Advertisement

Ligand Engineering via Yeast Surface Display and Adherent Cell Panning

  • Lawrence A. Stern
  • Patrick S. Lown
  • Benjamin J. HackelEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 2070)

Abstract

High-throughput ligand discovery and evolution—via genotype-phenotype linkage strategies—empower molecularly targeted therapy, diagnostics, and fundamental science. Maintaining high-quality target antigen in these selections, particularly for membrane targets, is often a technical challenge. Panning yeast-displayed ligand libraries on intact mammalian cells expressing the molecular target has emerged as an effective strategy. Herein we describe the techniques used to select target-binding ligands via this approach including the use of target-negative cells to deplete non-specific binders and avidity reduction to preferentially select high-affinity ligands.

Key words

Avidity Cell panning Depletion Ligand Protein engineering Specificity Yeast surface display 

Notes

Acknowledgments

This chapter describes work funded by the American Cancer Society (130418-RSG-17-110-01-TBG to B.J.H.), the National Institutes of Health (R01 EB023339 to B.J.H.), and the California Tobacco-Related Disease Research Grants Program Office of the University of California (28FT-0072 to L.A.S.).

References

  1. 1.
    James ML, Gambhir SS (2012) A molecular imaging primer: modalities, imaging agents, and applications. Physiol Rev 92:897–965PubMedCrossRefGoogle Scholar
  2. 2.
    Dijkstra S, Mulders PFA, Schalken JA (2014) Clinical use of novel urine and blood based prostate cancer biomarkers: a review. Clin Biochem 47:889–896PubMedCrossRefGoogle Scholar
  3. 3.
    Husseinzadeh N (2011) Status of tumor markers in epithelial ovarian cancer has there been any progress? A review. Gynecol Oncol 120:152–157PubMedCrossRefGoogle Scholar
  4. 4.
    Yotsukura S, Mamitsuka H (2015) Evaluation of serum-based cancer biomarkers: a brief review from a clinical and computational viewpoint. Crit Rev Oncol Hematol 93:103–115PubMedCrossRefGoogle Scholar
  5. 5.
    Leader B, Baca QJ, Golan DE (2008) Protein therapeutics: a summary and pharmacological classification. Nat Rev Drug Discov 7:21–39PubMedCrossRefGoogle Scholar
  6. 6.
    Mäbert K, Cojoc M, Peitzsch C et al (2014) Cancer biomarker discovery: current status and future perspectives. Int J Radiat Biol 90:659–677PubMedCrossRefGoogle Scholar
  7. 7.
    Zahnd C, Amstutz P, Plückthun A (2007) Ribosome display: selecting and evolving proteins in vitro that specifically bind to a target. Nat Methods 4:269–279CrossRefGoogle Scholar
  8. 8.
    Mattheakis LC, Bhatt RR, Dower W (2012) An in vitro polysome display system for identifying ligands from very large peptide libraries. Proc Natl Acad Sci 91:9022:9026CrossRefGoogle Scholar
  9. 9.
    Dreier B, Pluckthun A (2012) Rapid selection of high affinity binders using ribosome display. Methods Mol Biol 805:261-286Google Scholar
  10. 10.
    Josephson K, Ricardo A, Szostak JW (2014) MRNA display: from basic principles to macrocycle drug discovery. Drug Discov Today 19:388–399PubMedCrossRefGoogle Scholar
  11. 11.
    Lipovsek D, Plückthun A (2004) In-vitro protein evolution by ribosome display and mRNA display. J Immunol Methods 290:51–67PubMedCrossRefGoogle Scholar
  12. 12.
    Roberts RW, Szostak JW (1997) RNA-peptide fusions for the in vitro selection of peptides and proteins. Proc Natl Acad Sci 94:12297–12302PubMedCrossRefGoogle Scholar
  13. 13.
    Smith GP (1985) Filamentous fusion phage: novel expression vectors that display cloned antigens on the viron surface. Science (80) 228:1315–1317CrossRefGoogle Scholar
  14. 14.
    Bratkovic T (2010) Progress in phage display: evolution of the technique and its application. Cell Mol Life Sci 67:749–767PubMedCrossRefGoogle Scholar
  15. 15.
    Sidhu SS, Lowman HB, Cunningham BC et al (2000) Phage display for selection of novel binding peptides. Methods Enzymol 328:333–363Google Scholar
  16. 16.
    Boder ET, Wittrup KD (1997) Yeast surface display for screening combinatorial polypeptide libraries. Nat Biotechnol 15:553–557PubMedPubMedCentralCrossRefGoogle Scholar
  17. 17.
    Gai SA, Wittrup KD (2007) Yeast surface display for protein engineering and characterization. Curr Opin Struct Biol 17:467–473CrossRefGoogle Scholar
  18. 18.
    Ho M, Pastan I (2009) Mammalian cell display for antibody engineering. Methods Mol Biol 525:337–351Google Scholar
  19. 19.
    Beerli RR, Bauer M, Buser RB et al (2008) Isolation of human monoclonal antibodies by mammalian cell display. Proc Natl Acad Sci U S A 105:14336–14341PubMedPubMedCentralCrossRefGoogle Scholar
  20. 20.
    Bowley DR, Labrijn AF, Zwick MB et al (2007) Antigen selection from an HIV-1 immune antibody library displayed on yeast yields many novel antibodies compared to selection from the same library displayed on phage. Protein Eng Des Sel 20:81–90CrossRefGoogle Scholar
  21. 21.
    Shusta EV, Kieke MC, Parke E et al (1999) Yeast polypeptide fusion surface display levels predict thermal stability and soluble secretion efficiency. J Mol Biol 292:949–956PubMedCrossRefGoogle Scholar
  22. 22.
    Swers JS, Kellogg BA, Wittrup KD (2004) Shuffled antibody libraries created by in vivo homologous recombination and yeast surface display. Nucleic Acids Res 32:e36CrossRefGoogle Scholar
  23. 23.
    Ackerman M, Levary D, Tobon G et al (2009) Highly avid magnetic bead capture: an efficient selection method for de novo protein engineering utilizing yeast surface display. Biotechnol Prog 25:774–783PubMedPubMedCentralCrossRefGoogle Scholar
  24. 24.
    Chen TF, de Picciotto PS, Hackel BJ et al (2013) Engineering fibronectin-based binding proteins by yeast surface display. Methods Enzymol 523:303–326Google Scholar
  25. 25.
    Gera N, Hussain M, Rao BM (2013) Protein selection using yeast surface display. Methods 60:15–26CrossRefGoogle Scholar
  26. 26.
    Friedman M, Nordberg E, Höidén-Guthenberg I et al (2007) Phage display selection of affibody molecules with specific binding to the extracellular domain of the epidermal growth factor receptor. Protein Eng Des Sel 20:189–199PubMedCrossRefGoogle Scholar
  27. 27.
    Stefan N, Martin-Killias P, Wyss-Stoeckle S et al (2011) DARPins recognizing the tumor-associated antigen EpCAM selected by phage and ribosome display and engineered for multivalency. J Mol Biol 413:826–843PubMedCrossRefGoogle Scholar
  28. 28.
    Kruziki MA, Bhatnagar S, Woldring DR et al (2015) A 45-amino-acid scaffold mined from the PDB for high-affinity ligand engineering. Chem Biol 22:946–956PubMedPubMedCentralCrossRefGoogle Scholar
  29. 29.
    Hackel BJ, Ackerman ME, Howland SW et al (2010) Stability and CDR composition biases enrich binder functionality landscapes. J Mol Biol 401:84–96PubMedPubMedCentralCrossRefGoogle Scholar
  30. 30.
    Horak E, Heitner T, Robinson MK et al (2005) Isolation of scFvs to in vitro produced extracellular domains of EGFR family members. Cancer Biother Radiopharm 20:603–613PubMedCrossRefGoogle Scholar
  31. 31.
    Zielonka S, Weber N, Becker S et al (2014) Shark attack: high affinity binding proteins derived from shark vNAR domains by stepwise in vitro affinity maturation. J Biotechnol 191:236–245CrossRefGoogle Scholar
  32. 32.
    Iwasaki K, Goto Y, Katoh T et al (2015) A fluorescent imaging probe based on a macrocyclic scaffold that binds to cellular EpCAM. J Mol Evol 81:210–217PubMedCrossRefGoogle Scholar
  33. 33.
    Mazzei GJ, Edgerton MD, Losberger C et al (1995) Recombinant soluble trimeric CD40 ligand is biologically active. J Biol Chem 270:7025–7028PubMedCrossRefGoogle Scholar
  34. 34.
    Singer E, Landgraf R, Horan T et al (2001) Identification of a heregulin binding site in HER3 extracellular domain. J Biol Chem 276:44266–44274PubMedCrossRefGoogle Scholar
  35. 35.
    Stern LA, Lown PS, Kobe AC et al (2019) Cellular-based selections aid yeast-display discovery of genuine cell-binding ligands: targeting oncology vascular biomarker CD276. ACS Comb Sci 21:207–222PubMedCrossRefGoogle Scholar
  36. 36.
    Zorniak M, Clark PA, Umlauf BJ, et al (2017) Yeast display biopanning identifies human antibodies targeting glioblastoma stemlike cells. Sci Rep 7:1–12Google Scholar
  37. 37.
    Tillotson BJ, Cho YK, Shusta EV. (2013) Cells and cell lysates: A direct approach for engineering antibodies against membrane proteins using yeast surface display. Methods 60:27–37PubMedCrossRefGoogle Scholar
  38. 38.
    Williams RM, Hajiran CJ, Nayeem S, Sooter LJ (2014) Identification of an antibody fragment specific for androgen-dependentprostate cancer cells. BMC Biotechnol 14:81PubMedPubMedCentralCrossRefGoogle Scholar
  39. 39.
    Dangaj D, Lanitis E, Zhao A, et al (2013) Novel recombinant human B7-H4 antibodies overcome tumoral immune escape to potentiate T-cell antitumor responses. Cancer Res 73:4820–4829PubMedPubMedCentralCrossRefGoogle Scholar
  40. 40.
    Wang XX, Cho YK, Shusta E V (2007) Mining a yeast library for brain endothelial cell-binding antibodies. Nat Methods 4:143–145PubMedPubMedCentralCrossRefGoogle Scholar
  41. 41.
    Wang XX, Shusta E V (2005) The use of scFv-displaying yeast in mammalian cell surface selections. J Immunol Methods 304:30–42PubMedCrossRefGoogle Scholar
  42. 42.
    Even-desrumeaux K, Chames P (2012) Phage Display and Selections in Cells. In: Antibody Engineering. pp 225–235CrossRefGoogle Scholar
  43. 43.
    Watters JM, Telleman P, Junghans RP (1997) An optimized method for cell-based phage display panning. Immunotechnology 3:21–29CrossRefGoogle Scholar
  44. 44.
    Jones AR, Stutz CC, Zhou Y, et al (2014) Identifying blood-brain-barrier selective single-chain antibody fragments. Biotechnol J 9:664–674Google Scholar
  45. 45.
    Newton J and Deutscher SL (2008) Phage peptide display. in Handbook of Experimental Pharmacology 145–163CrossRefGoogle Scholar
  46. 46.
    Sanchez-Martin D, Sorensen MD, Lykkemark S, et al (2015) Selection strategies for anticancer antibody discovery: searching off the beaten path. Trends Biotechnol 33:292–301PubMedPubMedCentralCrossRefGoogle Scholar
  47. 47.
    Stern LA, Schrack IA, Johnson SM et al (2016) Geometry and expression enhance enrichment of functional yeast-displayed ligands via cell panning. Biotechnol Bioeng 113:2328–2341PubMedPubMedCentralCrossRefGoogle Scholar
  48. 48.
    Watters JM, Telleman P, Junghans RP (1997) An optimized method for cell-based phage display panning. Immunotechnology 3:21–29CrossRefGoogle Scholar
  49. 49.
    Stern LA, Csizmar CM, Woldring DR et al (2017) Titratable avidity reduction enhances affinity discrimination in mammalian cellular selections of yeast-displayed ligands ACS Comb Sci. 19(5):315–323PubMedPubMedCentralCrossRefGoogle Scholar
  50. 50.
    Even-Desrumeaux K, Chames P (2012) Phage display and selections in cells. Methods Mol Biol 907:225–235PubMedCrossRefPubMedCentralGoogle Scholar
  51. 51.
    Patrick WM, Firth AE, Blackburn JM (2003) User-friendly algorithms for estimating completeness and diversity in randomized protein-encoding libraries. Protein Eng Des Sel 16:451–457CrossRefGoogle Scholar
  52. 52.
    Stevens R, Stevens L, Price N (1983) The stabilities of various thiol compounds used in protein purifications. Biochem Educ 11:70CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2020

Authors and Affiliations

  • Lawrence A. Stern
    • 1
  • Patrick S. Lown
    • 2
  • Benjamin J. Hackel
    • 2
    Email author
  1. 1.Department of Hematology and Hematopoietic Cell TransplantationBeckman Research Institute of the City of HopeDuarteUSA
  2. 2.Department of Chemical Engineering and Materials ScienceUniversity of Minnesota—Twin CitiesMinneapolisUSA

Personalised recommendations