Biomedical Engineering Reference
In-Depth Information
lag to routine manufacture can, therefore, be almost 3 years in the case of cows or 7 months
in the case of rabbits. Furthermore, if the original transgenic embryo turns out to be male, a
further delay is encountered, as this male must breed in order to pass on the desired gene to
daughter animals, who will then eventually produce the desired product in their milk.
Another general disadvantage of this approach relates to the use of the microinjection technique
to introduce the desired gene into the pronucleus of the fertilized egg. This approach is ineffi cient
and time consuming. There is no control over issues such as if/where in the host genomes the
injected gene will integrate. Overall, only a modest proportion of manipulated embryos will cul-
minate in the generation of a healthy biopharmaceutical-producing animal.
A number of alternative approaches are being developed that may overcome some of these is-
sues. Replication-defective retroviral vectors are available that will more consistently (a) deliver a
chosen gene into cells and (b) ensure chromosomal integration of the gene. A second innovation
is the application of nuclear transfer technology.
Nuclear transfer entails substituting the genetic information present in an unfertilized egg with do-
nor genetic information. The best-known product of this technology is 'Dolly' the sheep, produced by
substituting the nucleus of a sheep egg with a nucleus obtained from an adult sheep cell. (Genetically,
therefore, Dolly was a clone of the original 'donor' sheep.) An extension of this technology applicable
to biopharmaceutical manufacture entails using a donor cell nucleus previously genetically manipu-
lated so as to harbour a gene coding for the biopharmaceutical of choice. The technical viability of this
approach was proven in the late 1990s upon the birth of two transgenic sheep, 'Polly' and 'Molly'. The
donor nucleus used to generate these sheep harboured an inserted (human) blood factor IX gene under
the control of a milk protein promoter. Both produced signifi cant quantities of human factor IX in their
milk. The fi rst (and only, at the time of writing) such product to gain approval anywhere in the world
is 'Atryn', a recombinant human antithrombin that is produced in the milk of transgenic goats. Atryn is
used to treat thromboembolism in surgery of people with congenital antithrombin defi ciency.
In addition to milk, a range of recombinant proteins have been expressed in various other tar-
geted tissues/fl uids of transgenic animals. Antibodies and other proteins have been produced in
the blood of transgenic pigs and rabbits. This mode of production, however, is unlikely to be pur-
sued industrially for a number of reasons:
•
Only relatively low volumes of blood can be harvested from the animal at any given time point.
•
Serum is a complex fl uid, containing a variety of native proteins. This renders purifi cation of the
recombinant product more complex.
•
Many proteins are poorly stable in serum.
•
The recombinant protein could have negative physiological side effects on the producer animal.
Therapeutic proteins have also been successfully expressed in the urine and seminal fl uid of
various transgenic animals. Again, issues of sample collection, volume of collected fl uid and the
appropriateness of these systems render unlikely their industrial-scale adoption. One system that
does show industrial promise, however, is the targeted production of recombinant proteins in
the egg white of transgenic birds. Targeted production is achieved by choice of an appropriate
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