Questions on pharming, recombinant medicines and synthetic biology

Question 1

A company will synthesise DNA of any sequence to order and sell it to research laboratories, and has patented the engineered yeast it uses in its own drug production. Evaluate the biosecurity and ownership concerns raised by work of this kind.

Accepted Answer

B1 the biosecurity worry is concrete: poliovirus was assembled from mail-ordered DNA fragments in 2002 and an extinct relative of smallpox was synthesised in 2017 for around one hundred thousand dollars, so recreating a dangerous pathogen needs money and a published sequence rather than rare skill B1 against that, the same openness lets a public-health laboratory synthesise a new pathogen's genes within days of its sequence appearing, which is how vaccine development now begins B1 the case for patents is that development costs run to hundreds of millions, and a period of exclusivity pays for the engineering, the animals and the trials B1 the case against is access and displacement: a patented medicine is priced by its owner and the diseases involved are concentrated where patients are poorest, and a fermenter making artemisinic acid competes with the smallholders who grow wormwood for the same drug B1 a judgement supported by those points, for example that the service should continue with every order screened against the sequences of known dangerous pathogens and the customer verified, while noting that such screening is voluntary and not every supplier carries it out

Question 2

Describe how a goat is engineered to secrete a human protein into its milk, and describe why the protein appears there and nowhere else in the animal.

Accepted Answer

B1 the human gene is joined downstream of the promoter of a milk protein gene, such as the promoter of β-lactoglobulin B1 the construct is microinjected into the nucleus of a fertilised egg, or introduced into cultured cells from which an embryo is cloned by nuclear transfer B1 the embryo is implanted into a surrogate, and the offspring are tested for the transgene; a transgenic female carries the gene in every cell of her body B1 the transcription factors that switch that promoter on are present only in mammary gland cells during lactation, so only those cells transcribe the gene and the protein is purified from milk rather than from blood

Question 3

Discuss the welfare arguments for and against producing medicines in the milk of transgenic livestock.

Accepted Answer

B1 against: founding the line is inefficient, because microinjection succeeds in only a small percentage of eggs, so each transgenic animal costs many surrogate pregnancies, failed implantations and discarded embryos, and cloning adds its own late pregnancy failures B1 against: redesigning an animal's genome so that it serves as production equipment treats a sentient creature as an instrument, whatever its day-to-day conditions B1 for: once the founder exists the producing herd is bred normally and milking does not harm the animal, and the commercial need for clean, consistent milk means the animals are housed and fed to a high standard B1 for: the alternative sources are pooled human plasma, with the infections it transmitted, or mammalian cell culture at a price some health systems will not pay, so the welfare cost has to be weighed against the patients who would otherwise go untreated

Question 4

Compare producing a human protein in mammalian cell culture with producing the same protein in the milk of a transgenic goat, referring to typical yield and to the starting material that has to be purified.

Accepted Answer

B1 mammalian cell culture is effective but expensive, requiring the cells to be grown slowly in a rich sterile medium, and yielding only milligrams per litre for a complex protein such as factor VIII B1 a lactating transgenic animal can secrete the protein into its milk at reported yields that vary widely between constructs and species, from milligrams up to several grams per litre B1 milk is a simpler starting material to purify than cell culture medium, since routine collection by milking replaces the sterile, continuous maintenance a cell culture demands B1 whichever route is used, a claimed yield should be quoted with its source rather than as a general figure, because it varies so widely between different constructs and species

Question 5

Describe how the artemisinin supply chain changed once an engineered yeast pathway became available, referring to what the yeast is engineered to produce and to what happens to that product afterwards.

Accepted Answer

B1 before the engineered pathway, artemisinin was extracted from sweet wormwood, a crop whose yield and price varied substantially between years depending on the harvest B1 a pathway of enzymes, some from the plant and some derived from yeast's own metabolism, was assembled in brewer's yeast B1 the engineered yeast converts sugar into artemisinic acid rather than the finished drug itself B1 industrial chemical steps then convert the artemisinic acid into artemisinin, and production at this scale began in 2013, giving a supply that did not depend solely on the harvest while it ran

Question 6

Discuss the argument that patenting an engineered organism or a standard genetic part slows down, rather than speeds up, the wider benefit of synthetic biology.

Accepted Answer

B1 for slowing benefit: a patented medicine or process is priced by its owner, and the diseases synthetic biology could treat most cheaply are often concentrated where patients are poorest B1 for slowing benefit: fermenter production of a compound such as artemisinic acid competes with existing small-scale producers, such as the farmers who grow wormwood, displacing an established supply rather than only adding a new one B1 against, for speeding benefit: patents are the usual justification for development costs running to hundreds of millions, and without a period of exclusivity a company has less reason to fund the engineering, the animals and the trials at all B1 the boundary is not fully settled in law either: an engineered gene sequence can be patented, though a naturally occurring human gene has not been patentable in the United States since 2013, but whether the overall design of an entire organism can be owned in the same way remains an open question

Question 7

The enzyme taliglucerase alfa, a glycoprotein used to treat Gaucher disease, is made in genetically modified carrot cells. Explain the advantages of producing a human glycoprotein in genetically modified plant cells rather than in bacteria or in the milk of transgenic animals.

Accepted Answer

B1 plant cells are eukaryotic, so they fold the protein and add sugar chains to it, which bacteria cannot do B1 no human or animal virus or prion can grow in plant cells, so the product cannot be contaminated by one, unlike a product from animals or from blood B1 plants or plant cells are cheap to grow and production can be scaled up easily B1 there is no need to produce and keep a herd of transgenic animals, so the welfare objections to founding and maintaining one do not arise

Question 8

State the condition treated with recombinant factor VIII, state the condition treated with recombinant adenosine deaminase, and state what recombinant factor VIII replaced.

Accepted Answer

B1 factor VIII is the clotting protein deficient in haemophilia A, the commonest severe haemophilia B1 adenosine deaminase is the enzyme missing in ADA-deficient severe combined immunodeficiency B1 recombinant factor VIII replaced factor VIII concentrated from donated blood plasma, pooled from thousands of donors per batch, which transmitted HIV and hepatitis C to a large fraction of the people treated in the early 1980s

Question 9

Insulin is manufactured in genetically modified bacteria, but factor VIII is made in cultured mammalian cells. Explain why factor VIII cannot be produced usefully in Escherichia coli.

Accepted Answer

B1 factor VIII is a large glycoprotein: it must be folded with help, have its disulfide bridges formed in the right places, and have sugar chains added in the Golgi apparatus B1 a prokaryote can transcribe and translate the human gene but has no endoplasmic reticulum and no Golgi apparatus, so it cannot carry out that post-translational modification B1 what accumulates has the right primary structure and the wrong tertiary structure and no glycosylation, so it is inactive, whereas insulin is small enough to need almost none of that modification

Question 10

A transgenic goat secretes antithrombin at 2.5 g per dm³ of milk and yields 800 dm³ of milk in a year. A mammalian cell culture yields 50 mg of the same protein per dm³ of medium. Calculate the mass of antithrombin one goat supplies in a year, and calculate the volume of culture medium needed to match it.

Accepted Answer

M1 mass from the goat is the concentration multiplied by the volume of milk, so 2.5 × 800 A1 2000 g, which is 2.0 kg of antithrombin a year A1 the culture yields 0.050 g per dm³, so 2000 ÷ 0.050 = 40 000 dm³ of medium would be needed

Question 11

Outline what synthetic biology adds to genetic engineering, referring to standard parts, genome synthesis and engineered pathways.

Accepted Answer

B1 standard parts are promoters, ribosome binding sites, genes and terminators catalogued with defined, measured behaviour and designed so that any part joins to any other, so a device is assembled from a registry rather than hunted for one sequence at a time B1 genome synthesis means whole genomes built chemically from sequence data: a bacterial genome of about a million base pairs was synthesised and installed in a recipient cell in 2010, and the cell ran on it B1 an engineered pathway assembles several genes from several organisms so that a cell runs a production line it never had, as in the yeast that ferments sugar into artemisinic acid, a few chemical steps short of the antimalarial artemisinin

Question 12

Describe why founder transgenic animals must be screened for mosaicism before a production line is established.

Accepted Answer

B1 microinjection of the DNA construct into a fertilised egg can produce a mosaic founder, in which only some of the animal's cells carry the inserted gene B1 an animal that is mosaic in the cells relevant to expression, such as the mammary gland, may secrete little or none of the protein even though the construct was successfully introduced B1 offspring are therefore tested individually for the transgene, and a confirmed line, descended from an animal shown to express the gene reliably, is established before large-scale production begins

Question 13

A regulator requires every batch of a drug produced in the milk of a transgenic herd to be tested, and requires the herd itself to be certified free of particular infections. Explain the reason for each requirement.

Accepted Answer

B1 milk composition varies with the individual animal, the season and the feed, so a batch collected at one time is not guaranteed to be identical to the last, and each must be purified and tested to prove that it is B1 an animal-derived product can carry pathogens the animal itself is infected with, so the herd must be certified free of infections, and of prion disease in particular, that could pass into the product B1 both requirements exist because a drug from a living animal poses risks that a batch produced by a sterile industrial process from defined chemical starting materials does not pose in the same way

Question 14

Explain why the goats used to produce a licensed drug in their milk must be kept in a closed, contained herd rather than allowed to breed with ordinary goat populations.

Accepted Answer

B1 a transgenic goat carries the human gene in every cell of its body and could pass it on to any offspring it breeds with B1 if a transgenic animal bred into an ordinary herd, the gene would spread into a population from which it could never be recalled or removed B1 containment, keeping the herd closed and its animals tracked individually, prevents that spread while the drug continues to be produced from the certified line

Question 15

A synthesis company screens every DNA order against a database of known dangerous pathogen sequences before manufacturing it. Suggest what this screening is intended to prevent, and suggest one limitation of relying on it.

Accepted Answer

B1 the screening is intended to stop a customer using the service to obtain the DNA needed to assemble a dangerous pathogen, such as poliovirus or an extinct relative of smallpox, both of which have been assembled from synthesised DNA fragments B1 screening only catches an order if the sequence matches something already in the database, so a novel or disguised dangerous sequence could still be missed B1 screening is voluntary and coverage differs between suppliers, so a customer refused by one company could in principle approach a supplier that does not screen orders

Question 16

State what is meant by pharming, and state one advantage milk has as a starting material for purification compared with mammalian cell culture medium.

Accepted Answer

B1 pharming is the use of genetically modified organisms, including plants and animals, to produce pharmaceutical substances B1 milk is a simpler starting material to purify than cell culture medium, and is collected non-invasively by the routine act of milking

Question 17

Suggest why many plant-made medicines are produced in tobacco relatives grown in greenhouses rather than in food crops grown in open fields.

Accepted Answer

B1 pollen from an open field could carry the transgene into other plants, and seed could end up in the human or animal food supply, so people could eat the medicine unknowingly B1 a non-food plant in a closed greenhouse cannot be confused with food and its pollen and seed are contained, so the transgene does not escape