arrow-pointing-right-300x251Golimumab

 

The apDia Golimumab ELISA is an enzyme linked immunosorbent assay intended for the quantitative determination of Golimumab ( Simponi®, anti- TNFα) in human serum and plasma.

 

BACKGROUND AND DIAGNOSTIC VALUE

 

Golimumab (GLM) is a human monoclonal antibody, derived from TNF-alpha immunized transgenic mice engineered to express human IgGs. Golimumab binds to both the soluble and transmembrane bioactive forms of human TNF-alpha, giving rise to stable high-affinity complexes thereby preventing the binding of TNF-alpha to its receptors. Golimumab has been approved for the treatment of various chronic immune-mediated inflammatory disorders in which TNF-alpha plays an important role, including rheumatoid arthritis (RA), psoriatic arthritis, ankylosing spondylitis and ulcerative colitis (UC).

 

A drug can only exert its pharmacologic effect when adequate concentrations are

achieved in the circulation. The serum concentration of golimumab just before the

next infusion, defined as the trough concentration, has been used for therapeutic drug

monitoring (TDM). Recent data on TDM have shown a positive relation between GLM trough serum concentrations and clinical outcomes in patients with RA and UC. TDM may therefore be very instrumental to optimize treatment.

 

 

The apDia Golimumab ELISA uses a highly specific monoclonal antibody – clone 171D8, developed at the KU Leuven – that only detects golimumab (Simponi®). Other anti-TNF drugs (like infliximab, adalimumab) do not interfere with the measurement. As an example of TDM, the use of GLM trough concentration measurements in UC is described.

 

Ulcerative colitis

In most European countries, patients receive the same induction treatment in daily clinical practice (200 mg at week 0 and 100 mg at week 2) followed by a body weight based dose stratification during maintenance, i.e. 50 mg every four weeks for patient with a body weight of less than 80 kg and 100 mg golimumab every four weeks for patients with a body weight of at least 80 kg. Results from PURSUIT and real-world observational studies reported clinical response rates of around 50% after golimumab induction therapy. An exposure-response relationship was observed, as patients with higher drug exposure were more likely to achieve improved outcomes. GLM trough concentration measurements during or shortly after induction may thus be used to identify undertreated patients. In PURSUIT-M, week 6 non-responders had lower serum drug levels compared to responders at week 6. These early non-responders received 100 mg golimumab maintenance, and their drug exposure was increased to levels comparable with that of responders by week 14. Since July 2018, the posology has therefore changed and patients with an inadequate response to induction can be dose increased to 100 mg at week 6 and every four weeks thereafter. Regularly checking GLM trough concentrations during induction or maintenance therapy may thus be useful to evaluate the GLM treatment schedule.

 

Immunogenicity

Currently, it is unclear if loss of response to GLM is due to formation of anti-drug antibodies since studies have reported a low rate of immunogenicity. However, 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 GOLIMUMAB ELISA

 

 

Microtiterstrips coated with TNF-alpha are incubated with calibrators, controls and diluted patient samples. During this incubation step golimumab 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 171D8, developed at the KU Leuven) directed to golimumab.
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 golimumab in patient samples is determined by interpolation from the calibration curve.

 

Coming soon!

arrow-pointing-right-300x251Vedolizumab

 

Vedolizumab (VDZ) is a humanised monoclonal antibody that binds exclusively to the lymphocyte integrin α4β7. VDZ inhibits the interaction of α4β7-expressing cells with mucosal addressin cell adhesion molecule-1 on endothelial cells, thereby hampering the infiltration of the α4β7-expressing cells into the gastrointestinal mucosa and gut-associated lymphoid tissue. VDZ suppresses gut inflammation and has therefore been approved for the treatment of patients with moderate to severely ulcerative colitis (UC)and Crohn’s disease (CD). It has been shown that VDZ can induce clinical remission and improve the patient’s quality of life.

Diagnostic Value

A drug can only exert its pharmacologic effect when adequate concentrations are achieved in the circulation. The serum concentration of biologicals just before their next infusion, defined as trough concentration, has been used for therapeutic drug monitoring (TDM). Recent data on TDM have shown a positive relationship between VDZ trough serum concentrations and clinical outcomes in patients with UC and CD. TDM may therefore be very instrumental to optimize treatment.

The apDia Vedolizumab ELISA uses highly specific monoclonal antibodies developed at the KU Leuven. Anti-TNF drugs (like infliximab, adalimumab, golimumab) do not interfere with the measurement.

As an example of TDM, the use of VDZ trough concentration measurements in UC and CD is described.

Ulcerative colitis

VDZ 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 exposure-efficacy relationships of VDZ evaluated in GEMINI 1 revealed a positive exposure-response relationship for clinical remission, clinical response, and mucosal healing for VDZ induction therapy in UC. VDZ trough concentration measurements during or shortly after induction may thus be used to identify undertreated patients.

It has been demonstrated that higher VDZ concentrations are associated with deep remission in patients with UC on maintenance therapy. Thus, regularly checking VDZ trough concentrations during maintenance therapy may be useful to evaluate the VDZ treatment schedule.

Crohn’s Disease

VDZ 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 exposure-efficacy relationships of VDZ evaluated in GEMINI 2 and 3 revealed a modest positive exposure-response relationship. Clinical remission rates were higher at week 10 than at week 6 in both studies. The European Medicines Agency allows an additional dose at week 10 before assessment of an induction response at week 14.

Due to the dosing regimen, trough concentrations during induction at week 2, week 6, week 10 (CD) & 14 (CD) are higher compared to trough concentrations during maintenance when VDZ is given every 8 weeks.

Immunogenicity

Secondary loss of response is often due to the development of anti-drug antibodies. The immunogenicity rate during treatment with VDZ is very low (4%).

Principle of the Vedolizumab ELISA

The apDia Vedolizumab ELISA uses highly specific monoclonal antibodies – clones 6F3 and 6E6, developed at the KU Leuven.

Microtiterstrips coated with anti-vedolizumab monoclonal antibody clone 6F3 are incubated with calibrators, controls and diluted patient samples. During this incubation step VDZ is captured specifically by the antibodies 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 6E6) directed to VDZ.
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 VDZ in patient samples is determined by interpolation from the calibration curve.

arrow-pointing-right-300x251Anti-Adalimumab

Adalimumab
Adalimumab (ADM) is a fully human antibody that targets the pro-inflammatory cytokine TNF-alpha. The introduction of therapeutic antibodies has revolutionized the treatment of chronic inflammatory diseases like inflammatory bowel disease (IBD), rheumatoid arthritis (RA), spondyloarthritis and plaque psoriasis. It has been shown that adalimumab can induce deep remission and improve the patient’s quality of life. Some patients do not respond to adalimumab therapy upon induction (primary non-responders), while others lose response over time (secondary nonresponders).
 
Immunogenicity
Secondary loss of response is often due to the development of anti-adalimumab antibodies (ATA), because of the immunogenic character of the drug. ATA can develop in any patient undergoing adalimumab therapy and are primarily neutralizing the activity of adalimumab through immunocomplex formation.
In addition, these immunocomplexes are rapidly cleared from the system. Analytically, they are responsible for subtherapeutic adalimumab concentrations. Therefore, in the case of very low trough concentrations of adalimumab (< 1 μg/mL), subsequent measurement of ATA may be helpful to determine the optimal treatment strategy.
 
Diagnostic Value
The diagnostic value of the Anti-Adalimumab ELISA lies in its ability to stratify patients with subtherapeutic adalimumab concentrations (< 1 μg/mL) in patients who need dose intensification or a drug (class) switch. Patients with low adalimumab concentrations (< 1 μg/mL) and low ATA titers can benefit from adalimumab dose intensification, as shown in several studies.
However, the ATA titers of patients with low ATA titers undergoing a dose intensified treatment regimen must be adequately monitored. Patients that have high ATA titers are preferably switched to another drug, both within class or out of class.
Note. The Anti-Adalimumab ELISA is not capable of measuring ATA in the presence of high concentrations of adalimumab. It should only be used when < 1 μg/mL adalimumab is quantified in the sample using the apDia Adalimumab ELISA.
 
Principle of the ATA ELISA
The apDia Anti-Adalimumab ELISA uses a highly specific monoclonal antibody – clone 6A10, developed at the KU Leuven – that only bridges adalimumab.
Microtiter strips coated with adalimumab (Humira®) are incubated with calibrators, controls and diluted patient samples. During this incubation step ATA binds specifically to the adalimumab on the solid phase. After removal of the unbound serum proteins by a washing procedure, the strips are incubated with biotin conjugated adalimumab (Humira®), binding directly to the antigen-antibody complex. After removal of the unbound biotin conjugate, the strips are incubated with peroxidase conjugated streptavidin. After removal of the unbound peroxidase 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 ATA in patient samples is determined by interpolation from the calibration curve.

 

arrow-pointing-right-300x251Adalimumab

Therapeutic Drug Monitoring
Adalimumab (ADM) is a fully human antibody that targets the pro-inflammatory cytokine TNF-alpha and is used to treat chronic inflammatory diseases like inflammatory bowel disease, rheumatoid arthritis, spondyloarthritis and plaque psoriasis. It has been shown that adalimumab can induce deep remission and improve the patient’s quality of life. Some patients do not respond to ADM 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 adalimumab just before the next injection, 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 inflammatory bowel disease and rheumatoid arthritis patients. TDM may therefore be very instrumental to optimize treatment and to overcome secondary loss of response. 
The apDia ADM ELISA uses a highly specific monoclonal antibody – Clone 40D8, developed at the K.U. Leuven – that only detects adalimumab (Humira?). Other anti- TNF drugs (for example infliximab and golimumab) do no interfere with the measurement.
As an example of TDM, the use of adalimumab trough concentration measurements in inflammatory bowel disease patients is described.
 
Inflammatory bowel disease
Induction therapy of adalimumab consists of a subcutaneous dose of 160 mg at week 0, followed by 80 mg at week 2 and 40 mg every other week from week 4 onwards. Upon good clinical response at week 12-14, treatment is continued (maintenance).
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 ADM trough concentrations during maintenance therapy may be useful to evaluate the ADM treatment schedule and make adjustments when necessary.
Patients with low or undetectable drug concentrations may benefit from a dose increase or interval shortening, while the interval in patients with very high ADM concentrations can be safely prolonged.
Due to the dosing regimen, trough concentrations during induction at w2 and w4 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.
 
Immunogenicity
Secondary loss of response is often due to the development of anti- drug antibodies, which have been observed despite of the fully human character of the drug. In case of undetectable trough concentrations, subsequent measurement of anti-drug antibodies may be helpful to determine the optimal treatment strategy.
 
Principle of the ADM ELISA
The apDia Adalimumab ELISA uses a highly specific monoclonal antibody – clone 40D8, developed at the KU Leuven - that only detects adalimumab (Humira®). Other anti-TNF drugs (like infliximab, golimumab) do not interfere with the measurement.
 
Microtiterstrips coated with TNF-alpha are incubated with calibrators, controls and diluted patient samples. During this incubation step ADM 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 40D8, developed at the KU Leuven) directed to ADM.
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 ADM in patient samples is determined by interpolation from the calibration curve.