Immunologic Research

, Volume 62, Issue 3, pp 377–385 | Cite as

A novel anti-TNF scFv constructed with human antibody frameworks and antagonistic peptides

  • Shusheng Geng
  • Hong Chang
  • Weisong Qin
  • Ming Lv
  • Yan Li
  • Jiannan Feng
  • Beifen Shen


The introduction of TNF inhibitors has revolutionized the treatment of some chronic inflammatory diseases, e.g., rheumatoid arthritis and Crohn’s disease. However, immunogenicity is one of the important mechanisms behind treatment failure, and generally, switching to another TNF inhibitor will be the first choice for patients and doctors, which results in unmet need for novel anti-TNF agents. Small antibody molecules with less number of epitope may be valuable in less immunogenicity. In this study, with the help of computer-guided molecular design, single-chain variable fragment (scFv) TSA2 was designed using consensus frameworks of human antibody variable region as scaffold to display anti-TNF antagonistic peptides. TSA2 showed evidently improved bioactivity over TSA1 (anti-TNF scFv explored before) and almost similar activity as S-Remicade (the scFv form of Remicade, anti-TNF antibody approved by FDA), especially in inhibiting TNF-induced cytotoxicity and NF-κB activation. Human antibody consensus frameworks with less immunogenicity have been used in the designing of VH domain antibody, scFv TSA1 and TSA2. A serial of TNF-related works convinced us that the novel design strategy was feasible and could be used to design inhibitors targeting more other molecules than TNF-α. More importantly, these designed inhibitors derived from computer modeling may form a virtual antibody library whose size depends on the number of candidate antagonistic peptides. It will be molecular-targeted virtual antibody library because of the specific antagonistic peptides and the potential antibodies could be determined by virtual screening and then confirmed by biologic experiments.


Antibody consensus frameworks Antagonist peptides Computer modeling TNF-α Virtual antibody library 



Monoclonal antibody


Single-chain variable fragment




Complementarity-determining regions


CDR of heavy chain


CDR of light chain


TNF-α receptor


Recombinant human TNF-α


scFv form of Remicade





S.G.& H.C. designed and performed experiments & data analysis, wrote the paper. W.Q. designed the PCR primers, offer suggestions for the experiments. M.L. assisted with experiments. J.F. performed computer modeling. B.S., Y.L. and J.F. contributed to conception, design and final approval of the paper. This work is supported by National 863 Fund (No. 2012AA02A302), National Sciences Fund (No. 31200701) and Hebei Sciences Fund (No. C2013206353) of China.

Conflict of interest



  1. 1.
    Palladino MA, Bahjat FR, Theodorakis EA, Moldawer LL. Anti-TNF-alpha therapies: the next generation. Nat Rev Drug Discov. 2003;2:736–46.PubMedCrossRefGoogle Scholar
  2. 2.
    Atzeni F, Talotta R, Salaffi F, Cassinotti A, Varisco V, Battellino M, et al. Immunogenicity and autoimmunity during anti-TNF therapy. Autoimmun Rev. 2013;12:703–8.PubMedCrossRefGoogle Scholar
  3. 3.
    Bendtzen K. Personalized medicine: theranostics (therapeutics diagnostics) essential for rational use of tumor necrosis factor-alpha antagonists. Discov Med. 2013;15:201–11.PubMedGoogle Scholar
  4. 4.
    Vincent FB, Morand EF, Murphy K, Mackay F, Mariette X, Marcelli C. Antidrug antibodies (ADAb) to tumour necrosis factor (TNF)-specific neutralising agents in chronic inflammatory diseases: a real issue, a clinical perspective. Ann Rheum Dis. 2013;72:165–78.PubMedCrossRefGoogle Scholar
  5. 5.
    Thalayasingam N, Isaacs JD. Anti-TNF therapy. Best Pract Res Clin Rheumatol. 2011;25:549–67.PubMedCrossRefGoogle Scholar
  6. 6.
    Ben-Horin S, Kopylov U, Chowers Y. Optimizing anti-TNF treatments in inflammatory bowel disease. Autoimmun Rev. 2014;13:24–30.PubMedCrossRefGoogle Scholar
  7. 7.
    Khanna R, Feagan BG. Ustekinumab for the treatment of Crohn’s disease. Immunotherapy. 2013;5:803–15.PubMedCrossRefGoogle Scholar
  8. 8.
    Spinelli FR, Valesini G. Immunogenicity of anti-tumour necrosis factor drugs in rheumatic diseases. Clin Exp Rheumatol. 2013;31:954–63.PubMedGoogle Scholar
  9. 9.
    Alawadhi A, Alawneh K, Alzahrani ZA. The effect of neutralizing antibodies on the sustainable efficacy of biologic therapies: what’s in it for African and Middle Eastern rheumatologists. Clin Rheumatol. 2012;31:1281–7.PubMedCrossRefGoogle Scholar
  10. 10.
    van Schouwenburg PA, Rispens T, Wolbink GJ. Immunogenicity of anti-TNF biologic therapies for rheumatoid arthritis. Nat Rev Rheumatol. 2013;9:164–72.PubMedCrossRefGoogle Scholar
  11. 11.
    Lowenberg M, de Boer N, Hoentjen F. Golimumab for the treatment of ulcerative colitis. Clin Exp Gastroenterol. 2014;7:53–9.PubMedCentralPubMedCrossRefGoogle Scholar
  12. 12.
    Holliger P, Hudson PJ. Engineered antibody fragments and the rise of single domains. Nat Biotechnol. 2005;23:1126–36.PubMedCrossRefGoogle Scholar
  13. 13.
    Worn A, Pluckthun A. Stability engineering of antibody single-chain Fv fragments. J Mol Biol. 2001;305:989–1010.PubMedCrossRefGoogle Scholar
  14. 14.
    Krenova Z, Pavelka Z, Lokaj P, Skotakova J, Kocmanova I, Teyschl O, et al. Successful treatment of life-threatening Candida peritonitis in a child with abdominal non-Hodgkin lymphoma using Efungumab and amphotericin B colloid dispersion. J Pediatr Hematol Oncol. 2010;32:128–30.PubMedCrossRefGoogle Scholar
  15. 15.
    Bagai A, Armstrong PW, Stebbins A, Mahaffey KW, Hochman JS, Weaver WD, et al. Prognostic implications of left ventricular end-diastolic pressure during primary percutaneous coronary intervention for ST-segment elevation myocardial infarction: findings from the assessment of Pexelizumab in acute myocardial infarction study. Am Heart J. 2013;166:913–9.PubMedCrossRefGoogle Scholar
  16. 16.
    Agunanne E, Horvat D, Uddin MN, Puschett J. The treatment of preeclampsia in a rat model employing Digibind. Am J Perinatol. 2010;27:299–305.PubMedCrossRefGoogle Scholar
  17. 17.
    Shao EH, Sivagnanavel V, Dabbagh A, Dave R, Tempest-Roe S, Tam FW, et al. Multiphasic changes in systemic VEGF following intravitreal injections of ranibizumab in a child. Eye (Lond). 2015;. doi: 10.1038/eye.2014.343.Google Scholar
  18. 18.
    Chan FK, Chun HJ, Zheng L, Siegel RM, Bui KL, Lenardo MJ. A domain in TNF receptors that mediates ligand-independent receptor assembly and signaling. Science. 2000;288:2351–4.PubMedCrossRefGoogle Scholar
  19. 19.
    De Genst E, Areskoug D, Decanniere K, Muyldermans S, Andersson K. Kinetic and affinity predictions of a protein-protein interaction using multivariate experimental design. J Biol Chem. 2002;277:29897–907.PubMedCrossRefGoogle Scholar
  20. 20.
    Steed PM, Tansey MG, Zalevsky J, Zhukovsky EA, Desjarlais JR, Szymkowski DE, et al. Inactivation of TNF signaling by rationally designed dominant-negative TNF variants. Science. 2003;301:1895–8.PubMedCrossRefGoogle Scholar
  21. 21.
    Feng J, Li Y, Zhang W, Shen B. Rational design of potent mimic peptide derived from monoclonal antibody: antibody mimic design. Immunol Lett. 2005;98:311–6.PubMedCrossRefGoogle Scholar
  22. 22.
    Qin W, Feng J, Li Y, Lin Z, Shen B. De novo design TNF-alpha antagonistic peptide based on the complex structure of TNF-alpha with its neutralizing monoclonal antibody Z12. J Biotechnol. 2006;125:57–63.PubMedCrossRefGoogle Scholar
  23. 23.
    Qin W, Feng J, Li Y, Lin Z, Shen B. Fusion protein of CDR mimetic peptide with Fc inhibit TNF-alpha induced cytotoxicity. Mol Immunol. 2006;43:660–6.PubMedCrossRefGoogle Scholar
  24. 24.
    Qin W, Feng J, Li Y, Lin Z, Shen B. A novel domain antibody rationally designed against TNF-alpha using variable region of human heavy chain antibody as scaffolds to display antagonistic peptides. Mol Immunol. 2007;44:2355–61.PubMedCrossRefGoogle Scholar
  25. 25.
    Chang H, Qin W, Li Y, Zhang J, Lin Z, Lv M, et al. A novel human scFv fragment against TNF-alpha from de novo design method. Mol Immunol. 2007;44:3789–96.PubMedCrossRefGoogle Scholar
  26. 26.
    Knappik A, Ge L, Honegger A, Pack P, Fischer M, Wellnhofer G, et al. Fully synthetic human combinatorial antibody libraries (HuCAL) based on modular consensus frameworks and CDRs randomized with trinucleotides. J Mol Biol. 2000;296:57–86.PubMedCrossRefGoogle Scholar
  27. 27.
    Zhang W, Feng J, Shen B. Identification of binding epitope of a monoclonal antibody (Z12) against human TNF-alpha using computer modeling and deletion mutant technique. Sci China C Life Sci. 2004;47:279–86.PubMedGoogle Scholar
  28. 28.
    Upchurch KS, Kay J. Evolution of treatment for rheumatoid arthritis. Rheumatology. 2012;51(6):vi28–36.PubMedGoogle Scholar
  29. 29.
    Hoogenboom HR. Selecting and screening recombinant antibody libraries. Nat Biotechnol. 2005;23:1105–16.PubMedCrossRefGoogle Scholar
  30. 30.
    Paul S, Kolla RV, Sidney J, Weiskopf D, Fleri W, Kim Y, et al. Evaluating the immunogenicity of protein drugs by applying in vitro MHC binding data and the immune epitope database and analysis resource. Clin Dev Immunol. 2013;2013:467852.PubMedCentralPubMedCrossRefGoogle Scholar
  31. 31.
    Kim Y, Ponomarenko J, Zhu Z, Tamang D, Wang P, Greenbaum J, et al. Immune epitope database analysis resource. Nucleic Acids Res. 2012;40:W525–30.PubMedCentralPubMedCrossRefGoogle Scholar
  32. 32.
    Salimi N, Fleri W, Peters B, Sette A. The immune epitope database: a historical retrospective of the first decade. Immunology. 2012;137:117–23.PubMedCentralPubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  1. 1.Department of Molecular ImmunologyInstitute of Basic Medical SciencesBeijingPeople’s Republic of China
  2. 2.College of Basic Medical SciencesHebei University of Chinese MedicineShijiazhuangPeople’s Republic of China
  3. 3.Hebei Key Laboratory of Chinese Medicine Research on Cardio-cerebrovascular DiseaseShijiazhuangPeople’s Republic of China
  4. 4.Antibody Preparation Research Center of Hebei Province UniversityShijiazhuangPeople’s Republic of China
  5. 5.National Clinical Research Center of Kidney Diseases, Jinling HospitalNanjingPeople’s Republic of China

Personalised recommendations