Small Miracles

FOR PEDIATRIC SURGEON Bill Peranteau ’97, this November afternoon starts with a success story. His first patient is an energetic preschooler wearing light-up Spider-Man shoes. When the little boy’s mother was pregnant, a routine ultrasound showed that he had a hole in the diaphragm, the muscle that separates the abdominal and lung cavities, which would permit abdominal organs to crowd out the lungs and threaten their development. When the child was still a newborn, Peranteau operated to close the hole. Now the family is back at the Children’s Hospital of Philadelphia (CHOP) for an annual follow-up visit, and the little boy is thriving; he has recently discovered the joys of jumping on the bed. 

Over the next couple of hours, Peranteau, an attending surgeon in CHOP’s division of general, thoracic, and fetal surgery, sees other children whose small bodies he’s repaired, sometimes when they were just days old. Under certain circumstances, when a structural defect could cause extensive or deadly damage before birth, he also performs surgery on fetuses — a previously unimaginable scenario pioneered at CHOP over the past few decades. But other kids remain beyond his reach — among them, those with genetic defects causing potentially fatal or severe health problems that begin and worsen during pregnancy. That’s where Peranteau’s current research interests come in. He and his colleagues are working to tackle those problems prenatally using CRISPR, the gene-editing technology that over the past several years has accumulated a growing list of possible applications, from creating hardier crops to eliminating disease-spreading mosquitoes to altering human DNA in pursuit of better health. 

There’s been a great deal of hype about CRISPR, and Peranteau doesn’t want to add to it. Speaking slowly and deliberately about his work, he describes two recent high-profile publications detailing successes in animal models as “very early proof of concept.” He’s right to be cautious: Plenty of experimental treatments have cured lab mice but not humans. Yet if things go as Peranteau hopes, surgically delivering CRISPR in utero could correct mutations in a single gene in the most serious of conditions — those that can’t be effectively treated after birth or that cause death or irreversible damage even in the womb. Fitting that general description are diseases of metabolism, neurodegeneration, and lung dysfunction. (The list of targets could be expanded if the concept proves safe and effective, he says.)

There’s a lot of experimental ground to be covered before that vision is a reality, and Peranteau, with his feet planted both in clinical practice and research, is in the thick of it. With these new technologies, “you have to have someone who’s the champion of the cause,” says Simon Waddington, professor of gene therapy at University College London. Peranteau “understands the science, and he sees the patients,” he says. N. Scott Adzick, the surgeon-in-chief at CHOP, calls him a “rare triple threat”: a stellar surgeon, researcher, and teacher. “He’s going to lead the field.”

Peranteau went to Princeton thinking he was interested in science; while there, that sharpened into a desire to become a physician. After doing lab work in immunology for his senior thesis in molecular biology, he decided to add research to the mix. He headed to the University of Pennsylvania, where he planned to get both M.D. and Ph.D. degrees.

During medical school, Peranteau opted to do surgery as the first of his clinical rotations, solely to get it out of the way. The stereotype of the brusque, arrogant surgeon didn’t appeal, and Peranteau originally thought he’d do his due diligence and move on to another specialty. But in the OR, he says he found a team-like atmosphere that recalled his years as a diver at Princeton. And he loved working with his hands and seeing the immediate impact of his work. He decided to pursue surgery and clinical research, scrapping his plans for a doctorate and instead doing two stints of focused research at CHOP’s Center for Fetal Research during his medical training. That experience, he says, gave him a model for his career aspirations: “doing very impactful research while at the same time doing major surgeries.” He was drawn to pediatric surgery in particular for the research questions it poses about abnormal development and for the chance to work with children and their families. It was at CHOP that he was exposed to fetal surgery, a still-developing field that was thought to be impossible just decades ago. 

WHEN THE IDEA OF OPERATING on a fetus was first proposed, it sounded “crazy” to many people, Adzick, a pioneer of the field, recalled in the 2015 PBS documentary series Twice Born. That’s in part because it exposes the mother to the risks of surgery even though she’s not the one who physically benefits. Early results didn’t show a benefit to the fetus, either. But as surgical techniques and criteria improved, so did outcomes. It’s now an option for a dozen or so conditions that pose deadly or devastating consequences to the fetus. 

Those conditions include carefully selected cases of the most common and serious form of spina bifida, a condition in which the fetus’s developing spinal column fails to form normally, leaving a hole in the back that exposes the spinal cord and nerves to damage from amniotic fluid and puts the baby at risk of serious motor and cognitive problems. To address the condition through fetal surgery, doctors operate on the mother to reveal the uterus. They use ultrasound to position incisions so as not to harm the baby or interfere with the placenta, then use uterine stapling to open the walls of the uterus just enough to expose the developing fetus’s tiny back and bottom. The surgeons sew up the hole in the back tightly enough to form a watertight seal, so that damaging amniotic fluid can’t get into the spinal cavity and cerebrospinal fluid can’t get out. The uterus is sewn up; the mother spends the rest of her pregnancy living close to CHOP; and the baby is delivered later by cesarean section. It’s delicate work, yet if all goes well, the surgery can be done in as little as an hour, thanks to a tightly organized team of surgeons, nurses, and other clinicians accustomed to working together, says Peranteau.

To be sure, prenatal spina bifida surgery — which unlike other fetal surgeries is done to improve quality of life rather than to save it — is not a guaranteed fix. Some damage has often occurred by the time surgery happens, and there are risks to both the mother and fetus, including premature birth. It doesn’t work for everyone. But a landmark trial led by Adzick and published in 2011 in The New England Journal of Medicine showed that the prenatal surgery led to a reduced need for shunting (inserting a tube into the skull to drain fluid) and improved motor outcomes at 30 months compared to surgery after birth. CHOP is one of the major centers for fetal surgery, and Peranteau and his colleagues counsel about 1,500 pregnant women a year, about 150 or 200 of whom end up having some kind of fetal surgical procedure, including removing tumors and placing shunts. Many of the others have a specialized delivery at CHOP and then surgery early in the baby’s life.

 As he took part in this new field of surgery, Peranteau became interested in an even more cutting-edge line of research: gene therapy, which involves replacing defective, disease-causing genes with healthy ones ferried into cells by a viral vector. It was a hot research area in the 1990s, but in 1999, an 18-year-old volunteer in a University of Pennsylvania trial of a treatment for a genetic disorder died due to an overwhelming inflammatory response against the modified cold virus that was used to transport the healthy gene into his body. Progress ground to a halt for years as researchers worked to make procedures safer. Slowly, the field came back, and these days gene therapy is again fertile ground for research, with hundreds of clinical trials accepting or planning to accept patients. “The delay, and the [current] excitement, are both appropriate,” says Peranteau. The U.S. Food and Drug Administration has already approved a handful of products, including a gene therapy for an inherited form of retinal blindness that was first developed by researchers at CHOP and the University of Pennsylvania and then by Spark Therapeutics, a biotech company spun off from CHOP. During his two research fellowships at CHOP, Peranteau studied using gene therapy in utero.

When he got his own lab several years later, a new, related technology caught his eye: CRISPR, which is short for “clustered regularly interspaced short palindromic repeats.” Unlike traditional gene therapy, which introduces new copies of healthy genes that don’t always integrate into the cell’s DNA, CRISPR seeks to change or edit the DNA, meaning changes would persist for the lifetime of the edited cell and be passed on to cells that arise from it. It also has the potential to be more precise. CRISPR was derived from the immune response of bacteria, and the original iteration has been joined by increasingly precise versions. 

This 2019 photo of CHOP’s “fetal family reunion” shows the children who have been cared for by the Center for Fetal Diagnosis and Treatment, with family members and staff. Peranteau is pictured at the right in the front row, with his son on his lap.

Center for Fetal Diagnosis and Treatment at Children’s Hospital of Philadelphia

Peranteau wants to use CRISPR as a tool to treat defects in a single gene prenatally, before irreversible damage accumulates. He describes several potential benefits of intervening at that stage. The small size of the fetus means smaller doses of the CRISPR components, so there’s a bigger bang for the therapeutic buck. Progenitor cells — rapidly dividing cells that are more differentiated than stem cells but still have the capacity to become one or more different cell types — are more abundant, potent, and accessible in a developing fetus. And because the fetal immune system isn’t fully mature, the chance of an immune reaction is lower. 

Peranteau emphasizes that this research is not the same as using CRISPR to genetically edit embryos in vitro, as was reported in China in late 2018. That research involved using CRISPR to edit the genes in a single cell, at the earliest stages of development, and then inserting the embryo into the uterus to continue its development. But editing the embryo so early means all the cells in the fetus will carry the change, including the egg and sperm cells — so changes are carried down to future generations. That, along with the fact that the editing wasn’t done to treat an existing disease, prompted widespread condemnation by the scientific community. In Peranteau’s vision, he would wait until the fetus is more fully developed and a disease is diagnosed, then treat only that affected organ. Edits would not carry down to any future offspring.

IN THE LAST TWO YEARS, Peranteau and his colleagues published two studies that showed their idea has promise. In one study, published in 2018 in Nature Medicine, Peranteau teamed up with researchers including Kiran Musunuru, a cardiologist at the University of Pennsylvania. Building on Musunuru’s expertise with heart disease and gene editing and Parenteau’s with fetal therapies, the researchers used CRISPR to do in fetal mice what Musunuru has studied for potential use in humans — targeting the liver to edit the PCSK9 gene into a form that lowers cholesterol levels. Musunuru “had already worked out protocols to treat after birth, so we used our model to see if we could get it to work in utero,” says Peranteau. The researchers operated on a pregnant mouse, accessed the developing fetus, and injected CRISPR components into a vein that leads to the liver, then returned the fetus and allowed the pregnancy to proceed as usual. It worked. About 15 percent of liver cells were edited, but that was enough to produce very low cholesterol in the baby — and then later, adult — mice. 

The experiment showed that the concept worked, but there’s no strong medical rationale for treating high cholesterol in the womb as opposed to in children or adults. So the researchers turned to a disease that was more clinically relevant. Certain mutations in a gene called FAH cause a rare metabolic disease called hereditary tyrosinemia type 1 that causes a buildup of the amino acid tyrosine, potentially leading to liver failure and cancer. It can be deadly unless treated with a drug and a strict diet. “Bill and I asked, what if we tried to cure this before birth?” says Musunuru. “It could be a one-time treatment, fix [the problem] permanently, and prevent early damage from happening.” They used CRISPR in fetal mice not to alter FAH itself, but to disable another gene, HPD, which stopped the disease process. 

This effort, too, worked; again, only a minority of liver cells were edited, but the positive results persisted as the mice aged. The mice that were treated prenatally not only survived but thrived better than the ones who had the medication after birth. Because there is a viable medication, however, it’s not clear that this particular disease is one to prioritize, says Peranteau. As with fetal surgery, researchers will first seek to treat the deadliest diseases for which there are no current options, perhaps something like the severe form of alpha thalassemia, a blood disease that usually results in death before or shortly after birth, he says.

In 2019, Peranteau, Musunuru, and colleagues published a study in Science Translational Medicine that used CRISPR to target a mouse model of a deadly lung disease. This time they surgically introduced it into the amniotic sac of the developing mouse, which then inhaled the CRISPR along with amniotic fluid. (The procedure was performed four days before the mouse pup’s birth, which is the equivalent of the third trimester in a human pregnancy.) All the mice that had not been treated died soon after birth. With treatment, however, the lungs showed less disease, and about 22 percent of mice pups survived. It’s another proof of concept, but it has relevance for a host of human lung diseases, including cystic fibrosis. Importantly, in both studies researchers reported no dangerous side effects for the mothers. 

Of course, many things will have to happen before CRISPR becomes part of Peranteau’s everyday arsenal. The scenario requires an early diagnosis, before the disease starts wreaking havoc. Noninvasive, blood-based fetal screening tests currently disclose certain chromosomal abnormalities but aren’t now used to look for the deadly mutations that CRISPR would target. That will likely change; Musunuru envisions that the kind of screening panels for specific harmful mutations that are now done in newborns eventually will be done prenatally. Also, CRISPR-based therapy in adults has only recently reached human clinical trials (for cancer, blood disorders, and a form of blindness), and delivering treatments to fetuses in utero is even more technologically and ethically complex. 

As with fetal surgery, there are two patients, even if only one needs help. “The mother is an innocent bystander,” says Peranteau, which means there is a very high bar for her safety. His research used adenoviruses to most efficiently deliver the CRISPR to the appropriate organ. But given the potential for a severe immune response, safer alternatives will need to be developed, he says. Those could include different viruses or nanoparticles. The next step, he says, is to try the approach on large animals to prove safety in both fetus and mom, even if the disease model isn’t directly applicable to humans. If all goes well, Peranteau estimates the approach could work for some diseases in some patients within a decade. 

Since 1997, CHOP has been holding annual “fetal family reunions” of families and kids who have been cared for by the Center for Fetal Diagnosis and Treatment. A lineup of photographs hangs in one of the hospital’s hallways. In the first photo, the babies and small children in laps are vastly outnumbered by parents and CHOP caregivers. By 2014 the photo session was moved outdoors to accommodate everyone. In 2019, there were 2,500 attendees; the photographer needed to perch high in a cherry-picker to capture the entire scene. If you look closely, you can see Bill Peranteau in the front row.

Peranteau doesn’t know what it’s like to have a child undergo lifesaving surgery before or after birth. But as a parent (he has two children with his wife, Jennifer Usas Peranteau ’99), he knows the anxiety provoked by threats to a child’s health, and says he hopes that allows him to be more patient and compassionate in his work. The reunion lets him see some of his former patients, many of whom are now able to run around, play in the bouncy house, and eat cotton candy just like kids who didn’t face serious problems so early in their lives. “It reminds me how fortunate we all are, to be a part of this group and have the trust of the parents and their families to help take care of their kids,” he says. “It’s awesome.”  

Katherine Hobson ’94 is a freelance journalist focusing on health and science.