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Therapeutic TNF Inhibitors can Differentially Stabilize Trimeric TNF by Inhibiting Monomer Exchange

View Article: PubMed Central - PubMed

ABSTRACT

Tumor necrosis factor (TNF) is a homotrimeric cytokine that is a key mediator of inflammation. It is unstable at physiological concentrations and slowly converts into an inactive form. Here, we investigated the mechanism of this process by using a Förster resonance energy transfer (FRET) assay that allowed monitoring of monomeric subunit exchange in time. We observed continuous exchange of monomeric subunits even at concentrations of TNF high enough to maintain its bioactivity. The kinetics of this process closely corresponds with the appearance of monomeric subunits and disappearance of trimeric TNF in time at ng/ml concentrations as monitored by high-performance size-exclusion chromatography (HP-SEC). Furthermore, of the five therapeutic TNF inhibitors that are currently used in the clinic, three (adalimumab, infliximab, etanercept) were found to completely inhibit the monomer exchange reaction and stabilize TNF trimers, whereas golimumab and certolizumab could not prevent monomer exchange, but did slow down the exchange process. These differences were not correlated with the affinities of the TNF inhibitors, measured with both surface plasmon resonance (SPR) and in fluid phase using fluorescence-assisted HP-SEC. The stabilizing effect of these TNF inhibitors might result in prolonged residual TNF bioactivity under conditions of incomplete blocking, as observed in vitro for adalimumab.

No MeSH data available.


Structure of the different TNF inhibitors.Adalimumab (ADL) is a fully human IgG1 kappa antibody, as is golimumab (GOL). Certolizumab pegol (CZP) is a pegylated humanized Fab’ fragment, infliximab (IFX) a chimeric antibody, and etanercept (ETN) a receptor-Fc construct. Human origin is shown in grey, murine origin in black.
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f1: Structure of the different TNF inhibitors.Adalimumab (ADL) is a fully human IgG1 kappa antibody, as is golimumab (GOL). Certolizumab pegol (CZP) is a pegylated humanized Fab’ fragment, infliximab (IFX) a chimeric antibody, and etanercept (ETN) a receptor-Fc construct. Human origin is shown in grey, murine origin in black.

Mentions: TNF is also an important mediator of a number of inflammatory auto-immune disorders including rheumatoid arthritis, Crohn’s disease, and psoriasis1. In many patients, the inhibition of TNF activity via a blocking antibody or antibody-receptor fusion protein has proven to successfully suppress disease activity, and may even lead to clinical remission91011. There are currently five TNF inhibitors approved for use in the clinic: three full-length antibodies (infliximab, adalimumab, and golimumab), a PEGylated Fab fragment (certolizumab pegol), and a receptor-Fc construct (etanercept), see Fig. 11213. All TNF inhibitors block the binding of TNF to its receptors, which explains the mechanism of action. However, little is known about the fate of TNF-anti-TNF complexes, their stability, size, rate of clearance, and uptake by antigen-presenting cells, which might contribute to the immunogenic potential of the TNF inhibitors.


Therapeutic TNF Inhibitors can Differentially Stabilize Trimeric TNF by Inhibiting Monomer Exchange
Structure of the different TNF inhibitors.Adalimumab (ADL) is a fully human IgG1 kappa antibody, as is golimumab (GOL). Certolizumab pegol (CZP) is a pegylated humanized Fab’ fragment, infliximab (IFX) a chimeric antibody, and etanercept (ETN) a receptor-Fc construct. Human origin is shown in grey, murine origin in black.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5015024&req=5

f1: Structure of the different TNF inhibitors.Adalimumab (ADL) is a fully human IgG1 kappa antibody, as is golimumab (GOL). Certolizumab pegol (CZP) is a pegylated humanized Fab’ fragment, infliximab (IFX) a chimeric antibody, and etanercept (ETN) a receptor-Fc construct. Human origin is shown in grey, murine origin in black.
Mentions: TNF is also an important mediator of a number of inflammatory auto-immune disorders including rheumatoid arthritis, Crohn’s disease, and psoriasis1. In many patients, the inhibition of TNF activity via a blocking antibody or antibody-receptor fusion protein has proven to successfully suppress disease activity, and may even lead to clinical remission91011. There are currently five TNF inhibitors approved for use in the clinic: three full-length antibodies (infliximab, adalimumab, and golimumab), a PEGylated Fab fragment (certolizumab pegol), and a receptor-Fc construct (etanercept), see Fig. 11213. All TNF inhibitors block the binding of TNF to its receptors, which explains the mechanism of action. However, little is known about the fate of TNF-anti-TNF complexes, their stability, size, rate of clearance, and uptake by antigen-presenting cells, which might contribute to the immunogenic potential of the TNF inhibitors.

View Article: PubMed Central - PubMed

ABSTRACT

Tumor necrosis factor (TNF) is a homotrimeric cytokine that is a key mediator of inflammation. It is unstable at physiological concentrations and slowly converts into an inactive form. Here, we investigated the mechanism of this process by using a Förster resonance energy transfer (FRET) assay that allowed monitoring of monomeric subunit exchange in time. We observed continuous exchange of monomeric subunits even at concentrations of TNF high enough to maintain its bioactivity. The kinetics of this process closely corresponds with the appearance of monomeric subunits and disappearance of trimeric TNF in time at ng/ml concentrations as monitored by high-performance size-exclusion chromatography (HP-SEC). Furthermore, of the five therapeutic TNF inhibitors that are currently used in the clinic, three (adalimumab, infliximab, etanercept) were found to completely inhibit the monomer exchange reaction and stabilize TNF trimers, whereas golimumab and certolizumab could not prevent monomer exchange, but did slow down the exchange process. These differences were not correlated with the affinities of the TNF inhibitors, measured with both surface plasmon resonance (SPR) and in fluid phase using fluorescence-assisted HP-SEC. The stabilizing effect of these TNF inhibitors might result in prolonged residual TNF bioactivity under conditions of incomplete blocking, as observed in vitro for adalimumab.

No MeSH data available.