Boston Strategics Tapboost

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)


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


mock

TAPBOOSTTM

>20x

TAPBOOSTTM

Target Specificity of

TAPBOOSTTM


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.


Target Protein; Factor VIII-Fc fusion protein

TAPBOOSTTM enhances the production of Factor VIII-Fc drastically


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


• 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)


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


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

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8

0

1

2

3

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5

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7

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


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


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


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%

Boston Strategics Tapboost

Technology

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