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Alternative Binding Scaffolds: Multipurpose Binders for Applications in Basic Research and Therapy

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Introduction to Antibody Engineering

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What You Will Learn in This Chapter

Next to monoclonal antibodies (mAbs), alternative binding scaffolds have proven to be powerful tool reagents and therapeutic entities over the past decades. In contrast to their hetero-tetrameric, macromolecular counterparts, these affinity reagents are based on non-antibody structures and provide several advantages, like a smaller size, improved biophysical properties as well as the possibility to engage difficult-to-address target proteins through surface-exposed binding sites. With three alternative binding scaffolds being approved by the US Food and Drug Administration (FDA) to date and many more currently investigated in clinical trials, scaffold proteins are on a rise to become promising next-generation therapeutics. Additionally, these binders have successfully been employed (pre)clinically in a variety of applications, e.g., as imaging reagents for the detection and monitoring of various cancers, given that their small format enables an efficient distribution in cancerous tissue while at the same time ensuring a rapid elimination from the system by the kidney. In addition, alternative scaffolds are often more robust than mAbs and have also proven to be versatile and efficient affinity reagents for applications in basic research, such as affinity purifications and structural biology. Advances in high-throughput screening using different display technologies and protein engineering have facilitated the generation of novel binding scaffolds with high affinities and specificities and have paved the way for the expansion of the alternative binding scaffold toolbox.

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References

  1. Köhler G, Milstein C. Continuous cultures of fused cells secreting antibody of predefined specificity. Nature. 1975;256:495–7. https://doi.org/10.1038/256495a0.

    Article  PubMed  Google Scholar 

  2. Kaplon H, Muralidharan M, Schneider Z, Reichert JM. Antibodies to watch in 2020. MAbs. 2020;12:1703531. https://doi.org/10.1080/19420862.2019.1703531.

    Article  CAS  PubMed  Google Scholar 

  3. Ansar W, Ghosh S. Monoclonal antibodies: a tool in clinical research. Indian J Clin Med. 2013;4:S11968. https://doi.org/10.4137/IJCM.S11968.

    Article  Google Scholar 

  4. Gao Y, Huang X, Zhu Y, Lv Z. A brief review of monoclonal antibody technology and its representative applications in immunoassays. J Immunoass Immunochem. 2018;39:351–64. https://doi.org/10.1080/15321819.2018.1515775.

    Article  CAS  Google Scholar 

  5. Moser AC, Hage DS. Immunoaffinity chromatography: an introduction to applications and recent developments. Bioanalysis. 2010;2:769–90. https://doi.org/10.4155/bio.10.31.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Pyzik M, Rath T, Lencer WI, et al. FcRn: the architect behind the immune and nonimmune functions of IgG and albumin. J Immunol. 2015;194:4595–603. https://doi.org/10.4049/jimmunol.1403014.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Li Z, Krippendorff B-F, Sharma S, et al. Influence of molecular size on tissue distribution of antibody fragments. MAbs. 2016;8:113–9. https://doi.org/10.1080/19420862.2015.1111497.

    Article  CAS  PubMed  Google Scholar 

  8. Shah DK, Betts AM. Antibody biodistribution coefficients: inferring tissue concentrations of monoclonal antibodies based on the plasma concentrations in several preclinical species and human. MAbs. 5:297–305. https://doi.org/10.4161/mabs.23684.

  9. Schaefer JV, Sedlák E, Kast F, et al. Modification of the kinetic stability of immunoglobulin G by solvent additives. MAbs. 2018;10:607–23. https://doi.org/10.1080/19420862.2018.1450126.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Ministro J, Manuel AM, Goncalves J. Therapeutic antibody engineering and selection strategies. Adv Biochem Eng Biotechnol. 2019;171:55–86.

    Google Scholar 

  11. Traxlmayr MW, Kiefer JD, Srinivas RR, et al. Strong enrichment of aromatic residues in binding sites from a charge-neutralized hyperthermostable Sso7d scaffold library. J Biol Chem. 2016;291:22496–508. https://doi.org/10.1074/jbc.M116.741314.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. McCrary BS, Edmondson SP, Shriver JW. Hyperthermophile protein folding thermodynamics: differential scanning calorimetry and chemical denaturation of Sac7d. J Mol Biol. 1996;264:784–805. https://doi.org/10.1006/jmbi.1996.0677.

    Article  CAS  PubMed  Google Scholar 

  13. Colgrave ML, Craik DJ. Thermal, chemical, and enzymatic stability of the cyclotide kalata B1: the importance of the cyclic cystine knot †. Biochemistry. 2004;43:5965–75. https://doi.org/10.1021/bi049711q.

    Article  CAS  PubMed  Google Scholar 

  14. Kintzing JR, Cochran JR. Engineered knottin peptides as diagnostics, therapeutics, and drug delivery vehicles. Curr Opin Chem Biol. 2016;34:143–50. https://doi.org/10.1016/j.cbpa.2016.08.022.

    Article  CAS  PubMed  Google Scholar 

  15. Vazquez-Lombardi R, Phan TG, Zimmermann C, et al. Challenges and opportunities for non-antibody scaffold drugs. Drug Discov Today. 2015;20:1271–83. https://doi.org/10.1016/j.drudis.2015.09.004.

    Article  CAS  PubMed  Google Scholar 

  16. Skerra A. Alternative non-antibody scaffolds for molecular recognition. Curr Opin Biotechnol. 2007;18:295–304. https://doi.org/10.1016/j.copbio.2007.04.010.

    Article  CAS  PubMed  Google Scholar 

  17. Simeon R, Chen Z. In vitro-engineered non-antibody protein therapeutics. Protein Cell. 2018;9:3–14. https://doi.org/10.1007/s13238-017-0386-6.

    Article  CAS  PubMed  Google Scholar 

  18. Lipovšek D, Carvajal I, Allentoff AJ, et al. Adnectin–drug conjugates for Glypican-3-specific delivery of a cytotoxic payload to tumors. Protein Eng Des Sel. 2018;31:159–71. https://doi.org/10.1093/protein/gzy013.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Wang W, Lu P, Fang Y, et al. Monoclonal antibodies with identical fc sequences can bind to FcRn differentially with pharmacokinetic consequences. Drug Metab Dispos. 2011;39:1469–77. https://doi.org/10.1124/dmd.111.039453.

    Article  CAS  PubMed  Google Scholar 

  20. Strohl WR. Fusion proteins for half-life extension of biologics as a strategy to make biobetters. BioDrugs. 2015;29:215–39. https://doi.org/10.1007/s40259-015-0133-6.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Kontermann RE. Half-life extended biotherapeutics. Expert Opin Biol Ther. 2016;16:903–15. https://doi.org/10.1517/14712598.2016.1165661.

    Article  CAS  PubMed  Google Scholar 

  22. JevsÌŒevar S, Kunstelj M, Porekar VG. PEGylation of therapeutic proteins. Biotechnol J. 2010;5:113–28. https://doi.org/10.1002/biot.200900218.

    Article  CAS  Google Scholar 

  23. Schlapschy M, Binder U, Borger C, et al. PASylation: a biological alternative to PEGylation for extending the plasma half-life of pharmaceutically active proteins. Protein Eng Des Sel. 2013;26:489–501. https://doi.org/10.1093/protein/gzt023.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Capon DJ, Chamow SM, Mordenti J, et al. Designing CD4 immunoadhesins for AIDS therapy. Nature. 1989;337:525–31. https://doi.org/10.1038/337525a0.

    Article  CAS  PubMed  Google Scholar 

  25. Kim J, Bronson CL, Hayton WL, et al. Albumin turnover: FcRn-mediated recycling saves as much albumin from degradation as the liver produces. Am J Physiol Liver Physiol. 2006;290:G352–60. https://doi.org/10.1152/ajpgi.00286.2005.

    Article  CAS  Google Scholar 

  26. Sleep D, Cameron J, Evans LR. Albumin as a versatile platform for drug half-life extension. Biochim Biophys Acta – Gen Subj. 2013;1830:5526–34. https://doi.org/10.1016/j.bbagen.2013.04.023.

    Article  CAS  Google Scholar 

  27. Nygren P-Å, Skerra A. Binding proteins from alternative scaffolds. J Immunol Methods. 2004;290:3–28. https://doi.org/10.1016/j.jim.2004.04.006.

    Article  CAS  PubMed  Google Scholar 

  28. Storz U. Intellectual property protection. MAbs. 2011;3:310–7. https://doi.org/10.4161/mabs.3.3.15530.

    Article  PubMed  PubMed Central  Google Scholar 

  29. Koide A, Bailey CW, Huang X, Koide S. The fibronectin type III domain as a scaffold for novel binding proteins. J Mol Biol. 1998;284:1141–51. https://doi.org/10.1006/jmbi.1998.2238.

    Article  CAS  PubMed  Google Scholar 

  30. Bloom L, Calabro V. FN3: a new protein scaffold reaches the clinic. Drug Discov Today. 2009;14:949–55. https://doi.org/10.1016/j.drudis.2009.06.007.

    Article  CAS  PubMed  Google Scholar 

  31. Gulyani A, Vitriol E, Allen R, et al. A biosensor generated via high-throughput screening quantifies cell edge Src dynamics. Nat Chem Biol. 2011;7:437–44. https://doi.org/10.1038/nchembio.585.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Gross GG, Junge JA, Mora RJ, et al. Recombinant probes for visualizing endogenous synaptic proteins in living neurons. Neuron. 2013;78:971–85. https://doi.org/10.1016/j.neuron.2013.04.017.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Nord K, Gunneriusson E, Ringdahl J, et al. Binding proteins selected from combinatorial libraries of an α-helical bacterial receptor domain. Nat Biotechnol. 1997;15:772–7. https://doi.org/10.1038/nbt0897-772.

    Article  CAS  PubMed  Google Scholar 

  34. Gunneriusson E, Nord K, Uhlén M, Nygren P-Å. Affinity maturation of a Taq DNA polymerase specific affibody by helix shuffling. Protein Eng Des Sel. 1999;12:873–8. https://doi.org/10.1093/protein/12.10.873.

    Article  CAS  Google Scholar 

  35. Orlova A, Magnusson M, Eriksson TLJ, et al. Tumor imaging using a picomolar affinity HER2 binding affibody molecule. Cancer Res. 2006;66:4339–48. https://doi.org/10.1158/0008-5472.CAN-05-3521.

    Article  CAS  PubMed  Google Scholar 

  36. Löfblom J, Feldwisch J, Tolmachev V, et al. Affibody molecules: engineered proteins for therapeutic, diagnostic and biotechnological applications. FEBS Lett. 2010;584:2670–80. https://doi.org/10.1016/j.febslet.2010.04.014.

    Article  CAS  PubMed  Google Scholar 

  37. Frejd FY, Kim K-T. Affibody molecules as engineered protein drugs. Exp Mol Med. 2017;49:e306. https://doi.org/10.1038/emm.2017.35.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Gera N, Hussain M, Wright RC, Rao BM. Highly stable binding proteins derived from the hyperthermophilic Sso7d scaffold. J Mol Biol. 2011;409:601–16. https://doi.org/10.1016/j.jmb.2011.04.020.

    Article  CAS  PubMed  Google Scholar 

  39. Mouratou B, Schaeffer F, Guilvout I, et al. Remodeling a DNA-binding protein as a specific in vivo inhibitor of bacterial secretin PulD. Proc Natl Acad Sci. 2007;104:17983–8. https://doi.org/10.1073/pnas.0702963104.

    Article  PubMed  Google Scholar 

  40. Kauke MJ, Traxlmayr MW, Parker JA, et al. An engineered protein antagonist of K-Ras/B-Raf interaction. Sci Rep. 2017;7:5831. https://doi.org/10.1038/s41598-017-05889-7.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Gocha T, Rao BM, DasGupta R. Identification and characterization of a novel Sso7d scaffold-based binder against Notch1. Sci Rep. 2017;7:12021. https://doi.org/10.1038/s41598-017-12246-1.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Goux M, Becker G, Gorré H, et al. Nanofitin as a new molecular-imaging agent for the diagnosis of epidermal growth factor receptor over-expressing tumors. Bioconjug Chem. 2017;28:2361–71. https://doi.org/10.1021/acs.bioconjchem.7b00374.

    Article  CAS  PubMed  Google Scholar 

  43. Kalichuk V, Renodon-Cornière A, Béhar G, et al. A novel, smaller scaffold for Affitins: showcase with binders specific for EpCAM. Biotechnol Bioeng. 2018;115:290–9. https://doi.org/10.1002/bit.26463.

    Article  CAS  PubMed  Google Scholar 

  44. Gebauer M, Skerra A. Engineering of binding functions into proteins. Curr Opin Biotechnol. 2019;60:230–41. https://doi.org/10.1016/j.copbio.2019.05.007.

    Article  CAS  PubMed  Google Scholar 

  45. Kalichuk V, Kambarev S, Béhar G, et al. Affitins: ribosome display for selection of Aho7c-based affinity proteins. Methods Mol Biol. 2020;2070:19–41.

    Article  CAS  Google Scholar 

  46. Silverman J, Lu Q, Bakker A, et al. Multivalent avimer proteins evolved by exon shuffling of a family of human receptor domains. Nat Biotechnol. 2005;23:1556–61. https://doi.org/10.1038/nbt1166.

    Article  CAS  PubMed  Google Scholar 

  47. Weidle UH, Auer J, Brinkmann U, et al. The emerging role of new protein scaffold-based agents for treatment of cancer. Cancer Genomics Proteomics. 2013;10:155–68.

    CAS  PubMed  Google Scholar 

  48. Moore SJ, Cochran JR. Engineering knottins as novel binding agents. Methods Enzymol. 2012;503:223–51.

    Article  CAS  Google Scholar 

  49. Avrutina O. Synthetic cystine-knot miniproteins – valuable scaffolds for polypeptide engineering. Adv Exp Med Biol. 2016;917:121–44.

    Article  CAS  Google Scholar 

  50. Werle M, Schmitz T, Huang H-L, et al. The potential of cystine-knot microproteins as novel pharmacophoric scaffolds in oral peptide drug delivery. J Drug Target. 2006;14:137–46. https://doi.org/10.1080/10611860600648254.

    Article  CAS  PubMed  Google Scholar 

  51. Miao Z, Ren G, Liu H, et al. An engineered knottin peptide labeled with 18 F for PET imaging of integrin expression. Bioconjug Chem. 2009;20:2342–7. https://doi.org/10.1021/bc900361g.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Plückthun A. Designed Ankyrin repeat proteins (DARPins): binding proteins for research, diagnostics, and therapy. Annu Rev Pharmacol Toxicol. 2015;55:489–511. https://doi.org/10.1146/annurev-pharmtox-010611-134654.

    Article  CAS  PubMed  Google Scholar 

  53. Jost C, Plückthun A. Engineered proteins with desired specificity: DARPins, other alternative scaffolds and bispecific IgGs. Curr Opin Struct Biol. 2014;27:102–12. https://doi.org/10.1016/j.sbi.2014.05.011.

    Article  CAS  PubMed  Google Scholar 

  54. Schilling J, Schöppe J, Plückthun A. From DARPins to LoopDARPins: novel LoopDARPin design allows the selection of low picomolar binders in a single round of ribosome display. J Mol Biol. 2014;426:691–721. https://doi.org/10.1016/j.jmb.2013.10.026.

    Article  CAS  PubMed  Google Scholar 

  55. Wurch T, Pierré A, Depil S. Novel protein scaffolds as emerging therapeutic proteins: from discovery to clinical proof-of-concept. Trends Biotechnol. 2012;30:575–82. https://doi.org/10.1016/j.tibtech.2012.07.006.

    Article  CAS  PubMed  Google Scholar 

  56. Mittal A, Böhm S, Grütter MG, et al. Asymmetry in the homodimeric ABC transporter MsbA recognized by a DARPin. J Biol Chem. 2012;287:20395–406. https://doi.org/10.1074/jbc.M112.359794.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Pecqueur L, Duellberg C, Dreier B, et al. A designed ankyrin repeat protein selected to bind to tubulin caps the microtubule plus end. Proc Natl Acad Sci. 2012;109:12011–6. https://doi.org/10.1073/pnas.1204129109.

    Article  PubMed  Google Scholar 

  58. Kummer L, Parizek P, Rube P, et al. Structural and functional analysis of phosphorylation-specific binders of the kinase ERK from designed ankyrin repeat protein libraries. Proc Natl Acad Sci. 2012;109:E2248–57. https://doi.org/10.1073/pnas.1205399109.

    Article  PubMed  Google Scholar 

  59. Kummer L, Hsu C-W, Dagliyan O, et al. Knowledge-based design of a biosensor to quantify localized ERK activation in living cells. Chem Biol. 2013;20:847–56. https://doi.org/10.1016/j.chembiol.2013.04.016.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  60. Amstutz P, Binz HK, Parizek P, et al. Intracellular kinase inhibitors selected from combinatorial libraries of designed Ankyrin repeat proteins. J Biol Chem. 2005;280:24715–22. https://doi.org/10.1074/jbc.M501746200.

    Article  CAS  PubMed  Google Scholar 

  61. Parizek P, Kummer L, Rube P, et al. Designed Ankyrin repeat proteins (DARPins) as novel isoform-specific intracellular inhibitors of c-Jun N-terminal kinases. ACS Chem Biol. 2012;7:1356–66. https://doi.org/10.1021/cb3001167.

    Article  CAS  PubMed  Google Scholar 

  62. Brauchle M, Hansen S, Caussinus E, et al. Protein interference applications in cellular and developmental biology using DARPins that recognize GFP and mCherry. Biol Open. 2014;3:1252–61. https://doi.org/10.1242/bio.201410041.

    Article  PubMed  PubMed Central  Google Scholar 

  63. Martin-Killias P, Stefan N, Rothschild S, et al. A novel fusion toxin derived from an EpCAM-specific designed Ankyrin repeat protein has potent antitumor activity. Clin Cancer Res. 2011;17:100–10. https://doi.org/10.1158/1078-0432.CCR-10-1303.

    Article  PubMed  Google Scholar 

  64. Goldstein R, Sosabowski J, Livanos M, et al. Development of the designed ankyrin repeat protein (DARPin) G3 for HER2 molecular imaging. Eur J Nucl Med Mol Imaging. 2015;42:288–301. https://doi.org/10.1007/s00259-014-2940-2.

    Article  CAS  PubMed  Google Scholar 

  65. Schlatter D, Brack S, Banner DW, et al. Generation, characterization and structural data of chymase binding proteins based on the human Fyn kinase SH3 domain. MAbs. 2012;4:497–508. https://doi.org/10.4161/mabs.20452.

    Article  PubMed  PubMed Central  Google Scholar 

  66. Dennis MS, Lazarus RA. Kunitz domain inhibitors of tissue factor-factor VIIa. I. Potent inhibitors selected from libraries by phage display. J Biol Chem. 1994;269:22129–36.

    CAS  PubMed  Google Scholar 

  67. Roberts BL, Markland W, Ley AC, et al. Directed evolution of a protein: selection of potent neutrophil elastase inhibitors displayed on M13 fusion phage. Proc Natl Acad Sci. 1992;89:2429–33. https://doi.org/10.1073/pnas.89.6.2429.

    Article  CAS  PubMed  Google Scholar 

  68. Roberts BL, Markland W, Siranosian K, et al. Protease inhibitor display M13 phage: selection of high-affinity neutrophil elastase inhibitors. Gene. 1992;121:9–15. https://doi.org/10.1016/0378-1119(92)90156-J.

    Article  CAS  PubMed  Google Scholar 

  69. Röttgen P, Collins J. A human pancreatic secretory trypsin inhibitor presenting a hypervariable highly constrained epitope via monovalent phagemid display. Gene. 1995;164:243–50. https://doi.org/10.1016/0378-1119(95)00441-8.

    Article  PubMed  Google Scholar 

  70. Richter A, Eggenstein E, Skerra A. Anticalins: exploiting a non-Ig scaffold with hypervariable loops for the engineering of binding proteins. FEBS Lett. 2014;588:213–8. https://doi.org/10.1016/j.febslet.2013.11.006.

    Article  CAS  PubMed  Google Scholar 

  71. Hosse RJ. A new generation of protein display scaffolds for molecular recognition. Protein Sci. 2006;15:14–27. https://doi.org/10.1110/ps.051817606.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  72. Barkovskiy M, Ilyukhina E, Dauner M, et al. An engineered lipocalin that tightly complexes the plant poison colchicine for use as antidote and in bioanalytical applications. Biol Chem. 2019;400:351–66. https://doi.org/10.1515/hsz-2018-0342.

    Article  CAS  PubMed  Google Scholar 

  73. Schlehuber S, Beste G, Skerra A. A novel type of receptor protein, based on the lipocalin scaffold, with specificity for digoxigenin. J Mol Biol. 2000;297:1105–20. https://doi.org/10.1006/jmbi.2000.3646.

    Article  CAS  PubMed  Google Scholar 

  74. Schonfeld D, Matschiner G, Chatwell L, et al. An engineered lipocalin specific for CTLA-4 reveals a combining site with structural and conformational features similar to antibodies. Proc Natl Acad Sci. 2009;106:8198–203. https://doi.org/10.1073/pnas.0813399106.

    Article  PubMed  Google Scholar 

  75. Dauner M, Eichinger A, Lücking G, et al. Reprogramming human siderocalin to neutralize petrobactin, the essential iron scavenger of anthrax bacillus. Angew Chem Int Ed. 2018;57:14619–23. https://doi.org/10.1002/anie.201807442.

    Article  CAS  Google Scholar 

  76. Edwardraja S, Eichinger A, Theobald I, et al. Rational design of an anticalin-type sugar-binding protein using a genetically encoded boronate side chain. ACS Synth Biol. 2017;6:2241–7. https://doi.org/10.1021/acssynbio.7b00199.

    Article  CAS  PubMed  Google Scholar 

  77. Umscheid CA, Margolis DJ, Grossman CE. Key concepts of clinical trials: a narrative review. Postgrad Med. 2011;123:194–204. https://doi.org/10.3810/pgm.2011.09.2475.

    Article  PubMed  PubMed Central  Google Scholar 

  78. Linaclotide. LiverTox: cinical and research information on drug-induced liver injury. National Institute of Diabetes and Digestive and Kidney Diseases, Bethesda (MD) [Updated 2019 May 13]. https://www.ncbi.nlm.nih.gov/books/NBK548021/.

  79. Cicardi M, Levy RJ, McNeil DL, et al. Ecallantide for the treatment of acute attacks in hereditary angioedema. N Engl J Med. 2010;363:523–31. https://doi.org/10.1056/NEJMoa0905079.

    Article  CAS  PubMed  Google Scholar 

  80. Chua F. Neutrophil elastase: mediator of extracellular matrix destruction and accumulation. Proc Am Thorac Soc. 2006;3:424–7. https://doi.org/10.1513/pats.200603-078AW.

    Article  CAS  PubMed  Google Scholar 

  81. Dunlevy FK, Martin SL, de Courcey F, et al. Anti-inflammatory effects of DX-890, a human neutrophil elastase inhibitor. J Cyst Fibros. 2012;11:300–4. https://doi.org/10.1016/j.jcf.2012.02.003.

    Article  CAS  PubMed  Google Scholar 

  82. Binz HK, Bakker TR, Phillips DJ, et al. Design and characterization of MP0250, a tri-specific anti-HGF/anti-VEGF DARPin® drug candidate. MAbs. 2017;9:1262–9. https://doi.org/10.1080/19420862.2017.1305529.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  83. Miller KL. Do investors value the FDA orphan drug designation? Orphanet J Rare Dis. 2017;12:114. https://doi.org/10.1186/s13023-017-0665-6.

    Article  PubMed  PubMed Central  Google Scholar 

  84. Link A, Hepp J, Reichen C, et al. Abstract 3752: preclinical pharmacology of MP0310: a 4-1BB/FAP bispecific DARPin drug candidate promoting tumor-restricted T-cell costimulation. In: Immunology. American Association for Cancer Research, 2018. p 3752.

    Google Scholar 

  85. Mross K, Richly H, Fischer R, et al. First-in-human phase I study of PRS-050 (angiocal), an anticalin targeting and antagonizing VEGF-A, in patients with advanced solid tumors. PLoS One. 2013;8:e83232. https://doi.org/10.1371/journal.pone.0083232.

    Article  PubMed  PubMed Central  Google Scholar 

  86. Rothe C, Skerra A. Anticalin® proteins as therapeutic agents in human diseases. BioDrugs. 2018;32:233–43. https://doi.org/10.1007/s40259-018-0278-1.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  87. Renders L, Budde K, Rosenberger C, et al. First-in-human Phase I studies of PRS-080#22, a hepcidin antagonist, in healthy volunteers and patients with chronic kidney disease undergoing hemodialysis. PLoS One. 2019;14:e0212023. https://doi.org/10.1371/journal.pone.0212023.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  88. Samkoe KS, Shahzad Sardar H, Gunn JR, et al. Measuring microdose ABY-029 fluorescence signal in a primary human soft-tissue sarcoma resection. In: Pogue BW, Gioux S, editors. Molecular-guided surgery: molecules, devices, and applications V: SPIE; 2019. p. 38.

    Google Scholar 

  89. Schiff D, Kesari S, de Groot J, et al. Phase 2 study of CT-322, a targeted biologic inhibitor of VEGFR-2 based on a domain of human fibronectin, in recurrent glioblastoma. Investig New Drugs. 2015;33:247–53. https://doi.org/10.1007/s10637-014-0186-2.

    Article  CAS  Google Scholar 

  90. Mullard A. Nine paths to PCSK9 inhibition. Nat Rev Drug Discov. 2017;16:299–301. https://doi.org/10.1038/nrd.2017.83.

    Article  CAS  PubMed  Google Scholar 

  91. Nord K, Nord O, Uhlén M, et al. Recombinant human factor VIII-specific affinity ligands selected from phage-displayed combinatorial libraries of protein A. Eur J Biochem. 2001;268:4269–77. https://doi.org/10.1046/j.1432-1327.2001.02344.x.

    Article  CAS  PubMed  Google Scholar 

  92. Grönwall C, Sjöberg A, Ramström M, et al. Affibody-mediated transferrin depletion for proteomics applications. Biotechnol J. 2007;2:1389–98. https://doi.org/10.1002/biot.200700053.

    Article  CAS  PubMed  Google Scholar 

  93. Rönnmark J, Grönlund H, Uhlén M, Nygren P-Å. Human immunoglobulin A (IgA)-specific ligands from combinatorial engineering of protein A. Eur J Biochem. 2002;269:2647–55. https://doi.org/10.1046/j.1432-1033.2002.02926.x.

    Article  CAS  PubMed  Google Scholar 

  94. Rönnmark J, Kampf C, Asplund A, et al. Affibody-β-galactosidase immunoconjugates produced as soluble fusion proteins in the Escherichia coli cytosol. J Immunol Methods. 2003;281:149–60. https://doi.org/10.1016/j.jim.2003.06.001.

    Article  CAS  PubMed  Google Scholar 

  95. Lundberg E, Höidén-Guthenberg I, Larsson B, et al. Site-specifically conjugated anti-HER2 Affibody® molecules as one-step reagents for target expression analyses on cells and xenograft samples. J Immunol Methods. 2007;319:53–63. https://doi.org/10.1016/j.jim.2006.10.013.

    Article  CAS  PubMed  Google Scholar 

  96. Rönnmark J, Hansson M, Nguyen T, et al. Construction and characterization of affibody-Fc chimeras produced in Escherichia coli. J Immunol Methods. 2002;261:199–211. https://doi.org/10.1016/S0022-1759(01)00563-4.

    Article  PubMed  Google Scholar 

  97. Renberg B, Nordin J, Merca A, et al. Affibody molecules in protein capture microarrays: evaluation of multidomain ligands and different detection formats. J Proteome Res. 2007;6:171–9. https://doi.org/10.1021/pr060316r.

    Article  CAS  PubMed  Google Scholar 

  98. Lyakhov I, Zielinski R, Kuban M, et al. HER2- and EGFR-specific affiprobes: novel recombinant optical probes for cell imaging. Chembiochem. 2010;11:345–50. https://doi.org/10.1002/cbic.200900532.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  99. Engfeldt T, Renberg B, Brumer H, et al. Chemical synthesis of triple-labelled three-helix bundle binding proteins for specific fluorescent detection of unlabelled protein. Chembiochem. 2005;6:1043–50. https://doi.org/10.1002/cbic.200400388.

    Article  CAS  PubMed  Google Scholar 

  100. Bernhagen D, De Laporte L, Timmerman P. High-affinity RGD-knottin peptide as a new tool for rapid evaluation of the binding strength of unlabeled RGD-peptides to α v β 3 , α v β 5 , and α 5 β 1 integrin receptors. Anal Chem. 2017;89:5991–7. https://doi.org/10.1021/acs.analchem.7b00554.

    Article  CAS  PubMed  Google Scholar 

  101. Muñoz-Maldonado C, Zimmer Y, Medová M. A comparative analysis of individual RAS mutations in cancer biology. Front Oncol. 2019;9:1088. https://doi.org/10.3389/fonc.2019.01088.

    Article  PubMed  PubMed Central  Google Scholar 

  102. Spencer-Smith R, Koide A, Zhou Y, et al. Inhibition of RAS function through targeting an allosteric regulatory site. Nat Chem Biol. 2017;13:62–8. https://doi.org/10.1038/nchembio.2231.

    Article  CAS  PubMed  Google Scholar 

  103. Wojcik J, Hantschel O, Grebien F, et al. A potent and highly specific FN3 monobody inhibitor of the Abl SH2 domain. Nat Struct Mol Biol. 2010;17:519–27. https://doi.org/10.1038/nsmb.1793.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  104. Skerra A. Alternative binding proteins: anticalins – harnessing the structural plasticity of the lipocalin ligand pocket to engineer novel binding activities. FEBS J. 2008;275:2677–83. https://doi.org/10.1111/j.1742-4658.2008.06439.x.

    Article  CAS  PubMed  Google Scholar 

  105. Theurillat J-P, Dreier B, Nagy-Davidescu G, et al. Designed ankyrin repeat proteins: a novel tool for testing epidermal growth factor receptor 2 expression in breast cancer. Mod Pathol. 2010;23:1289–97. https://doi.org/10.1038/modpathol.2010.103.

    Article  CAS  PubMed  Google Scholar 

  106. Sennhauser G, Grütter MG. Chaperone-assisted crystallography with DARPins. Structure. 2008;16:1443–53. https://doi.org/10.1016/j.str.2008.08.010.

    Article  CAS  PubMed  Google Scholar 

  107. Mittl PR, Ernst P, Plückthun A. Chaperone-assisted structure elucidation with DARPins. Curr Opin Struct Biol. 2020;60:93–100. https://doi.org/10.1016/j.sbi.2019.12.009.

    Article  CAS  PubMed  Google Scholar 

  108. Bandeiras TM, Hillig RC, Matias PM, et al. Structure of wild-type Plk-1 kinase domain in complex with a selective DARPin. Acta Crystallogr Sect D Biol Crystallogr. 2008;64:339–53. https://doi.org/10.1107/S0907444907068217.

    Article  CAS  Google Scholar 

  109. Schweizer A, Roschitzki-Voser H, Amstutz P, et al. Inhibition of caspase-2 by a designed Ankyrin repeat protein: specificity, structure, and inhibition mechanism. Structure. 2007;15:625–36. https://doi.org/10.1016/j.str.2007.03.014.

    Article  CAS  PubMed  Google Scholar 

  110. Sennhauser G, Amstutz P, Briand C, et al. Drug export pathway of multidrug exporter AcrB revealed by DARPin inhibitors. PLoS Biol. 2006;5:e7. https://doi.org/10.1371/journal.pbio.0050007.

    Article  CAS  PubMed Central  Google Scholar 

  111. Gumpena R, Lountos GT, Waugh DS. MBP-binding DARPins facilitate the crystallization of an MBP fusion protein. Acta Crystallogr Sect F Struct Biol Commun. 2018;74:549–57. https://doi.org/10.1107/S2053230X18009901.

    Article  CAS  Google Scholar 

  112. Batyuk A, Wu Y, Honegger A, et al. DARPin-based crystallization chaperones exploit molecular geometry as a screening dimension in protein crystallography. J Mol Biol. 2016;428:1574–88. https://doi.org/10.1016/j.jmb.2016.03.002.

    Article  CAS  PubMed  Google Scholar 

  113. Wu Y, Honegger A, Batyuk A, et al. Structural basis for the selective inhibition of c-Jun N-terminal kinase 1 determined by rigid DARPin–DARPin fusions. J Mol Biol. 2018;430:2128–38. https://doi.org/10.1016/j.jmb.2017.10.032.

    Article  CAS  PubMed  Google Scholar 

  114. Ernst P, Honegger A, van der Valk F, et al. Rigid fusions of designed helical repeat binding proteins efficiently protect a binding surface from crystal contacts. Sci Rep. 2019;9:16162. https://doi.org/10.1038/s41598-019-52121-9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  115. Schütz M, Batyuk A, Klenk C, et al. Generation of fluorogen-activating designed ankyrin repeat proteins (FADAs) as versatile sensor tools. J Mol Biol. 2016;428:1272–89. https://doi.org/10.1016/j.jmb.2016.01.017.

    Article  CAS  PubMed  Google Scholar 

  116. Xu S, Hu H-Y. Fluorogen-activating proteins: beyond classical fluorescent proteins. Acta Pharm Sin B. 2018;8:339–48. https://doi.org/10.1016/j.apsb.2018.02.001.

    Article  PubMed  PubMed Central  Google Scholar 

  117. Wang Y, Prosen DE, Mei L, et al. A novel strategy to engineer DNA polymerases for enhanced processivity and improved performance in vitro. Nucleic Acids Res. 2004;32:1197–207. https://doi.org/10.1093/nar/gkh271.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  118. Béhar G, Renodon-Cornière A, Mouratou B, Pecorari F. Affitins as robust tailored reagents for affinity chromatography purification of antibodies and non-immunoglobulin proteins. J Chromatogr A. 2016;1441:44–51. https://doi.org/10.1016/j.chroma.2016.02.068.

    Article  CAS  PubMed  Google Scholar 

  119. Kruziki MA, Bhatnagar S, Woldring DR, et al. A 45-amino-acid scaffold mined from the PDB for high-affinity ligand engineering. Chem Biol. 2015;22:946–56. https://doi.org/10.1016/j.chembiol.2015.06.012.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  120. Kruziki MA, Case BA, Chan JY, et al. 64 cu-labeled Gp2 domain for PET imaging of epidermal growth factor receptor. Mol Pharm. 2016;13:3747–55. https://doi.org/10.1021/acs.molpharmaceut.6b00538.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  121. Strauch E-M, Fleishman SJ, Baker D. Computational design of a pH-sensitive IgG binding protein. Proc Natl Acad Sci. 2014;111:675–80. https://doi.org/10.1073/pnas.1313605111.

    Article  CAS  PubMed  Google Scholar 

  122. Reichen C, Hansen S, Plückthun A. Modular peptide binding: from a comparison of natural binders to designed armadillo repeat proteins. J Struct Biol. 2014;185:147–62. https://doi.org/10.1016/j.jsb.2013.07.012.

    Article  CAS  PubMed  Google Scholar 

  123. Coates J. Armadillo repeat proteins: beyond the animal kingdom. Trends Cell Biol. 2003;13:463–71. https://doi.org/10.1016/S0962-8924(03)00167-3.

    Article  CAS  PubMed  Google Scholar 

  124. Parmeggiani F, Pellarin R, Larsen AP, et al. Designed Armadillo repeat proteins as general peptide-binding scaffolds: consensus design and computational optimization of the hydrophobic core. J Mol Biol. 2008;376:1282–304. https://doi.org/10.1016/j.jmb.2007.12.014.

    Article  CAS  PubMed  Google Scholar 

  125. Reichen C, Hansen S, Forzani C, et al. Computationally designed armadillo repeat proteins for modular peptide recognition. J Mol Biol. 2016;428:4467–89. https://doi.org/10.1016/j.jmb.2016.09.012.

    Article  CAS  PubMed  Google Scholar 

  126. Sassoon I, Blanc V. Antibody–drug conjugate (ADC) clinical pipeline: a review. Methods Mol Biol. 2013;1045:1–27.

    Article  Google Scholar 

  127. Birrer MJ, Moore KN, Betella I, Bates RC. Antibody-drug conjugate-based therapeutics: state of the science. JNCI J Natl Cancer Inst. 2019;111:538–49. https://doi.org/10.1093/jnci/djz035.

    Article  CAS  PubMed  Google Scholar 

  128. Merten H, Schaefer JV, Brandl F, et al. Facile site-specific multiconjugation strategies in recombinant proteins produced in bacteria. Methods Mol Biol. 2019;2033:253–73.

    Article  CAS  Google Scholar 

  129. Zhang Y, Auger S, Schaefer JV, et al. Site-selective enzymatic labeling of designed Ankyrin repeat proteins using protein farnesyltransferase. Methods Mol Biol. 2019;2033:207–19.

    Article  CAS  Google Scholar 

  130. Wang D, Tai PWL, Gao G. Adeno-associated virus vector as a platform for gene therapy delivery. Nat Rev Drug Discov. 2019;18:358–78. https://doi.org/10.1038/s41573-019-0012-9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  131. Dreier B, Honegger A, Hess C, et al. Development of a generic adenovirus delivery system based on structure-guided design of bispecific trimeric DARPin adapters. Proc Natl Acad Sci. 2013;110:E869–77. https://doi.org/10.1073/pnas.1213653110.

    Article  PubMed  Google Scholar 

  132. Hartmann J, Münch RC, Freiling R-T, et al. A library-based screening strategy for the identification of DARPins as ligands for receptor-targeted AAV and lentiviral vectors. Mol Ther Methods Clin Dev. 2018;10:128–43. https://doi.org/10.1016/j.omtm.2018.07.001.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  133. Hanzl A, Winter GE. Targeted protein degradation: current and future challenges. Curr Opin Chem Biol. 2020;56:35–41. https://doi.org/10.1016/j.cbpa.2019.11.012.

    Article  CAS  PubMed  Google Scholar 

  134. Chamberlain PP, Hamann LG. Development of targeted protein degradation therapeutics. Nat Chem Biol. 2019;15:937–44. https://doi.org/10.1038/s41589-019-0362-y.

    Article  CAS  PubMed  Google Scholar 

  135. Fulcher LJ, Hutchinson LD, Macartney TJ, et al. Targeting endogenous proteins for degradation through the affinity-directed protein missile system. Open Biol. 2017;7:170066. https://doi.org/10.1098/rsob.170066.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Authors and Affiliations

  1. Department of Antibody-Drug Conjugates and Targeted NBE Therapeutics, Merck KGaA, Darmstadt, Germany

    Doreen Koenning

  2. Chemical Biology and Therapeutics (CBT), Novartis Institutes for BioMedical Research, Basel, Switzerland

    Jonas V. Schaefer

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  1. Doreen Koenning
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  2. Jonas V. Schaefer
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Correspondence to Doreen Koenning or Jonas V. Schaefer .

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  1. Institute of Molecular Biotechnology, Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna (BOKU), Vienna, Austria

    Florian Rüker

  2. Christian Doppler Laboratory for Innovative Immunotherapeutics, Institute of Molecular Biotechnology, Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna (BOKU), Vienna, Austria

    Gordana Wozniak-Knopp

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Koenning, D., Schaefer, J.V. (2021). Alternative Binding Scaffolds: Multipurpose Binders for Applications in Basic Research and Therapy. In: Rüker, F., Wozniak-Knopp, G. (eds) Introduction to Antibody Engineering. Learning Materials in Biosciences. Springer, Cham. https://doi.org/10.1007/978-3-030-54630-4_9

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