Sources and upstream processing - Pharmaceutical Biotechnology: Concepts and Aapplications - page 122

Silkworms reared in
cabinets

Innoculation of silkworms
with engineered silkworm
virus

Continue rearing of
(innoculated) silkworms (5
days)

Product extraction in
0.1 M HCl

Mechanical disruption of
silkworms

Neutralization of extract
with NaOH

Clarification of solution
(filtration/centrifugation)

Dye affinity
chromatography

Immobilized metal affinity
chromatography Filling and freeze drying Adjustment of potency and

excipient addition

Desalting (gel filtration)

Figure 5.4 Overview of the industrial manufacture of the IFN- ω product 'Vibragen Omega'. Refer to text for

details

An alternative insect cell-based system used to achieve recombinant protein production entails
the use of live insects. Most commonly, live caterpillars or silkworms are injected with the engi-
neered baculovirus vector, effectively turning the whole insect into a live bioreactor. One veteri-
nary biopharmaceutical, Vibragen Omega, is manufactured using this approach, and an overview
of its manufacture is outlined in Figure 5.4. Briefl y, whole, live silkworms are introduced into
pre-sterile cabinets and reared on laboratory media. After 2 days, each silkworm is inoculated
with engineered virus using an automatic microdispenser. This engineered silkworm polyhedrosis
virus harbours a copy of cDNA coding for feline IFN- . During the subsequent 5 days of rearing,

a viral infection is established and, hence, recombinant protein synthesis occurs within the silk-
worms. After acid extraction, neutralization and clarifi cation, the recombinant product is purifi ed
chromatographically. A two-step affi nity procedure using blue sepharose dye affi nity and copper
chelate sepharose chromatography is employed. After a gel fi ltration step, excipients (sorbitol and
gelatin) are added and the product is freeze-dried after fi lling into glass vials.

ω

5.3 Upstream processing

Biopharm production can be divided into 'upstream' and 'downstream' processing ( Figure 5.5). Up-
stream processing refers to the initial fermentation process that results in the initial generation of prod-
uct, i.e. the product biosynthesis phase. Downstream processing refers to the actual purifi cation of the
protein product and generation of fi nished product format (i.e. fi lling into its fi nal product containers,