Bench to Bedside

February/March 2017

Brain Biomarkers May Be Useful for Early Diagnosis of Autism


A gaze into your eyes. A smile back. A gurgle and babble. These are treasured moments between parents and their babies. Yet, for families with a high familial risk of autism spectrum disorder (ASD), these precious moments may be elusive. Researchers have long suspected that autism emerges in gradual ways during the first year of life, but it has been impossible to predict with confidence which high-risk infants are likely to be diagnosed with this complex developmental disability.

In a novel study conducted by the multi-center Infant Brain Imaging Study (IBIS) network that includes the Center for Autism Research (CAR) at Children’s Hospital of Philadelphia, scientists found a brain biomarker that could help to identify children with ASD earlier in life, with the help of a computer-generated algorithm. Their findings, reported in the journal Nature, suggest that brain changes may precede behavioral manifestations of ASD.

“The results of this study are a real breakthrough for early diagnosis of autism,” said Robert Schultz, PhD, who directs CAR and led the CHOP study site. “Currently, the earliest clinicians can diagnose a child with autism is around 2 or 3 years old, because that is when clinicians can observe the earliest behavioral presentations of autism. This research opens the door to be able to identify those needing intervention during the first year of life, before the full emergence of autism. Delivering early intervention offers hope that we can blunt the development of autism and set the stage to dramatically improve long-term outcomes.”

Earliest Detection of Enlarged Brain Size May Define the Core of Autism

Behavioral symptoms of ASD are characterized by difficulties in social interaction, verbal and nonverbal communication, repetitive behaviors, and restricted interests. By the time autism is diagnosed in the preschool years — on average around age 4 in the U.S. — their brains have already changed substantially and tend to be enlarged. Increased brain size was one of the earliest brain markers discovered in autism, but until now, brain size has only been studied in children and adults after the full onset of autism. The development of differences in brain anatomy before the first outward manifestations of autism appear has been a mystery.

“Brain imaging studies in older children have taught us a great deal about structural and functional differences in the way a child on the autism spectrum’s brain develops, but a key question has always been: What are the first changes in the brain, and how do neuroanatomical differences unfold during early development?” Dr. Schultz said. “These earliest developmental differences in the brain are likely the most important for understanding autism, for defining the core of autism.”

In the current study, the IBIS investigators from four sites in the U.S. used magnetic resonance imaging (MRI) to look for anatomical differences in brain development at 6 months, 12 months, and 24 months of age in 106 children at high risk for developing autism by virtue of having an older sibling with ASD. At each age, they measured brain volume and specific structural characteristics of the brain, in particular the thickness and surface area of the cerebral cortex in each of its many subregions. The researchers compared the measurements to a group of 42 low-risk infants without any family history of autism. The results showed that the babies who developed autism experienced much more rapid growth of the brain’s surface area between the ages of 6 and 12 months compared to babies who did not develop autism by 24 months of age. These findings suggest that the cerebral cortex expands first, and then enlarged brain size follows.

“The rate of cortical surface area expansion in the second half of the first year of life predicted with high accuracy which babies would later develop autism and which would not,” Dr. Schultz said. “This surface area expansion also helped us to understand why, on average, children with autism have bigger brains. Although brain size was not yet enlarged by age 1, the cortical surface growth rate from 6 to 12 months accurately predicted which children would show significantly larger brains at 2 years of age among the children who did develop autism.”

Honing in on Differences in ‘Social’ Areas of the Brain

While the entire cortical sheet expanded between 6 and 12 months, the greatest enlargement was confined to specific parts of the temporal and frontal lobes of the brain for children who were later diagnosed with autism. These brain areas are involved in language and nonverbal social understanding like facial expressions and complex social thinking. The researchers noted that these are the same brain regions that always show the greatest structural and functional differences in older children with autism.

Scientists do not fully understand why bigger is not better for the brains of children with autism. During normal development, there is significant pruning of neuronal connections called synapses that presumably enables improved cognition and social functioning, Dr. Schultz explained. It has been hypothesized that inefficient culling of connections that are not essential and perhaps not needed at all is the reason why brain size is enlarged in autism.

Applying ‘Smart’ Data to Neuroscience

The study team used a type of machine learning algorithm known as deep learning to establish their prediction. This process enabled more than 80 percent accuracy in predicting which babies would develop autism by age 2 and more than 95 percent accuracy in predicting which babies at high risk for developing autism would not develop autism.

“While the machine learning algorithm that predicted autism at age 2 drew upon many brain features, the prediction was largely carried by the growth rate of the surface area at 6 to 12 months,” Dr. Schultz said. “This was the single biggest part of our predictor for who is going to have autism at 2 years of age.”

On the Horizon: Translating Earlier Diagnosis to Earlier Intervention

The researchers envision that this kind of procedure might one day be part of routine clinical care for high-risk infants. However, the research results will first need to be reproduced in additional studies in order to be ready for clinical uses.

The community of researchers, clinicians, and families are excited by these findings because the ability to identify autism risk during infancy could lead to the development of valuable early interventions. Research has shown that children with autism who receive the earliest treatment tend to reap the most benefits.

For example, a specific social behavior called joint attention is impaired in children with ASD. Joint attention is the coordination of attention to objects between two people for the purpose of sharing. It’s motivated by the desire to share. Children with autism don’t readily initiate joint attention; they don’t use nonverbal cues to seek for others to attend to what is capturing their attention.

Joint attention emerges toward the end of the first year of life, and these interactions foster a common frame of reference critical for learning language. It is also a core building block for social development.  Joint attention training, which has been very successful in preschoolers and school age children, is one example of the kind of intervention that now might be used before autism emerges, Dr. Schultz explained.

In a related IBIS study that CAR investigators also were involved in, the study team used functional MRI scans to identify brain networks involved in initiation of joint attention. They evaluated the strength of connections between the brain’s vision, attention, and default modes to identify patterns of neural activity that also may allow for earlier diagnosis. Those findings appeared in the January issue of the journal Cerebral Cortex.

The IBIS study has been underway for a decade, and Dr. Schultz is thrilled that the study teams are on the cusp of reporting many more significant findings that also may have predictive value for earlier diagnosis of autism.

“We’ve known for a long time that ASD symptoms emerge over the first two years, but it has been difficult to find evidence of those symptoms in the first year of life,” said Dr. Schultz, who also is R.A.C. Endowed Professor, Department of Pediatrics, Perelman School of Medicine at the University of Pennsylvania. “These studies are hard to do because they take a long time, but now the results are coming in spades, and it’s wonderful.”

Researchers at the Carolina Institute for Developmental Disabilities at the University of North Carolina, which is directed by the Nature study’s senior author, Joseph Piven, MD, worked on the study. Other data collection sites included CHOP, the University of Washington, and Washington University in St. Louis. Investigators at McGill University, the University of Alberta, the College of Charleston, and New York University were involved in data analyses and interpretation.

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Peanut Power: New Guidelines Encourage Early Introduction of Peanuts Into Diets


Parents have feared peanut-containing foods for many years and excluded them from their infants’ diets, but new guidelines say it’s time to make friends once again with our favorite lowly legume. Yes, it’s true — peanuts are not nuts, after all. Now that we’ve completely turned your world upside down, here’s the science behind the latest recommendations.

Jonathan Spergel, MD, PhD, Allergy Section chief at Children’s Hospital of Philadelphia and professor of Pediatrics at the Perelman School of Medicine at the University of Pennsylvania, participated on the expert panel sponsored by the National Institute of Allergy and Infectious Diseases (NIAID) that issued the guidelines to prevent peanut allergy. Studies show the number of children living with peanut allergy tripled between 1997 and 2008, according to the national organization Food Allergy Research & Education. Peanuts, out of all food allergies, most often cause the reactions that lead to fatal food anaphylaxis, so people living with peanut allergy must be vigilant about ingredient labels on the foods they eat.

However, scientific evidence gathered through observational studies over the last decade has shown that early exposure to peanuts actually makes you less likely to have allergy, Dr. Spergel said. A large-scale randomized trial of introducing peanuts early into the diet called Learning Early About Peanut Allergy (LEAP) gave scientists even more food for thought. Results from LEAP reported in 2015 showed that regular consumption of a peanut-containing snack begun in early infancy and continued until age 5 reduced the rate of peanut allergy in at-risk infants by 80 percent compared to non-peanut-consumers.

Professor Gideon Lack at Kings College London got the idea to conduct the LEAP study based on an observation that a Jewish population in England who weren’t fed peanut products during infancy had a very high rate of food allergy compared to a Jewish population in Israel who used a popular teething biscuit made with peanut butter and had a very low rate of food allergy. He wondered if the Israeli babies’ early exposure to peanuts was training their immune systems not to overreact. The research, which was published in The New England Journal of Medicine, helped to spur the fundamental change in the latest peanut allergy prevention guidelines.

“Early is better,” Dr. Spergel said, summing up the new peanut paradigm. “Avoiding things probably leads to more allergy. The big challenge is that there is a food allergy phobia we will need to help parents get over so that they will let their kids eat foods with peanuts.”

The recommendations are tailored to three levels of infants’ risk for developing peanut allergy: high risk (infants who have severe eczema, egg allergy, or both); moderate risk (infants who have mild or moderate eczema); and low risk (infants who do not have eczema or any food allergy).

Parents should remember two other important peanut pointers: All babies should try other solid foods before peanut-containing ones. And infants and small children should never be given whole peanuts due to the risk of choking.

While these new guidelines apply to peanut allergy, Dr. Spergel suggested that the next big research question will be to figure out if other foods, such as milk and eggs, that are known to trigger allergic responses also can be introduced early into babies’ diets. And since, the LEAP study was performed with 600 children from England, the study needs to be replicated to demonstrate that the findings are translatable to a broader population.

One of the practical issues that the guidelines took under consideration, Dr. Spergel noted, is that some communities have a shortage of allergy specialists, which is partly why it was important for the expert panel to limit the allergy screening recommendation to those infants with the highest risk.

“These guidelines were written not only for the U.S. but worldwide, and for most of the rest of the world, even with just the high-risk group, there aren’t enough allergists to do the testing,” Dr. Spergel said. “Getting all those patients in early, hopefully within the first eight months of life, is a challenge.”

The expert panel was comprised of specialists from a variety of clinical, scientific, and public health arenas who used a literature review of food allergy prevention research and their own expert opinions to prepare the guidelines. They appear in the Journal of Allergy and Clinical Immunology, and resources including a summary for clinicians and a summary for parents and caregivers are available on the NIAID website.

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Oligodendrocytes Yield New Insights for Myelin Repair in Multiple Sclerosis


Cells with a funny name called oligodendrocytes make myelin in your central nervous system, which includes your brain, spinal cord, and optic nerves. “Oligo” means “small.” “Dendros” are “branches” that come out of a cell. And “cytes” are “cells.” So oligodendrocytes are very small cells with many lacey arms. The lab of Judith Grinspan, PhD, research professor of Neurology at Children’s Hospital of Philadelphia and the Pereleman School of Medicine at the University of Pennsylvania, has a particular expertise in starting oligodendrocytes as precursor cells in a lab dish and observing their maturation to better understand the myelin-making process. Her study team’s observations are yielding clues on how to encourage myelin to regenerate in people who have multiple sclerosis.

Oligodendrocytes’ arms, also known as processes, produce the myelin sheath, which is a membrane rich in lipids, or fat, that coats our nerves and allows for conduction of nervous impulses. Myelin functions like the insulation on an electrical wire. If the insulation is not intact, the electricity does not get where it needs to go. So, if part of your myelin sheath is missing, the messages sent by nerves in your brain — telling your big toe to wiggle, for example — don’t reach your muscles.

Myelin is destroyed in multiple sclerosis, and depending on which areas of the brain have demyelination, patients may experience various symptoms, such as movement problems, vision loss, and fatigue. About 450,000 people in U.S. and Canada are living with multiple sclerosis, and studies suggest that 2 to 5 percent have a history of symptom onset before age 18.

While the cause of multiple sclerosis remains unknown, it is considered to be an autoimmune disease, in which the immune system recognizes the central nervous system as foreign and attacks the myelin. Researchers have helped to identify drug therapies for multiple sclerosis that target the immune system; however, little progress has been made on finding factors at the cellular level that could improve myelin repair.

“Researchers like me who work on myelination want to know everything about how these oligodendrocytes tick and how they make myelin,” Dr. Grinspan said.

In a paper published in January by the Journal of Neurochemistry, Dr. Grinspan and her colleagues reported on a novel insight they gained about this process, albeit in a serendipitous way. They were working with Kelly Jordan-Sciutto, PhD, chair and professor of Pathology at Penn’s School of Dental Medicine, on a study looking at the effect of the antiretroviral drugs that are used to treat HIV on oligodendrocytes and myelin. The study team noted that one of the drugs dysregulated a protein called sterol regulatory element binding protein (SREBP), which is thought to control the synthesis of fat all over the body. SREBP raised the investigators’ curiosity even higher when they realized that no one had ever studied it in oligodendrocytes.

Hubert Monnerie, PhD, a post-doc in the department of Neurology who at the time was working with Dr. Grinspan, performed a series of lab experiments that proved SREBP indeed was present on the oligodendrocytes, and levels of SREBP increased as the cells matured. Next, he explored SREBP’s role in the cells by using a chemical to block the SREBP. When SREBP was inhibited, the oligodendrocytes did not form new myelin.

“What we saw was really a remarkable phenomenon,” Dr. Grinspan said. “When we inhibited our favorite protein, SREBP, there were no processes. They were gone. It’s as if the cells curled them up instead of extending them. We saw it all over the dish. And then we wanted to see if this was reversible, and sure enough, if we washed out the blocking agent, the processes were back.”

The investigators performed more preliminary experiments, with the assistance of John Millar, PhD, director of the Metabolic Tracer Resource at Penn, that suggest SREBP controls the synthesis of cholesterol in oligodendrocytes that is necessary for the cells to make a proper myelin sheath. This is an important finding, not just because it provides new insights about how oligodendrocytes form myelin. Dr. Grinspan pointed out that on the therapeutic side, one idea being discussed in the multiple sclerosis field is the use of a class of drugs called statins because they have anti-inflammatory properties; however, statins also lower cholesterol, which could perhaps hinder lipid synthesis in oligodendrocytes and myelin regeneration.

Future research is needed to better understand exactly how SREBP may act as a lipid control factor in oligodendrocytes and if the protein has other fundamental effects on the cells’ development. With the help of a grant from the National Multiple Sclerosis Society, Dr. Grinspan is working to establish an animal model in which SREBP is inactivated in oligodendrocytes. This may reveal novel ways to create conditions in the brain that are beneficial for myelin repair and ease symptoms of multiple sclerosis.

“We’ll learn more about all the factors that you need to have in one place to promote myelination,” Dr. Grinspan said. “I am extremely grateful to the Multiple Sclerosis Society for their support.”

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CHOP Named Rett Syndrome Clinical Research Center of Excellence


Although they aren’t able to talk, girls with Rett syndrome have 14 Rett Syndrome Clinical Research Centers of Excellence who are speaking up for them by accelerating research through a natural history study and other projects aimed at improving care and finding a cure. One of those Centers of Excellence, which were designated by the nonprofit organization, is at Children’s Hospital of Philadelphia, under the leadership of Eric Marsh, MD, director of the Neurogenetics Program at CHOP.

“It’s an honor to be chosen, and it shows that we’re doing good work,” said Dr. Marsh a pediatric neurologist with expertise in neurogenetic conditions and genetic epilepsies at CHOP and assistant professor of Neurology at the Perelman School of Medicine at the University of Pennsylvania. “There is a lot of research activity going on that we hope to be a part of into the future to really see a change for the care of these girls and kids with related diseases too.”

In 1999, a research team at Baylor University in Waco, Texas, discovered that mutations on the MECP2 gene, located at the Xq28 site on the X chromosome, cause Rett syndrome. It’s a chance mutation that happens in DNA, and children almost never inherit the faulty gene from their parents. Rett syndrome occurs in an estimated one of every 10,000 female births.

Researchers suggest that MECP2 provides instructions for making a protein called MeCP2, which has an important role in maintaining neurons responsible for cell-to-cell communication and normal brain function. Children with Rett syndrome have a deficiency of MeCP2 protein. While basic scientists have revealed the genetic underpinnings of Rett syndrome, they don’t yet fully understand how mutations in the MECP2 gene disrupt the function of neurons and other cells in the brain and lead to the unique pattern of symptoms seen in the disease. Related disorders are caused by duplications of MECP2 (MECP2 duplication syndrome) and mutations in the CDKL5 or FOXG1 genes.

Girls with Rett syndrome usually seem to be developing normally until symptoms appear between 6 and 19 months of age. Their communication skills regress, and they lose the functional use of their hands because they constantly rub or wring them. They also tend to have trouble walking and have a small head size. Other problems include seizures, disorganized breathing, scoliosis, and sleep disturbances.

Despite having the same MECP2 mutation, girls with Rett syndrome can appear quite differently because the disease has a great deal of clinical variability. For example, when Dr. Marsh sees patients with Rett syndrome in his clinical practice, some girls are able to walk and express themselves with their eyes and use augmentative communication devices, while others are immobile and noncommunicative. Knowing the path someone with Rett syndrome generally follows if they have milder or more severe forms of the disease is one of the goals of the Rett Syndrome, MECP2 Duplication and Rett-Related Disorders Natural History Study that CHOP is participating in.

“By gathering all this natural history data, we hope to have a good baseline from which to compare people to, especially as research gets closer to finding possible treatments,” Dr. Marsh said.

A subproject of the natural history study that Dr. Marsh launched in January with the help of Timothy Roberts, PhD, vice chair of research in the department of Radiology and professor of Radiology at Penn, is taking a close look at evoked potentials in patients with Rett syndrome. Evoked potential tests are harmless and involve electrodes placed on the scalp to measure the electrical activity of the brain in response to stimulation of specific sensory nerve pathways. In this study, the researchers will focus on visual and auditory evoked potentials to analyze the spectrum of brain waves in girls with Rett syndrome and identify biomarkers of the disease.

“We know from some preliminary studies that the way the brain responds in Rett syndrome is different than in healthy, age-matched individuals,” Dr. Marsh said. “We’ll be testing evoked potentials at different time points with children with different severity of illness to see if we can see any changes in the evoked potentials that would predict or demonstrate a change in severity or change over time. Our hypothesis is that the signature will be more different and get worse over time. If that does happen, you can imagine that if you give a patient a particular drug and you see the signature go closer toward the normal response, then it would suggest that your intervention is working.”

CHOP and four other sites have begun to enroll 120 girls with Rett syndrome, and they plan to follow them yearly for at least three years. The study participants, ages 2 to 15, will be grouped into different age brackets. For the visual evoked potentials, the girls will be placed in a comfortable chair and watch a checkerboard on a big screen. The spaces will flash in a repeated pattern, and the researchers will monitor how the girls’ brains respond to that stimuli. Testing the girls’ auditory evoked potentials involves them wearing headphones and listening to a series of tones while they are watching a movie without sound.

So far, the investigators have received an exceptionally favorable response from study participants, according to Dr. Marsh, who gives much of the credit to excellent patient advocacy groups like for emphasizing the importance of families being at the forefront of research that has the potential to improve their daughters’ quality of life. Moving forward, CHOP also will be involved in a clinical trial of a pharmaceutical agent for Rett syndrome and a blood-based biomarker study.

In an announcement of the Centers of Excellence,’s Chief Science Officer Steve Kaminsky, PhD, stated, “As a rare disease, the Rett community is fortunate to have these 14 Clinical Research Centers of Excellence. The clinicians, nurses, and the medical team are deeply committed to the care of individuals with Rett syndrome, while being heavily involved in clinical research through the Rett Consortium and Natural History Study. These clinics understand our mission and make it possible to accelerate research and empower families.”

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Ring Chromosome Study to Address Neurodevelopment and DNA Looping


Picture a human chromosome. Chances are, the image in your mind looks like a tiny rod in the shape of a letter “I” cinched in at the waist. Or, if you are picturing a chromosome in the process of copying itself for cell division, it looks like a letter “X” or “Y.” But what if one of those tiny letters, each of which is actually a single long string of DNA coiled and bundled up, was tied to itself end-to-end to make an “O”? This is exactly what happens in a rare and mysterious group of cytogenetic disorders known as ring chromosome syndromes.

“Seizures are the big problem for these kids,” said Nancy Spinner, PhD, chief of the division of Genomic Diagnostics at Children’s Hospital of Philadelphia and professor of Pathology and Genetics at the Perelman School of Medicine at the University of Pennsylvania. Dr. Spinner was awarded a grant from the National Institute of Neurological Disorders and Stroke (NINDS) to develop a neurological model for studying Ring 14 syndrome to better understand how gene expression changes as a result of the chromosome’s changed shape.

Amid her other full-time projects, Dr. Spinner has studied ring chromosomes since 2008 with funding from philanthropic organizations including Ring 14 International and the Ring Chromosome 20 Foundation. The NINDS grant will allow her lab to dig deeper into how these chromosomes cause impairments in children. In addition to experiencing seizures from a young age, children with Ring 14 syndrome also have intellectual disability and developmental delay.

Working in partnership with Co-Principal Investigator Stewart Anderson, MD, and collaborators Jason Mills, PhD, Laura Conlin, PhD, and Deborah McEldrew, the team is inducing the growth of cells from Ring 14 patients from induced pluripotent stem cells into neuron cells. They are using a method that matures the cells without causing them to divide and reproduce. This approach overcomes a challenge with using stem cell methods that require reproduction: Stem cells in vitro tend to self-correct the ring chromosome by losing it and replacing it with a duplication of the other copy of chromosome 14. The researchers are also attempting to develop an in vitro cerebral cortex organoid model to attempt to understand how embryonic brain development is altered by the ring chromosome.

This exploratory work should help identify which questions are most promising for further study to someday move toward targeted therapies for Ring 14 syndrome. Current knowledge is limited.

“We can see the rings, we can map the rings, we know what their gene content is, and we know that they don’t grow very well,” Dr. Spinner said. “We know some facts about them, but we don’t yet know enough about specific pathways or specific genes, so that it’s not clear how we would target them. It hasn’t been a simple answer so far.”

While these first fundamental steps toward future targeted therapies for Ring 14 syndrome are important, another, more immediate possibility in this project is that it could offer new insights into emerging scientific questions about how the physical folding and looping of DNA influences genes’ expression.

This concept of the “chromatin landscape” or “loopscape” is built on the fact that DNA inside the nucleus of a cell does not normally look like an alphabet soup of I-shaped rods — with a rare “O” in the case of ring chromosomes. Those shapes are only formed for cellular reproduction purposes as a kind of travel packaging. In the normal life of a cell, each long string of DNA is unraveled from the rod shape and re-coiled into looser bundles.

“If you take a ball of string and just ball it up in your hand, before the cat gets to it, you might think, well it’s random, which little piece of the string is going to be right next to another piece of the string, which, when pulled apart, could be a meter away,” said Dr. Anderson, a research psychiatrist at CHOP and associate professor of Psychiatry at the Perelman School of Medicine. “But it turns out that this kind of bundling of DNA is not random, and beyond that, it’s highly regulated and very specific.”

If two ends of that ball of string are tied together, as in a ring chromosome, the researchers hypothesize that that regulated and specific organization could be disrupted. Detailing the nature of those disruptions could not only help explain the mechanism of disease impairments in ring syndromes, but it could also help scientists understand more about the way DNA packaging influences gene expression in normal chromosomes.

The researchers also hope to bring more attention to ring chromosomes to rally the attention of other scientists to help.

“The families are eager to get attention because their children have serious needs and the conditions are so rare,” Dr. Spinner said. “Part of what I feel like we’re doing when we talk about this among scientists and clinicians is proselytizing. We’re trying to expose people to an interesting problem.”

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Kassa Darge, MD, PhD, Expands What Ultrasound Contrast Agents Can Do


The Food and Drug Administration’s latest approval of a contrast agent for use in ultrasonography of the urinary tract in pediatric patients marks the end of an important chapter in the career of Kassa Darge, MD, PhD. A new one, though, has just begun. In December, Dr. Darge was named chair of the Department of Radiology and Radiologist-in-Chief at Children’s Hospital of Philadelphia, following an extensive national search.

For years, Dr. Darge has strived to obtain FDA clearance for new and expanded indications of Lumason (known globally as SonoVue® and manufactured by Bracco Diagnostics), by presenting his research to the agency and increasing awareness among his peers. Even his PhD work focused on contrast agents for intravesical application.

“It’s just amazing,” said a proud Dr. Darge of the FDA’s decision announced in January.

The agent now can be used to evaluate suspected or known vesicoureteral reflux (VUR) in children. VUR is a condition when urine flows up the ureters back toward the kidneys where the urine came from. VUR allows bacteria that may be in the bladder to travel with the refluxing urine to the kidney, which can cause recurrent urinary tract infections and renal damage.

Conventional methods to detect kidney reflux include fluoroscopy and scintigraphy. This FDA approval now gives physicians greater access to a modality that eliminates radiation. In addition, a contrast agent, basically microbubbles significantly smaller than red blood cells (1 mL of suspension of Lumason contains roughly 600 to 800 million microbubbles), improves the echogenicity of a patient’s blood or urine and results in better visualization and assessment.

This particular approval of Lumason “opens the floodgates” for research in other related areas, said Dr. Darge, who served as the pediatric radiology department chair at the University of Wuerzburg in Germany before coming to CHOP in 2007. “More indications need to be studied — each and every one.”

Fortunately, he hasn’t encountered any lack of excitement from fellow researchers looking to collaborate. “I get so many emails now,” he said with a laugh. “It’s so overwhelming.”

To further spread the word of the potential, he will be presenting on ultrasound contrast this year at the meetings of the Society of Pediatric Radiology, American Institute of Ultrasound in Medicine, and the International Contrast Ultrasound Society.

One area of interest for him zeros in on ultrasound contrast and focal lesions of the liver, which signal a possible mass. When conventional ultrasound is performed, physicians see these either as focal “discoloration” of the liver and/or distinct mass.

“Many times, just from the grayscale and Doppler ultrasound images, we cannot tell what these lesions are,” Dr. Darge explained. “When we inject ultrasound contrast agent, we have the opportunity to witness how blood flows in and out of the lesion, if at all. From the pattern of enhancement or no enhancement and washout, we have more information to classify the focal lesion in the liver.”

Similarly, future research will investigate focal lesions in the kidneys, as well as evaluation of the bowel in patients with inflammatory bowel disease, such as Crohn’s disease. Ultrasound contrast will allow for a better depiction of the inflamed bowel to gauge the severity.

“We will be able to decide if it’s fibrotic or just inflamed,” Dr. Darge said.

Another practical application will examine the benefits of ultrasound contrast during hip dislocation surgery, he said. The standard (and lengthy) procedure brings the patient out of the operating room to the magnetic resonance imaging machine (MRI) to check for proper alignment and to confirm blood flow isn’t compromised. Using ultrasound contrast in the operating room streamlines the process for everyone.

“If there is any problem, you can correct it then and there,” Dr. Darge said.

While CHOP leads the charge with ultrasound contrast research, Dr. Darge also heads investigation efforts in several other areas, including ultrasound elastography for evaluation of the liver and spleen. Elastography is a method to assess the mechanical properties of tissue. The spleen’s stiffness has been shown to correlate with the presence and degree of portal hypertension, he said. Further avenues of ultrasound elastography research involve the evaluation of the bladder to see if patients with neurogenic or abnormal bladder function have possible fibrotic change in the bladder wall.

“We also have started to look at the utility of elastography in boys with varicocele with the aim of possibly finding a parameter to help with the decision for surgery and its timing,” Dr. Darge noted.

And with magnetic resonance elastography, he’s systematically evaluating patients born with autosomal recessive polycystic kidney who develop liver fibrosis. Unlike ultrasound, magnetic resonance elastography allows for the evaluation of the whole liver, said Dr. Darge, adding he’s mentoring other research projects that compare the utility of positron emission tomography-MRI to the more established method of positron emission tomography-computed tomography in children.

All this work will continue to add to Dr. Darge’s impressive research portfolio, which encompasses nearly three decades with more than 200 publications and numerous grants. He first gained an interest in radiology while practicing tropical medicine in West Africa. Dr. Darge spent several months collaborating with the University of Heidelberg pediatric radiology department on the utilization of ultrasound to monitor drug effects on filarial worms, a creature that causes onchocerciasis (river blindness). Dr. Darge’s work on the project impressed the chief of radiology, and he offered him a radiology residency soon after.

Now, years later, Dr. Darge still has the same excitement and passion about improving patient outcomes with his research. Dr. Darge is also a professor of Pediatrics and professor of Radiology in Surgery at the Perelman School of Medicine at the University of Pennsylvania.

“I enjoy seeing how such advances facilitate and make easier diagnostic imaging in children,” he said.

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Powering Up Mitochondria Could Boost Military and Civilian Health


For a month in 1962, India and China fought a war in the Himalayan highlands. In the high-altitude, frigid conditions, physical endurance was at least as much of a threat to the soldiers on each side as the opposing army. Perhaps one reason that China won is that many of the soldiers on their side were native to the region and well conditioned to handle the cold and low-oxygen environment. If you could look inside the cells of those soldiers’ lungs and other tissues, perhaps these soldiers had an arsenal of microscopic secret weapons that made their physical endurance possible.

Those secret weapons are mitochondria, according to Liming Pei, PhD, and Douglas Wallace, PhD, of the Center for Mitochondrial and Epigenomic Medicine (CMEM) at Children’s Hospital of Philadelphia and Pathology and Laboratory Medicine at the Perelman School of Medicine at the University of Pennsylvania. In the early 1990s, Dr. Wallace became interested in studying whether mitochondria — the organelles inside cells that generate 90 percent of the body’s energy — were responsible for different populations’ abilities to withstand high-altitude conditions. He and several U.S. colleagues partnered with the Third Military Medical University in China to analyze the DNA within the mitochondria (mitochondrial DNA), comparing samples from people native to Tibet in the Himalayas with those from lowland Han Chinese populations.

They found a variant in the mitochondrial DNA that was frequent among Tibetans and rare in Han Chinese — a tantalizing clue that this variant was under positive selection in the highlands, and that mitochondrial function could make a difference in performance in that setting, perhaps even tipping the balance in a battle.

Dr. Pei and Dr. Wallace are now returning to the study of mitochondrial function in a military setting with a grant from the U.S. Army. Their powerful collaboration builds on the strength of both scientists’ labs. Dr. Pei specializes in the study of how mitochondrial genes are transcribed and function within the cell, and Dr. Wallace is an expert in mitochondrial genetics and mitochondrial diseases and a founder of the field of mitochondrial medicine.

“Mitochondrial disease affects many parts of the body, and the parts most affected are the brain, heart, and skeletal muscles, because those parts of the body have a lot of mitochondria and use a lot of energy,” Dr. Pei said. “For proof of principle, we are focusing on the heart because you can evaluate cardiac function fairly well in animal models.”

Their project could help improve mitochondrial function for the benefit of U.S. service members and their families, veterans, and civilians, including children and adults with mitochondrial diseases. Mitochondrial DNA is distinct from the DNA in the cell’s nucleus. Mitochondrial diseases are inherited conditions caused by a number of different mutations in mitochondrial DNA and in nuclear genes that are involved in the functions of mitochondria.

Over the last few years, Dr. Pei identified a family of transcription factor proteins to be essential for the production of mitochondria and of proteins involved in energy generation in neurons (ERR gamma) and heart cells (ERR alpha and ERR gamma).

“Our idea was that, if you increase the level of these particular proteins, that would cause the cell to make more mitochondria, and that might in fact then increase the energy output of the cell and make the cell healthier,” Dr. Pei said.

While other efforts to develop therapies for mitochondrial diseases take a precision approach, Drs. Wallace and Pei aim to power up mitochondria broadly, regardless of the underlying mutation that might cause dysfunction or disease. This is a plausible option because patients with mitochondrial disease often have a combination of some damaged mitochondria and some healthy ones. If their method increases the number of healthy mitochondria or increases healthy proteins to aid the function of unhealthy mitochondria, the net effect could be improvements in energy production.

Their study uses three different preclinical models of cardiomyopathy due to mitochondrial dysfunction. Two are mouse models with different mitochondrial mutations that result in cardiomyopathy. The third is induced pluripotent stem cells derived from a large mitochondrial cardiomyopathy lineage that their collaborator, Xilma Ortiz-Gonzalez, MD, PhD, of the division of Pediatric Neurology at CHOP, has successfully grown into human cardiomyocytes (heart cells).

They will test gene therapy approaches to increase the expression of the ERR alpha or ERR gamma transcription factor and pharmacologic interventions to increase that protein’s activity. In their proof-of-principle approach, they hope to show that increasing these proteins boosts the number and function of mitochondria therapeutically for cell and animal models, and potentially to show the safety of the approach in animals.

If the principle proves successful, it opens the possibility that this type of method for boosting mitochondrial function could benefit many people with mitochondrial dysfunction, since mitochondrial defects are being associated with a broad range of common diseases such as diabetes, obesity, and neurological diseases. For military personnel and veterans, these approaches might ameliorate some of the negative effects of conflict toxicity such as exposure to Agent Orange and conditions such as Gulf War syndrome. Improving mitochondrial function could benefit people who are not sick — such as soldiers fighting in the mountains.

“It would be amazing, for instance, if we could use ERR alpha/gamma as a preventative therapy for high altitude or even just increase the mitochondrial level of people, making them more resistant to stress, for instance, able to exercise longer, or able to recover from toxins,” Dr. Wallace said. “There are many potential applications long down the road.”

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Philadelphia Coalition for a Cure: Re-defining Cancer Research Through Collaboration


The newly launched Philadelphia Coalition for a Cure (PC4C) received a warm welcome at Children’s Hospital of Philadelphia in honor of World Cancer Day. Five regional cancer academic treatment centers and two pediatric hospitals met hospitals met Feb. 3 to announce the collaboration that is the nation’s first city-wide brain tumor precision-medicine research partnership to benefit both adult and pediatric brain tumor patients. In time, the PC4C plans to expand treatment to include additional tumor and cancer types.

“Together we’re embarking on an incredibly exciting journey toward revolutionizing cancer care,” said Jay Storm, MD, chief of the division of Neurosurgery at CHOP.

The PC4C is a first-in-kind cooperative clinical diagnostics and research initiative focused on assessing and developing leading-edge technologies and diagnostic platforms through shared initiatives to benefit adult and pediatric brain tumor patients. The collaboration is committed to streamlining research and precision medicine efforts. They will work together and with commercial partners and payers to advance data-driven discovery through the rapid sharing and release of data to the entire research community through open science initiatives.

“Brain tumors are the leading cause of disease-related death in children and more than 20,000 adults are diagnosed each year,” Dr. Storm said. “Working with PC4C, we hope to define a new collaborative clinical and research ecosystem that harnesses partnerships among leading academic centers, commercial partners, and insurers to identify therapies and accelerate discovery.”

PC4C is empowering data-driven discovery and improving treatments for brain tumors through therapies that are individually tailored and specific to the biology of each patient’s tumor, young or old, with the aim of reducing toxic side effects and increasing the therapeutic effectiveness of targeted approaches. This new targeted approach will provide researchers with access to GPS Cancer, a comprehensive molecular profiling and diagnostics test which integrates whole genome (DNA) sequencing, whole transcriptome (RNA) sequencing, and quantitative proteomics to provide oncologists with a comprehensive molecular profile of a patient’s cancer. This clinical test, funded in-part by grants to Children’s Hospital of Orange County and supplied by NantHealth Inc., can inform personalized treatment strategies and identify therapies that may have clinical benefits for the patient.

Healthcare insurance provider Independence Blue Cross (IBC) is also at the forefront of this new initiative, by providing full-coverage for GPS Cancer testing of all IBC-insured patients at each of the participating PC4C member institutions. For non-IBC insured brain tumor patients, additional grant support will be provided to cover the cost for the GPS Cancer test.

All patient-consented data for the PC4C will be accessible to the research community via Cavatica, a biomedical data analysis and storage platform that, for the first time, will integrate adult and pediatric brain tumor data. These efforts leverage the recently launched Children’s Brain Tumor Atlas initiative, another large-scale Cancer Breakthrough 2020 data initiative.

“The PC4C is made up of many individuals and institutions who are coming together to change the way we care for cancer patients of all ages by changing the culture to one of openness, sharing, and collaboration,” Dr. Storm said. “That is going to lead to new innovations that will occur quickly. One of the mandates of the PC4C is that all the data has to be shared immediately, in real time. There is going to be no data embargo. As soon as it’s generated, it’s going to be made available to the entire scientific community in a protected cloud environment that anyone can access. This kind of sharing is clearly a paradigm shift for academic medicine.”

PC4C member institutions include the division of Neurosurgery and Center for Data Driven Discovery at CHOP, Lewis Katz School of Medicine at Temple University, The Perelman School of Medicine at the University of Pennsylvania, Sidney Kimmel Medical College at Thomas Jefferson University, Cooper Medical School of Rowan University, Drexel Neurosciences Institute at Drexel University College of Medicine and The Hyundai Cancer Institute at The Children’s Hospital of Orange County.

“There have not been many breakthroughs for children with brain tumors in many, many years, and the Philadelphia Coalition for a Cure is going to change that for all of us,” said Madeline Bell, CEO of CHOP. “We can really make a difference for all of our patients.”

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Produced by The Children’s Hospital of Philadelphia Research Institute.

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