Induction of cross-reactive antibodies in mice immunized with conserved influenza A virus neuraminidase-derived linear B-cell epitopes

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Introduction. Influenza is a socially significant infection that causes profound damage to populational health and national economy annually. Preventive vaccination is the most effective means to manage influenza and its complications. Diverse influenza vaccines exist, but their common drawback is the narrow specificity, the need for annual renewal of the strain composition, not always satisfactory immunogenicity and effectiveness. In this regard, close attention is paid to developing universal influenza vaccines aimed to induce cross-reactive immune-related cues against most conserved parts of viral proteins. Antibodies against neuraminidase (NA) are able to provide heterosubtypic protection, which is important due to potential threat posed by influenza viruses differed in hemagglutinin and neuraminidase sequence in comparison to currently circulating viruses. The present study is aimed to search for new and analysis of earlier predicted NA linear B-cell epitopes conserved among all influenza A virus subtypes. Results. Eight conserved linear B-cell epitopes were identified around the active site of neuraminidase, three of which (MNPNQKIITIGS, ILRTQESEC, and DNWKGSNRP) were synthesized de novo, conjugated with bovine serum albumin to be next used for mouse immunization. IgG antibodies were detected by ELISA in the sera of immunized mice. Antibodies were found to specifically bind to various influenza A viruses containing NA subtypes N1, N2, N3, and N9. Immunization with NA peptides did not protect mice from substantial body weight loss after infection with lethal H1N1 influenza virus. However, all immunized mice survived during the observation period, whereas in the control group the survival rate was as low as 28.6%. Analyzing lung viral load in the mice infected with the H1N1 virus revealed no differences in virus titers on either day 4 or 8 post-infection. Nevertheless, the protective effect lacked after the mice were challenged with lethal H7N9 influenza virus: Mortality rate, body weight loss, and lung virus titers were comparable in immunized and control mice. Conclusions. The data obtained evidenced about serum cross-reactive anti-NA antibodies induced by immunization with NA peptides, as well as protective efficacy against infection caused by H1N1 virus, but not H7N9 virus. Such results hold promise and indicate that NA linear B-cell epitopes can be used for designing epitope-directed influenza vaccines, but a deeper and more comprehensive study on the specificity of conserved NA epitopes, as well as optimization of immunization schemes for achieving higher protective efficacy are required.

About the authors

I. A. Sychev

Institute of Experimental Medicine

Author for correspondence.
Email: sychev.ia@iemspb.ru

Ivan A. Sychev - Junior Researcher, A.A. Smorodintsev Department of Virology, Institute of Experimental Medicine.

197376, St. Petersburg, Pavlov str., 12.

Phone: +7 904 638-04-18 (mobile)

Россия

P. M. Kopeikin

Institute of Experimental Medicine

Email: pmkopeikin@mail.ru

Junior Researcher, Department of General Pathology and Pathophysiology, Institute of Experimental Medicine.

St. Petersburg.

Россия

E. V. Tsvetkova

Institute of Experimental Medicine; St. Petersburg State University

Email: evtsvetkova72@mail.ru

PhD (Biology), Associate Professor, Department of Biochemistry, St. Petersburg State University; Senior Researcher, Department of General Pathology and Pathophysiology, Institute of Experimental Medicine.

St. Petersburg.

Россия

K. V. Cheredova

Institute of Experimental Medicine

Email: xeniya.cheredova@yandex.ru

Investigator (Biologist), Department of General Pathology and Pathophysiology, Institute of Experimental Medicine.

St. Petersburg.

Россия

B. L. Milman

Institute of Experimental Medicine

Email: bormilman@yandex.ru

PhD, MD (Chemistry), Head of the Laboratory for Mass Spectrometry, Institute of Experimental Medicine.

St. Petersburg.

Россия

O. V. Shamova

Institute of Experimental Medicine

Email: oshamova@yandex.ru

PhD, MD (Biology), Head of the Department of General Pathology and Pathophysiology, Institute of Experimental Medicine.

St. Petersburg.

Россия

I. N. Isakova-Sivak

Institute of Experimental Medicine

Email: isakova.sivak@gmail.com

PhD, MD (Biology), Head of the Laboratory of Immunology and Prevention of Viral Diseases, A.A. Smorodintsev Department of Virology, Institute of Experimental Medicine.

St. Petersburg.

Россия

Y. A. Desheva

Institute of Experimental Medicine; St. Petersburg State University

Email: desheva@mail.ru

PhD, MD (Medicine), Associate Professor, Leading Researcher, A.A. Smorodintsev Department of Virology, Institute of Experimental Medicine; Professor of the Department of Fundamental Problems of Medicine and Medical Technologies, St. Petersburg State University.

St. Petersburg.

Россия

References

  1. О мерах по дальнейшему совершенствованию организационных форм работы с использованием экспериментальных животных: Приказ Министерства здравоохранения СССР № 755 от 12.08.1977 г. URL: https://docplayer.ru/31723947-Ministerstvo-zdravoohraneniya-sssr-prikaz-12-avgusta-1977-g-n-755.html (20.11.2020)
  2. Патент № 2428476C Российская Федерация, МПК C12N 7/00 (2006.01), A61K 39/145 (2006.01). Реассортантный штамм вируса гриппа RN 1/09-Swine A(H7N1) для определения антител к нейраминидазе при гриппозной инфекции и вакцинации: № 2010125452/10; заявлено 2010.06.21: опубликовано 2011.09.10 / Дешева Ю.А., Смолоногина Т.С., Руденко Л.Г., Киселева И.В., Ларионова Н.В. Патентообладатель: НИИ экспериментальной медицины Северо-Западного отделения РАМН. 8 с.
  3. Патент № 2464312C1 Российская Федерация, МПК C12N 7/00 (2006.01), C12R 1/93 (2006.01), G01N 33/53 (2006.01). Реассортантный штамм вируса гриппа RN2/57-Human A(H7N2) для определения антител к нейраминидазе при гриппозной инфекции и вакцинации: № 2011124663/10; заявлено 2011.06.16: опубликовано 2012.10.20 / Дешева Ю.А., Смолоногина Т.С., Руденко Л.Г. Патентообладатель: ФГБУ НИИ экспериментальной медицины Северо-Западного отделения РАМН. 8 с.
  4. Патент № 2507256C2 Российская Федерация, МПК C12N 7/00 (2006.01), A61K 39/145 (2006.01), A61P 31/00 (2006.01). Штамм вируса гриппа A/17/rnallard/IIugepjaiigbi/0O/95(II7N3) для производства живой и производства инактивированной гриппозных вакцин: № 2012108866/10; заявлено 2012.03.07: опубликовано 2013.09.20 / Дешева Ю.А., Руденко Л.Г., Александрова Г.И. Патентообладатель: ФГБУ НИИ экспериментальной медицины Северо-Западного отделения РАМН. 8 с.
  5. Патент № 2587629C1 Российская Федерация, МПК C12N 7/00 (2006.01), C12Q 1/00 (2006.01). Реассортантный штамм вируса гриппа RN9/13-Hunan A(H6N9) для определения антител к нейраминидазе при гриппозной инфекции и вакцинации: № 2014154110/10; заявлено 2014.12.29: опубликовано 2016.06.20 / Смолоногина Т.С., Дешева Ю.А. Патентообладатель: ФГБНУ Институт экспериментальной медицины. 11 с.
  6. Bao Y., Bolotov P., Dernovoy D., Kiryutin B., Zaslavsky L., Tatusova T., Ostell J., Lipnan D. The influenza virus resource at the National Center for Biotechnology Information. J. Virol., 2008, vol. 82, no. 2, pp. 596— 601. doi: 10.1128/JVI.02005-07
  7. Bradford M.M. A rapid and sensitive method for the quantitation of microgran quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem, 1976, vol. 72, pp. 248—254. doi: 10.1006/abio.1976.9999
  8. Bui C., Bethmont A., Chughtai A.A., Gardner L., Sarkar S., Hassan S., Seale H., MacIntyre C.R. A systematic review of the comparative epidemiology of avian and human influenza A H5N1 and H7N9 — lessons and unanswered questions. Transbound Emerg. Dis., 2016, vol. 63, no. 6, pp. 602—620. doi: 10.1111/tbed.12327
  9. Chen J., Liu H., Yang J., Chou K.C. Prediction of linear B-cell epitopes using amino acid pair antigenicity scale. Amino Acids, 2007, vol. 33, no. 3,pp. 423-428. doi: 10.1007/s00726-006-0485-9
  10. Chen Y.Q., Wohlbold T.J., Zheng N.Y., Huang M., Huang Y., Neu K.E., Lee J., Wan H., Rojas K.T., Kirkpatrick E., Henry C., Palm A.E., Stamper C.T., Lan L.Y., Topham D.J., Treanor J., Wrammert J., Ahmed R., Eichelberger M.C., Georgiou G., Krammer F., Wilson P.C. Influenza infection in humans induces broadly cross-reactive and protective neuraminidase-reactive antibodies. Cell, 2018, vol. 173, no. 2, pp. 417-429. doi: 10.1016/j.cell.2018.03.030
  11. Davydov Y.I., Tonevitsky A. Prediction of linear B-cell epitopes. Mol. Biol., 2009, vol. 43, no. 1, pp. 150-158. doi: 10.1134/S0026893309010208
  12. Desheva J.A., Lu X.H., Rekstin A.R., Rudenko L.G., Swayne D.E., Cox N.J., Katz J.M., Klimov A.I. Characte rization of an influenza A H5N2 reassortant as a candidate for live-attenuated and inactivated vaccines against highly pathogenic H5N1 viruses with pandemic potential. Vaccine, 2006, vol. 24, no. 47-48, pp. 6859- 6866. doi: 10.1016/j.vaccine.2006.06.023
  13. Dowdle W.R. Influenza anti-neuraminidase: the second best antibody. N. Engl. J. Med., 1972, vol. 286, no. 25, pp. 1360-1361. doi: 10.1056/NEJM197206222862511
  14. Doyle T.M., Hashem A.M., Li C., Van Domselaar G., Larocque L., Wang J., Smith D., Cyr T., Farnsworth A., He R., Hurt A.C., Brown E.G., Li X. Universal anti-neuraminidase antibody inhibiting all influenza A subtypes. Antiviral Res., 2013, vol. 100, no. 2, pp. 567-574. doi: 10.1016/j.antiviral.2013.09.018
  15. Eichelberger M.C., Monto A.S. Neuraminidase, the forgotten surface antigen, emerges as an influenza vaccine target for broadened protection. J. Infect. Dis., 2019, vol. 219, suppl. 1, pp. 75-80. doi: 10.1093/infdis/jiz017
  16. Eichelberger M.C., Morens D.M., Taubenberger J.K. Neuraminidase as an influenza vaccine antigen: a low hanging fruit, ready for picking to improve vaccine effectiveness. Curr. Opin. Immunol., 2018, vol. 53, pp. 38-44. doi: 10.1016/j.coi.2018.03.025
  17. Fmoc Solid-Phase Peptide Synthesis: A Practical Approach. Ed. by W.C. Chan, P.D. White. Oxford: Oxford University Press, 2000. 346 p.
  18. Gottlieb T., Ben-Yedidia T. Epitope-based approaches to a universal influenza vaccine. J. Autoimmun., 2014, vol. 54, pp. 15-20. doi: 10.1016/j.jaut.2014.07.005
  19. Gravel C., Li C., Wang J., Hashem A.M., Jaentschke B., Xu K.W., Lorbetskie B., Gingras G., Aubin Y., Domselaar G.V., Girard M., He R., Li X. Qualitative and quantitative analyses of virtually all subtypes of influenza A and B viral neuraminidases using antibodies targeting the universally conserved sequences. Vaccine, 2010, vol. 28, no. 36, pp. 5774-5784. doi: 10.1016/j.vaccine.2010.06.075
  20. Herrera-Rodriguez J., Meijerhof T., Niesters H.G., Stjernholm G., Hovden A.O., S0rensen B., Okvist M., Sommerfelt M.A., Huckriede A. A novel peptide-based vaccine candidate with protective efficacy against influenza A in a mouse model. Virology, 2018, vol. 515, pp. 21-28. doi: 10.1016/j.virol.2017.11.018
  21. Huang P., Xu Y., Ni H., Zhong J., Zhang X., Tan S., Wu D., Qiu B., Guan D., Wen M., Yan J., Zhang Y. Linear B-cell epitope mapping of neuraminidases of the 2009 A H1N1 viruses based on immunoinformatics. Vaccine, 2011, vol. 29, no. 6, pp. 12781282. doi: 10.1016/j.vaccine.2010.11.080
  22. Krammer F., Fouchier R.A.M., Eichelberger M.C., Webby R.J., Shaw-Saliba K., Wan H., Wilson P.C., Compans R.W., Skountzou I., Monto A.S. NAction! How can neuraminidase-based immunity contribute to better influenza virus vaccines? mBio, 2018, vol. 9, no. 2: e02332-17. doi: 10.1128/mBio.02332-17
  23. Larsen J.E., Lund O., Nielsen M. Improved method for predicting linear B-cell epitopes. Immunome Res., 2006, vol. 2: 2. doi: 10.1186/1745-7580-2-2
  24. Murphy B.R., Kasel J.A., Chanock R.M. Association of serum anti-neuraminidase antibody with resistance to influenza in man. N. Engl. J. Med., 1972, vol. 286, no. 25, pp. 1329-1332. doi: 10.1056/NEJM197206222862502
  25. Nichol K.L. Efficacy and effectiveness of influenza vaccination. Vaccine, 2008, vol. 26, suppl. 4, pp. 17-22. doi: 10.1016/j.vaccine.2008.07.048
  26. Okonechnikov K., Golosova O., Fursov M., UGENE team. Unipro UGENE: a unified bioinformatics toolkit. Bioinformatics, 2012, vol. 28, no. 8, pp. 1166-1167. doi: 10.1093/bioinformatics/bts091
  27. Pace C.N. Evaluating contribution of hydrogen bonding and hydrophobic bonding to protein folding. Methods Enzymol., 1995, vol. 259, pp. 538-554. doi: 10.1016/0076-6879(95)59060-9
  28. Ponomarenko J., Bui H.H., Li W., Fusseder N., Bourne P.E., Sette A., Peters B. ElliPro: a new structure-based tool for the prediction of antibody epitopes. BMC Bioinformatics, 2008, vol. 9: 514. doi: 10.1186/1471-2105-9-514
  29. Quan F.S., Kim M.C., Lee B.J., Song J.M., Compans R.W., Kang S.M. Influenza M1 VLPs containing neuraminidase induce heterosubtypic cross-protection. Virology, 2012, vol. 430, no. 2, pp. 127-135. doi: 10.1016/j.virol.2012.05.006
  30. Reed L.J., Muench H.A. A simple method of estimating fifty per cent endpoints. Am. J. Epidemiol., 1938, vol. 27, no.3, pp. 493497. doi: 10.1093/oxfordjournals.aje.a118408
  31. Rubinstein N.D., Mayrose I., Martz E., Pupko T. Epitopia: a web-server for predicting B-cell epitopes. BMC Bioinformatics, 2009, vol. 10: 287. doi: 10.1186/1471-2105-10-287
  32. Saha S., Raghava G.P. Prediction of continuous B-cell epitopes in an antigen using recurrent neural network. Proteins, 2006, vol. 65, no. 1, pp. 40-48. doi: 10.1002/prot.21078
  33. Schulman J.L., Khakpour M., Kilbourne E.D. Protective effects of specific immunity to viral neuraminidase on influenza virus infection of mice. J. Virol., 1968, vol. 2, no. 8, pp. 778-786. doi: 10.1128/JVI.2.8.778-786.1968
  34. Soema P.C., Rosendahl Huber S.K., Willems G.J., Jacobi R., Hendriks M., Soethout E., Jiskoot W., de Jonge J., van Beek J., Kersten G.F.A., Amorij J.P. Whole-inactivated influenza virus is a potent adjuvant for influenza peptides containing CD8(+) T cell epitopes. Front. Immunol., 2018, vol. 9: 525. doi: 10.3389/fimmu.2018.00525
  35. Stadlbauer D., Zhu X., McMahon M., Turner J.S., Wohlbold T.J., Schmitz A.J., Strohmeier S., Yu W., Nachbagauer R., Mudd P.A., Wilson I.A., Ellebedy A.H., Krammer F. Broadly protective human antibodies that target the active site of influenza virus neuraminidase. Science, 2019, vol. 366, no. 6464, pp. 499-504. doi: 10.1126/science.aay0678
  36. Sweredoski M.J., Baldi P. COBEpro: a novel system for predicting continuous B-cell epitopes. Protein Eng. Des. Sel., 2009, vol. 22, no. 3, pp. 113-120. doi: 10.1093/protein/gzn075
  37. Wohlbold T.J., Krammer F. In the shadow of hemagglutinin: a growing interest in influenza viral neuraminidase and its role as a vaccine antigen. Viruses, 2014, vol. 6, no. 6, pp. 2465-2494. doi: 10.3390/v6062465
  38. Xiao J., Zhang L., Wang Z., Xiang W., Lu P., Zhao Y., Han M., Ma A., Qi P., Wang M., Gao G.F., Liu W.J. Conserved peptides enhance immune efficiency of inactive vaccines against emerging avian influenza viruses in chicken. Science China Life Sciences, 2017, vol. 60, no. 12, pp. 1340-1347. doi: 10.1007/s11427-017-9153-2

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Copyright (c) 2020 Sychev I.A., Kopeikin P.M., Tsvetkova E.V., Cheredova K.V., Milman B.L., Shamova O.V., Isakova-Sivak I.N., Desheva Y.A.

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