ObjectiveTo investigate whether antidrug antibodies and/or drug non‐trough levels predict the long‐term treatment response in a large cohort of patients with rheumatoid arthritis (RA) treated with adalimumab or etanercept and to identify factors influencing antidrug antibody and drug levels to optimize future treatment decisions.MethodsA total of 331 patients from an observational prospective cohort were selected (160 patients treated with adalimumab and 171 treated with etanercept). Antidrug antibody levels were measured by radioimmunoassay, and drug levels were measured by enzyme‐linked immunosorbent assay in 835 serial serum samples obtained 3, 6, and 12 months after initiation of therapy. The association between antidrug antibodies and drug non‐trough levels and the treatment response (change in the Disease Activity Score in 28 joints) was evaluated.ResultsAmong patients who completed 12 months of followup, antidrug antibodies were detected in 24.8% of those receiving adalimumab (31 of 125) and in none of those receiving etanercept. At 3 months, antidrug antibody formation and low adalimumab levels were significant predictors of no response according to the European League Against Rheumatism (EULAR) criteria at 12 months (area under the receiver operating characteristic curve 0.71 [95% confidence interval (95% CI) 0.57, 0.85]). Antidrug antibody–positive patients received lower median dosages of methotrexate compared with antidrug antibody–negative patients (15 mg/week versus 20 mg/week; P = 0.01) and had a longer disease duration (14.0 versus 7.7 years; P = 0.03). The adalimumab level was the best predictor of change in the DAS28 at 12 months, after adjustment for confounders (regression coefficient 0.060 [95% CI 0.015, 0.10], P = 0.009). Etanercept levels were associated with the EULAR response at 12 months (regression coefficient 0.088 [95% CI 0.019, 0.16], P = 0.012); however, this difference was not significant after adjustment. A body mass index of ≥30 kg/m2 and poor adherence were associated with lower drug levels.ConclusionPharmacologic testing in anti–tumor necrosis factor–treated patients is clinically useful even in the absence of trough levels. At 3 months, antidrug antibodies and low adalimumab levels are significant predictors of no response according to the EULAR criteria at 12 months.

Traditional methods of avian transgenesis involve complex manipulations involving either retroviral infection of blastoderms or the ex vivo manipulation of primordial germ cells (PGCs) followed by injection of the cells back into a recipient embryo. Unlike in mammalian systems, avian embryonic PGCs undergo a migration through the vasculature on their path to the gonad where they become the sperm or ova producing cells. In a development which simplifies the procedure of creating transgenic chickens we have shown that PGCs are directly transfectable in vivo using commonly available transfection reagents. We used Lipofectamine 2000 complexed with Tol2 transposon and transposase plasmids to stably transform PGCs in vivo generating transgenic offspring that express a reporter gene carried in the transposon. The process has been shown to be highly effective and as robust as the other methods used to create germ-line transgenic chickens while substantially reducing time, infrastructure and reagents required. The method described here defines a simple direct approach for transgenic chicken production, allowing researchers without extensive PGC culturing facilities or skills with retroviruses to produce transgenic chickens for wide-ranging applications in research, biotechnology and agriculture.

Newly hatched chickens are highly susceptible to infection by opportunistic pathogens during the first 1 or 2 weeks of life. The use of cytokines as therapeutic agents has been studied in animal models as well as in immunosuppressed patients. This approach has become more feasible in livestock animals, in particular poultry, with the recent cloning of cytokine genes and the development of new technologies, such as live delivery vectors. We have recently cloned the gene for chicken interferon-gamma (Ch-IFN-gamma). Poly-HIS-tagged recombinant Ch-IFN-gamma was expressed in Escherichia coli, was purified by Ni chromatography, and was found to be stable at 4 degrees C and an ambient temperature for at least several months and Several weeks, respectively. Ch-IFN-gamma was capable of protecting chick fibroblasts from undergoing virus-mediated lysis, induced nitrite secretion from chicken macrophages in vitro, and enhanced MHC class II expression on macrophages. Administration of recombinant Ch-IFN-gamma to chickens resulted in enhanced weight gain over a 12-day period. Furthermore, the therapeutic potential of Ch-IFN-gamma was assessed using a coccidial challenge model. Birds were treated with Ch-IFN-gamma or a diluent control and then infected with Eimeria acervulina. Infected birds treated with Ch-IFN-gamma showed improved weight gain relative to noninfected birds. The ability of Ch-IFN-gamma to enhance weight gain in the face of coccidial infection makes it an excellent candidate as a therapeutic agent.

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