Process design for the production of Fab fragment for therapy Group 6 Shiang-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)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 separation Your product is in periplasm So first step should be 1.Separation of cells by centrifugation/microfiltation 2.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 antigens 2.Other antigens (Gammabind G Sepharose) Barbas et al. (2001).

Antibody Purification - Use Device: Gradiflow

Products Recoveries: Affinity Chromatography vs Gradiflow

Products Duration