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Updated: Sep 30, 2026

Determination of the Relative Potency of an Anti-TNF Monoclonal Antibody (mAb) by Neutralizing TNF Using an In Vitro Bioanalytical Method
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Determination of the Relative Potency of an Anti-TNF Monoclonal Antibody (mAb) by Neutralizing TNF Using an In Vitro Bioanalytical Method

Published on: September 16, 2017

Endogenous membrane tumor necrosis factor (TNF) is a potent amplifier of TNF receptor 1-mediated apoptosis

Monika Weingärtner1, Daniela Siegmund, Ulrich Schlecht

  • 1Institute of Cell Biology and Immunology, University of Stuttgart, Allmandring 31, 70569 Stuttgart, Germany.

The Journal of Biological Chemistry
|July 10, 2002
PubMed

Insights

Heat shock protein 90 inhibitor geldanamycin and proteasome inhibitor MG-132 blocked TNF-R1-induced apoptosis, indicating NF-kappaB dependence. TNF-R1 triggers apoptosis via membrane TNF and TNF receptor cooperation, bypassing soluble TNF requirements.

Area of Science:

  • Cell Biology
  • Immunology
  • Molecular Biology

Background:

  • Tumor Necrosis Factor Receptor 1 (TNF-R1) plays a critical role in apoptosis induction.
  • NF-kappaB activation is a key pathway in cellular responses to TNF-R1 signaling.
  • The interplay between TNF receptors and endogenous TNF in apoptosis is not fully elucidated.

Purpose of the Study:

  • To investigate the mechanisms of TNF-R1-induced apoptosis in Kym-1 cells.
  • To determine the role of NF-kappaB activation in TNF-R1-mediated cell death.
  • To explore the involvement of endogenous TNF and TNF receptor cooperation in this process.

Main Methods:

  • Treatment of Kym-1 cells with geldanamycin, MG-132, agonistic antibodies for TNF-R1 and TNF-R2, and neutralizing TNF antibodies.
  • Assessment of apoptosis induction and NF-kappaB activation.
  • Analysis of endogenous TNF expression and localization.

Main Results:

  • Geldanamycin and MG-132 inhibited TNF-R1-induced apoptosis but not TRAIL-induced apoptosis, suggesting NF-kappaB dependence.
  • TNF-R1 triggering induced endogenous TNF and apoptosis, which was blocked by anti-TNF antibodies.
  • Endogenous TNF was primarily membrane-bound, acting in its cell-associated form.
  • Geldanamycin did not inhibit apoptosis induced by combined TNF-R1 and TNF-R2 antibodies, indicating receptor cooperation.
  • TNF-R1 stimulation rapidly induced apoptosis via membrane TNF and TNF-R1/TNF-R2 cooperation, independent of prolonged soluble TNF.

Conclusions:

  • TNF-R1-induced apoptosis in Kym-1 cells is dependent on NF-kappaB activation.
  • A mechanism involving membrane-bound endogenous TNF and cooperation between TNF-R1 and TNF-R2 mediates rapid apoptosis.
  • This pathway circumvents the need for sustained exogenous soluble TNF for TNF-R1-mediated apoptosis.

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