A new cloning experiment conducted in China in 2022 and described this month in Nature Communications marks the first successful cloning of a rhesus monkey. It was achieved using a slightly different approach to the conventional cloning technique used to clone Dolly the sheep and other mammals, including long-tailed macaques (Macaca fascicularis), which were the first primates to be cloned.

The experiment, which has just been made public, involves obtaining a specimen of a rhesus monkey, a primate genetically very close to the human species, following a somatic cell nuclear transfer (SCNT) experiment.

Previously, in 2018, the birth of a macaque also generated by cloning was reported, although the current case presents differences that we shall explain later.

What is cloning?

SCNT technology consists of producing individuals of a species whose genome comes from an adult cell of another, being injected into a previously enucleated oocyte (an immature egg). In this way, genetically identical individuals are obtained.

This technique has been widely used for cloning several mammalian species, including sheep, cows, mice, pigs, goats, rabbits, and dogs. The aforementioned case of macaque cloning in 2018 has meant a significant qualitative change in attempts at reproductive cloning, given the genetic complexity of primates and their proximity to humans. These hitherto insurmountable difficulties were overcome by applying epigenetic regulators during the preimplantation process. This allowed the expression of certain genes to be regulated, ultimately enabling the birth of the first cloned primate. During this previous macaque cloning experiment, 109 cloned embryos were created, with nearly three-quarters of them implanted in 21 surrogate monkeys, resulting in six pregnancies. Just two of the monkeys survived the birth.

In humans, therapeutic non-reproductive cloning has been carried out with the intention of obtaining stem cells from the cloned embryos that can be used in regenerative medicine, i.e., to obtain cloned embryos from an individual from which stem cells would be extracted that, appropriately manipulated, could contribute to the regeneration of organs or tissues of that same individual, thus eliminating the problems associated with immunological rejection in transplant patients.

Efficiency of the technique

The live birth rate in the case of previous cloning of macaques was 2% on average. In conventional cloning methods, live birth rates for most mammalian species are between 1% and 3%, with slightly higher rates observed for cattle (5%-20%). It should also be noted that developmental abnormalities occur, particularly in extraembryonic lineages, such as placental tissues.

In the recent case of the rhesus monkey experiment, a single live cloned specimen was born from 113 initial embryos, which translates to an efficiency of less than 1%.

One more step

The study now published has investigated the genetic differences in embryos obtained by in vitro fertilization (IVF), specifically through intracytoplasmic sperm injection (ICSI), with others obtained through cloning processes (SCNT). The researchers found important differences in the epigenetics of both groups, consisting of a widespread decrease in DNA methylation and the loss of maternal imprinting in the genes of the cloned embryos.

These changes in the epigenetic information translate into histological modifications in the placentas of the cloned embryos, which show noticeable hyperplasia and calcification.

The investigators devised a novel procedure to “replace” the placental tissues of cloned embryos with those obtained from non-cloned embryos resulting from IVF techniques, specifically ICSI. They designed a procedure called “trophoblast replacement” (TR), in which they combined cells from both types of embryo: those that would develop the fetus came from the cloned embryo, while those that would give rise to the placenta came from the uncloned embryo, eventually leading to the birth of a healthy male rhesus monkey that has survived for more than 2 years at the time the paper was prepared for publication. The procedure consisted of injecting the internal cell mass (ICM) derived from cloned embryos (SCNT) into the blastoceles (without the ICM) derived from embryos produced with ICSI.

The birth of a rhesus monkey cloned with somatic cells by trophoblast replacement

Credit: Liao, Z., Zhang, J., Sun, S., et al. Reprogramming mechanism dissection and trophoblast replacement application in monkey somatic cell nuclear transfer. Nat Commun 15, 5 (2024).

Consequences of this new discovery

These findings provide valuable insight into the genetic reprogramming mechanism that occurs following monkey cloning, and pave the way for the expansion of primate cloning, offering new possibilities for the alarming prospect of human reproductive cloning.

Mu-ming Poo, director of the Institute of Neuroscience at the Chinese Academy of Sciences in Shanghai, said “We could produce a large number of genetically uniform monkeys that can be used for drug-efficacy tests”, eliminating differences in responses attributable to genetic variability in the study specimens.

Bioethical assessment

Obtaining individuals of one species through SCNT (cloning) whose genome is an identical copy of that of another individual constitutes an alternative that violates the natural process by which living beings that use sexual reproduction perpetuate their species. Through sexual reproduction, nature has ensured genetic variability, allowing offspring to inherit the genetic material of their parents combined in an original and unrepeatable way. This genetic variability is what ensures the evolution of the species we know today. Furthermore, reproductive cloning techniques, such as the one used in the recently published case of the rhesus monkey, have a low success rate, thus requiring the creation of numerous embryos and the use of many pregnant females. Additionally, the process fails in most cases due to the genetic anomalies that accumulate during the manipulation to which these embryos are subjected. In conventional cloning methods, live birth rates for most mammalian species are extremely low, ranging from 1% to 3%, with slightly higher rates observed for cattle (5%-20%). This involves huge efforts with poor results and the loss of numerous embryos in the process. As already mentioned, the birth of a macaque in 2018 was a first step in the race to overcome the difficulties of achieving the birth of a live cloned human.

The monkey currently in the news is genetically closer to humans than macaques, raising fears that reproductive cloning may be carried out in humans within a relatively short period of time. Obtaining cloned human individuals by manipulation presents ethical difficulties that are difficult to overcome, since it implies the genetic pre-design of the individual and possibilities for their instrumentalization, which violate their dignity as a unique and inimitable human being.

Unlike in China, reproductive cloning experiments with primates are currently not permitted in the European Union due to their genetic proximity to humans, unless the experiment is aimed at investigating a serious, fatal disease that affects humans or the primate species itself, which is not the case with this experiment.

However, it should be clarified that human cloning has been carried out for more than a decade, starting with the experiments of Mitalipov, although these were intended for research and not reproductive purposes. The laws of the countries that regulate these types of trials allow human embryos to be obtained by cloning for use in research, albeit with limitations, and they must be destroyed early in their evolution, when they contain up to about 200 cells. These early embryos are also individuals of the human species, artificially produced to be destroyed. It is not easy to justify them ethically either, especially when the number of embryos generated and destroyed is massive, and the results limited.

The clinical application of regenerative therapies based on stem cells obtained from cloned embryos is far from being a reality in medicine. The current progress of research into therapies with induced pluripotent cells, known as iPS cells, makes it unnecessary to undertake the costly and inefficient process of cloning and the bioethically unacceptable creation and destruction of human embryos that it entails.

Although some Chinese companies have already stated their intention to clone human beings for reproductive purposes, the technical difficulties to achieve this are insurmountable at the present time. Nevertheless, the rapid progress of gene editing techniques and the deepening knowledge of the functioning of our genome and the epigenetic mechanisms on which the development of the embryo depends makes us fearful that reproductive cloning in humans may become a reality before long.

Not everything that can be done in research should be done. Reverting sexual reproduction through the use of cloning techniques is not an evolutionary advance, but a regression. Utilitarian scientists for whom the end justifies the means often forget that some ends presented as scientific advances are not actually so in reality, but rather a step backwards. A thorough analysis of the consequences of these experiments, their usefulness and their risks, should inform the necessary regulation of these research practices, in defense of our species, our ecosystem and our dignity as persons.

Julio Tudela

Bioethics Observatory – Institute of Life Sciences

Catholic University of Valencia

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