Expression and Manufacture of Pharmaceutical Proteins in Genetically Engineered Horticultural Plants - Transgenic Horticultural Crops: Challenges and Opportunities - page 105

The.amount.of.these.compounds.varies.between.plant.species.. Tobacco.and.related.
Nicotiana .plant.leaves.generally.contain.high.levels.of.phenolics.and.toxic.alkaloids..
However,.tobacco.cultivars.with.low-alkaloid.and.other.toxic.metabolites.have.been.
developed. for. pharmaceutical. protein. production. 230 . Protein. stability. is. one. of. the.
major.issues.for.leafy.crops..Transgenic.proteins.expressed.in.leaves.accumulate.in.a.
more.aqueous.environment.and.thus.tend.to.be.unstable..This.translates.into.a.short.
target.protein.shelf.life.in.harvested.green.biomass.due.to.proteolysis..They,.therefore,.
must.be.quickly.transported.to.a.processing.facility.at.a.controlled.temperature.to.be.
extracted.for.target.protein.or.to.be.processed.by.freezing,.drying,.or.other.means.for.
short-term.storage..In.contrast,.pharmaceutical.proteins.produced.in.cereal.seeds.can.
remain.stable.for.a.long.period.at.ambient.temperature.(see.below)..Regulatory.con-
cerns.for.nonfood,.nonfeed.pharmaceutical.producing.leafy.crops.include.the.potential.
leaching.of.recombinant.products.into.the.environment.and.the.exposure.of.herbivores.
to.transgenic.proteins..Again,.this.type.of.risks.can.be.reduced.by.using.inducible.pro-
moters.to.drive.the.expression.of.the.transgene.so.that.transgenic.proteins.can.only.be.
synthesized.in.a.controlled.environment.post.biomass.harvest. 229,231
S eed c ropS
Expression. of. pharmaceutical. protein. has. been. explored. in. seeds. of. several. crops.
including.maize,.rice,.wheat,.barley,.tobacco,.oilseed.rape,.and.a.variety.of.beans. 232-234 .
For. certain. transgenic. proteins. that. required. long-term. storage. in. biomass. before.
processing,.expression.in.seeds.driven.by.seed-speciic.promoters.is.preferred.over.
leafy.expression.systems..In.contrast.to.the.“watery”.leaves,.seeds.are.natural.desic-
cators,.which.preserve.functional.proteins.by.a.drying.process.during.their.matura-
tion. and,. therefore,. allow. long-term. protein. stability. during. storage.. Studies. have.
shown.that.mAbs.produced.in.corn.seeds.remain.stable.after.3.years.of.storage.at.
room.temperature.and.retain.their.biological.activities. 235 .Phenolic.and.alkaloid.com-
pounds.are.not.present.in.cereal.crop.seeds,.thus.simplifying.downstream.puriica-
tion.process.and.reducing.the.overall.production.cost.accordingly..Cereal.seeds.do.
contain.higher.levels.of.lectins.that.could.be.dificult.to.remove.from.the.inal.prod-
uct.especially.if. the. target.proteins.are.glycosylated..Legumes.and.other.oil.crops.
such. as. oilseed.rape.are.useful. alternative.systems.for. pharmaceutical.production,.
because.their.oil.bodies.can.be.exploited.to.facilitate.target.protein.puriication..For.
example,.SemBioSys.Genetics.has.developed.an.oleosin-fusion.technology.to.target.
pharmaceutical.proteins.to.oil.bodies..As.a.result,.these.proteins.can.be.puriied.by.
a. simple. extraction. process. 236 . The. level. of. expression. of. recombinant. proteins. in.
seeds.is.generally.lower.than.that.in.tobacco.leaves..This.is.mostly.due.to.the.fact.
that.many.of.the.seed.expression.systems.are.still.at.early.developing.stage.and.have.
ample.rooms.for.further.optimization. 234 .An.inherent.disadvantage.of.producing.pro-
tein.in.seeds.is.that.it.can.only.be.evaluated.until.seeds.are.produced.after.a.long.
growth.cycle..Within.the.three.expression.platforms,.only.nuclear.stable.transforma-
tion.is.feasible.for.seed.expression..Transient.expression.techniques.for.seed.expres-
sion. have. not. been. well. established,. therefore,. hindering. the. quick. assessment. of.
expression.feasibility.for.a.given.product..Seed.crops.offer.both.biosafety.advantages.
and.drawbacks.for.expression.of.recombinant.proteins..On.one.hand,.it.reduces.the.