The AAPS Journal

, Volume 13, Issue 1, pp 99–110 | Cite as

Bioanalytical Approaches to Quantify “Total” and “Free” Therapeutic Antibodies and Their Targets: Technical Challenges and PK/PD Applications Over the Course of Drug Development

  • Jean W. Lee
  • Marian Kelley
  • Lindsay E. King
  • Jihong Yang
  • Hossein Salimi-Moosavi
  • Meina T. Tang
  • Jian-Feng Lu
  • John Kamerud
  • Ago Ahene
  • Heather Myler
  • Cindy Rogers
White Paper

Abstract

The predominant driver of bioanalysis in supporting drug development is the intended use of the data. Ligand-binding assays (LBA) are widely used for the analysis of protein biotherapeutics and target ligands (L) to support pharmacokinetics/pharmacodynamics (PK/PD) and safety assessments. For monoclonal antibody drugs (mAb), in particular, which non-covalently bind to L, multiple forms of mAb and L can exist in vivo, including free mAb, free L, and mono- and/or bivalent complexes of mAb and L. Given the complexity of the dynamic binding equilibrium occurring in the body after dosing and multiple sources of perturbation of the equilibrium during bioanalysis, it is clear that ex vivo quantification of the forms of interest (free, bound, or total mAb and L) may differ from the actual ones in vivo. LBA reagents and assay formats can be designed in principle to measure the total or free forms of mAb and L. However, confirmation of the forms being measured under the specified conditions can be technically challenging. The assay forms and issues must be clearly communicated and understood appropriately by all stakeholders as the program proceeds through the development process. This paper focuses on monoclonal antibody biotherapeutics and their circulatory L that are either secreted as soluble forms or shed from membrane receptors. It presents an investigation into the theoretical and practical considerations for total/free analyte assessment to increase awareness in the scientific community and offer bioanalytical approaches to provide appropriate PK/PD information required at specific phases of drug development.

Key words

ligand-binding assays monoclonal antibody biotherapeutics PK/PD target biomarkers total and free assays 

Abbreviations

ADA

Anti-drug antibodies

Anti-id

Anti-idiotypic

CDR

Complementarity-determining regions

ELISA

Enzyme-linked immunosorbent assay

FIH

First in human

Fc

Fragment cystallizable region

IC50

The concentration of the interfering compound at which the analyte recovery is 50% of that in the absence of the interfering compound (50% inhibition point)

IgG

Immunoglobulin G

Kd

Dissociation equilibrium constant

LBA

Ligand-binding assay

L

Target ligand

Lfree

Target ligand not bound to drug

Ltotal

Sum of bound and unbound target ligand

mAb

Monoclonal antibody

mAbfree

The unbound and partially free forms of the monoclonal antibody drug that are able to bind to the target ligand to mediate biological actions

mAbtotal

Sum of bound partially bound and unbound forms of the monoclonal antibody drug

mAb*L

Complex of monoclonal antibody drug and target ligand

pAb

Polyclonal antibodies

PD

Pharmacodynamics

PK

Pharmacokinetics

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Copyright information

© American Association of Pharmaceutical Scientists 2011

Authors and Affiliations

  • Jean W. Lee
    • 1
  • Marian Kelley
    • 2
  • Lindsay E. King
    • 3
  • Jihong Yang
    • 4
  • Hossein Salimi-Moosavi
    • 1
  • Meina T. Tang
    • 4
  • Jian-Feng Lu
    • 1
  • John Kamerud
    • 5
  • Ago Ahene
    • 6
  • Heather Myler
    • 7
  • Cindy Rogers
    • 8
  1. 1.Amgen IncThousand OaksUSA
  2. 2.MKelley Consulting LLCWest ChesterUSA
  3. 3.Pfizer IncGrotonUSA
  4. 4.GenentechSouth San FranciscoUSA
  5. 5.CovancePrincetonUSA
  6. 6.Pharmargo LLCUnion CityUSA
  7. 7.Bristol-Myers SquibbPrincetonUSA
  8. 8.GenzymeFraminghamUSA

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