Process design for the production of Fab fragment for therapy Group 6 Shiang-Yuan Hsieh, Yun-Hsuan Liu, Chao-Ming Yen.

Process design for the production of Fab fragment for therapy

Group 6Shiang-Yuan Hsieh, Yun-Hsuan Liu, Chao-Ming Yen

Antibody therapy

Bevacizumab v.s Ranibizumab

• Anti-angiogenic drugs: block the formation of new blood vessels to inhibit tumor growth

• By targeting and inhibiting the function of a natural protein called vascular endothelial growth factor (VEGF)

• The whole antibody versus a antibody fragment

Antibody• Also called immunoglobulins (Ig) that used by

immune system.• Five classes of Ig: IgA, IgE, IgG, IgD, IgM.

Fragment antigen binding(Fab fragment)

• A region on an antibody which binds to antigens

• Composed of one constant and one variable domain of each of the heavy and the light chain

Why make Fab Fragments?

• Smaller size that can promote the efficiency of penetration of tissue section.

• Reduce nonspecific binding that results from Fc interactions.

• Potentially higher sensitivity in antigen.• Reduce the risks from immune response.• Production is more economical.

Production large scale up- Use Hollow-Fiber Bioreactor

Production large scale up- Use Hollow-Fiber Bioreactor

Results of production reactor• 100 g dry weight of cells/L • Periplasm production of Fab fragments

– (50g /l protein, 3 g Fab/l)

This is the starting material of your separationYour product is in periplasmSo first step should be 1. Separation of cells by centrifugation/microfiltation2. loosely breakup cells

potentially use osmotic shock (high concentration of guanidine HCL or urea)

3. Ion exchange for enriching Fab fragment

Antibody Purification - Affinity Chromatography

1. Use the original VEGF as the antigens2. Other antigens (Gammabind G Sepharose) Barbas et al. (2001).

Antibody Purification - Use Device: Gradiflow

Products Recoveries: Affinity Chromatography vs Gradiflow

Products Duration