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Updated: Sep 2, 2026
Detection of Antibodies That Neutralize the Cellular Uptake of Enzyme Replacement Therapies with a Cell-based Assay
Published on: September 10, 2018
STAT3 phosphorylation as an endpoint in a functional cell-based neutralizing antibody assay with potential for broad
Michael Luong1, Vitaly Ablamunits2, Brianna McPhee2
1Translational Clinical Sciences, Pfizer, Inc., Andover, MA, USA.
Background:
PF-07314470 is an antibody that was tested in a first-in-human study to evaluate its safety, pharmacokinetics, and pharmacodynamics. Because PF-07314470 is an interleukin-27 receptor (IL-27 R) agonist, the feasibility of a cell-based neutralizing antibody (NAb) assay was assessed as a tool to characterize anti-drug antibodies (ADA). Identifying a suitable assay endpoint is critical to NAb assay development. Phosphorylation of signal transducer and activator of transcription 3 (STAT3) was chosen, because it reflected activation of IL-27 R and could be measured by different means.
Methods:
PF-07314470 was used to activate IL-27 R on engineered Chinese Hamster ovary (CHO) cells, and resultant phospho-STAT3 (pSTAT3) levels were measured using a Meso Scale Discovery (MSD) assay. Experiments were conducted to confirm specificity of the pSTAT3 response and its inhibition by a NAb positive control. Biotin-Drug Extraction and Acid Dissociation (BEAD) was used to enhance drug tolerance.
Results:
PF-07314470 induced pSTAT3 that was reliably measured by MSD. The pSTAT3 response was specific to PF-07314470 and could only be inhibited by a positive control against PF-07314470. Assay sensitivity and drug tolerance were achieved through implementation of BEAD.
Conclusions:
Experiments demonstrated the feasibility of a cell-based NAb assay for PF-07314470. The assay system could be used for other biotherapeutics that modulate pSTAT3.
