Lulu, Nana, and a third girl were born as a result of an illegal genetic experiment in China in 2018. They edited their genome with the CRISPR technique. They are now three years old
Lluís Montoliu , National Center for Biotechnology (CNB – CSIC)
It has been three years since the world discovered that the researcher He Jiankui, from Schenzen (China), had crossed all the red lines when genetically editing with the CRISPR tools various human embryos and transfer them to various women for gestation.
From that irresponsible experiment two girls were born, twins, whom we met as Lulu and Nana (fictitious names) and later we learned of the gestation of a third girl.
This was an irresponsible experiment because it was still not safe then (and it is still not safe today, three years later) to guarantee the safety and efficacy of CRISPR tools in human embryos.
From similar experiments in embryos of other mammalian species (mice) we already knew that it was possible to obtain unexpected results, not always innocuous. For example, the alteration of genomic sequences similar to but different from those planned and the generation of multiple allelic variants after cleavage produced by the Cas9 nuclease –what is known as genetic mosaicism–.
There are no countries where germline gene editing is directly licensed . Nor in China, and it is explicitly prohibited in all the signatories of the Asturias Convention of 1997 , such as Spain, which in its article 13 states that we cannot modify the genome of the offspring .
The exclusive of the century
The birth of the first human beings with their genome edited with CRISPR tools is one of the news of the century. The publication of this news catapulted the publication of a series of videos from the laboratory of researcher He Jiankui , where he proudly described his feat, which he considered a scientific advance comparable to fertilization in vitro .
The dates chosen for this news (late November 2018) were not the result of chance. From 27 to 29 November, the second international summit on gene editing in humans was announced in Hong-Kong. In it, He Jiankui was one of the invited researchers and his exhibition was scheduled for Wednesday, November 28, 2018. He had anticipated to the organizers that he would talk about his results in vitro editing primate and human embryos.
His presentation broke all the forecasts of the summit and became probably one of the most viewed scientific talks in history . His only and probably last talk on this subject can be remembered in this video , starting at minute 75.
The discovery of Chinese twins with their edited genome caused the almost unanimous rejection of the scientific community , with some exceptions such as that of the expert researcher in gene editing George Church (Harvard), who he claimed his right to have a balanced opinion .
Daydreams that ended behind bars
Many institutions condemned the abominable experiment that should not have been carried out. Any possible reasonable explanation of He Jiankui’s intentions was blown up when fragments of the manuscript were published that He Jiankui had tried, unsuccessfully, to publish in top-tier journals.
Their messianic reverie was evident by trying to generate a line of boys and girls immune to the acquired immunodeficiency syndrome (AIDS), resistant to infection by the human immunodeficiency virus (HIV), after inactivating the gene CCR5 which encodes the coreceptor that the virus uses to enter lymphocytes. It did not succeed.
At the end of 2019, we learned of He Jiankui’s fate, after months without knowing his whereabouts and circumstances. He was sentenced by the Chinese authorities to financial penalties, jail time and disqualification from investigating , he and his most direct collaborators. The success dreamed of by He Jiankui, who expected to be applauded and honored as the savior of humanity, ended up behind bars, with the loss of all prestige and the general rejection of practically the entire scientific community.
What happened to the three girls?
These three girls are both famous and victimized, but we still don’t know much about them. The current ethical debate resides in whether we should have the data generated (even if it is to avoid redoing it without consent before being able to guarantee its safety and efficacy) from an experiment carried out without the proper ethical permissions. Also if we must assume, once carried out, that its results should be accessible to the rest of the scientific community.
Much has been written about He Jiankui’s experiment. But what do we know about the three CRISPR girls who were born with their genome edited?
This is what the journalist Vivien Marx wondered when the third anniversary of the news approached. He set out to gather as much information as he could and update what little we know about these CRISPR girls and how much we still do not know. The results of their work have been published in the journal Nature Biotechnology .
We know that the two twins were born a few weeks before the news broke, apparently prematurely.
We know that the third girl must have been born in the spring of 2019, and that the gestation of this new baby was already underway at the time of the international summit, as He Jiankui himself announced to the surprise of all those attending the scientific meeting.
We know that He Jiankui ignored a fundamental precept in the development of any therapy: the existence of a medical need. This was not the case, as there were assisted reproduction procedures designed for couples carrying the HIV virus who wanted to have a biological child. In addition, there is already a chronic treatment (antiretroviral cocktail) that makes it possible to keep the virus at bay throughout a person’s life.
We know that He Jiankui tricked his gynecologist colleagues into implanting the edited human embryos by pointing out that they were embryos obtained by a standard in vitro fertilization procedure , which was true, but without alerting them that they had been genetically edited. This, although not explicitly regulated at the time, was not allowed in China.
We know that He Jiankui deceived the parents of all the couples who participated in his experiment, giving them to understand that treatment with CRISPR by trying to inactivate the CCR5 gene was the only possible way to avoid that their children will become infected with HIV and develop AIDS.
He did not explain to them that the recommended medical procedure in these cases (carrier father, healthy mother) was in vitro fertilization , after washing the father’s sperm to eliminate all traces of the HIV virus. This procedure, by the way, was used by He Jiankui in his experiment, as deduced from his revealed and never published manuscript ).
We know that He Jiankui knew (from at least the two twins) that the experiment had not been successful by analyzing a biopsy of the edited and grown embryos in vitro before they were implanted. Despite this, he went ahead and had the embryos implanted for their gestation. Personally, I think this fact is the most despicable and irresponsible of all the wrong decisions that He Jiankui made.
We know that the Chinese authorities first applauded and praised the work of He Jiankui and, a few hours later, erase these initial manifestations and join the criticism and rejection without palliative that was mounted from all over the world. This initial support suggests that somehow the Chinese authorities probably knew of He Jiankui’s intentions.
We know that the Chinese authorities formed a committee of experts to investigate this case.
We know that He Jiankui had promised parents medical follow-up of girls born until they were 18 years old, at which point they could decide for themselves whether they wanted to continue medical supervision or not.
We know that the Chinese authorities have promised, for the time being, to closely monitor these three girls up to 5 years of age.
We know, from unidentified sources close to He Jiankui, that, apparently, these medical check-ups have taken place at one month, six months and one year after birth, with new check-ups pending in the near future. The cost of these reviews is apparently being paid by people close to He Jiankui.
We know that none of the three girls reproduced the deletion that occurs naturally in the human population in the gene CCR5 (delta32) that blocks coreceptor function and prevents entry of HIV into lymphocytes . People who carry this homozygous delta32 mutation are naturally immune to HIV (although they may be more sensitive to influenza, dengue and Nile fever viruses, among other alterations in their immune system). Nor was that the intention of He Jiankui in his experiment: he did not intend to reproduce the delta32 mutation and, obviously, he did not succeed.
We know that the CRISPR-Cas9 strategy used was aimed at producing a cut at the end of the region delimited by the delta32 deletion.
We know that the girl known as Lulu had 15 base pairs deleted in one of the two alleles of the CCR5 gene (multiple of three, which could be associated with a loss of function of the encoded protein, or not, if the five amino acids removed were not relevant to the function of the protein), while the other allele appeared to be intact, and was therefore functional.
We know that the girl nicknamed Nana had an insertion of one base pair in one of the alleles and a deletion of four base pairs in the other (neither is a multiple of three), which could suggest that both alleles would be non-functional due to alteration of the encoded sequence in the following triplets. However, Nana’s mosaicism would suggest that a percentage of her cells may not have been edited, so that if they were, Nana would still be infected by the HIV virus.
We know that the third girl apparently had only one of the two alleles of the CCR5 gene edited.
We know that the three girls are presumably mosaics , by analogy with other experiments in other mammalian species and by analysis of the chromatograms of the DNA sequences of the twins that He Jiankui made public in his presentation. That is, not all cells in your body have the same allelic versions of the CCR5 gene.
We know that at least one unwanted genomic alteration was found in the Lulu genome in an intergenic zone of chromosome 1.
We know that He Jiankui probably falsified or altered the ethical approval of his experiments at the university or hospital where he performed them.
We know that both the US National Academy of Sciences ( in its report published in September 2020 ) and the World Health Organization ( in its report published in July of 2021 ) have delimited when it would be acceptable to carry out an experiment similar to the one carried out by He Jiankui and what multiple conditions should be fulfilled before being authorized. None of them were fulfilled in He Jiankuui’s experiment.
Finally, due to all the above, we know that this experiment should not have been carried out.
What do we ignore about the three CRISPR girls?
We do not know where they were born, the city and the hospital where the birth occurred.
We do not know his real names and those of his parents. We also don’t know where they currently live. The Chinese authorities argue for the utmost respect for privacy and the legal impossibility of sharing these confidential details of these patients in order not to share this data.
We do not know his current state of health. We only know, from the Chinese authorities, that their health is being monitored and supervised by the country’s health authorities (at least up to 5 years, or until 18, according to He Jiankui’s promise to all participating parents in his experiments).
We do not know what type of medical tests have been carried out on these three girls and the results obtained. We also do not know the tests that will be carried out in the short, medium and long term.
We do not know what were the conclusions and the report on this case of the committee of experts convened by the Chinese authorities.
We ignore the real effect of any of the allelic edits detected in the alleles of the CCR5 gene of the three girls, their functional significance, and their possible involvement of the immune system.
We ignored all possible unwanted alterations in similar genes in the genomes of the three girls.
We ignore the probable existence of unwanted genomic alterations affecting large chromosomal regions , such as those detected in similar experiments on human embryos performed in vitro , without implantation.
We do not know if the mental health of these girls is being assessed, given their uniqueness and the potential stigma of discovering that their genomes have been genetically edited with unforeseeable consequences.
We do not know whether the knowledge that Lulu and Nana’s parents presumably have of their likely different susceptibilities to the HIV virus will determine whether they treat their twin daughters the same or differently.
We do not know the percentage and extent of genomic mosaicism in the three girls. To find out, it would be necessary to obtain biopsies of different tissues and derive DNA from all of them to be completely sequenced, in search of variations, polymorphisms, in certain coordinates.
We ignore the complete manuscript that He Jiankui wrote for publication in a first-rate journal, and of which only fragments are known. We do not know if it will be made public in full. The scientific community is divided on this. Their considerable and obvious ethical problems would prevent their publication in regular journals, but the doctors in charge of supervising the health of these three girls would benefit from knowing exactly the genomic modifications that each one of them presents and that were described in the manuscript.
We do not know if biological samples (biopsies, cells, DNA) from any of the three girls have been deposited in a biobank to be analyzed by other teams that could independently validate what He Jiankui reported.
We do not know how involved the Chinese authorities were in this experiment. If it was the idea of an isolated researcher or if he received institutional and financial support to be the first scientist to perform a genetic edition on human embryos with the result of an edited baby.
We do not know how involved some foreign collaborators were in the success of this experiment.
We do not know why none of the various foreign collaborators to whom He Jiankui openly confessed his intention to generate edited babies born from genetically edited human embryos with CRISPR decided to go to the Chinese or national authorities and alert the scientific community of the ethically inappropriate experiment that was to take place imminently.
A version of this article was published on the author’s blog . 
Lluís Montoliu , Scientific researcher at the CSIC, National Center for Biotechnology (CNB – CSIC)
This article was originally published in The Conversation . Read the original .