Therapeutic Drug Monitoring
Infliximab (IFX) is a chimeric antibody that targets the pro-inflammatory cytokine TNF-alpha. The introduction of infliximab has revolutionized the treatment of chronic inflammatory diseases like inflammatory bowel disease (IBD), rheumatoid arthritis (RA) and spondyloarthritis. It has been shown that infliximab can induce deep remission and improve the patient’s quality of life. Some patients do not respond to IFX therapy upon induction (primary non-responders), while others lose response over time (secondary non-responders).
A drug can only exert its pharmacologic effect when adequate concentrations are achieved in the circulation. The serum concentration of infliximab just before the next infusion, defined as the trough concentration, has been used for therapeutic drug monitoring (TDM). Recent data on TDM have shown that a good clinical response is associated with adequate trough concentrations in IBD and RA patients. TDM may therefore be very instrumental to optimize treatment and to overcome secondary loss of response.
The apDia Infliximab ELISA uses a highly specific monoclonal antibody – clone 6B7, developed at the KU Leuven - that only detects infliximab (Remicade®). Other anti-TNF drugs (like adalimumab, golimumab) do not interfere with the measurement6. Biosimilars of Remicade® (Remsima®, Inflectra®) are equally well quantified in the apDia Infliximab ELISA.
As an example of TDM, the use of infliximab trough concentration measurements in inflammatory bowel disease patients is described.
Inflammatory bowel disease
Infliximab is given at week 0, week 2 and week 6 (induction) and upon good clinical response at week 14, treatment is continued by infusions every 8 weeks (maintenance). The diagnostic value of therapeutic drug monitoring in IBD patients is described hereunder for both the induction as well as the maintenance phase. 
Induction phase: It has been demonstrated that postinduction IFX trough concentrations (week 14) are associated with a sustained clinical response. Infliximab trough concentration measurements during or shortly after induction may thus be used to identify undertreated patients and dose-optimize them.
Maintenance phase: It has been shown that patients on maintenance therapy having sustained trough concentrations, are more likely to remain in remission than patients with undetectable trough concentrations. Thus, regularly checking IFX trough concentrations during maintenance therapy may be useful to evaluate the IFX treatment schedule and make adjustments when necessary. On top, it has been shown that patients on maintenance therapy who lost response to infliximab have more benefit from individualized treatment based on the measured IFX serum concentrations than from an empirical strategy that uses all other available therapeutic options.
Due to the dosing regimen, trough concentrations during induction w2 and w6 are higher and serum samples need to be diluted more compared to the maintenance phase in which trough concentrations between 0.5-12 μg/ml are common.
Secondary loss of response is often due to the development of anti-drug antibodies, because of the immunogenic character of the drug. In the case of undetectable trough concentrations, subsequent measurement of anti-drug antibodies may be helpful to determine the optimal treatment strategy.
Principle of the IFX ELISA
Microtiterstrips coated with TNF-alpha are incubated with calibrators, controls and diluted patient samples. During this incubation step IFX binds specifically to the TNF-alpha on the solid phase. After removal of the unbound serum proteins by a washing procedure, the antigen-antibody complex in each well is detected with specific peroxidase-conjugated monoclonal antibody (clone 6B7, developed at the KU Leuven) directed to IFX.
After removal of the unbound conjugate, the strips are incubated with a chromogenic solution containing tetramethylbenzidin and hydrogen peroxide: a blue colour develops in proportion to the amount of immunocomplex bound to the wells of the strips. The enzymatic reaction is stopped by the addition of 0.5M H2SO4 and the absorbance values at 450 nm are determined.
A standard curve is obtained by plotting the absorbance values versus the corresponding calibrator values. The concentration of IFX in patient samples is determined by interpolation from the calibration curve.