ISIS Report 22/07/05
GM Pharmaceuticals from Common Green Alga
Amid widespread protests against using crop plants to produce genetically
modified pharmaceuticals, companies are turning to the single-cell green alga
Chlamydomonas touted as a "safe" alternative; but not when it is to be
grown in largescale outdoor bioreactors. Prof.
Joe Cummins and Dr. Mae-Wan Ho
A fully referenced version
of this article is posted on ISIS members’ website. Details here
Largescale production of GM algae producing human proteins
The Hawaii Department of
Agriculture received an application from Mera Pharmaceuticals, originally filed
1 November 2004, for a permit to begin large-scale production of a genetically modified (GM) alga, Chlamydomonas reinhardtii, producing a
human immunoglobulin-A protein against a variant of the herpes simplex virus.
On 17 April 2005, Mera Pharmaceuticals revised their application to include
seven additional GM strains expressing a range of human antibodies,
interleukins and nerve growth factors (see Box) . The case for the first GM
strain was heard on 25 May 2005, and the Plant Quarantine Honolulu Office did
not grant the permit.
The application for the seven
other GM strains had been scheduled for 7 October 2005, but was brought forward to
29 June. On that occasion, the state Board of Agriculture gave its approval
. Mera Pharmaceuticals will begin the process of importing the microalgae
from The Scripps Institute, La Jolla, California, even as opponents are seeking to block the
permit, which was approved on a six-to-two vote after nearly three hours of
testimony and an hour of discussion.
Single cell green algae are
serving as green cell factories for producing pharmaceuticals, and have been
touted as a safe alternative to producing them in crop plants, because they can
be contained in the laboratory [3, 4]. Unfortunately, Mera’s proposed
large-scale cultivation is not contained. Anything from 500 to 26 000 litres of
culture are housed in transparent “Outdoor Photobioreactors”, which are cooled
with cold seawater with added chlorine . The exposed facilities are prone to
weather, storm and other damages, resulting in immediate massive contamination
of the marine environment. In addition, the use of chlorinated seawater for
cooling will be expected to impact on marine life. Neither of these concerns
appeared to have been addressed.
Henry Curtis, executive
director of Life of the Island, told the Board that the nonprofit organization will file
for a contested case hearing.
GM Chlamydomonas strains to
be imported into large-scale culture facility in Hawaii
1. Strain Hsv8, producing a full-length human immunoglobulin-A against
a variant of the herpes simplex virus
2. Strain aFceR 1r-1, producing
a protein targeting the Fc portion of the IgE molecule, thereby
limiting the interaction between circulating IgE molecules and receptors on
mast cells, which is turn limits the release of histamines and reduces
3. Strain aTNFr-1, producing IgG1 anti-tumor necrosis factor antibody.
4. Strain a TNr-1, producing IgG1 anti-microbial antibody.
5. Strain aCRr-1, producing IgG1d anti-cell proliferation antibody.
6. Strain aBSSsr-1, producing anti-cancer cell specific antibody.
7. Strain aIL 10r-1, producing various interleukins (including
interleukin 10, interleukin 13. interleukin 5 and interleukin 3)
8. Strain aARTr-1, producing neurotrophic factors to stimulate the
growth of new nerve tissue.
The GM algae
will be imported into Kona on the island of Hawaii to be
grown in the outdoor bioreactor system at Keahole Point at the state’s aquaculture
park Nelha. As Nancy Redfeather, Director of Hawaii Genetic Engineering Action
Network (Hawaii GEAN ) points out, this type of “field trial” of a
biopharmaceutical algae has never been attempted before in the United States, and all government agencies,
the Food and Drug Administration (FDA), the US Department of Agriculture (USDA)
and Environment Protection Agency (EPA) had waived oversight of the trial. The
native Hawaii algal systems
have not been well documented; nor has the ecology of Chlamydomonas itself. Apart from some last
minute attempt to conduct basic environmental experiments of the survival of GM
algae, there are no peer-review studies, or studies of any kind to back up Mera
Pharmaceutical’s claim of “no harm to the environment or human health.”
Written and oral testimony by the public was overwhelmingly opposed to the
project. The board also ignored testimony and reports by a number of local algae
experts from the University of Hawaii, Manoa, the State Biologist and Maui Country
District Health Officer. Other scientists providing testimony include R. Malcolm
Brown Jr., the Johnson and Johnson Centennial Chair in Plant Cell Biology at
the University of Texas at Austin; Marti Crouch, Doug Sherman from Friends of
the Earth, and Joe Cummins and Mae-Wan Ho from ISIS.
Malcolm Brown’s message to the board was, “Hawaii is still the supreme ecosystem
on earth to understand the dynamics of evolution and natural selection. Let’s
not forever lose this opportunity because a few commercial operations thoughtlessly
tried to construct mass scaleup of genetically modified organisms in Hawaii.”
Nancy Redfeather said, “It was indeed a sad day for the native algae of Hawaii
The Chlamydomonas reinhardtii transformation
reinhardtii is a preferred organism for molecular pharming by chloroplast transformation because its nuclear and chloroplast
genomes have both been sequenced, and it has a long history of laboratory
culture. In addition, it has a
single chloroplast, which makes it easy to produce a uniformly transformed
culture. One technical drawback with the alga is codon bias related to the high
GC content of the algal DNA, To achieve significant production, the code of human and
many other genes must be altered to fit the bias of the algal cell, For that
reason the human pharmaceutical products are produced from synthetic
approximations of the human gene . The synthetic human DNA in the alga, and
all the more so, the expressed transgene, should not be deemed equivalent to
the human gene and gene product until they have been tested for untoward
effects on humans and other organisms in the environment. It is already known
that the proteins are not subject to post-translational modifications  as
they would be in human cells, and hence likely to be treated as ‘foreign’ by
the human immune system.
Mayfield and Franklin described
construction of transgenic Chlamydomonas
reinhardtii whose chloroplasts had been modified to express human
antibodies . The human genes were extensively
adjusted for codon bias. Either the rbcl
(ribulose-1,5-bisphosphate carboxylase of the chloroplast) promoter or the atpA (alpha-subunit of the chloroplast ATP
synthase of the chloroplast) promoter were used to drive the antibody gene, with the rbcL transcription terminator following the human gene. A
16S ribosomal subunit with resistance to the antibiotic accompanied the human
gene transformation. Using this system, IgA antibody directed against herpes
simplex virus were produced , as well as single chain antibodies against the
herpes virus . The numerous codon alterations to optimize production of
recombinant protein in the alga have been described . Code optimization need
not change the amino acid sequence of the protein produced from
the recombinant gene, but Mera’s petition contained no proof that the protein
produced from the synthetic gene was identical to the original human gene, nor the fact that the
transgenes were synthetic approximations to the human genes.
Although not prominently stated, the GM
strains probably all contained in addition a kanamycin resistance marker gene
, which is necessary to select for the transformed cells.
Risks from the GM alga
The claim that the risk from
contact with the recombinant products was negligible even in the worst case is groundless, because no experiments were reported
to support that conclusion.
As pointed out
already (“Molecular pharming by chloroplast transformation”, this series),
producing pharmaceuticals in chloroplasts entails specific risks due to the
large quantities of transgenic proteins produced and the hazards of horizontal
gene transfer to bacteria due to homologies between the chloroplast and
Large quantities of transgenic proteins are
produced from multiple copies of transgenes present, in the case of Chlamydomonas, about 50 to 100 per cell.
Strains 2-6 (see Box) produce antibodies that bind to immunologically active
proteins that could lead to anaphylaxis (severe life-threatening allergic
reaction) following repeated exposure. Strain 7 produce interleukins, potent
regulators of immune functions active in minute quantities. Pulmonary exposure
to interleukin 13, for example, causes inflammation, mucus hypersecretion,
physiologic abnormalities associated with asthma , while interleukin 10 is a
powerful immune suppressant . Strain 8 produces unspecified neurotrophic
factors to stimulate growth of nerve tissue, again, potent molecules active at
very low concentrations, whose effects, especially at high concentrations are
An additional hazard from the gene products
is that they are in all cases not the same as the human protein, because of the
changes in making the synthetic gene copies, and because there is no
post-translational processing. They may hence be treated as immunologically
‘foreign’ by the human immune system, resulting in dangerous complications.
Horizontal transfer of
the GM Chlamydomonas transgenes
are likely to occur in all environments, particularly in the soil, where Chlamydomonas is commonly found, but also
in the marine environment and in the gastrointestinal tract of all animals.
Horizontal transfer of transgenes can occur both from the accidental release of
genetically modified Chlamydomonas
reinhardtii itself, or from the intentional release of transgenic
DNA in the large amounts of transgenic wastes that are likely to be discharged
from the large scale culture facilities into the environment.
As already mentioned,
horizontal transfer and recombination of transgenes could create new bacteria
and viruses that cause diseases and spread antibiotic resistance marker genes
to the pathogens