An in utero stem cell transplant for alpha thalassemia

Stephanie joined Drug Discovery News as an Assistant Editor in 2021. She earned her PhD from the University of California Los Angeles in 2019 and has written for Discover Magazine,...

When soon-to-be parents meet with Billie Lianoglou, a genetic counselor at the University of California, San Francisco (UCSF), it's often the first time they’ve ever heard of alpha thalassemia. The recessive genetic disease arises from the deletion of the genes coding for alpha globin, a vital component of both fetal and adult hemoglobin.

These fetuses get sick in the pregnancy. When [they’re] not making red blood cells that work, the fetuses become hypoxic. They don't have oxygen. The oxygen isn't going to the developing bone and tissue. – Billie Lianoglou, University of California, San Francisco

Healthy people typically have four copies of the alpha globin gene — two from mom and two from dad. If people are missing one or two copies of the gene, they appear perfectly fine, but they are slightly anemic. Lacking two alpha globin genes does confer protection against malaria, so carrying this alpha globin deletion is common in places where malaria is endemic, including Africa, Southeast Asia, southern China, the Middle East, and some Mediterranean areas (1). But, if a fetus lacks all four alpha globin genes, they cannot make functional hemoglobin.

These fetuses get sick in the pregnancy. When [they’re] not making red blood cells that work, the fetuses become hypoxic. They don't have oxygen. The oxygen isn't going to the developing bone and tissue. – Billie Lianoglou, University of California, San Francisco

"Often, when families would receive this diagnosis, the first thing that they would hear from their providers is this is a fatal condition," said Emma Canepa, a clinical trial program manager at UCSF and one of Lianoglou's colleagues.

Supplying the fetus with working red blood cells, however, can prevent this outcome. Over the past thirty years or so, doctors have increasingly given fetuses with alpha thalassemia healthy red blood cells via in utero transfusions (IUT), allowing them to survive to birth. But not all doctors know about this treatment option, and many who do worry about how IUT might affect the child's future development. Furthermore, IUT only treats the symptoms of alpha thalassemia; when the children are born, they still need monthly blood transfusions for the rest of their lives.

With expertise in maternal-fetal medicine and alpha thalassemia in particular, researchers at UCSF and their collaborators have established a registry of patients with alpha thalassemia to study the long-term effects of IUT. Based on the success of IUT in treating fetuses with alpha thalassemia, they have embarked on a groundbreaking clinical trial to try to cure alpha thalassemia while the fetus is still in the womb, potentially revolutionizing what it means to treat fetal diseases.

Before an embryo even has a heart to pump them, primitive red blood cells made up of embryonic hemoglobin shuttle oxygen to and from developing tissues. This early hemoglobin, consisting of two zeta and two epsilon globin subunits, has a higher affinity for oxygen than adult hemoglobin (2). This ensures that the embryo receives enough oxygen from its mother's red blood cells to grow into a healthy fetus.

After the first eight weeks of gestation, fetal hemoglobin — two subunits of alpha and gamma globin each — replaces embryonic hemoglobin. In alpha thalassemia, where there are no alpha globin subunits to partner with gamma globin, the gamma globin subunits latch onto each other in groups of four to form a nonfunctional form of hemoglobin called Bart's hemoglobin (3).

"It has an infinite oxygen affinity, which means it binds to oxygen and never lets go, so tissues never get oxygen," said Elliott Vichinsky, a hematologist and leader of the Northern California Sickle Cell and Thalassemia Center at UCSF Benioff Children's Hospital Oakland. "Very rarely will one survive to birth without intervention, and those who do often have suffered irreversible hypoxic damage."

If parents don't know that they are alpha thalassemia carriers, it is difficult to diagnose a fetus with alpha thalassemia right away. Typically, the first symptom is hydrops fetalis, the abnormal swelling or collection of fluid in fetal tissues such as the liver, spleen, or heart.

"Deductively backwards, [doctors would] figure out, ‘Oh, maybe the parents are carriers.’ And then they'll start testing the parents for carrier status, then offering an amniocentesis, and all of that takes weeks and weeks of time that you're waiting," said Lianoglou. "The baby's just getting sicker and sicker."

For a long time, doctors had no treatment for alpha thalassemia. They could only offer genetic counseling and pregnancy prevention. Once clinicians diagnosed a fetus with alpha thalassemia, the standard of care was pregnancy termination.

There were families who had already gone through a couple of miscarriages [due to alpha thalassemia], and they saw what was happening in beta thalassemia. They said, ‘If my baby could have that quality of life now, I want to have the pregnancy.’ - Elliott Vichinsky, UCSF Benioff Children's Hospital Oakland

As a hematologist at UCSF Benioff Children's Hospital Oakland in the early 1990s, Vichinsky noticed that as immigration from Southeast Asia started to increase in the United States and in Northern California especially, more and more thalassemia patients arrived at his door.

Many of his patients had the related genetic disease beta thalassemia, which results from the loss of the beta globin gene. Two subunits of alpha globin and two subunits of beta globin make up adult hemoglobin, which children start to produce soon after birth. Because beta globin is only needed to make adult hemoglobin, children with beta thalassemia survive gestation, but they need blood transfusions every three to four weeks once they are born. While beta thalassemia requires lifelong care, it is survivable.

"There were families who had already gone through a couple of miscarriages [due to alpha thalassemia], and they saw what was happening in beta thalassemia. They said, ‘If my baby could have that quality of life now, I want to have the pregnancy,’" said Vichinsky. He reasoned that perhaps IUT would allow fetuses with alpha thalassemia to survive pregnancy and grow up into kids who could manage their conditions with regular blood transfusions.

IUT is the bread and butter of fetal therapy. "It's kind of the first fetal therapy, and honestly, if you have any place that calls themselves a fetal treatment center, usually that's baseline what they can do," said Lianoglou.

Albert William Liley, a clinician and senior research fellow at the National Women's Hospital, performed the first successful IUT in 1963 to treat a fetus with hemolytic disease, which occurs when maternal immune cells cross the placenta and attack fetal red blood cells (4). In 1981, scientists improved the technique by transfusing blood directly into the umbilical vein rather than into the abdomen as Liley had done (5,6). In the mid-1990s, doctors, including Vichinsky, began using IUT to treat alpha thalassemia (7,8).

Because alpha thalassemia is a relatively rare condition in the United States, a lot of physicians don't know that IUT is a potential treatment option for these patients. "Still today, people will reach out to us to find out if their patient is eligible for transfusion, so there's still some education needed for the obstetrics and maternal-fetal medicine community regarding the value of transfusions for this diagnosis," said Lianoglou.

Over the past thirty or so years, anecdotal reports of how the fetuses treated with IUT for alpha thalassemia faired as kids have been generally positive. However, "There hasn't been a real large series put together to show the value of this intervention," Lianoglou said, so she and her colleagues established a registry of patients with alpha thalassemia to do just that.

Lianoglou along with Tippi Mackenzie, a maternal-fetal medicine researcher and pediatric surgeon at UCSF Benioff Children's Hospital and its Fetal Treatment Center, collected data from their patients at UCSF and doctors who treat alpha thalassemia all over the world. They tallied how many IUT each fetus received and how early in gestation they received them (9). They then assessed how those factors impacted the children's neurodevelopmental outcomes later in life.

When you're told your baby's going to die, and then you have a baby who's doing great… it's way different than what you expected. My theory is that your impression of their quality of life is pretty awesome because you thought they wouldn't have a life. – Billie Lianoglou, University of California, San Francisco

"We were able to demonstrate that those children who received two or more transfusions, they're all scoring average normal, which turns out, that's what you want! You can't have them all go to Harvard," Lianoglou laughed.

What intrigued Lianoglou the most was that the parents of the kids who had received IUT for alpha thalassemia rated their children's quality of life higher than the parents of kids with other chronic conditions and even parents of age-matched healthy kids.

"When you're told your baby's going to die, and then you have a baby who's doing great… it's way different than what you expected. My theory is that your impression of their quality of life is pretty awesome because you thought they wouldn't have a life," said Lianoglou.

While these children still need monthly blood transfusions, the IUT they received as fetuses have allowed them to grow up into happy and thriving children. "It's a miracle," said Vichinsky. "They’re regular kids."

Lianoglou added, "We've gone from terminate because we just think it's going to be bad to having data to say, 'Well, if you don't terminate, there is hope.' And for patients who don't find termination as an acceptable course, they deserve an option and hope."

IUT has made alpha thalassemia a survivable diagnosis, but Mackenzie, Vichinsky, and their teams at UCSF didn't want to stop there. By using IUT as a starting point and leveraging a unique quirk of the fetal immune system, these researchers may have found a way to cure this disease before the child is even born.

A fetus with alpha thalassemia can survive to term with IUT, but the only way to completely cure the disease is with a stem cell transplant after the child is born.

"A traditional bone marrow transplant has complicated risks, including finding a donor [and] the risk of becoming very sick from having your immune system ablated," said Lianoglou. Scientists estimate that only about 30 percent of people have a donor match, and even in that group, there is still the chance that the body will reject the donor stem cells (10).

Compounding that risk, doctors often use chemotherapy to kill the patient's disordered bone marrow cells to make room for the healthy donor cells. Chemotherapy also suppresses the patient's immune system, which allows the donor cells to engraft in the bone marrow more easily. Many people who have a donor match are not recommended to undergo chemotherapy because the high doses of chemotherapy necessary for engraftment can lead to serious organ toxicity.

One way to overcome the risks associated with chemotherapy and the lack of a perfect donor match would be to give a recipient a stem cell transplant before their immune system fully matures and before their bone marrow fills up with their own stem cells — so, during gestation.

By understanding that nuance in the fetal immune system, they proposed that using the mom as a stem cell donor would potentially optimize this protocol. You're transplanting cells the fetus is already potentially exposed to and may already have induced tolerance to. – Billie Lianoglou, University of California, San Francisco

In 1953, researchers discovered that a fetus's immune system could tolerate the introduction of foreign cells, suggesting that in utero hematopoietic stem cell transplants might be able to fix genetic diseases before a child is born (11). So far, in utero stem cell transplants have only completely succeeded in diseases where the fetus lacks a functioning immune system: bare lymphocyte syndrome and X-linked severe combined immunodeficiency (SCID) (12-14).

The main barriers to a successful in utero stem cell transplant are the maternal and fetal immune systems. Working in mice, Mackenzie and her team discovered that maternal T cells are the main antagonists to the donor stem cells trying to establish themselves in a fetus's bone marrow (15). They noticed, however, that if the donor cells matched the mother's cells, they engrafted into the fetal bone marrow much more easily, and the fetal immune system tolerated the cells completely. In a follow up study, Mackenzie and her team discovered that fetal regulatory T cells lead to tolerance of the maternal cells (16).

"By understanding that nuance in the fetal immune system, they proposed that using the mom as a stem cell donor would potentially optimize this protocol," said Lianoglou. "You're transplanting cells the fetus is already potentially exposed to and may already have induced tolerance to."

Based on this unique maternal-fetal tolerance, Mackenzie and her team initiated a phase 1 clinical trial to assess the safety of an IUT of maternal stem cells at the same time as a red blood cell transfusion, the standard IUT treatment, to treat fetuses with alpha thalassemia.

Alpha thalassemia, Lianoglou added, "is a condition that already requires you to intervene and accept a risk of that intervention. So, it's just really piggybacking: You're giving red blood cells, but you'll also give them maternal stem cells."

When they launched their clinical trial in 2016, Mackenzie and her team, which includes Canepa, Lianoglou, and Vichinsky among many other experts in bone marrow transplants and maternal-fetal medicine, had a difficult time finding alpha thalassemia patients to enroll.

"It was really just a matter of outreach, outreach, outreach and reminding folks that we're here," said Canepa. "It feels like we hit a bit of a tipping point in the past couple of years where folks know about us, which is amazing… We'll often receive emails from a provider when they have a family who they potentially think might be coming back with this diagnosis."

Sometimes, families who previously had a pregnancy affected by alpha thalassemia or who are having another pregnancy that they think may be affected directly reach out to Mackenzie's research team.

Once families have a confirmed alpha thalassemia diagnosis, Mackenzie and her team chat with them over a video call about the clinical trial and whether they might be interested in participating. After that call, families will often hop on a plane and arrive at UCSF in as few as one to two days.

"The sooner that families do get here and consider participating, the thinking is, the more effective the potential therapy would be," said Canepa. "It certainly can be a whirlwind."

When the families arrive, they meet with all of the specialists involved in the clinical trial, and the mother undergoes a series of screening tests to make sure it would be safe for both her and her fetus to participate in the trial. The researchers treat the fetuses between 18 and 26 weeks of pregnancy. Before 18 weeks, the procedure may put the pregnancy at risk, and after 26 weeks, the fetus may require more stem cells for a successful treatment than doctors can collect from the mother. If the team thinks it's safe to move forward and the patient agrees, then the researchers schedule the mother for the stem cell transplant as soon as possible.

The day of the stem cell transplant is a big day, Canepa said. "We've had one where we started at seven AM, and the team didn't leave the hospital until two in the morning that same day. There's a lot of waiting for the processing to be done and for the cells to be ready."

Pregnant women are usually not eligible to serve as stem cells donors because the process of collecting stem cells may lead to bleeding, which could put mothers at risk of anemia. During the bone marrow collection, the team collects as many stem cells from the mother as possible to give the transplant the highest chance of success without causing too much bleeding.

After delivering the maternal stem cells and blood via an IUT to the fetus, mothers will continue receiving regular IUT every two to three weeks until birth. When the child is born, the researchers collect the child's cord blood to see how many of the mother's cells successfully engrafted into the fetus's bone marrow. They also look for evidence that the baby's immune system still tolerates its mother's cells.

So far, Mackenzie and her team have enrolled and treated six patients. They hope to enroll 10 patients total in the trial.

"Five have been born. One is going to be born next week," Lianoglou said. But, she said, "with the cases that we've transfused so far, there hasn't been a cure."

While some of the maternal stem cells engrafted into the fetus's bone marrow, the kids did not produce enough healthy hemoglobin from their mother's stem cells, Vichinsky said. These children do, however, still have immune tolerance to their mother's cells. This tolerance means that the researchers may be able to repeat the stem cell transplant with the mother's stem cells even after the child is born.

"The goal is to be able to offer a booster transplant," said Lianoglou. "The baby's already been tolerized to the mom's cells. They have had some exposure, and then mom could serve as a donor again."

Because these children already tolerate their mothers’ stem cells, the procedure to prepare the children's bone marrow for more maternal stem cells will require very little chemotherapy, making the procedure much safer than a standard bone marrow transplant. The research team has not administered these booster transplants yet, but Vichinsky said that they hope to begin them soon.

"In addition to the clinical trial for stem cell transplantation, we're exploring potentially expanding the indications for that trial beyond alpha thalassemia for other conditions that may benefit, including Fanconi anemia, for example, or beta thalassemia," said Lianoglou. Mackenzie and her team are also working on gene therapies that they could deliver in utero as another approach for curing alpha thalassemia.

For the research team, the most rewarding aspect of working on this clinical trial has been getting to know the families and eventually the kids themselves once they’re born.

"We constantly get pictures of these babies as they grow and as they thrive, and just being able to see that and be a part of that is really, really special," said Canepa. "Both those who choose to participate and those who choose not to, we learn something from every single family that comes through our doors. They're absolutely incredible."

Stephanie joined Drug Discovery News as an Assistant Editor in 2021. She earned her PhD from the University of California Los Angeles in 2019 and has written for Discover Magazine,...

June 2023 Issue