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Advancing targeted co-stimulation with antibody-fusion proteins by introducing TNF superfamily members in a

Sina Fellermeier1, Nadine Beha1, Jan-Erik Meyer1

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

Oncoimmunology
|December 22, 2016
PubMed

Insights

Engineered single-chain co-stimulatory ligands (scTNFSF) fused to antibodies offer enhanced stability and bioavailability for targeted cancer immunotherapy. This novel format improves co-stimulatory activity, showing promise for future therapeutic fusion protein development.

Area of Science:

  • Immunology
  • Protein Engineering
  • Biotechnology

Background:

  • Co-stimulation via tumor necrosis factor superfamily (TNFSF) receptors is a key strategy for enhancing antitumor immune responses.
  • Antibody-fusion proteins targeting co-stimulatory ligands to tumor sites offer a promising approach for focused immunotherapy.
  • Engineering TNFSF ligands into stable, functional fusion proteins presents significant protein engineering challenges due to their natural trimeric structure.

Purpose of the Study:

  • To develop a novel, stable format for antibody-fusion proteins incorporating co-stimulatory TNFSF ligands.
  • To evaluate the co-stimulatory activity, stability, and bioavailability of engineered fusion proteins.
  • To assess the therapeutic potential of this new format for cancer immunotherapy.

Main Methods:

  • Generation of single-chain TNFSF ligands (scTNFSF) designed to form intramolecular trimers.
  • Fusion of tumor-directed single-chain variable fragment (scFv) antibody fragments against EpCAM or FAP to scTNFSF molecules (sc4-1BBL, scOX40L, scGITRL, scLIGHT).
  • Characterization of the resulting monomeric scFv-scTNFSF fusion proteins for co-stimulatory activity, serum stability, and in vivo bioavailability in mice.

Main Results:

  • The novel scFv-scTNFSF format demonstrated equal or enhanced co-stimulatory activity compared to traditional scFv-TNFSF formats, despite reduced antibody binding avidity.
  • Enhanced serum stability and improved bioavailability were observed for the scFv-scTNFSF fusion proteins in mouse models.
  • The scFv-scTNFSF format proved versatile, applicable to various TNFSF members, and exhibited targeting-dependent co-stimulatory activity.

Conclusions:

  • The scFv-scTNFSF format represents a simplified and stable approach to engineering co-stimulatory antibody-fusion proteins.
  • This format overcomes challenges associated with TNFSF ligand trimerization and offers improved pharmacokinetic properties.
  • The scFv-scTNFSF fusion proteins show significant potential for advancing targeted cancer immunotherapy development.
Keywords:
Antibody-fusion proteinsTNFSF ligandscancer immunotherapyco-stimulation

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