Discovery of a Non-competitive TNFR1 Antagonist Affibody with Picomolar Monovalent Potency That Does Not Affect TNFR2 Function

Nagamani Vunnam, Mary Jane Been, Evan Huber, Carolyn Paulson, Sophia Szymonski, Benjamin J. Hackel, Jonathan N Sachs

Research output: Contribution to journalArticlepeer-review


Tumor necrosis factor (TNF) is a key regulator of immune responses and plays a significant role in the initiation and maintenance of inflammation. Upregulation of TNF expression leads to several inflammatory diseases, such as Crohn’s, ulcerative colitis, and rheumatoid arthritis. Despite the clinical success of anti-TNF treatments, the use of these therapies is limited because they can induce adverse side effects through inhibition of TNF biological activity, including blockade of TNF-induced immunosuppressive function of TNFR2. Using yeast display, we identified a synthetic affibody ligand (ABYTNFR1-1) with high binding affinity and specificity for TNFR1. Functional assays showed that the lead affibody potently inhibits TNF-induced NF-κB activation (IC50 of 0.23 nM) and, crucially, does not block the TNFR2 function. Additionally, ABYTNFR1-1 acts non-competitively─it does not block TNF binding or inhibit receptor-receptor interactions in pre-ligand-assembled dimers─thereby enhancing inhibitory robustness. The mechanism, monovalent potency, and affibody scaffold give this lead molecule uniquely strong potential as a therapeutic candidate for inflammatory diseases.

Original languageEnglish (US)
Pages (from-to)1884-1897
Number of pages14
JournalMolecular pharmaceutics
Issue number4
StatePublished - Apr 3 2023

Bibliographical note

Funding Information:
This work was supported by the National Institutes of Health [grants to J.N.S. (R01EB028274 and R35GM131814) and B.J.H. (R01EB023339 and R01EB028274)].

Publisher Copyright:
© 2023 American Chemical Society.


  • affibody
  • inflammatory diseases
  • NF-κB activation
  • TNF
  • TNFR1
  • yeast surface display

PubMed: MeSH publication types

  • Journal Article
  • Research Support, N.I.H., Extramural


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