urchins, particularly the long-spined Diadema setosum, as keystone species essential for the thriving equilibrium of coral reefs. They express a pressing concern, stating, "It must be understood that the threat to coral reefs is already at an all-time peak, and now a previously unknown variable has been added. This situation is unprecedented in the documented history of the Gulf of Eilat."

According to the researchers' hypothesis, the cause of the deadly epidemic can be attributed to a pathogenic ciliate parasite that has spread from the Mediterranean to the Red Sea. In response to the gravity of the situation, an urgent report outlining the current state has been submitted to the Israel Nature and Parks Authority, instigating deliberation on emergency measures to safeguard Israel's coral reefs.

"Sea urchins in general, and Diadema setosum in particular, are considered key species essential for the healthy functioning of coral reefs. The sea urchins are the reef's 'gardeners' – they feed on the algae and prevent them from taking over and suffocating the corals that compete with them for sunlight." – Dr. Omri Bronstein

Dr. Omri Bronstein and a dying sea urchin

"At first we thought it was some kind of pollution or poisoning, or a local chemical spill, from the industry and hotels in the north of the Gulf of Eilat, but when we examined additional sites in Eilat, Jordan, and Sinai, we quickly realized that this was not a local incident," explains Dr. Bronstein. "All findings pointed to a rapidly spreading epidemic. Similar reports are coming in from colleagues in Saudi Arabia. Even sea urchins that we grow for research purposes in our aquariums at the Interuniversity Institute, and sea urchins at the Underwater Observatory Marine Park in Eilat, contracted the disease and died, probably because the pathogen got in through the pumping systems."

Dr. Bronstein describes it as a fast and violent death: "Within just two days a healthy sea urchin becomes a skeleton with massive tissue loss. While some corpses are washed ashore, most sea urchins are devoured while they are dying and unable to defend themselves, which could speed up contagion by the fish who prey on them."

Invasion and Vanishing Species

In recent years, Dr. Bronstein's research group has dedicated their efforts to the investigation of marine invasions, with a specific focus on the long-spined Diadema setosum. "Until recently, the black sea urchins with long spines, familiar to many of us, was one of the dominant species in Eilat's coral reef," reflects Dr. Bronstein. "Sea urchins in general, and Diadema setosum in particular, are considered key species essential for the healthy functioning of coral reefs. The sea urchins are the reef's 'gardeners' – they feed on the algae and prevent them from taking over and suffocating the corals that compete with them for sunlight. Regrettably, these once-thriving sea urchins have vanished from the Gulf of Eilat and are quickly disappearing from constantly expanding parts of the Red Sea further to the south," shares Dr. Bronstein with a sense of lament.

A dying urchin in the Mediterranean Sea (photo: Dr. Omri Bronstein)

Several months ago, Dr. Bronstein was alerted to the initial reports of widespread mortality by colleagues in Greece and Turkey, where the sea urchins had invaded, likely via the Suez Canal. "In 2006, the first sighting of this species of sea urchin occurred in the southern regions of Turkey," Dr. Bronstein adds. This phenomenon, known as biological invasion, carries far-reaching ecological implications, pervasively affecting the eastern Mediterranean, particularly along Israel's coastline. "We have been monitoring the dynamics of this species' invasion in the Mediterranean since its first emergence," he shares. 

In 2016, they discovered the first Diadema setosum sea urchin along Israel’s Mediterranean coastline – a lone urchin sighted at Gordon Beach in Tel Aviv. For over a decade since the first discovery in Turkey, the Mediterranean populations of these sea urchins remained small and usually hidden. However, since 2018 the sea urchin population in the Mediterranean has been growing exponentially, reaching a state of population explosion – with giant populations of thousands and even tens of thousands found in Greece and Turkey.

"The window of opportunity for preserving a thriving population of this species in Eilat has regrettably closed. To establish a safeguard population, we must act without delay, by preserving healthy individuals from the Israeli Mediterranean before the encroaching disease from the north reaches this region." Dr. Omri Bronstein

"However, during the course of our research, while scrutinizing the invasion of sea urchins in the Mediterranean, we began to receive reports on sudden extensive mortality," says Dr. Bronstein. "While the extinction of an invasive species is supposedly not a bad thing, we must be aware of two major risks: Firstly, we don't yet know how this mortality and its causes might impact local species in the Mediterranean. Secondly, and of far greater significance, the geographic proximity shared by the eastern Mediterranean and the Red Sea provides a potential conduit for the swift transmission of the pathogen into the Red Sea. As we feared and predicted, this is what appears to have happened."

Dr. Bronstein and his research team (photo: courtesy of Dr. Omri Bronstein)

A Reminiscent Crisis

The massive loss of sea urchins reminded the TAU researchers of one of the most devastating events in marine ecology: the disappearance of the sea urchins in the Caribbean. Until 1983, the Caribbean coral reef thrived as a vibrant tropical ecosystem, much like the one in the Gulf of Eilat. But as the sea urchins vanished, the uncontrollable growth of algae took over, blocking sunlight from reaching the corals and forever altering the reef into a sea of algae.

Dr. Bronstein reveals, "Just last year, the Caribbean experienced another outbreak of the disease, resulting in the demise of the remaining urchin populations. However, unlike previous incidents, we now possess advanced scientific and technological resources to analyze the forensic evidence. Researchers from Cornell University successfully pinpointed the cause of mortality in the Caribbean: a pathogenic ciliate parasite. The identical pathology observed in the dying sea urchins of Greece, Turkey, and the Red Sea corroborates this finding."

Dr. Bronstein's pioneering research not only identified the unprecedented mass mortality of an invasive species in the Mediterranean but also shed light on the alarming decline of the widely prevalent sea urchin species, Diadema setosum. In a groundbreaking study, Dr. Bronstein issued a a warning that the epidemic plaguing the Mediterranean could extend its reach to the nearby Red Sea. Sadly, this cautionary prediction has become a disheartening reality.

Urgent Measures and Closing Window

"The gravity of the situation cannot be understated: the Red Sea is witnessing an alarming surge in mortality, surpassing the extent observed in the Mediterranean. Looming in the background is an ominous uncertainty: What is the exact cause of the sea urchin die-offs? Is it the same Caribbean pathogen or an entirely new and unfamiliar factor? Regardless, it is evident that this pathogen spreads through water, and we anticipate a rapid escalation of sickness and demise among the entire population of these sea urchins in both the Mediterranean and the Red Sea."

"In my view, it is imperative that we swiftly establish a safeguard population for these sea urchins, ensuring the potential for their reintroduction into the wild. Similar to the approach taken with COVID-19, the trajectory of this epidemic remains uncertain. Will it eventually subside on its own, or persist for years, radically transforming coral reefs? However, unlike the COVID-19 pandemic, there are no available vaccines or treatments for the afflicted sea urchins. Hence, our efforts must be steadfastly directed towards prevention. The window of opportunity for preserving a thriving population of this species in Eilat has regrettably closed. To establish a safeguard population, we must act without delay, by preserving healthy individuals from the Israeli Mediterranean before the encroaching disease from the north reaches this region. While this is a complex undertaking, it is imperative if we aspire to secure the future of this unique species, which plays a critical role in the destiny of coral reefs," concludes Dr. Bronstein.

The breakthrough was achieved by a TAU research team led by Prof. Dan Peer of The Shmunis School of Biomedicine and Cancer Research, a global pioneer in the development of RNA-based drugs, Head of the Laboratory of Precision Nanomedicine, and TAU's VP for R&D; and by Dr. Sushmita Chatterjee, post-doctoral student from India at Prof. Peer’s lab, in collaboration with Prof. David Sprinzak of The George S. Wise Faculty of Life Sciences and Prof. Ronen Zaidel-Bar of the Sackler Faculty of Medicine. The study was funded by the Rivkin Foundation for Ovarian Cancer Research and the Shmunis Family Foundation. The results were published in the leading scientific journal Science Advances.

"The protein CKAP5 has never been studied with relation to the fight against cancer, simply because there was no known way to silence it," explains Dr. Chatterjee. "The lipid nanoparticles developed by Prof. Peer enabled us for the first time to silence this protein through targeted delivery of an RNA drug. We proved that CKAP5, a protein responsible for the cell's stability, can be silenced, and that this procedure collapses and destroys the entire cancer cell."

Prof. Dan Peer

"Something Like a Dominoes Game"

At the second stage of the study the researchers tested the new CKAP5-silencing RNA drug on 20 types of cancer. Some cancer cells proved more sensitive than others to this procedure. Cancers displaying high genetic instability, which are usually highly resistant to chemotherapy, were found to be especially sensitive to the silencing of CKAP5.

"As researchers, we are involved in something like a dominoes game: we always look for the one piece in the cancer's structure that is so important, that if we pull it out the entire cell will collapse. CKAP5 is such a domino piece, and we are already working on more applications (…)" - Prof. Dan Peer

"All cancer cells are genetically unstable," says Dr. Chatterjee. "Otherwise, they would be healthy, not cancerous. However, there are different levels of genetic instability. We found that cancer cells that are more unstable, are also more affected by damage to CKAP5.  Our drug pushed them to their limit, and essentially destroyed their structure. Our idea was to turn the trait of genetic instability into a threat for these cells, by using RNA to silence the flawed protein. We demonstrated for the first time that CKAP5 can be used to kill cancer cells, and then observed the biological mechanism that causes the cancer cells to collapse in the protein's absence."

Equipped with these insights, the researchers tested the new drug in an animal model for ovarian cancer, achieving a survival rate of 80%.

"We chose ovarian cancer because it's a good target," explains Prof. Peer. "While highly resistant to both chemotherapy and immunotherapy, this type of cancer is very sensitive to the silencing of CKAP5. It should be emphasized that the CKAP5 protein is a new target in the fight against cancer. Targeting cell division is not new, but using RNA to target proteins that make up the cell's skeleton (cytoskeleton) – this is a new approach and a new target that must be further investigated. As researchers, we are involved in something like a dominoes game: we always look for the one piece in the cancer's structure that is so important, that if we pull it out the entire cell will collapse. CKAP5 is such a domino piece, and we are already working on more applications, this time in blood cancers."