[en] Snakebite envenoming can be a life-threatening medical emergency that requires prompt medical intervention to neutralise the effects of venom toxins. Each year up to 138,000 people die from snakebites and threefold more victims suffer life-altering disabilities. The current treatment of snakebite relies solely on antivenom—polyclonal antibodies isolated from the plasma of hyperimmunised animals—which is associated with numerous deficiencies. The ADDovenom project seeks to deliver a novel snakebite therapy, through the use of an innovative protein-based scaffold as a next-generation antivenom. The ADDomer is a megadalton-sized, thermostable synthetic nanoparticle derived from the adenovirus penton base protein; it has 60 high-avidity binding sites to neutralise venom toxins. Here, we outline our experimental strategies to achieve this goal using state-of-the-art protein engineering, expression technology and mass spectrometry, as well as in vitro and in vivo venom neutralisation assays. We anticipate that the approaches described here will produce antivenom with unparalleled efficacy, safety and affordability.
Disciplines :
Biochemistry, biophysics & molecular biology
Author, co-author :
Menzies, Stefanie K. ; Centre for Snakebite Research & Interventions, Liverpool School of Tropical Medicine, Pembroke Place, Liverpool L3 5QA, UK ; Centre for Drugs & Diagnostics, Liverpool School of Tropical Medicine, Pembroke Place, Liverpool L3 5QA, UK
Arinto-Garcia, Raquel ; iBET, Instituto de Biologia Experimental e Technológica, Apartado 12, 2781-901 Oeiras, Portugal ; ITQB NOVA, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa, Av. da República, 2780-157 Oeiras, Portugal
Gobbi Amorim, Fernanda ; Université de Liège - ULiège > Département de chimie (sciences) > Laboratoire de spectrométrie de masse (L.S.M.)
Cardoso, Iara Aimê ; Centre for Snakebite Research & Interventions, Liverpool School of Tropical Medicine, Pembroke Place, Liverpool L3 5QA, UK
Abada, Camille; Centre for Snakebite Research & Interventions, Liverpool School of Tropical Medicine, Pembroke Place, Liverpool L3 5QA, UK
Crasset, Thomas ; Université de Liège - ULiège > Faculté des Sciences > Master en sc. chimiques, à fin.
Durbesson, Fabien; Architecture et Fonction des Macromolécules Biologiques, CNRS, Aix-Marseille Université, 13009 Marseille, France
Edge, Rebecca J.; Centre for Snakebite Research & Interventions, Liverpool School of Tropical Medicine, Pembroke Place, Liverpool L3 5QA, UK
El-Kazzi, Priscila; Architecture et Fonction des Macromolécules Biologiques, CNRS, Aix-Marseille Université, 13009 Marseille, France
Hall, Sophie; School of Biochemistry, University of Bristol, Biomedical Sciences Building, University Walk, Bristol BS8 1TD, UK ; Max Planck Bristol Centre for Minimal Biology, Cantock’s Close, Bristol BS8 1TS, UK
Redureau, Damien ; Université de Liège - ULiège > Département de chimie (sciences) > Laboratoire de spectrométrie de masse (L.S.M.)
Stenner, Richard; School of Biochemistry, University of Bristol, Biomedical Sciences Building, University Walk, Bristol BS8 1TD, UK ; Max Planck Bristol Centre for Minimal Biology, Cantock’s Close, Bristol BS8 1TS, UK
Boldrini-França, Johara; School of Biochemistry, University of Bristol, Biomedical Sciences Building, University Walk, Bristol BS8 1TD, UK ; Max Planck Bristol Centre for Minimal Biology, Cantock’s Close, Bristol BS8 1TS, UK
Sun, Huan; School of Biochemistry, University of Bristol, Biomedical Sciences Building, University Walk, Bristol BS8 1TD, UK ; Max Planck Bristol Centre for Minimal Biology, Cantock’s Close, Bristol BS8 1TS, UK
Roldão, António ; iBET, Instituto de Biologia Experimental e Technológica, Apartado 12, 2781-901 Oeiras, Portugal ; ITQB NOVA, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa, Av. da República, 2780-157 Oeiras, Portugal
Alves, Paula M. ; iBET, Instituto de Biologia Experimental e Technológica, Apartado 12, 2781-901 Oeiras, Portugal ; ITQB NOVA, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa, Av. da República, 2780-157 Oeiras, Portugal
Harrison, Robert A.; Centre for Snakebite Research & Interventions, Liverpool School of Tropical Medicine, Pembroke Place, Liverpool L3 5QA, UK ; Centre for Drugs & Diagnostics, Liverpool School of Tropical Medicine, Pembroke Place, Liverpool L3 5QA, UK
Vincentelli, Renaud ; Architecture et Fonction des Macromolécules Biologiques, CNRS, Aix-Marseille Université, 13009 Marseille, France
Berger, Imre ; School of Biochemistry, University of Bristol, Biomedical Sciences Building, University Walk, Bristol BS8 1TD, UK ; Max Planck Bristol Centre for Minimal Biology, Cantock’s Close, Bristol BS8 1TS, UK
Quinton, Loïc ; Université de Liège - ULiège > Département de chimie (sciences) > Chimie biologique
Casewell, Nicholas R. ; Centre for Snakebite Research & Interventions, Liverpool School of Tropical Medicine, Pembroke Place, Liverpool L3 5QA, UK ; Centre for Drugs & Diagnostics, Liverpool School of Tropical Medicine, Pembroke Place, Liverpool L3 5QA, UK
Schaffitzel, Christiane ; School of Biochemistry, University of Bristol, Biomedical Sciences Building, University Walk, Bristol BS8 1TD, UK
Harrison R.A. Hargreaves A. Wagstaff S.C. Faragher B. Lalloo D.G. Snake Envenoming: A Disease of Poverty PLoS Negl. Trop. Dis. 2009 3 e569 10.1371/journal.pntd.0000569
Laustsen A.H. María Gutiérrez J. Knudsen C. Johansen K.H. Bermúdez-Méndez E. Cerni F.A. Jürgensen J.A. Ledsgaard L. Martos-Esteban A. Øhlenschlæger M. et al. Pros and cons of different therapeutic antibody formats for recombinant antivenom development Toxicon 2018 146 151 175 10.1016/j.toxicon.2018.03.004 29534892
Chippaux J.P. Williams V. White J. Snake venom variability: Methods of study, results and interpretation Toxicon 1991 29 1279 1303 10.1016/0041-0101(91)90116-9 1814005
Harrison R.A. Casewell N.R. Ainsworth S.A. Lalloo D.G. The time is now: A call for action to translate recent momentum on tackling tropical snakebite into sustained benefit for victims Trans. R. Soc. Trop. Med. Hyg. 2019 113 835 838 10.1093/trstmh/try134
World Health Organization Guidelines for the Production, Control and Regulation of Snake Antivenom Immunoglobulins 2017 Available online: https://cdn.who.int/media/docs/default-source/biologicals/blood-products/document-migration/antivenomglrevwho_trs_1004_web_annex_5.pdf?sfvrsn=ef4b2aa5_3&download=true (accessed on 3 November 2023)
Abubakar S.B. Abubakar I.S. Habib A.G. Nasidi A. Durfa N. Yusuf P.O. Larnyang S. Garnvwa J. Sokomba E. Salako L. et al. Pre-clinical and preliminary dose-finding and safety studies to identify candidate antivenoms for treatment of envenoming by saw-scaled or carpet vipers (Echis ocellatus) in northern Nigeria Toxicon 2010 55 719 723 10.1016/j.toxicon.2009.10.024
Segura Á. Herrera M. Villalta M. Vargas M. Gutiérrez J.M. León G. Assessment of snake antivenom purity by comparing physicochemical and immunochemical methods Biologicals 2013 41 93 97 10.1016/j.biologicals.2012.11.001
Rawat S. Laing G. Smith D.C. Theakston D. Landon J. A new antivenom to treat eastern coral snake (Micrurus fulvius fulvius) envenoming Toxicon 1994 32 185 190 10.1016/0041-0101(94)90107-4
Herrera M. Paiva O.K. Pagotto A.H. Segura Á. Serrano S.M.T. Vargas M. Villalta M. Jensen S.D. León G. Williams D.J. et al. Antivenomic Characterization of Two Antivenoms Against the Venom of the Taipan, Oxyuranus scutellatus, from Papua New Guinea and Australia Am. J. Trop. Med. Hyg. 2014 91 887 894 10.4269/ajtmh.14-0333
Harrison R.A. Gutiérrez J.M. Priority Actions and Progress to Substantially and Sustainably Reduce the Mortality, Morbidity and Socioeconomic Burden of Tropical Snakebite Toxins 2016 8 351 10.3390/toxins8120351
Pucca M.B. Cerni F.A. Janke R. Bermúdez-Méndez E. Ledsgaard L. Barbosa J.E. Laustsen A.H. History of Envenoming Therapy and Current Perspectives Front. Immunol. 2019 10 1598 10.3389/fimmu.2019.01598
Gutiérrez J.M. Albulescu L.-O. Clare R.H. Casewell N.R. Abd El-Aziz T.M. Escalante T. Rucavado A. The Search for Natural and Synthetic Inhibitors That Would Complement Antivenoms as Therapeutics for Snakebite Envenoming Toxins 2021 13 451 10.3390/toxins13070451
Jenkins T.P. Fryer T. Dehli R.I. Jürgensen J.A. Fuglsang-Madsen A. Føns S. Laustsen A.H. Toxin Neutralization Using Alternative Binding Proteins Toxins 2019 11 53 10.3390/toxins11010053 30658491
Lewin M.R. Carter R.W. Matteo I.A. Samuel S.P. Rao S. Fry B.G. Bickler P.E. Varespladib in the Treatment of Snakebite Envenoming: Development History and Preclinical Evidence Supporting Advancement to Clinical Trials in Patients Bitten by Venomous Snakes Toxins 2022 14 783 10.3390/toxins14110783 36422958
Abouyannis M. FitzGerald R. Ngama M. Mwangudzah H. Nyambura Y. Ngome S. Riako D. Babu L. Lewa F. Else L. et al. TRUE-1: Trial of Repurposed Unithiol for snakebite Envenoming phase 1 (safety, tolerability, pharmacokinetics and pharmacodynamics in healthy Kenyan adults) Wellcome Open Res. 2022 7 90 10.12688/wellcomeopenres.17682.1 35372700
Vragniau C. Bufton J.C. Garzoni F. Stermann E. Rabi F. Terrat C. Guidetti M. Josserand V. Williams M. Woods C.J. et al. Synthetic self-assembling ADDomer platform for highly efficient vaccination by genetically encoded multiepitope display Sci. Adv. 2019 5 eaaw2853 10.1126/sciadv.aaw2853 31620562
Besson S. Vragniau C. Vassal-Stermann E. Dagher M.C. Fender P. The Adenovirus Dodecahedron: Beyond the Platonic Story Viruses 2020 12 718 10.3390/v12070718 32630840
Chevillard C. Amen A. Besson S. Hannani D. Bally I. Dettling V. Gout E. Moreau C.J. Buisson M. Gallet S. et al. Elicitation of potent SARS-CoV-2 neutralizing antibody responses through immunization with a versatile adenovirus-inspired multimerization platform Mol. Ther. 2022 30 1913 1925 10.1016/j.ymthe.2022.02.011
Luo C. Yan Q. Huang J. Liu J. Li Y. Wu K. Li B. Zhao M. Fan S. Ding H. et al. Using Self-Assembling ADDomer Platform to Display B and T Epitopes of Type O Foot-and-Mouth Disease Virus Viruses 2022 14 1810 10.3390/v14081810
Buzas D. Bunzel H.A. Staufer O. Milodowski E.J. Edmunds G.L. Bufton J.C. Mateo B.V.V. Yadav S.K.N. Gupta K. Fletcher C. et al. In vitro generated antibodies guide thermostable ADDomer nanoparticle design for nasal vaccination and passive immunization against SARS-CoV-2 Antib. Ther. 2023 tbad024 10.1093/abt/tbad024
Zochowska M. Paca A. Schoehn G. Andrieu J.-P. Chroboczek J. Dublet B. Szolajska E. Adenovirus Dodecahedron, as a Drug Delivery Vector. Jagetia GC, editor PLoS ONE 2009 4 e5569 10.1371/journal.pone.0005569 19440379
Buzas D. Sun H. Toelzer C. Yadav S.K.N. Borucu U. Gautam G. Gupta K. Bufton J. Capin J. Sessions R.B. et al. Engineering the ADDobody protein scaffold for generation of high-avidity ADDomer super-binders BioRxiv. 2023 10.1101/2023.09.09.556966
Gutiérrez J.M. Theakston R.D.G. Warrell D.A. Confronting the Neglected Problem of Snake Bite Envenoming: The Need for a Global Partnership PLoS Med. 2006 3 e150 10.1371/journal.pmed.0030150 16729843
Warrell D.A. Davidson N.M.c.D. Greenwood B.M. Ormerod L.D. Pope H.M. Watkins B.J. Prentice C.R.M. Poisoning by bites of the saw-scaled or carpet viper (Echis carinatus) in Nigeria Q. J. Med. 1977 46 33 62 866568
Dingwoke E.J. Adamude F.A. Mohamed G. Klein A. Salihu A. Abubakar M.S. Sallau A.B. Venom proteomic analysis of medically important Nigerian viper Echis ocellatus and Bitis arietans snake species Biochem. Biophys. Rep. 2021 28 101164 10.1016/j.bbrep.2021.101164
Rowan E.G. Harvey A.L. Snake toxins from mamba venoms: Unique tools for the physiologist Acta Chim. Slov. 2011 58 689 692
Lauridsen L.P. Laustsen A.H. Lomonte B. Gutiérrez J.M. Toxicovenomics and antivenom profiling of the Eastern green mamba snake (Dendroaspis angusticeps) J. Proteom. 2016 136 248 261 10.1016/j.jprot.2016.02.003
Laustsen A.H. Lomonte B. Lohse B. Fernández J. Gutiérrez J.M. Unveiling the nature of black mamba (Dendroaspis polylepis) venom through venomics and antivenom immunoprofiling: Identification of key toxin targets for antivenom development J. Proteom. 2015 119 126 142 10.1016/j.jprot.2015.02.002
Quinton L. Demeure K. Dobson R. Gilles N. Gabelica V. De Pauw E. New Method for Characterizing Highly Disulfide-Bridged Peptides in Complex Mixtures: Application to Toxin Identification from Crude Venoms J. Proteome Res. 2007 6 3216 3223 10.1021/pr070142t
Dunbar J.P. Fort A. Redureau D. Sulpice R. Dugon M.M. Quinton L. Venomics Approach Reveals a High Proportion of Lactrodectus-like Toxins in the Venom of the Noble False Widow Spider Steatoda nobilis Toxins 2020 12 402 10.3390/toxins12060402
Calvete J.J. Sanz L. Angulo Y. Lomonte B. Gutiérrez J.M. Venoms, venomics, antivenomics FEBS Lett. 2009 583 1736 1743 10.1016/j.febslet.2009.03.029 19303875
Wilson D. Daly N.L. Venomics: A Mini-Review High Throughput 2018 7 19 10.3390/ht7030019 30041430
Oliveira A.L. Viegas M.F. da Silva S.L. Soares A.M. Ramos M.J. Fernandes P.A. The chemistry of snake venom and its medicinal potential Nat. Rev. Chem. 2022 6 451 469 10.1038/s41570-022-00393-7 37117308
Casewell N.R. Wagstaff S.C. Wüster W. Cook D.A.N. Bolton F.M.S. King S.I. Pla D. Sanz L. Calvete J.J. Harrison R.A. Medically important differences in snake venom composition are dictated by distinct postgenomic mechanisms Proc. Natl. Acad. Sci. USA 2014 111 9205 9210 10.1073/pnas.1405484111 24927555
Morsa D. Baiwir D. La Rocca R. Zimmerman T.A. Hanozin E. Grifnée E. Longuespée R. Meuwis M.-A. Smargiasso N. De Pauw E. et al. Multi-Enzymatic Limited Digestion: The Next-Generation Sequencing for Proteomics? J. Proteome Res. 2019 18 2501 2513 10.1021/acs.jproteome.9b00044 31046285
Ma B. Zhang K. Hendrie C. Liang C. Li M. Doherty-Kirby A. Lajoie G. PEAKS: Powerful software for peptide de novo sequencing by tandem mass spectrometry Rapid Commun. Mass Spectrom. 2003 17 2337 2342 10.1002/rcm.1196 14558135
Ainsworth S. Petras D. Engmark M. Süssmuth R.D. Whiteley G. Albulescu L.-O. Kazandjian T.D. Wagstaff S.C. Rowley P. Wüster W. et al. The medical threat of mamba envenoming in sub-Saharan Africa revealed by genus-wide analysis of venom composition, toxicity and antivenomics profiling of available antivenoms J. Proteom. 2018 172 173 189 10.1016/j.jprot.2017.08.016 28843532
Gutiérrez J.M. Lomonte B. León G. Alape-Girón A. Flores-Díaz M. Sanz L. Angulo Y. Calvete J.J. Snake venomics and antivenomics: Proteomic tools in the design and control of antivenoms for the treatment of snakebite envenoming J. Proteom. 2009 72 165 182 10.1016/j.jprot.2009.01.008
Hanes J. Schaffitzel C. Knappik A. Plückthun A. Picomolar affinity antibodies from a fully synthetic naive library selected and evolved by ribosome display Nat. Biotechnol. 2000 18 1287 1292 10.1038/82407
Dreier B. Plückthun A. Rapid Selection of High-Affinity Antibody scFv Fragments Using Ribosome Display Methods Mol. Biol. 2018 1827 235 268 10.1007/978-1-4939-8648-4_13 30196501
Zimmermann I. Egloff P. Hutter C.A.J. Kuhn B.T. Bräuer P. Newstead S. Dawson R.J.P. Geertsma E.R. Seeger M.A. Generation of synthetic nanobodies against delicate proteins Nat. Protoc. 2020 15 1707 1741 10.1038/s41596-020-0304-x 32269381
Binz H.K. Amstutz P. Kohl A. Stumpp M.T. Briand C. Forrer P. Grütter M.G. Plückthun A. High-affinity binders selected from designed ankyrin repeat protein libraries Nat. Biotechnol. 2004 22 575 582 10.1038/nbt962 15097997
Zahnd C. Spinelli S. Luginbühl B. Amstutz P. Cambillau C. Plückthun A. Directed in Vitro Evolution and Crystallographic Analysis of a Peptide-binding Single Chain Antibody Fragment (scFv) with Low Picomolar Affinity J. Biol. Chem. 2004 279 18870 18877 10.1074/jbc.M309169200 14754898
Rivera-de-Torre E. Rimbault C. Jenkins T.P. Sørensen C.V. Damsbo A. Saez N.J. Duhoo Y. Hackney C.M. Ellgaard L. Laustsen A.H. Strategies for Heterologous Expression, Synthesis, and Purification of Animal Venom Toxins Front. Bioeng. Biotechnol. 2022 9 811905 10.3389/fbioe.2021.811905 35127675
Simonato M. Morbiato L. Zorzi V. Caccin P. Fernández J. Massimino M.L. de Laureto P.P. Tonello F. Production in Escherichia coli, folding, purification and characterization of notexin with wild type sequence and with N-terminal and catalytic site mutations Toxicon 2014 88 11 20 10.1016/j.toxicon.2014.06.009 24951874
Chiou Y.-L. Lin S.-R. Chang L.-S. Mutations on N-terminal region of Taiwan cobra phospholipase A2 result in structurally distorted effects J. Pept. Sci. 2008 14 890 897 10.1002/psc.1020 18288787
Turchetto J. Sequeira A.F. Ramond L. Peysson F. Brás J.L.A. Saez N.J. Duhoo Y. Blémont M. Guerreiro C.I.P.D. Quinton L. et al. High-throughput expression of animal venom toxins in Escherichia coli to generate a large library of oxidized disulphide-reticulated peptides for drug discovery Microb. Cell Fact. 2017 16 6 10.1186/s12934-016-0617-1
Sequeira A.F. Turchetto J. Saez N.J. Peysson F. Ramond L. Duhoo Y. Blémont M. Fernandes V.O. Gama L.T. Ferreira L.M.A. et al. Gene design, fusion technology and TEV cleavage conditions influence the purification of oxidized disulphide-rich venom peptides in Escherichia coli Microb. Cell Fact. 2017 16 4 10.1186/s12934-016-0618-0
Fairhead M. Howarth M. Site-specific biotinylation of purified proteins using BirA Methods Mol. Biol. 2015 1266 171 184 10.1007/978-1-4939-2272-7_12
Nguyen V.D. Hatahet F. Salo K.E. Enlund E. Zhang C. Ruddock L.W. Pre-expression of a sulfhydryl oxidase significantly increases the yields of eukaryotic disulfide bond containing proteins expressed in the cytoplasm of E. coli Microb. Cell Fact. 2011 10 1 10.1186/1475-2859-10-1
Fitzgerald D.J. Berger P. Schaffitzel C. Yamada K. Richmond T.J. Berger I. Protein complex expression by using multigene baculoviral vectors Nat. Methods 2006 3 1021 1032 10.1038/nmeth983 17117155
Tessier D.C. Thomas D.Y. Khouri H.E. Laliberté F. Vernet T. Enhanced secretion from insect cells of a foreign protein fused to the honeybee melittin signal peptide Gene 1991 98 177 183 10.1016/0378-1119(91)90171-7 2016060
Wagstaff S.C. Laing G.D. Theakston R.D.G. Papaspyridis C. Harrison R.A. Bioinformatics and Multiepitope DNA Immunization to Design Rational Snake Antivenom PLoS Med. 2006 3 e184 10.1371/journal.pmed.0030184 16737347
Gutiérrez J.M. Vargas M. Segura Á. Herrera M. Villalta M. Solano G. Sánchez A. Herrera C. León G. In Vitro Tests for Assessing the Neutralizing Ability of Snake Antivenoms: Toward the 3Rs Principles Front. Immunol. 2021 11 617429 Available online: https://www.frontiersin.org/articles/10.3389/fimmu.2020.617429 (accessed on 13 December 2022) 10.3389/fimmu.2020.617429 33505403
Albulescu L.-O. Xie C. Ainsworth S. Alsolaiss J. Crittenden E. Dawson C.A. Softley R. Bartlett K.E. Harrison R.A. Kool J. et al. A therapeutic combination of two small molecule toxin inhibitors provides broad preclinical efficacy against viper snakebite Nat. Commun. 2020 11 6094 10.1038/s41467-020-19981-6
Macêdo J.K.A. Fox J.W. Biological Activities and Assays of the Snake Venom Metalloproteinases (SVMPs) Venom Genomics and Proteomics Gopalakrishnakone P. Calvete J.J. Springer Dordrecht, The Netherlands 2016 211 238 10.1007/978-94-007-6416-3_21
Conlon J.M. Attoub S. Arafat H. Mechkarska M. Casewell N.R. Harrison R.A. Calvete J.J. Cytotoxic activities of [Ser49]phospholipase A2 from the venom of the saw-scaled vipers Echis ocellatus, Echis pyramidum leakeyi, Echis carinatus sochureki, and Echis coloratus Toxicon 2013 71 96 104 10.1016/j.toxicon.2013.05.017
Still K.B.M. Nandlal R.S.S. Slagboom J. Somsen G.W. Casewell N.R. Kool J. Multipurpose HTS Coagulation Analysis: Assay Development and Assessment of Coagulopathic Snake Venoms Toxins 2017 9 382 10.3390/toxins9120382
Navred K. Martin M. Ekdahl L. Zetterberg E. Andersson N.G. Strandberg K. Norstrom E. A simplified flow cytometric method for detection of inherited platelet disorders—A comparison to the gold standard light transmission aggregometry PLoS ONE 2019 14 e0211130 10.1371/journal.pone.0211130
Kolvekar N. Bhattacharya N. Sarkar A. Chakrabarty D. How snake venom disintegrins affect platelet aggregation and cancer proliferation Toxicon 2023 221 106982 10.1016/j.toxicon.2022.106982
Patel R.N. Clare R.H. Ledsgaard L. Nys M. Kool J. Laustsen A.H. Ulens C. Casewell N.R. An in vitro assay to investigate venom neurotoxin activity on muscle-type nicotinic acetylcholine receptor activation and for the discovery of toxin-inhibitory molecules Biochem. Pharmacol. 2023 216 115758 10.1016/j.bcp.2023.115758 37604290
Slagboom J. Otvos R.A. Cardoso F.C. Iyer J. Visser J.C. van Doodewaerd B.R. McCleary R.J. Niessen W.M. Somsen G.W. Lewis R.J. et al. Neurotoxicity fingerprinting of venoms using on-line microfluidic AChBP profiling Toxicon 2018 148 213 222 10.1016/j.toxicon.2018.04.022 29730150
Servent D. Winckler-Dietrich V. Hu H.-Y. Kessler P. Drevet P. Bertrand D. Ménez A. Only Snake Curaremimetic Toxins with a Fifth Disulfide Bond Have High Affinity for the Neuronal α7 Nicotinic Receptor J. Biol. Chem. 1997 272 24279 24286 10.1074/jbc.272.39.24279 9305882
Harvey A.L. Twenty years of dendrotoxins Toxicon 2001 39 15 26 10.1016/S0041-0101(00)00162-8 10936620
Yajuan X. Xin L. Zhiyuan L. A Comparison of the Performance and Application Differences between Manual and Automated Patch-Clamp Techniques Curr. Chem. Genom. 2012 6 87 92 10.2174/1875397301206010087 23346269
Theakston R.D. Reid H.A. Development of simple standard assay procedures for the characterization of snake venom Bull. World Health Organ. 1983 61 949 956 6609011
Knudsen C. Casewell N.R. Lomonte B. Gutiérrez J.M. Vaiyapuri S. Laustsen A.H. Novel Snakebite Therapeutics Must Be Tested in Appropriate Rescue Models to Robustly Assess Their Preclinical Efficacy Toxins 2020 12 528 10.3390/toxins12090528
Ikonomou L. Schneider Y.-J. Agathos S.N. Insect cell culture for industrial production of recombinant proteins Appl. Microbiol. Biotechnol. 2003 62 1 20 10.1007/s00253-003-1223-9
Yee C.M. Zak A.J. Hill B.D. Wen F. The Coming Age of Insect Cells for Manufacturing and Development of Protein Therapeutics Ind. Eng. Chem. Res. 2018 57 10061 10070 10.1021/acs.iecr.8b00985
Maghodia A.B. Geisler C. Jarvis D.L. Characterization of an Sf-rhabdovirus-negative Spodoptera frugiperda cell line as an alternative host for recombinant protein production in the baculovirus-insect cell system Protein Expr. Purif. 2016 122 45 55 10.1016/j.pep.2016.02.014
Maghodia A.B. Geisler C. Jarvis D.L. A new nodavirus-negative Trichoplusia ni cell line for baculovirus-mediated protein production Biotechnol. Bioeng. 2020 117 3248 3264 10.1002/bit.27494 32662870
Fuenmayor J. Gòdia F. Cervera L. Production of virus-like particles for vaccines New Biotechnol. 2017 39 174 180 10.1016/j.nbt.2017.07.010 28778817
Roldão A. Mellado M.C.M. Castilho L.R. Carrondo M.J. Alves P.M. Virus-like particles in vaccine development Expert Rev. Vaccines 2010 9 1149 1176 10.1586/erv.10.115 20923267
Junne S. Neubauer P. How scalable and suitable are single-use bioreactors? Curr. Opin. Biotechnol. 2018 53 240 247 10.1016/j.copbio.2018.04.003 29753977
Gerstweiler L. Bi J. Middelberg A.P.J. Continuous downstream bioprocessing for intensified manufacture of biopharmaceuticals and antibodies Chem. Eng. Sci. 2021 231 116272 10.1016/j.ces.2020.116272
Gerstweiler L. Billakanti J. Bi J. Middelberg A.P.J. An integrated and continuous downstream process for microbial virus-like particle vaccine biomanufacture Biotech. Bioeng. 2022 119 2122 2133 10.1002/bit.28118
Mvundura M. Kien V.D. Nga N.T. Robertson J. Van Cuong N. Tung H.T. Hong D.T. Levin C. How much does it cost to get a dose of vaccine to the service delivery location? Empirical evidence from Vietnam’s Expanded Program on Immunization Vaccine 2014 32 834 838 10.1016/j.vaccine.2013.12.029
Mvundura M. Lorenson K. Chweya A. Kigadye R. Bartholomew K. Makame M. Lennon T.P. Mwangi S. Kirika L. Kamau P. et al. Estimating the costs of the vaccine supply chain and service delivery for selected districts in Kenya and Tanzania Vaccine 2015 33 2697 2703 10.1016/j.vaccine.2015.03.084
Kumar R. Srivastava V. Baindara P. Ahmad A. Thermostable vaccines: An innovative concept in vaccine development Expert Rev. Vaccines 2022 21 811 824 10.1080/14760584.2022.2053678
Daniell H. Rai V. Xiao Y. Cold chain and virus-free oral polio booster vaccine made in lettuce chloroplasts confers protection against all three poliovirus serotypes Plant Biotechnol. J. 2019 17 1357 1368 10.1111/pbi.13060
Guo M. Li J. Teng Z. Ren M. Dong H. Zhang Y. Ru J. Du P. Sun S. Guo H. Four Simple Biomimetic Mineralization Methods to Improve the Thermostability and Immunogenicity of Virus-like Particles as a Vaccine against Foot-and-Mouth Disease Vaccines 2021 9 891 10.3390/vaccines9080891 34452016
Cardoso F.M.C. Petrovajová D. Horňáková T. Viral vaccine stabilizers: Status and trends Acta Virol. 2017 61 231 239 10.4149/av_2017_301 28854787