Licensed Biological Products with Structural Heterogeneity

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Licensed Biological Products with Structural
Heterogeneity

• Andrew C. Chang, Ph.D.
• Associate Director for Policy and Regulation
• Division of Hematology, CBER, FDA
• BPAC, Gaithersburg, MD
• November 4, 2005

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TOPICS TO BE COVERED

• Structural Heterogeneity of Biological Products
  Case Studies
• Plasma derived products
• - Andrew Chang, Ph.D. OBRR/CBER
• Recombinant products and monoclonal antibodies
• - Kurt Brorson, Ph.D. OPS/CDER

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Structural Heterogeneity of Biological Products

• Factors contributing to structural heterogeneity
• Biosynthetic processes used by living organisms
• Manufacture and/or storage of the drug substance
  and drug product
• Control of structural heterogeneity
• To demonstrate consistency of the heterogeneity
  pattern of commercial lots with that of the lots
  used in preclinical and clinical studies
• To assure lot-to-lot consistency, the kind and
  extent of this heterogeneity should be
  characterized and controlled

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Structural Heterogeneity of Biological Products
(Cont)
Examples of potential structural modifications

• Acylation
• Amidation or deamidation
• Carbamylation
• Carboxylation
• Formylation
• Formation of gamma carboxyglutamic acid
• Methylation
• Succinimide forms
• Aspartate isomerization
• Disulfide linkage

• Oxidation
• Phosphorylation
• Sulphation
• Proteolysis (terminal or domain deletion)
• Glycosylation (N-linked, O-linked, site
  occupancy, terminal groups, fucosylation)
• Aggregation
• High order structural change (conformational
  change or denaturization)

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Case Study (I)

• Charge analysis of N-linked Oligosaccharides from
  FVIII Standards, Recombinant and Plasma Derived
  FVIII Products

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Hemostasis
XII
Intrinsic Pathway
Contact
Kallikrein
XIIa HMWK
Extrinsic Pathway
PK
XIa Ca
Vascular Injury
Thrombin
X
IXa or Xa
VIIa
IXa
VII Tissue Factor
Ca
TF
VIIIa
Ca
Ca, PL
Prothrombin
Xa
Va
V
Ca, PL
Ca, PL
Thrombin
Fibrinogen
Fibrin
XL-Fibrin
XIIIa
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• FVIII
• Multi-domain glycoprotein (Mr 264,763)
• Potential 25 N-linked Oligosaccharides

A1
A2
B
A3
C1
C2
N
C
1648/1649
IC Protease
210 kDa
Heavy Chain
Light Chain
FVIII
90-210 kDa
740/741
1689/1690
Thrombin
vWF
372/373
Activated FVIII
50 kDa
43 kDa
72 kDa
vWF
372/373
1689/1690
Light Chain
Heavy Chain
B-Domain Deleted FVIII Toole et al., PNAS 1986
83 5939-5942
A1
A2
A3
C1
C2
N
C
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Charge Analysis of N-linked Oligosaccharides from
FVIII Standards, Recombinant and Plasma Derived
FVIII ProductsSchilow et al., Thromb Haemost
2004 92427-428
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Conclusion

• The charge analysis demonstrated heterogeneity of
  the glycoforms of the tested FVIII products.

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Case Study (II)

• Characterization of five von Willebrand Factor
  (VWF) concentrates produced by five different
  manufacturing processes

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von Willebrand Factor
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Variation of vWF Multimers Among Five Different
VWF Concentrates
1 Gel
2 Gel
IS P C-1 C-2 C-3 C-4 C-5
IS P C-1 C-2 C-3 C-4 C-5
Chang et al., Blood 2000 96567a, 2434
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VWF Activities from 5 Different VWF Concentrates
Chang et al., Blood 2000 96567a, 2434
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Conclusion

• The characteristics (e.g., Multimeric pattern and
  specific activity) of von Willebrand factor
  concentrates depend on the manufacturing
  processes.