ceng 109 class 5

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CENG 109 class 5

Bioprocess Engineering

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Class announcement Class project

Team forming situation Schedule for library training will be posted on

class web site. Attendance is mandatory

Make-up class in early October (to

replacing missing class on October 24)

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Outline for this class

The definition and scope of bioprocess

engineering

The technical challenges for bioprocess

engineers

Case studies of early biotechnology products-Insulin

-tPA

The economic impacts of bioprocessengineering

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Science and the application of

science are linked as the fruitis to the tree

Louis Pasteur

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Definition of

Bioprocess Engineering

Discovery in

Basic Science

Bioprocess Engineering PracticalProducts,

Processes

or Systems

Market Force

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What is bioprocess engineering about?

A bioprocess includes an

upstream process and adownstream process

Upstream: product production

in cell cultures/fermentations

Downstream: isolation of

product and its formulation

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What is bioprocess engineering? the design and construction of a

bioreactors (fermentation tanks) to growcells

the design and utilization of different

equipments to isolate the products

traditional engineering approaches for

scaling up and minimizing costs

the design and construction of living

organisms to optimize product production

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Technical Challenges

Number of protein products:from almost none to many

Complexity of protein products: High Liability of protein products: High

Purity requirement: High Scale of production and product

pricing: Highly variable

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Unit value and production quantities

of early biotech products

0.24840,000Erythropoietin

8.723,000Tissue plasminogen

activator (tPA)

5.735,000Growth Hormone

530375Human insulin

20,00010Human serum albumin

Amount of Productfor $200M in sales,

kg

Approx. selling price$/g

Product

Source: National Academy of Sciences, Committee on Bioprocess Engineering (1992)

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Insulin case studyThe first production process of human insulin involved

separate expression of A and B chains.

To stablize each chain, each chain is fused to a largerprotein to form insoluble granules. The fusion proteins are

subsequently removed and the two chains recombined .

Fi ure extracted from Biose arations En ineerin : Princi les Practice and Economics b Ladish

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Upstream process made more

efficient by mimicking nature

This figure

Illustrates

insulin synthesisin pancreas

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The proinsulin method is adopted

to improve production efficiency


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The proinsulin method is adopted

to improve production efficiency

The one-chain proinsulinmethod has been adopted

to improve the yield of

correctly folded proteins.

What kinds of expertise

are required to achievethis improvement?

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Isolation of human insulin requires

27 stepsWhy multiple steps are

needed?

Principles of orthogonalityProduct is the minority, to

be selected among many

different biomolecules and

cell debris

Similarity bewteen product

and

JunksRegulations require high

product purity (contaminants

below parts per millionrange)


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Product yield loss amplified by the

number of purification steps

Overall Product Yield = (1- L%)n x 100%

Lets calculate the yield for a 10-step processwith a step loss of 5%

The early insulin production process has a

overall yield of 29%=> Lower product yield is translated to higher

manufacturing cost

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tPA case study Tissue plasminogen

activator can dissolveblood clots --- is atherapeutic for treatingheart attacks and strokes

Sold as Activase, its

sale accounted for 30%of total sales ofGENENTECH, andabout 1/3 of theworldwide sales in bloodclot dissolvingtherapeutics in 1995

Unlike insulin, it isregulated as a biologic,not a drug

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Production of tPA: a unique opportunity for

the application of recombinant technology

50,000L of blood required to extract single

dose of tPA (~100mg) not practical

Recombinant technology provides afeasible approach

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4NoneGlycosylation

sites

123Disulphide

bonds

66,000 Da5,600 DaSize

tPAInsulin

But tPA is much more

complicated than insulin

.and achieving correct protein folding is more difficult

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Molecular arrangement of tPA

Lets count the no. of disulphide bridges (solid bars), potential cleavage sites

(arrows), glycosylation sites (zigzag lines) and short peptide bridge (dotted line)

Figure extracted from Bioseparations Engineering: Principles, Practice and Economics by Ladish

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Upstream process can be made

more efficient by mimicking nature

YesNoSecrete products tomedium?

ExtensiveNonePosttranslationalmodification?

YesNoRemove introns?

HighLowMaintenance cost

LowHighReplication rate

Mammalian Cells

(e.g. Chinese

Hamster Ovary Cells)

Bacteria

(E.Coli)

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Mammalian cell culture creates

new downstream challenges

Fear of transfer of infectious agents

Fear of genetic materials from cell linescausing cancer

Purification process must be able to removecells, virus and DNA

Analytical assays must be able to detectvery low level of cells, virus and DNA

Lead to advance in downstream processingunits, particularly membrane andchromatography applications

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Get it fast and get it right

the first time

Bioprocess cost is secondary to being firstto market

But cost of product is important in

subsequent competition with rival products For biologic, the license provision requires

not just the product, but also the

manufacturing process/facility be approved

A huge cost barrier exists to implement

process changes after license is granted

ceng 109 class 5

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