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[SANET-MG] algal recombinant pharmaceuticals in Hawaii
June 18, 2005
Prof. Joe Cummins
“Algal recombinant pharmaceuticals in Hawaii”
The Hawaii Department of Agriculture ,on 25 May 2005, heard a proposal
from Mera Pharmaceuticals for a permit to begin large scale production
of a genetically modified alga, Chlamydomonas reinhardtii, modified to
express a full length immunoglobulin-A moleculae against a variant of
the herpes simplex virus .the permit was denied by thee Plant Quarantine
Honolulu Office. However another application from Mera Pharmaceutical
has been scheduled for October 7,2005, it calls for an additional 7
strains to be imported and produced in the Mera facility in Hawaii. The
strains included the following:
1. Strain aFceR 1r-1:Molecule targeting the Fc
portion of the IgE molecule limiting the interaction
between circulating IgE molecules and receptors on
mast cells, which is turn limits the release of
histamines and reduces inflammation.
2. Strain aTNFr-1: IgG1 anti-tumor necrosis factor
antibody.
3. Strain a TNr-1: IgG1 anti-microbial antibody.
4. Strain aCRr-1: IgG1d anti-cell proliferation
antibody.
5. Strain aBSSsr-1: Anti-Cancer cell specific
antibody.
6. Strain aIL 10r-1: Various interleukins (various including
interleukin 10, interleukin 13. interleukin 5 and interleukin 3)
7. Strain aARTr-1: Neurotrophic factors to stimulate
frowth of new nerve tissue.
The petition from Mera Pharmaceutical described the Chlamydomas strains
in broad detail with little or no molecular detail describing the actual
genetic makeup of the strains. The petition provided some broad and
groundless assurances such as in the “worst case scenario” in a
complete failure of there reactor growth facility the risk from a health
perspective would be negligible because the strains , as well as their
expressed proteins are completely harmless unless injected. Since the
petition did not provide adequate information about the genetic and
molecular modifications of the alga the following discussion will deal
with some considerations about the genetic modification Chalamydomonas
and actual concerns about the products produced in the modified algae.
Transgenic microalgae serve as green cell factories for producing
pharmaceuticals. Either chromosomal or chloroplast transgenes can be
produced with ease and a number of transgenic algae have been prepared.
Chlamydomonas reinhardii is a preferred organism for such experiments
because its genome has been sequenced and it has a long history of
laboratory culture One technical drawback with the alga is codon bias
related to the high GC content in 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(1). The synthetic human DNA in the alga should not be deemed
equivalent until it has been tested for untoward effects on humans and
the environmental biota.
Mayfield and Franklin described construction of transgenic
Chlamydomonas reinhardii whose chloroplasts had been modified for
expression of human antibodies. The human genes were synthetic
approximation extensively adjusted for codon bias. Either the rbcl
promoter (ribulose-1,5-bisphosphate carboxylase of the chloroplast) or
the atpA promoter (alpha -subunit of the chloroplast ATP synthase of
the chloroplast) were used to drive the antibody gene. The rbcL
transcription terminator followed the human gene. A 16s (small)
ribosomal subunit with resistance to the antibiotic accompanied the
human gene transformation(2). IgA antibody directed against herpes
simplex virus were produced in Chlamydomonas reinhardii (3). Single
chain antibodies against the herpes virus were produced in Chlamydomonas
reinhardii (4). In order to achieve adequate production of the human
gene products the DNA code of the human gene the genetic code must be
optimized for the codon bias of Chlamymononas. The code letter
alterations are rather extensive so that the synthetic human DNA
inserted into Chlamydomonas bears little resemblance to the original
human gene. The numerous codon alterations to optimized production of
recombinant protein in the alga have been described (5). Of course,
code optimization need not lead to alteration in the amino acid sequence
of the protein produced from the recombinant gene but the protein
produced from the synthetic gene must be proved to be identical to the
protein produced from the original human gene. In the petition for
production both codon optimization by producing synthetic gene and the
proof that the protein produced from the synthetic gene was identical
to the original human gene was not mentioned.
The petition failed to provide minimal information about the genetic
makeup of the strains to be introduced into Hawaii. On top of that the
fact that synthetic approximations of human genes was not mentioned. The
claim that the risk of contacting the recombinant products was
negligible even in the worst case seems to have been groundless because
there were no experiments reported that support that conclusion.
For example, strain aFceR produces 1r-1 molecule targeting the IgE to
reduce inflammation. The petition did not provide evidence showing that
the protein was inactivated on ingestion nor did it discuss problematic
side effects. Strain TNFr-1 produces anti-tumour necrosis factor
antibody, as above there was no evidence that the protein was
inactivated on ingestion nor was there any discussion of the a potential
immune response leading to anaphylaxis following repeated exposure.
Strain TNr-1 produces IgG1d antimicrobial antibody, that antibody seems
unlikely to be inactivated by ingestion and repeated exposure is likely
to result in anaphylaxis. Strain aCRr-1 produces anti-cancer
proliferation antibody, there was no evidence presented on the
destruction of the antibody on ingestion nor of the problem of
anaphylaxis following repeated exposure. Strain aIL 10r-1produces
various interleukins (various including interleukin 10, interleukin 13.
interleukin 5 and interleukin 3). Interleukins are very potent
regulators of the immune system and they are active in minute
quantities. The petition failed to provide evidence that the
interleukins were destroyed on ingestion but pulmonary exposure to
interleukin 13 , for example, causes inflammation, mucus hypersecretion
, physiologic abnormalities associated with asthma (6). Interleukin 10
is a powerful immune suppressant (7).The many adverse effects of the
interleukins should have been discussed in the application but were not.
Strain aARTr-1 produced unspecified neurotrophic factors to stimulate
growth of nerve tissue. Since the factors were ill defined it is not
possible to evaluate any effects of the nerve factors or their stability.
In conclusion, the Mera Pharmaceuticals application for a permit to
produce recombinant pharmaceuticals in Hawaii employing recombinant
Chlamydomonas algae was defiecient in the main areas related to the
safety of the recombinant microbes. The molecular genetic description
of the genetic modification were simply ignored, there was no
acknowledgement that the strains were modified with synthetic
approximations of the human genes and no effort was made to insure that
the products of the synthetic genes were identical to the human proteins
that they approximated. Even if the products of the synthetic genes can
be shown to be identical to the products that they approximate the claim
that such products are inactive on ingestion should have been documented
with experiments. It staggers the mind to think that an application
lacking fundamental safety information might be approved anywhere.
References
1. Leon-Banares R, Gonzalez-Ballester D, Galvan A. and Fernandez E.
Transgenic microalgae as green cell-factories. Trends Biotechnol. 2004
Jan;22(1):45-52
2. Mayfield,S. and Franklin,S. Expression of human antibodies in
eukaryotic micro-algae Vaccine 2005 ,23,1828-32
3. Franklin S and Mayfield S . Recent developments in the production of
human therapeutic proteins in eukaryotic algae. Expert Opin Biol Ther.
2005 Feb;5(2):225-35
4. Mayfield SP, Franklin SE and Lerner RA. Expression and assembly of
a fully active antibody in algae. Proc Natl Acad Sci U S A. 2003 Jan
21;100(2):438-42
5. Franklin S, Ngo B, Efuet E and Mayfield S. Development of a GFP
reporter gene for Chlamydomonas reinhardtii chloroplast. Plant J. 2002
Jun;30(6):733-44
6. Zhu Z, Homer RJ, Wang Z, Chen Q, Geba GP, Wang J, Zhang Y and Elias
J Pulmonary expression of interleukin-13 causes inflammation, mucus
hypersecretion, subepithelial fibrosis, physiologic abnormalities, and
eotaxin production. J Clin Invest. 1999 Mar;103(6):779-88
7. Kamradt T, Goggel R and Erb K Induction, exacerbation and inhibition
of allergic and autoimmune diseases by infection. Trends Immunol. 2005
May;26(5):260-7
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