biopreparatБИОпрепараты. Профилактика, диагностика, лечениеBiological Products. Prevention, Diagnosis, Treatment2221-996X2619-1156Scientific Centre for Expert Evaluation of Medicinal Products10.30895/2221-996X-2024-24-3-294-311biopreparat-576Research ArticleТЕМА НОМЕРА: РАЗРАБОТКА ДИАГНОСТИЧЕСКИХ, ЛЕЧЕБНЫХ И ПРОФИЛАКТИЧЕСКИХ ПРЕПАРАТОВ ПРОТИВОВИРУСНОГО ДЕЙСТВИЯISSUE TOPIC: DEVELOPMENT OF MEDICINAL PRODUCTS FOR DIAGNOSIS, TREATMENT, AND PREVENTION OF VIRAL INFECTIONSИммуногенность различных вариантов гена гликопротеина вирусов Эбола и Марбург в составе рекомбинантных аденовирусных векторовImmunogenicity of various variants of Ebola and Marburg virus glycoprotein genes in recombinant adenoviral vectorshttps://orcid.org/0000-0001-7147-1553ОжаровскаяТ. А.OzharovskaiaT. A.

Ожаровская Татьяна Андреевна, канд. биол. наук

ул. Гамалеи, д. 18, Москва, 123098

Tatiana A. Ozharovskaia, Cand. Sci. (Biol.)

18 Gamaleya St., Moscow 123098

t.ozh@yandex.ruhttps://orcid.org/0000-0001-7893-8419ЗубковаО. В.ZubkovaO. V.

Зубкова Ольга Вадимовна, канд. биол. наук

ул. Гамалеи, д. 18, Москва, 123098

Olga V. Zubkova, Cand. Sci. (Biol.)

18 Gamaleya St., Moscow 123098

olga-zubkova@yandex.ruhttps://orcid.org/0000-0003-3248-1227ПоповаО.PopovaO.

Попова Ольга

ул. Гамалеи, д. 18, Москва, 123098

Olga Popova

18 Gamaleya St., Moscow 123098

olga.popova31@yandex.ruhttps://orcid.org/0000-0002-8757-7026КовыршинаА. В.KovyrshinaA. V.

Ковыршина Анна Витальевна

ул. Гамалеи, д. 18, Москва, 123098

Anna V. Kovyrshina

18 Gamaleya St., Moscow 123098

anna-kovyrshina@mail.ruhttps://orcid.org/0009-0000-1965-0482ГолдовскаяП. П.GoldovskayaP. P.

Голдовская Полина Павловна

ул. Гамалеи, д. 18, Москва, 123098

Polina P. Goldovskaya

18 Gamaleya St., Moscow 123098

goldovskaya00@mail.ruhttps://orcid.org/0009-0008-6272-0368ВавиловаИ. В.VavilovaI. V.

Вавилова Ирина Викторовна

ул. Гамалеи, д. 18, Москва, 123098

Irina V. Vavilova

18 Gamaleya St., Moscow 123098

tigr-11@mail.ruhttps://orcid.org/0000-0003-2548-6142ДолжиковаИ. В.DolzhikovaI. V.

Должикова Инна Вадимовна, канд. биол. наук

ул. Гамалеи, д. 18, Москва, 123098

Inna V. Dolzhikova, Cand. Sci. (Biol.)

18 Gamaleya St., Moscow 123098

iv.dolzhikova@yandex.ruhttps://orcid.org/0000-0003-2704-6730ЕрмоловаЕ. И.ErmolovaE. I.

Ермолова Екатерина Ивановна, канд. биол. наук

ул. Гамалеи, д. 18, Москва, 123098

Ekaterina I. Ermolova, Cand. Sci. (Biol.)

18 Gamaleya St., Moscow 123098

aksenova16@yandex.ruhttps://orcid.org/0000-0003-1945-0397КундаМ. С.KundaM. S.

Кунда Марина Сергеевна, канд. биол. наук

ул. Гамалеи, д. 18, Москва, 123098

Marina S. Kunda, Cand. Sci. (Biol.)

18 Gamaleya St., Moscow 123098

markunda99@gmail.comhttps://orcid.org/0000-0001-5361-870XРыжоваН. Н.RyzhovaN. N.

Рыжова Наталья Николаевна, канд. биол. наук

ул. Гамалеи, д. 18, Москва, 123098

Natalia N. Ryzhova, Cand. Sci. (Biol.)

18 Gamaleya St., Moscow 123098

rynatalia@yandex.ruhttps://orcid.org/0000-0001-7206-3594ВоронинаО. Л.VoroninaO. L.

Воронина Ольга Львовна, канд. биол. наук, доц.

ул. Гамалеи, д. 18, Москва, 123098

Olga L. Voronina, Cand. Sci. (Biol.), Assoc. Prof.

18 Gamaleya St., Moscow 123098

olv550@gmail.comhttps://orcid.org/0000-0002-8518-1669ЩербининД. Н.ShcherbininD. N.

Щербинин Дмитрий Николаевич, канд. биол. наук

ул. Гамалеи, д. 18, Москва, 123098

Dmitry N. Shcherbinin, Cand. Sci. (Biol.)

18 Gamaleya St., Moscow 123098

dim284@inbox.ruhttps://orcid.org/0000-0002-1289-3411ЩебляковД. В.ShcheblyakovD. V.

Щебляков Дмитрий Викторович, канд. биол. наук

ул. Гамалеи, д. 18, Москва, 123098

Dmitry V. Shcheblyakov, Cand. Sci. (Biol.)

18 Gamaleya St., Moscow 123098

sdmitryv@yahoo.comhttps://orcid.org/0000-0003-4035-6581ЛогуновД. Ю.LogunovD. Y.

Логунов Денис Юрьевич, д-р биол. наук, академик РАН

ул. Гамалеи, д. 18, Москва, 123098

Denis Y. Logunov, Dr. Sci. (Biol.), Academician of the Russian Academy of Sciences

18 Gamaleya St., Moscow 123098

ldenisy@yahoo.comhttps://orcid.org/0000-0003-1769-5059ГинцбургА. Л.GintsburgA. L.

Гинцбург Александр Леонидович, д-р биол. наук, академик РАН, профессор

ул. Гамалеи, д. 18, Москва, 123098

Alexander L. Gintsburg, Dr. Sci. (Biol.), Academician of the Russian Academy of Sciences, Prof.

18 Gamaleya St., Moscow 123098

gintsburg@gamaleya.orgФедеральное государственное бюджетное учреждение «Национальный исследовательский центр эпидемиологии и микробиологии имени почетного академика Н.Ф. Гамалеи» Министерства здравоохранения Российской ФедерацииРоссияNational Research Center for Epidemiology and Microbiology named after the honorary academician N. F. GamaleyaRussian Federation202403102024243Разработка диагностических, лечебных и профилактических препаратов противовирусного действия294311Copyright © Ожаровская Т.А., Зубкова О.В., Попова О., Ковыршина А.В., Голдовская П.П., Вавилова И.В., Должикова И.В., Ермолова Е.И., Кунда М.С., Рыжова Н.Н., Воронина О.Л., Щербинин Д.Н., Щебляков Д.В., Логунов Д.Ю., Гинцбург А.Л., 20242024Ожаровская Т.А., Зубкова О.В., Попова О., Ковыршина А.В., Голдовская П.П., Вавилова И.В., Должикова И.В., Ермолова Е.И., Кунда М.С., Рыжова Н.Н., Воронина О.Л., Щербинин Д.Н., Щебляков Д.В., Логунов Д.Ю., Гинцбург А.Л.Ozharovskaia T.A., Zubkova O.V., Popova O., Kovyrshina A.V., Goldovskaya P.P., Vavilova I.V., Dolzhikova I.V., Ermolova E.I., Kunda M.S., Ryzhova N.N., Voronina O.L., Shcherbinin D.N., Shcheblyakov D.V., Logunov D.Y., Gintsburg A.L.Данная работа распространяется под лицензией Creative Commons Attribution 4.0.This work is licensed under a Creative Commons Attribution 4.0 License.https://www.biopreparations.ru/jour/article/view/576ВВЕДЕНИЕ

ВВЕДЕНИЕ. Вирусы Марбург и Эбола вызывают тяжелую геморрагическую лихорадку у людей и приматов. В настоящее время не зарегистрированы вакцины для одновременной профилактики вызываемых этими филовирусами заболеваний, способные предотвратить распространение заболевания или снизить его тяжесть. Исследования, направленные на выбор наиболее иммуногенной формы протективного антигена, являются необходимым этапом при разработке эффективных профилактических вакцин.

ЦЕЛЬ

ЦЕЛЬ. Оценка индукции гуморального иммунного ответа на введение животным рекомбинантных аденовирусных векторов, экспрессирующих различные формы гликопротеина вирусов Эбола и Марбург.

МАТЕРИАЛЫ И МЕТОДЫ

МАТЕРИАЛЫ И МЕТОДЫ. Рекомбинантные аденовирусы человека 5 серотипа получали методом гомологичной рекомбинации в клетках Escherichia coli. Культивирование аденовирусов проводили в культуре клеток HEK293 с последующей очисткой методом ультрацентрифугирования в ступенчатом градиенте плотности цезия хлорида. Полученные препараты аденовирусов были охарактеризованы на подлинность методом ПЦР и полногеномным секвенированием; по количеству вирусных частиц (спектрофлуориметрия) и инфекционных вирусных частиц (метод ТЦД50). Оценку титра гликопротеин-специфических IgG антител в сыворотке крови иммунизированных животных (мыши) проводили методом иммуноферментного анализа.

РЕЗУЛЬТАТЫ

РЕЗУЛЬТАТЫ. Получены рекомбинантные аденовирусы человека 5 серотипа, содержащие в своем геноме кассету с одной из форм гена гликопротеина, кодирующего полноразмерный гликопротеин (GP), GP с удаленным муциноподобным доменом, GP с удаленными гликановым кэпом и муциноподобным доменом. Каждую из форм изучали на примере GP четырех представителей филовирусов: вируса Эбола видов Заир, Судан, Бундибугио и вируса Марбург. Показано, что форма GP не оказывает критического влияния на репликативную способность рекомбинантного аденовируса. Установлено, что через 3 нед. после иммунизации наибольший уровень продукции антиген-специфичных антител вызывают аденовирусы, кодирующие полноразмерный GP и GP с удаленным муциноподобным доменом. Вне зависимости от вида филовируса антиген, представленный GP с удаленными гликановым кэпом и муциноподобным доменом, оказался наименее иммуногенным.

ВЫВОДЫ

ВЫВОДЫ. Наиболее перспективными для разработки филовирусных вакцин на основе рекомбинантных аденовирусных векторов являются конструкции, включающие ген полноразмерного GP и GP с удаленным муциноподобным доменом.

INTRODUCTION

INTRODUCTION. Marburg and Ebola viruses cause severe haemorrhagic fever in humans and primates. Currently, there are no licensed prophylactic vaccines that can simultaneously prevent the spread or reduce the severity of both diseases caused by these filoviruses. The development of effective prophylactic vaccines requires studies aimed at selecting the most immunogenic forms of protective antigens.

AIM

AIM. This study aimed to evaluate humoral immune induction in animals after administration of recombinant adenoviral vectors expressing various forms of Ebola and Marburg virus glycoproteins (GPs).

MATERIALS AND METHODS

MATERIALS AND METHODS. Samples of recombinant human adenovirus type 5 (rAd5) were obtained using homologous recombination in Escherichia coli, growth in HEK293 cells, and purification by CsCl gradient ultracentrifugation. The resulting rAd5 samples were characterised in terms of their identity (PCR and whole-genome sequencing), the concentration of viral particles (fluorescence spectroscopy), and the concentration of infectious viral particles (TCID50 assay). Enzyme-linked immunosorbent assay (ELISA) was used to evaluate the GP-specific IgG titres in the sera of immunised mice.

RESULTS

RESULTS. The authors constructed rAd5 samples, and each construct contained an expression cassette with a GP gene form encoding a full-length GP, a GP without the mucin-like domain, or a GP without both the glycan cap and the mucin-like domain. Each of these forms was studied using the GPs of four filoviruses, including Zaire Ebola virus, Sudan Ebola virus, Bundibugyo Ebola virus, and Marburg virus. Neither of the forms had a critical effect on the rAd5 replicative capacity. Three weeks after immunisation, the highest GP-specific IgG production was induced by the rAd5 samples encoding either the full-length GP or the GP without the mucin-like domain. The GP without both the glycan cap and the mucin-like domain was the least immunogenic antigen regardless of the filovirus species.

CONCLUSIONS

CONCLUSIONS. The most promising constructs for the development of filovirus vaccines based on recombinant adenoviral vectors are the constructs that include the genes encoding the fulllength GP or the GP without the mucin-like domain.

иммуногенностьварианты гена гликопротеинавирус Эболавирус Марбургаденовирус человека 5 серотипарекомбинантные аденовирусные векторыболезньвызванная вирусом Эболаболезньвызванная вирусом Марбурггуморальный иммунный ответimmunogenicityglycoprotein gene variantsEbola virusMarburg virushuman adenovirus type 5recombinant adenoviral vectorsEbola virus diseaseMarburg virus diseasehumoral immune responseРабота выполнена в рамках государственного задания Минздрава России «Разработка пан-филовирусной вакцины (для профилактики болезни, вызванной вирусом Эбола, и геморрагической лихорадки Марбург)» (рег. номер АААА-А20-120022890039-0)This study was funded by the Ministry of Health of the Russian Federation under the State Assignment “Development of a pan-filovirus vaccine (for the prevention of Ebola virus disease and Marburg haemorrhagic fever)” (R&D Registry No. AAAA-A20-120022890039-0)ReferencesDolzhikova IV, Zubkova OV, Tukhvatulin AI, Dzharullaeva AS, Tukhvatulina NM, Shcheblyakov DV, et al. Safety and immunogenicity of GamEvac-Combi, a heterologous VSVand Ad5-vectored Ebola vaccine: An open phase I/II trial in healthy adults in Russia. Hum Vaccin Immunother. 2017;13(3):613–20. https://doi.org/10.1080/21645515.2016.1238535Dolzhikova IV, Zubkova OV, Tukhvatulin AI, Dzharullaeva AS, Tukhvatulina NM, Shcheblyakov DV, et al. Safety and immunogenicity of GamEvac-Combi, a heterologous VSVand Ad5-vectored Ebola vaccine: An open phase I/II trial in healthy adults in Russia. Hum Vaccin Immunother. 2017;13(3):613–20. https://doi.org/10.1080/21645515.2016.1238535Zhang Z, Zhao Z, Wang Y, Wu S, Wang B, Zhang J, et al. Comparative immunogenicity analysis of intradermal versus intramuscular immunization with a recombinant human adenovirus type 5 vaccine against Ebola virus. Front Immunol. 2022;13:963049. https://doi.org/10.3389/fimmu.2022.963049Zhang Z, Zhao Z, Wang Y, Wu S, Wang B, Zhang J, et al. Comparative immunogenicity analysis of intradermal versus intramuscular immunization with a recombinant human adenovirus type 5 vaccine against Ebola virus. Front Immunol. 2022;13:963049. https://doi.org/10.3389/fimmu.2022.963049Ahmed I, Salsabil L, Hossain MJ, Shahriar M, Bhuiyan MA, Islam MR. The recent outbreaks of Marburg virus disease in African countries are indicating potential threat to the global public health: Future prediction from historical data. Health Sci Reports. 2023;6(7):e1395. https://doi.org/10.1002/hsr2.1395Ahmed I, Salsabil L, Hossain MJ, Shahriar M, Bhuiyan MA, Islam MR. The recent outbreaks of Marburg virus disease in African countries are indicating potential threat to the global public health: Future prediction from historical data. Health Sci Reports. 2023;6(7):e1395. https://doi.org/10.1002/hsr2.1395Rojas M, Monsalve DM, Pacheco Y, Acosta-Ampudia Y, Ramírez-Santana C, Ansari AA, et al. Ebola virus disease: An emerging and re-emerging viral threat. J Autoimmun. 2020;106:102375. https://doi.org/10.1016/j.jaut.2019.102375Rojas M, Monsalve DM, Pacheco Y, Acosta-Ampudia Y, Ramírez-Santana C, Ansari AA, et al. Ebola virus disease: An emerging and re-emerging viral threat. J Autoimmun. 2020;106:102375. https://doi.org/10.1016/j.jaut.2019.102375Jones SM, Feldmann H, Ströher U, Geisbert JB, Fernando L, Grolla A, et al. Live attenuated recombinant vaccine pro tects nonhuman primates against Ebola and Marburg vi ruses. Nat Med. 2005;11(7):786–90. https://doi.org/10.1038/nm1258Jones SM, Feldmann H, Ströher U, Geisbert JB, Fernando L, Grolla A, et al. Live attenuated recombinant vaccine pro tects nonhuman primates against Ebola and Marburg vi ruses. Nat Med. 2005;11(7):786–90. https://doi.org/10.1038/nm1258Swenson DL, Wang D, Luo M, Warfield KL, Woraratanadharm J, Holman DH, et al. Vaccine to confer to nonhuman primates complete protection against multistrain Ebola and Marburg virus infections. Clin Vaccine Immunol. 2008;15(3):460–7. https://doi.org/10.1128/CVI.00431-07Swenson DL, Wang D, Luo M, Warfield KL, Woraratanadharm J, Holman DH, et al. Vaccine to confer to nonhuman primates complete protection against multistrain Ebola and Marburg virus infections. Clin Vaccine Immunol. 2008;15(3):460–7. https://doi.org/10.1128/CVI.00431-07Afolabi MO, Ishola D, Manno D, Keshinro B, Bockstal V, Rogers B, et al. Safety and immunogenicity of the twodose heterologous Ad26.ZEBOV and MVA-BN-Filo Ebola vaccine regimen in children in Sierra Leone: a ran domised, double-blind, controlled trial. Lancet Infect Dis. 2022;22(1):110–22. https://doi.org/10.1016/S1473-3099(21)00128-6Afolabi MO, Ishola D, Manno D, Keshinro B, Bockstal V, Rogers B, et al. Safety and immunogenicity of the twodose heterologous Ad26.ZEBOV and MVA-BN-Filo Ebola vaccine regimen in children in Sierra Leone: a ran domised, double-blind, controlled trial. Lancet Infect Dis. 2022;22(1):110–22. https://doi.org/10.1016/S1473-3099(21)00128-6Peng W, Rayaprolu V, Parvate AD, Pronker MF, Hui S, Parekh D, et al. Glycan shield of the ebolavirus envelope glycoprotein GP. Commun Biol. 2022;5(1):785. https://doi.org/10.1038/s42003-022-03767-1Peng W, Rayaprolu V, Parvate AD, Pronker MF, Hui S, Parekh D, et al. Glycan shield of the ebolavirus envelope glycoprotein GP. Commun Biol. 2022;5(1):785. https://doi.org/10.1038/s42003-022-03767-1Ahmad I, Fatemi SN, Ghaheri M, Rezvani A, Khezri DA, Natami M, et al. An overview of the role of Niemann– Pick C1 (NPC1) in viral infections and inhibition of viral infections through NPC1 inhibitor. Cell Commun Signal. 2023;21(1):352. https://doi.org/10.1186/s12964-023-01376-xAhmad I, Fatemi SN, Ghaheri M, Rezvani A, Khezri DA, Natami M, et al. An overview of the role of Niemann– Pick C1 (NPC1) in viral infections and inhibition of viral infections through NPC1 inhibitor. Cell Commun Signal. 2023;21(1):352. https://doi.org/10.1186/s12964-023-01376-xMurin CD, Gilchuk P, Crowe JE, Ward AB. Structural biology illuminates molecular determinants of broad ebolavirus neutralization by human antibodies for pan-ebolavirus therapeutic development. Front Immunol. 2022;12:808047. https://doi.org/10.3389/fimmu.2021.808047Murin CD, Gilchuk P, Crowe JE, Ward AB. Structural biology illuminates molecular determinants of broad ebolavirus neutralization by human antibodies for pan-ebolavirus therapeutic development. Front Immunol. 2022;12:808047. https://doi.org/10.3389/fimmu.2021.808047Зубкова ОВ, Ожаровская ТА, Должикова ИВ, Попова О, Щебляков ДВ, Гроусова ДМ и др. Экспрессионный вектор для создания иммунобиологического средства для индукции специфического иммунитета против вируса тяжелого острого респираторного синдрома SARS-CoV-2 (варианты). Патент Российской Федерации № 2731356 C9; 2021. EDN: KSWWVXZubkova OV, Ozharovskaia TA, Dolzhikova IV, Popova O, Shcheblyakov DV, Grousova DM, et al. Expression vector for creating immunobiological agent for inducing specific immunity to virus of severe acute respiratory syndrome SARS-COV-2 (embodiments). Patent of the Russian Federa tion No. 2731356 C9; 2021 (In Russ.). EDN: KSWWVXLock M, Korn M, Wilson J, Sena-Esteves M, Gao G. Measuring the infectious titer of recombinant adenovirus using tissue culture infection dose 50% (TCID50) end-point dilution and quantitative polymerase chain reaction (qPCR). Cold Spring Harb Protoc. 2019;2019(8):pdb.prot095562. https://doi.org/10.1101/pdb.prot095562Lock M, Korn M, Wilson J, Sena-Esteves M, Gao G. Measuring the infectious titer of recombinant adenovirus using tissue culture infection dose 50% (TCID50) end-point dilution and quantitative polymerase chain reaction (qPCR). Cold Spring Harb Protoc. 2019;2019(8):pdb.prot095562. https://doi.org/10.1101/pdb.prot095562Whitt MA. Generation of VSV pseudotypes using recombinant ΔG-VSV for studies on virus entry, identification of entry inhibitors, and immune responses to vaccines. J Virol Methods. 2010;169(2):365–74. https://doi.org/10.1016/j.jviromet.2010.08.006Whitt MA. Generation of VSV pseudotypes using recombinant ΔG-VSV for studies on virus entry, identification of entry inhibitors, and immune responses to vaccines. J Virol Methods. 2010;169(2):365–74. https://doi.org/10.1016/j.jviromet.2010.08.006Ожаровская ТА, Попова О, Зубкова ОВ, Вавилова ИВ, Почтовый АА, Щебляков ДЕ и др. Резработка и характеристика векторной системы на основе аденовируса обезьян 25-го серотипа. Вестник Российского государственного медицинского университета. 2023;(1):4–11. https://doi.org/10.24075/vrgmu.2023.006Ozharovskaia TA, Popova O, Zubkova OV, Vavilova IV, Pochtovyy AA, Shcheblyakov DE, et al. Development and characterization of a vector system based on the simian adenovirus type 25. Bull Russ State Med Univ. 2023;(1):4-11 (In Russ.). https://doi.org/10.24075/vrgmu.2023.006Carroll SA, Towner JS, Sealy TK, McMullan LK, Khris tova ML, Burt FJ, et al. Molecular evolution of viruses of the family Filoviridae based on 97 whole-genome sequences. J Virol. 2013;87(5):2608–16. https://doi.org/10.1128/JVI.03118-12Carroll SA, Towner JS, Sealy TK, McMullan LK, Khris tova ML, Burt FJ, et al. Molecular evolution of viruses of the family Filoviridae based on 97 whole-genome sequences. J Virol. 2013;87(5):2608–16. https://doi.org/10.1128/JVI.03118-12Yang ZY, Duckers HJ, Sullivan NJ, Sanchez A, Nabel EG, Nabel GJ. Identification of the Ebola virus glycoprotein as the main viral determinant of vascular cell cytotoxicity and injury. Nat Med. 2000;6(8):886–9. https://doi.org/10.1038/78645Yang ZY, Duckers HJ, Sullivan NJ, Sanchez A, Nabel EG, Nabel GJ. Identification of the Ebola virus glycoprotein as the main viral determinant of vascular cell cytotoxicity and injury. Nat Med. 2000;6(8):886–9. https://doi.org/10.1038/78645Zhao Y, Ren J, Harlos K, Jones DM, Zeltina A, Bowden TA, et al. Toremifene interacts with and destabilizes the Ebola virus glycoprotein. Nature. 2016;535(7610):169–72. https://doi.org/10.1038/nature18615Zhao Y, Ren J, Harlos K, Jones DM, Zeltina A, Bowden TA, et al. Toremifene interacts with and destabilizes the Ebola virus glycoprotein. Nature. 2016;535(7610):169–72. https://doi.org/10.1038/nature18615Cottingham MG, Carroll F, Morris SJ, Turner AV, Vaughan AM, Kapulu MC, et al. Preventing spontaneous genetic rearrangements in the transgene cassettes of adenovirus vec tors. Biotechnol Bioeng. 2012;109(3):719–28. https://doi.org/10.1002/bit.24342Cottingham MG, Carroll F, Morris SJ, Turner AV, Vaughan AM, Kapulu MC, et al. Preventing spontaneous genetic rearrangements in the transgene cassettes of adenovirus vec tors. Biotechnol Bioeng. 2012;109(3):719–28. https://doi.org/10.1002/bit.24342Gallais Y, Sierocki R, Lhomme G, Sivelle C, Kiseljak D, Wurm F, et al. Large-scale mapping of the Ebola NP and GP proteins reveals multiple immunoprevalent and conserved CD4 T-cell epitopes. Cell Mol Immunol. 2021;18(5):1323–5. https://doi.org/10.1038/s41423-020-0455-2Gallais Y, Sierocki R, Lhomme G, Sivelle C, Kiseljak D, Wurm F, et al. Large-scale mapping of the Ebola NP and GP proteins reveals multiple immunoprevalent and conserved CD4 T-cell epitopes. Cell Mol Immunol. 2021;18(5):1323–5. https://doi.org/10.1038/s41423-020-0455-2Bhatia B, Furuyama W, Hoenen T, Feldmann H, Marzi A. Ebola virus glycoprotein domains associated with protective efficacy. Vaccines. 2021;9(6):630. https://doi.org/10.3390/vaccines9060630Bhatia B, Furuyama W, Hoenen T, Feldmann H, Marzi A. Ebola virus glycoprotein domains associated with protective efficacy. Vaccines. 2021;9(6):630. https://doi.org/10.3390/vaccines9060630Meyer M, Yoshida A, Ramanathan P, Saphire EO, Collins PL, Crowe JE, et al. Antibody repertoires to the same Ebola vaccine antigen are differentially affected by vaccine vectors. Cell Rep. 2018;24(7):1816–29. https://doi.org/10.1016/j.celrep.2018.07.044Meyer M, Yoshida A, Ramanathan P, Saphire EO, Collins PL, Crowe JE, et al. Antibody repertoires to the same Ebola vaccine antigen are differentially affected by vaccine vectors. Cell Rep. 2018;24(7):1816–29. https://doi.org/10.1016/j.celrep.2018.07.044Kimble JB, Malherbe DC, Meyer M, Gunn BM, Karim MM, Ilinykh PA, et al. Antibody-mediated protective mecha nisms induced by a trivalent parainfluenza virus-vectored ebolavirus vaccine. J Virol. 2019;93(4):e01845–18. https://doi.org/10.1128/JVI.01845-18Kimble JB, Malherbe DC, Meyer M, Gunn BM, Karim MM, Ilinykh PA, et al. Antibody-mediated protective mecha nisms induced by a trivalent parainfluenza virus-vectored ebolavirus vaccine. J Virol. 2019;93(4):e01845–18. https://doi.org/10.1128/JVI.01845-18Murin CD, Fusco ML, Bornholdt ZA, Qiu X, Olinger GG, Zeitlin L, et al. Structures of protective antibodies reveal sites of vulnerability on Ebola virus. Proc Natl Acad Sci USA. 2014;111(48):17182–7. https://doi.org/10.1073/pnas.1414164111Murin CD, Fusco ML, Bornholdt ZA, Qiu X, Olinger GG, Zeitlin L, et al. Structures of protective antibodies reveal sites of vulnerability on Ebola virus. Proc Natl Acad Sci USA. 2014;111(48):17182–7. https://doi.org/10.1073/pnas.1414164111Saphire EO, Schendel SL, Fusco ML, Gangavarapu K, Gunn BM, Wec AZ, et al. Systematic analysis of monoclonal antibodies against Ebola virus GP defines features that contribute to protection. Cell. 2018;174(4):938–952.e13. https://doi.org/10.1016/j.cell.2018.07.033Saphire EO, Schendel SL, Fusco ML, Gangavarapu K, Gunn BM, Wec AZ, et al. Systematic analysis of monoclonal antibodies against Ebola virus GP defines features that contribute to protection. Cell. 2018;174(4):938–952.e13. https://doi.org/10.1016/j.cell.2018.07.033Bornholdt ZA, Turner HL, Murin CD, Li W, Sok D, Souders CA, et al. Isolation of potent neutralizing antibodies from a survivor of the 2014 Ebola virus outbreak. Science. 2016;351(6277):1078–83. https://doi.org/10.1126/science.aad5788Bornholdt ZA, Turner HL, Murin CD, Li W, Sok D, Souders CA, et al. Isolation of potent neutralizing antibodies from a survivor of the 2014 Ebola virus outbreak. Science. 2016;351(6277):1078–83. https://doi.org/10.1126/science.aad5788Towner JS, Sealy TK, Khristova ML, Albariño CG, Conlan S, Reeder SA, et al. Newly discovered Ebola virus associated with hemorrhagic fever outbreak in Uganda. PLoS Pathog. 2008;4(11):e1000212. https://doi.org/10.1371/journal.ppat.1000212Towner JS, Sealy TK, Khristova ML, Albariño CG, Conlan S, Reeder SA, et al. Newly discovered Ebola virus associated with hemorrhagic fever outbreak in Uganda. PLoS Pathog. 2008;4(11):e1000212. https://doi.org/10.1371/journal.ppat.1000212Hensley LE, Mulangu S, Asiedu C, Johnson J, Honko AN, Stanley D, et al. Demonstration of cross-protective vaccine immunity against an emerging pathogenic Ebolavirus species. PLoS Pathog. 2010;6(5):e1000904. https://doi.org/10.1371/journal.ppat.1000904Hensley LE, Mulangu S, Asiedu C, Johnson J, Honko AN, Stanley D, et al. Demonstration of cross-protective vaccine immunity against an emerging pathogenic Ebolavirus species. PLoS Pathog. 2010;6(5):e1000904. https://doi.org/10.1371/journal.ppat.1000904Hevey M, Negley D, Pushko P, Smith J, Schmaljohn A. Marburg virus vaccines based upon alphavirus replicons protect guinea pigs and nonhuman primates. Virology. 1998;251(1):28–37. https://doi.org/10.1006/viro.1998.9367Hevey M, Negley D, Pushko P, Smith J, Schmaljohn A. Marburg virus vaccines based upon alphavirus replicons protect guinea pigs and nonhuman primates. Virology. 1998;251(1):28–37. https://doi.org/10.1006/viro.1998.9367Zhu F-C, Guan X-H, Li Y-H, Huang J-Y, Jiang T, Hou L-H, et al. Immunogenicity and safety of a recombinant adenovirus type-5-vectored COVID-19 vaccine in healthy adults aged 18 years or older: a randomised, double-blind, placebocontrolled, phase 2 trial. Lancet. 2020;396(10249):479–88. https://doi.org/10.1016/S0140-6736(20)31605-6Zhu F-C, Guan X-H, Li Y-H, Huang J-Y, Jiang T, Hou L-H, et al. Immunogenicity and safety of a recombinant adenovirus type-5-vectored COVID-19 vaccine in healthy adults aged 18 years or older: a randomised, double-blind, placebocontrolled, phase 2 trial. Lancet. 2020;396(10249):479–88. https://doi.org/10.1016/S0140-6736(20)31605-6Flickinger JC, Singh J, Carlson R, Leong E, Baybutt TR, Barton J, et al. Chimeric Ad5.F35 vector evades anti-adenovirus serotype 5 neutralization opposing GUCY2C-targeted antitumor immunity. J Immunother Cancer. 2020;8(2):e001046. https://doi.org/10.1136/jitc-2020-001046Flickinger JC, Singh J, Carlson R, Leong E, Baybutt TR, Barton J, et al. Chimeric Ad5.F35 vector evades anti-adenovirus serotype 5 neutralization opposing GUCY2C-targeted antitumor immunity. J Immunother Cancer. 2020;8(2):e001046. https://doi.org/10.1136/jitc-2020-001046Flickinger JC, Staudt RE, Singh J, Carlson RD, Barton JR, Baybutt TR, et al. Chimeric adenoviral (Ad5.F35) and listeria vector prime-boost immunization is safe and effective for cancer immunotherapy. NPJ Vaccines. 2022;7(1):61. https://doi.org/10.1038/s41541-022-00483-zFlickinger JC, Staudt RE, Singh J, Carlson RD, Barton JR, Baybutt TR, et al. Chimeric adenoviral (Ad5.F35) and listeria vector prime-boost immunization is safe and effective for cancer immunotherapy. NPJ Vaccines. 2022;7(1):61. https://doi.org/10.1038/s41541-022-00483-zMitchell DAJ, Dupuy LC, Sanchez-Lockhart M, Palacios G, Back JW, Shimanovskaya K, et al. Epitope mapping of Ebola virus dominant and subdominant glycoprotein epitopes facilitates construction of an epitope-based DNA vaccine able to focus the antibody response in mice. Hum Vaccin Immunother. 2017;13(12):2883–93. https://doi.org/10.1080/21645515.2017.1347740Mitchell DAJ, Dupuy LC, Sanchez-Lockhart M, Palacios G, Back JW, Shimanovskaya K, et al. Epitope mapping of Ebola virus dominant and subdominant glycoprotein epitopes facilitates construction of an epitope-based DNA vaccine able to focus the antibody response in mice. Hum Vaccin Immunother. 2017;13(12):2883–93. https://doi.org/10.1080/21645515.2017.1347740

The authors declare that there are no conflicts of interest present.