In Vitro ELISA and Cell-Based Assays Confirm the Low Immunogenicity of VNAR Therapeutic Constructs in a Mouse Model of Human RA: An Encouraging Milestone to Further Clinical Drug Development

Figure 5

In vitro functional assessment of serum samples of mice treated with anti-hTNF-α drugs. (a–e) Determination of residual neutralising capacity of serum anti-hTNF-α drugs in a classical in vitro hTNF-α-induced cytotoxicity assay using a sensitised L929 cell line. (a and b) Shown are 1 mg/kg and 3 mg/kg Quad-X™-treated serum samples, respectively (Quad-X™-treated sera vs. D-PBS-treated sera, ). (c and d) Shown are 1 mg/kg and 3 mg/kg Humira®-treated serum samples, respectively (Humira®-treated sera vs. D-PBS-treated sera, for all 1 mg/kg treated sera except sera 5 () and for 3 mg/kg treated sera 4 and 5). (e) Shown are 30 mg/kg D1-NDure™-C4-treated serum samples ( for all D1-NDure™-C4-treated sera vs. D-PBS-treated sera). Exogenous recombinant hTNF-α at 0.3 ng/mL and 1 μg/mL actinomycin-D were used in all treatment groups except where otherwise stated. No exogenous hTNF-α and actinomycin-D were added to untreated cells and D-PBS-treated sera (nil hTNF) control groups. Quad-X™ Std and Humira® Std represent standard known concentrations of the drugs. The results shown are the ( with three replicates per experiment). Results were analysed statistically using a two-way ANOVA with multiple comparison post hoc test ( is considered not significant).

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