Safety, PK, and immunogenicity of i.d. and s.c. adalimumab
administration
Adverse events were summarized by treatment group, in subsets of all
treatment-emergent AEs, and separately for treatment-related AEs.
Clinical laboratory and vital sign measurements were summarized by
treatment and change from baseline was recorded. Summary statistics
included number of subjects, mean, median, minimum, and maximum values
(with standard deviation [SD]). Immunogenicity, i.e.,
anti-adalimumab antibodies, was reported descriptively.
For PK analyses, serum adalimumab concentrations were assessed in blood
collected in 4 mL plain tubes (BD) after coagulation (30-60 minutes) and
centrifugation (2000G for 10 minutes at 4°C), from day 1 (pre-dose) till
day 71 post-dose. Adalimumab levels were quantified by fully automated
ELISA as described (16) . Briefly, TNF was indirectly coated on
microtitre plates. Serum was added and incubated. Immobilized adalimumab
was subsequently detected using biotinylated rabbit anti-idiotype. The
LOD for this assay is 10 ng/mL.
Anti-adalimumab antibodies were measured using a semi-quantitative
radioimmunoassay as previously described (16). Briefly, samples were
incubated with Sepharose-immobilized protein A (1.0 mg/test; Pharmacia
Uppsala, Sweden) on its surface to capture IgG. After washing,
radioactive iodine labelled F(ab’)2 fragments of adalimumab were added
to detect drug-specific antibodies. The LOD for this assay 12 AU/mL.
Ex vivo whole blood challenge was performed to assess the effect
of adalimumab on the release of cytokines by circulating immune cells
and activation of these cells. Blood (6 mL) was collected in sodium
heparin tubes (Becton Dickinson, NJ, USA) followed by stimulation with 2
ng/mL lipopolysaccharide (Sigma-Aldrich, Deisenhofen, Germany) and 25
µg/mL aluminium hydroxide (Alhydrogel 2%, Invivogen, Toulouse, France)
for 24 hours at 37°C, 5% CO2. Culture supernatants were
assayed for release of pro-inflammatory cytokines TNFα, IL-6, IL-1β,
IFNγ and IL-8 using the Mesoscale Discovery multiplex immunoassay
platform.