STAT article First human-pig chimera created, sparking hopes for transplantable organs — and debate by Sharon Begley

These human-pig “chimeras” were not allowed to develop past the fetal stage, but the experiment suggests such creations could eventually be used to grow fully human organs for transplant, easing the fatal shortage of organs: 120,000 people in the United States are waiting for lifesaving transplants, but every day two dozen die before they get them.

Human-pig chimeras could also be used for research into prenatal development and to test experimental drugs. A human lung in a pig might show more accurately the effect of a compound intended to treat, say, cystic fibrosis than today’s lab animals.

Scientists not involved in the chimera experiment, reported in Cell, said it answered a long-standing question. “What would happen if human stem cells were implanted in the early embryo of a large animal, not a mouse or rat?” said Dr. Sean Wu, of Stanford University, who studies congenital heart disease. “Now we have the first answer: You can get some human cells, though not a lot. It’s a tremendous accomplishment.”

But it’s one that bioethicists have warned about for at least a dozen years, since advances in stem cell biology made it easier to produce chimeras. The creation of “intermediate forms” of life is thought by some critics to “denigrate human dignity and blur the line between what is human and what is not, especially if you believe that we were created in the image of God,” said bioethicist and legal scholar Hank Greely of Stanford.

Much of the bioethics focus has been on what would happen if an animal had enough human brain cells to think and feel like a person — but a person inside the body of a monkey, pig, rat, or mouse. That and other concerns led the National Institutes of Health to announce in 2015 that it would not fund experiments that put human pluripotent stem cells, those with the ability to morph into almost any kind of tissue or organ, into the early embryos of other animals. (It wasn’t funding such research at the time.) This is how the human-pig chimera was created, with funding largely from foundations.

The NIH proposed lifting that moratorium last August, requiring additional oversight of chimera experiments and barring the use of human cells to create chimeras. But the proposal was not finalized before the start of the Trump administration, which this week directed federal agencies not to issue final guidelines or regulations on any topic, so the 2015 ban stands. That has no effect on experiments supported with private or state money. But a bill introduced late in the last Congress would prohibit chimera research, with penalties of up to $1 million and 10 years in prison.

It’s anyone’s guess as to what the new administration thinks about chimeras. A White House spokesman did not immediately respond to a request for comment.

Policymakers likely have a lot of time to figure out their position. The attempts at human-pig chimeras failed more often than they succeeded. And even the successes carried very few human cells. “The overall human contribution was very low, with what we estimate is less than 1 human cell per 100,000 pig cells,” and no human cells in the chimeras’ brains, biologist Jun Wu of the Salk Institute of Biological Studies, lead author of the Cell paper, said in an interview.

But that’s likely a floor, not a ceiling. In the same paper, the scientists — working under Salk’s Juan Carlos Izpisua Belmonte — reported making rat-mouse chimeras, too. Injecting rat pluripotent stem cells into mouse embryos produced rat-mouse chimeras. By age 2, the mice had some rat cells in their kidney, lung, pancreas, liver, and brain. The heart had the most rat cells — 10 percent.

The credit for that likely goes to CRISPR, the powerful new genome-editing technology. “To enrich the donor cells in the host, you need to disable the genetic program in the host embryo that gives rise to a particular organ,” Salk’s Wu said. He and his colleagues therefore used CRISPR to knock out, in fertilized mouse eggs, at least one gene crucial for the development of a particular organ, like the heart. Then, when the rat stem cells are injected, the ones fated to become heart cells had less competition and outcompeted mouse cells to form the particular organ, including the heart, eyes, and pancreas.

The Salk team did not report using CRISPR in the human-pig chimeras to help the pigs develop more humanlike organs. But those experiments are underway, Wu said.

The same technique — injecting pluripotent stem cells into early embryos — failed with other combinations: The scientists couldn’t create rat-pig chimeras, and although they produced human-cow chimeric embryos, they did not transfer them into cows to develop into fetuses.

And the human-pig chimeras proved much harder to create than the scientists expected, taking four years instead of the expected one. They first created “induced pluripotent stem cells” by turning back the calendar on adult cells until they were embryo-like. Wu and his team injected three to 10 of the human pluripotent stem cells into 1,506 pig embryos, each a few days old.

After growing the embryos in dishes for a few days, the scientists transferred them to 41 surrogate mother sows — 30 to 50 embryos each. Because ethical guidelines advise against letting chimeras develop completely, the scientists gathered the 186 surviving embryos after 21 to 28 days. Pigs have a 114-day gestation period.

Chimera
CELLCells derived from rat pluripotent stem cells were enriched in the developing heart of a genetically modified mouse embryo using CRISPR.

That’s a low success rate. But the cloning that led to the creation of Dolly the sheepin 1996 also failed far more often than it succeeded, and now cattle, sheep, pigs, and some pets are routinely cloned. If the marriage of stem cells and CRISPR follows a similar path, it might not be long before pigs have enough Homo sapiens in them not only to grow human hearts, lungs, livers, and kidneys for transplant but also to model human diseases more closely than current lab animals do and to test experimental drugs.

For that to happen, of course, the chimeras will need to gestate completely, be born, and grow up. Of the human-pig chimeras, those with the most human cells were the most underdeveloped; those with the fewest seemed to be developing more normally. “A lot of people are not optimistic that you can have full-term births of chimeras when the two species are so far apart evolutionarily,” said Wu.

Pigs and humans shared a common ancestor about 96 million years ago. There has been greater progress in creating chimeras from more closely related animals. On Wednesday, scientists reported in Nature that they had created mouse-rat chimeras — also starting with mouse pluripotent stem cells and fertilized rat eggs — in which the pancreases were sufficiently mouse-like that, when cells from them were transplanted into mice with diabetes, they churned out insulin and reversed the disease.

Any human organs growing in chimeras that scientists want to transplant or just study will need to be very human. It’s not clear if using CRISPR to knock out genes involved in making the host animal’s organ will be enough. But making chimeras with human organs whose development can be studied is more likely to succeed than the technique researchers have been trying for years: coaxing stem cells growing in lab dishes to become three-dimensional, functional tissues and organs.

“Chimeras involving human cells can provide a unique window into human development,” said Paul Knoepfler, a stem cell biologist at the University of California, Davis.

With chimeras, added Wu, “we might have a chance to see how the human cells develop, how organs form, and how mutations can cause [such serious malformations] that there’s a miscarriage.”

He and other scientists questioned how soon, if ever, chimeras would serve as sources for human organs, however. “Human cells are unlikely to behave entirely normally within a chimera,” said Knoepfler; cells from the host species are far more prevalent, providing the blood supply and connective tissue, for instance. “The odds are against it working any time soon,” he said, “or possibly at all.”

And if it does work? Knoepfler said that a chimera that’s human enough to have transplantable organs could also have “too many” human cells in its brain. “How many human cells are ‘too many’ in a chimera’s brain? How many human cells are too many in a chimera overall?” he asked. “Nobody really knows.”

Stanford’s Greely predicts that ethical concerns will abate. “In American bioethics, ‘cures’ is the trump card,” he said. “You play that angle” — such as by saying chimeras will provide transplantable organs to dying patients — “and, politically, you almost always win.”