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Enhanced Protein Production
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Enhanced Protein ProductionTAPTAPBOOSTBOOST
TM
Degradation
RNA
Gene
Misfolding
folding
polypeptide
Functional
Aggregation
Abnormal Function
Unfolding Protein Response
Secretion
· Translation
· Expression of Chaperones
· Apoptosis
Recombinant ProteinsProtein SecretionStabilityFunctional Recombinant ProteinsImmunogenicity
Alzheimer's diseaseParkinson's diseaseHuntington's disease etc.
Impact of Protein Folding
Conformational Diseases
Protein Folding regulates many events in protein production
TAPBOOSTTM (Targeted Protein Boosting Technology)
Proprietary sequence that facilitates protein production
Binding domain specific for targeted protein
BSC2
TAPBOOSTERTM
BSC1
TAPBOOSTTM Technology
TAPBOOSTERTM
ER
Targeted Protein
Golgi
Insolubility of expressed proteins is, however, one of the biggest problems of Bacterial expression platformsTAPBOOOSTTM Technology significantly enhances solubility of expressed proteins.
Bacterial Expression Platform
Difficult-to-Express Protein Platform
TAPBOOOSTTM Technology enhances active antibody production in mammalian expression platform.
Market for therapeutic recombinant protein is huge (>$100 billions) with tremendous growth potential. However, difficulties in the production of therapeutic recombinant proteins make protein-based therapies the most expensive treatment in the world.
Antibody Expression Platform
TAPBOOSTTM
Technology
BacterialExpression
AntibodyExpression
Tough ProteinExpression
TAPBOOSTTM Technology (Targeted Protein Boosting Technology)
TAPBOOSTTM Technology
Model Protein; IL-13 receptor 2 (IL13R2)-Fc fusion protein
The expression levels of IL13R2 are poor, and the protein is prone to misfolding and aggregation.
Cell
IL13R2-Fc
ERNu
• The receptor of IL-13, a key mediator of allergic inflammation
Aggregation of IL13R2-Fc
Only properly folded IL13R2-Fc secrets
• Fc fusion protein with sIL13R2 (sIL13R2-Fc) has been developed for the treatment of asthma
• Production of sIL13R2-Fc using human cells is inefficient due to its instability
• Soluble IL13R2 (sIL13R2) blocks the signal from IL-13 as a decoy receptor
Difficult-to-Express Protein Expression Platform
SecretedIL13R2-Fc
mock
IL13R2-FcInside Cells
mock
TAPBOOSTTM enhances the production of IL13R2-Fc drastically
Cell
IL13R2-Fc
ERNu
Aggregation of IL13R2-Fc
Cell
TAPBOOSTTM TechnologyERNu
Secreted IL13R2-FcIL13R2-Fc TAPBOOSTERTM
Difficult-to-Express Protein Expression Platform
mock
TAPBOOSTTM
>20x
TAPBOOSTTM
Target Specificity of
TAPBOOSTTM
TAPBOOSTTM Technology
TAPBOOSTERTM
ER
Targeted Protein
Target Protein(A or B)
A A A B B B B BTarget Protein
Model Protein; Factor VIII-Fc fusion protein• Factor VIII is the responsible protein for hemophilia A, disorder that causes serious bleeds and can lead to degenerative joint disease.
• The primary treatment for hemophilia A is replacement therapy utilizing infusions of clotting factors, mainly Factor VIII.
•Manufacture of Factor VIII is extremely difficult due to protein folding problem. Therefore, yields are low and often inconsistent. • The average annual cost for hemophilia treatment in 2008 was estimated at $155,000, with clotting factor concentrates accounting for 70-82% of total costs.
Difficult-to-Express Protein Expression Platform
Target Protein; Factor VIII-Fc fusion protein
TAPBOOSTTM enhances the production of Factor VIII-Fc drastically
Difficult-to-Express Protein Expression Platform
Fo
ld in
duc
tion
FVIII Production
0
2
4
6
8
10
12
14
16
18
20
FVIII-Fc
TAPBOOSTTMF
old
ind
uctio
n
FVIII Activity
0
2
4
6
8
10
12
14
16
18
20
22
24
FVIII-Fc
TAPBOOSTTM
Antibody Expression Platform
• Monoclonal antibodies are composed of heavy and light polypeptide chains bridged through disulfide bonds. Due to nature of this structural complexity, monoclonal antibodies are expressed in mammalian expression systems that could benefit from an increase in the yield of fully active antibodies
• The combined sales of 25 actively marketed mAb therapeutics in 2010 was $43 billion.
Brand Sales ($m)
Market (%)
Avastin 6973 16.02Rituxan 6859 15.76Humira 6548 15.04Remicade 6520 14.98Herceptin 5859 13.46Lucentis 3106 7.14Erbitux 1791 4.12Tysabri 1230 2.83Synagis 1038 2.38Xolair 750 1.72
Sales of 10 actively marketed monoclonal antibody therapeutics in 2010
International Journal of Pharmaceutics (2013)
Antibody Expression Platform
anti-IL8 Ab(DP12)
TAPBOOSTTM enhances the production of all three tested antibodies
1 2 3
anti-VEGF Ab(Avastin)
No antibody expressed
antibody without TAPBOOSTTM
1 2 3
anti-TNF Ab(Humira)
1 2 3
antibody with TAPBOOSTTM
Antibody Expression Platform
anti-TNF Ab(Humira)
anti-VEGF Ab(Avastin)
anti-IL8 Ab(DP12)
0
0.5
1
1.5
2
2.5
3
3.5
4
4.5
5
0
1
2
3
4
5
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8
0
1
2
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8
Antibodies produced by TAPBOOSTTM hold the activity
Fol
d in
duct
ion
Avastin
TAPBOOSTTM
Fol
d in
duct
ion
Fol
d in
duct
ion
Humira
TAPBOOSTTM
DP12
TAPBOOSTTM
Bacterial Expression Platform
Pharma Manufacturing (2012)
Bacterial Expression Platforms
Expression Systems used for Manufacture of Approved Drugs
Major problem in Bacterial Expression System
Soluble protein
bacteria
Bacterial Genome
plasmid
<Example of Insoluble Protein>
SolubleFraction
InsolubleFraction
Expressedprotein
Mammalian
E.coli
S. cerevisiae (yeast)
Direct synthesis in
transgenic animals
Other
inclusion body
Bacterial Expression Platform
Soluble protein (SP)
Inclusion Body (IB)
bacteria
Bacterial Genome
plasmid
We customized TAPBOOSTTM technology for bacterial expression system by using a proprietary algorithms and intensive search for specific amino acid sequences involved in protein folding.
SP IB
TAPBOOSTTM Technology
SP IB
Test Protein A
Test Protein B
Test Protein C
Summary
• Any kind of protein can be expressed by TAPBOOSTTM technology
• Easy implementation into your current bioprocess
• Combine TAPBOOSTTM with any other technology for synergistic effect
Advantages of TAPBOOSTTM
Increased Productivity by TAPBOOSTTM (%)Target proteins
TNFRI-Fc (Enbrel) 600%
Factor IX-Fc 320%
Factor FVII-Fc 1000%
IL13R2-Fc protein 2000%
Factor VIII-Fc 2100%
Humira (anti-TNFα antibody)
Avastin (anti-VEGFa antibody)
DP12 (anti-IL8 antibody)
600%
420%
730%