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Efficient expression of single chain variable fragment antibody against paclitaxel using the Bombyx mori nucleopolyhedrovirus bacmid DNA system and its characterizations

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Abstract

A single chain variable fragment (scFv), the smallest unit of functional recombinant antibody, is an attractive format of recombinant antibodies for various applications due to its small fragment and possibility of genetic engineering. Hybridoma clone 3A3 secreting anti-paclitaxel monoclonal antibody was used to construct genes encoding its variable domains of heavy (VH) and light (VL) chains. The VH and VL domains were linked to be the PT-scFv3A3 using flexible peptide linker in a format of VH-(GGGGS)5-VL. The PT-scFv3A3 was primarily expressed using the pET28a(+) vector in the Escherichia coli system, and was then further expressed by using the Bombyx mori nucleopolyhedrovirus (BmNPV) bacmid DNA system. Interestingly, the reactivity of PT-scFv3A3 expressed in the hemolymph of B. mori using the BmNPV bacmid DNA system was much higher than that expressed in the E. coli system. Using indirect competitive enzyme-linked immunosorbent assay (icELISA), the PT-scFv3A3 (B. mori) reacted not only with immobilized paclitaxel, but also with free paclitaxel in a concentration-dependent manner, with the linear range of free paclitaxel between 0.156 and 5.00 µg/ml. The PT-scFv3A3 (B. mori) exhibited less cross-reactivity (%) than its parental MAb clone 3A3 against paclitaxel-related compounds, including docetaxel (31.1 %), 7-xylosyltaxol (22.1 %), baccatin III (<0.68 %), 10-deacetylbaccatin III (<0.68 %), 1-hydroxybaccatin I (<0.68 %), and 1-acetoxy-5-deacetylbaccatin I (<0.68 %). With the exception of cephalomannine, the cross-reactivity was slightly increased to 8.50 %. The BmNPV bacmid DNA system was a highly efficient expression system of active PT-scFv3A3, which is applicable for PT-scFv3A3-based immunoassay of paclitaxel. In addition, the PT-scFv3A3 can be applied to evaluate its neutralizing property of paclitaxel or docetaxel toxicity.

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References

  1. de Marco A (2011) Biotechnological applications of recombinant single-domain antibody fragments. Microb Cell Fact 10:44

    Article  PubMed  PubMed Central  Google Scholar 

  2. Hoogenboom HR (2005) Selecting and screening recombinant antibody libraries. Nat Biotechnol 23:1105–1116

    Article  CAS  PubMed  Google Scholar 

  3. Boonrod K, Galetzka D, Nagy PD, Conrad U, Krczal G (2004) Single-chain antibodies against a plant viral RNA-dependent RNA polymerase confer virus resistance. Nat Biotechnol 22:856–862

    Article  CAS  PubMed  Google Scholar 

  4. Plana E, Moreno MJ, Montoya Á, Manclús JJ (2014) Development and application of recombinant antibody-based immunoassays to tetraconazole residue analysis in fruit juices. Food Chem 143:205–213

    Article  CAS  PubMed  Google Scholar 

  5. Leivo J, Lamminmäki U, Lövgren T, Vehniäinen M (2013) Multiresidue detection of fluoroquinolones: specificity engineering of a recombinant antibody with oligonucleotide-directed mutagenesis. J Agric Food Chem 61:11981–11985

    Article  CAS  PubMed  Google Scholar 

  6. Holliger P, Hudson PJ (2005) Engineered antibody fragments and the rise of single domains. Nat Biotechnol 23:1126–1136

    Article  CAS  PubMed  Google Scholar 

  7. Hudson PJ, Souriau C (2003) Engineered antibodies. Nat Med 9:129–134

    Article  CAS  PubMed  Google Scholar 

  8. Sakamoto S, Pongkitwitoon B, Sasaki-Tabata K, Putalun W, Maenaka K, Tanaka H, Morimoto S (2011) A fluorescent single domain antibody against plumbagin expressed in silkworm larvae for fluorescence-linked immunosorbent assay (FLISA). Analyst 136:2056–2063

    Article  CAS  PubMed  Google Scholar 

  9. Motohashi T, Shimojima T, Fukagawa T, Maenaka K, Park EY (2005) Efficient large-scale protein production of larvae and pupae of silkworm by Bombyx mori nuclear polyhedrosis virus bacmid system. Biochem Biophys Res Commun 326:564–569

    Article  CAS  PubMed  Google Scholar 

  10. Kato T, Kajikawa M, Maenaka K, Park EY (2010) Silkworm expression system as a platform technology in life science. Appl Microbiol Biotechnol 85:459–470

    Article  CAS  PubMed  Google Scholar 

  11. Sung LY, Chen CL, Lin SY, Li KC, Yeh CL, Chen GY, Lin CY, Hu YC (2014) Efficient gene delivery into cell lines and stem cells using baculovirus. Nat Protoc 9:1882–1899

    Article  CAS  PubMed  Google Scholar 

  12. Chan BS, Buckley NA (2014) Digoxin-specific antibody fragments in the treatment of digoxin toxicity. Clin Toxicol (Phila) 52:824–836

    Article  CAS  Google Scholar 

  13. Butler VP Jr, Schmidt DH, Smith TW, Haber E, Raynor BD, Demartini P (1977) Effects of sheep digoxin-specific antibodies and their Fab fragments on digoxin pharmacokinetics in dogs. J Clin Invest 59:345–359

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Baud FJ, Sabouraud A, Vicaut E, Taboulet P, Lang J, Bismuth C, Rouzioux JM, Scherrmann JM (1995) Brief report: treatment of severe colchicine overdose with colchicine-specific Fab fragments. N Engl J Med 332:642–645

    Article  CAS  PubMed  Google Scholar 

  15. Peake PW, Pianta TJ, Succar L, Fernando M, Buckley NA, Endre ZH (2015) Fab fragments of ovine antibody to colchicine enhance its clearance in the rat. Clin Toxicol (Phila) 53:427–432

    Article  CAS  Google Scholar 

  16. Heard K, Dart RC, Bogdan G, O’Malley GF, Burkhart KK, Donovan JW, Ward SB (2006) A preliminary study of tricyclic antidepressant (TCA) ovine FAB for TCA toxicity. Clin Toxicol (Phila) 44:275–281

    Article  CAS  Google Scholar 

  17. Yalindağ-Oztürk N, Goto CS, Shepherd G, Torres ON, Giroir B (2010) A pilot pharmacokinetic study of tricyclic antidepressant ovine Fab for TCA poisoning in children. Clin Toxicol (Phila) 48:418–423

    Article  Google Scholar 

  18. Bignami GS, Mooberry SL (1998) Monoclonal antibodies to taxanes that neutralize the biological activity of paclitaxel. Cancer Lett 126:127–133

    Article  CAS  PubMed  Google Scholar 

  19. Poi MJ, Berger M, Lustberg M, Layman R, Shapiro CL, Ramaswamy B, Mrozek E, Olson E, Wesolowski R (2013) Docetaxel-induced skin toxicities in breast cancer patients subsequent to paclitaxel shortage: a case series and literature review. Support Care Cancer 21:2679–2686

    Article  PubMed  PubMed Central  Google Scholar 

  20. Ziske CG, Schöttker B, Gorschlüter M, Mey U, Kleinschmidt R, Schlegel U, Sauerbruch T, Schmidt-Wolf IG (2002) Acute transient encephalopathy after paclitaxel infusion: report of three cases. Ann Oncol 13:629–631

    Article  CAS  PubMed  Google Scholar 

  21. Chao Z, Tan M, Paudel MK, Sakamoto S, Ma L, Sasaki-Tabata K, Tanaka H, Shoyama Y, Xuan L, Morimoto S (2013) Development of an indirect competitive enzyme-linked immunosorbent assay (icELISA) using highly specific monoclonal antibody against paclitaxel. J Nat Med 67:512–518

    Article  CAS  PubMed  Google Scholar 

  22. Krebber A, Bornhauser S, Burmester J, Honegger A, Willuda J, Bosshard HR, Plückthun A (1997) Reliable cloning of functional antibody variable domains from hybridomas and spleen cell repertoires employing a reengineered phage display system. J Immunol Methods 201:35–55

    Article  CAS  PubMed  Google Scholar 

  23. Sakamoto S, Pongkitwitoon B, Nakamura S, Maenaka K, Tanaka H, Morimoto S (2010) Efficient silkworm expression of single-chain variable fragment antibody against ginsenoside Re using Bombyx mori nucleopolyhedrovirus bacmid DNA system and its application in enzyme-linked immunosorbent assay for quality control of total ginsenosides. J Biochem 148:335–340

    Article  CAS  PubMed  Google Scholar 

  24. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  CAS  PubMed  Google Scholar 

  25. Bu D, Zhou Y, Tang J, **g F, Zhang W (2013) Expression and purification of a novel therapeutic single-chain variable fragment antibody against BNP from inclusion bodies of Escherichia coli. Protein Expr Purif 92:203–207

    Article  CAS  PubMed  Google Scholar 

  26. Weiler EW, Zenk MH (1976) Radioimmunoassay for the determination of digoxin and related compounds in Digitalis lanata. Phytochemistry 15:1537–1545

    Article  CAS  Google Scholar 

  27. Muraki M, Honda S (2011) Improved isolation and purification of functional human Fas receptor extracellular domain using baculovirus-silkworm expression system. Protein Expr Purif 80:102–109

    Article  CAS  PubMed  Google Scholar 

  28. Hirohata S, Wang LW, Miyagi M, Yan L, Seldin MF, Keene DR, Crabb JW, Apte SS (2002) Punctin, a novel ADAMTS-like molecule, ADAMTSL-1, in extracellular matrix. J Biol Chem 277:12182–12189

    Article  CAS  PubMed  Google Scholar 

  29. Du D, Kato T, Suzuki F, Park EY (2009) Expression of protein complex comprising the human prorenin and (pro)renin receptor in silkworm larvae using Bombyx mori nucleopolyhedrovirus (BmNPV) bacmids for improving biological function. Mol Biotechnol 43:154–156

    Article  CAS  PubMed  Google Scholar 

  30. Maesaki R, Satoh R, Taoka M, Kanaba T, Asano T, Fujita C, Fujiwara T, Ito Y, Isobe T, Hakoshima T, Maenaka K, Mishima M (2014) Efficient and cost effective production of active-form human PKB using silkworm larvae. Sci Rep 4:6016

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Tang L, Meibohm B (2006) Pharmacokinetics of peptides and proteins. In: Meibohm B (ed) Pharmacokinetics and pharmacodynamics of biotech drugs: principles and case studies in drug development. Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim, pp 15–43

    Chapter  Google Scholar 

  32. Miner JH (2008) Glomerular filtration: the charge debate charges ahead. Kidney Int 74:259–261

    Article  PubMed  Google Scholar 

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Acknowledgments

We acknowledge the financial support from the Japan Society for the Promotion of Science (no. 22501046 and no. 19590119) and from Takeda Science Foundation. We also appreciate the technical support from the Research Support Center, Graduate School of Medical Sciences, Kyushu University, Japan.

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Authors and Affiliations

  1. Department of Pharmacognosy, Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka, 812-8582, Japan

    Gorawit Yusakul, Seiichi Sakamoto, Hiroyuki Tanaka & Satoshi Morimoto

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  1. Gorawit Yusakul
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  2. Seiichi Sakamoto
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  3. Hiroyuki Tanaka
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  4. Satoshi Morimoto
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Correspondence to Hiroyuki Tanaka.

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Yusakul, G., Sakamoto, S., Tanaka, H. et al. Efficient expression of single chain variable fragment antibody against paclitaxel using the Bombyx mori nucleopolyhedrovirus bacmid DNA system and its characterizations. J Nat Med 70, 592–601 (2016). https://doi.org/10.1007/s11418-016-0981-5

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  • DOI: https://doi.org/10.1007/s11418-016-0981-5

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