Xenotransplantation How Bad Science and Big Business Put the World at Risk
from Viral Pandemics
ISIS Sustainable Science Audit #2 Mae-Wan Ho and Joe Cummins
Xenotransplantation - the transplant of animal organs into human beings
- is a multi-billion dollar business venture built on the anticipated sale
of patented techniques and organs, as well as drugs to overcome
organ-rejection (1). It has received strong criticism and opposition from
scientists warning of the risks of new viruses crossing from animal organs
to human subjects and from there to infect the population at large. But
regulators are adopting a permissive attitude for clinical trials to go
ahead. Scientific reports of virus crossing from pig to human cells (2)
and of viral infections in humans subjects transplanted with baboon livers
(3) are being ignored or dismissed, while inconclusive, widely faulted
papers are taken as evidence that no viruses are found in xenotransplant
patients (4). This audit exposes the shoddy science that puts the world at
risk of viral pandemics for the sake of corporate profit, and concludes
that xenotranplantation should not be allowed to continue in any form.
Instead, effort should be devoted to developing safer, more sustainable
and affordable alternatives that are already showing promise and will be
more likely to benefit society as a whole in the industrialized west as
well as in the Third World.
Xenotransplantation - the transplant of organs or tissues between
species has become a major issue within the past ten years. Biotech
companies are developing genetically engineered humanized pigs
to meet the demand for spare body parts in the industrialized world. A
multi-billion dollar market is anticipated from the sale of patented
techniques and organs, as well as existing and new drugs to overcome
organ-rejection (1). Still at the experimental phase, it has received
strong criticism and opposition from mainstream scientists warning of the
risks of new viruses crossing from animal donor organs to human subjects,
and from there to infect the population at large. But these warnings have
done little to dampen the enthusiasm for continued research well into
The world-leader in xenotransplant research is the UK biotech company
Imutran based in Cambridge, now a subsidiary of Novartis. Novartis already
own the rights to Cyclosporine A, the main anti-rejection drug given to
transplant patients to suppress the immune system. Since acquiring
Imutran, Novartis have pledged $1 billion for research in
xenotransplantation, and thereby to dominate a projected $11billion a year
market for organs and associated immune-suppressive drugs.
An estimated 10 000 pigs and nearly five hundred primates have been in
the UK, with very little accomplished. Xenotransplantation is in crisis.
At the bottom of the crisis lies some shoddy science that puts the world
at risk of viral pandemics for the sake of profit. At least one company,
PPL, which produced Dolly the cloned sheep, is reported to be winding up
xenotransplantation research, on the possibility that pig virus could
infect humans (5).
Humanized transgenic pigs as organ donors
Transgenic pigs, rather than our close relatives primates, were
considered as organ donors because there are greater ethical objections to
using primates, many of which are endangered protected species (6). As
pigs are already farmed for food, it was thought that there would be less
ethical concern, and that pigs could also be more easily controlled for
viral infections and consistent quality. Nevertheless, large numbers of
primates are exploited and made to suffer as experimental transplant
recipients; and primate to human transplant clinical trials have been
authorized in the United States.
The first hurdle in transplanting organs between distant species, as in
the case of pig to human, is hyperacute rejection (HAR) of the donor organ
by the host. This reaction is swift and severe, and depends on naturally
occurring, pre-existing antibodies. Naturally occurring human anti-pig
antibodies predominantly recognize the carbohydrate antigen
galactose-alpha-(1,3)-galactose attached to cell-surface proteins. Both
IgM and IgG (different classes of immunoglobulins or antibodies) in the
human blood contain antibodies that bind this antigen; which may comprise
up to 1% of the IgG. The enzyme for making the
galactose-alpha-(1,3)-galactose exists in all mammals except humans, old
world monkeys and the great apes. The binding of these antibodies to the
antigens triggers a cascade of reactions complement activation
that results in destruction of the donor organ and cells within minutes.
Induced antibodies against the foreign graft, xenograft, are responsible
for organ rejection in the longer term.
There are three possible ways to block HAR: by depleting the
pre-existing antibodies, by reducing antigen expression in the donor
cells, and by inhibiting complement activation. Of these, the last option
appeared to be the only clinically viable strategy in combating HAR. One
of the five candidate proteins that proved most promising is the decay
accelerating factor (DAF), which blocks an early step in complement
activation. Transgenic pigs containing hDAF were therefore produced.
Lack of documentation and molecular characterization of the transgenic
The first and only report in the scientific literature on the experiment
creating the transgenic pigs with hDAF was a note (7) less than one and a
half pages long, published in Transplantation Proceedings in 1994.
It contained no molecular genetic documentation of the construct such as
genetic map to indicate whether unknown sequences are present, or the
promoter-enhancer sequences used. It did not state how the minigene
construct was introduced, whether by itself, or spliced into a vector.
Later papers (6, 8, 9, 10) up to year 2000, all referred back to the same
experiment with no further elaboration.
About 2500 fertilized eggs were injected with the minigene. Of 85
surrogate mothers implanted with embryos, 49 delivered litters with 311
piglets, 49 of which were transgenic, ie, contained human DNA, with one to
30 copies of the gene. Only 33 expressed the gene for hDAF, however. The
rate of success is thus no better than 1.3%.
There was considerable variability in expression of hDAF in the
transgenic animals, not only between animals, but also between organs from
the same animal (6). Liver expression was found in 90% of transgenic pigs,
and expression in the heart was the least frequently detected (18/30). But
high expression did not guarantee expression in endothelial cells (cells
on the surfaces of the organ). No correlation was found between the number
of copies of the gene integrated and hDAF expression. Animals with the
highest gene copy number (13 copies) expressed very low levels of hDAF in
all transplantable tissues. The two most promising lines incorporated
between 6-8copies of the gene and expressed hDAF on parenchyma (inside)
and endothelium (surfaces) of all the transplantable organs. In 75% of the
organs, gene expression levels of hDAF was greater than in equivalent
These results underscore the unpredictable, uncontrollable nature of the
transgenic process and the low rate of success. There were no attempts to
characterise the transgenic inserts, nor to create stable transgenic lines
before transplant experiments were carried out; thus compromising
not only the reproducibility of the experimental findings, but also the
safety of the procedure, particularly with regard to the stability of the
transgenic inserts (see box 1) and the potential for creating new viruses
(see box 3). Plans were made (8, 9) to use whole yeast artificial
chromosomes (YACs) containing large segments of the human genome to
optimize gene expression, but it is not clear if such procedures have been
carried out. Introducing YACs may mean that uncharacterized human genome
DNA, including endogenous human viruses (see box 3), will be transferred
into the transgenic pigs, which could increase the potential for
generating new recombinant viruses (see box 3).
The 1997 review (6) admitted that production of an ideal hDAF expressing
pig was not complete, and that all organs used for xenotransplantation
were derived from heterozygous pigs, ie, pigs having hDAF gene(s) on one
of a pair of chromosomes. The review did not state whether these were
zero-generation transgenic pigs, or transgenic pigs from later generations
bred from the original. The ideal pig, according to the authors, would
express high levels of hDAF on organs and cells lining the organs, and
would be bred to homozygosity, ie, having hDAF genes on both of a pair of
chromosomes. That means the transgenic pigs would breed true.
Only one line was reported to fulfill these criteria in 1997. But the
authors pointed out that "breeding to homozygosity might cause
undesirable effects on the stability and health of the pig". This
conceals a major technical problem with creating transgenic lines.
Transgenic organisms, plants as well as animals, are genetically unstable
and do not breed true (see box 1).
Transgenesis is unpredictable and uncontrollable,
and trangenes are unstable
The instability of transgenic plants is well-known and actively
researched. Transgenic constructs typically integrate at random into
the host genome, and in a scrambled configuration, consisting of
repeats, rearrangements and deletions (11). There is no reason to
expect transgenic animals to be different. Indeed, integration of
transgenic construct was reported to be random in the transgenic pigs,
and the expression of the transgene depended on the site of
Transgene integration was examined by fluorescence in situ
hybridization (FISH) (12), a technique that enables the inserts to be
seen on chromosomes. Routine slot blot analysis of total
transgenic pig DNA was done first to identify pigs with hDAF DNA.
According to the strength of the signal, one line, E14, was estimated
to contain 40 copies of the hDAF transgene; while another, A74,
contained 6 copies, and a third, C50, two copies. These were referred
to as "heterozygous founders lines". Again, it is not stated
how many generations were bred after transgenesis. And it is not clear
why the "lines" are heterozygous and not homozygous.
In the cross E14 x A74, one from a litter of 13, and another from a
litter of four were the only piglets that showed a signal similar to
that of one parent. In the cross C50 x A74, three out of 9, and 4 out
of 11, respectively showed signals that were similar to those of the
parents. But when analyzed by FISH, only 4 piglets were actually found
to have inherited any transgenes from their parents. Thus, the actual
transmission of transgenes is 4/37 or 10.8%. This is much lower than
the 75% predicted (assuming both parents were heterozygous), and is
typical of the instability of transgenic inserts, which can become
lost in subsequent generations. This raises the question as to whether
the lost insert can be tranferred again, unintentionally, to unrelated
species, a process referred to as horizontal gene transfer, with its
own attendant hazards (13).
Intended xenotransplant recipient animals were not screened for viruses
and postmortems of transplant recipients did not include examination for
In the first experiment, eight hearts from transgenic pigs were
transplanted into non-immune suppressed cynomolgus monkeys (Macaca
fascicularis) of unspecified origins, and without pre-screening for
viral infections or endogenous viruses. The median survival was 5.1 days
(97-126h) with no reported hyperacute rejection. Five of the ten controls
that received control hearts survived a surprising mean of 86.4h, while
the other five survived for only 2.6h, as typical of HAR.
In a second experiment, ten cynomolgus monkeys receiving heart
xenotransplants from transgenic pigs were dosed with a regimen of immune
suppressing drugs: 80-180mg/kg/day of cyclosporine and 10-20mg/kg of
cyclophosphamide on alternate days. Methylprednisolone was also
administered at 1mg/kg. This regimen produced median survival of 40 days
(2 to 62 days). The five non-transgenic hearts were rejected hyperacutely
(median 55 mins). Five animals from the transgenic heart group had to be
euthanased (killed out of compassion, to relieve suffering) due to
gastrointestinal toxicity, resulting in severe diarrhoea. All hearts were
reported to be normal with no evidence of complement or immunoglobulin
In immune-suppressed animals, rejection was considered not the primary
cause of graft failure. Only two out of ten were due to rejection, while
drug toxicity resulted in 50% having to be euthanized. That accounted for
seven of the ten xenotransplant recipients. So, what did the remaining
three die of? The report did not specify. There was no indication that
post-mortem examination for viral infections had been carried out.
An experiment involving a single transgenic pig heart transplanted to a
baboon was described in a subsequent paper (10). The animal survived 39
days with an immunosupressive regimen of cyclophosphamide, cyclosporine A,
mycophenolate mofetila and cortecosteroids. It was reported to be active
and energetic until day 39, when it underwent sudden and rapid decline,
leading to collapse almost immediately following the routine
administration of drugs. The cause of death was recorded as "unclear".
Postmortem examination was limited to ascertaining that organ rejection
was not to blame. Again, no investigations for viral infections were
Hazards of cross-species viruses arising from xenotransplantation
The problem of infectious viruses arising from xenotransplantation was
first raised by Robin Weiss and his coworkers at the Institute of Cancer
Research (2). They showed that a pig endogenous retrovirus (see Box 2) can
infect cultured human cells. And once the virus has gone through a
complete life-cycle in human cells, it is then able to infect a wide range
of other human cells. Many copies of pig endogenous retroviruses (PERV)
exist in the pig genome and it will be extremely difficult, if not
impossible, to breed pigs free of PERV. Robin Weiss argued that accidents
have already occurred (reported in ref. 1). Millions have become infected
with the monkey SV40 virus through polio and adenovirus vaccines made in
monkey kidney cells. Many viruses lying dormant in animals, in particular
herpes viruses and retroviruses, can become activated and deadly in
humans. Activation of animal viruses might be favoured under transplant
conditions, which compromise many barriers to natural infection. Robin
Weiss stressed that virus adaptation or recombination with other
retroviruses in the new host cannot be dismissed.
What are endogenous retroviruses and why are they
A retrovirus is a RNA virus that is reverse-transcribed into
complementary DNA (cDNA) and integrated into the host cell genome to
replicate and complete its life-cycle. Endogenous retroviruses (ERVs)
are elements in the genomes of all higher organisms including human
beings, which are very similar to the genomes of retroviruses. They
are flanked by long terminal repeats (LTRs) and carry genes coding for
structural and coat proteins of the virus as well as the reverse
transcriptase and integrase enzymes (required for
reverse-transcription and integration of the viral genome into the
host genome) (14).
There are two theories on how ERVs may have evolved. Howard Temin,
Nobel laureate who co-discovered the enzyme reverse transcriptase,
suggested that they have evolved from retro-transposons mobile
genetic elements with reverse transcriptase - which are part of the
genomes of all higher organisms. Alternatively, ERVs may have evolved
from exogenous viruses, foreign viruses that have become integrated
into the genome. There is no reason to believe that these alternatives
are mutually exclusive. Exogenous viruses, which may have arisen from
retrotransposons, can indeed re-invade the genome of higher organisms
to become endogenous retroviruses. In general, most endogenous
retroviruses appear to have been acquired millions of years ago, but
there is evidence that new retroviruses can be acquired. Under certain
circumstances, endogenous retroviruses can also give rise to
infectious retroviruses, although most ERVs are in a dormant,
Some ERVs have retained their ability to code for virus that can
infect the cells of other species, a phenomenon known as xenotropism,
and this is of particular safety concern with regard to
xenotransplantation. For example, xenotropic retroviruses in mice have
been described that cannot replicate in mouse cells, but can propagate
profusely in human cells in culture. Also, chick and pig ERVs rarely
replicate in their own species but readily infect cultured cells of
other species, including those of humans. Likewise, a cat ERV
replicates in human cells, as does one from baboon, although neither
replicates in its own host species.
Another important safety consideration is that the creation of
transgenic pigs with human genes, such as hDAF, to suppress hyperacute
rejection, actually increases the potential for creating infectious
cross-species viruses. It suppresses the bodys defense against
bacteria and viral infections, and also provides more opportunities for
the viruses to gain access to the host cells (see Box 3).
Transgenic pigs increase the likelihood of
generating cross-species viruses
Robin Weiss (15) points out that many animal viruses with lipid
envelopes are sensitive to inactivation by the human complement
cascade. The virus undergoes lysis (breaking open), triggered by the
binding of anti-alpha-Gal antibodies to alpha-Gal on the viral
envelope. Viruses grown in non-primate cells are sensitive to
inactivation by fresh human serum, whereas the same viruses propagated
in human cells are not because they have lost the alpha-Gal. Other
enveloped viruses grown in animal cells are also sensitive to lysis by
human complement, including arenavirus, paramyxovirus, alphavirus and
the rhabdoviral pig pathogen, vesicular stomatitis virus. If alpha-Gal
is on the host cell, then the viral envelope becomes sensitive to
rapid lysis by human serum. In other words, virus inactivation occurs
by precisely the same mechanism as hyperacute rejection of xenograft.
So, modifications to make pig xenografts resistant to hyperacute
rejection may also make any enveloped viruses of pigs similarly
resistant to breakdown in the human host.
The key proteins are CD46 (membrane cofactor protein, MCP-1), CD55
(decay accelerating factor, DAF) and CD59 (prolectin). They all
inhibit downstream steps in the complement cascade, and several
transgenic pig herds have been developed expressing one or more of
these human genes. All of these are present in the envelope of HIV,
the AIDS virus, and protect the virus from lysis.
CD46 is the cell-surface receptor for measles virus and CD55 can
serve as a binding receptor for Echo and Coxsackie B picornaviruses.
Coxsackie B virus causes myocarditis and might endanger the pig heart
in an immune suppressed recipient of a xenograft. Transgenic pigs may
therefore also provide an opportunity for animal viruses to adapt to a
human host range. Coxsackie B virus, for example, can be adapted to
grow in mice, and in some human cell cultures, it increases its
infectivity a million-fold by adopting the CD55 receptor. If pigs were
to harbour picornaviruses that use the pig equivalent of CD55, such
viruses may readily adapt to recognize human CD55 in transgenic pigs
that express both pig and human equivalents. These viruses would then
be pre-adapted to transmit to the xenograft recipient and to be
transmitted from human-to-human. There is already concern that mice
transgenic for human poliovirus receptor should not escape and become
a non-human reservoir for a human pathogen.
Animal morbilliviruses (measles-related viruses such as canine
distemper virus and rinderpest virus) might become pre-adapted for
human transmission in CD46 transgenic pigs. Morbilliviruses are known
to jump host species as in the recent epidemic in seals and dolphins.
In Australia, a vet and a stable-hand died after an autopsy of a horse
with a new type of morbillivirus which in turn was probably acquired
from fruit bats.
Human tumour tissue transplanted into immunodeficient mice
frequently becomes infected by endogenous xenotropic mouse retrovirus.
Two or three distinct pig retroviruses can infect some human cells in
Researchers are identifying many new pig viruses. One pig virus,
closely related to human hepatitis virus E (16), was found in the
majority of pigs, three months or older, in herds from mid-western
United States. This raised concerns over the creation of cross-species
pathogens in xenotransplantation.
Risks considered disproportionate to benefits by many scientists
Jonathan Allen, a virologist on FDAs advisory subcommittee,
accused the FDA of the failure to adhere to the precautionary principle
(see ref. 1). It may take decades for a xenozoonosis infectious
diseases arising from cross-species viruses - like the AIDs virus or Human
T-cell Leukemia Virus to spread and become detected. The FDAs
requirement that all future xenotransplant recipients be monitored for
infectious diseases over their life time, and prohibiting them and their
close contacts from donating blood, amount to shutting the barn door after
the horse has bolted.
The American Society of Transplant Physicians also want tougher
guidelines, and accuse the capital-hungry biotech companies of excessive
hype, and creating unrealistic expectations among patients, fuelling
pressure to proceed to clinical trials.
Fritz Bach, xenotransplant scientist from Harvard among others, called
for a moratorium in 1998, as potential risk of xenotransplants would
affect the general public who are being exposed without informed consent.
He argued for a wide "informed" public debate on whether such
trials should be allowed to proceed at all, as it is an ethical question.
According to the United Network for Organ Sharing, the number of
transplants increased from 12 000 to 20 000 between 1988 and 1996; while
the number on the waiting list soared from 16 000 to 50 000 and the number
of deaths rose from about 1 000 to 3 000 (17). David Sachs of Harvard
Medical School estimated that more than 400 000 could benefit from heart
transplants when the official waiting list in 1996 was 3 698. Many on the
waiting list are for repeat procedures to replace failed transplants. Was
Sachs estimate overblown? Did it reflect the over-enthusiasm on the
part of the medical establishment for spare-organ trafficking rather than
real demand or benefit? Chronic rejection is the major cause of the loss
of allotransplants from unrelated human donors. So it can be predicted
that xenotransplants will be much worse.
A new study published in the British Medical Journal suggests that even
transplants from unrelated humans save lives only in patients on the verge
of death (18). The study was carried out in Germany. Researchers looked
back at 889 patients listed for a first heart transplant in 1997. The
patients were categorised into groups with low, medium or high risk of
dying and compared the mortality of those on the waiting list with those
who had a transplant. It turns out that there were no differences in
mortality for the low and medium risk groups. Only in the sickest patients
was there an improvement in survival due to the transplant.
There is evidence for cross-species viruses in xenotransplant
Evidence that baboon viruses have arisen in two human subjects
transplanted with baboon livers emerged two years ago (3). DNA of two
retroviruses, the simian foamy virus (SFV) and baboon endogenous virus
(BaEV), were found in many tissues of the patients. The presence of baboon
mitochondrial DNA (evidence of baboon cells) were also founded in the same
tissues, suggesting that baboon leukocytes harboring latent or active
viral infections had migrated from the xenografts to distant sites in the
human transplant recipients. The authors stressed, "The persistence
of SFV and BaEV in human recipients throughout the posttransplant period
underscores the potential infectious risks associated with
These were the first baboon-to-human liver transplants. One was
performed in June 1992 in an HIV-infected 35-yr old man, who survived 70
days, and the second, in Jan. 1993, in a 62-yr man who also received donor
bone marrow intravenously and survived for 27 days. Both patients had
hepatitis B virus-associated liver cirrhosis. Neither transplanted baboon
liver functioned normally. In addition, both patients developed kidney
failures and multiple post-transplant infectious complications. Both
received an immunosuppressive regimen of FD-506 prednisone and
cyclophosphamide. The two adult male baboon donors were screened against a
panel of simian and human viruses and were negative for Simian T-cell
Leukemia Virus, Simian Immunodeficiency Virus and simian retrovirus.
Antibodies against SFV were detected in samples from both donor baboon
samples prior to transplantation, whereas the human patients were
non-reactive during several time points after receiving the transplant.
However, a faint positive result was recorded at day 22 in patient 2. The
absence of antiviral antibody in the patients may be due to insufficient
time in case of patient 2, and AIDs in patient 1. Furthermore,
immunosuppressants may have suppressed the antiviral response.
Nevertheless, SFV DNA was detected by PCR (Polymerase Chain Reaction)
probes in tissues from both patients. In patient 2, SFV DNA was detected
in the liver graft on day 24 but not day 12. The liver sample from patient
1 on day 16 was negative for the viral DNA, but positive results in both
lymph node and kidney were obtained on the day 70. DNA sequence analyses
confirmed that the SFV in the transplant recipients were closely related
to the baboon virus rather than those of other primates. The life cycle of
SFV includes integration of viral DNA into the host genome.
Baboon mitochondrial DNA and BaEV were simultaneously detected in every
sample in which SFV was present.
The authors stated, "These findings demonstrate the potential for
both exogenous and endogenous viruses to reside in human recipients of
animal organs for a significant period after transplantation. It is
possible that these circulating xenogeneic cells could also act as
conduits for new human infections .Since retroviruses commonly exist
as persistent latent infections, with an incidence of disease that varies
because of both host and viral factors, the possibility that baboon foamy
viruses might cause disease in humans remains a consideration in
discussing future animal sources for xenotransplantation. Theoretically,
other yet to be characterized viruses carried by baboons might also be
transmitted to human recipients." (p.824).
As mentioned earlier, none of the published papers up to year 2000 from
the Imutran group gave any indication that post-mortem pathological
investigations included tests for viral infections.
A brief note (less than one page) from Imutran-Novartis, published later
in the same year, reported an experiment in which pig alveolar macrophages
(PAM) from pig blood, infected with pig cytomegalovirus, PCMV, were
cultivated for up to 15 passages together with human cell lines, and
monitored for the presence of PCMV (19) at three time points: passages 5,
10 and 15. It reported "no evidence of PCMV infection of the human
cells at passage 15, the farthest time point in this study, despite
evidence that PAM and PCMV were present in the co-culture up to at least
passage 10. On the basis of this evidence, PCMV is unlikely to be a
significant zoonotic agent in clinical xenotransplantation of pig organs
The experimental results were equivocal, to say the least. It is bad
science to draw any conclusions on the basis of such limited, inconclusive
data. Cell culture conditions are obviously different from the conditions
in which a xenograft is transplanted into a living body. Furthermore,
positive indications for PCMV were obtained at both passages 5 and 10.
Clinical trials to go ahead based on faulted study
White and Nicholson (20) reviewed xenotransplantation research at the
end of 1999, and concluded that xenograft rejection cannot be prevented
without significant immune suppression and toxic side-effects. They
highlighted the risk of pig endogenous retrovirus transmission, but state
that some of the important issues will never be solved "until
carefully regulated clinical trials are allowed to begin." They take
at face-value a report (4) published by Imutran/Novartis and other biotech
companies claiming no retroviral cross-infection in patients exposed to
pig tissues or receiving pig xenografts; and which has been criticized by
The study tracked 160 patients in 9 countries exposed to living pig
tissue over a 12-year period. One hundred and thirty one patients had
their blood "filtered" and re-circulated through pig spleens,
kidneys, livers, or devices made with pig liver cells; 15 received pig
skin grafts for burns, and 14 received injections of pig pancreas cells
As pointed out by Peter Collignon of the Infectious Diseases Unit,
Canberra Hospital, Australia (21), pig endogenous viral (PERV) genes were
detected in 30 of the patients, and pig cells persisted in 23
xenotransplant recipients for up to 8.5 years. Although the authors found
no active infection, the possibility of infection remains in the four
patients with positive antibodies to PERV, and in another four patients
with unexplained symptoms (skin rashes). In addition, lack of antibodies
to PERV may not exclude the existence of infection, as for example, prion
diseases (which include mad cow disease) cannot be detected by antibody or
cellular immune responses. Immune suppressive drugs could also prevent the
development of anti-viral antibodies (3). Collignon asked, "Who would
have predicted that so many patients only transiently exposed to pig
tissue would have persistent pig cells (and PERV) in their blood?"
Even though the authors claim that there is no conclusive evidence of
human infection by PERV (4), they admit that "PERV infection [cannot]
Emanuel Goldman, Professor of Microbiology and Molecular Genetics at New
Jersey Medical School in Newark noted that a majority of the samples
tested were from patients whose blood had been flushed through pig
organs/tissues, and recirculated into their bodies for very short periods
- of the order of minutes to hours. Such data are hardly relevant to the
kinds of conditions that would apply in whole organ xenotransplants. Data
from the 14 subjects who received pig pancreatic islet cells could be
taken more seriously. But, as with the burn victims, important information
about these patients exposure times to the xenografts and health and
immunological status was missing (22).
Moreover, Goldman pointed out that the patients in the study were
treated, and serum samples handled and stored in 9 separate countries,
making quality control almost impossible. Looking for PERV RNA is always
suspect with serum stored for several years. Plasma samples are frozen at
-70C and thawed at very high temperatures. Many viruses are very unstable;
it is unknown whether such extreme temperature changes might alter PERV
and affect test results.
Another problem with the study is that the PCR probes are only good for
two genes of one PERV, and will not detect other viruses, such as
Hepatitis E virus or Cytomegalovirus, nor recombinant viruses, which are
hybrids of pig and human viruses. Finally, none of the patients have been
exposed to transgenic pig tissues. And it has already been pointed out
that transgenic pig tissue may be more likely to give rise to new viruses
(see Box 3).
To address the risks of infection, the US Food and Drug Administration
(FDA) established an Advisory Panel on Xenotransplantation, and the
British government set up the UK Xenotransplantation Interim Regulatory
Authority (UKXIRA) in 1997.
The report (4) on the lack of evidence for PERV infection in
xenotransplant recipients allows the Novartis/Centers for Disease Control
teams to conclude that only cautious progress in closely monitored,
prospective clinical trials will help to assess the safety and efficacy of
xenotransplantation. Both the FDA and UKXIRA are taking this same
attitude, and are ready to approve small-scale human trials of pig cell
therapy. To proceed on this basis not only exhibits flagrant violation of
the precautionary principle, it is to adopt the anti-precautionary
approach (23), where failure to rule out viral infection (due to faults in
data collection or handling) is taken as evidence that there is no risk of
Robin Weiss (24) compares the present situation in xenotransplantation
to the short-lived Asilomar moratorium on genetic engineering declared in
mid 1970s. The parallel is closer than perhaps he thinks, as some of us
have indeed questioned whether the exponential growth in genetic
engineering biotechnology since the 1970s may have contributed to the
recent resurgence of drug and antibiotic resistance diseases (25). In
genetic engineering as in xenotransplantation, species barriers are
undermined, and conditions are created which favour the generation of new
viruses through horizontal gene transfer and recombination.
It turns out that PERVs not only infect human cells but produce products
of the infection that inhibit human immune cell functions. Thus, PERV
infection in transplant recipients could lead to an immunodeficiency
disease (26). The suitability of baboons as models for human
transplantation was previously questioned on grounds that pig cells do not
release PERVS when they contact baboon cells or following pig to baboon
cell transplants (27). However, a subsequent study showed that human,
gorilla, and Papio hamadryas primary skin fibroblasts, as well as
baboon B-cell lines, are permissive for PERV infection (28). There are
probably no barriers to the transfer of viruses across species under
conditions of co-culture of cells, or xenotransplantation of tissues and
British scientists have now found that cancer-causing retroviruses can
also spread relatively easily across species in the wild (29). Mouse
leukaemia viruses, close relatives of the cancer retroviruses known to
infect pigs, were found in a range of mammalian species, suggesting that
pig retroviruses may also be capable of infecting other animals -
including humans - with relative ease. This has prompted the Western
health authorities to impose a moratorium on all xenotransplant surgery.
Professor George Griffen, a member of the UK Xenotransplantation Interim
Regulatory Authority, admitted that viruses jumping species from
xenotransplant organs is possible, but draws attention to the fact
that "hundreds of pre-moratorium xenotransplant recipients have yet
to show reactions to retroviruses."
Governments disregard scientific evidence to put their citizens at risk
from cross-species viral pandemics
In January 2000, the US FDAs Xenotransplant Subcommittee met in
Gaithersburg, Maryland to review its proposed guidelines to "indefinitely
defer" blood and plasma donations from xenotransplant recipients and
their "close contacts" (29).
Phil Noguchi, Director of FDAs Division of Cellular and Gene
Therapies, acknowledged that xenotransplantation is "fraught with
danger." Yet he revealed that there are currently 12 FDA-approved
xenotransplant clinical trials going on in the U.S. Most, if not all, are
industry-sponsored, and involve the use of pig cells to treat diabetes and
neurological diseases, and whole pig livers and cells to perfuse the blood
of patients with acute liver failure.
In order to perform such trials, companies must submit an
Investigational New Drug (IND) application. But Jay Siegel, Director of
FDAs Office of Therapeutics Research and Review indicated that he
would be shocked if there werent activities being done that are not
under IND that should be.
Genzyme, a Cambridge, Massachusetts-based biotech company, had been
treating about 100 burn patients per year since 1987 with a xenotransplant
product called Epicel, regulated as a device. The company use
3T3 mouse cells to grow layers of human skin,which are then applied to the
patient. The mouse cells are allegedly irradiated to prevent them, and any
viruses, from proliferating; though when pressed, Genzymes President
admitted that the company was still assessing the efficacy of its
irradiation method. And, it had not performed FDA-required tests to
determine whether its mouse cells could infect human cells. Most shocking
was the companys admission that it had not kept a registry of the
patients it treated, nor followed up to see whether any of them might have
developed signs of illness or infection. Genzyme said it would be "impractical"
to try to find these patients. The FDA seemed to have no knowledge of this
Andrew Dayton of the FDAs Division of Transfusion Transmitted
Diseases, and architect of the guidelines, acknowledged that if a
xenotransplant-related virus entered the blood supply by mistake, the
results would be "disastrous" and the necessary withdrawal of
contaminated blood products would cause serious blood shortages.
While some Subcommittee members seemed to downplay the threat of
infection by pig viruses, virologist Jonathan Allan commented that, for
FDA to recognize infectious disease risks in non-human primates, but not
in pigs, is arbitrary. Prem Paul, a veterinary researcher at Iowa State
University, warned that new pig viruses were continually being discovered;
they had not been extensively studied; and the potential existed for them
to mutate and infect humans.
British veterinary pathologist David Onions concurred. He warned that
pig parvovirus can change hosts and escape inactivation treatments; and
has already been found in Porcine Factor 8 used to treat hemophiliacs.
In May, 2000, a new US Public Health Service (PHS) Guideline on
Infectious Disease Issues in Xenotransplantation was published (30). It
involves a complex series of measures to store tissue samples for future
study and to establish a national xenotransplant database something
that should have been done before clinical trials were approved. As it is,
they will only serve to detect disease and virus after it is too late.
The PHS acknowledge that viruses from animals used in xenotranplantation
could infect patients, their offspring, health workers and the general
public. And even admits that, "all xenotransplantation products pose
a risk of infection and disease to humans", "baboon endogenous
retrovirus in human recipients of baboon [livers] has been documented",
"new viruses capable of infecting humans have been identified in pigs",
"all species pose infectious disease risks", and "[xenotransplant
recipients] may represent a biohazard to healthy livestock".
According to the PHS guidelines, the sponsors are entrusted to design
and monitor xenotransplant trials, tailor complex informed consent
documents, educate workers, to effectively screen source animals for
viruses, maintain proper documentation, and reliably report crucial
information about patient and animal health to federal agencies. There is
no mention of who will be held responsible if a novel virus is unleashed,
and no emergency procedures to deal with an outbreak have been proposed,
even though the PHS acknowledges that "airborne transmission of
infectious agents" is possible.
PHS further suggests that some animals from xenotransplant facilities
may be considered "safe for human food use or as feed ingredients",
in flagrant disregard of the fact that the safety of transgenic food is
yet to be established, and the international community has found it
necessary to negotiate and agree a Biosafety Protocol regulating the safe
use and transfer of genetic engineered products under the UN Convention on
If xenotransplantation is to go ahead, it will involve levels of animal
suffering unacceptable to the majority of people. As it is extremely
inefficient, it will also generate many abnormal failures and surplus
animals which have to be disposed of safely. There is as yet no
documented, true-breeding transgenic line established to-date.
PHS, in their current guidelines, state that Americans have neither
endorsed nor rejected xenotransplantation. But documents obtained through
the Freedom of Information Act reveal otherwise. In response to its 1996
draft guideline, PHS received over 160 comments: 115 against
xenotransplantation, 29 in favor, and 19 neither for nor against, with 8
of these strongly opposing the use of nonhuman primates. Furthermore, the
Food and Drug Administration received almost 6,000 postcards, and over 350
letters protesting its April 1999 guidelines on the use of non-human
primates in xenotransplant trials.
The Campaign for Responsible Transplantation (CRT), an international
coalition of physicians, scientists, and 90 public interest groups, have
denounced the PHS Guidelines as irrational and in violation of the
Conclusion: stop xenotransplantation for safer, more humane and
Our investigations have revealed how bad science has been involved in
the xenotransplant project from the start:
lack of proper documentation of the transgenic process and
characterization of the transgenic pigs
lack of quality control
failure to obtain well-characterized stable transgenic lines before
transplantation experiments were attempted
failure to screen for viral infections in experimental xenograft
use of inconclusive studies to push for clinical trials in humans
systematic disregard of existing scientific evidence of cross-species
viruses arising from xenografts
It is nothing short of a scandal to allow xenotranplantation to go ahead
in the light of existing scientific evidence, especially when there are
safer, more humane and effective alternatives (17).
Much can be done to increase human organ donation in the short term,
especially if an assurance can be made to the donor that the organ will be
offered free of commercial interest to the recipient. The use of
artificial organs and human cells and tissues will both avoid the risk of
cross-species viral epidemics.
One of the most exciting recent development is the possibility of
regenerating organs and tissues from the patients own stem cells
(31), cells which retain the ability to multiply and differentiate into a
number of different cell types even in the adult. This would avoid immune
rejection as well as viral epidemics. We reject the claim that human embryonic
stem cells have to be used, which are obtained from human embryos
created solely for the purpose. It has now been demonstrated that adult
human liver cells can be derived from stem cells originating in the bone
marrow (which normally produce blood cells) or circulating outside the
liver. This raises the possibility that bone-marrow stem cells, either
from a donor or from the patient could be used to generate liver cells for
replacing damaged tissue, thus obviating the need for organ transplant
altogether (32). Better yet, why not find out how to encourage adult stem
cells to regenerate in situ?These alternatives are
infinitely preferable to xenotransplantation in being safe, humane,
sustainable and affordable; and hence more likely to benefit society as a
whole in the industrialized west as well as in the Third World.
References and notes
Butler, D. (1998). Last chance to stop and think on risks of
xenotransplants. Nature 391, 320-324.
Patience, C., Takeuchi, Y., and Weiss, R.A. (1997). Infection of
human cells by an endogenous retrovirus of pigs. Nature Med 3,
Allan, J.S., Broussard, S.R., Michaels, M.G., Starzl, T.E., Leighton,
K.L., Whitehead, E.M., Comuzzie, A.F., Lanford, R.E., Leland, M.M.,
Switzer, W.M. and Heneine, W. (1998). Amplification of simian retroviral
sequences from human recipients of baboon liver transplants. AIDS
RESEARCH AND HUMAN RETROVIRUSES 14, 821-824.
Paradis, K., Langford, G., Long, Z., Heneine, W., Sanstrom, P.,
Switzer, W.M., Chapman, L.E., Lockey, C., Onions, D., The Xen 111 study
Group and Otta, E. (1999). Search for Cross-Species Transmission of
Porcine Endogenous Retrovirus in Patients Treated with Living Pig
Tissue. Science 285, 1236-1241.
BBC Radio 4 Today, August 13, 2000 "Scientists
to stop experiments involving genetically modifying pigs" The
Guardian August 14, 2000.
Van den Bogaerde, J. and White, D.J.G. (1997). Xenogeneic
transplantation. British Medical Bulletin 53, 904-920.
Langford, G.A., Yannoutsos, N., Cozzi, E., Lancaster, R., Elsome, K.,
Chen, P., Richards, A. and White, D.J.G. (1994). Production of pigs
transgenic for human decay accelerating factor. Transplantation
Proceedings 26, 1400-1401.
Cozzi, E. and White, D.J.G. (1995). The generation of transgenic pigs
as potential organ donors for humans. Nature Medicine 1,
Wallwork J. (1997). Current status of xenotransplantation. International
Journal of Cardiology 62, S37-38.
Vial, C.M., Ostlie, D.J., Bhatti, F.N.K., Cozzi, E., Goddard, M.,
Chavez, G.P., Wallwork, J., Shite, D.J.G. and Dunning, J.J. (2000). The
Journal of Heart and Lung Transplantation 19, 224-229.
See for example, Kumpatla, S.P. and Hall, T.C. (1999). Organizational
complexity of a rice transgenic locus susceptible to methylation-based
silencing. IUBMB Life 48, 459-467.
Kuipers, H.W., Langford, G.A., and White, D.J.G. (1997). Analysis of
transgene integration sites in transgenic pigs by fluorescence in situ
hybridization. Transgenic Research 6, 253-259.
Meng, X.J., Purcell, R.H., Halbur, P.G., Lehman, J.R., Webb, D.M.,
Tsareva, T.S., Haynes, J.S., Thacker, B.J. and Emerson, S.U. (1997). A
novel virus in swine is closely related to the human hepatitis E virus,
Butler, D. (1998). Alternative ways of meeting demand. Nature
391, 325, 1998
"Heart transplants: medical miracle may be a waste of time and
money" Sarah Boseley, The Guardian, 1 September , 2000.
Tucker, A.S., Galbraith, D., McEwan, P. and Onions, D. (1999).
Evaluation of procine cytomegalovirus as a potential zoonotic agent in
xenotransplantation. Transplantation Proceedings 31, 915.
White, S.A. and Nicholson, M.L. (1999). Xenotransplantation. British
Journal of Surgery 86, 1499-1514.
Collignon, P. (1999). Transplants from pigs. Science 286,
"Coalition exposes flaws in pig transplant study" Campaign
for Responsible Transplantation Press Release, August 23, 1999.
Weiss, R.A. (1999). Xenografts and retroviruses. Science 285,
Ho, M.W., Traavik, T., Olsvik, R., Tappeser, B., Howard, V., von
Weizsacker, C. and McGavin, G. (1998). Gene Technology and Gene Ecology
of Infectious Diseases. Microbial Ecology in Health and Disease 10,
Tackle, S., Kurth, R. and Denner, J. (2000). Porcine endogenous
retroviruses inhibit human immune cell function : risk for
xenotransplantation? Virology 268, 87-93.
Martin, U., Steinhoff, G., Kiessig, V., Chickobava, M., Anssar, M.,
Morschheuse, T., Lapin, G., and Haverich, A. (1999). Porcine endogenous
retrovirus is transmitted neither in vivo nor in vitro from porcine
endothelial cells to baboons. Transplant Proc. 1, 913-914.
Blusch, J.H. Patience, C., Takeuchi, Y., Templin, C., Roos, C., Von
Der Helm, K., Steinhoff, G., Martin, U. (2000). Baboons unsuitable model
for pig human transplants: Infection of nonhuman primate cells by pig
endogenous retrovirus. J. Virol. 74, 7687-7690
"Cancer peril of animal organ transplants" Robin McKie,
The Observer, July 23, 2000.
"Public Meeting Reveals Weak Oversight of Xenotransplantation"
Campaign for Responsible Transplantation Press Release, Jan 17, 2000.
"ANTI-XENOTRANSPLANTATION COALITION DENOUNCES NEW FEDERAL
GUIDELINE" Campaign for Responsible Transplantation Press Release,
May 31, 2000.
See Radford, T. (1998). 'Test-tube' thumb helps surgeons move towards
own-body tissue. The Guardian, November 2; See the 25 February
2000 issue of Science, which contains many articles and features
on both embryonic and adult stem cells.
Alison, M.R., Poulsom, R., Jeffery, R., Dhillon, A.P., Quaglia, A.,
Jacob, J., Novelli, M., Prentice, G., Williamson, J. and Wright, N.A.
(2000). Cell differentiation: Hepatocytes from non-hepatic adult stem
cells. Nature 406, 257.